1- Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD:-
Review of published literature spanning 20 years from 2000 to 2020. EBV-positive post-transplant lymphoproliferative disease (PTLD) is a major complication of hematopoietic stem cell and solid organ transplantation.
In clinical trials, adoptive cellular therapy using EBVSTs has shown impressive results in immunocompromised patients.
The Epstein-Barr virus (EBV) is a ubiquitous member of.the gamma herpes virus family that is associated with avariety of lymphomas and lymphoproliferative disorders.
Primary infection occurs in childhood as an asymptomatic or mild infection and/or may result in a more florid infectious mononucleosis syndrome in teenagers and young adults.
The Ebola virus (EBV) causes the production and proliferation of type 3 latency EBV-infected B cells resulting in EBV-associated lymphoproliferative disease (LPD and malignancies).
These B-lymphocytes present several antigenic viral proteins including EBNA 1-3, LMP1 and LMP2 that induce potent EBVST responses.
Such immune responses are lacking in immunosuppressed individuals leading to the proliferation of infected B cells.
EBV-associated post-transplant lymphoproliferative disease (PTLD) occurs in less than 1% to 25% of HSCT recipients.
The biggest risk factor for developing PTLD is EBV seropositivity at the time of transplant.
Rituximab, monoclonal antibody targeting CD20 present on B cells has been an effective monotherapy
EBVSTs can be expanded via ex-ex vivo expansion of T cells targeting viral antigens via T cell receptors.
LCL-activated EBV transformed lymphoblastoid cell lines (LCL) express high levels of class I and class II HLA and co-stimulatory molecules.
================================================================= Antigen-specific T cell selection
The rapid selection of CD8 + T cells with high affinity to a specific viral epitope/peptide in an HLA-restricted manner is one of the fastest and most effective ways to identify new drug candidates for use in the field of immunotherapy.
Autologous stem cell therapy and multimer-based approaches have been developed in the past, but their development has been hampered by a number of challenges, such as the HLA-restriction requirement and selection of a purely CD8 + T cell product that lacks CD4+ T cell help.
However, using both of these techniques, GMP-grade EBVST products can be made quickly from EBV seropositive autologous and allogeneic donors for clinical use.
==================================================================== Post-HSCT donor-derived T cell therapy
Donor lymphocyte infusions
This therapy carries a significant risk of graft-versus-host disease, or graft versus lymphoma (GVHD), and strategies have been employed to lessen the risk.
Donor-derived EBVSTs
Therapy was well tolerated with remarkable reduction of EBV viral copy numbers within 4 weeks.
None of the 101 patients who received donor derived EBVSTs as prophylaxis developed PTLD.
Of the 13 patients with active PTLD, 11 patients achieved a complete remission with evidence of VST persistence up to 9 years after transplant.
Autologous EBVSTs
Administration of autologous EBVSTs has been used for patients with EBV-associated malignancies outside the context of allogeneic HSCT and in SOT recipients with PTLD.
In the SOT setting, the production is technically more challenging because of the ongoing immunosuppression.
Autologous EBVSTs have been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy or in relapsed patients.
Such tumors are more difficult to treat because of the reduced expression of immunogenic viral antigens.
EBVST products produced by LCL stimulation alone have shown promising results in patients with type II latency-type malignancies (50,51).
A pilot study of 14 patients with relapsed EBV + HL reported 5 patients who maintained a CR for up to 40 months.
Production of EBVSTs was successful in 91% of patients, with LMP1 and/or LMP2-specificity detected in 66% of products .
A total of 50 patients with EBV-associated HL or Non-Hodgkin lymphoma were treated.
Of 29 patients receiving latent membrane protein (LMP)-specific T cells as adjuvant therapy after chemotherapy, 28 patients remained in a complete remission.
Adjuvant treatment with LMP-specific T cell therapy for patients with metastatic Non Hodgkin’s lymphoma (NPC) led to a significant increase in survival compared to a control group, according to a study.
The study was conducted at the Queensland Institute of Medical Research (QIMR) (Brisbane, Australia).
Third-party T cells
Patients with rapidly progressive disease may not be able to access patient-specific T cell therapies due to procurement times of donor or patient cells.
Patient-specific EBVST (autologous or allogeneic) manufacture can still be prolonged when procurement times are considered.
For these reasons, a readily available T cell therapy product is often the preferred treatment.
Thirty-three transplant recipients (stem cell, heart, kidney, liver, 10; liver and small bowel, 3; lung, 2; heart and lung, 1) with refractory PTLD between the age of 1–76 years received partially HLA matched EBVSTs.
Overall, the response rate (CR and PR) was 64% at 5 weeks and 52% at 6 months.
Chiou et al. from Birmingham, United Kingdom published experience in 10 pediatric SOT recipients with PTLD and reported an ORR of 80% (8 out of 10) .
This may indicate possible differences in the biology of EBV-driven PTLD in this population who is often EBV naïve at the time of transplant.
The BCM group has published extensively on their third-party EBVST products including the use of off-the-shelf VSTs for patients with severe refractory viral disease after BMT.
In a multicenter study, 15 products were given to 50 HSCT recipients with EBV, CMV, adenovirus, BKV and human herpes virus 6 (HHV6) .
The Memorial Sloan Kettering experience utilizing a third-party cell bank comprising 330 GMP-grade EBVSTs was reported by Prockop et al.
A total of 46 patients post-HSCT (33) or SOT (13) and PTLD were treated with three weekly infusions of the products.
Third-party EBVSTs have been mostly used in the post- transplant setting.
There has been no published experience with patients with HIV-associated lymphomas.
No standardization of EBV viral load measurements by polymerase chain reaction makes comparisons between different laboratories impossible.
Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo
TGF-β can be released into the tumor microenvironment by tumor cells, fibroblasts and immune cells and creates a immunosuppressive environment by impeding T-cell activation, proliferation and migration .
In addition, it affects macrophage antigen presentation and chemoattractant presentation.
Modification of EBVSTs to overcome these immune suppressive effects has led to increased activity in patients with latency type I and II.
The potential to overcome the immunosuppressive properties of TGF-β has been studied in EBV-positive patients.
The Baylor group developed an EBV/LMP-specific LST product expressing a dominant negative TGF-beta receptor type II (DNRII).
Calcineurin resistance to enhance EBVSTs
Some groups have explored gene-engineering of virus specific T cells to render them resistant to immune suppressive agents including steroids and calcineurin inhibitors.
In mouse xenograft models bearing human B-cell lymphoma, treatment with CNI-resistant EBVSTs produced enhanced activity in the presence of CNI compared to control EBVS.
Chimeric antigen receptor T cells
(CARTs) have shown impressive efficacy in acute B-lymphoblastic leukemia and nin B-cell lymphomas.
In patients with Epstein-Barr Virus-Lymphoma (EBV-LD, or PTLD), the wider use of CARTs will likely require an off- the-shelf product.
The use of adoptive immunotherapy, particularly using third-party “off the shelf” EBVSTs for the treatment of EBV-LD has shown promise in several studies.
Preclinical work and early clinical trials are in process exploring various gene engineering and combination therapy approaches to improve the potency of these therapies in vitro.
2- What are the indications of adoptive immunotherapy?
1- If conventional tratment fails.
2- It uses EBV-specific cytotoxic T lymphocytes(EBV-CTLs)or donor lymphocyte infusion(DLI)in attempt to about the EBV-driven proliferation of B cells in EBV -association PTLD.
3- Prevention and remmision of EBV-induced PTLD have been achieved in 52 to 75% of pateint .
3- an adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
1- Adoptive cellular treatment using EBVSTs has produced outstanding outcomes in clinical trials for immunocompromised patients with EBV-LD, and there is potential for wider application.
2- The focus of current research is on genetic engineering and combination therapy to increase EBVST potency in vivo.
EBV is a virus related to lymphoproliferative disorders, it infects 90% of adults . Primary infection is mainly asymptomatic or may lead to infectious mononucleosis.
In healthy seropositive individuals, virus neutralizing antibodies control the spread of infectious virus particles and EBV specific HlA class I restricted CD8 cytotoxic T lymphocyte specific to the early lytic cycle proteins kill cells entering the lytic cycle before they are able to release infectious viral particles.
In immunocompetent individuals, the frequency of EBV infected B cells remain low and stable by a potent EBVST ( EBV specific T lymphocytes ) response.
In immunocompromised ones, the lack of apotent EBVS..T. response can lead to uncontrolled proliferation of type 3 latency EBV infected B cells lead to EBV associated LPD.
The rationale of adoptive transfer of EBVSTs is based on harnessing the immunogenicity of type III latency malignancy to control the proliferation of the latent infected B cells
PTLD is highly immunogenic and amenable to immunotherapy with EBVSTs
Manufacturing EBVSTs
Methods
1. Ex vivo expansion of EBVSTs
2. Antigen specific T cell selection
Post HSCT donor derived T cell therapy
1. Donor lymphocyte infusi ons
2. Donor derived EBVSTs
3. Autologous EBVSTs
4. Third party T cell
Modification of EBVSTs to enhance activity
1. Overcoming the immune suppressi effects of TGF-B to enhance EBVSTs activity in vivo
2. Calcineurin resistance to enhance EBVSTs activity
3. Chimeric antigen receptor T cells
Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance EBVSTs activity in vivo
. Demethylating agents
. Checkpoint inhibitors
. BCL-2 inhibitors
Manufacturing of EBVSTs
●EBVSTs can be readily produced from EBV-positive donors
●Donor types include autologous and allogeneic
●The most commonly utilized methods consist of ex vivo expansion of VSTs versus antigen-specific T cell selection [e.g., interferon γ (IFN-γ) capture]
Ex vivo expansion of EBVSTs
○ EBVSTs were selectively expanded utilizing irradiated EBV transformed (LCL) (that express a type III latency pattern of EBV antigen expression) as (APCs) to selectively expand EBVSTs
○LCL-activated EBVSTs consist of a product with activity against early lytic antigens and EBNA 3A, 3B and 3C but unreliable activity towards LMP1 and LMP2
○To enhance the specificities to LMP1 and LMP2, several groups have made further modifications to this approach by transducing DC and LCLs with adenovirus vectors to overexpress LMP1 or LMP2
○ this approach is complex and time consuming
○Subsequently the use of APC such as (DC) can facilitate the rapid (10–21 days as opposed to 2–3 months).
○A further benefit of ex vivo expansion approaches that utilize whole antigen, is that EBVSTs can be manufactured from individuals irrespective of their HLA type.
Antigen-specific T cell selection Other rapid methods include:
(I) (MHC)
(II) IFN-γ capture approaches
They require seropositive donors with high frequency of circulating antigen/epitope specific T cells which may be technically challenging in the autologous setting.
Post-HSCT donor-derived T cell therapy Donor lymphocyte infusions
●This therapy carries a significant risk of (GVHD)
●Donor-derived EBVSTs
For the prevention and treatment of EBV+ PTLD in the post-HSCT setting
● This therapy was well tolerated without significant complications with remarkable reduction in EBV viral copy numbers within 4 weeks, including in a patient with immunoblastic lymphoma.
●This pivotal study established the safety and early evidence of efficacy of donor-derived EBVSTs for the treatment and prophylaxis of PTLD in transplant recipients
● GVHD rates in this study were low
Studies using allogeneic VSTs in EBV+ PTLD in HSCT recipients
□BCM, Houston, TX, USA
□MSKCC, New York,
□Karolinska Institute, Stockholm, Sweden
□Children’s Research Hospital, Kyoto, Japan (47)
Autologous EBVSTs
◇ It is used outside the context of allogeneic HSCT and in SOT recipients with PTLD where donors usually are not available because of the use of cadaveric grafts.
◇It is technically more challenging because of the ongoing immunosuppression but this can be overcome with modern manufacturing approaches
◇In theory, autologous EBVSTs are preferrable to donor EBVSTs even if available in the post-SOT setting because PTLD is usually of recipient origin and solid organ grafts are not routinely HLA matched to the recipient.
◇the challenges
the production time that will lead to delays in initiation of therapy in patients with a rapidly progressive disease
◇Autologous EBVSTs have also been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients.
◇Such tumors are more challenging targets because of the reduced expression of immunogenic viral antigens in these type II latency tumors which express a more restricted array of antigens (e.g., LMP1, LMP2, EBNA1 and BARF1) compared to type III latency tumors.
Third-party T cells
●It needs a prolonged time
Donor cells may not be available (e.g., recipients of umbilical cord blood transplants or cadaveric organ transplants
●For the above reasons, a readily available “off the shelf” T cell therapy product is desirable.
The first third-party EBVST bank of 60 EBVST products was established by Haque et al. in the United Kingdom
no significant toxicities were observed,
●The UK group established a cell bank of 25 donors with HLA alleles prevalent at high frequencies in individuals of European descent
●Chiou et al. from Birmingham, United Kingdom published their experience in 10 pediatric SOT recipients with PTLD and reported an ORR of 80% (8 out of 10)
●This favorable response in a pediatric population may indicate differences in the biology of EBV-driven PTLD in this population who is often EBV naïve at the time of transplant and develops PTLD in the earlier post-transplant period compared to the adult population.
Conclusions .
Further work is however needed to create widely available and commercialized third-party cell banks .
Strategies need to be explored to enhance the anti-tumor activity of EBVST therapies especially for the less immunogenic type I and II latency tumors.
Preclinical work and early clinical trials are in process exploring various gene engineering and combination therapy approaches to improve the potency of EBVSTs in vivo.
The GMP (good manufacturing-practice) grade EBV-specific T-cell manufacturing involve mononuclear cells which are harvested from the peripheral blood of a donor. B-lymphocytes are infected with laboratory strain EBV and transduced with an adenoviral vector expressing latent membrane proteins: LMP1 and LMP2 and irradiated. Monocytes are separately transduced with the same adenoviral vector and cocultured with T cells followed by a second stimulation by the transduced lymphoblastoid cell lines to create and further expand LMP-specific T-cell product. Methods are:
1-Ex vivo expansion of T cells targeting viral antigens via native T cell receptors was established initially by Smith et al. where EBVSTs were selectively expanded utilizing irradiated EBV transformed lymphoblastoid cell lines as antigen presenting cells to selectively expand EBVSTs.
2-major histocompatibility complex (MHC) multimer selection where oligomeric forms of MHC molecules are designed and conjugated to magnetic beads to isolate typically CD8+ T cells with high affinity to a specific viral epitope/peptide in an HLA restricted manner.
3- IFN-γ capture approaches where mononuclear cells are pulsed with antigen [e.g., single peptides, overlapping peptides (pepmixes), to isolate CD8+ and CD4+ T cells which secrete IFN-γ in response to the viral antigens.
Indications:
prevention and treatment of EBV+ PTLD in the post-HSCT
EBV-associated malignancies outside the context of allogeneic HSCT and in SOT recipients with PTLD
Autologous EBVSTs have also been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients
Side effects
1- graft-versus-host disease
2- graft versus lymphoma
Latency III is a stage in which B-lymphocytes that have been infected with EBV produce certain proteins that are part of the EBV genome. These proteins include the EBV nuclear antigens EBNA1, 2, 3, EBNALP, LMP1, 2, and BARF1. This high antigenic expression is linked to a powerful T cell response, which is responsible for limiting the growth of B cells to less than 2%. Latency III stage is connected with lymphoproliferative illness and malignancy after the patient is immunosuppressed, which causes the B cells to start developing uncontrolled, unchecked proliferation. PTLD is the most prevalent form of malignancy that is associated with this stage.
On the other hand, the latency II and I stages are less immunogenic. This is because, during these stages, there is a downregulation of the immunodominant antigens, such as EBNA3 and EBNA2. Latency I and II stages are associated with the development of lymphoproliferative diseases and malignancies in patients who are immune-competent as well as immune-suppressed. These lymphoproliferative diseases and malignancies include Hodgkin’s lymphoma, natural killer/T-cell lymphoma, nasopharyngeal carcinoma (Latency II), and Burkitt lymphoma or gastric carcinoma (Latency I).
Post-transplant lymphoproliferative disorder (PTLD) is a dangerous complication that can arise after solid organ transplantation. The activation of EBV viruses picked up during infancy accounts for about 95% of cases, and 95% of cases are of the B cell type, while only 5% of cases are of the T cell lymphoma variety.
After the initial stage of EBV infection, the virus infects B cells and progresses through the next two key stages:
Latency stage in which the virus becomes latent in B cells for life, and at this stage the patient is asymptomatic but at risk of developing certain lymphoproliferative diseases. At this stage, several stages of latency develop (latency III, II, and I). Lytic stage may be asymptomatic, or the patient may develop mild symptoms or frank infectious mononucleosis. Latency stage, in which the virus becomes latent in B cells life long and at this stage the patient is a
Q1: EBV is a member of herpes viruses causing a primary disease such as asymptomatic or infectious mononucleosis. After primary infection, EBV remains in B cells and oral epithelial cells as latent viruses. Viral antigens like EBNA 1-3, and LMP 1-2, are expressed during type III latency. After entrance to lymphoid follicles, infected B cells downregulate the expression of these proteins, so that not to recognize by EBV-specific T cells (a kind of cytotoxic T cells or, EBVSTs). In immunocompetent persons, these infected B cells are controlled by EBVSTs response lifelong. However, in immunosuppressed transplanted recipients this response is lacking and results in excessive proliferation of infected cells and lymphoproliferative disease (LPD). seronegative children before TX are at the highest risk for these LPDS and in more than 90% of cases their origin is CD20-positive B cells. In HSCT, the vast majority respond to rituximab, but in SOT response rate is about 50%. Another regime including cyclophosphamide with prednisone and rituximab is effective but has a low tolerance. Hence, there is need for another treatment. In the past donor lymphocyte infusions (DLI) were used in HSCT or allogenic virus-specific T cells (VSTs) for SOT, but nowadays manufacturing EBVSTs are produced by GMP methodologies against EBV latency proteins. There are two methods: 1. Ex vivo production of EBVSTs 2. Antigon-specific T cell selection by MHC multimer selection or IFN-Ɣ capture methods to isolate CD8+ and CD4+ cells selecting IFN-Ɣ due to viral Ags. DLC caused a significant risk of GVHD. EBVSTs could be donor-derived or autologous or third-party. Donor-derived EBVSTs have been used with success in HSCTs. Autologous EBVSTs have been used in SOT recipients for prevention and treatment with successful results in type III latency and acceptable results in type II latency as adjustment therapy which has lower immunogenicity. third-party EBVSTs, therapy, or even covering CMV, adenovirus, BKV, and HHV6 have produced with acceptable results. Bank of these cells is providing and reduces EBV viral load in these patients. Other productions are available. One of them uses DNPII to overcome the immunosuppressive effect of TC-F-β with downregulation of TGF-β receptors with promising results. Another way was gene engineering of VST to resist them against CNIs and steroids with good results in mice. CD19- chimeric antigen T cell (CD19-CART) is another way but due to cytokine release syndrome and other adverse reactions, is not routinely used. Other combinations of EBVST with other modalities have been used such as demethylating agents to express more EBV antigens or checkpoint inhibitors. To inhibit PD-L1 in classic HL and NHL combined with EBVSTs. In addition, antiapoptotic proteins like BCL-2 are upregulated by LMP1 EBV antigens. Therefore, pretreatment with BCL-2 inhibitor combined with CD19-CART is studied. Hence, these investigations will increase the use of EBVST in the future. Q2: 1. Prevention or treatment of PTLD in HSCT or SOT 2. Treatment of resistant viral infections 3. In combination with other treatments for resistant cases of PTLD or even other malignancies Q3: EBVST could be used for the treatment of a wide variety of viral infections and malignancies, alone or combined with other methods. Q4: 1. Acute and chronic GVHD especially with DLL 2. Cytokine release syndrome in CD19-CART 3. Other side effects such as itching, skin rash, allergic reactions, nausea, vomiting
Level 5 evidence
Introduction:
EBV infection usually occurs in childhood as mild clinical presentation and maintains a lifelong latency in B cells .
The adoptive immunotherapy in the treatment of PTLD in SOT has demonstrated a better clinical outcome with less toxicity .
Adoptive immunosuppressive Side effects :Graft versus host disease ,Ig-E-mediated anaphylaxis reaction ,Flu-like symptoms.
Post-transplant lymphoproliferative disorder (PTLD) is a dangerous complication that can arise after solid organ transplantation. The activation of EBV viruses picked up during infancy accounts for about 95% of cases, and 95% of cases are of the B cell type, while only 5% of cases are of the T cell lymphoma variety.
After the initial stage of EBV infection, the virus infects B cells and progresses through the next two key stages:
Latency stage in which the virus becomes latent in B cells for life, and at this stage the patient is asymptomatic but at risk of developing certain lymphoproliferative diseases. At this stage, several stages of latency develop (latency III, II, and I). Lytic stage, which may be asymptomatic, or the patient may develop mild symptoms or frank infectious mononucleosis. Latency stage, in which the virus becomes latent in B cells life long and at this stage the patient is a
Latency III is a stage in which B-lymphocytes that have been infected with EBV produce certain proteins that are part of the EBV genome. These proteins include the EBV nuclear antigens EBNA1, 2, 3, EBNALP, LMP1, 2, and BARF1. This high antigenic expression is linked to a powerful T cell response, which is responsible for limiting the growth of B cells to less than 2%. Latency III stage is connected with lymphoproliferative illness and malignancy after the patient is immunosuppressed, which causes the B cells to start developing uncontrolled, unchecked proliferation. PTLD is the most prevalent form of malignancy that is associated with this stage.
On the other hand, the latency II and I stages are less immunogenic. This is because, during these stages, there is a downregulation of the immunodominant antigens, such as EBNA3 and EBNA2. Latency I and II stages are associated with the development of lymphoproliferative diseases and malignancies in patients who are immune-competent as well as immune-suppressed. These lymphoproliferative diseases and malignancies include Hodgkin’s lymphoma, natural killer/T-cell lymphoma, nasopharyngeal carcinoma (Latency II), and Burkitt lymphoma or gastric carcinoma (Latency I).
Adoptive immunotherapy is a way to treat PTLD that uses donor lymphocytes or EBV-specific cytotoxic T lymphocytes to kill B cells that are dividing and contain the virus.
In what circumstances does adoptive immunotherapy make sense to use?
Cases of EBV+ PTLD that are resistant to treatment and refractory to treatment are treated as follows:
Treatment of specific types of solid cancers, such as glioblastoma and gastrointestinal tract cancers
Treatment of certain persistent viral infections, such as HIV, employs Treg adoptive immunotherapy; autoimmune diseases treated include type 1 diabetes and systemic lupus erythematosus.
Is there a possibility that adoptive immunotherapy could be used to treat other illnesses that are associated with organ transplantation?
Treg cell infusions have the potential to be employed as a therapy for the treatment of acute rejection.
The treatment of CMV instances that are resistant as well as BK nephropathy
What sorts of adverse reactions are associated with adoptive immunotherapy?
The most serious adverse impact of this treatment is an increased risk of developing graft-versus-host disease. This risk is reduced when EBV-specific cytotoxic T cells are used, but it is extremely high when donor-derived lymphocytes are used.
Reactions caused by allergies
Symptoms that are similar to those of the influenza virus, including a fever, headache, malaise, fatigue, and myalgia
GI tract (GIT) adverse effects can include nausea, vomiting, and diarrhea.
The adoptive cellular immunotherapy for Epstein-Barr virus-associated lymphoproliferative disease (EBV-LD) are going in advance and more widely available including in industry led multi-centre studies and consortium and cooperative group studies. EBV-positive post-transplant lymphoproliferative disease (PTLD) is a major complication of hematopoietic stem cell and solid organ transplantation.
First successes with adoptive immunotherapy were described using donor lymphocyte infusion for the treatment of EBV+ PTLD in hematopoietic stem cell transplantation (HSCT) recipients but can lead to high risk of graft-versus-host disease (GVHD).
EBV Specific T lymphocytes( EBVSTs) can be readily produced from EBV-positive
donors using good-manufacturing-grade (GMP) compliant methodologies . Donor types include autologous and allogeneic (including third party) sources.
Over the years, several strategies have been developed to manufacture EBVST products with minimal alloreactivity and broad specificity against EBV latency proteins or as a
multi-virus specific product with activity against multiple viruses . The most commonly utilized methods consist of ex vivo expansion of VSTs versus antigen-specific T cell selection.
Indications of adoptive immunotherapy include:
1- EBV associated malignancies like PTLD
2- EBV severe infection and complications in immuno-compromised patients
3- Refractory PTLD post organ transplantation
The main side effect is graft versus host disease (GVHD)
The article discusses the Epstein-Barr virus (EBV), a member of the gamma herpes virus family, and its association with various lymphomas and lymphoproliferative disorders (LPD). EBV infects a large percentage of the adult population worldwide and primarily causes asymptomatic or mild infections in childhood. In individuals with a healthy immune system, virus-neutralizing antibodies and specific cytotoxic T lymphocytes (CTLs) control the virus’s spread and kill infected cells. EBV establishes lifelong latency in B cells and oral epithelial cells.
During primary infection, EBV replicates in the oropharynx’s epithelial cells and infects B lymphocytes. Infected B cells express a range of viral proteins, enter lymphoid follicles, downregulate immunogenic proteins, and persist as latent infections in memory B cells. In healthy individuals, the frequency of EBV-infected B cells remains controlled by an effective EBV-specific T-lymphocyte response. However, in individuals with weakened immune systems, such as those with primary immunodeficiency or HIV infection, or recipients of stem cell or solid organ transplants, uncontrolled proliferation of EBV-infected B cells can occur, leading to EBV-associated LPD and malignancies.
The article discusses how the adoptive transfer of EBV-specific T cells (EBVSTs) can be a potential therapeutic approach. EBV-associated LPD, which presents highly immunogenic viral proteins, can be controlled by harnessing the immunogenicity of type III latency malignancies.
The article mentions that EBVSTs have been evaluated for over two decades, and various strategies have been explored, including donor lymphocyte infusions, donor-derived multi-antigen specific VSTs, and autologous or allogeneic VSTs. The review aims to define current manufacturing strategies, summarize clinical experiences, explore opportunities to expand the use of EBV-specific VSTs, and discuss future strategies to enhance their efficacy.
Adoptive immunotherapy has various indications, including the treatment of post-transplant lymphoproliferative disorder (PTLD), prophylaxis and reduction of Epstein-Barr virus (EBV) viral load, and management of refractory cytomegalovirus (CMV), EBV, or adenovirus infections in transplant recipients.
In addition to these indications, adoptive immunotherapy shows potential in the treatment of other conditions related to organ transplantation. It may be explored as a therapeutic option for autoimmune skin diseases and autoimmune conditions such as systemic lupus erythematosus (SLE), type 1 diabetes mellitus (DM I), multiple sclerosis (MS), and inflammatory diseases like atherosclerosis. Furthermore, it could be considered for managing Hodgkin’s lymphoma and nasopharyngeal carcinoma.
As with any medical intervention, adoptive immunotherapy carries the risk of side effects. These can include neurological symptoms such as confusion, headache, delirium, convulsions, and cerebral edema. Anaphylactic reactions are possible in some cases, and patients may experience flu-like symptoms, generalized fatigue, palpitations, and weight gain. There is also a risk of graft-vs-host disease, cytokine release syndrome, and acute kidney injury (AKI) associated with adoptive immunotherapy.
Briefly summarize the principle of adoptive immunotherapy in the treatment of PTLDThis article has a level V evidence because it is narrative review.90% of general population are affected with EBV, in immune-competent persons, EBV infection is well controlled, rarely it presents with fulminant disease, in other cases EBV will stay in latent infection in B-cells. In immune-compromised subjects like solid organ transplant recipients, EBV infection can lead to lympho-proliferative disorders.EBV viral proteins (such as EBNA, LMP1, and LMP2) will be presented on the surface of B-cells in EBV- associated lympho-proliferative disease.Malignancies associated with type II latency include: Hodgkin’s lymphoma, natural killer T-cell lymphoma and Nasopharyngeal carcinoma.Malignancies associated with type I latency tumors include: Burkitt’s lymphoma and gastric carcinoma.PTLD is commonly seen in EBV negative recipients receiving organs from EBV positive donors.EBV-specific T-cells (EBVSTs) were studied for prevention and treatment of EBV related cancers. EBVSTs are produced from EBV positive donors who can be autologous or allogenic sources.EBVSTs can be manufactured via the following strategies:1- Ex-vivo expansion of VSTs vs Antigen-specific T-cell selection.2- MHC molecules are formed and attached to magnetic beads to separate typical CD8+ T-cells specific to viral antigens in HLA restricted manner. 3- IFN-gamma capture uses mononuclear cells pulsed with antigen to separate CD8+ &CD4+ T-cells that respond to viral antigens by secreting IFN-gamma.
What are the indications of adoptive immunotherapy?
a- PTLD.
b- Treatment prophylaxis and EBV viral load reduction.
c- Refractory CMV or EBV or adenovirus infection.
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
4- Auto-immune skin diseases.
5- Auto-immune conditions such as SLE, DM I, and MS.
6- Inflammatory diseases, atherosclerosis.
7- Hodgkin’s Lymphoma.
8- Nasopharyngeal carcinoma..
What are the side effects of adoptive immunotherapy?
1- Confusion, headache, delirium, convulsion, and cerebral edema.
Epstein-Barr virus (EBV) is a ubiquitous member of the gamma herpes virus family that is associated with a variety of lymphomas and lymphoproliferative disorders (LPD). It infects more than 90% of the adult population worldwide. Primary infection occurs in childhood as a asymptomatic or mild infection and/or may result in a more florid infectious mononucleosis syndrome in teenagers and young adults.
Regardless of the initial infection, EBV maintains a lifelong latency in B cells and oral epithelial cells. During primary infection, EBV enters the oropharynx replicating within the epithelial cells and infect transiting B-lymphocytes (primarily due to their expression of CD21 which is the major receptor for the virus). EBV can also infect epithelial cells via transfer from infected B cells and other processes.
Donor lymphocyte infusions
The earliest reported experience of cellular therapy for the treatment of PTLD utilized unmodified DLI derived from the patient’s EBV seropositive HSCT donor which contained effector cells with activity against EBV (37). Although effective in inducing remissions, this therapy carries a significant risk of graft-versus-host disease (GVHD).
Donor-derived EBVSTs
There is extensive reported experience using donor derived EBVSTs for the prevention and treatment of EBV+ PTLD in the post-HSCT setting when the donor is available.
In the original report using ex vivo expanded EBVSTs from healthy seropositive donors, the team at St. Jude’s Research Hospital treated ten allogeneic HSCT recipients, three with evidence of EBV reactivation and seven at high risk of reactivation.
Demethylating agents
Newly EBV-infected B-cells express up to 90 viral genes; however rapid CpG-methylation of viral antigens leads to downregulation of viral protein expression and the latency .
Azacytidine and decitabine are potent CpG demethylating agents. In a mouse xenograft of latency type I Burkitt lymphoma, pretreatment with decitabine induced expression of LMP1 and ENBA3 associated with latency type 3 which sensitized tumor cells to subsequent therapy with EBVSTs (86). In contrast, azacytidine did not increase expression of those proteins.
Conclusions
The use of adoptive immunotherapy, particularly using third-party “off the shelf” EBVSTs for the treatment of EBV-LD has shown promise in several studies conducted at specialized centers .More recently, EBVSTs have become more widely available including in industry led multi-center studies and consortium and cooperative group studies (20,66). Further work is however needed to create widely available and commercialized third-party cell banks to broaden the applicability of this approach beyond boutique centers. In addition, strategies need to be explored to enhance the anti-tumor activity of EBVST therapies especially for the less immunogenic type I and II latency tumors. Preclinical work and early clinical trials are in process exploring various gene engineering and combination therapy approaches to improve the potency of EBVSTs in vivo.
The principle of adoptive immunotherapy in PTLD treatment:
It involves generating EBV-specific T lymphocytes (EBVSTs) from EBV-positive autologous or allogenic donors with limited alloreactivity and wide virus-fighting activity.
It is necessary to use ex vivo virus-specific T cell growth or antigen-specific T cell selection.
MHC selection involves isolating CD8+ T cells with high affinity to a specific viral epitope or peptide in an HLA-restricted manner; IFN- capturing involves isolating CD8+ and CD4+ T cells that release IFN- in response to viral antigens.
Technically challenging approaches.
Post-HSCT donor-derived T cells
Donor-lymphocyte infusions
Unmodified DLI from PTLD patients’ EBV-positive HSCT donors induces remission.
Donor-derived EBVSTs may prevent and treat EBV+ PTLD in post-HSCT patients without reactivation or GVHD by lowering EBV burden.
EBV-transformed lymphoblastoid cell lines (LCLs) are used to selectively grow EBV-specific T lymphocytes as antigen-presenting cells.
DC and LCLs were transduced with adenovirus vectors to overexpress LMP1 or LMP2 to increase specificity for less immunogenic EBV antigens, but it is difficult and wastes time.
What are the indications for adoptive immunotherapy?
-EBSTs and multi-VSTs provided by third parties in PTLD
-recipient with immunosuppression and latent EBV infection
-Treatment for prevention and a decrease in the patient’s EBV load
-Treatment resistant and refractory cases of PTLD
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
T-cell adoptive immunotherapy for BK nephropathy in renal transplantation
The treatment of lymphoproliferative diseases connected to viral (EBV) loads
What are the side effects of adoptive immunotherapy?
Briefly summarize the principle of adoptive immunotherapy in the treatment of PTLD:
Adoptive immunotherapy is a principle of treatment used to treat post-transplant lymphoproliferative disorders which developed in transplant recipients due to uncontrolled proliferation of lymphocytes.
.Adoptive T-cell immunotherapy with donor derived EBV-specific T cells focuses on administering EBV-specific T cells, which are specific to immunodominant latency-associated antigens expressed in PTLD, such as Epstein-Barr nuclear Antigens EBNA1,2,3 And membrane proteins LMP1,2A,2B . What are the indications of adoptive immunotherapy?
1) Some cancers like: Leukemia, lymphoma, melanoma, lung cancer, and kidney cancer.
2) Viral infection in immunocompromised patients like: Post-transplant lymphoproliferative disorders, CMV infection, and HIV.
3) Immunotherapy-resistant or relapsed diseases Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Yes we can use it in resistant viral infections including BKV, CMV, Adenovirus, and HHV 6 What are the side effects of adoptive immunotherapy?
A. Risk of graft vs host disease.
B. Cytokine release syndrome.
C. Neurotoxicity. Level of evidence is 5
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
Adoptive immunotherapy is a treatment approach used to treat post-transplant lymphoproliferative disorders (PTLD), a group of diseases that can develop in transplant recipients due to uncontrolled proliferation of lymphocytes. The principle behind adoptive immunotherapy is the isolation, expansion, and reinfusion of immune cells, such as T-cells, that are specifically designed or selected to target and eliminate cancerous or infected cells in the patient.
In the context of PTLD, adoptive immunotherapy often involves the use of donor-derived or modified T-cells that recognize and target Epstein-Barr virus (EBV)-infected B-cells, since EBV infection is a common cause of PTLD. This approach helps boost the patient’s immune system to recognize and eliminate malignant or infected cells, leading to better disease control and potentially improved outcomes.
What are the indications of adoptive immunotherapy?
Adoptive immunotherapy has been explored as a treatment option for various medical conditions, particularly in the context of cancer and viral infections. The main indications for adoptive immunotherapy include:
Cancer: Adoptive immunotherapy has been used in the treatment of various cancers, including but not limited to:
Hematological malignancies, such as leukemia and lymphoma
Solid tumors, such as melanoma, lung cancer, and kidney cancer
Viral infections: Adoptive immunotherapy can be used to treat certain viral infections, particularly in immunocompromised patients, by boosting the immune system to fight the infection more effectively. Examples include:
Post-transplant lymphoproliferative disorders (PTLD) associated with Epstein-Barr virus (EBV) infection
Cytomegalovirus (CMV) infection in transplant recipients
Human immunodeficiency virus (HIV) infection
Immunotherapy-resistant or relapsed diseases: In cases where conventional therapies have failed, adoptive immunotherapy may be considered to improve disease control and outcomes.
It is important to note that the specific indications for adoptive immunotherapy may vary depending on the type of immune cells being utilized (e.g., T-cells, natural killer cells), the targeted antigen or receptor, and the patient population. The use of adoptive immunotherapy may be limited by factors such as availability, cost, and potential side effects. Always consult with a healthcare professional to determine the most appropriate treatment options for a specific medical condition.
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Yes, adoptive immunotherapy has potential applications in the treatment of various conditions related to organ transplantation. While its primary focus has been on addressing post-transplant lymphoproliferative disorders (PTLD) and viral infections in transplant recipients, researchers are exploring its potential use in other transplantation-related conditions:
Prevention and treatment of graft-versus-host disease (GVHD): GVHD is a complication that can occur after allogeneic hematopoietic stem cell transplantation (HSCT) when donor immune cells recognize the recipient’s tissues as foreign and attack them. Adoptive immunotherapy with regulatory T-cells (Tregs) or other immune cells with immunosuppressive properties is being investigated to prevent or treat GVHD.
Tolerance induction: Adoptive immunotherapy can be explored to induce tolerance to the transplanted organ, reducing the need for long-term immunosuppressive medications and lowering the risk of chronic rejection. This approach often involves the use of Tregs or other immune cells that promote tolerance.
Treatment of chronic rejection: Chronic rejection is a long-term complication of organ transplantation that can lead to graft dysfunction or failure. Adoptive immunotherapy using immune cells that can modulate the immune response or specifically target the pathological processes driving chronic rejection might be a potential therapeutic option.
Infections in immunocompromised transplant recipients: Adoptive immunotherapy can be used to treat or prevent viral infections, such as cytomegalovirus (CMV) or BK virus, in transplant recipients who are at high risk due to their immunosuppressed state.
These applications are mostly in experimental stages and require further clinical research to determine their safety, efficacy, and optimal usage. However, they highlight the potential of adoptive immunotherapy in improving transplant outcomes and addressing various challenges in the field of organ transplantation.
Summary of Article
EBV positive PTLD is a major complication of solid organ transplantation and Hematopoietic stem cell transplant.Although use of Rituximab ,Chemotherapy and reduction of immunosuppression have increased remission of PTLD,and increased survival of patient ,poor response to existing therapy in many cases needs new therapy.Adoptive immunotherapy is one such therapy.Adoptive T-cell immunotherapy with donor derived EBV-specific T cells focuses on administering EBV-specific T cells, which are specific to immunodominant latency-associated antigens expressed in PTLD, such as Epstein-Barr nuclear Antigens EBNA1,2,3 And membrane proteins LMP1,2A,2B .In Primary EBV infection ,most viral antigens are expressed during lytic stage while virus is replicating .In EBV infected B cells ,there is subsequent downregulation of viral antigen expression from latency type 3 to latency type 0,which allows escape from immune surveillance .It is the latency period which lead to Malignancy such as PTLD,Hodgkin and non hodgkin lymphoma.
Manufacturing of EBV specific T cell manufacturing(EBVSTs)-Although there are number of methods for manufacturing EBVSTs ,most common utilised is Ex vivo Expansion of EBVSTs.In this method mononuclear cells are harvested from the peripheral blood of the donor.After that B lymphocytes are infected with lab strain of EBV .Then they transduced with adenoviral vector expressing LMP1 and LMP2 and then irradiated.Monocytes are separately transduced with same adenoviral vector ,cultured with T Cells followed by a second stimulation by transduced Lymphoblastoid cells lines (LCL) to create Latent membrane protein specific T cell product.
Donor lymphocyte infusion( DLI) in an attempt to kill dividing B cells in EBV-associated PTLD, is a effective method to induce remission. Although both DLI and EBV specific T cytotoxic lymphocyte are almost equally effective in inducing remission,DLI is associated with Graft versus host disease,so not preferred .
Indications of Adoptive immunotherapy-
(A)PTLD–Due to non-availability of EBV specific cytotoxic lymphocytes at many centers and association with Graft versus host disease ,It is reserved for PTLD who failed to resolve with other therapy.
(B) Other disease -Treatment of difficult to treat viral infection, Progressive Multiple sclerosis .,Immune related Brain tumors
Other conditions related to organ transplantation
(1) Treatment of Drug resistant CMV infection
(2) Treatment of BK Virus nephropathy
(3)Prevention and Treatment of Rejection
(4) Development of immune tolerance via regulatory T cells (Tregs)
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
Post-transplant PTLD is strongly associated with latent EBV infection being reactivated due to immunosuppressive therapy leading to suppression of T cell activity and suppression of EBV infected B cells. In this regard the use of EBV specific T cells transference (allogenic or autologous) can be used to keep the EBV infected B cell population within control and reduce risk of PTLD development. What are the indications of adoptive immunotherapy?
· EBV associated PTLD (treatment and prophylaxis)
· Immunoblastic Lymphoma
· EBV associated Hodgkin lymphoma
· EBV associated NK/T Lymphoma
· EBV associated nasopharyngeal carcinoma Can adoptive immunotherapy be used in the treatment of other conditions related to organtransplantation?
Refractory Viral infections including BKV, CMV, Adenovirus, and HHV 6. What are the side effects of adoptive immunotherapy?
· Risk of graft vs host disease (acute and chronic)
· Infusion associated reactions including cytokine storm Level of Evidence:
Level 5 as it is a review article based on multiple non RCTs as well as authors’ personal experience and data.
1. Briefly summarize the article
Principle of adoptive immunotherapy in the treatment of PTLD
Review aricle about adoptive cellular immunotherapy for Epstein-Barr virus-associated lymphoproliferative disease (EBV-LD) .
Introduction:
When infection occur , EBV remain latent in B cells, oral epithelial cells and lymphoid follicles by downregulating the immunogenic proteins and get saved from immune surveillance [invisible to EBV-specific T-lymphocytes (EBVSTs)].
Competent T cell immunity control the infected B cell amount at <2% ; but in case of reduced immunity, with absence of EBVST response and uncontrolled proliferation of EBV infected B cells (mainly type3 latency) results in EBV-associated LPD and malignancies.
Manufacturing of EBVSTs:
1- Ex vivo expansion: of T cells targeting viral antigens via native T cell receptors where EBVSTs were selectively expanded utilizing irradiated EBV transformed lymphoblastoid cell lines (LCL) as antigen presenting cells (APCs) to selectively expand EBVSTs
2- Antigen specific T cell selection :
EBVSTs can be easily produced from seropositive blood using good-manufacturing-practice (GMP)-compliant protocols and cryopreserved for future use.
Adoptive cell therapy with Virus-specific T cells (VSTs) – reports impressive results in immunocompromised candidate with EBV-LD studies .
donor lymphocyte infusions (DLI) and donor derived multi-antigen specific VSTs in the HSCT – produce ORR of about 55-67%
Post-HSCT donor-derived T cell therapy:
1- Donor lymphocyte infusions
2- Donor-derived EBVSTs
In a study from (USA) : safety and efficacy of donor-derived EBVSTs for treatment and prophylaxis of PTLD post transplantation has been reported .
In another study :
– 114 patients received donor derived EBVSTs after allogeneic HSCT
– as a prophylaxis of 101 pt – No PTLD
– as a Treatment of 13 PTLD pt — 11 patients underwent complete remission.
(MSKCC) group result shows – (ORR) were 72% with DLI and 68% with EBVSTs
Strategies to enhance EBVST potency in vivo through genetic engineering as well as combination therapies:
Modification of EBVSTs to enhance activity
1. Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo – dominant negative TGF-β receptor type II (DNRII)
2. Calcineurin resistance to enhance EBVSTs – EBVSTs were genetically engineered to express a mutant form of calcineurin thus rendering them calcineurin inhibitor (CNI) resistant
3. Chimeric antigen receptor T cells — CD19-chimeric antigen T cells (CD19-CART) have shown impressive efficacy in acute B-lymphoblastic leukemia and in B-cell lymphomas
4. Combination of EBVSTs + modalities to enhance EBVST activity in vivo:
Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance
EBVST activity in vivo–
– Demethylating agents
– Checkpoint inhibitors
– BCl2 inhibitors
Conclusions:
Adoptive immunotherapy, particularly third-party “off the shelf” EBVSTs for treatment of EBV-LD is a promising approach.
But it is not widely available at time being.
Further work and Strategies needed to improve the anti-tumor activity of EBVST therapies for the less immunogenic type I and II latency tumors .
2. What are the indications of adoptive immunotherapy?
EBVSTs have been used as an adjunctive therapy to chemotherapy in
· Hodgkin lymphoma
· NK/T Cell lymphoma
· Nasopharyngeal carcinoma
· Burkitt lymphoma side effects :
1- Cytokine release syndrome (CRS)
2- Neurotoxicity
3- Acute kidney injury
4- Risk of graft rejection and autoimmunity.
3. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Treatment and prophylaxis to CMV, EBV, and Adenovirus.
Treatment of malignancies – Melanoma, Leukemias, Adenocarcinoma of Prostate
Referactory cases of EBV positive PTLD.
Level of evidence : 5
Adoptive immunmotherapy has good result in EBV related PTLD. The rationale behind using adoptive transfer of EBVSTs is based on harnessing immunogenicity of type III latency malignancies to control proliferation of latently infected B cells.
EBV associated malignancies with type II latency can develop in immunocompetent and immune deficient individuals and are much less immunogenic due to down regulation of the immunodominant antigens.
Indications of adoptive immunotherapy
EBV
CMV
Recurrent infection
Recurrent lymphoma
Recurrent prostate cancer
MS
Lupus
Adoptive therapy in treatment of conditions assoc. with transplant
Briefly summarize the principle of adoptive immunotherapy in the treatment of PTLD:
· Adoptive immunotherapeutic approaches to re-establish viral-specific immunity are an appealing alternative since viral problems in these patients are unmistakably linked to the absence of recovery of virus-specific cellular immune responses.
· The CD8+ cytotoxic T lymphocytes (CTLs), which identify peptides produced from viral proteins complexed to major histocompatibility complex (MHC) class I molecules, are the most crucial part of the cellular immune response that regulates the majority of viral infections.
· Furthermore, CD4 virus-specific T cells are crucial for supporting CD8+ cells and generating long-lasting protection.
· Researchers have looked into whether adoptive transfer of virus-specific CTLs can protect patients from such viruses because unmanipulated T-cell populations also contain alloreactive T cells.
What are the indications of adoptive immunotherapy:
· Treatment and prevention of Viral infections like CMV, EBV, and Adenovirus.
· Number of malignancies ranging from melanoma to certain types of leukaemia, as well as prostate cancer
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation:
· Recurrent leukemic after allogeneic bone marrow transplantation. · Treatment against CMV infection in SOT recipients. · Treatment of PTLD caused by Epstein–Barr virus.
What are the side effects of adoptive immunotherapy:
· General Fatigue · Fever · elevation of hepatobiliary enzymes, · cytokine release syndrome (also known as cytokine storm), · neurotoxicity · Graft vs Host Disease
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
EBV positive PTLD is a major complication of hematopoietic stem cell & solid organ transplantation. Two types of adoptive immunotherapy i) Donor lymphocyte infusion & ii) EBV specific T- lymphocytes (EBVSTs)
Individuals with weakened immune system, such as recipients of HSCT or SOT (solid organ transplant), the lack of a robust EBVST response can lead to uncontrolled proliferation of type 3 latency EBV – infected B cells resulting in EBV associated LPD & malignancies. The transformed B-cells in EBV- LD associated withlatency type III present several antigenic viral protein that induce potent EBVDT responses. Such potent T cell immunity maintains the infected B cell pool at <2% of total B cells in immunocompetent individuals but is lacking in immunosuppressef individuals leading to uncontrolled lymphoproliferation of the infected B cells.
The adoptive transfer of EBVSTs is based on harnessing the immunogenecity of type III latency malignancies to control the proliferation of the latency infected B cells.
What are the indications of adoptive immunotherapy?
– EBV associated PTLD in HSCT & SOT recipients.
-HIV lymphoma
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Prevention of EBV associated PTLD in high risk transplant recipients
What are the side effects of adoptive immunotherapy?
Acute & chronic graft versus host disease ( GVHD), which is more common with donor lymphocyte infusion.
1. Briefly summarize the article Principle of adoptive immunotherapy in the treatment of PTLD This is a review of published literature on adoptive cellular immunotherapy for Epstein-Barr virus-associated lymphoproliferative disease (EBV-LD) in last 20 years. Introduction: After initial infection (lytic phase), EBV maintains a lifelong latency in B cells, oral epithelial cells and lymphoid follicles by downregulating the immunogenic proteins and get rescue from immune surveillance [invisible to EBV-specific T-lymphocytes (EBVSTs)]. potent T cell immunity maintains the infected B cell pool at <2%; but in patients with weak immunity, lack of robust EBVST response and uncontrolled proliferation of EBV infected B cells (mainly type3 latency) results in EBV-associated LPD and malignancies. Rationale for adoptive transfer of EBVSTs is based on harnessing the immunogenicity of type III latency malignancies to control the proliferation of the infected B cells. Objectives:
– Define current best manufacturing strategies for EBVST
– Summarize the clinical experience on their use in EBV-related LPD,
– Opportunities to broaden the applicability of this approach
– Explore future strategies to enhance their efficacy
Manufacturing of EBVSTs: Ex vivo expansion Antigen specific T cell selection EBVSTs can be readily produced from seropositive donors using good-manufacturing-practice (GMP)-compliant protocols and cryopreserved for future use.
Adoptive cell therapy with Virus-specific T cells (VSTs) have been used for more than two decades – has shown impressive results in immunocompromised patients with EBV-LD in clinical trials.
· donor lymphocyte infusions (DLI) and donor derived multi-antigen specific VSTs in the HSCT – ORR 55-67%
· autologous & allogeneic VSTs in the SOT – EBVST infusion did not consistently decrease EBV viremia, but no patient after prophylaxis developed PTLD EBVST from LCL used for Latency type2 malignancies (relapsed EBV-HL) – OR 60-70%
Post-HSCT donor-derived T cell therapy:
· Donor lymphocyte infusions
· Donor-derived EBVSTs St. Jude’s (USA) preliminary study: safety and efficacy of donor-derived EBVSTs for treatment and prophylaxis of PTLD in transplant recipients has been seen in many studies. Large multicenter study:
– 114 patients received donor derived EBVSTs after allogeneic HSCT
– prophylaxis in 101 patients – No PTLD
– Treatment in 13 patients with PTLD — 11 patients achieved complete remission; persisted 9 years (MSKCC) group – (ORR) were 72% with DLI and 68% with EBVSTs
Third Party T cells readily available “off the shelf” T cell therapy product is desirable. UK- 60 products bank 33 Tx recipients – ORR 64 % in HSCT; 52% in SOT Birmingham, United Kingdom published their experience in 10 pediatric SOT recipients with PTLD and reported an ORR of 80% (8 out of 10)
Strategies to enhance EBVST potency in vivo through genetic engineering as well as combination therapies: Modification of EBVSTs to enhance activity
1. Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo – dominant negative TGF-β receptor type II (DNRII)
2. Calcineurin resistance to enhance EBVSTs – EBVSTs were genetically engineered to express a mutant form of calcineurin thus rendering them calcineurin inhibitor (CNI) resistant
3. Chimeric antigen receptor T cells — CD19-chimeric antigen T cells (CD19-CART) have shown impressive efficacy in acute B-lymphoblastic leukemia and in B-cell lymphomas
4. Combination of EBVSTs + modalities to enhance EBVST activity in vivo:
Conclusions: Adoptive immunotherapy, particularly third-party “off the shelf” EBVSTs for treatment of EBV-LD has shown promise in several studies conducted at specialized centers, also is being available for multi-center studies, consortium and cooperative group studies. Further work is needed to create widely available and commercialized third-party cell banks to broaden the applicability of this approach beyond specialized centers. Strategies to enhance the anti-tumor activity of EBVST therapies for the less immunogenic type I and II latency tumors, need to be explored.
2. What are the indications of adoptive immunotherapy? PTLD in SOT – post Retuximab + Chemotherapy – refractory / relapse – post Retuximab monotherapy relapse patients not tolerating Chemotherapy Post-HSCT: prophylaxis for PTLD and viral associated malignancies in children, post Stem Cell Transplant Malignancies secondary to EBV. Resistant immunological diseases. 3. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation? Treatment and prevention of Viral illness due to CMV, EBV, and Adenovirus. Malignancies – Melanoma, Leukemias, Adenocarcinoma Prostate EBV positive PTLD – refractory and relapse EBV associated Hodgkin’s lymphoma, NK cell Lymphoma, nasopharyngeal carcinoma Autoimmune diseases like lupus or Multiple Sclerosis 4. What are the side effects of adoptive immunotherapy?
GVHD
Cytokine release syndrome
Neurotoxicity.
Discussion question Level of evidence – none levelled
Adoptive cellular immunotherapy for Epstein-Barr virus-associated lymphoproliferative disease. Introduction.
Epstein-Barr virus (EBV) is associated with a variety of lymphomas and lymphoproliferative disorders (LPD). It infects more than 90% of the adult population worldwide, body produces CD8+ cytotoxic T lymphocytes (CTL) specific to the early lytic cycle proteins kill cells entering the lytic cycle before they are able to release infectious virus particles, EBV infected B lymphocytes per 1,000,000 peripheral blood mononuclear cells (PBMCs) over their lifetime, controlled at these levels by a potent EBVST response, in immunocompromised situation such as post organ transplantation, the lack of a robust EBVST response can lead to uncontrolled proliferation of type 3 latency EBV-infected B cells resulting in EBV-associated LPD and malignancies.
The role for adoptive transfer of EBVSTs is to keep the immunogenicity of type III latency malignancies to control the proliferation of the latently infected B cells, the biggest risk factor for developing EBV-associated PTLD post-SOT is EBV seronegativity at the time of transplant.
Rituximab, targeting CD20 present on the B cells has been an effective monotherapy with response rates of 55% to 100% in HSCT recipient and so response rates to rituximab monotherapy in SOT recipients are generally lower and around 50%, and other treatment option is immunotherapy with EBVSTs. Adoptively transferred virus-specific T cells (VSTs) have been evaluated for more than two decades. Manufacturing of EBVSTs.
1-ex vivo expansion of VSTs versus antigen-specific.
2-T cell selection [e.g., interferon γ (IFN-γ) capture]. Post-HSCT donor-derived T cell therapy. Donor lymphocyte infusions.
Utilized unmodified DLI derived from the patient’s EBV seropositive HSCT donor which contained effector cells with activity against EBV. Although effective in inducing remissions, this therapy carries a significant risk of graft-versus-host disease (GVHD). Donor-derived EBVSTs.
Patients who received the EBVSTs as prophylaxis had any EBV reactivation or GVHD and there was evidence of persistence of EBVSTs by tracking of genetic markers on the T cells for a median of 10 weeks. Autologous EBVSTs.
When the donor is not available such as cadaveric organ transplantation, autologous EBVSTs have also been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy. Third-party T cells.
Used with refractory PTLD between the age of 1–76 years received partially HLA matched EBVSTs. HLA matches ranged from 2 to 5/6 HLA alleles and there was a statistically significant association of better outcome with higher HLA matches. Overall, the response rate (CR and PR) was 64% at 5 weeks and 52% at 6 months. Modification of EBVSTs to enhance activity =Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo.
The DNRII-LSTs were resistant to otherwise inhibitory concentrations of TGF-β. Calcineurin resistance to enhance EBVSTs.
EBVSTs were genetically engineered to express a mutant form of calcineurin thus rendering them calcineurin inhibitor (CNI) resistant. Chimeric antigen receptor T cells.
CD19-chimeric antigen T cells (CD19-CART) have shown impressive efficacy in acute B-lymphoblastic leukemia and in B-cell lymphomas. Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance EBVST activity in vivo. –Demethylation agents.
-Checkpoint inhibitors.
-BCL-2 inhibitors. Conclusions.
The use of adoptive immunotherapy, EBVSTs for the treatment of EBV-LD has shown promise in several studies conducted
at specialized centers, and need more studies for combination therapy to increase their efficacy. Indications:
·Post-transplant lymphoproliferative diseases.
·Malignancies secondary to EBV.
·Resistant immunological diseases. Other Indications: ·EBV associated malignancies e.g., Hodgkin’s lymphoma, natural killer cell lymphoma, nasopharyngeal carcinoma
·Viral associated infections -CMV,EBV
· Autoimmune diseases like lupus or MS Side effects:
·Cytokine release syndrome
·GVHD
· Neurotoxicity.
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
In individuals with weakened immune systems, such as patients with solid organ transplant (SOT), the lack of a robust EBVST response can lead to uncontrolled proliferation of type 3 latency EBV-infected B cells resulting in EBV-associated LPD and malignancies. Adoptive immunotherapy involves isolating EBV specific T-lymphocytes (EBVSTs) from a patient with EBV-seropositive PTLD patients (these cells can be isolated from a resected tumor tissue or peripheral blood then they undergo ex vivo (in the laboratory) activation and expansion using good-manufacturing-practice (GMP)-compliant protocols)
Autologous EBVSTs have also been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients or the third-party EBVSTs can be used After that infused back into the patient in large numbers where they can now help the patient’s immune system to destroy the cancer cells. Adoptive immunotherapy is reserved for EBV-positive patients who fail to respond to initial therapy.
What are the indications of adoptive immunotherapy? – Management of resistant PTLD
– EBV-associated malignacy
– Refractory/ resistant disease/cancer
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation? – resistant EBV specific tumors – resistant Skin Melanoma – resistant Autoimmune disease – resistant BK nephropathy
What are the side effects of adoptive immunotherapy? – GVHD – Cytokine storm (cytokine release syndrome) – Neurotoxicity – Graft rejection
Disscussion question Level of evidence – none leveled.
FIRST B ELLS GET INFECTED WITH EBV, EBVinfected B-lymphocytes then enter the lymphoid follicles
and downregulate the immunogenic proteins
CD8 PLUS T CELLS ( VIRUS SPECIFIC T CELLS, VST ) ARE INVOLVED IN DOWNREGULATIMH EBV INFECTED B CELLS WHICH CAN FORM EBV RELATED TUMORS
WHEN SUCH T CELLS ARE LESS IN NUMBER , THERE IS HIGH CHANCE OF EBV RELALATED LYMPHOPROILIFERATIVE DISEASE
TYPE 1 AND 2 LATENCY TUMOR ARE LESS IMMUNOGENIC WHILE TYPE 3 IS MORE IMMUNOGENIC AND IS THE TARGET OF ADAPTIVE IMMUNOTHERAPY AS IT INDICES STRONG EBV RELATED T CELL RESPONSE
INDICATION-
FOR PTLD IN POST TRANSPLANT SITUATION WHEN RITUXIMAB AND OR CHEMO IS ADVISABLE BUT IT IS MORE TOXIC
THIS IS NEW AND EVOLVING THERAPY FOR PTLD AND FOUND TO BE USEFUL IN PEDIATRCI PATIENTS AND PTLD AFTER STEM CELL TRANSPLANT
FOR SOLID ORGAN TRANSPLANT , TRAILS ARE GOING ON WITH THIRD PARTY T CELL ADAPTIVE IMMUNOTHERAPY
MINIMAL SIDE EFFECT CAN BE THEIR ADVANTAGE
SOURCE OF T CELL-
AUTOLOGOUS T CELL
OWN T CELLS AUGMENTATED IN LAB
DONOR SPECIFIC , HLA MATCHED T CELLS
OFF THE SHELF , THIRD PARTY T CELLS- EASILY AVAILABE ON TIME , COMMON IN RESEARCH TRIALS
OTEHR USES-
TO IMPROVE THE IMMUNITY OF TRANSPLANT PATIENT
TO COMBAT OTHR FORM OF TUMOR WHICG ARE STILL COMMON AFTER TRANSPLANT
Briefly summaries the principle of adoptive immunotherapy in the treatment of PTLD
Epstein-Barr virus (EBV) is a member of the gamma herpes virus family that is associated with a variety of lymphomas and lymphoproliferative disorders (LPD).
It infects more than 90% of the adult population worldwide.
have different clinical presentations include asymptomatic infection to symptomatic disease.
normally in healthy person, virus neutralizing antibodies control the spread of infection by EBV-specific CD8 + cytotoxic T lymphocytes (CTL) specific to the early lytic cycle proteins that kill cells entering the lytic cycle before they are able to release infectious virus particles.
What are the indications of adoptive immunotherapy?
PTLD associated with and without EBV. malignancy other than PTLD that associate with EBV.
treatment of PTLD.
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Treatment against CMV infection in SOT recipients. Treatment of PTLD caused by Epstein–Barr virus.
recurrent leukemia.
What are the side effects of adoptive immunotherapy?
Briefly summarize the principle of adoptive immunotherapy in the treatment of PTLD
· Immunotherapeutic approaches to reestablish viral-specific immunity are an appealing alternative since viral problems in these patients are unmistakably linked to the absence of recovery of virus-specific cellular immune responses.
· The CD8+ cytotoxic T lymphocytes (CTLs), which identify peptides produced from viral proteins complexed to major histocompatibility complex (MHC) class I molecules, are the most crucial part of the cellular immune response that regulates the majority of viral infections.
· Furthermore, CD4 virus-specific T cells are crucial for supporting CD8+ cells and generating long-lasting protection.
· Researchers have looked into whether adoptive transfer of virus-specific CTLs can protect patients from such viruses because unmanipulated T-cell populations also contain alloreactive T cells.
===================================== What are the indications of adoptive immunotherapy?
· Treatment and prevention of Viral infections like CMV, EBV, and Adenovirus.
· Number of malignancies ranging from melanoma to certain types of leukemia, as well as prostate cancer
====================================== Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
· Recurrent leukemia after allogeneic bone marrow transplantation.
· Treatment against CMV infection in SOT recipients.
· Treatment of PTLD caused by Epstein–Barr virus.
======================================= What are the side effects of adoptive immunotherapy?
· Fever,
· general fatigue.
· elevation of hepatobiliary enzymes,
· cytokine release syndrome (also known as cytokine storm),
· neurotoxicity
· GVHD
· Epstein Bar Virus causes widely spread virus associated with lymphoprolifrative disease in SOT and hematopoietic stem cell transplant.
· This virus infects more than 90% of adult population and tends to have different clinical presentations include asymptomatic infection to full blown disease.
· First successes with adoptive immunotherapy were described using donor lymphocyte infusion for the treatment of EBV+ PTLD in hematopoietic stem cell transplantation (HSCT) recipients · Adoptive immunotherapy basically involves isolating EBV specific T-lymphocytes (EBVSTs) from a patient with PTLD i.e., EBV-seropositive patients. · These cells can be isolated from a resected tumor tissue or peripheral blood then they undergo ex vivo (in the laboratory) activation and expansion using good-manufacturing-practice (GMP)-compliant protocols. · After that infused back into the patient in large numbers where they can now help the patient’s immune system to destroy the cancer cells. · Adoptive immunotherapy is reserved for EBV-positive patients who fail to respond to initial therapy. · Adoptive immunotherapy using EBVSTs has shown good results in EBV related Lymphoprolifrative disorders.
Indications: · Treatment of post transplant lymphoproliferative diseases. · Malignancies secondary to EBV. · Resistant diseases.
Other Indications: · EBV associated malignancies e.g., Hodgkin’s lymphoma, natural killer cell lymphoma, nasopharyngeal carcinoma · Skin cancer-Melanoma. · Viral associated infections -CMV,EBV · Autoimmune diseases like lupus or MS
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
EBV-positive post-transplant PTLD is one of the major complication of solid organ transplantation.
objectives of this review are:
(I) manufacturing strategies of EBV specific T-lymphocytes (EBVSTs);
(II) to review different EBVST donor options
(III) to summarize current experience with adoptive cellular therapy
(IV) to explore the potential for combination therapies with other immunotherapeutic strategies.
Epstein-Barr virus (EBV) is a member of the gamma herpes virus family. . It infects more than 90% of the adult population worldwide. normally
In healthy individuals, virus neutralizing antibodies control the spread of infection by EBV-specific CD8 + cytotoxic T lymphocytes (CTL) specific to the early lytic cycle proteins that kill cells entering the lytic cycle before they are able to release infectious virus particles.
EBV-infected naïve B-lymphocytes express proteins comprising the entire EBV genome including the EBV nuclear antigens EBNA1, EBNA2, EBNA3, and EBNALP, membrane proteins LMP1, and LMP2 as well as BARF1 and two small non-translated ribonucleic acids (RNA) (Type III latency) .
EBV infected B-lymphocytes then enter the lymphoid follicles and downregulate the immunogenic proteins to express less immunogenic type II latency proteins (EBNA1, LMP1 and LMP2) and thus rescue them into the memory compartment where the virus persists in latently infected B-lymphocytes by further downregulating expression of viral proteins so as to become invisible to EBV-specific T-lymphocytes (EBVSTs).
EBV-associated malignancies are associated with type II latency [e.g., Hodgkin’s lymphoma (HL), natural killer (NK)/T-cell lymphoma, nasopharyngeal carcinoma (NPC)] or type I latency tumors (e.g., Burkitt lymphoma or gastric carcinoma).
EBV-associated post-transplant lymphoproliferative disease (PTLD) occurs between 1% -25% of HSCT recipients depending on the serostatus of the donor and patient. and the level of post-HSCT immunosuppression
biggest risk factor for PTLD post-SOT is EBV seronegativity at the time of transplant. Since most children are transplanted at a young age while still being EBV seronegative and convert to EBV seropositivity within 2 years of transplant, EBV driven PTLD is much more common in pediatric SOT recipients because of the fact that at a young age while still being EBV seronegative and convert to EBV seropositivity within 2 years of transplant,
More than 90% of EBV-associated PTLD is of B-cell origin which are CD20+ .Rituximab are effective monotherapy with response rates of 55% to 100% in HSCT recipients Third-party EBVSTs have been mostly used in the posttransplant setting. But not all patients treated with third-party EBVST’s had detectable viral loads at start of therapy .
Moreover, clinical response and reduction of EBV viremia correlated with an increase of EBVSTs .
therefore it was conculded that the use of adoptive immunotherapy, particularly using third-party EBVSTs for the treatment of EBV-LD has shown promise in several studies Further work is however needed to create widely available and commercialized third-party cell banks.
What are the indications of adoptive immunotherapy?
treatment of treatment of difficult to treat viral infections
treatment of EBV viral specific cancers i,e. leukemia lymphomas
lymphoma treatment in immunocompromised patients.
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
utilized for type II latency EBV associated malignancies including HL, T cell lymphoma, NPC as adjunctive therapy.
treatment of autoimmune diseases and has been used in BK virus nephropathy, melanoma and EBV specific tumors.
What are the side effects of adoptive immunotherapy?
1. Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD. Introduction; Epstein Barr virus is a gamma virus family almost infects >90% of population worldwide. Usually asymptomatic and involves at childhood. But can present with mild and florid disease like IMS. Despite of initial infection the virus maintains a lifelong latency in B cell and oral epithelial cells. The major receptor for EBV which is present on B-cell and remain latent. The infected B- cells then express protein comprises of EBNA1, 2, 3 and EBNALP, membrane protein LMP1, 2 as well as well BARF1. They survive in cells by down regulating the immune system. When there is immunodeficiency and lack of EBVCT response in an individual then there is uncontrolled proliferation of virus infected b-cells and resulted EBV associated malignancies. .. The adaptive immunotherapy is based on harnessing the immunogenicity of the type III latency malignancies to control its latent B-cell proliferation. Adaptive immunotherapy involves isolating EBVST from a seropositive patient from peripheral blood then is being activated (ex vivo) using good manufacturing protocol, MHC multicenter selection and the most commonly used mathod interferon-y capture approach and these expended effector cells are re-infused, where they can boast immune system against the viruses, tumor cells. Adaptive immunity as a prophylactic and therapeutic the success rate is around 80- 100%. For this purpose usually the donors are autologous and allogeneic. 2. What are the indications of adoptive immunotherapy? It’s being used for type III, II, and type I related carcinomas and EBV- associated lymphproliferative diseases, Treatment autoimmune diseases, Treatment of refractory viral infections like CMV, Hodgkin lymphoma, Burkit lymphoma, Nasopharyngeal carcinoma, NK/T cell lymphoma. 3. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation? Autoimmune disease like lupus, MS, Chronic hypersensitivity reaction, Other inflammatory disorders, 4. What are the side effects of adoptive immunotherapy?
Cytokine release syndrome,
Acute kidney injury,
Thrombotic microangiopathy,
Hypersensitivity like reaction like fever, edema, rash,
Neurotoxicity.
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
EBV-positive post-transplant lymphoproliferative disease (PTLD) is a major complication of hematopoietic stem cell and solid organ transplantation.
objectives of this review are: (I) to describe different manufacturing strategies of EBV specific T-lymphocytes (EBVSTs); (II) to review different EBVST donor options and their advantages and limitations; (III) to summarize current clinical experience with adoptive cellular therapy for EBV-LD and finally; (IV) to explore the potential for combination therapies with other immunotherapeutic strategies.
Epstein-Barr virus (EBV) is a ubiquitous member of the gamma herpes virus family that is associated with a variety of lymphomas and lymphoproliferative disorders (LPD). It infects more than 90% of the adult population worldwide
In healthy seropositive individuals, virus neutralizing antibodies control the spread of infectious virus particles and EBV-specific, human leukocyte antigen (HLA) class I restricted, CD8 + cytotoxic T lymphocytes (CTL) specific to the early lytic cycle proteins kill cells entering the lytic cycle before they are able to release infectious virus particles.
EBV-infected naïve B-lymphocytes express proteins comprising the entire EBV genome including the EBV nuclear antigens EBNA1, EBNA2, EBNA3, and EBNALP, membrane proteins LMP1, and LMP2 as well as BARF1 and two small non-translated ribonucleic acids (RNA) (Type III latency) .
EBVinfected B-lymphocytes then enter the lymphoid follicles and downregulate the immunogenic proteins to express less immunogenic type II latency proteins (EBNA1, LMP1 and LMP2) and thus rescue them into the memory compartment where the virus persists in latently infected B-lymphocytes by further downregulating expression of viral proteins so as to become invisible to EBV-specific T-lymphocytes (EBVSTs).
EBV-associated malignancies associated with type II latency [e.g., Hodgkin’s lymphoma (HL), natural killer (NK)/T-cell lymphoma, nasopharyngeal carcinoma (NPC)] or type I latency tumors (e.g., Burkitt lymphoma or gastric carcinoma) can develop in immunocompetent and immune deficient individuals and are much less immunogenic due to downregulation of the immunodominant (e.g., EBNA3 and EBNA2) antigens.
EBV-associated post-transplant lymphoproliferative disease (PTLD) occurs in less than 1% to 25% of HSCT recipients depending on the serostatus of the donor and patient, the degree to which the graft is T-cell depleted, and the post-HSCT immunosuppression
biggest risk factor for developing EBV-associated PTLD post-SOT is EBV seronegativity at the time of transplant. Since most children are transplanted at a young age while still being EBV seronegative and convert to EBV seropositivity within 2 years of transplant, EBV driven PTLD is much more common in pediatric SOT recipients .
More than 90% of EBV-associated PTLD is of mature B-cell origin with cell surface expression of CD20 .
Rituximab, monoclonal antibody targeting CD20 present on the B cells has been an effective monotherapy with response rates of 55% to 100% in HSCT recipients
Third-party EBVSTs have been mostly used in the posttransplant setting. There has been no published experience in patients with HIV-associated lymphomas because in the modern era of highly active anti-retroviral therapy (HAART) the incidence in the Western word has decreased and the logistical support and specialized experience needed does not make EBVSTs an easily accessible option for the treatment of patients in the developing world.
not all patients treated with third-party EBVST’s had detectable viral loads at start of therapy .
Leen et al. also showed reduction of EBV viral load correlated with response in HSCT recipients with EBV-associated disease treated with third-party multi virus specific VSTs .
Moreover, clinical response and reduction of EBV viremia correlated with an increase of EBVSTs .
Conclusion
The use of adoptive immunotherapy, particularly using third-party “off the shelf” EBVSTs for the treatment of EBV-LD has shown promise in several studies conducted at specialized centers .
Further work is however needed to create widely available and commercialized third-party cell banks to broaden the applicability of this approach beyond boutique centers.
In addition, strategies need to be explored to enhance the anti-tumor activity of EBVST therapies especially for the less immunogenic type I and II latency tumors.
What are the indications of adoptive immunotherapy?
Prophylactic treatment of EBV load
Refractory EBV and CMV
PTLD associated with and without EBV
malignancy other than PTLD that associate withEBV
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Autoimmune diseases
Skin diseases
What are the side effects of adoptive immunotherapy?
GVHD
neurotoxicity
Acute rejection
Allergic reaction
PRINCIPLES OF ADOPTIVE IMMUNOTHERAPY IN TREATING POST TRANSPLANT PTLD.
EBV infection is common in the general population and mostly latent affecting more than 90% of the world population. It has is associated with lymphoproliferative diseases post transplant ;SOT and HSCT. Irrespective of initial mode of infection, maintains a lifelong latency in oral epithelial cells and will cause an infection once immunity gets compromised either by chronic infections like HIV, malignancies or immunosuppressive medications.
Enhanced immunogenicity of type 3 latency malignancies reduces proliferation of latent B cells thus reducing propensity of EBV to be active/malignant post transplant. EBV malignancies(Type 2 eg HL,NK Cell lymphoma and NPC) can in both healthy and immunocompromised population and a re not very immunogenic due to decreased expression of EBNA3 and EBNA2 antigens.
EBV associated LD ,type 3 present antigenic proteins that stimulate strong EBVST response, This strong resultant immunity helps keep infected B cell pool at a low nadir, less than 2% of total B cells in healthy people a factor not present in the immunosuppressed post transplant, Exploration of this aspect may lead to a decrease in proliferation of infected B cells and a decline in incidences of EBV associated malignancies post transplant.
EBV Production methods;
1.EX vivo expansion of EBVST.
2.Antigen specific T cells.
3.Autologous EBVST.
4.3rd Party T cells.
5.Allogenic lymphocytes.
EBVST enhancing Methods;
1.Surmount TGF Beta immunosuppressive effects.
2.Chimeric AG receptor T cell.
3.Calcineurin inhibitors to enhance EBVST.
INDICATIONS OF ADOPTIVE IMMUNOTHERAPY.
1.Treatment of post transplant lymphoproliferative diseases.
2,Malignancies secondary to EBV.
3.Resistant diseases.
OTHER INDICATIONS OF ADOPTIVE IMMUNOTHERAPY.
1.Skin cancer-Melanoma.
2.Viral associated infections -CMV,EBV
3.Autoimmune diseases like lupus or MS
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD????
Epstein- Bar Virus Is A Worldwide Widely Spread Gamma Herpes Virus Family That Has Been Associated With Lymphoproliferative Disease In Solid Organ Transplant And Hematopoietic Stem Cell Transplant,
Adoptive Immunotherapy Involves Isolation And Infusion Of T Lymphocytes In A Patient For Treatment/ Prevention Of A Disease
EBV Infects More Than 90% Of The Adult Population.
Primary Infection Mainly Occurs In Childhood Leading To Asymptomatic/Mild Infection/Infectious Mononucleosis.
EBV Enters The Oropharynx Replicating In The Epithelial Cells And Infecting Transiting B Lymphocytes.
The Infected B Lymphocytes Express Proteins Representing The Whole EBV Genome. They Then Enter The Lymphoid Follicles Where They Down Regulate The Immunogenic Proteins To Express Less Immunogenic Type 2 Latency Proteins.
They Thus Enter The Memory Compartment Where The Virus Persist In Latently Infected B Lymphocytes.
The Frequency Of EBV Infected B Lymphocytes Remain Stable In A Healthy Individual. However, In An Immunocompromised Individual, Lack Of A Robust EBV Specific T Lymphocytes Leads To Uncontrolled Proliferation Of Type 3 Latency EBV Leading To LPD And Malignancies.
In healthy individuals, the rate of EBV infection remains stable at 0.1-50 EBV-infected B lymphocytes per 1,000,000 peripheral blood mononuclear cell
EBV associated PTLD is a well described complication after both hematopoietic stem cell (HPSC) and solid organ transplantation (SOT) in about 1-25% of cases.
The risk of PTLD is dependent on the EBV serostatus of the donor (higher risk with positive donor and negative recipient) and the degree of immunosuppression (higher risk with ATG induction and TAC based maintenance therapy).
Risk of EBV-PTLD is higher in pediatric SOT (as they are transplanted early before acquiring infection) while > 90 % of adult donors are seropositive.
In Case Of Immunodeficient Individuals As Primary Immune Deficiency (PID), HIV Or SOT: The Absence Of EBV Specific T Lymphocytes (EBVSTS) Allows The Uncontrolled Expansion And Proliferation Of EBV Infected B Lymphocytes That Leads To EBV- Lymphoproliferative Diseases.
EBV Infection Is Then Associated With Many Diseases As PTLD, Hodgkin Lymphoma, Nk And T Cell Lymphoma, Burkitt’s Lymphoma, Nasopharyngeal And Gastric Carcinoma.
Most Of EBV-PTLD (90%) Are Of B Cell Origin. Rituximab Is An Effective Monotherapy In Case Of HSCT (50-100 % Of Cases) And Effective In 50 % Of Sot Cases. In Children, Combination Of Rituximab, Steroids And Cyclophosphamide Is Effective In Treatment But It Increases The Risk Of Infections, Recurrence Of Disease & Severe Toxicity
Hence, The Need For The Development Of Medication That Addresses The Underlying Immune Defect With Little To No Toxicity
EBV specific T lymphocytes (EBVSTs) inhibit the entry of EBV infected B cells into the lymphoid follicles. In transplant recipients, due to weakened immune system, EBVST response is lacking, leading to uncontrolled proliferation of EBV-infected B cells and resultant EBV-associated post-transplant lymphoproliferative disorders (PTLD) and malignancies. So, infusion of EBVSTs help in reducing the latently infected B cell pool.
Autologous or allogeneic (including third party) source of EBVSTs are used to manufacture them using either ex vivo expansion or antigen-specific T cell selection interferon gamma capture).
Manufacturing EBV-specific T-lymphocytes (EBVST) Is an adoptive transfer of virus-specific T cell use of donor lymphocyte infusions (DLI) to donor-derived multi-antigen specific VSTs in the HSCT setting
It is produced from EBV-positive donors using good manufacturing grade, and the donor type could be;( autologous, allogeneic)
Methods used in manufacturing EBVST products
–Ex vivo expansion EBVSTs
–Antigen-specific T cell
Post-HSCT donor-derived T-cell therapy
1- Donor lymphocytic infusion effective in remission but with attendant GVHD
2- Donor-derived EBVSTs 67% response rate but a very GVHD effect was seen among those treated with it
Autologous EBVSTs used in HSCT and SOT recipients where their donors are not available in theory, it is a preferred method used for PTLD Delay in the production and administration of the drug because its autologous Has been used in the treatment of type 11 latency EBV-associated malignancy
Third-party T cell
Designed to overcome the delay that may come with either autologous or allogenic methods of producing EBVSTs It is a readily available off-the-shelf T-cell therapy product Its response rate (CR and PR) was 64% at 5 weeks and 52% at 6 months with no significant toxicity observed In the pediatric age group, it has a more favourable outcome Many study groups have reported a significant reduction in EBV viral loads following treatment with third-party T cell
Modification of EBVSTs treatment to enhance the activity Overcoming the immune suppressive effects of TGF- beta to enhance EBVSTs activity in vivo Calcineurin resistance to enhance EBVSTs Chimeric antigen receptor T-cell
In Summary :
The adoptive immunotherapy in the treatment of PTLD in SOT or HSTC especially the third- party T- cell has demonstrated a better clinical outcome with less toxicity and alleviating the concern of GVHD
What are the indications of adoptive immunotherapy?
-immunocompromised patient with EBV-negative lymphoproliferative disease
-Those that failed to respond to Rituximab monotherapy or plus steroid and cyclophosphamide
-Treatment of viral infections that are difficult to cure like CMV, EBV.
–Immunocompromised patient with EBV-negative lymphoproliferative disease.
–Treatment of cancers that are caused by specific viruses, leukemias, DLBC lymphoma.
–Treatment of autoimmune diseases like lupus.
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
-Yes, in prevention of EBV related PTLD
–Treatment of cancers that are caused by specific viruses, leukemias, DLBC lymphoma.
–Treatment of autoimmune diseases like lupus.
What are the side effects of adoptive immunotherapy?
-Graft-versus-host disease
–Graft rejection
–IgE mediated anaphylactic reaction
–Neurotoxicity
–Cytokine release syndrome
*The article is a narrative review: Level of evidence – Level 5
I. Adoptive cellular immunotherapy for Epstein-Barr virus-associated lymphoproliferative disease
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
Adoptive immunotherapy basically involves isolating EBV specific T-lymphocytes (EBVSTs) from a patient with PTLD i.e., EBV-seropositive patients. These cells can be isolated from a resected tumor tissue or peripheral blood then they undergo ex vivo (in the laboratory) activation and expansion using good-manufacturing-practice (GMP)-compliant protocols. Thereafter these expanded effector cells are infused back into the patient in large numbers where they can now help the patient’s immune system fight/ destroy the cancer cells. Adoptive immunotherapy in PTLD is both therapeutic and prophylactic with 80% and 100% success respectively. The transferred EBVSTs can persist for up to nine years. Adoptive immunotherapy is reserved for EBV-positive patients who fail to respond to initial therapy.
What are the indications of adoptive immunotherapy? – Therapeutic and prophylactic management of PTLD
– EBV-associated malignancies
– Refractory/ resistant disease
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
– EBV associated malignancies e.g., Hodgkin’s lymphoma, natural killer cell lymphoma, nasopharyngeal carcinoma
– Skin cancer
– Autoimmune disease
– Viral infections e.g., BK virus, CMV
What are the side effects of adoptive immunotherapy?
– Acute and chronic graft-versus-host-disease (GVHD)
– Cytokine storm (cytokine release syndrome) due to an overactive immune response leading to excessive inflammation
– Neurotoxicity due to inflammation of the brain
– Others: – AKI, graft rejection, bleeding episodes, arrhythmias, delirium, dizziness, edema, altered bowel patterns, hypotension, hypoxia, tremors, tachycardia, infections
Introduction
Epstein Bar Virus is a world widely spread virus associated with lymphoprolifrative disease in SOT and hematopoietic stem cell transplant. It infects more than 90% of adult population. This virus tends to have
· Primary infection usually asymptomatic or infection mononucleosis like symptoms.
· Lifelong potency in B cells
· Progression to latency type I where EBV infected B cells would be invisible to EBSVTs.
· Uncontrolled type III latency
Donor lymphocyte infusion was first successful adoptive immunotherapy for EBV related PTLD though associated with risk of GVHD. Several studies has shown benefits of adoptive immunotherapy using EBVST “of the shelf”. Clinical Experience
Adoptive immunotherapy using EBVSTs has shown good results in EBV related Lymphoprolifrative disorders. EBVST Donor Options
· Seropositive donor with cryopreserved
· Autologous and allogeneic EBVST manufacturing
· Ex-vivo expansion of VSTs Vs antigen specific
· MHC multimer selection( to isolate CD8+ T cell)
· Interferon –y capture approach Combination therapy
Using genetic engineering technique to enhance EBVST. Indications
EBVSTs have been used as an adjunctive therapy to chemotherapy in
· Hodgkin lymphoma
· NK/T Cell lymphoma
· Nasopharyngeal carcinoma
· Burkitt lymphoma Adverse association
· Cytokine release syndrome
· AKI
· Rejection and autoimmunity
· Neurotoxicity Level of evidence V
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
The scientific rationale for adoptive transfer of EBVSTs is based on harnessing the immunogenicity of type III latency malignancies to control the proliferation of the latently infected B cells. EBV-associated malignancies associated with type II latency [e.g., Hodgkin’s lymphoma (HL), natural killer (NK)/T-cell lymphoma, nasopharyngeal carcinoma (NPC)] or type I latency tumors (e.g., Burkitt lymphoma or gastric carcinoma) can develop in immunocompetent and immune deficient individuals and are much less immunogenic due to downregulation of the immunodominant (e.g., EBNA3 and EBNA2) antigens.
-The transformed B-cells in EBV-associated lymphoproliferative disease (EBV-LD)
associated with latency type III present several antigenic viral proteins including EBNA 1-3, LMP1 and LMP2 that induce potent EBVST responses. Such potent T cell immunity maintains the infected B cell pool at <2% of total B cells in immunocompetent individuals but is lacking in immunosuppressed individuals leading to uncontrolled lymphoproliferation of the infected B cells. What are the indications of adoptive immunotherapy?
EBV-associated lymphoproliferative disease Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
It can be used as prevention of EBV related PTLD What are the side effects of adoptive immunotherapy?
-Graft-versus-host disease
· The gamma herpes virus family member Epstein-Barr virus (EBV) is common and linked to a number of lymphomas and lymphoproliferative diseases (LPD). More than 90% of adults globally are infected with it.
· Primary infection typically manifests in childhood as asymptomatic or mild clinical presentation or during adolescence as florid mononucleosis-like symptoms.
· It remains its potency in B cells or oral epithelial cells throughout life.
· EBV-infected naive B-lymphocyte nuclear antigens expressed proteins like EBNA1, EBNA2, EBNA3, and EBNALP, membrane proteins LMP1, and LMP2 as well as BARF1 and two small non-translated ribonucleic acids (RNA) (Type III latency)
· In healthy individuals, the rate of EBV infection remains stable at 0.1-50 EBV-infected B lymphocytes per 1,000,000 peripheral blood mononuclear cell (PBMCs).
· In individuals with weakened immune systems, such as patients with primary immunodeficiency (PID) or infection with human immunodeficiency virus (HIV), recipients of hematopoietic stem cell transplantation (HSCT) or solid organ transplant (SOT), the lack of a robust EBVST response can lead to uncontrolled proliferation of type 3 latency EBV-infected B cells resulting in EBV-associated LPD and malignancies.
· Manufacturing EBV-specific T-lymphocytes (EBVST) is an adoptive transfer of virus-specific T cell . The use of donor lymphocyte infusions (DLI) to donor-derived multi-antigen specific VSTs in the HSCT setting.
· Manufacturing EBVST products as ex vivo expansion EBVSTs with antigen-specific T cell.
· Autologous EBVSTs used in HSCT and SOT recipients where their donors are not available and it is a preferred method used for PTLD.
· Third-party T cell designed to overcome the delay that may come with either autologous or allogenic methods of producing EBVSTs and readily available “off the shelf” T-cell therapy product. Many studies have reported a significant reduction in EBV viral loads following treatment with third-party T cell.
· Modification of EBVSTs treatment to enhance the activity which overcome the immune suppressive effects of TGF- -β to enhance EBVSTs activity in vivo. Calcineurin resistance to enhance EBVSTs and Chimeric antigen receptor T cells.
· Combination therapy of EBVSTs with other therapeutic modalities in vivo as demethylating agents, checkpoint inhibitors and BCL-2 inhibitors.
· The use of adoptive immunotherapy, particularly using third-party “off the shelf” EBVSTs for the treatment of EBV-LD has shown promise in several studies conducted at specialized centers . More recently, EBVSTs have become more widely available including in industry led multi-center studies and consortium and cooperative group studies.
Indications for adoptive immunotherapy:
· Treatment of viral infections that are difficult to cure like CMV, EBV. · Immunocompromised patient with EBV-negative lymphoproliferative disease. · Treatment of cancers that are caused by specific viruses, leukemias, DLBC lymphoma. · Treatment of autoimmune diseases like lupus.
Use of adoptive immunosuppressive for treatment of other conditions related to organ transplant:
· It has been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma, and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients. · Can be used in BK virus nephropathy in transplant recepients, melonoma, EB virus associated malignancy. Side effects · Graft versus host disease · Graft rejection · IgE mediated anaphylactic reaction · Neurotoxicity · Cytokine release syndrome
1.Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
EBV is linked to a number of cancers that affect B-cells, T-cells, NK cells, epithelial cells, and smooth muscle. Although each of these is connected to the latent EBV life cycles, the pattern of latency-associated viral antigens produced in tumor cells varies depending on the kind of tumor.
HSCT and SOT are significantly complicated by EBV-positive PTLD.
Donor lymphocyte infusion was the first adoptive immunotherapy treatment to be successful in treating EBV+ PTLD in HSCT patients, although it was associated with a significant risk of GVHD.
Subsequently EBV-specific T cells (EBVSTs) have been studied as a prevention and treatment for EBV-associated cancers, including PTL.
The use of adoptive immunotherapy, particularly using third-party “off the shelf” EBVSTs for the treatment of EBV-LD has shown promise in several studies conducted at specialized centers.
Current clinical experience with adoptive cellular therapy for EBV-LD:
In clinical trials, adoptive cellular therapy using EBVSTs has shown impressive results in immunocompromised patients with EBV-LD and there is the potential for wider use.
EBVST donor options:
EBVSTs can be produced from seropositive donors using good-manufacturing-practice-compliant protocols and cryopreserved for future use.
Donor types include autologousand allogeneic (including third party) sources.
EBVST manufacturing strategies:
ex vivo expansion of VSTs versus antigen-specific T cell selection [e.g., interferon γ (IFN-γ) capture]
MHC multimer selection where oligomeric forms of MHC molecules are designed and conjugated to magnetic beads to isolate typically CD8+ T cells with high affinity to a specific viral epitope/peptide in an HLA restricted manner
IFN-γ capture approaches where mononuclear cells are pulsed with antigen (e.g., single peptides), etc.] to isolate CD8+ and CD4+ T cells which secrete IFN-γ in response to the viral antigens.
Combination therapies with other immunotherapeutic approaches:
Current research is focused on strategies to enhance EBVST potency in vivo through genetic engineeringas well as combination therapies.
============================== 2. What are the indications of adoptive immunotherapy?
Treatment of newly diagnosed PTLD in pediatric SOT recipients.
Refractory CMV, EBV and or adenoviral infections post HSCT in pediatric patients.
For pediatric patients with PID disorders suffering from refractory viral infections prior to HSCT.
============================== 3. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation? Yes
EBVSTs have been used, as adjunctive therapy to chemotherapy and/or in relapsed patients, to treat type II latency EBV-associated malignancies including:
Hodgkin’s lymphoma
Natural killer (NK)/T-cell lymphoma
Nasopharyngeal carcinoma
EBVSTs have also been used to treat type I latency tumors (e.g., Burkitt lymphoma or gastric carcinoma)
============================== 4. What are the side effects of adoptive immunotherapy?
Introduction
Ebstein- Bar virus is a worldwide widely spread gamma herpes virus family that has been associated with lymphoproliferative disease in solid organ transplant and hematopoietic stem cell transplant. The virus infects more than 90% of the population and it has the following characteristics in the mode of operation.
Primary infection usually occurs in childhood as asymptomatic/mild clinical presentation or with florid mononucleosis-like symptoms during the teenage years
It maintains a lifelong potency in B cells or in oral epithelia cell
EBV-infected naive B-lymphocyte nuclear antigens proteins like EBNA1, EBNA2, EBNA3, and EBNALP, membrane proteins LMP1, and LMP2 as well as BARF1 and two small non-translated ribonucleic acids (RNA) (Type III latency)
In healthy individuals, the rate of EBV infection remains stable at 0.1-50 EBV-infected B lymphocytes per 1,000,000 peripheral blood mononuclear cell
Rituximab use, either as monotherapy or in combination with steroids or cyclophosphamide for the treatment of HSCT has been limited by high infection rate, recurrence of disease, and severe toxicity known with those medications.
Hence, the need for the development of medication that addresses the underlying immune defect with little to no toxicity
Manufacturing EBV-specific T-lymphocytes (EBVST)
Is an adoptive transfer of virus-specific T cell
use of donor lymphocyte infusions (DLI) to donor-derived multi-antigen specific VSTs in the HSCT setting
It is produced from EBV-positive donors using good manufacturing grade, and the donor type could be;
– autologous
-allogeneic
Methods used in manufacturing EBVST products
Ex vivo expansion EBVSTs
Antigen-specific T cell
Post-HSCT donor-derived T-cell therapy
a) Donor lymphocytic infusion
effective in remission but with attendant GVHD
b) Donor-derived EBVSTs
67% response rate but a very GVHD effect was seen among those treated with it
Autologous EBVSTs
used in HSCT and SOT recipients where their donors are not available
in theory, it is a preferred method used for PTLD
Delay in the production and administration of the drug because its autologous
Has been used in the treatment of type 11 latency EBV-associated malignancy
Third-party T cell
Designed to overcome the delay that may come with either autologous or allogenic methods of producing EBVSTs
It is a readily available off-the-shelf T-cell therapy product
Its response rate (CR and PR) was 64% at 5 weeks and 52% at 6 months with no significant toxicity observed
In the pediatric age group, it has a more favourable outcome
Many study groups have reported a significant reduction in EBV viral loads following treatment with third-party T cell
Modification of EBVSTs treatment to enhance the activity
Overcoming the immune suppressive effects of TGF- beta to enhance EBVSTs activity in vivo
Calcineurin resistance to enhance EBVSTs
Chimeric antigen receptor T-cell
Combination therapy of EBVSTs with other therapeutic modalities
Demethylating agents
Checkpoint inhibitors
BCL-2 inhibitors
Conclusion
The adoptive immunotherapy in the treatment of PTLD in SOT or HSTC especially the third- party T- cell has demonstrated a better clinical outcome with less toxicity and alleviating the concern of GVHD
Indications for adoptive immunotherapy
immunocompromised patient with EBV-negative lymphoproliferative disease
Those that failed to respond to Rituximab monotherapy or plus steroid and cyclophosphamide
Use of adoptive immunosuppressive for treatment of other conditions related to organ transplant
It has been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma, and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients
Autoimmune diseases like MS, or lupus
Use in the treatment of inflammatory disease
Side effects
Graft versus host disease especially with the use of DLI
Principles of adoptive immunotherapy:
In healthy seropositive individuals, virus neutralizing Ab control the spread of the infectious virus particles and the EBV specific HLA class I restricted, CD8 T lymphocytes specific to the early lytic cycle proteins kill cells entering the lytic cycle before they are able to release infectious virus particles
EBV maintains a lifelong latency in B cells and oral epithelial cells. CD 21 found on the B cells is the major receptor for the virus. EBV infected naive B lymphocytes express proteins comprising the EBV genome including:
EBV nuclear Ag
EBV membrane proteins
2 small non-translated RNA
This is type III latency pattern
The EBV infected B lymphocytes then enter the lymphoid follicles and downregulate the immunogenic proteins to express less immunogenic type II latency proteins – EBNA 1, LMP1 and LMP2. This is termed as latency type II pattern
The virus persists in latently infected B cells by further downregulating expression of viral proteins – latency type I – so as to become invisible to EBSVTs
In individuals with a weakened immune system, the lack of a robust EBSVT response can lead to uncontrolled proliferation of type III latent EBV infected B cells resulting in EBV associated LPDs and malignancies
The latent type III B cells that cause LPDs present several antigenic viral proteins including EBNA 1-3, and LMP 1 and 2 that induce potent EBSVT response. Such potent T cell immunity maintains the infected B cell pool at less than 2% of total B cells in immunocompetent individuals but is lacking in immunosuppressed individuals leading to uncontrolled proliferation of the infected B cells.
The scientific rationale for the adoptive transfer of EBSVTs id based on harnessing the immunogenicity of type III latent B cells to control the proliferation of these cells that cause the LPDs and other malignancies.
EBV associated malignancies associated with type II latency (Hodgkins lymphoma, NK/T cell lymphoma) or type I latency (Burkitts lymphoma or gastric carcinoma) can develop in immunocompetent and immunosuppressed individuals and are much less immunogenic due to downregulation of the immunodominant Ag.
Manufacturing of EBSVTs will require lymphocytes which can be:
Autologous
Allogeneic
Third party
There are different ways to manufacture the EBSVTs:
Ex vivo expansion of the EBSVTs
Antigen specific T cell selection
Allogeneic lymphocytes: Can be donor lymphocyte infusions or donor derived EBSVTs
Autologous EBSVTs
Third Party T cells
There are several ways of enhancing the activity of EBSVTs:
Overcoming the immunosuppressant effects of TGF beta to enhance EBSVT activity
Calcineurin resistance to enhance EBSVTs
Chimeric Ag receptor T cells
One can also administer EBSVTs with other therapeutic modalities to enhance EBSVT activity in vivo:
Demethylating agents
Checkpoint inhibitors
BCL 2 inhibitors
Indications of Adoptive Immunotherapy:
Treatment of viral infections that are difficult to cure like CMV, EBV
Treatment of cancers that are caused by specific viruses, leukemias, DLBC lymphoma
Treatment of autoimmune diseases
Treatment of chronic hypersensitivity reactions
Other Uses Of Adoptive Immunotherapy In Organ Transplant:
In GVHD disease (although it is one of the adverse effects of this therapy
Infections post transplant like CMV that is resistant to antiviral treatment
Side Effects:
In HSCT patients, it can cause GVHD
It can lead to hypersensitivity reactions
Can cause immediate infusion reactions like fevers, pruritus
It can elicit a strong immune response which theoretically can lead to acute rejection
That is an excellent summary and very well structured reply. I wish you could type the heading of first para as Introduction of Principles of Immunotherapy, and then you could type sub-headings in bold or underline or in italics. That would make it easier to read. What is your analysis of level of evidence of this study?
2) To enhance the specificities to less immunogenic EBV antigens by transducing DC and LCLs with adenovirus vectors to overexpress LMP1 or LMP2.
3) The novel antigen-presenting complex (KATpx)] can facilitate the rapid (10–21 days as opposed to 2–3 months) expansion of T cells targeting EBNA1, LMP1 and LMP2 from healthy donors and from patients with type 2 latency EBV-associated malignancies.
b) Antigen-specific T cell selection [e.g., interferon γ (IFN-γ) capture].
1) major histocompatibility complex (MHC) multimer selection where oligomeric forms of MHC molecules are designed and conjugated to magnetic beads to isolate typically CD8+ T cells with high affinity to a specific viral epitope/peptide in an HLA restricted manner.
2) IFN-γ capture approaches where mononuclear cells are pulsed with antigen to isolate CD8+ and CD4+ T cells which secrete IFN-γ in response to the viral antigens.
c) Post-HSCT donor-derived T cell therapy
1) Donor lymphocyte infusions.
2) Donor-derived EBVSTs.
d) Autologous EBVSTs: has been used for patients with EBV-associated malignancies outside the context of allogeneic HSCT and in SOT recipients with PTLD where donors usually are not available because of the use of cadaveric grafts.
e) Third-party EBVSTs: a readily available “off the shelf” T cell therapy product is desirable as patient-specific products may not be possible or may be so delayed.
f) Modification of EBVSTs to enhance activity
1) Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo.
2) Calcineurin resistance to enhance EBVSTs:Given the concerns regarding EBVST persistence in patients who require ongoing immune suppression.
3) Chimeric antigen receptor T cells: using CD19-chimeric antigen T cells (CD19-CART).
g) Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance EBVST activity in vivo
1) Demethylating agents; Azacytidine and decitabine are potent CpG demethylating agents.
2) Checkpoint inhibitors the risk of graft rejection and autoimmunity limits their use in that setting.
3) BCL-2 inhibitors; BCL-2 inhibitor induce remission in approximately 70% of mice PTLD xenografts.
Indications of adoptive immunotherapy:
Immunotherapy is used to upregulate or downregulate the immune system to achieve a therapeutic effect in immunological mediated disorders:
· Hyper-sensitivity reaction.
· Auto-immune diseases.
· Tissue and organ transplantation.
· Malignancies; Type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients.
· Inflammatory disorders.
· Infectious diseases.
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Autologous EBVSTs have also been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients.
The side effects of adoptive immunotherapy:
1) Donor lymphocyte infusions carries a significant risk of graft-versus-host disease (GVHD).
2) Selective depletion of T-regulatory cells (Tregs) prior to infusion enhances the graft versus lymphoma (GVL) effect.
3) Serious adverse events in relation to third-party EBVSTs are TMA and GI hemorrhage(all deemed unrelated).
4) Adverse effects in relation to CART therapy include cytokine release syndrome (CRS), neurotoxicity and acute kidney injury.
5) Checkpoint inhibitors: the risk of graft rejection and autoimmunity limits their use.
Briefly summarize the principle of adoptive immunotherapy in the treatment of PTLD. Adaptive immunotherapy involves isolation of specific T lymphocytes which can be used to target infection and malignancies. Due to immunosuppressive medication in post-transplant patients, the cytotoxic T lymphocyte response is weakened, which leads to uncontrolled proliferation of cancerous cells and viral infections such EBV. Adaptive immunotherapy can be used as means of prevention or treatment against EBV and PTLD in transplant recipients. What are the indications of adoptive immunotherapy? It can be used for the treatment and prophylaxis of EBV related PTLD in transplant patients. And also, in treatment of type 2 latency EBV associated cancers in relapsed patients. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation? Skin cancers, melanoma, EBV associated PTLD. BK virus nephropathy What are the side effects of adoptive immunotherapy? Neurotoxicity, Acute Kidney Injury GVHD, TMA Cytokine release syndrome.
GIT hemorrhage, IgE mediated anaphylactic reaction.
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
Adoptive immunotherapy involves isolation of T lymphocytes and then using then to treat cancers or other diseases. Antibodies which neutralise viral particles can control spread of viral disease. also EBV specific cytotoxic T lymphocytes can kill cells before release of viral particles. EBVSTs- EBV specific T lymphocytes are important in immune response. In transplant recipient , EBVSTs response is weak due to weak immune system . This can cause proliferation of B cells. This will lead to PTLD.
The scientific rationale for adoptive transfer of EBVSTs is based on harnessing the immunogenicity of type III latency malignancies to control the proliferation of the latently infected B cells.
What are the indications of adoptive immunotherapy?
It can be used in Type 2 latency EBV associated malignancy with chemotherapy or relapse.
It is also used in prophylaxis of EBV related PTLD
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
It can be used in BK virus nephropathy in transplant recepients, melonoma, EB virus associated malignancy.
What are the side effects of adoptive immunotherapy?
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
Adoptive immunotherapy involves isolation and infusion of T lymphocytes in a patient for treatment/ prevention of a disease. Epstein-Barr virus (EBV) infects more than 90% of adult population. Virus neutralizing antibodies control spread of infectious viral particles and EBV-specific CD8+ cytotoxic T lymphocytes kill cells before they release viral particles.
EBV specific T lymphocytes (EBVSTs) inhibit the entry of EBV infected B cells into the lymphoid follicles. In transplant recipients, due to weakened immune system, EBVST response is lacking, leading to uncontrolled proliferation of EBV-infected B cells and resultant EBV-associated post-transplant lymphoproliferative disorders (PTLD) and malignancies. So, infusion of EBVSTs help in reducing the latently infected B cell pool.
Autologous or allogeneic (including third party) source of EBVSTs are used to manufacture them using either ex vivo expansion or antigen-specific T cell selection )interferon gamma capture).
2. What are the indications of adoptive immunotherapy?
Adoptive immunotherapy is indicated in prevention and treatment of EBV-associated lymphoproliferative disease, and in patients with elevated EBV viral load.
3. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Adoptive immunotherapy has been utilized in BK virus nephropathy in transplant recipients (1). They have also been utilized in melanoma.
4. What are the side effects of adoptive immunotherapy?
Side effects of adoptive immunotherapy include GVHD (graft versus host disease), acute kidney injury, gastrointestinal hemorrhage, and thrombotic microangiopathy. Chills, fever, hypotension, oliguria, edema (due to capillary leak syndrome), uveitis, cytokine release syndrome, confusion, delirium, seizures, cerebral edema, and IgE mediated anaphylactic reaction have been also seen with them (2).
The article is a narrative review: Level of evidence – Level 5
References:
1) Jahan S, Scuderi C, Francis L, Neller MA, Rehan S, Crooks P, Ambalathingal GR, Smith C, Khanna R, John GT. T-cell adoptive immunotherapy for BK nephropathy in renal transplantation. Transpl Infect Dis. 2020 Dec;22(6):e13399. doi: 10.1111/tid.13399. Epub 2020 Jul 14. PMID: 32608543; PMCID: PMC7816252.
2) Rohaan MW, Wilgenhof S, Haanen JBAG. Adoptive cellular therapies: the current landscape. Virchows Arch. 2019 Apr;474(4):449-461. doi: 10.1007/s00428-018-2484-0. Epub 2018 Nov 23. PMID: 30470934; PMCID: PMC6447513.
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD.
EBV infects more than 90% of the adult population. Primary infection mainly occurs in childhood leading to either asymptomatic/mild infection/infectious mononucleosis.
EBV enters the oropharynx replicating in the epithelial cells and infecting transiting B lymphocytes. The infected B lymphocytes express proteins representing the whole EBV genome. They then enter the lymphoid follicles where they down regulate the immunogenic proteins to express less immunogenic type 2 latency proteins. They thus enter the memory compartment where the virus persist in latently infected B lymphocytes.
The frequency of EBV infected B lymphocytes remain stable in a healthy individual. However, in an immunocompromised individual, lack of a robust EBV specific T lymphocytes leads to uncontrolled proliferation of type 3 latency EBV leading to LPD and malignancies. The rationale of adoptive immune transfer is based on harnessing the immunogenicity of type 3 latency malignancies to control the proliferation of latently infected B cells.
What are the indications of adoptive immunotherapy?
Treatment and prophylaxis of EBV related PTLD in transplant recipients.
Treatment of type 2 latency EBV associated malignancies as an adjuvant to chemotherapy or in relapsed patients.
Other conditions
Type 2 latency EBV associated malignancy
Skin cancer-melanoma
Side effects
GVHD
Cytokine release syndrome
Acute kidney injury
Neurotoxicity
1-Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
Introduction:
– EBV-positive PTLD is a major complication of HSCT and SOT.
– EBV-infected naïve B-lymphocytes express proteins comprising the entire EBV genome
– After primary EBV infection, most viral antigens are expressed during the lytic stage while the virus is replicating.
– In EBV-infected B-cells, there is subsequent downregulation of viral antigen expression from latency type III to latency type 0 which allows escape from immune surveillance.
-Periodic expansion of EBV-infected B-cells requires re-expression of viral antigens which re-stimulates the EBVST response.
– The lack of a robust EBVST response can lead to uncontrolled proliferation of type 3 latency EBV-infected B cells resulting in EBV-associated LPD and malignancies.
– PTLD is highly immunogenic and amenable to immunotherapy with EBVSTs.
-The scientific rational to adoptive transfer of EBVSTs is based on harnessing the immunogenicity of type III latency malignancies to control the proliferation of the latently infected B cells.
Manufacturing of EBVSTs
It can be produced from EBV-positive donors using good-manufacturing-grade (GMP) compliant methodologies with minimal alloreactivity and broad specificity against EBV latency proteins or as a multi-virus specific product with activity against multiple viruses.
Donor types; autologous and allogeneic (including third party).
The most common strategies: Ex vivo expansion of EBVSTs:
-Utilizing irradiated EBV transformed lymphoblastoid cell lines (LCL) as antigen presenting cells (APCs) to selectively expand EBVSTs.
– LCL-activated EBVSTs consist of a product with activity against early lytic antigens and EBNA 3A, 3B and 3C but unreliable activity towards LMP1 and LMP2.
– To enhance the specificities to less immunogenic EBV antigens LMP1 and LMP2, transducing DC and LCLs with adenovirus vectors to overexpress LMP1 or LMP2.
– Utilize whole antigen
Antigen-specific T cell selection:(IFN-γ) capture
– Selection and isolation of CD8+ T cells with high affinity to a specific viral epitope/peptide in an HLA restricted manner.
– Mononuclear cells are pulsed with antigen to isolate CD8+ and CD4+ T cells which secrete IFN-γ in response to the viral antigens.
– Require seropositive donors with high frequency of circulating antigen/epitope specific T cells.
Post-HSCT donor-derived T cell therapy Donor lymphocyte infusions DLI.
-Utilized unmodified DLI derived from the patient’s EBV seropositive HSCT donor which contained effector cells with activity against EBV.
– Carries a significant risk of GVHD, selective depletion of T-regulatory cells (Tregs) reduces the risk.
Donor-derived EBVSTs
-Using donor derived EBVSTs for the prevention and treatment of EBV+ PTLD in the post-HSCT setting when the donor is available
Autologous EBVSTs
– Used for patients with EBV-associated malignancies outside the context of allogeneic HSCT and in SOT recipients with PTLD. – In SOT it is preferred to donor EBVSTs even if available – The challenges in production and the production time leading to delays in initiation of therapy in patients with a rapidly progressive disease.
– Used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients.
– Such tumors are more challenging targets because of the reduced expression of immunogenic viral antigens which express a more restricted array of antigens compared to type III.
– Better responses were observed with EBVST products that included T cells with activity against LMP1 and LMP2.
Third-party T cells – Readily available T cell therapy; EBVST bank with HLA alleles prevalent at high frequencies in individuals community.
– No significant toxicities were observed, alleviating concerns of graft rejection and/or GVHD with this therapy which fueled the broadened applicability of this approach.
– Multiple studies used third-party multi-VSTs for EBV-associated PTLD, no significant adverse events attributable to
the product were observed.
– Third-party EBVSTs have been mostly used in the posttransplant setting.
– No published experience in patients with HIV-associated lymphomas.
Modification of EBVSTs to enhance activity
The efficacy of EBVSTs is dependent on the expression of viral antigens and limited by the paucity of EBV antigen expression in malignancies of latency type I and II.
Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo
– In the immunocompetent host, TGF-β can be released into the tumor microenvironment by tumor cells, fibroblasts and immune cells and creates an immunosuppressive environment by impeding T-cell activation and it affects antigen presenting cells and suppresses functional antitumor T cell responses in vivo – The potential to overcome the immunosuppressive properties of TGF-β has been studied in EBV-positive HL. Calcineurin resistance to enhance EBVSTs;
-As there is a concerns regarding EBVST persistence in patients who require ongoing immune suppression SOT, EBVSTs were genetically engineered to express a CNI-resistant EBVSTs than can persisted with enhanced activity in the presence of CNI.
Chimeric antigen receptor T cells:
-CD19-chimeric antigen T cells (CD19-CART) have shown impressive efficacy in acute B-lymphoblastic leukemia and
in B-cell lymphomas. In addition to viral antigens, EBV-LD expresses a variety of B-cell antigens targetable by
CARTs including CD19, CD20 and CD30.
-However, their use in EBV-lymphoproliferation is limited by the patient’s immunosuppressive state impeding T cell manufacture and the length of production time. Combination strategies :EBVSTs with other modalities to enhance EBVST activity in vivo Demethylating agents:
– Newly EBV-infected B-cells express viral genes. Rapid CpG-methylation of viral antigens leads to downregulation of viral protein expression and the latency. Checkpoint inhibitors
-EBV-positive diffuse large B-cell lymphoma (DLBCL) and EBV-positive PTLD were found to express PD-L1.
– LMP1 induce expression of the checkpoint protein PD-L1
– Using checkpoint PD-L1 blockade.
BCL-2 inhibitors
-Latently EBV-infected cells inhibit proapoptotic signals thus ensuring immortality.
– LMP1 upregulates the expression of the anti-apoptotic protein BCL-2.
– BCL-2 inhibitor increased proapoptotic proteins and sensitivity to CD19-CART;
Conclusion:
-The use of third-party EBVSTs for the treatment of EBV-LD has shown promise in several studies.
– Further Preclinical work and early clinical trials is needed.
2-What are the indications of adoptive immunotherapy?
-Treatment and prophylaxis of PTLD in transplant recipients HSCT, SOT.
-In refractory cases.
-EBV- associated malignancies.
3-Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation? -Skin cancer; metastatic melanoma.
– EBV-associated malignancies.
– Autoimmune diseases. 4-What are the side effects of adoptive immunotherapy?
-Most people have a mild form of cytokine release syndrome. But in some people, it may be severe or life-threatening.
– GVHD.
– Capillary leak syndrome.
– Fever.
– Flue like illness.
– Skin rash.
– Neurotoxicity
– Gastrointestinal symptoms; diarrhea, nausea, vomiting.
– Acute kidney injury.
–Graft rejection and autoimmune complications.
– TMA, GI hemorrhage.
Briefly summaries the principle of adoptive immunotherapy in the treatment of PTLD
Introduction: EBV, a common viral infection in childhood, affecting 90% of the adult population. Normally the immune system contains the virus and protect the body against spreading of this infection, this causes EBV to undergo into a dormant phase called as latent phase in the B cells.
But all the patients with the defective immune system, i.e., HIV. HSCT. SOT. can lead to uncontrolled infection and proliferation which can lead to EBV- associated PTLD and malignancies.
The transformed B-cells in EBV-associated Lymphoproliferative Disease associated with type III latency, present Viral proteins: EBNA 1-3. LMP1 and 2 , such antigens in intact immune systems, T-cells can control B cells and maintain infected B cells in the range of < 2%. Type II latency associated with malignancies: NHL, Hodgkins’s lymphoma. Natural killer T-cell lymphoma. Nasopharyngeal carcinoma, NPC. Type I latency tumors:Burkitt’s lymphoma. Gastric carcinoma.
The occurrence of PTLD in HSCT ranges from 1-25%, depending on the following:
Serostatus of the donor.
The degree of patient T- cell depletion.
Post-HSCT immunosuppressants.
Sero negativity in a transplant recipient is the top determinant factor of the development of post-transplant LPD. PTLDs is usually seen more commonly in pediatric patients. Monotherapy with Rituximab anti-CD20 is more effective in HSCT with a response rate of 55 -100%, while combination therapy (prednisolone/cyclophosphamide/rituximab) is shown to better survival up to 725.
#EBVSTs manufacturing:
Ex vivo expansion of EBVSTs.
Antigen-specific T cell selection.
#Post-HSCT donor-derived T-cell therapy:
Donor Lymphocyte infusion.
Donor-derived EBVSTs.
# Autologous EBVSTs.
# Third-party T cells.
the purpose of Modification of EBVST is to enhance activity of the immune suppressive effects of TGF-B in vivo, Calcineurin resistance to enhance EBVSTs and Chimeric antigen receptor T cells.
different Combination strategies are adopted, with EBVSTs which for example could be use of Demethylating agents, use of Checkpoint inhibitors, use of BCL-2 inhibitors.
Conclusion:
Promised result of treatment with adoptive immunotherapy has been shown, using third-party EBVSTs for the treatment of EBV related PTLD.
EBVSTs should become widely available, however, more effort is needed to make third-party cell banks broadly available and easily applicable.
Indication of adoptive immunotherapy: Immunocompromised recipient with EBV-PTLD. and Prophylactic treatment in reduction of EBV load.
Uses of adoptive immunotherapy other than cancer treatment:
Antirejection therapy.
Other autoimmune conditions; Lupus, Type 1 DM, Multiple sclerosis.
Inflammatory diseases; atherosclerosis and hardening of arteries.
Summary
PTLD is a major complication of solid organ donation caused by childhood EBV viral activation. EBV infects more than 90% of the adult population worldwide. PTLD is classified into B cell lymphoma (90%) and T cell lymphoma accounts for 10%.
EBV infects B cells in two key stages after primary infection:
Lytic stage, where the patient may be asymptomatic or develop mild symptoms or frank infectious mononucleosis; latency stage, where the virus becomes latent in B cells lifelong and the patient is asymptomatic but at risk of developing certain lymphoproliferative diseases; and several stages of latency develop (Latency III, II, I).
During latency III, EBV-infected B-lymphocytes express the EBV nuclear antigens EBNA1, 2, 3, EBNALP, LMP1, 2, and BARF1. Due to excessive antigenic expression, T lymphocytes restrict B cell proliferation to 2%. Because immunosuppression leads B lymphocytes to proliferate uncontrollably, the Latency III stage is connected with lymphoproliferative diseases including cancer. The most prevalent malignancy at this stage is PTLD.
In Latency II and I, EBNA3 and EBNA2 are downregulated, rendering them less immunogenic. Latency I and II stages are associated with lymphoproliferative diseases and cancers in immune-competent and immune-compromised patients, such as Hodgkin’s lymphoma, natural killer/T-cell lymphoma, nasopharyngeal carcinoma, Burkitt lymphoma, and gastric carcinoma (Latency I).
In PTLD adoptive immunotherapy, EBV-specific cytotoxic T lymphocytes or donor lymphocyte infusion kill dividing EBV-positive B cells.
What are the indications of adoptive immunotherapy?
Treatment of EBV+ PTLD cases that are resistant or refractory
Treatment of some solid tumors, such as glioblastoma and GIT tumors
Treg adoptive immunotherapy is used to treat certain chronic viral infections such as HIV, as well as autoimmune diseases such as type I DM and SLE.
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Through the infusion of Treg cells, adoptive immunotherapy can be used to treat acute rejection refractory to standardized treatment.
Adoptive therapy can also be used in the treatment of autoimmune diseases such as SLE and viral infections such as BK virus and CMV infection.
What are the side effects of adoptive immunotherapy?
The adverse effects of adoptive treatment depend on the type of cells employed and the reason for infusion. The following are typical adverse effects: constitutional symptoms during infusion is common, Chronic and acute graft-versus-host disease, Cytokine storm, Neurotoxicity
That is an excellent summary and very well structured reply. I wish you could type the heading of first para as Introduction. Under this main heading, I wish you could type sub-headings in bold or underline or in italics. That would make it easier to read. What is your analysis of level of evidence of this study?
Epstein-Barr virus (EBV) is a member of gamma herpes family associated with various lymphomas and lymphoproliferative disorders (LPD).
It infects more than 90% of the adult population worldwide.
Primary infection occurs in childhood as an asymptomatic or mild infection and may result in a more florid infectious mononucleosis syndrome in teenagers and young adults
In healthy seropositive individuals:
Virus-neutralizing antibodies control the spread of infectious virus particles, and EBV-specific, human leukocyte antigen (HLA) class I restricted, CD8+ cytotoxic T lymphocytes (CTL) specific to the early lytic cycle proteins kill cells entering the lytic cycle before they can release infectious virus particles
In individuals with weakened immune systems,
Such as patients with primary immunodeficiency (PID) or infection with human immunodeficiency virus (HIV), recipients of hematopoietic stem cell transplantation (HSCT) or solid organ transplant (SOT), the lack of a robust EBVST response can lead to uncontrolled proliferation of type 3 latency EBV-infected B cells resulting in EBV-associated LPD and malignancies.
EBV-associated malignancies associated with :
Type II latency [e.g., Hodgkin’s lymphoma (HL), natural killer (NK)/T-cell lymphoma, nasopharyngeal carcinoma (NPC)]
Type I latency tumors (e.g., Burkitt lymphoma or gastric carcinoma) can develop in immunocompetent, and immune deficient individuals and are much less immunogenic due to downregulation of the immunodominant (e.g., EBNA3 and EBNA2) antigens
Type 3 latency EBV-infected B cells resulting in EBV-associated LPD and malignancies
Manufacturing of EBVSTs:
Donor types include :
Autologous
Allogeneic (including third-party) sources.
Ex vivo expansion of EBVSTs:
GMP-grade EBV-specific T-cell manufacturing. Mononuclear cells are harvested from the peripheral blood of a donor. B-lymphocytes are infected with laboratory strain EBV, transduced with an adenoviral vector expressing LMP1 and LMP2, and irradiated. Next, monocytes are separately transduced with the same adenoviral vector and cocultured with T cell, followed by a second stimulation by the transduced LCL to create and further expand the LMP-specific T-cell product.
Antigen-specific T-cell selection:
Post-HSCT donor-derived T-cell therapy:
Ø Remissions,
Ø This therapy carries a significant risk of graft-versus-host disease (GVHD)
this article elucidates the novel therapy used for EBV related post-transplant lymphoproliferative disease EBV-LPD. EBV:
It’s an extensively common virus belongs to herpes virus family. It was found to be linked to variable types of lymphomas and lymphoproliferative disorders LPD. its prevalent in 90% of adult people. Its inhabitant of oropharyngeal epithelial cells and infecting trafficking B-lymphocytes as they have a specific receptor for EBV, that is CD21. Post transplant LPD :
Part of pre transplant preparation is ascertaining EBV serology status of donor and recipient, as it’s a common risk factor for post transplant LPD , particularly in the context of donor positive recipient negative serology for EBV.
in the context of competent immune system T lymphocytes recognized B lymphocytes infected with EBV, as those B lymphocytes are expressing EBV related protein on surface membrane, full virus genome is expressed in B-lymphocytes surface membrane type III latency, some Lymphocytes might undergo downregulation of antigen presentation type II latency , hence the percentage of B-lymphocytes infected with EBV is generally maintained at maximum of 2 % of total B lymphocytes, a status that is entirely deranged in immunocompromised condition such as HIV infected patients, and post transplant status where B lymphocytes proliferate out of control resultant in PTLD. Type III latency associated tumors:
PTLD Type II latency associated tumors:
Hodgkins Lymphoma and nasopharyngeal carcinoma. Type 1 latency associated tumors:
Burkitts lymphoma and gastric carcinoma. EBVSTs:
those T lymphocytes are sensitive to EBV infected B lymphocytes and tagged as EBVSTs. This clone of T lymphocytes made the basics for adoptive transfer of EBVSTs to the patient with PTLD to control unchecked proliferation of B lymphocytes.
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD:
Introduction: Epstein-Barr virus – a widespread (almost more than 90% of the population infected) gamma herpes virus family, associated with lymphomas and lymphoproliferative diseases. Childs usually asymptomatic or with mild symptoms, adolescents may have severe infectious mononucleosis syndrome. in seropositive people the antibodies control the spread of virus by various immune system actios: – Cytotoxic T cells kills the lysed cells before releasing the viral particles. – EBV remains latents in B cells and oral epithelial cells. – These infected B cells can rescue the virus by memory cells and down regulate the viral expression so the cytotoxic T (EBV specific T lymphocytes) cells cannot identify and attack it by various actions. Breakdown of this immune regulation, due to immune deficiencies – inherited or acquired (ie HIV, Immunosuppressive medications) results in activation of virus, and leads to a lymphoproliferative disorders and lymphoma. Type I latency tumors are: Burkitt’s lymphoma.Gastric carcinoma. Type II latency associated with malignancies are: HL, Hodgkins’s lymphoma, Natural killer T-cell lymphoma, and Nasopharyngeal carcinoma, NPC. Type III latency associated with PTLD: due to presenting cell proteins (EBNA1-3, LMP, and LMP2). 1-25% of hematopiotic stem cell transplant patients experience a PTLD through the following: – The degree of patients T cell depletion. – Seronegativity status. – The immunosuppressant’s. PTLD is highly immunogenic and amenable to immunotherapy with EBVSTs. Adoptively transferred virus-specific T cells (VSTs) have been evaluated for more than two decades, ranging from the use of donor lymphocyte infusions (DLI) to donor derived multi-antigen specific VSTs in the HSCT setting to autologous as well as allogeneic VSTs in the SOT setting.
Manufacturing of EBVSTs: – Ex vivo expansion of EBVSTs. – Antigen-specific T cell selection. Post-HSCT donor-derived T-cell therapy: – Donor Lymphocyte infusion. – Donor-derived EBVSTs. Autologous EBVSTs. Third-party T cells. Modification of EBVSTs to enhance activity:
Overcoming the immune suppressive effects of TGF-B to enhance EBVST activity in vivo.
Calcineurin resistance to enhance EBVSTs.
Chimeric antigen receptor T cells.
Combination strategy, with EBVSTs:
Demethylating agents.
Checkpoint inhibitors.
BCL-2 inhibitors.
Conclusion: Use of adoptive immunotherapy using a third party “off the shelf” EBVSTs for the treatment of EBV associated LD is promising with limited use in specialized centers, that should be widely available. EBVSTs anti-tumor activity for type I and type II latency associated tumors should be explored.
Level of evidence is V erratic review.
What are the indications of adoptive immunotherapy?
Treatment and prevention of lymphoproliferative disorders in immunosuppressed patients. Reduction in EBV viral load.
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Can be used to control other viral infection such as CMV, adenovirus.. etc Can be used to prevent viral induced skin diseases or cancers. Can be used to treat malignancies other than PTLD. Example glioblastoma and hepatic cancers. Can be used in treating vasculitis and connective tissue disease.
What are the side effects of adoptive immunotherapy? Graft versus host disease. Allergic reaction, flue like illness, fever, palpitation and shortness of breath. Neurotoxicity. Cytokine release syndrome.
1- Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD:-
Review of published literature spanning 20 years from 2000 to 2020. EBV-positive post-transplant lymphoproliferative disease (PTLD) is a major complication of hematopoietic stem cell and solid organ transplantation.
In clinical trials, adoptive cellular therapy using EBVSTs has shown impressive results in immunocompromised patients.
The Epstein-Barr virus (EBV) is a ubiquitous member of.the gamma herpes virus family that is associated with avariety of lymphomas and lymphoproliferative disorders.
Primary infection occurs in childhood as an asymptomatic or mild infection and/or may result in a more florid infectious mononucleosis syndrome in teenagers and young adults.
The Ebola virus (EBV) causes the production and proliferation of type 3 latency EBV-infected B cells resulting in EBV-associated lymphoproliferative disease (LPD and malignancies).
These B-lymphocytes present several antigenic viral proteins including EBNA 1-3, LMP1 and LMP2 that induce potent EBVST responses.
Such immune responses are lacking in immunosuppressed individuals leading to the proliferation of infected B cells.
EBV-associated post-transplant lymphoproliferative disease (PTLD) occurs in less than 1% to 25% of HSCT recipients.
The biggest risk factor for developing PTLD is EBV seropositivity at the time of transplant.
Rituximab, monoclonal antibody targeting CD20 present on B cells has been an effective monotherapy
EBVSTs can be expanded via ex-ex vivo expansion of T cells targeting viral antigens via T cell receptors.
LCL-activated EBV transformed lymphoblastoid cell lines (LCL) express high levels of class I and class II HLA and co-stimulatory molecules.
================================================================= Antigen-specific T cell selection
The rapid selection of CD8 + T cells with high affinity to a specific viral epitope/peptide in an HLA-restricted manner is one of the fastest and most effective ways to identify new drug candidates for use in the field of immunotherapy.
Autologous stem cell therapy and multimer-based approaches have been developed in the past, but their development has been hampered by a number of challenges, such as the HLA-restriction requirement and selection of a purely CD8 + T cell product that lacks CD4+ T cell help.
However, using both of these techniques, GMP-grade EBVST products can be made quickly from EBV seropositive autologous and allogeneic donors for clinical use.
==================================================================== Post-HSCT donor-derived T cell therapy
Donor lymphocyte infusions
This therapy carries a significant risk of graft-versus-host disease, or graft versus lymphoma (GVHD), and strategies have been employed to lessen the risk.
Donor-derived EBVSTs
Therapy was well tolerated with remarkable reduction of EBV viral copy numbers within 4 weeks.
None of the 101 patients who received donor derived EBVSTs as prophylaxis developed PTLD.
Of the 13 patients with active PTLD, 11 patients achieved a complete remission with evidence of VST persistence up to 9 years after transplant.
Autologous EBVSTs
Administration of autologous EBVSTs has been used for patients with EBV-associated malignancies outside the context of allogeneic HSCT and in SOT recipients with PTLD.
In the SOT setting, the production is technically more challenging because of the ongoing immunosuppression.
Autologous EBVSTs have been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy or in relapsed patients.
Such tumors are more difficult to treat because of the reduced expression of immunogenic viral antigens.
EBVST products produced by LCL stimulation alone have shown promising results in patients with type II latency-type malignancies (50,51).
A pilot study of 14 patients with relapsed EBV + HL reported 5 patients who maintained a CR for up to 40 months.
Production of EBVSTs was successful in 91% of patients, with LMP1 and/or LMP2-specificity detected in 66% of products .
A total of 50 patients with EBV-associated HL or Non-Hodgkin lymphoma were treated.
Of 29 patients receiving latent membrane protein (LMP)-specific T cells as adjuvant therapy after chemotherapy, 28 patients remained in a complete remission.
Adjuvant treatment with LMP-specific T cell therapy for patients with metastatic Non Hodgkin’s lymphoma (NPC) led to a significant increase in survival compared to a control group, according to a study.
The study was conducted at the Queensland Institute of Medical Research (QIMR) (Brisbane, Australia).
Third-party T cells
Patients with rapidly progressive disease may not be able to access patient-specific T cell therapies due to procurement times of donor or patient cells.
Patient-specific EBVST (autologous or allogeneic) manufacture can still be prolonged when procurement times are considered.
For these reasons, a readily available T cell therapy product is often the preferred treatment.
Thirty-three transplant recipients (stem cell, heart, kidney, liver, 10; liver and small bowel, 3; lung, 2; heart and lung, 1) with refractory PTLD between the age of 1–76 years received partially HLA matched EBVSTs.
Overall, the response rate (CR and PR) was 64% at 5 weeks and 52% at 6 months.
Chiou et al. from Birmingham, United Kingdom published experience in 10 pediatric SOT recipients with PTLD and reported an ORR of 80% (8 out of 10) .
This may indicate possible differences in the biology of EBV-driven PTLD in this population who is often EBV naïve at the time of transplant.
The BCM group has published extensively on their third-party EBVST products including the use of off-the-shelf VSTs for patients with severe refractory viral disease after BMT.
In a multicenter study, 15 products were given to 50 HSCT recipients with EBV, CMV, adenovirus, BKV and human herpes virus 6 (HHV6) .
The Memorial Sloan Kettering experience utilizing a third-party cell bank comprising 330 GMP-grade EBVSTs was reported by Prockop et al.
A total of 46 patients post-HSCT (33) or SOT (13) and PTLD were treated with three weekly infusions of the products.
Third-party EBVSTs have been mostly used in the post- transplant setting.
There has been no published experience with patients with HIV-associated lymphomas.
No standardization of EBV viral load measurements by polymerase chain reaction makes comparisons between different laboratories impossible.
Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo
TGF-β can be released into the tumor microenvironment by tumor cells, fibroblasts and immune cells and creates a immunosuppressive environment by impeding T-cell activation, proliferation and migration .
In addition, it affects macrophage antigen presentation and chemoattractant presentation.
Modification of EBVSTs to overcome these immune suppressive effects has led to increased activity in patients with latency type I and II.
The potential to overcome the immunosuppressive properties of TGF-β has been studied in EBV-positive patients.
The Baylor group developed an EBV/LMP-specific LST product expressing a dominant negative TGF-beta receptor type II (DNRII).
Calcineurin resistance to enhance EBVSTs
Some groups have explored gene-engineering of virus specific T cells to render them resistant to immune suppressive agents including steroids and calcineurin inhibitors.
In mouse xenograft models bearing human B-cell lymphoma, treatment with CNI-resistant EBVSTs produced enhanced activity in the presence of CNI compared to control EBVS.
Chimeric antigen receptor T cells
(CARTs) have shown impressive efficacy in acute B-lymphoblastic leukemia and nin B-cell lymphomas.
In patients with Epstein-Barr Virus-Lymphoma (EBV-LD, or PTLD), the wider use of CARTs will likely require an off- the-shelf product.
The use of adoptive immunotherapy, particularly using third-party “off the shelf” EBVSTs for the treatment of EBV-LD has shown promise in several studies.
Preclinical work and early clinical trials are in process exploring various gene engineering and combination therapy approaches to improve the potency of these therapies in vitro.
2- What are the indications of adoptive immunotherapy?
1- If conventional tratment fails.
2- It uses EBV-specific cytotoxic T lymphocytes(EBV-CTLs)or donor lymphocyte infusion(DLI)in attempt to about the EBV-driven proliferation of B cells in EBV -association PTLD.
3- Prevention and remmision of EBV-induced PTLD have been achieved in 52 to 75% of pateint .
3- an adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
1- Adoptive cellular treatment using EBVSTs has produced outstanding outcomes in clinical trials for immunocompromised patients with EBV-LD, and there is potential for wider application.
2- The focus of current research is on genetic engineering and combination therapy to increase EBVST potency in vivo.
1. Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD EBV maintains a lifelong latency in B cells and oral epithelial cells.
During primary infection, EBV enters the oropharynx replicating within the epithelial cells and infect B-lymphocytes subsequently it undergoes several steps from lytic, latency III to latency 0.
each step can result in different types of malignancies
· lytic none
· latency III PTLD, HIV lymphoma
· latency II hodgkins lymphoma, DLBCL, diffuse large B-cell lymphoma; NK, T cell lymphoma, nasopharangeal carcinoma
· latency I Burkitt lymphoma, Gastric carcinoma
· latency 0 none
EBV-infected B-lymphocytes in latency III express proteins EBNA1, EBNA2, EBNA3, and EBNALP, LMP1, and LMP2, BARF1. then it enters the lymphoid follicles and downregulate the immunogenic proteins to express less immunogenic proteins and save them into the memory compartment then it continue to further downregulating expression of viral proteins so as to become invisible to EBV-specific T-lymphocytes (EBVSTs).
In individuals with weakened immune systems, such as
(I) patients with primary immunodeficiency (PID) or
(II) infection with human immunodeficiency virus (HIV),
(III) recipients of hematopoietic stem cell transplantation (HSCT) or
(IV) solid organ transplant (SOT),
the lack of a robust EBVST response can lead to uncontrolled proliferation of type 3 latency EBV-infected B cells resulting in
· EBV-associated LPD and · Malignancies
The scientific rationale for adoptive transfer of EBVSTs is based on harnessing the immunogenicity of type III latency malignancies to control the proliferation of the latently infected B cells. 2. What are the indications of adoptive immunotherapy?
Either prophylaxis or treatment of EBV-associated LPD and malignancies in the following populations:
· patients with chronic immunosuppression
· against EBV latency proteins
· as a multi-virus specific product with activity against multiple viruses · patients with primary immunodeficiency (PID) · infection with human immunodeficiency virus (HIV) · recipients of hematopoietic stem cell transplantation (HSCT) · solid organ transplant (SOT) · post-HSCT (33) or SOT (13) and PTLD 3. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients.
refractory PTLD
treatment of multi-VSTs [EBV, cytomegalovirus (CMV), adenovirus, +/− BK virus (BKV) and human herpes virus 6 (HHV6)] products
human B-cell lymphoma
acute B-lymphoblastic leukemia 4. What are the side effects of adoptive immunotherapy?
skin GVHD
TMA
GI hemorrhage
Adoptive cellular immunotherapy for EBV-associated PTLD Introduction:
· EBV associated PTLD is a well described complication after both hematopoietic stem cell (HPSC) and solid organ transplantation (SOT) in about 1-25% of cases.
· The risk of PTLD is dependent on the EBV serostatus of the donor (higher risk with positive donor and negative recipient) and the degree of immunosuppression (higher risk with ATG induction and TAC based maintenance therapy).
· Risk of EBV-PTLD is higher in pediatric SOT (as they are transplanted early before acquiring infection) while > 90 % of adult donors are seropositive.
· EBV is one of herpes virus family and present in >90 % of adult population. It is either asymptomatic if acquired in early childhood or manifests as IMN when acquired in adolescents.
· The degree of viral replication and antigen presentation is maximal during the lytic stage then decrease with time in latency stage as occurs in healthy individuals with intact immune surveillance.
· Usually lytic stage is asymptomatic or causing IMN, while latency phase is associated with risk of LPD.
· After the initial infection and viral replications, it remains dormant in B cell and oral epithelial cells.
· In case of immunodeficient individuals as primary immune deficiency (PID), HIV or SOT: the absence of EBV specific T lymphocytes (EBVSTs) allows the uncontrolled expansion and proliferation of EBV infected B lymphocytes that leads to EBV- lymphoproliferative diseases.
· EBV infection is then associated with many diseases as PTLD, Hodgkin lymphoma, NK and T cell lymphoma, Burkitt’s lymphoma, nasopharyngeal and gastric carcinoma.
· Most of EBV-PTLD (90%) are of B cell origin.
· Rituximab is an effective monotherapy in case of HSCT (50-100 % of cases) and effective in 50 % of SOT cases. In children, combination of rituximab, steroids and cyclophosphamide is effective in treatment but it increases the risk of infections,
1. Summarize principles of adaptive immunotherapy
· Immunotherapy includes:
1. Use of Donor lymphocytes infusion (derived from EBV seropositive donors) was utilized in treatment of PTLD after HSCT. However, it was associated with higher risk of graft versus host disease (GVHD).
2. EBVSTs which are prepared from seropositive donors and cro-preserved for future use.
· Genetic engineering and new methods targeting enhanced capacity of EBVST are investigated to improve outcome of patients with PTLD and EBV related lymphoproliferative disease (EBVLD).
· Methods of preparation of EBVSTs are either Ex-vivo expansion or use of antigen specific T cell selection:
· Ex-vivo expansion of EBVSTs:
o It depends on many steps for good manufacturing-practice GMP-grade EBV-STs.
o Mononuclear cells are isolated from the peripheral blood of donor.
o Then the B-lymphocytes are infected with laboratory strain EBV and transduced with an adenoviral vector expressing latent membrane protein LMP1 and LMP2 and then irradiated.
o Monocytes are separately transduced with the same adenoviral vector and cocultured with T cells followed by a second stimulation by the transduced lymphoblastoid cell lines (LCL) to create and further expand LMP-specific T-cell product.
· Use of antigen specific T cell selection in EBVSTs:
o Using IFN-γ capture approaches where mononuclear cells are pulsed with antigen to isolate CD8+ and CD4+ T cells which secrete IFN-γ in response to the viral antigens.
o Then use of the HLA-restriction requirement to purely select CD8+ T cell product that lacks CD4+T cell (which is necessary for a sustained immune response) and limits the overall applicability of the multimer-selection approach.
· EBVSTs can be either allogenic (T cell are derived from the donor as in case of available living donor) or autologous (used in case of unavailable donor as in deceased organ transplantation or 3rd party (off the shelf) T cell therapy.
· Autologous is more difficult due to immunosuppressed state of transplant recipients. However, it seems more beneficial as mostly PTLD is of recipient origin.
· As regard the outcome of allogenic EBVST immunotherapy in HSCT in either prophylaxis or treatment of EBV-PTLD; it helps in prophylaxis against PTLD and decrease viral load. However, GVHD was a well-known adverse effect. It was also used in SOT and was effective. (mainly for prophylaxis, few cases for treatment of PTLD).
· While autologous EBVST immunotherapy was effective in prophylaxis against PTLD after SOT.
· Type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC required autologous EBVST immunotherapy as an adjunctive therapy to chemotherapy and/or in relapsed patients as they have more restrictive antigen presentation than in type III latency tumors as Burkitt Lymphoma and Gastric Carcinoma.
· 3rd party cell bank (off the shelf T cell therapy):
o Available banked EBVSTs with use of HLA matched (2-5 out of 6) to those with refractory PTLD with reasonable success rate (50-60%.
o The response was better in pediatric population (negative EBV at time of transplantation and has early onset disease).
o It shows reasonable results in BMT and HSCT in children with refractory viral infections (EBV, CMV) and PTLD.
o Its advantage is time saving and early availability without waiting time to access the donor or recipient derived cells.
· The response to EBVSTs can be assessed through measuring the decrease in EBV viral load. However, the PCR is not standardized in all labs (measured in plasma vs whole blood).
· Methods to enhance In-Vivo activity of EBVSTs especially in less antigenic cells of type I and II latency stages:
o Producing TGF-β receptor negative cells to avoid the inhibitory effect of TGF-β secreted by tumor cells.
o Genetic engineering to modify EBVSTs to be CNI and steroid resistant to allow its persistent activity in spite of use of immunosuppressive therapy in SOT.
o Chimeric antigen receptor T cells (CD19) is being investigated in SOT, however nephrotoxicity and neurotoxicity were main side effects.
o Demethylating agents as Azacytidine and decitabine which prevent downregulation of viral antigen expression by EBV-infected B lymphocytes. 2. Indications of its use
o Treatment of PTLD in case of:
§ Refractory cases to standard immunosuppressive medication reduction and chemotherapy as steroids and cyclophosphamide.
§ Advanced and late onset cases.
§ Recurrent cases.
§ EBV +ve cases.
§ Still just tried in case reports and series (few number of cases) and mainly in HSCT not in SOT.
o Treatment of glioblatoma and GIT malignancies.
o Treatment of chronic HIV infection.
o Treatment of autoimmune diseases as type 1 DM and SLE.
3. Can it be used in ttt of other conditions related to SOT
· Ttt of resistant viral infections as CMV and BK.
· Immunomodulation through T reg in treatment of acute rejection. 4. Side effects of adaptive immunotherapy
· DLI is associated with GVHD. However, EBVSTs therapy is not associated with this side effets.
· Generally, may be associated with allergic reactions, GIT upset and flu like symptoms.
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
PTLD is a serious complication occurring after solid organ transplantation, around 95% occur due to activation of EBV viruses acquired in childhood, and 95% of cases are B cell type, T cell lymphoma accounts for 5%
After the primary infection EBV, the virus infect B cells and passes into 2 main stages :
Lytic stage which may be asymptomatic or the patient may develop mild symptoms or frank infectious mononucleosis and
Latency stage in which the virus become latent in B cells life long and at this stage the patient is asymptomatic but at risk of development of certain lymphoproliferative diseases and, several stages of latency develop (Latency III, II, I)
Latency III is a stage in which EBV-infected B-lymphocytes express some protein that comprise the EBV genome such as EBV nuclear antigens EBNA1, 2, 3, EBNALP, LMP1, 2 and BARF1. this high antigenic expression is associated with potent T cell response that limit proliferation of B cells to < 2%. Latency III stage is associated with lymphoproliferative disease and malignancy once the patient is immunosuppressed so the B cell start to develop uncontrolled, unchecked proliferation, the most common form of malignancy associated with this stage is PTLD.
On the other hand, Latency II and I stages are less immunogenic , in which there is down regulation of the immunodominant antigens such as EBNA3 and EBNA2. Latency I and II stages are associated with the development of lymphoproliferative diseases and malignancies in both immune-competent and immune-suppressed patients such Hodgkin’s lymphoma, natural killer /T-cell lymphoma, nasopharyngeal carcinoma (Latency II) and Burkitt lymphoma or gastric carcinoma (Latency I)
The principle of adoptive immunotherapy in treatment of PTLD is using EBV specific cytotoxic T lymphocytes or donor lymphocyte infusion to kill dividing B cells containing EBV. What are the indications of adoptive immunotherapy?
Treatment of resistant and refractory cases of EBV+ PTLD cases
Treatment of certain solid malignancies such as glioblastoma, GIT malignancies
Treatment of certain chronic viral infections such as HIV
Autoimmune disease such as type 1 DM, SLE using Treg adoptive immunotherapy
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
May be used for the treatment of acute rejection through infusing Treg cells
Treatment of resistant cases of CMV, BK nephropathy
What are the side effects of adoptive immunotherapy?
The main side effect of this therapy is the development of graft versus host disease which is less common if using EBV specific cytotoxic T lymphocytes , and very common when using donor derived lymphocytes
Allergic reactions
General flue like symptoms in the form of headache, malaise, fatigue, myalgia, fever
GIT side effects including nausea, vomiting, diarhhia
Introduction:
EBV is the main cause of PTLD.
When the virus enter the oropharynx in primary infection ,it replicate in B-lymphocyte ,which produce different protein comprises wide range of viral genome , EBV nuclear antigens EBNA1, EBNA2, EBNA3, and EBNALP, membrane proteins LMP1, and LMP2 as well as BARF1 and two small non-translated ribonucleic acids (RNA) (Type III latency).
EBV infected B-lymphocytes then downregulate the immunogenic proteins to express less immunogenic type II latency proteins (EBNA1, LMP1 and LMP2) and thus rescue them into the memory compartment where the virus persists in latently infected B-lymphocytes by further downregulating expression of viral proteins so as to become invisible to EBV-specific T-lymphocytes (EBVSTs).
The frequency of EBV-infected B cells in a healthy person remains stable [approximately 0.1–50 EBV infected B lymphocytes per 1,000,000 peripheral blood mononuclear cells (PBMCs)].
EBV-infected B cells, controlled by a potent EBVST response.
Individuals with weakened immune systems like solid organ transplant, the lack of a robust EBVST response can lead to uncontrolled proliferation of type 3 latency EBV-infected B cells resulting in EBV-associated LPD and malignancies.
The transformed B-cells in EBV-associated lymphoproliferative disease associated with latency type III presents several antigenic viral proteins including EBNA 1-3, LMP1 and LMP2 that induce potent EBVST responses. Such potent T cell immunity maintains the infected B cell pool at less than 2%
B cells in immunocompetent individuals but is lacking in immunosuppressed individuals leading to uncontrolled lymph proliferation of the infected B cells. The scientific rationale:
Adoptive transfer of EBVSTs is based on harnessing the immunogenicity of type III latency malignancies to control the proliferation of the latently infected B cells. Manufacturing of EBVSTs:
EBVSTs can be readily produced from EBV-positive donors.
Methods: Ex vivo expansion of EBVSTs:
Where EBVSTs were selectively expanded utilizing irradiated EBV transformed lymphoblastoid cell lines (LCL) as antigen presenting cells (APCs) to selectively expand EBVSTs.
Adenovirus vectors also used to enhance the specificities.
Disadvantage:
Complex.
Time consuming.
APC such as dendritic cell (DC) pulsed with overlapping peptide libraries spanning whole antigen coupled with the use of artificial APCs [e.g., the novel antigen-presenting complex (KATpx)] can facilitate the rapid (10–21 days) expansion. Antigen-specific T cell selection:
Products can be rapidly produced.
(I) major histocompatibility complex (MHC) multimer selection where oligomeric forms of MHC molecules are designed and conjugated to magnetic beads to isolate typically CD8+ T cells with high affinity to a specific viral epitope/peptide in an HLA restricted manner .
(II) IFN-γ capture approaches where mononuclear cells are pulsed with antigen to isolate CD8+ and CD4+ T cells which secrete IFN-γ in response to the viral antigens.
Post-HSCT donor-derived T cell therapy: Donor lymphocyte infusions: Unmodified DLI derived from the patient’s EBV seropositive HSCT donor which contained effector cells with activity against EBV. Donor-derived EBVSTs: Donor derived EBVSTs for the prevention and treatment of EBV+ PTLD in the post-HSCT setting when the donor is available. Autologous EBVSTs
Autologous EBVSTs has been used for patients with EBV-associated malignancies in SOT recipients with PTLD where donors usually are not available because of the use of cadaveric grafts.
Autologous EBVSTs have also been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patient Third-party T cells:
Off the shelf” T cell therapy products. Many bank have being created to reduce need for donors for patient-specific EBVST product (autologous or allogeneic). Modification of EBVSTs to enhance activity: Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo:
EBV/LMP-specific T-cell (LST) product expressing a dominant negative TGF-β receptor type II (DNRII) .
Calcineurin resistance to enhance EBVSTs: EBVSTs were genetically engineered to express a mutant form of calcineurin thus rendering them calcineurin inhibitor (CNI) resistant. Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance EBVST activity in vivo Demethylatingagent:
Pretreatment with decitabine induced expression of LMP1 and ENBA3 associated with latency type 3 which sensitized tumor cells to subsequent therapy with EBVSTs. Conclusions:
The use of adoptive immunotherapy, particularly using third-party “off the shelf” EBVSTs for the treatment of EBV-LD has shown promise in several studies conducted at specialized center. 1. What are the indications of adoptive immunotherapy?
Prophylaxis; treatment of EBV virus-associated lymphoproliferative disease. 1. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Natural killer /T-cell lymphoma, nasopharyngeal carcinoma or type I latency tumors (e.g., Burkitt lymphoma or gastric carcinoma) 1. What are the side effects of adoptive immunotherapy? GVHD. Anaphylactic.
That is an excellent summary and very well structured reply. Under the heading of Introduction, I wish you could type sub-headings in bold or underline or in italics. That would make it easier to read. What is your analysis of level of evidence of this study?
The major complication of EBV positive post transplant is PTLD in HSCT and solid organ transplantation. Less than 25% of HSCT recipient have PTLD and is associated with EBV sero status, degree of T-cell depletion, and immunosupression. Principals;
PTLD is highly immunogenic and responded to immunotherapy with EBVSTs.
EBVSTs can be produced from EBV-positive donors using good manufacturing grade compliant methodology.
The commonly used methods consist of ex vivo expansion of VSTs versus antigen specific.T-cell selection
The major risk factor for developing EBV-associated PTLD post-SOT(solid organ transplant) is EBV
seronegativity at the time of transplant. Since most children are transplanted at a young age while still being EBV seronegative and convert to EBV seropositivity within 2 years of transplant, EBV driven PTLD is much more common in paediatric SOT recipient
Adoptive cellular therapy using EBVSTs (EBV specific T-lymphocyte) has shown impressive results in immunocompromised patients with
Adoptive immunotherapy of autologous or allogeneic EBV-specific CTLs has and will have an important role in establishing scientific principles and might find clinical application in conjunction with conventional therapies
Recent research is focused on strategies to enhance EBVST potency in vivo through genetic engineering as well as combination therapy
. What are the indications for adoptive immunotherapy?
-EBSTs and multi-VSTs presumed by third parties in PTLD
-recipient with immunosuppression and latent EBV infection
To decrease in patients EBV load and prevention of treatment
In refractory and treatment resistant cases of PTLD
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Autoimmune disease
Anti rejection treatment
It can be used as a anti fibrotic therapy Side effect of adoptive therapy
Fever, chills, flue
Kidney injury
Neurological side effect
GVHD
1. Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
rituximab: – anti cd20 – response rate 55-100% in HSCT& around 50% in SOT – in pediatric SOT recipients, a combination regimen using – low dose cyclophosphamide with prednisone and rituximab – has shown event-free survival (EFS) rates of 72%.
Adoptively transferred virus-specific T cells (VSTs): – ranging from the use of donor lymphocyte infusions (DLI) to donor-derived multi-antigen specific VSTs – can be readily produced from EBV-positive donors using good-manufacturing-grade (GMP) compliant methodologies – Ex vivo expansion of EBVSTs – Antigen-specific T-cell selection – Post-HSCT donor-derived T cell therapy ( Donor lymphocyte infusions, Donor-derived EBVSTs ) – Autologous EBVSTs – Third-party T cells modification of EBVSTs to enhance the activity: – Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo – Calcineurin resistance to enhance EBVSTs – Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance EBVST activity in vivo (Demethylating agents, Checkpoint inhibitors, BCL-2 inhibitors)
2. What are the indications of adoptive immunotherapy? – EBV-associated PTLD in HSCT & SOT as prophylaxis to reduce the viral load, induction of remission & decrease relapses 3. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation? Trials showing promising efficacy in: – Auto-immune triggered skin disease(pemphigus) – autoimmune diseases: Lupus, Multiple sclerosis. 4. What are the side effects of adoptive immunotherapy? – Flu-like symptoms: chills, fever – Neurological toxicity: delirium, headache, confusion. – GVHD – AKI – Anaphylactic reaction, palpitation
1. Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD?
EBV infection has different phases from lytic phase (when the virus express all the antigens) to different phases of latency periods which allows virus escaping from immune surveillance. There is associated malignancies with every virus phase, PTLD is associated with latency phase III (LMP1, 2A, 2B virus antigens).
Immunosuppressed population lack of a robust EBV specific T cell (EBVST) response can lead to uncontrolled proliferation of type 3 latency EBV-infected B cells resulting in EBV-associated LPD and malignancies. The scientific rationale for adoptive transfer of EBVSTs is based on harnessing the immunogenicity of type III latency malignancies to control the proliferation of the latently infected B cells. 2. What are the indications of adoptive immunotherapy?
EBV-associated PTLD that persists following initial therapy 3. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
It varies according to the type and target, it includes: acute and chronic graft-versus host disease (GVHD) cytokine storm Neurotoxicity Others: AKI, Arrhythmia, GI upset, neutropenia, infections and constitutional symptoms
This is a systematic review is based on a review of published literature spanning 20 years from 2000 to 2020 in addition to personal experience and data from the authors
EBV is prevalent in adult population by more than 90% infection acquired during childhood where it get attach to B lymphocyte through specific site ( CD21 ) and stimulate the cell protein factory to produce its antigens (3 nuclear antigen and three surface membrane protein) that stimulate Cytotoxic T lymphocyte to control the infection . hence the virus within the infected slave B lymphocyte enter in latent phase inside lymphoid tissue so long the immune system is potent . under any weakness of immune system ( HIV infection or solid organ transplantation SOT) latent antigenic machine inside B lymphocyte will be active and can cause lymphoproliferative disease
So the rule of r adoptive transfer of EBV specific T lymphocytes (EBVSTs) is to control the proliferation of the latently infected B cells
In solid organ recipients, incidence of PTLD ranges from 2% to 25% depending on
– The organ transplanted,
– Passenger lymphocytes in the transplanted organ
– Type of immunosuppression
– EBV seronegativity at the time of transplant
Manufacturing of EBVSTs 1-Ex vivo expansion of EBVST: Disadvantage: long production time 2- Antigen-specific T cell selection
– Advantages: less time consuming
– Disadvantages: difficulties in finding donor with specific cells 3- Autologous EBVSTs indication of adoptive immunotherapy: prophylactic to reduce EBV viral load therapeutic : immunosuppressed recipient with EBV- latent disease causing PTLD Uses of adoptive immunotherapy other than cancer treatment:
Autoimmune disease side effects
GVHD acute kidney injury, Heart : heart arrhythmias, tachycardia, hypotension chills, fever GIT: constipation, diarrhea nausea, vomiting decreased appetite, Chest : cough, cytokine release syndrome (cytokine storm), Neurological : delirium, , dizziness, headache, encephalopathy, fatigue, tremors, Hematology: febrile neutropenia, , hypogammaglobulinemia, bleeding episodes,
That is an excellent summary and very well structured reply. I wish you could type sub-headings in bold or underline or in italics. You have used too much of bold type in one heading. Typing in bold or capitals amounts to shouting! What is your analysis of level of evidence of this study?
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
————————————————————————————————————
PTLD is highly immunogenic and amenable to immunotherapy with EBVST . Adoptively transferred virus-specific T cells (VSTs) have been evaluated for more than two decades, ranging from the use of donor lymphocyte infusions (DLI) to donor derived multi-antigen specific VSTs in the HSCT setting to autologous as well as allogeneic VSTs in the SOT setting .
Donor lymphocyte infusions;
———————————————————
The earliest reported experience of cellular therapy for the treatment of PTLD utilized unmodified DLI derived from the patient’s EBV seropositive HSCT donor which contained effector cells with activity against EBV . Although effective in inducing remissions, this therapy carries a significant risk of graft-versus-host disease (GVHD) .
Donor-derived EBVSTs; ———————————————–
There is extensive reported experience using donor derived EBVSTs for the prevention and treatment of EBV+ PTLD in the post-HSCT setting when the donor is available . This therapy was well tolerated without significant complications with remarkable reductionin EBV viral copy numbers within 4 weeks, including in a patient with immunoblastic lymphoma .
Autologous EBVST; ——————————————
Autologous EBVSTs are preferrable to donor EBVSTs even if available in the post-SOT setting because PTLD is usually of recipient origin and solid organ grafts are not routinely HLA matched to the recipient. However, the challenges in production and the production time leading to delays in initiation of therapy in patients with a rapidly progressive disease impede the routine use of these products in this setting .
Several strategies have been developed to manufacture EBVST products with minimal alloreactivity and broad specificity against EBV latency proteins or as a multi-virus specific product with activity against multiple viruses . The most commonly utilized methods consist of ex vivo expansion of VSTs versus antigen-specific T cell selection [e.g., interferon γ (IFN-γ) capture.
1-Ex vivo expansion of EBVSTs; ——————————————–
Ex vivo expansion of T cells targeting viral antigens via native T cell receptor .
2-Antigen-specific T cell selection; —————————————————–
1-major histococomplex (MHC) multimer selection; where oligomeric forms of MHC molecules are designed and conjugated to magnetic beads to isolate typically CD8 + T cells with high affinity to a specific viral epitope/peptide in an HLA restricted manner .
2- IFN- γ capture approaches; where mononuclear cells are pulsed with antigen to isolate CD8 + and CD4 + T cells which secrete IFN- γ in response to the viral antigens.
Third-party T cells ; ————————————–
It is a readily available “off the shelf” T cell therapy product .The feasibility of third-party cell banks able to cover a majority of the referred patient population had been demonstrated in different reports with impressive response rates .
Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo; —————————————————
TGF-β can be released into the tumor microenvironment by tumor cells, fibroblasts and immune cells and creates an immunosuppressive environment by impeding T-cell activation, proliferation and migration. In addition, it affects DC and macrophage antigen presentation and chemo taxis .
Calcineurin resistance to enhance; ——————————————————
Given the concerns regarding EBVST persistence in patients who require ongoing immune suppression , several groups have explored gene engineering of virus specific T cells to render them resistant to immune suppressive agents including steroids and calcineurin inhibitor .
Chimeric antigen receptor T cells ; ——————————————————-
CD19-chimeric antigen T cells (CD19-CART) have shown impressive efficacy in acute B-lymphoblastic leukemia and in B-cell lymphomas . In addition to viral antigens, EBV-LD expresses a variety of B-cell antigens targetable by CARTs including CD19, CD20 and CD30. However, their use in EBV-lymphoproliferation is limited by the patient’s immunosuppressive state impeding T cell manufacture and the length of production time.
.
Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance EBVST activity in vivo; ———————————————————————— 1-Demethylating agent;
Newly EBV-infected B-cells express up to 90 viral genes; however rapid CpG methylation of viral antigens leads to downregulation of viral protein expression and the latency . Azacytidine and decitabine are potent CpG demethylating agent .
2-Checkpoint inhibitors ;
LMP1 has been shown to induce expression of the checkpoint protein PD-L1 in classic HL (CHL) without 9p24.1 alteration . PD-L1 is expressed in 73% of EBV-positive PTLD . . There have been several trials using checkpoint blockade in DLBCL hinting at single agent activity . A phase I study at BCM is exploring combination therapy of checkpoint inhibitors with EBV directed T cells for EBV + HL and NHL .
3-BCL-2 inhibitotors;
In preclinical, in vitro studies, pretreatment of malignant B-cell lines with the BCL-2 inhibitor venetoclax increased proapoptotic proteins and sensitivity to CD19.
2-What are the indications of adoptive immunotherapy?
——————————————————————————————–
1- Inducing remission of PTLD .
2- Prevention and treatment of EBV+ PTLD in the post-HSC.
3-Treatment of type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients .
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
—————————————————————————
Autologous EBVSTs have also been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients .
What are the side effects of adoptive immunotherapy?
———————————————————————————
1-The risk of graft-versus-host disease (GVHD)
2-The risk of graft rejection and autoimmunity .
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
The major risk factor for developing EBV-associated PTLD post-SOT is EBV seronegativity at the time of transplant
More than 90% of EBV-associated PTLD is of B-cell origin with cell surface expression of CD20
PTLD is highly immunogenic and adjustable to immunotherapy with EBVSTs
Donor lymphocyte infusion or EBV-specific T-lymphocytes (EBVSTs) in specialized centers
The aims of this study are to define current best manufacturing strategies for EBV-specific VSTs, clinical experience of their use, discuss opportunities to broaden the applicability of this approach and to explore future strategies to enhance their efficacy
Manufacturing of EBVSTs
Produced from EBV-positive donors using good-manufacturing-grade (GMP) compliant methodologies
Donor type source: autologous and allogeneic (including third party)
Two methods: ex vivo expansion of VSTs and antigen-specific T cell selection Ex vivo expansion of EBVSTs:complex and time consuming. Can be manufactured from individuals irrespective of their HLA type. Can be produced from EBV seronegative donors Antigen-specific T cell selection [interferon γ (IFN-γ)]: EBVST products can be rapidly produced (in 48 hours) from EBV seropositive autologous and allogeneic donors
Donor-derived T cell therapy (Post-HSCT) Donor lymphocyte infusions: produced from EBV seropositive donor. Effective but high risk of GVHD Donor-derived EBVSTs: safe and rare GVHD
Autologous EBVSTs
Technically challenging (ongoing immunosuppression)
Used in case of no available donor (cadaveric grafts). Effective and rare PTLD
Third-party T cells
“Off the shelf” T cell therapy product. Mostly used in the post-transplant setting
Modification of EBVSTs to enhance activity
1. Overcoming the immune suppressive effects of TGF-β
2. Calcineurin resistance
3. Chimeric antigen receptor T cells
4. combination strategies administering EBVSTs with other therapeutic modalities
(a) Demethylating agents
(b) Checkpoint inhibitors
(c) BCL-2 inhibitors
Conclusions
Use of adoptive immunotherapy for the treatment of EBV-LD is promising in several studies (EBVSTs
Is now widely available)
What are the indications of adoptive immunotherapy?
Patients with EBV-associated PTLD who fail conventional treatments or cannot tolerate it (RI, rituximab, and chemotherapy)
Treatment of PTLD with EBV-specific CTLs should be considered where available with refractory or relapsed (R/R) EBV-positive PTLD (1C)]
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
CMV, BK virus, and rejection What are the side effects of adoptive immunotherapy?
Vary according to the type of adoptive immunotherapy, location of cancer, type of cancer, and patient overall health
In most cases side effects can be managed safely when recognized early
Overactive immune response (cytokine release syndrome), and neurotoxicity, AKI, bleeding disorders, arrhythmias, chills, GIT symptoms, febrile neutropenia, and hypotension
1. The principle of adoptive immunotherapy in PTLD treatment:
It involves generating EBV-specific T lymphocytes (EBVSTs) from EBV-positive autologous or allogenic donors with limited alloreactivity and wide virus-fighting activity.
It is necessary to use ex vivo virus-specific T cell growth or antigen-specific T cell selection.
MHC selection involves isolating CD8+ T cells with high affinity to a specific viral epitope or peptide in an HLA-restricted manner; IFN- capturing involves isolating CD8+ and CD4+ T cells that release IFN- in response to viral antigens.
Technically challenging approaches.
Post-HSCT donor-derived T cells
Donor-lymphocyte infusions
Unmodified DLI from PTLD patients’ EBV-positive HSCT donors induces remission.
Donor-derived EBVSTs may prevent and treat EBV+ PTLD in post-HSCT patients without reactivation or GVHD by lowering EBV burden.
EBV-transformed lymphoblastoid cell lines (LCLs) are used to selectively grow EBV-specific T lymphocytes as antigen-presenting cells.
DC and LCLs were transduced with adenovirus vectors to overexpress LMP1 or LMP2 to increase specificity for less immunogenic EBV antigens, but it is difficult and wastes time.
2. What are the indications for adoptive immunotherapy?
-EBSTs and multi-VSTs provided by third parties in PTLD
-recipient with immunosuppression and latent EBV infection
-Treatment for prevention and a decrease in the patient’s EBV load
-Treatment resistant and refractory cases of PTLD
3-Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
T-cell adoptive immunotherapy for BK nephropathy in renal transplantation
The treatment of lymphoproliferative diseases connected to viral (EBV) loads
4—What are the side effects of adoptive immunotherapy?
1. Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD.
-EBV-positive post-transplant lymphoproliferative disease (PTLD) is a major complication of hematopoietic stem cell and solid organ transplantation,
– The biggest risk factor for developing EBV-associated PTLD post-SOT is EBV seronegativity at the time of transplant. Since most children are transplanted at a young age while still being EBV seronegative and convert to EBV seropositivity within 2 years of transplant, EBV driven PTLD is much more common in pediatric SOT recipients,
-Adoptive cellular therapy using EBVSTs has shown impressive results in immunocompromised patients with EBV-LD and there is the potential for wider use.
– Adoptive immunotherapy uses EBV-specific cytotoxic T lymphocytes (EBV-CTLs) (autologous or allogeneic) or donor lymphocyte infusion (DLI) in an attempt to kill dividing B cells in EBV-associated PTLD,
–“off the shelf” T cell therapy product is desirable. Because donor cells may not be available (e.g., recipients of umbilical cord blood transplants or cadaveric organ transplants). Hence, manufacture of patient-specific products may not be possible or may be so delayed that patients with rapidly progressive disease are not able to access these therapies,
-Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance EBVST activity in vivo,
-Demethylating agents
-Checkpoint inhibitors
-BCL-2 inhibitors
2- What are the indications of adoptive immunotherapy? –Allogenic virus-specific T cells in PTLD in HSCT recipients, -Autologous EBVSTs for EBV PTLD in SOT recipients, -Third-party EBSTs and multi-VSTs in PTLD,
-Type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients. Such tumors are more challenging targets because of the reduced expression of immunogenic viral antigens in these type II latency tumors which express a more restricted array of antigens compared to type III.
3- Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
-Autologous EBVSTs have also been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients.
4- What are the side effects of adoptive immunotherapy? -The major complication of adoptive immunotherapy is acute and chronic graft-versus-host disease (GVHD) . DLI may produce GVHD, which does not appear to be a problem with EBV-CTLs. -Impaired Graft Function -CRS
– EBV-infected B-lymphocytes pass through out many phases: lytic phase with high antigen expression, Latency III with less expression, Latency II, Latency I and Latency 0 with no anitigen expression (dormant infection) and become invisible to specific T cells.
-Impaired immunity—replication and proliferation of B cells and cause
EBV- malignancies which are:
1- type III latency: PTLD
2- Type II latency : HL, natural killer (NK)/T-cell lymphoma, nasopharyngeal
carcinoma (NPC)
3- type I latency: Burkitt lymphoma or gastric carcinoma
presence of potent T-Cell immunity maintain proliferating infected B cell <2%
– So, he rational for adoptive immunotherapy is to attack the proliferating B-cells with EBV-specific T cells.
2- Indications of adoptive immunotherapy:
EBV-lymph proliferative diseases either post transplantation (HSCT or SOT) or not.
3- Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation: prophylaxis against EBV-related PTLD
4- side effects: Graft versus host disease impaired graft function non-response
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
It’s mainly for eradication of Epstein-Barr virus (EBV), to prevent risk of lymphoma and lymph proliferative disorders in solid organ transplant.
The incidence of PTLD is 1-25% and mainly express CD20 on the surface of B cell. Only 10% is T cell and responded well to rituximab in case of HSCT but it’s limited in use because risk of infection and response low in solid organ transplant. Post-HSCT donor-derived T cell therapy
Donor lymphocyte infusions:
Enhance remission by eradicate of EBV but carrying high risk of GVHD, However it’s limited by depletion of T cell regulators before infusion.
Donor-derived EBVSTs:
prevent and treat of EBV related PTLD; with no risk of GVHD.
Autologous EBVSTs:
It’s used to treat type II latency EBV-associated malignancies including Hodgkin’s lymphoma and natural killer lymphoma, with complete remission but not reduce viral load of EBV.
Third-party T cells:
This therapy use to treatment of lymphoproliferative disorder related to refractory infection like EBV and CMV and adenovirus by reducing viral load.
It’s has good effects.
Modification of EBVSTs to enhance activity
Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo:
Calcineurin resistance to enhance EBVSTs
Chimeric antigen receptor T cells Combination therapy of EBVSTs with other therapy like
Demethylating agents
Checkpoint inhibitors
BCL-2 inhibitors
What are the indications of adoptive immunotherapy?
Treatment of lymph proliferation disease related to viral load especially EBV
Prevention and reduce virama of EBV
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Prophylactic and treat viral load
Autoimmune disease
What are the side effects of adoptive immunotherapy?
Adoptive cellular immunotherapy for Epstein-Barr virus-associated lymphoproliferative disease. Summary of the article: Introduction: EBV, the worldwide common viral infection in childhood, affects 90% of the adult population. In healthy individuals, the immune system analyses the virus and protects against spreading infection, although rare cases can go into fulminant disease. EBV goes into a dormant phase latent in the B cells. In contrast to healthy individuals, the patients with the defective immune system, include:
HIV.
HSCT.
SOT.
can lead to uncontrolled infection and proliferation which can lead to EBV- associated LPD or malignancies. The transformed B-cells in EBV-associated LD, associated with latency type III, present Viral proteins:
EBNA, 1-3.
LMP1.
LMP2.
such antigens in intact immune systems, T-cells can control B cells and maintain infected B cells < 2%. Type II latency associated with malignancies:
HL, Hodgkins’s lymphoma.
Natural killer T-cell lymphoma.
Nasopharyngeal carcinoma, NPC.
Type I latency tumors:
Burkitt’s lymphoma.
Gastric carcinoma.
The occurrence of PTLD in HSCT ranges from 1-25%, depending on the following:
Serostatus of the donor.
The degree of patient T- cell depletion.
Post-HSCT immunosuppressants.
Sero negativity is the top determinant factor of the development of post-transplant LPD. Since pediatric patients are being still seronegative as they transplant earlier at a younger age, PTLDs are more common in pediatric patients. Monotherapy with Rituximab anti-CD20 is more effective in HSCT with a response rate of 55 -100%, while combination therapy (prednisolone/cyclophosphamide/rituximab) is shown to better survival up to 725. #EBVSTs manufacturing:
Ex vivo expansion of EBVSTs.
Antigen-specific T cell selection.
#Post-HSCT donor-derived T-cell therapy:
Donor Lymphocyte infusion.
Donor-derived EBVSTs.
# Autologous EBVSTs. # Third-party T cells.
Modification of EBVSTs to enhance activity:
Overcoming the immune suppressive effects of TGF-B to enhance EBVST activity in vivo.
Calcineurin resistance to enhance EBVSTs.
Chimeric antigen receptor T cells.
Combination strategy, with EBVSTs:
Demethylating agents.
Checkpoint inhibitors.
BCL-2 inhibitors.
Conclusion:
Promised result with treatment with adoptive immunotherapy, using third-party EBVSTs for the treatment of EBV-LD.
EBVSTs become widely available, however, more effort is needed to make third-party cell banks broadly available and easily applicable.
Indication of adoptive immunotherapy:
Immunocompromised recipient with EBV-LD.
Prophylactic treatment and reduction of EBV load.
Uses of adoptive immunotherapy other than cancer treatment:
Auto-immune triggered skin disease.
Other autoimmune conditions; Lupus, Type 1 DM, Multiple sclerosis.
Inflammatory diseases; atherosclerosis and hardening of arteries.
1- The principal of adoptive immunotherapy in PTLD treatment EBVSTs Manufacture
It includes producing EBV specific T lymphocytes (EBVSTs) from EBV positive donors either autologous or allogenic donors with minimal alloreactivity and broad spectrum activity against multiple viruses.
The methodology involve either ex vivo expansion of virus specific T cells or antigen-specific
T cell selection. Ex vivo expansion of EBVST
By using irradiated EBV transformed lymphoblastoid cell lines(LCLs) expressing high levels of class I and class II HLA and co-stimulatory molecules acting as antigen presenting cells to selectively expand EBV specific T lymphocytes.
DC and LCLs were transduced with adenovirus vectors to overexpress LMP1 or LMP2 to strengthen the specificity to less immunogenic EBV antigens but in fact it is difficult and wastes time on the other hand it enhances rapid increase of T cells targeting EBNA1, LMP1 and LMP2 from healthy donors and from cases with type 2 latency EBV-associated malignancies, also EBVTs can be derived from EBV seronegative donors. Antigen specific T cell selection
-MHC selection by isolating CD8+T cells having high affinity to a specific viral epitope/peptide in an HLA restricted manner
– IFN-γ capturing by isolating CD8+and CD4+ T cells that secrete IFN-γ in response to the viral antigens
Those methods are difficult technically. Post-HSCT donor-derived T cell therapy Donor lymphocyte infusions
Used in the treatment of PTLD by unmodified DLI taken from patient’s EBV seropositive HSCT donor having activity against EBV, inducing remission.
Donor-derived EBVSTs
Using ex vivo expanded EBVSTs derived from seropositive donors for the prevention and treatment of EBV+ PTLD in the post-HSCT efficiently without reactivation nor occurrence of GVHD , through decreasing EBV load in cases with reactivation or high risk for reactivation.
Autologous EBVSTs
It was used for treatment of PLTD in SOT patients from cadavers ,which is challenging due to the immunosuppression and that can delay starting therapy.
EBVST produced by LCL stimulation consist of T cells specific to early lytic antigens and the immunodominant EBNA3 antigens but less active towards viral antigens expressed
in latency type II.
Meanwhile in a trial latent membrane protein (LMP)-specific T cells were used as adjuvant therapy with acceptable outcomes with type II latency malignancies Third party T cells
Donor cells may not be available therefore off the shelf T cells will be needed.
33 transplant recipients of different organs ,having refractory PTLD were given partially HLA matched EBVSTs and the cases with the higher HLA match had a better response with less rejection and GVHD risk.
Third-party multi-VSTs use for EBV-associated PTLD had acceptable outcomes.
Third party EBV gave promising results for CNS PLTD cases EBVSTs modification to increase efficiency
-by inhibiting TGFB immunosuppressive activity
In EBV positive HL ,EBV/LMP-specific T-cell (LST) product expressing a dominant negative TGF-β receptor type II (DNRII) were resistant to inhibitory concentrations of TGF-β produced by tumor cells with favourable response – Calcineurin resistance to enhance EBVSTs
Gene engineering of virus specific T cells to resist immune suppressives
– Chimeric antigen receptor T cells –Combining EBVSTs with other therapies as
· Demethylating agents that can sensitize tumor cells to EBVST
· EBV-positive diffuse large B-cell lymphoma (DLBCL) and 73% of EBV-positive PTLD express PD-L1 but combining checkpoint inhibitors with EBV directed T cells for EBV+ PTLD had side effects
· BCL-2 inhibitors ,as BCL-2 is an antiapoptotic protein for latent EBV infected cells upregulated by LMP1 protein.
Combing BCL-2 inhibitors with CD 19 CART for treatment of EBV lymphoproliferative is under trial
2-Adoptive therapy indications are EBV positive PLTD in HSCT and in SOT recipients, EBV-associated malignancies as HL, T/NK lymphoma and NPC and in BMT recipeints infected with EBV,CMV ,HHV6,and BKV
3- Autologous EBVSTswas involved in treat of type II latency EBV-associated malignancies as HL, T/NK lymphoma and NPC with chemotherapy and/or in relapsed patients.
–Multi-VSTs [EBV, cytomegalovirus (CMV), adenovirus, +/− BK virus (BKV) and human herpes virus 6 (HHV6)] products were given to HSCT recipients having refractory viral disease after BMT targeting EBV, CMV and/or adenovirus Third party EBV was given to post-HSCT or SOT cases ,no published data on it’s use in HIV-associated lymphoma EBV/LMP-specific T-cell (LST) use in EBV positive HL CNI-resistant EBVST for human B cell lymphoma bared by a mouse (under trial) CD19-chimeric antigen T cells in acute B-lymphoblastic leukemia and in B-cell lymphomas EBVSTs with demethylating agent in Burkitt lymphoma Combining checkpoint inhibitors with EBV directed T cells for EBV+ HL and NHL under trial
4- Side effects of adoptive immunotherapy
-DLI can lead to GVHD through selective depletion of T reg, while depleting
naïve T cells can decrease the GVHD risk
-Multi VST can lead to TMA and GI hemorrhage
– CD19-chimeric antigen T cells (CD19-CART) caused cytokine release syndrome (CRS), neurotoxicity and acute kidney injury.
– Combining checkpoint inhibitors with EBV directed T cells for EBV+PTLD lead to graft rejection and autoimmune complications.
EBV-specific T-lymphocytes (EBVSTs): EBVSTs were selectively expanded utilizing irradiated EBV transformed lyphmoblastoid cell lines (LCL) as APC to selectively expand EBVSTs. LCL-activated EBVSTs work against early lytic antigens and viral proteins EBNA 3A,3B, &3C and may not be active against LMP1 and LMP2 (their over expression is engineered with the use of adenovirus vectors).
Donor lymphocyte infusions: This is a form of cellular therapy which utilized un-modified DLI obtained from the patient’s EBV seropositive HSCT donor, which contained effector cells against EBV
Combination therapy other therapeutic modalities to enhance EBVST activity in vivo: e.g., demethylating agents, check point inhibitors, & BCL-2 inhibitors
2.Indications of adoptive immunotherapy: Prophylaxis and treatment
Allogenic virus-specific T cells in PTLD in HSCT recipients
Autologous EBVSTs for EBV PTLD in SOT recipients
Third-party EBSTs and multi-VSTs in PTLD
3.Can adoptive immunotherapy be used in the treatment of other conditions?
In clinical trials, adoptive cellular therapy using EBVSTs has shown impressive results in immunocompromised patients with EBV-LD and there is the potential for wider use. Current research is focused on strategies to enhance EBVST potency in vivo through genetic engineering as well as combination therapies.
4.What are the side effects of adoptive immunotherapy?
There is a significant risk of graft-versus host disease (GVHD) with donor lymphocyte infusions therapy. This risk is not seen with EBV specific T-lymphocytes (EBVSTs)
1- Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD:-
====================================================================
Introduction
====================================================================
Manufacturing of EBVSTs
====================================================================
Ex vivo expansion of EBVSTs
=================================================================
Antigen-specific T cell selection
====================================================================
Post-HSCT donor-derived T cell therapy
Donor lymphocyte infusions
Donor-derived EBVSTs
Autologous EBVSTs
Third-party T cells
====================================================================
Modification of EBVSTs to enhance activ
Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo
Calcineurin resistance to enhance EBVSTs
Chimeric antigen receptor T cells
====================================================================
Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance EBVST activity in vivo
Demethylating agents
Checkpoint inhibitors
BCL-2 inhibitors
====================================================================
Conclusions
====================================================================
2- What are the indications of adoptive immunotherapy?
1- If conventional tratment fails.
2- It uses EBV-specific cytotoxic T lymphocytes(EBV-CTLs)or donor lymphocyte infusion(DLI)in attempt to about the EBV-driven proliferation of B cells in EBV -association PTLD.
3- Prevention and remmision of EBV-induced PTLD have been achieved in 52 to 75% of pateint .
====================================================================
3- an adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
1- Adoptive cellular treatment using EBVSTs has produced outstanding outcomes in clinical trials for immunocompromised patients with EBV-LD, and there is potential for wider application.
2- The focus of current research is on genetic engineering and combination therapy to increase EBVST potency in vivo.
====================================================================
4- What are the side effects of adoptive immunotherapy?
1- Acute and chronic graft -versus-host disease(GVHD).
2- DLI may produce GVHD,which dose not appear to be problem with EBV-CTLs.
EBV is a virus related to lymphoproliferative disorders, it infects 90% of adults . Primary infection is mainly asymptomatic or may lead to infectious mononucleosis.
In healthy seropositive individuals, virus neutralizing antibodies control the spread of infectious virus particles and EBV specific HlA class I restricted CD8 cytotoxic T lymphocyte specific to the early lytic cycle proteins kill cells entering the lytic cycle before they are able to release infectious viral particles.
In immunocompetent individuals, the frequency of EBV infected B cells remain low and stable by a potent EBVST ( EBV specific T lymphocytes ) response.
In immunocompromised ones, the lack of apotent EBVS..T. response can lead to uncontrolled proliferation of type 3 latency EBV infected B cells lead to EBV associated LPD.
The rationale of adoptive transfer of EBVSTs is based on harnessing the immunogenicity of type III latency malignancy to control the proliferation of the latent infected B cells
PTLD is highly immunogenic and amenable to immunotherapy with EBVSTs
Manufacturing EBVSTs
Methods
1. Ex vivo expansion of EBVSTs
2. Antigen specific T cell selection
Post HSCT donor derived T cell therapy
1. Donor lymphocyte infusi ons
2. Donor derived EBVSTs
3. Autologous EBVSTs
4. Third party T cell
Modification of EBVSTs to enhance activity
1. Overcoming the immune suppressi effects of TGF-B to enhance EBVSTs activity in vivo
2. Calcineurin resistance to enhance EBVSTs activity
3. Chimeric antigen receptor T cells
Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance EBVSTs activity in vivo
. Demethylating agents
. Checkpoint inhibitors
. BCL-2 inhibitors
Manufacturing of EBVSTs
●EBVSTs can be readily produced from EBV-positive donors
●Donor types include autologous and allogeneic
●The most commonly utilized methods consist of ex vivo expansion of VSTs versus antigen-specific T cell selection [e.g., interferon γ (IFN-γ) capture]
Ex vivo expansion of EBVSTs
○ EBVSTs were selectively expanded utilizing irradiated EBV transformed (LCL) (that express a type III latency pattern of EBV antigen expression) as (APCs) to selectively expand EBVSTs
○LCL-activated EBVSTs consist of a product with activity against early lytic antigens and EBNA 3A, 3B and 3C but unreliable activity towards LMP1 and LMP2
○To enhance the specificities to LMP1 and LMP2, several groups have made further modifications to this approach by transducing DC and LCLs with adenovirus vectors to overexpress LMP1 or LMP2
○ this approach is complex and time consuming
○Subsequently the use of APC such as (DC) can facilitate the rapid (10–21 days as opposed to 2–3 months).
○A further benefit of ex vivo expansion approaches that utilize whole antigen, is that EBVSTs can be manufactured from individuals irrespective of their HLA type.
Antigen-specific T cell selection Other rapid methods include:
(I) (MHC)
(II) IFN-γ capture approaches
They require seropositive donors with high frequency of circulating antigen/epitope specific T cells which may be technically challenging in the autologous setting.
Post-HSCT donor-derived T cell therapy
Donor lymphocyte infusions
●This therapy carries a significant risk of (GVHD)
●Donor-derived EBVSTs
For the prevention and treatment of EBV+ PTLD in the post-HSCT setting
● This therapy was well tolerated without significant complications with remarkable reduction in EBV viral copy numbers within 4 weeks, including in a patient with immunoblastic lymphoma.
●This pivotal study established the safety and early evidence of efficacy of donor-derived EBVSTs for the treatment and prophylaxis of PTLD in transplant recipients
● GVHD rates in this study were low
Studies using allogeneic VSTs in EBV+ PTLD in HSCT recipients
□BCM, Houston, TX, USA
□MSKCC, New York,
□Karolinska Institute, Stockholm, Sweden
□Children’s Research Hospital, Kyoto, Japan (47)
Autologous EBVSTs
◇ It is used outside the context of allogeneic HSCT and in SOT recipients with PTLD where donors usually are not available because of the use of cadaveric grafts.
◇It is technically more challenging because of the ongoing immunosuppression but this can be overcome with modern manufacturing approaches
◇In theory, autologous EBVSTs are preferrable to donor EBVSTs even if available in the post-SOT setting because PTLD is usually of recipient origin and solid organ grafts are not routinely HLA matched to the recipient.
◇the challenges
the production time that will lead to delays in initiation of therapy in patients with a rapidly progressive disease
◇Autologous EBVSTs have also been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients.
◇Such tumors are more challenging targets because of the reduced expression of immunogenic viral antigens in these type II latency tumors which express a more restricted array of antigens (e.g., LMP1, LMP2, EBNA1 and BARF1) compared to type III latency tumors.
Third-party T cells
●It needs a prolonged time
Donor cells may not be available (e.g., recipients of umbilical cord blood transplants or cadaveric organ transplants
●For the above reasons, a readily available “off the shelf” T cell therapy product is desirable.
The first third-party EBVST bank of 60 EBVST products was established by Haque et al. in the United Kingdom
no significant toxicities were observed,
●The UK group established a cell bank of 25 donors with HLA alleles prevalent at high frequencies in individuals of European descent
●Chiou et al. from Birmingham, United Kingdom published their experience in 10 pediatric SOT recipients with PTLD and reported an ORR of 80% (8 out of 10)
●This favorable response in a pediatric population may indicate differences in the biology of EBV-driven PTLD in this population who is often EBV naïve at the time of transplant and develops PTLD in the earlier post-transplant period compared to the adult population.
Conclusions .
Further work is however needed to create widely available and commercialized third-party cell banks .
Strategies need to be explored to enhance the anti-tumor activity of EBVST therapies especially for the less immunogenic type I and II latency tumors.
Preclinical work and early clinical trials are in process exploring various gene engineering and combination therapy approaches to improve the potency of EBVSTs in vivo.
What are the indications of adoptive immunotherapy?
PTLD (SOT- HSCT)
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
HHV6- HCV – HBC –
The most important side effect is GVHD
The GMP (good manufacturing-practice) grade EBV-specific T-cell manufacturing involve mononuclear cells which are harvested from the peripheral blood of a donor. B-lymphocytes are infected with laboratory strain EBV and transduced with an adenoviral vector expressing latent membrane proteins: LMP1 and LMP2 and irradiated. Monocytes are separately transduced with the same adenoviral vector and cocultured with T cells followed by a second stimulation by the transduced lymphoblastoid cell lines to create and further expand LMP-specific T-cell product. Methods are:
1-Ex vivo expansion of T cells targeting viral antigens via native T cell receptors was established initially by Smith et al. where EBVSTs were selectively expanded utilizing irradiated EBV transformed lymphoblastoid cell lines as antigen presenting cells to selectively expand EBVSTs.
2-major histocompatibility complex (MHC) multimer selection where oligomeric forms of MHC molecules are designed and conjugated to magnetic beads to isolate typically CD8+ T cells with high affinity to a specific viral epitope/peptide in an HLA restricted manner.
3- IFN-γ capture approaches where mononuclear cells are pulsed with antigen [e.g., single peptides, overlapping peptides (pepmixes), to isolate CD8+ and CD4+ T cells which secrete IFN-γ in response to the viral antigens.
Indications:
prevention and treatment of EBV+ PTLD in the post-HSCT
EBV-associated malignancies outside the context of allogeneic HSCT and in SOT recipients with PTLD
Autologous EBVSTs have also been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients
Side effects
1- graft-versus-host disease
2- graft versus lymphoma
Q1: EBV is a member of herpes viruses causing a primary disease such as asymptomatic or infectious mononucleosis. After primary infection, EBV remains in B cells and oral epithelial cells as latent viruses.
Viral antigens like EBNA 1-3, and LMP 1-2, are expressed during type III latency. After entrance to lymphoid follicles, infected B cells downregulate the expression of these proteins, so that not to recognize by EBV-specific T cells (a kind of cytotoxic T cells or, EBVSTs). In immunocompetent persons, these infected B cells are controlled by EBVSTs response lifelong. However, in immunosuppressed transplanted recipients this response is lacking and results in excessive proliferation of infected cells and lymphoproliferative disease (LPD). seronegative children before TX are at the highest risk for these LPDS and in more than 90% of cases their origin is CD20-positive B cells. In HSCT, the vast majority respond to rituximab, but in SOT response rate is about 50%. Another regime including cyclophosphamide with prednisone and rituximab is effective but has a low tolerance. Hence, there is need for another treatment. In the past donor lymphocyte infusions (DLI) were used in HSCT or allogenic virus-specific T cells (VSTs) for SOT, but nowadays manufacturing EBVSTs are produced by GMP methodologies against EBV latency proteins. There are two methods:
1. Ex vivo production of EBVSTs
2. Antigon-specific
T cell selection by MHC multimer selection or IFN-Ɣ capture methods to isolate CD8+ and CD4+ cells selecting IFN-Ɣ due to viral Ags.
DLC caused a significant risk of GVHD. EBVSTs could be donor-derived or autologous or third-party.
Donor-derived EBVSTs have been used with success in HSCTs. Autologous EBVSTs have been used in SOT recipients for prevention and treatment with successful results in type III latency and acceptable results in type II latency as adjustment therapy which has lower immunogenicity. third-party EBVSTs, therapy, or even covering CMV, adenovirus, BKV, and HHV6 have produced with acceptable results. Bank of these cells is providing and reduces EBV viral load in these patients. Other productions are available. One of them uses DNPII to overcome the immunosuppressive effect of TC-F-β with downregulation of TGF-β receptors with promising results. Another way was gene engineering of VST to resist them against CNIs and steroids with good results in mice.
CD19- chimeric antigen T cell (CD19-CART) is another way but due to cytokine release syndrome and other adverse reactions, is not routinely used. Other combinations of EBVST with other modalities have been used such as demethylating agents to express more EBV antigens or checkpoint inhibitors. To inhibit PD-L1 in classic HL and NHL combined with EBVSTs. In addition, antiapoptotic proteins like BCL-2 are upregulated by LMP1 EBV antigens. Therefore, pretreatment with BCL-2 inhibitor combined with CD19-CART is studied. Hence, these investigations will increase the use of EBVST in the future.
Q2:
1. Prevention or treatment of PTLD in HSCT or SOT
2. Treatment of resistant viral infections
3. In combination with other treatments for resistant cases of PTLD or even other malignancies
Q3: EBVST could be used for the treatment of a wide variety of viral infections and malignancies, alone or combined with other methods.
Q4:
1. Acute and chronic GVHD especially with DLL
2. Cytokine release syndrome in CD19-CART
3. Other side effects such as itching, skin rash, allergic reactions, nausea, vomiting
Level 5 evidence
Introduction:
EBV infection usually occurs in childhood as mild clinical presentation and maintains a lifelong latency in B cells .
The adoptive immunotherapy in the treatment of PTLD in SOT has demonstrated a better clinical outcome with less toxicity .
Adoptive immunosuppressive Side effects :Graft versus host disease ,Ig-E-mediated anaphylaxis reaction ,Flu-like symptoms.
In what circumstances does adoptive immunotherapy make sense to use?
Is there a possibility that adoptive immunotherapy could be used to treat other illnesses that are associated with organ transplantation?
What sorts of adverse reactions are associated with adoptive immunotherapy?
Level of evidence is 5
The adoptive cellular immunotherapy for Epstein-Barr virus-associated lymphoproliferative disease (EBV-LD) are going in advance and more widely available including in industry led multi-centre studies and consortium and cooperative group studies. EBV-positive post-transplant lymphoproliferative disease (PTLD) is a major complication of hematopoietic stem cell and solid organ transplantation.
First successes with adoptive immunotherapy were described using donor lymphocyte infusion for the treatment of EBV+ PTLD in hematopoietic stem cell transplantation (HSCT) recipients but can lead to high risk of graft-versus-host disease (GVHD).
EBV Specific T lymphocytes( EBVSTs) can be readily produced from EBV-positive
donors using good-manufacturing-grade (GMP) compliant methodologies . Donor types include autologous and allogeneic (including third party) sources.
Over the years, several strategies have been developed to manufacture EBVST products with minimal alloreactivity and broad specificity against EBV latency proteins or as a
multi-virus specific product with activity against multiple viruses . The most commonly utilized methods consist of ex vivo expansion of VSTs versus antigen-specific T cell selection.
Indications of adoptive immunotherapy include:
1- EBV associated malignancies like PTLD
2- EBV severe infection and complications in immuno-compromised patients
3- Refractory PTLD post organ transplantation
The main side effect is graft versus host disease (GVHD)
The article discusses the Epstein-Barr virus (EBV), a member of the gamma herpes virus family, and its association with various lymphomas and lymphoproliferative disorders (LPD). EBV infects a large percentage of the adult population worldwide and primarily causes asymptomatic or mild infections in childhood. In individuals with a healthy immune system, virus-neutralizing antibodies and specific cytotoxic T lymphocytes (CTLs) control the virus’s spread and kill infected cells. EBV establishes lifelong latency in B cells and oral epithelial cells.
During primary infection, EBV replicates in the oropharynx’s epithelial cells and infects B lymphocytes. Infected B cells express a range of viral proteins, enter lymphoid follicles, downregulate immunogenic proteins, and persist as latent infections in memory B cells. In healthy individuals, the frequency of EBV-infected B cells remains controlled by an effective EBV-specific T-lymphocyte response. However, in individuals with weakened immune systems, such as those with primary immunodeficiency or HIV infection, or recipients of stem cell or solid organ transplants, uncontrolled proliferation of EBV-infected B cells can occur, leading to EBV-associated LPD and malignancies.
The article discusses how the adoptive transfer of EBV-specific T cells (EBVSTs) can be a potential therapeutic approach. EBV-associated LPD, which presents highly immunogenic viral proteins, can be controlled by harnessing the immunogenicity of type III latency malignancies.
The article mentions that EBVSTs have been evaluated for over two decades, and various strategies have been explored, including donor lymphocyte infusions, donor-derived multi-antigen specific VSTs, and autologous or allogeneic VSTs. The review aims to define current manufacturing strategies, summarize clinical experiences, explore opportunities to expand the use of EBV-specific VSTs, and discuss future strategies to enhance their efficacy.
Adoptive immunotherapy has various indications, including the treatment of post-transplant lymphoproliferative disorder (PTLD), prophylaxis and reduction of Epstein-Barr virus (EBV) viral load, and management of refractory cytomegalovirus (CMV), EBV, or adenovirus infections in transplant recipients.
In addition to these indications, adoptive immunotherapy shows potential in the treatment of other conditions related to organ transplantation. It may be explored as a therapeutic option for autoimmune skin diseases and autoimmune conditions such as systemic lupus erythematosus (SLE), type 1 diabetes mellitus (DM I), multiple sclerosis (MS), and inflammatory diseases like atherosclerosis. Furthermore, it could be considered for managing Hodgkin’s lymphoma and nasopharyngeal carcinoma.
As with any medical intervention, adoptive immunotherapy carries the risk of side effects. These can include neurological symptoms such as confusion, headache, delirium, convulsions, and cerebral edema. Anaphylactic reactions are possible in some cases, and patients may experience flu-like symptoms, generalized fatigue, palpitations, and weight gain. There is also a risk of graft-vs-host disease, cytokine release syndrome, and acute kidney injury (AKI) associated with adoptive immunotherapy.
Briefly summarize the principle of adoptive immunotherapy in the treatment of PTLDThis article has a level V evidence because it is narrative review.90% of general population are affected with EBV, in immune-competent persons, EBV infection is well controlled, rarely it presents with fulminant disease, in other cases EBV will stay in latent infection in B-cells. In immune-compromised subjects like solid organ transplant recipients, EBV infection can lead to lympho-proliferative disorders.EBV viral proteins (such as EBNA, LMP1, and LMP2) will be presented on the surface of B-cells in EBV- associated lympho-proliferative disease.Malignancies associated with type II latency include: Hodgkin’s lymphoma, natural killer T-cell lymphoma and Nasopharyngeal carcinoma.Malignancies associated with type I latency tumors include: Burkitt’s lymphoma and gastric carcinoma.PTLD is commonly seen in EBV negative recipients receiving organs from EBV positive donors.EBV-specific T-cells (EBVSTs) were studied for prevention and treatment of EBV related cancers. EBVSTs are produced from EBV positive donors who can be autologous or allogenic sources.EBVSTs can be manufactured via the following strategies:1- Ex-vivo expansion of VSTs vs Antigen-specific T-cell selection.2- MHC molecules are formed and attached to magnetic beads to separate typical CD8+ T-cells specific to viral antigens in HLA restricted manner. 3- IFN-gamma capture uses mononuclear cells pulsed with antigen to separate CD8+ &CD4+ T-cells that respond to viral antigens by secreting IFN-gamma.
What are the indications of adoptive immunotherapy?
a- PTLD.
b- Treatment prophylaxis and EBV viral load reduction.
c- Refractory CMV or EBV or adenovirus infection.
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
4- Auto-immune skin diseases.
5- Auto-immune conditions such as SLE, DM I, and MS.
6- Inflammatory diseases, atherosclerosis.
7- Hodgkin’s Lymphoma.
8- Nasopharyngeal carcinoma..
What are the side effects of adoptive immunotherapy?
1- Confusion, headache, delirium, convulsion, and cerebral edema.
2- Anaphylactic reaction.
3- Flu like symptoms.
4- Generalized fatigue, palpitation, weight gain
5- Graft vs host disease.
6- Cytokine Release Syndrome.
7- AKI.
Epstein-Barr virus (EBV) is a ubiquitous member of the gamma herpes virus family that is associated with a variety of lymphomas and lymphoproliferative disorders (LPD). It infects more than 90% of the adult population worldwide. Primary infection occurs in childhood as a asymptomatic or mild infection and/or may result in a more florid infectious mononucleosis syndrome in teenagers and young adults.
Regardless of the initial infection, EBV maintains a lifelong latency in B cells and oral epithelial cells. During primary infection, EBV enters the oropharynx replicating within the epithelial cells and infect transiting B-lymphocytes (primarily due to their expression of CD21 which is the major receptor for the virus). EBV can also infect epithelial cells via transfer from infected B cells and other processes.
Donor lymphocyte infusions
The earliest reported experience of cellular therapy for the treatment of PTLD utilized unmodified DLI derived from the patient’s EBV seropositive HSCT donor which contained effector cells with activity against EBV (37). Although effective in inducing remissions, this therapy carries a significant risk of graft-versus-host disease (GVHD).
Donor-derived EBVSTs
There is extensive reported experience using donor derived EBVSTs for the prevention and treatment of EBV+ PTLD in the post-HSCT setting when the donor is available.
In the original report using ex vivo expanded EBVSTs from healthy seropositive donors, the team at St. Jude’s Research Hospital treated ten allogeneic HSCT recipients, three with evidence of EBV reactivation and seven at high risk of reactivation.
Demethylating agents
Newly EBV-infected B-cells express up to 90 viral genes; however rapid CpG-methylation of viral antigens leads to downregulation of viral protein expression and the latency .
Azacytidine and decitabine are potent CpG demethylating agents. In a mouse xenograft of latency type I Burkitt lymphoma, pretreatment with decitabine induced expression of LMP1 and ENBA3 associated with latency type 3 which sensitized tumor cells to subsequent therapy with EBVSTs (86). In contrast, azacytidine did not increase expression of those proteins.
Conclusions
The use of adoptive immunotherapy, particularly using third-party “off the shelf” EBVSTs for the treatment of EBV-LD has shown promise in several studies conducted at specialized centers .More recently, EBVSTs have become more widely available including in industry led multi-center studies and consortium and cooperative group studies (20,66). Further work is however needed to create widely available and commercialized third-party cell banks to broaden the applicability of this approach beyond boutique centers. In addition, strategies need to be explored to enhance the anti-tumor activity of EBVST therapies especially for the less immunogenic type I and II latency tumors. Preclinical work and early clinical trials are in process exploring various gene engineering and combination therapy approaches to improve the potency of EBVSTs in vivo.
It involves generating EBV-specific T lymphocytes (EBVSTs) from EBV-positive autologous or allogenic donors with limited alloreactivity and wide virus-fighting activity.
It is necessary to use ex vivo virus-specific T cell growth or antigen-specific T cell selection.
MHC selection involves isolating CD8+ T cells with high affinity to a specific viral epitope or peptide in an HLA-restricted manner; IFN- capturing involves isolating CD8+ and CD4+ T cells that release IFN- in response to viral antigens.
Technically challenging approaches.
Post-HSCT donor-derived T cells
Donor-lymphocyte infusions
Unmodified DLI from PTLD patients’ EBV-positive HSCT donors induces remission.
Donor-derived EBVSTs may prevent and treat EBV+ PTLD in post-HSCT patients without reactivation or GVHD by lowering EBV burden.
EBV-transformed lymphoblastoid cell lines (LCLs) are used to selectively grow EBV-specific T lymphocytes as antigen-presenting cells.
DC and LCLs were transduced with adenovirus vectors to overexpress LMP1 or LMP2 to increase specificity for less immunogenic EBV antigens, but it is difficult and wastes time.
-EBSTs and multi-VSTs provided by third parties in PTLD
-recipient with immunosuppression and latent EBV infection
-Treatment for prevention and a decrease in the patient’s EBV load
-Treatment resistant and refractory cases of PTLD
T-cell adoptive immunotherapy for BK nephropathy in renal transplantation
The treatment of lymphoproliferative diseases connected to viral (EBV) loads
GVHD, acute kidney injury, bleeding episodes, heart arrhythmias, chills, constipation, cough, cytokine release syndrome (cytokine storm), decreased appetite, delirium, diarrhea, dizziness, edema, and encephalopathy.
Briefly summarize the principle of adoptive immunotherapy in the treatment of PTLD:
Adoptive immunotherapy is a principle of treatment used to treat post-transplant lymphoproliferative disorders which developed in transplant recipients due to uncontrolled proliferation of lymphocytes.
.Adoptive T-cell immunotherapy with donor derived EBV-specific T cells focuses on administering EBV-specific T cells, which are specific to immunodominant latency-associated antigens expressed in PTLD, such as Epstein-Barr nuclear Antigens EBNA1,2,3 And membrane proteins LMP1,2A,2B .
What are the indications of adoptive immunotherapy?
1) Some cancers like: Leukemia, lymphoma, melanoma, lung cancer, and kidney cancer.
2) Viral infection in immunocompromised patients like: Post-transplant lymphoproliferative disorders, CMV infection, and HIV.
3) Immunotherapy-resistant or relapsed diseases
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Yes we can use it in resistant viral infections including BKV, CMV, Adenovirus, and HHV 6
What are the side effects of adoptive immunotherapy?
A. Risk of graft vs host disease.
B. Cytokine release syndrome.
C. Neurotoxicity.
Level of evidence is 5
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
Adoptive immunotherapy is a treatment approach used to treat post-transplant lymphoproliferative disorders (PTLD), a group of diseases that can develop in transplant recipients due to uncontrolled proliferation of lymphocytes. The principle behind adoptive immunotherapy is the isolation, expansion, and reinfusion of immune cells, such as T-cells, that are specifically designed or selected to target and eliminate cancerous or infected cells in the patient.
In the context of PTLD, adoptive immunotherapy often involves the use of donor-derived or modified T-cells that recognize and target Epstein-Barr virus (EBV)-infected B-cells, since EBV infection is a common cause of PTLD. This approach helps boost the patient’s immune system to recognize and eliminate malignant or infected cells, leading to better disease control and potentially improved outcomes.
What are the indications of adoptive immunotherapy?
Adoptive immunotherapy has been explored as a treatment option for various medical conditions, particularly in the context of cancer and viral infections. The main indications for adoptive immunotherapy include:
It is important to note that the specific indications for adoptive immunotherapy may vary depending on the type of immune cells being utilized (e.g., T-cells, natural killer cells), the targeted antigen or receptor, and the patient population. The use of adoptive immunotherapy may be limited by factors such as availability, cost, and potential side effects. Always consult with a healthcare professional to determine the most appropriate treatment options for a specific medical condition.
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Yes, adoptive immunotherapy has potential applications in the treatment of various conditions related to organ transplantation. While its primary focus has been on addressing post-transplant lymphoproliferative disorders (PTLD) and viral infections in transplant recipients, researchers are exploring its potential use in other transplantation-related conditions:
These applications are mostly in experimental stages and require further clinical research to determine their safety, efficacy, and optimal usage. However, they highlight the potential of adoptive immunotherapy in improving transplant outcomes and addressing various challenges in the field of organ transplantation.
Summary of Article
EBV positive PTLD is a major complication of solid organ transplantation and Hematopoietic stem cell transplant.Although use of Rituximab ,Chemotherapy and reduction of immunosuppression have increased remission of PTLD,and increased survival of patient ,poor response to existing therapy in many cases needs new therapy.Adoptive immunotherapy is one such therapy.Adoptive T-cell immunotherapy with donor derived EBV-specific T cells focuses on administering EBV-specific T cells, which are specific to immunodominant latency-associated antigens expressed in PTLD, such as Epstein-Barr nuclear Antigens EBNA1,2,3 And membrane proteins LMP1,2A,2B .In Primary EBV infection ,most viral antigens are expressed during lytic stage while virus is replicating .In EBV infected B cells ,there is subsequent downregulation of viral antigen expression from latency type 3 to latency type 0,which allows escape from immune surveillance .It is the latency period which lead to Malignancy such as PTLD,Hodgkin and non hodgkin lymphoma.
Manufacturing of EBV specific T cell manufacturing(EBVSTs)-Although there are number of methods for manufacturing EBVSTs ,most common utilised is Ex vivo Expansion of EBVSTs.In this method mononuclear cells are harvested from the peripheral blood of the donor.After that B lymphocytes are infected with lab strain of EBV .Then they transduced with adenoviral vector expressing LMP1 and LMP2 and then irradiated.Monocytes are separately transduced with same adenoviral vector ,cultured with T Cells followed by a second stimulation by transduced Lymphoblastoid cells lines (LCL) to create Latent membrane protein specific T cell product.
Donor lymphocyte infusion( DLI) in an attempt to kill dividing B cells in EBV-associated PTLD, is a effective method to induce remission. Although both DLI and EBV specific T cytotoxic lymphocyte are almost equally effective in inducing remission,DLI is associated with Graft versus host disease,so not preferred .
Indications of Adoptive immunotherapy-
(A)PTLD–Due to non-availability of EBV specific cytotoxic lymphocytes at many centers and association with Graft versus host disease ,It is reserved for PTLD who failed to resolve with other therapy.
(B) Other disease -Treatment of difficult to treat viral infection, Progressive Multiple sclerosis .,Immune related Brain tumors
Other conditions related to organ transplantation
(1) Treatment of Drug resistant CMV infection
(2) Treatment of BK Virus nephropathy
(3)Prevention and Treatment of Rejection
(4) Development of immune tolerance via regulatory T cells (Tregs)
Side effects of Adoptive immunotherapy-
Graft versus host disease,
Cytokine release syndrome
Neurotoxicity
Level of evidence of Article– 5
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
Post-transplant PTLD is strongly associated with latent EBV infection being reactivated due to immunosuppressive therapy leading to suppression of T cell activity and suppression of EBV infected B cells. In this regard the use of EBV specific T cells transference (allogenic or autologous) can be used to keep the EBV infected B cell population within control and reduce risk of PTLD development.
What are the indications of adoptive immunotherapy?
· EBV associated PTLD (treatment and prophylaxis)
· Immunoblastic Lymphoma
· EBV associated Hodgkin lymphoma
· EBV associated NK/T Lymphoma
· EBV associated nasopharyngeal carcinoma
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Refractory Viral infections including BKV, CMV, Adenovirus, and HHV 6.
What are the side effects of adoptive immunotherapy?
· Risk of graft vs host disease (acute and chronic)
· Infusion associated reactions including cytokine storm
Level of Evidence:
Level 5 as it is a review article based on multiple non RCTs as well as authors’ personal experience and data.
1. Briefly summarize the article
Principle of adoptive immunotherapy in the treatment of PTLD
Review aricle about adoptive cellular immunotherapy for Epstein-Barr virus-associated lymphoproliferative disease (EBV-LD) .
Introduction:
When infection occur , EBV remain latent in B cells, oral epithelial cells and lymphoid follicles by downregulating the immunogenic proteins and get saved from immune surveillance [invisible to EBV-specific T-lymphocytes (EBVSTs)].
Competent T cell immunity control the infected B cell amount at <2% ; but in case of reduced immunity, with absence of EBVST response and uncontrolled proliferation of EBV infected B cells (mainly type3 latency) results in EBV-associated LPD and malignancies.
Manufacturing of EBVSTs:
1- Ex vivo expansion: of T cells targeting viral antigens via native T cell receptors where EBVSTs were selectively expanded utilizing irradiated EBV transformed lymphoblastoid cell lines (LCL) as antigen presenting cells (APCs) to selectively expand EBVSTs
2- Antigen specific T cell selection :
EBVSTs can be easily produced from seropositive blood using good-manufacturing-practice (GMP)-compliant protocols and cryopreserved for future use.
Adoptive cell therapy with Virus-specific T cells (VSTs) – reports impressive results in immunocompromised candidate with EBV-LD studies .
donor lymphocyte infusions (DLI) and donor derived multi-antigen specific VSTs in the HSCT – produce ORR of about 55-67%
Post-HSCT donor-derived T cell therapy:
1- Donor lymphocyte infusions
2- Donor-derived EBVSTs
In a study from (USA) : safety and efficacy of donor-derived EBVSTs for treatment and prophylaxis of PTLD post transplantation has been reported .
In another study :
– 114 patients received donor derived EBVSTs after allogeneic HSCT
– as a prophylaxis of 101 pt – No PTLD
– as a Treatment of 13 PTLD pt — 11 patients underwent complete remission.
(MSKCC) group result shows – (ORR) were 72% with DLI and 68% with EBVSTs
Strategies to enhance EBVST potency in vivo through genetic engineering as well as combination therapies:
Modification of EBVSTs to enhance activity
1. Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo – dominant negative TGF-β receptor type II (DNRII)
2. Calcineurin resistance to enhance EBVSTs – EBVSTs were genetically engineered to express a mutant form of calcineurin thus rendering them calcineurin inhibitor (CNI) resistant
3. Chimeric antigen receptor T cells — CD19-chimeric antigen T cells (CD19-CART) have shown impressive efficacy in acute B-lymphoblastic leukemia and in B-cell lymphomas
4. Combination of EBVSTs + modalities to enhance EBVST activity in vivo:
Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance
EBVST activity in vivo–
– Demethylating agents
– Checkpoint inhibitors
– BCl2 inhibitors
Conclusions:
Adoptive immunotherapy, particularly third-party “off the shelf” EBVSTs for treatment of EBV-LD is a promising approach.
But it is not widely available at time being.
Further work and Strategies needed to improve the anti-tumor activity of EBVST therapies for the less immunogenic type I and II latency tumors .
2. What are the indications of adoptive immunotherapy?
EBVSTs have been used as an adjunctive therapy to chemotherapy in
· Hodgkin lymphoma
· NK/T Cell lymphoma
· Nasopharyngeal carcinoma
· Burkitt lymphoma
side effects :
1- Cytokine release syndrome (CRS)
2- Neurotoxicity
3- Acute kidney injury
4- Risk of graft rejection and autoimmunity.
3. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Treatment and prophylaxis to CMV, EBV, and Adenovirus.
Treatment of malignancies – Melanoma, Leukemias, Adenocarcinoma of Prostate
Referactory cases of EBV positive PTLD.
Level of evidence : 5
Summary of principle
Adoptive immunmotherapy has good result in EBV related PTLD. The rationale behind using adoptive transfer of EBVSTs is based on harnessing immunogenicity of type III latency malignancies to control proliferation of latently infected B cells.
EBV associated malignancies with type II latency can develop in immunocompetent and immune deficient individuals and are much less immunogenic due to down regulation of the immunodominant antigens.
Indications of adoptive immunotherapy
Adoptive therapy in treatment of conditions assoc. with transplant
Side effects of adoptive immunotherapy
Briefly summarize the principle of adoptive immunotherapy in the treatment of PTLD:
· Adoptive immunotherapeutic approaches to re-establish viral-specific immunity are an appealing alternative since viral problems in these patients are unmistakably linked to the absence of recovery of virus-specific cellular immune responses.
· The CD8+ cytotoxic T lymphocytes (CTLs), which identify peptides produced from viral proteins complexed to major histocompatibility complex (MHC) class I molecules, are the most crucial part of the cellular immune response that regulates the majority of viral infections.
· Furthermore, CD4 virus-specific T cells are crucial for supporting CD8+ cells and generating long-lasting protection.
· Researchers have looked into whether adoptive transfer of virus-specific CTLs can protect patients from such viruses because unmanipulated T-cell populations also contain alloreactive T cells.
What are the indications of adoptive immunotherapy:
· Treatment and prevention of Viral infections like CMV, EBV, and Adenovirus.
· Number of malignancies ranging from melanoma to certain types of leukaemia, as well as prostate cancer
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation:
· Recurrent leukemic after allogeneic bone marrow transplantation.
· Treatment against CMV infection in SOT recipients.
· Treatment of PTLD caused by Epstein–Barr virus.
What are the side effects of adoptive immunotherapy:
· General Fatigue
· Fever
· elevation of hepatobiliary enzymes,
· cytokine release syndrome (also known as cytokine storm),
· neurotoxicity
· Graft vs Host Disease
EBV positive PTLD is a major complication of hematopoietic stem cell & solid organ transplantation. Two types of adoptive immunotherapy i) Donor lymphocyte infusion & ii) EBV specific T- lymphocytes (EBVSTs)
Individuals with weakened immune system, such as recipients of HSCT or SOT (solid organ transplant), the lack of a robust EBVST response can lead to uncontrolled proliferation of type 3 latency EBV – infected B cells resulting in EBV associated LPD & malignancies. The transformed B-cells in EBV- LD associated withlatency type III present several antigenic viral protein that induce potent EBVDT responses. Such potent T cell immunity maintains the infected B cell pool at <2% of total B cells in immunocompetent individuals but is lacking in immunosuppressef individuals leading to uncontrolled lymphoproliferation of the infected B cells.
The adoptive transfer of EBVSTs is based on harnessing the immunogenecity of type III latency malignancies to control the proliferation of the latency infected B cells.
– EBV associated PTLD in HSCT & SOT recipients.
-HIV lymphoma
Prevention of EBV associated PTLD in high risk transplant recipients
Acute & chronic graft versus host disease ( GVHD), which is more common with donor lymphocyte infusion.
1. Briefly summarize the article
Principle of adoptive immunotherapy in the treatment of PTLD
This is a review of published literature on adoptive cellular immunotherapy for Epstein-Barr virus-associated lymphoproliferative disease (EBV-LD) in last 20 years.
Introduction:
After initial infection (lytic phase), EBV maintains a lifelong latency in B cells, oral epithelial cells and lymphoid follicles by downregulating the immunogenic proteins and get rescue from immune surveillance [invisible to EBV-specific T-lymphocytes (EBVSTs)].
potent T cell immunity maintains the infected B cell pool at <2%; but in patients with weak immunity, lack of robust EBVST response and uncontrolled proliferation of EBV infected B cells (mainly type3 latency) results in EBV-associated LPD and malignancies.
Rationale for adoptive transfer of EBVSTs is based on harnessing the immunogenicity of type III latency malignancies to control the proliferation of the infected B cells.
Objectives:
– Define current best manufacturing strategies for EBVST
– Summarize the clinical experience on their use in EBV-related LPD,
– Opportunities to broaden the applicability of this approach
– Explore future strategies to enhance their efficacy
Manufacturing of EBVSTs:
Ex vivo expansion
Antigen specific T cell selection
EBVSTs can be readily produced from seropositive donors using good-manufacturing-practice (GMP)-compliant protocols and cryopreserved for future use.
Adoptive cell therapy with Virus-specific T cells (VSTs) have been used for more than two decades – has shown impressive results in immunocompromised patients with EBV-LD in clinical trials.
· donor lymphocyte infusions (DLI) and donor derived multi-antigen specific VSTs in the HSCT – ORR 55-67%
· autologous & allogeneic VSTs in the SOT – EBVST infusion did not consistently decrease EBV viremia, but no patient after prophylaxis developed PTLD
EBVST from LCL used for Latency type2 malignancies (relapsed EBV-HL) – OR 60-70%
Post-HSCT donor-derived T cell therapy:
· Donor lymphocyte infusions
· Donor-derived EBVSTs
St. Jude’s (USA) preliminary study: safety and efficacy of donor-derived EBVSTs for treatment and prophylaxis of PTLD in transplant recipients has been seen in many studies.
Large multicenter study:
– 114 patients received donor derived EBVSTs after allogeneic HSCT
– prophylaxis in 101 patients – No PTLD
– Treatment in 13 patients with PTLD — 11 patients achieved complete remission; persisted 9 years
(MSKCC) group – (ORR) were 72% with DLI and 68% with EBVSTs
Third Party T cells
readily available “off the shelf” T cell therapy product is desirable.
UK- 60 products bank
33 Tx recipients – ORR 64 % in HSCT; 52% in SOT
Birmingham, United Kingdom published their experience in 10 pediatric SOT recipients with PTLD and reported an ORR of 80% (8 out of 10)
Strategies to enhance EBVST potency in vivo through genetic engineering as well as combination therapies:
Modification of EBVSTs to enhance activity
1. Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo – dominant negative TGF-β receptor type II (DNRII)
2. Calcineurin resistance to enhance EBVSTs – EBVSTs were genetically engineered to express a mutant form of calcineurin thus rendering them calcineurin inhibitor (CNI) resistant
3. Chimeric antigen receptor T cells — CD19-chimeric antigen T cells (CD19-CART) have shown impressive efficacy in acute B-lymphoblastic leukemia and in B-cell lymphomas
4. Combination of EBVSTs + modalities to enhance EBVST activity in vivo:
– Demethylating agents
– Checkpoint inhibitors
– BCl2 inhibitors
Conclusions:
Adoptive immunotherapy, particularly third-party “off the shelf” EBVSTs for treatment of EBV-LD has shown promise in several studies conducted at specialized centers, also is being available for multi-center studies, consortium and cooperative group studies.
Further work is needed to create widely available and commercialized third-party cell banks to broaden the applicability of this approach beyond specialized centers.
Strategies to enhance the anti-tumor activity of EBVST therapies for the less immunogenic type I and II latency tumors, need to be explored.
2. What are the indications of adoptive immunotherapy?
PTLD in SOT – post Retuximab + Chemotherapy – refractory / relapse
– post Retuximab monotherapy relapse patients not tolerating Chemotherapy
Post-HSCT: prophylaxis for PTLD and viral associated malignancies in children, post Stem Cell Transplant
Malignancies secondary to EBV.
Resistant immunological diseases.
3. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Treatment and prevention of Viral illness due to CMV, EBV, and Adenovirus.
Malignancies – Melanoma, Leukemias, Adenocarcinoma Prostate
EBV positive PTLD – refractory and relapse
EBV associated Hodgkin’s lymphoma, NK cell Lymphoma, nasopharyngeal carcinoma
Autoimmune diseases like lupus or Multiple Sclerosis
4. What are the side effects of adoptive immunotherapy?
Discussion question
Level of evidence – none levelled
Adoptive cellular immunotherapy for Epstein-Barr virus-associated lymphoproliferative disease.
Introduction.
Epstein-Barr virus (EBV) is associated with a variety of lymphomas and lymphoproliferative disorders (LPD). It infects more than 90% of the adult population worldwide, body produces CD8+ cytotoxic T lymphocytes (CTL) specific to the early lytic cycle proteins kill cells entering the lytic cycle before they are able to release infectious virus particles, EBV infected B lymphocytes per 1,000,000 peripheral blood mononuclear cells (PBMCs) over their lifetime, controlled at these levels by a potent EBVST response, in immunocompromised situation such as post organ transplantation, the lack of a robust EBVST response can lead to uncontrolled proliferation of type 3 latency EBV-infected B cells resulting in EBV-associated LPD and malignancies.
The role for adoptive transfer of EBVSTs is to keep the immunogenicity of type III latency malignancies to control the proliferation of the latently infected B cells, the biggest risk factor for developing EBV-associated PTLD post-SOT is EBV seronegativity at the time of transplant.
Rituximab, targeting CD20 present on the B cells has been an effective monotherapy with response rates of 55% to 100% in HSCT recipient and so response rates to rituximab monotherapy in SOT recipients are generally lower and around 50%, and other treatment option is immunotherapy with EBVSTs. Adoptively transferred virus-specific T cells (VSTs) have been evaluated for more than two decades.
Manufacturing of EBVSTs.
1-ex vivo expansion of VSTs versus antigen-specific.
2-T cell selection [e.g., interferon γ (IFN-γ) capture].
Post-HSCT donor-derived T cell therapy.
Donor lymphocyte infusions.
Utilized unmodified DLI derived from the patient’s EBV seropositive HSCT donor which contained effector cells with activity against EBV. Although effective in inducing remissions, this therapy carries a significant risk of graft-versus-host disease (GVHD).
Donor-derived EBVSTs.
Patients who received the EBVSTs as prophylaxis had any EBV reactivation or GVHD and there was evidence of persistence of EBVSTs by tracking of genetic markers on the T cells for a median of 10 weeks.
Autologous EBVSTs.
When the donor is not available such as cadaveric organ transplantation, autologous EBVSTs have also been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy.
Third-party T cells.
Used with refractory PTLD between the age of 1–76 years received partially HLA matched EBVSTs. HLA matches ranged from 2 to 5/6 HLA alleles and there was a statistically significant association of better outcome with higher HLA matches. Overall, the response rate (CR and PR) was 64% at 5 weeks and 52% at 6 months.
Modification of EBVSTs to enhance activity
=Overcoming the immune suppressive effects of TGF-β to
enhance EBVST activity in vivo.
The DNRII-LSTs were resistant to otherwise inhibitory concentrations of TGF-β.
Calcineurin resistance to enhance EBVSTs.
EBVSTs were genetically engineered to express a mutant form of calcineurin thus rendering them calcineurin inhibitor (CNI) resistant.
Chimeric antigen receptor T cells.
CD19-chimeric antigen T cells (CD19-CART) have shown impressive efficacy in acute B-lymphoblastic leukemia and in B-cell lymphomas.
Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance EBVST activity in vivo.
–Demethylation agents.
-Checkpoint inhibitors.
-BCL-2 inhibitors.
Conclusions.
The use of adoptive immunotherapy, EBVSTs for the treatment of EBV-LD has shown promise in several studies conducted
at specialized centers, and need more studies for combination therapy to increase their efficacy.
Indications:
·Post-transplant lymphoproliferative diseases.
·Malignancies secondary to EBV.
·Resistant immunological diseases.
Other Indications:
·EBV associated malignancies e.g., Hodgkin’s lymphoma, natural killer cell lymphoma, nasopharyngeal carcinoma
·Viral associated infections -CMV,EBV
· Autoimmune diseases like lupus or MS
Side effects:
·Cytokine release syndrome
·GVHD
· Neurotoxicity.
In individuals with weakened immune systems, such as patients with solid organ transplant (SOT), the lack of a robust EBVST response can lead to uncontrolled proliferation of type 3 latency EBV-infected B cells resulting in EBV-associated LPD and malignancies.
Adoptive immunotherapy involves isolating EBV specific T-lymphocytes (EBVSTs) from a patient with EBV-seropositive PTLD patients (these cells can be isolated from a resected tumor tissue or peripheral blood then they undergo ex vivo (in the laboratory) activation and expansion using good-manufacturing-practice (GMP)-compliant protocols)
Autologous EBVSTs have also been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients or the third-party EBVSTs can be used
After that infused back into the patient in large numbers where they can now help the patient’s immune system to destroy the cancer cells.
Adoptive immunotherapy is reserved for EBV-positive patients who fail to respond to initial therapy.
What are the indications of adoptive immunotherapy?
– Management of resistant PTLD
– EBV-associated malignacy
– Refractory/ resistant disease/cancer
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
– resistant EBV specific tumors
– resistant Skin Melanoma
– resistant Autoimmune disease
– resistant BK nephropathy
What are the side effects of adoptive immunotherapy?
– GVHD
– Cytokine storm (cytokine release syndrome)
– Neurotoxicity
– Graft rejection
Disscussion question
Level of evidence – none leveled.
PRINCIPLE OF ADAPTIVE IMMUNOTHERAP-
FIRST B ELLS GET INFECTED WITH EBV, EBVinfected B-lymphocytes then enter the lymphoid follicles
and downregulate the immunogenic proteins
CD8 PLUS T CELLS ( VIRUS SPECIFIC T CELLS, VST ) ARE INVOLVED IN DOWNREGULATIMH EBV INFECTED B CELLS WHICH CAN FORM EBV RELATED TUMORS
WHEN SUCH T CELLS ARE LESS IN NUMBER , THERE IS HIGH CHANCE OF EBV RELALATED LYMPHOPROILIFERATIVE DISEASE
TYPE 1 AND 2 LATENCY TUMOR ARE LESS IMMUNOGENIC WHILE TYPE 3 IS MORE IMMUNOGENIC AND IS THE TARGET OF ADAPTIVE IMMUNOTHERAPY AS IT INDICES STRONG EBV RELATED T CELL RESPONSE
INDICATION-
FOR PTLD IN POST TRANSPLANT SITUATION WHEN RITUXIMAB AND OR CHEMO IS ADVISABLE BUT IT IS MORE TOXIC
THIS IS NEW AND EVOLVING THERAPY FOR PTLD AND FOUND TO BE USEFUL IN PEDIATRCI PATIENTS AND PTLD AFTER STEM CELL TRANSPLANT
FOR SOLID ORGAN TRANSPLANT , TRAILS ARE GOING ON WITH THIRD PARTY T CELL ADAPTIVE IMMUNOTHERAPY
MINIMAL SIDE EFFECT CAN BE THEIR ADVANTAGE
SOURCE OF T CELL-
AUTOLOGOUS T CELL
OWN T CELLS AUGMENTATED IN LAB
DONOR SPECIFIC , HLA MATCHED T CELLS
OFF THE SHELF , THIRD PARTY T CELLS- EASILY AVAILABE ON TIME , COMMON IN RESEARCH TRIALS
OTEHR USES-
TO IMPROVE THE IMMUNITY OF TRANSPLANT PATIENT
TO COMBAT OTHR FORM OF TUMOR WHICG ARE STILL COMMON AFTER TRANSPLANT
SIDE EFFECT-
GVHD , SKIN LESION
GI BLEEDING
TMA
Briefly summaries the principle of adoptive immunotherapy in the treatment of PTLD
Epstein-Barr virus (EBV) is a member of the gamma herpes virus family that is associated with a variety of lymphomas and lymphoproliferative disorders (LPD).
It infects more than 90% of the adult population worldwide.
have different clinical presentations include asymptomatic infection to symptomatic disease.
normally in healthy person, virus neutralizing antibodies control the spread of infection by EBV-specific CD8 + cytotoxic T lymphocytes (CTL) specific to the early lytic cycle proteins that kill cells entering the lytic cycle before they are able to release infectious virus particles.
What are the indications of adoptive immunotherapy?
PTLD associated with and without EBV.
malignancy other than PTLD that associate with EBV.
treatment of PTLD.
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Treatment against CMV infection in SOT recipients.
Treatment of PTLD caused by Epstein–Barr virus.
recurrent leukemia.
What are the side effects of adoptive immunotherapy?
fatigue.
elevation of hepatobiliary enzymes
fever
Briefly summarize the principle of adoptive immunotherapy in the treatment of PTLD
· Immunotherapeutic approaches to reestablish viral-specific immunity are an appealing alternative since viral problems in these patients are unmistakably linked to the absence of recovery of virus-specific cellular immune responses.
· The CD8+ cytotoxic T lymphocytes (CTLs), which identify peptides produced from viral proteins complexed to major histocompatibility complex (MHC) class I molecules, are the most crucial part of the cellular immune response that regulates the majority of viral infections.
· Furthermore, CD4 virus-specific T cells are crucial for supporting CD8+ cells and generating long-lasting protection.
· Researchers have looked into whether adoptive transfer of virus-specific CTLs can protect patients from such viruses because unmanipulated T-cell populations also contain alloreactive T cells.
=====================================
What are the indications of adoptive immunotherapy?
· Treatment and prevention of Viral infections like CMV, EBV, and Adenovirus.
· Number of malignancies ranging from melanoma to certain types of leukemia, as well as prostate cancer
======================================
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
· Recurrent leukemia after allogeneic bone marrow transplantation.
· Treatment against CMV infection in SOT recipients.
· Treatment of PTLD caused by Epstein–Barr virus.
=======================================
What are the side effects of adoptive immunotherapy?
· Fever,
· general fatigue.
· elevation of hepatobiliary enzymes,
· cytokine release syndrome (also known as cytokine storm),
· neurotoxicity
· GVHD
SUMMARY:
· Epstein Bar Virus causes widely spread virus associated with lymphoprolifrative disease in SOT and hematopoietic stem cell transplant.
· This virus infects more than 90% of adult population and tends to have different clinical presentations include asymptomatic infection to full blown disease.
· First successes with adoptive immunotherapy were described using donor lymphocyte infusion for the treatment of EBV+ PTLD in hematopoietic stem cell transplantation (HSCT) recipients
· Adoptive immunotherapy basically involves isolating EBV specific T-lymphocytes (EBVSTs) from a patient with PTLD i.e., EBV-seropositive patients.
· These cells can be isolated from a resected tumor tissue or peripheral blood then they undergo ex vivo (in the laboratory) activation and expansion using good-manufacturing-practice (GMP)-compliant protocols.
· After that infused back into the patient in large numbers where they can now help the patient’s immune system to destroy the cancer cells.
· Adoptive immunotherapy is reserved for EBV-positive patients who fail to respond to initial therapy.
· Adoptive immunotherapy using EBVSTs has shown good results in EBV related Lymphoprolifrative disorders.
Indications:
· Treatment of post transplant lymphoproliferative diseases.
· Malignancies secondary to EBV.
· Resistant diseases.
Other Indications:
· EBV associated malignancies e.g., Hodgkin’s lymphoma, natural killer cell lymphoma, nasopharyngeal carcinoma
· Skin cancer-Melanoma.
· Viral associated infections -CMV,EBV
· Autoimmune diseases like lupus or MS
Side effects:
· Cytokine release syndrome
· GVHD
· Graft rejection
· Others: AKI, dizziness, capillary leak syndrome, hypotension, infection, arrhythmia
EBV-positive post-transplant PTLD is one of the major complication of solid organ transplantation.
objectives of this review are:
(I) manufacturing strategies of EBV specific T-lymphocytes (EBVSTs);
(II) to review different EBVST donor options
(III) to summarize current experience with adoptive cellular therapy
(IV) to explore the potential for combination therapies with other immunotherapeutic strategies.
Epstein-Barr virus (EBV) is a member of the gamma herpes virus family. . It infects more than 90% of the adult population worldwide. normally
In healthy individuals, virus neutralizing antibodies control the spread of infection by EBV-specific CD8 + cytotoxic T lymphocytes (CTL) specific to the early lytic cycle proteins that kill cells entering the lytic cycle before they are able to release infectious virus particles.
EBV-infected naïve B-lymphocytes express proteins comprising the entire EBV genome including the EBV nuclear antigens EBNA1, EBNA2, EBNA3, and EBNALP, membrane proteins LMP1, and LMP2 as well as BARF1 and two small non-translated ribonucleic acids (RNA) (Type III latency) .
EBV infected B-lymphocytes then enter the lymphoid follicles and downregulate the immunogenic proteins to express less immunogenic type II latency proteins (EBNA1, LMP1 and LMP2) and thus rescue them into the memory compartment where the virus persists in latently infected B-lymphocytes by further downregulating expression of viral proteins so as to become invisible to EBV-specific T-lymphocytes (EBVSTs).
EBV-associated malignancies are associated with type II latency [e.g., Hodgkin’s lymphoma (HL), natural killer (NK)/T-cell lymphoma, nasopharyngeal carcinoma (NPC)] or type I latency tumors (e.g., Burkitt lymphoma or gastric carcinoma).
EBV-associated post-transplant lymphoproliferative disease (PTLD) occurs between 1% -25% of HSCT recipients depending on the serostatus of the donor and patient. and the level of post-HSCT immunosuppression
biggest risk factor for PTLD post-SOT is EBV seronegativity at the time of transplant. Since most children are transplanted at a young age while still being EBV seronegative and convert to EBV seropositivity within 2 years of transplant, EBV driven PTLD is much more common in pediatric SOT recipients because of the fact that at a young age while still being EBV seronegative and convert to EBV seropositivity within 2 years of transplant,
More than 90% of EBV-associated PTLD is of B-cell origin which are CD20+ .Rituximab are effective monotherapy with response rates of 55% to 100% in HSCT recipients Third-party EBVSTs have been mostly used in the posttransplant setting. But not all patients treated with third-party EBVST’s had detectable viral loads at start of therapy .
Moreover, clinical response and reduction of EBV viremia correlated with an increase of EBVSTs .
therefore it was conculded that the use of adoptive immunotherapy, particularly using third-party EBVSTs for the treatment of EBV-LD has shown promise in several studies Further work is however needed to create widely available and commercialized third-party cell banks.
utilized for type II latency EBV associated malignancies including HL, T cell lymphoma, NPC as adjunctive therapy.
treatment of autoimmune diseases and has been used in BK virus nephropathy, melanoma and EBV specific tumors.
1. Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD.
Introduction; Epstein Barr virus is a gamma virus family almost infects >90% of population worldwide.
Usually asymptomatic and involves at childhood.
But can present with mild and florid disease like IMS.
Despite of initial infection the virus maintains a lifelong latency in B cell and oral epithelial cells.
The major receptor for EBV which is present on B-cell and remain latent.
The infected B- cells then express protein comprises of EBNA1, 2, 3 and EBNALP, membrane protein LMP1, 2 as well as well BARF1.
They survive in cells by down regulating the immune system.
When there is immunodeficiency and lack of EBVCT response in an individual then there is uncontrolled proliferation of virus infected b-cells and resulted EBV associated malignancies.
.. The adaptive immunotherapy is based on harnessing the immunogenicity of the type III latency malignancies to control its latent B-cell proliferation.
Adaptive immunotherapy involves isolating EBVST from a seropositive patient from peripheral blood then is being activated (ex vivo) using good manufacturing protocol, MHC multicenter selection and the most commonly used mathod interferon-y capture approach and these expended effector cells are re-infused, where they can boast immune system against the viruses, tumor cells.
Adaptive immunity as a prophylactic and therapeutic the success rate is around 80- 100%.
For this purpose usually the donors are autologous and allogeneic.
2. What are the indications of adoptive immunotherapy?
It’s being used for type III, II, and type I related carcinomas and EBV- associated lymphproliferative diseases,
Treatment autoimmune diseases,
Treatment of refractory viral infections like CMV,
Hodgkin lymphoma,
Burkit lymphoma,
Nasopharyngeal carcinoma,
NK/T cell lymphoma.
3. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Autoimmune disease like lupus, MS,
Chronic hypersensitivity reaction,
Other inflammatory disorders,
4. What are the side effects of adoptive immunotherapy?
Cytokine release syndrome,
Acute kidney injury,
Thrombotic microangiopathy,
Hypersensitivity like reaction like fever, edema, rash,
Neurotoxicity.
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
EBV-positive post-transplant lymphoproliferative disease (PTLD) is a major complication of hematopoietic stem cell and solid organ transplantation.
objectives of this review are: (I) to describe different manufacturing strategies of EBV specific T-lymphocytes (EBVSTs); (II) to review different EBVST donor options and their advantages and limitations; (III) to summarize current clinical experience with adoptive cellular therapy for EBV-LD and finally; (IV) to explore the potential for combination therapies with other immunotherapeutic strategies.
Epstein-Barr virus (EBV) is a ubiquitous member of the gamma herpes virus family that is associated with a variety of lymphomas and lymphoproliferative disorders (LPD). It infects more than 90% of the adult population worldwide
In healthy seropositive individuals, virus neutralizing antibodies control the spread of infectious virus particles and EBV-specific, human leukocyte antigen (HLA) class I restricted, CD8 + cytotoxic T lymphocytes (CTL) specific to the early lytic cycle proteins kill cells entering the lytic cycle before they are able to release infectious virus particles.
EBV-infected naïve B-lymphocytes express proteins comprising the entire EBV genome including the EBV nuclear antigens EBNA1, EBNA2, EBNA3, and EBNALP, membrane proteins LMP1, and LMP2 as well as BARF1 and two small non-translated ribonucleic acids (RNA) (Type III latency) .
EBVinfected B-lymphocytes then enter the lymphoid follicles and downregulate the immunogenic proteins to express less immunogenic type II latency proteins (EBNA1, LMP1 and LMP2) and thus rescue them into the memory compartment where the virus persists in latently infected B-lymphocytes by further downregulating expression of viral proteins so as to become invisible to EBV-specific T-lymphocytes (EBVSTs).
EBV-associated malignancies associated with type II latency [e.g., Hodgkin’s lymphoma (HL), natural killer (NK)/T-cell lymphoma, nasopharyngeal carcinoma (NPC)] or type I latency tumors (e.g., Burkitt lymphoma or gastric carcinoma) can develop in immunocompetent and immune deficient individuals and are much less immunogenic due to downregulation of the immunodominant (e.g., EBNA3 and EBNA2) antigens.
EBV-associated post-transplant lymphoproliferative disease (PTLD) occurs in less than 1% to 25% of HSCT recipients depending on the serostatus of the donor and patient, the degree to which the graft is T-cell depleted, and the post-HSCT immunosuppression
biggest risk factor for developing EBV-associated PTLD post-SOT is EBV seronegativity at the time of transplant. Since most children are transplanted at a young age while still being EBV seronegative and convert to EBV seropositivity within 2 years of transplant, EBV driven PTLD is much more common in pediatric SOT recipients .
More than 90% of EBV-associated PTLD is of mature B-cell origin with cell surface expression of CD20 .
Rituximab, monoclonal antibody targeting CD20 present on the B cells has been an effective monotherapy with response rates of 55% to 100% in HSCT recipients
Third-party EBVSTs have been mostly used in the posttransplant setting. There has been no published experience in patients with HIV-associated lymphomas because in the modern era of highly active anti-retroviral therapy (HAART) the incidence in the Western word has decreased and the logistical support and specialized experience needed does not make EBVSTs an easily accessible option for the treatment of patients in the developing world.
not all patients treated with third-party EBVST’s had detectable viral loads at start of therapy .
Leen et al. also showed reduction of EBV viral load correlated with response in HSCT recipients with EBV-associated disease treated with third-party multi virus specific VSTs .
Moreover, clinical response and reduction of EBV viremia correlated with an increase of EBVSTs .
Conclusion
The use of adoptive immunotherapy, particularly using third-party “off the shelf” EBVSTs for the treatment of EBV-LD has shown promise in several studies conducted at specialized centers .
Further work is however needed to create widely available and commercialized third-party cell banks to broaden the applicability of this approach beyond boutique centers.
In addition, strategies need to be explored to enhance the anti-tumor activity of EBVST therapies especially for the less immunogenic type I and II latency tumors.
What are the indications of adoptive immunotherapy?
Prophylactic treatment of EBV load
Refractory EBV and CMV
PTLD associated with and without EBV
malignancy other than PTLD that associate withEBV
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Autoimmune diseases
Skin diseases
What are the side effects of adoptive immunotherapy?
GVHD
neurotoxicity
Acute rejection
Allergic reaction
Level of evidence 5
PRINCIPLES OF ADOPTIVE IMMUNOTHERAPY IN TREATING POST TRANSPLANT PTLD.
EBV infection is common in the general population and mostly latent affecting more than 90% of the world population. It has is associated with lymphoproliferative diseases post transplant ;SOT and HSCT. Irrespective of initial mode of infection, maintains a lifelong latency in oral epithelial cells and will cause an infection once immunity gets compromised either by chronic infections like HIV, malignancies or immunosuppressive medications.
Enhanced immunogenicity of type 3 latency malignancies reduces proliferation of latent B cells thus reducing propensity of EBV to be active/malignant post transplant. EBV malignancies(Type 2 eg HL,NK Cell lymphoma and NPC) can in both healthy and immunocompromised population and a re not very immunogenic due to decreased expression of EBNA3 and EBNA2 antigens.
EBV associated LD ,type 3 present antigenic proteins that stimulate strong EBVST response, This strong resultant immunity helps keep infected B cell pool at a low nadir, less than 2% of total B cells in healthy people a factor not present in the immunosuppressed post transplant, Exploration of this aspect may lead to a decrease in proliferation of infected B cells and a decline in incidences of EBV associated malignancies post transplant.
EBV Production methods;
1.EX vivo expansion of EBVST.
2.Antigen specific T cells.
3.Autologous EBVST.
4.3rd Party T cells.
5.Allogenic lymphocytes.
EBVST enhancing Methods;
1.Surmount TGF Beta immunosuppressive effects.
2.Chimeric AG receptor T cell.
3.Calcineurin inhibitors to enhance EBVST.
INDICATIONS OF ADOPTIVE IMMUNOTHERAPY.
1.Treatment of post transplant lymphoproliferative diseases.
2,Malignancies secondary to EBV.
3.Resistant diseases.
OTHER INDICATIONS OF ADOPTIVE IMMUNOTHERAPY.
1.Skin cancer-Melanoma.
2.Viral associated infections -CMV,EBV
3.Autoimmune diseases like lupus or MS
SIDE EFFECTS OF ADOPTIVE IMMUNOTHERAPY.
1.Cytokine storm- CART therapy.
2.GVHD- Donor lymphocyte infusion.
3.Graft dysfunction ;rejection -Check point inhibitors
4.Others – dizziness, capillary leak syndrome, hypotension, infection etc
Narration – Level 5 evidence.
Epstein- Bar Virus Is A Worldwide Widely Spread Gamma Herpes Virus Family That Has Been Associated With Lymphoproliferative Disease In Solid Organ Transplant And Hematopoietic Stem Cell Transplant,
Adoptive Immunotherapy Involves Isolation And Infusion Of T Lymphocytes In A Patient For Treatment/ Prevention Of A Disease
EBV Infects More Than 90% Of The Adult Population.
Primary Infection Mainly Occurs In Childhood Leading To Asymptomatic/Mild
Infection/Infectious Mononucleosis.
EBV Enters The Oropharynx Replicating In The Epithelial Cells And Infecting Transiting B Lymphocytes.
The Infected B Lymphocytes Express Proteins Representing The Whole EBV Genome. They Then Enter The Lymphoid Follicles Where They Down Regulate The Immunogenic Proteins To Express Less Immunogenic Type 2 Latency Proteins.
They Thus Enter The Memory Compartment Where The Virus Persist In Latently Infected B Lymphocytes.
The Frequency Of EBV Infected B Lymphocytes Remain Stable In A Healthy Individual. However, In An Immunocompromised Individual, Lack Of A Robust EBV Specific T Lymphocytes Leads To Uncontrolled Proliferation Of Type 3 Latency EBV Leading To LPD And Malignancies.
In healthy individuals, the rate of EBV infection remains stable at 0.1-50 EBV-infected B lymphocytes per 1,000,000 peripheral blood mononuclear cell
EBV associated PTLD is a well described complication after both hematopoietic stem cell (HPSC) and solid organ transplantation (SOT) in about 1-25% of cases.
The risk of PTLD is dependent on the EBV serostatus of the donor (higher risk with positive donor and negative recipient) and the degree of immunosuppression (higher risk with ATG induction and TAC based maintenance therapy).
Risk of EBV-PTLD is higher in pediatric SOT (as they are transplanted early before acquiring infection) while > 90 % of adult donors are seropositive.
In Case Of Immunodeficient Individuals As Primary Immune Deficiency (PID), HIV Or SOT: The Absence Of EBV Specific T Lymphocytes (EBVSTS) Allows The Uncontrolled Expansion And Proliferation Of EBV Infected B Lymphocytes That Leads To EBV- Lymphoproliferative Diseases.
EBV Infection Is Then Associated With Many Diseases As PTLD, Hodgkin Lymphoma, Nk And T Cell Lymphoma, Burkitt’s Lymphoma, Nasopharyngeal And Gastric Carcinoma.
Most Of EBV-PTLD (90%) Are Of B Cell Origin.
Rituximab Is An Effective Monotherapy In Case Of HSCT (50-100 % Of Cases) And Effective In 50 % Of Sot Cases. In Children, Combination Of Rituximab, Steroids And Cyclophosphamide Is Effective In Treatment But It Increases The Risk Of Infections, Recurrence Of Disease & Severe Toxicity
Hence, The Need For The Development Of Medication That Addresses The Underlying Immune Defect With Little To No Toxicity
EBV specific T lymphocytes (EBVSTs) inhibit the entry of EBV infected B cells into the lymphoid follicles. In transplant recipients, due to weakened immune system, EBVST response is lacking, leading to uncontrolled proliferation of EBV-infected B cells and resultant EBV-associated post-transplant lymphoproliferative disorders (PTLD) and malignancies. So, infusion of EBVSTs help in reducing the latently infected B cell pool.
Autologous or allogeneic (including third party) source of EBVSTs are used to manufacture them using either ex vivo expansion or antigen-specific T cell selection interferon gamma capture).
Manufacturing EBV-specific T-lymphocytes (EBVST) Is an adoptive transfer of virus-specific T cell use of donor lymphocyte infusions (DLI) to donor-derived multi-antigen specific VSTs in the HSCT setting
It is produced from EBV-positive donors using good manufacturing grade, and the donor type could be;( autologous, allogeneic)
Methods used in manufacturing EBVST products
–Ex vivo expansion EBVSTs
–Antigen-specific T cell
Post-HSCT donor-derived T-cell therapy
1- Donor lymphocytic infusion
effective in remission but with attendant GVHD
2- Donor-derived EBVSTs
67% response rate but a very GVHD effect was seen among those treated with it
Autologous EBVSTs
used in HSCT and SOT recipients where their donors are not available
in theory, it is a preferred method used for PTLD
Delay in the production and administration of the drug because its autologous
Has been used in the treatment of type 11 latency EBV-associated malignancy
Third-party T cell
Designed to overcome the delay that may come with either autologous or allogenic methods of producing EBVSTs
It is a readily available off-the-shelf T-cell therapy product
Its response rate (CR and PR) was 64% at 5 weeks and 52% at 6 months with no significant toxicity observed
In the pediatric age group, it has a more favourable outcome
Many study groups have reported a significant reduction in EBV viral loads following treatment with third-party T cell
Modification of EBVSTs treatment to enhance the activity
Overcoming the immune suppressive effects of TGF- beta to enhance EBVSTs activity in vivo
Calcineurin resistance to enhance EBVSTs
Chimeric antigen receptor T-cell
In Summary :
The adoptive immunotherapy in the treatment of PTLD in SOT or HSTC especially the third- party T- cell has demonstrated a better clinical outcome with less toxicity and alleviating the concern of GVHD
What are the indications of adoptive immunotherapy?
-immunocompromised patient with EBV-negative lymphoproliferative disease
-Those that failed to respond to Rituximab monotherapy or plus steroid and
cyclophosphamide
-Treatment of viral infections that are difficult to cure like CMV, EBV.
–Immunocompromised patient with EBV-negative lymphoproliferative disease.
–Treatment of cancers that are caused by specific viruses, leukemias, DLBC lymphoma.
–Treatment of autoimmune diseases like lupus.
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
-Yes, in prevention of EBV related PTLD
–Treatment of cancers that are caused by specific viruses, leukemias, DLBC lymphoma.
–Treatment of autoimmune diseases like lupus.
What are the side effects of adoptive immunotherapy?
-Graft-versus-host disease
–Graft rejection
–IgE mediated anaphylactic reaction
–Neurotoxicity
–Cytokine release syndrome
*The article is a narrative review: Level of evidence – Level 5
I. Adoptive cellular immunotherapy for Epstein-Barr virus-associated lymphoproliferative disease
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
Adoptive immunotherapy basically involves isolating EBV specific T-lymphocytes (EBVSTs) from a patient with PTLD i.e., EBV-seropositive patients. These cells can be isolated from a resected tumor tissue or peripheral blood then they undergo ex vivo (in the laboratory) activation and expansion using good-manufacturing-practice (GMP)-compliant protocols. Thereafter these expanded effector cells are infused back into the patient in large numbers where they can now help the patient’s immune system fight/ destroy the cancer cells. Adoptive immunotherapy in PTLD is both therapeutic and prophylactic with 80% and 100% success respectively. The transferred EBVSTs can persist for up to nine years. Adoptive immunotherapy is reserved for EBV-positive patients who fail to respond to initial therapy.
What are the indications of adoptive immunotherapy?
– Therapeutic and prophylactic management of PTLD
– EBV-associated malignancies
– Refractory/ resistant disease
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
– EBV associated malignancies e.g., Hodgkin’s lymphoma, natural killer cell lymphoma, nasopharyngeal carcinoma
– Skin cancer
– Autoimmune disease
– Viral infections e.g., BK virus, CMV
What are the side effects of adoptive immunotherapy?
– Acute and chronic graft-versus-host-disease (GVHD)
– Cytokine storm (cytokine release syndrome) due to an overactive immune response leading to excessive inflammation
– Neurotoxicity due to inflammation of the brain
– Others: – AKI, graft rejection, bleeding episodes, arrhythmias, delirium, dizziness, edema, altered bowel patterns, hypotension, hypoxia, tremors, tachycardia, infections
Level of evidence – Level V
Introduction
Epstein Bar Virus is a world widely spread virus associated with lymphoprolifrative disease in SOT and hematopoietic stem cell transplant. It infects more than 90% of adult population. This virus tends to have
· Primary infection usually asymptomatic or infection mononucleosis like symptoms.
· Lifelong potency in B cells
· Progression to latency type I where EBV infected B cells would be invisible to EBSVTs.
· Uncontrolled type III latency
Donor lymphocyte infusion was first successful adoptive immunotherapy for EBV related PTLD though associated with risk of GVHD. Several studies has shown benefits of adoptive immunotherapy using EBVST “of the shelf”.
Clinical Experience
Adoptive immunotherapy using EBVSTs has shown good results in EBV related Lymphoprolifrative disorders.
EBVST Donor Options
· Seropositive donor with cryopreserved
· Autologous and allogeneic
EBVST manufacturing
· Ex-vivo expansion of VSTs Vs antigen specific
· MHC multimer selection( to isolate CD8+ T cell)
· Interferon –y capture approach
Combination therapy
Using genetic engineering technique to enhance EBVST.
Indications
EBVSTs have been used as an adjunctive therapy to chemotherapy in
· Hodgkin lymphoma
· NK/T Cell lymphoma
· Nasopharyngeal carcinoma
· Burkitt lymphoma
Adverse association
· Cytokine release syndrome
· AKI
· Rejection and autoimmunity
· Neurotoxicity
Level of evidence V
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
The scientific rationale for adoptive transfer of EBVSTs is based on harnessing the immunogenicity of type III latency malignancies to control the proliferation of the latently infected B cells. EBV-associated malignancies associated with type II latency [e.g., Hodgkin’s lymphoma (HL), natural killer (NK)/T-cell lymphoma, nasopharyngeal carcinoma (NPC)] or type I latency tumors (e.g., Burkitt lymphoma or gastric carcinoma) can develop in immunocompetent and immune deficient individuals and are much less immunogenic due to downregulation of the immunodominant (e.g., EBNA3 and EBNA2) antigens.
-The transformed B-cells in EBV-associated lymphoproliferative disease (EBV-LD)
associated with latency type III present several antigenic viral proteins including EBNA 1-3, LMP1 and LMP2 that induce potent EBVST responses. Such potent T cell immunity maintains the infected B cell pool at <2% of total B cells in immunocompetent individuals but is lacking in immunosuppressed individuals leading to uncontrolled lymphoproliferation of the infected B cells.
What are the indications of adoptive immunotherapy?
EBV-associated lymphoproliferative disease
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
It can be used as prevention of EBV related PTLD
What are the side effects of adoptive immunotherapy?
-Graft-versus-host disease
Well done, WHAT IS THE LEVEL OF EVIDENCE PROVIDED BY THIS ARTICLE?
SUMMARY:
· The gamma herpes virus family member Epstein-Barr virus (EBV) is common and linked to a number of lymphomas and lymphoproliferative diseases (LPD). More than 90% of adults globally are infected with it.
· Primary infection typically manifests in childhood as asymptomatic or mild clinical presentation or during adolescence as florid mononucleosis-like symptoms.
· It remains its potency in B cells or oral epithelial cells throughout life.
· EBV-infected naive B-lymphocyte nuclear antigens expressed proteins like EBNA1, EBNA2, EBNA3, and EBNALP, membrane proteins LMP1, and LMP2 as well as BARF1 and two small non-translated ribonucleic acids (RNA) (Type III latency)
· In healthy individuals, the rate of EBV infection remains stable at 0.1-50 EBV-infected B lymphocytes per 1,000,000 peripheral blood mononuclear cell (PBMCs).
· In individuals with weakened immune systems, such as patients with primary immunodeficiency (PID) or infection with human immunodeficiency virus (HIV), recipients of hematopoietic stem cell transplantation (HSCT) or solid organ transplant (SOT), the lack of a robust EBVST response can lead to uncontrolled proliferation of type 3 latency EBV-infected B cells resulting in EBV-associated LPD and malignancies.
· Manufacturing EBV-specific T-lymphocytes (EBVST) is an adoptive transfer of virus-specific T cell . The use of donor lymphocyte infusions (DLI) to donor-derived multi-antigen specific VSTs in the HSCT setting.
· Manufacturing EBVST products as ex vivo expansion EBVSTs with antigen-specific T cell.
· Autologous EBVSTs used in HSCT and SOT recipients where their donors are not available and it is a preferred method used for PTLD.
· Third-party T cell designed to overcome the delay that may come with either autologous or allogenic methods of producing EBVSTs and readily available “off the shelf” T-cell therapy product. Many studies have reported a significant reduction in EBV viral loads following treatment with third-party T cell.
· Modification of EBVSTs treatment to enhance the activity which overcome the immune suppressive effects of TGF- -β to enhance EBVSTs activity in vivo. Calcineurin resistance to enhance EBVSTs and Chimeric antigen receptor T cells.
· Combination therapy of EBVSTs with other therapeutic modalities in vivo as demethylating agents, checkpoint inhibitors and BCL-2 inhibitors.
· The use of adoptive immunotherapy, particularly using third-party “off the shelf” EBVSTs for the treatment of EBV-LD has shown promise in several studies conducted at specialized centers . More recently, EBVSTs have become more widely available including in industry led multi-center studies and consortium and cooperative group studies.
Indications for adoptive immunotherapy:
· Treatment of viral infections that are difficult to cure like CMV, EBV.
· Immunocompromised patient with EBV-negative lymphoproliferative disease.
· Treatment of cancers that are caused by specific viruses, leukemias, DLBC lymphoma.
· Treatment of autoimmune diseases like lupus.
Use of adoptive immunosuppressive for treatment of other conditions related to organ transplant:
· It has been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma, and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients.
· Can be used in BK virus nephropathy in transplant recepients, melonoma, EB virus associated malignancy.
Side effects
· Graft versus host disease
· Graft rejection
· IgE mediated anaphylactic reaction
· Neurotoxicity
· Cytokine release syndrome
Level of evidence: Level 5
Well done
1.Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
Current clinical experience with adoptive cellular therapy for EBV-LD:
EBVST donor options:
EBVST manufacturing strategies:
Combination therapies with other immunotherapeutic approaches:
==============================
2. What are the indications of adoptive immunotherapy?
==============================
3. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Yes
==============================
4. What are the side effects of adoptive immunotherapy?
5. Level of evidence V
Well done
Thankyou prof
SUMMARY
Introduction
Ebstein- Bar virus is a worldwide widely spread gamma herpes virus family that has been associated with lymphoproliferative disease in solid organ transplant and hematopoietic stem cell transplant. The virus infects more than 90% of the population and it has the following characteristics in the mode of operation.
Rituximab use, either as monotherapy or in combination with steroids or cyclophosphamide for the treatment of HSCT has been limited by high infection rate, recurrence of disease, and severe toxicity known with those medications.
Hence, the need for the development of medication that addresses the underlying immune defect with little to no toxicity
Manufacturing EBV-specific T-lymphocytes (EBVST)
– autologous
-allogeneic
Methods used in manufacturing EBVST products
Post-HSCT donor-derived T-cell therapy
a) Donor lymphocytic infusion
b) Donor-derived EBVSTs
Autologous EBVSTs
Third-party T cell
Modification of EBVSTs treatment to enhance the activity
Combination therapy of EBVSTs with other therapeutic modalities
Conclusion
The adoptive immunotherapy in the treatment of PTLD in SOT or HSTC especially the third- party T- cell has demonstrated a better clinical outcome with less toxicity and alleviating the concern of GVHD
Indications for adoptive immunotherapy
Use of adoptive immunosuppressive for treatment of other conditions related to organ transplant
Side effects
Level of evidence
That is a very detailed summary and very well structured reply.
I like your analysis of level of evidence of this study.
Principles of adoptive immunotherapy:
In healthy seropositive individuals, virus neutralizing Ab control the spread of the infectious virus particles and the EBV specific HLA class I restricted, CD8 T lymphocytes specific to the early lytic cycle proteins kill cells entering the lytic cycle before they are able to release infectious virus particles
EBV maintains a lifelong latency in B cells and oral epithelial cells. CD 21 found on the B cells is the major receptor for the virus. EBV infected naive B lymphocytes express proteins comprising the EBV genome including:
This is type III latency pattern
The EBV infected B lymphocytes then enter the lymphoid follicles and downregulate the immunogenic proteins to express less immunogenic type II latency proteins – EBNA 1, LMP1 and LMP2. This is termed as latency type II pattern
The virus persists in latently infected B cells by further downregulating expression of viral proteins – latency type I – so as to become invisible to EBSVTs
In individuals with a weakened immune system, the lack of a robust EBSVT response can lead to uncontrolled proliferation of type III latent EBV infected B cells resulting in EBV associated LPDs and malignancies
The latent type III B cells that cause LPDs present several antigenic viral proteins including EBNA 1-3, and LMP 1 and 2 that induce potent EBSVT response. Such potent T cell immunity maintains the infected B cell pool at less than 2% of total B cells in immunocompetent individuals but is lacking in immunosuppressed individuals leading to uncontrolled proliferation of the infected B cells.
The scientific rationale for the adoptive transfer of EBSVTs id based on harnessing the immunogenicity of type III latent B cells to control the proliferation of these cells that cause the LPDs and other malignancies.
EBV associated malignancies associated with type II latency (Hodgkins lymphoma, NK/T cell lymphoma) or type I latency (Burkitts lymphoma or gastric carcinoma) can develop in immunocompetent and immunosuppressed individuals and are much less immunogenic due to downregulation of the immunodominant Ag.
Manufacturing of EBSVTs will require lymphocytes which can be:
There are different ways to manufacture the EBSVTs:
There are several ways of enhancing the activity of EBSVTs:
One can also administer EBSVTs with other therapeutic modalities to enhance EBSVT activity in vivo:
Indications of Adoptive Immunotherapy:
Other Uses Of Adoptive Immunotherapy In Organ Transplant:
In GVHD disease (although it is one of the adverse effects of this therapy
Infections post transplant like CMV that is resistant to antiviral treatment
Side Effects:
That is an excellent summary and very well structured reply. I wish you could type the heading of first para as Introduction of Principles of Immunotherapy, and then you could type sub-headings in bold or underline or in italics. That would make it easier to read.
What is your analysis of level of evidence of this study?
The principle of adoptive immunotherapy in the treatment of PTLD:
a) Ex vivo expansion of VSTs:
1) utilizing irradiated EBV transformed lymphoblastoid cell lines (LCL).
2) To enhance the specificities to less immunogenic EBV antigens by transducing DC and LCLs with adenovirus vectors to overexpress LMP1 or LMP2.
3) The novel antigen-presenting complex (KATpx)] can facilitate the rapid (10–21 days as opposed to 2–3 months) expansion of T cells targeting EBNA1, LMP1 and LMP2 from healthy donors and from patients with type 2 latency EBV-associated malignancies.
b) Antigen-specific T cell selection [e.g., interferon γ (IFN-γ) capture].
1) major histocompatibility complex (MHC) multimer selection where oligomeric forms of MHC molecules are designed and conjugated to magnetic beads to isolate typically CD8+ T cells with high affinity to a specific viral epitope/peptide in an HLA restricted manner.
2) IFN-γ capture approaches where mononuclear cells are pulsed with antigen to isolate CD8+ and CD4+ T cells which secrete IFN-γ in response to the viral antigens.
c) Post-HSCT donor-derived T cell therapy
1) Donor lymphocyte infusions.
2) Donor-derived EBVSTs.
d) Autologous EBVSTs: has been used for patients with EBV-associated malignancies outside the context of allogeneic HSCT and in SOT recipients with PTLD where donors usually are not available because of the use of cadaveric grafts.
e) Third-party EBVSTs: a readily available “off the shelf” T cell therapy product is desirable as patient-specific products may not be possible or may be so delayed.
f) Modification of EBVSTs to enhance activity
1) Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo.
2) Calcineurin resistance to enhance EBVSTs: Given the concerns regarding EBVST persistence in patients who require ongoing immune suppression.
3) Chimeric antigen receptor T cells: using CD19-chimeric antigen T cells (CD19-CART).
g) Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance EBVST activity in vivo
1) Demethylating agents; Azacytidine and decitabine are potent CpG demethylating agents.
2) Checkpoint inhibitors the risk of graft rejection and autoimmunity limits their use in that setting.
3) BCL-2 inhibitors; BCL-2 inhibitor induce remission in approximately 70% of mice PTLD xenografts.
Indications of adoptive immunotherapy:
Immunotherapy is used to upregulate or downregulate the immune system to achieve a therapeutic effect in immunological mediated disorders:
· Hyper-sensitivity reaction.
· Auto-immune diseases.
· Tissue and organ transplantation.
· Malignancies; Type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients.
· Inflammatory disorders.
· Infectious diseases.
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Autologous EBVSTs have also been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients.
The side effects of adoptive immunotherapy:
1) Donor lymphocyte infusions carries a significant risk of graft-versus-host disease (GVHD).
2) Selective depletion of T-regulatory cells (Tregs) prior to infusion enhances the graft versus lymphoma (GVL) effect.
3) Serious adverse events in relation to third-party EBVSTs are TMA and GI hemorrhage(all deemed unrelated).
4) Adverse effects in relation to CART therapy include cytokine release syndrome (CRS), neurotoxicity and acute kidney injury.
5) Checkpoint inhibitors: the risk of graft rejection and autoimmunity limits their use.
That is an excellent summary and very well structured reply.
What is your analysis of level of evidence of this study?
Briefly summarize the principle of adoptive immunotherapy in the treatment of PTLD.
Adaptive immunotherapy involves isolation of specific T lymphocytes which can be used to target infection and malignancies. Due to immunosuppressive medication in post-transplant patients, the cytotoxic T lymphocyte response is weakened, which leads to uncontrolled proliferation of cancerous cells and viral infections such EBV. Adaptive immunotherapy can be used as means of prevention or treatment against EBV and PTLD in transplant recipients.
What are the indications of adoptive immunotherapy?
It can be used for the treatment and prophylaxis of EBV related PTLD in transplant patients. And also, in treatment of type 2 latency EBV associated cancers in relapsed patients.
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Skin cancers, melanoma,
EBV associated PTLD.
BK virus nephropathy
What are the side effects of adoptive immunotherapy?
Neurotoxicity,
Acute Kidney Injury
GVHD, TMA
Cytokine release syndrome.
GIT hemorrhage,
IgE mediated anaphylactic reaction.
That is an excellent summary and very well structured reply.
What is your analysis of level of evidence of this study?
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
Adoptive immunotherapy involves isolation of T lymphocytes and then using then to treat cancers or other diseases. Antibodies which neutralise viral particles can control spread of viral disease. also EBV specific cytotoxic T lymphocytes can kill cells before release of viral particles. EBVSTs- EBV specific T lymphocytes are important in immune response. In transplant recipient , EBVSTs response is weak due to weak immune system . This can cause proliferation of B cells. This will lead to PTLD.
The scientific rationale for adoptive transfer of EBVSTs is based on harnessing the immunogenicity of type III latency malignancies to control the proliferation of the latently infected B cells.
What are the indications of adoptive immunotherapy?
It can be used in Type 2 latency EBV associated malignancy with chemotherapy or relapse.
It is also used in prophylaxis of EBV related PTLD
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
It can be used in BK virus nephropathy in transplant recepients, melonoma, EB virus associated malignancy.
What are the side effects of adoptive immunotherapy?
These include –
Neurotoxicity, GVHD, AKI, Cytokine release syndrome. GIT haemorrhage, Thrombotic microangiopathy, Fever , Odema, Uveitis, IgE mediated anaphylactic reaction.
That is an excellent summary and very well structured reply.
What is your analysis of level of evidence of this study?
Level of evidence is V
Adoptive immunotherapy involves isolation and infusion of T lymphocytes in a patient for treatment/ prevention of a disease. Epstein-Barr virus (EBV) infects more than 90% of adult population. Virus neutralizing antibodies control spread of infectious viral particles and EBV-specific CD8+ cytotoxic T lymphocytes kill cells before they release viral particles.
EBV specific T lymphocytes (EBVSTs) inhibit the entry of EBV infected B cells into the lymphoid follicles. In transplant recipients, due to weakened immune system, EBVST response is lacking, leading to uncontrolled proliferation of EBV-infected B cells and resultant EBV-associated post-transplant lymphoproliferative disorders (PTLD) and malignancies. So, infusion of EBVSTs help in reducing the latently infected B cell pool.
Autologous or allogeneic (including third party) source of EBVSTs are used to manufacture them using either ex vivo expansion or antigen-specific T cell selection )interferon gamma capture).
2. What are the indications of adoptive immunotherapy?
Adoptive immunotherapy is indicated in prevention and treatment of EBV-associated lymphoproliferative disease, and in patients with elevated EBV viral load.
3. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Adoptive immunotherapy has been utilized in BK virus nephropathy in transplant recipients (1). They have also been utilized in melanoma.
4. What are the side effects of adoptive immunotherapy?
Side effects of adoptive immunotherapy include GVHD (graft versus host disease), acute kidney injury, gastrointestinal hemorrhage, and thrombotic microangiopathy. Chills, fever, hypotension, oliguria, edema (due to capillary leak syndrome), uveitis, cytokine release syndrome, confusion, delirium, seizures, cerebral edema, and IgE mediated anaphylactic reaction have been also seen with them (2).
The article is a narrative review: Level of evidence – Level 5
References:
1) Jahan S, Scuderi C, Francis L, Neller MA, Rehan S, Crooks P, Ambalathingal GR, Smith C, Khanna R, John GT. T-cell adoptive immunotherapy for BK nephropathy in renal transplantation. Transpl Infect Dis. 2020 Dec;22(6):e13399. doi: 10.1111/tid.13399. Epub 2020 Jul 14. PMID: 32608543; PMCID: PMC7816252.
2) Rohaan MW, Wilgenhof S, Haanen JBAG. Adoptive cellular therapies: the current landscape. Virchows Arch. 2019 Apr;474(4):449-461. doi: 10.1007/s00428-018-2484-0. Epub 2018 Nov 23. PMID: 30470934; PMCID: PMC6447513.
That is a very detailed summary and very well structured reply.
I like your analysis of level of evidence of this study.
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD.
EBV infects more than 90% of the adult population. Primary infection mainly occurs in childhood leading to either asymptomatic/mild infection/infectious mononucleosis.
EBV enters the oropharynx replicating in the epithelial cells and infecting transiting B lymphocytes. The infected B lymphocytes express proteins representing the whole EBV genome. They then enter the lymphoid follicles where they down regulate the immunogenic proteins to express less immunogenic type 2 latency proteins. They thus enter the memory compartment where the virus persist in latently infected B lymphocytes.
The frequency of EBV infected B lymphocytes remain stable in a healthy individual. However, in an immunocompromised individual, lack of a robust EBV specific T lymphocytes leads to uncontrolled proliferation of type 3 latency EBV leading to LPD and malignancies.
The rationale of adoptive immune transfer is based on harnessing the immunogenicity of type 3 latency malignancies to control the proliferation of latently infected B cells.
What are the indications of adoptive immunotherapy?
Other conditions
Type 2 latency EBV associated malignancy
Skin cancer-melanoma
Side effects
GVHD
Cytokine release syndrome
Acute kidney injury
Neurotoxicity
That is an excellent summary and very well structured reply.
What is your analysis of level of evidence of this study?
Thankyou.
This was a narrative review hence level 5
1-Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
Introduction:
– EBV-positive PTLD is a major complication of HSCT and SOT.
– EBV-infected naïve B-lymphocytes express proteins comprising the entire EBV genome
– After primary EBV infection, most viral antigens are expressed during the lytic stage while the virus is replicating.
– In EBV-infected B-cells, there is subsequent downregulation of viral antigen expression from latency type III to latency type 0 which allows escape from immune surveillance.
-Periodic expansion of EBV-infected B-cells requires re-expression of viral antigens which re-stimulates the EBVST response.
– The lack of a robust EBVST response can lead to uncontrolled proliferation of type 3 latency EBV-infected B cells resulting in EBV-associated LPD and malignancies.
– PTLD is highly immunogenic and amenable to immunotherapy with EBVSTs.
-The scientific rational to adoptive transfer of EBVSTs is based on harnessing the immunogenicity of type III latency malignancies to control the proliferation of the latently infected B cells.
Manufacturing of EBVSTs
It can be produced from EBV-positive donors using good-manufacturing-grade (GMP) compliant methodologies with minimal alloreactivity and broad specificity against EBV latency proteins or as a multi-virus specific product with activity against multiple viruses.
Donor types; autologous and allogeneic (including third party).
The most common strategies:
Ex vivo expansion of EBVSTs:
-Utilizing irradiated EBV transformed lymphoblastoid cell lines (LCL) as antigen presenting cells (APCs) to selectively expand EBVSTs.
– LCL-activated EBVSTs consist of a product with activity against early lytic antigens and EBNA 3A, 3B and 3C but unreliable activity towards LMP1 and LMP2.
– To enhance the specificities to less immunogenic EBV antigens LMP1 and LMP2, transducing DC and LCLs with adenovirus vectors to overexpress LMP1 or LMP2.
– Utilize whole antigen
Antigen-specific T cell selection: (IFN-γ) capture
– Selection and isolation of CD8+ T cells with high affinity to a specific viral epitope/peptide in an HLA restricted manner.
– Mononuclear cells are pulsed with antigen to isolate CD8+ and CD4+ T cells which secrete IFN-γ in response to the viral antigens.
– Require seropositive donors with high frequency of circulating antigen/epitope specific T cells.
Post-HSCT donor-derived T cell therapy
Donor lymphocyte infusions DLI.
-Utilized unmodified DLI derived from the patient’s EBV seropositive HSCT donor which contained effector cells with activity against EBV.
– Carries a significant risk of GVHD, selective depletion of T-regulatory cells (Tregs) reduces the risk.
Donor-derived EBVSTs
-Using donor derived EBVSTs for the prevention and treatment of EBV+ PTLD in the post-HSCT setting when the donor is available
Autologous EBVSTs
– Used for patients with EBV-associated malignancies outside the context of allogeneic HSCT and in SOT recipients with PTLD.
– In SOT it is preferred to donor EBVSTs even if available
– The challenges in production and the production time leading to delays in initiation of therapy in patients with a rapidly progressive disease.
– Used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients.
– Such tumors are more challenging targets because of the reduced expression of immunogenic viral antigens which express a more restricted array of antigens compared to type III.
– Better responses were observed with EBVST products that included T cells with activity against LMP1 and LMP2.
Third-party T cells
– Readily available T cell therapy; EBVST bank with HLA alleles prevalent at high frequencies in individuals community.
– No significant toxicities were observed, alleviating concerns of graft rejection and/or GVHD with this therapy which fueled the broadened applicability of this approach.
– Multiple studies used third-party multi-VSTs for EBV-associated PTLD, no significant adverse events attributable to
the product were observed.
– Third-party EBVSTs have been mostly used in the posttransplant setting.
– No published experience in patients with HIV-associated lymphomas.
Modification of EBVSTs to enhance activity
The efficacy of EBVSTs is dependent on the expression of viral antigens and limited by the paucity of EBV antigen expression in malignancies of latency type I and II.
Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo
– In the immunocompetent host, TGF-β can be released into the tumor microenvironment by tumor cells, fibroblasts and immune cells and creates an immunosuppressive environment by impeding T-cell activation and it affects antigen presenting cells and suppresses functional antitumor T cell responses in vivo
– The potential to overcome the immunosuppressive properties of TGF-β has been studied in EBV-positive HL.
Calcineurin resistance to enhance EBVSTs;
-As there is a concerns regarding EBVST persistence in patients who require ongoing immune suppression SOT, EBVSTs were genetically engineered to express a CNI-resistant EBVSTs than can persisted with enhanced activity in the presence of CNI.
Chimeric antigen receptor T cells:
-CD19-chimeric antigen T cells (CD19-CART) have shown impressive efficacy in acute B-lymphoblastic leukemia and
in B-cell lymphomas. In addition to viral antigens, EBV-LD expresses a variety of B-cell antigens targetable by
CARTs including CD19, CD20 and CD30.
-However, their use in EBV-lymphoproliferation is limited by the patient’s immunosuppressive state impeding T cell manufacture and the length of production time.
Combination strategies :EBVSTs with other modalities to enhance EBVST activity in vivo
Demethylating agents:
– Newly EBV-infected B-cells express viral genes. Rapid CpG-methylation of viral antigens leads to downregulation of viral protein expression and the latency.
Checkpoint inhibitors
-EBV-positive diffuse large B-cell lymphoma (DLBCL) and EBV-positive PTLD were found to express PD-L1.
– LMP1 induce expression of the checkpoint protein PD-L1
– Using checkpoint PD-L1 blockade.
BCL-2 inhibitors
-Latently EBV-infected cells inhibit proapoptotic signals thus ensuring immortality.
– LMP1 upregulates the expression of the anti-apoptotic protein BCL-2.
– BCL-2 inhibitor increased proapoptotic proteins and sensitivity to CD19-CART;
Conclusion:
-The use of third-party EBVSTs for the treatment of EBV-LD has shown promise in several studies.
– Further Preclinical work and early clinical trials is needed.
2-What are the indications of adoptive immunotherapy?
-Treatment and prophylaxis of PTLD in transplant recipients HSCT, SOT.
-In refractory cases.
-EBV- associated malignancies.
3-Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
-Skin cancer; metastatic melanoma.
– EBV-associated malignancies.
– Autoimmune diseases.
4-What are the side effects of adoptive immunotherapy?
-Most people have a mild form of cytokine release syndrome. But in some people, it may be severe or life-threatening.
– GVHD.
– Capillary leak syndrome.
– Fever.
– Flue like illness.
– Skin rash.
– Neurotoxicity
– Gastrointestinal symptoms; diarrhea, nausea, vomiting.
– Acute kidney injury.
–Graft rejection and autoimmune complications.
– TMA, GI hemorrhage.
5-Level of evidence; narrative review level 5.
That is a very detailed summary and very well structured reply.
I like your analysis of level of evidence of this study.
Briefly summaries the principle of adoptive immunotherapy in the treatment of PTLD
Introduction:
EBV, a common viral infection in childhood, affecting 90% of the adult population.
Normally the immune system contains the virus and protect the body against spreading of this infection, this causes EBV to undergo into a dormant phase called as latent phase in the B cells.
But all the patients with the defective immune system, i.e., HIV. HSCT. SOT.
can lead to uncontrolled infection and proliferation which can lead to EBV- associated PTLD and malignancies.
The transformed B-cells in EBV-associated Lymphoproliferative Disease associated with type III latency, present Viral proteins: EBNA 1-3. LMP1 and 2 , such antigens in intact immune systems, T-cells can control B cells and maintain infected B cells in the range of < 2%.
Type II latency associated with malignancies: NHL, Hodgkins’s lymphoma. Natural killer T-cell lymphoma. Nasopharyngeal carcinoma, NPC.
Type I latency tumors: Burkitt’s lymphoma. Gastric carcinoma.
The occurrence of PTLD in HSCT ranges from 1-25%, depending on the following:
Sero negativity in a transplant recipient is the top determinant factor of the development of post-transplant LPD. PTLDs is usually seen more commonly in pediatric patients.
Monotherapy with Rituximab anti-CD20 is more effective in HSCT with a response rate of 55 -100%, while combination therapy (prednisolone/cyclophosphamide/rituximab) is shown to better survival up to 725.
#EBVSTs manufacturing:
#Post-HSCT donor-derived T-cell therapy:
# Autologous EBVSTs.
# Third-party T cells.
the purpose of Modification of EBVST is to enhance activity of the immune suppressive effects of TGF-B in vivo, Calcineurin resistance to enhance EBVSTs and Chimeric antigen receptor T cells.
different Combination strategies are adopted, with EBVSTs which for example could be use of Demethylating agents, use of Checkpoint inhibitors, use of BCL-2 inhibitors.
Conclusion:
Indication of adoptive immunotherapy:
Immunocompromised recipient with EBV-PTLD. and Prophylactic treatment in reduction of EBV load.
Uses of adoptive immunotherapy other than cancer treatment:
Side effects:
That is an excellent summary and very well structured reply.
What is your analysis of level of evidence of this study?
Summary
PTLD is a major complication of solid organ donation caused by childhood EBV viral activation. EBV infects more than 90% of the adult population worldwide. PTLD is classified into B cell lymphoma (90%) and T cell lymphoma accounts for 10%.
EBV infects B cells in two key stages after primary infection:
Lytic stage, where the patient may be asymptomatic or develop mild symptoms or frank infectious mononucleosis; latency stage, where the virus becomes latent in B cells lifelong and the patient is asymptomatic but at risk of developing certain lymphoproliferative diseases; and several stages of latency develop (Latency III, II, I).
During latency III, EBV-infected B-lymphocytes express the EBV nuclear antigens EBNA1, 2, 3, EBNALP, LMP1, 2, and BARF1. Due to excessive antigenic expression, T lymphocytes restrict B cell proliferation to 2%. Because immunosuppression leads B lymphocytes to proliferate uncontrollably, the Latency III stage is connected with lymphoproliferative diseases including cancer. The most prevalent malignancy at this stage is PTLD.
In Latency II and I, EBNA3 and EBNA2 are downregulated, rendering them less immunogenic. Latency I and II stages are associated with lymphoproliferative diseases and cancers in immune-competent and immune-compromised patients, such as Hodgkin’s lymphoma, natural killer/T-cell lymphoma, nasopharyngeal carcinoma, Burkitt lymphoma, and gastric carcinoma (Latency I).
In PTLD adoptive immunotherapy, EBV-specific cytotoxic T lymphocytes or donor lymphocyte infusion kill dividing EBV-positive B cells.
What are the indications of adoptive immunotherapy?
Treatment of EBV+ PTLD cases that are resistant or refractory
Treatment of some solid tumors, such as glioblastoma and GIT tumors
Treg adoptive immunotherapy is used to treat certain chronic viral infections such as HIV, as well as autoimmune diseases such as type I DM and SLE.
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Through the infusion of Treg cells, adoptive immunotherapy can be used to treat acute rejection refractory to standardized treatment.
Adoptive therapy can also be used in the treatment of autoimmune diseases such as SLE and viral infections such as BK virus and CMV infection.
What are the side effects of adoptive immunotherapy?
The adverse effects of adoptive treatment depend on the type of cells employed and the reason for infusion. The following are typical adverse effects: constitutional symptoms during infusion is common, Chronic and acute graft-versus-host disease, Cytokine storm, Neurotoxicity
That is an excellent summary and very well structured reply. I wish you could type the heading of first para as Introduction. Under this main heading, I wish you could type sub-headings in bold or underline or in italics. That would make it easier to read.
What is your analysis of level of evidence of this study?
Introduction:
Epstein-Barr virus (EBV) is a member of gamma herpes family associated with various lymphomas and lymphoproliferative disorders (LPD).
It infects more than 90% of the adult population worldwide.
Primary infection occurs in childhood as an asymptomatic or mild infection and may result in a more florid infectious mononucleosis syndrome in teenagers and young adults
In healthy seropositive individuals:
Virus-neutralizing antibodies control the spread of infectious virus particles, and EBV-specific, human leukocyte antigen (HLA) class I restricted, CD8+ cytotoxic T lymphocytes (CTL) specific to the early lytic cycle proteins kill cells entering the lytic cycle before they can release infectious virus particles
In individuals with weakened immune systems,
Such as patients with primary immunodeficiency (PID) or infection with human immunodeficiency virus (HIV), recipients of hematopoietic stem cell transplantation (HSCT) or solid organ transplant (SOT), the lack of a robust EBVST response can lead to uncontrolled proliferation of type 3 latency EBV-infected B cells resulting in EBV-associated LPD and malignancies.
EBV-associated malignancies associated with :
Type II latency [e.g., Hodgkin’s lymphoma (HL), natural killer (NK)/T-cell lymphoma, nasopharyngeal carcinoma (NPC)]
Type I latency tumors (e.g., Burkitt lymphoma or gastric carcinoma) can develop in immunocompetent, and immune deficient individuals and are much less immunogenic due to downregulation of the immunodominant (e.g., EBNA3 and EBNA2) antigens
Type 3 latency EBV-infected B cells resulting in EBV-associated LPD and malignancies
Manufacturing of EBVSTs:
Donor types include :
Autologous
Allogeneic (including third-party) sources.
Ex vivo expansion of EBVSTs:
GMP-grade EBV-specific T-cell manufacturing. Mononuclear cells are harvested from the peripheral blood of a donor. B-lymphocytes are infected with laboratory strain EBV, transduced with an adenoviral vector expressing LMP1 and LMP2, and irradiated. Next, monocytes are separately transduced with the same adenoviral vector and cocultured with T cell, followed by a second stimulation by the transduced LCL to create and further expand the LMP-specific T-cell product.
Antigen-specific T-cell selection:
Post-HSCT donor-derived T-cell therapy:
Ø Remissions,
Ø This therapy carries a significant risk of graft-versus-host disease (GVHD)
That is an excellent summary and very well structured reply.
What is your analysis of level of evidence of this study?
this article elucidates the novel therapy used for EBV related post-transplant lymphoproliferative disease EBV-LPD.
EBV:
It’s an extensively common virus belongs to herpes virus family. It was found to be linked to variable types of lymphomas and lymphoproliferative disorders LPD. its prevalent in 90% of adult people. Its inhabitant of oropharyngeal epithelial cells and infecting trafficking B-lymphocytes as they have a specific receptor for EBV, that is CD21.
Post transplant LPD :
Part of pre transplant preparation is ascertaining EBV serology status of donor and recipient, as it’s a common risk factor for post transplant LPD , particularly in the context of donor positive recipient negative serology for EBV.
in the context of competent immune system T lymphocytes recognized B lymphocytes infected with EBV, as those B lymphocytes are expressing EBV related protein on surface membrane, full virus genome is expressed in B-lymphocytes surface membrane type III latency, some Lymphocytes might undergo downregulation of antigen presentation type II latency , hence the percentage of B-lymphocytes infected with EBV is generally maintained at maximum of 2 % of total B lymphocytes, a status that is entirely deranged in immunocompromised condition such as HIV infected patients, and post transplant status where B lymphocytes proliferate out of control resultant in PTLD.
Type III latency associated tumors:
PTLD
Type II latency associated tumors:
Hodgkins Lymphoma and nasopharyngeal carcinoma.
Type 1 latency associated tumors:
Burkitts lymphoma and gastric carcinoma.
EBVSTs:
those T lymphocytes are sensitive to EBV infected B lymphocytes and tagged as EBVSTs. This clone of T lymphocytes made the basics for adoptive transfer of EBVSTs to the patient with PTLD to control unchecked proliferation of B lymphocytes.
What are the conclusions of your summary, Dr Wael?
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD:
Introduction:
Epstein-Barr virus – a widespread (almost more than 90% of the population infected) gamma herpes virus family, associated with lymphomas and lymphoproliferative diseases.
Childs usually asymptomatic or with mild symptoms, adolescents may have severe infectious mononucleosis syndrome.
in seropositive people the antibodies control the spread of virus by various immune system actios:
– Cytotoxic T cells kills the lysed cells before releasing the viral particles.
– EBV remains latents in B cells and oral epithelial cells.
– These infected B cells can rescue the virus by memory cells and down regulate the viral expression so the cytotoxic T (EBV specific T lymphocytes) cells cannot identify and attack it by various actions.
Breakdown of this immune regulation, due to immune deficiencies – inherited or acquired (ie HIV, Immunosuppressive medications) results in activation of virus, and leads to a lymphoproliferative disorders and lymphoma.
Type I latency tumors are:
Burkitt’s lymphoma.Gastric carcinoma.
Type II latency associated with malignancies are:
HL, Hodgkins’s lymphoma, Natural killer T-cell lymphoma, and Nasopharyngeal carcinoma, NPC.
Type III latency associated with PTLD: due to presenting cell proteins (EBNA1-3, LMP, and LMP2).
1-25% of hematopiotic stem cell transplant patients experience a PTLD through the following:
– The degree of patients T cell depletion.
– Seronegativity status.
– The immunosuppressant’s.
PTLD is highly immunogenic and amenable to immunotherapy with EBVSTs. Adoptively transferred virus-specific T cells (VSTs) have been evaluated for more than two decades, ranging from the use of donor lymphocyte infusions (DLI) to donor derived multi-antigen specific VSTs in the HSCT setting to autologous as well as allogeneic VSTs in the SOT setting.
Manufacturing of EBVSTs:
– Ex vivo expansion of EBVSTs.
– Antigen-specific T cell selection.
Post-HSCT donor-derived T-cell therapy:
– Donor Lymphocyte infusion.
– Donor-derived EBVSTs.
Autologous EBVSTs.
Third-party T cells.
Modification of EBVSTs to enhance activity:
Combination strategy, with EBVSTs:
Conclusion:
Use of adoptive immunotherapy using a third party “off the shelf” EBVSTs for the treatment of EBV associated LD is promising with limited use in specialized centers, that should be widely available.
EBVSTs anti-tumor activity for type I and type II latency associated tumors should be explored.
Level of evidence is V erratic review.
What are the indications of adoptive immunotherapy?
Treatment and prevention of lymphoproliferative disorders in immunosuppressed patients.
Reduction in EBV viral load.
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Can be used to control other viral infection such as CMV, adenovirus.. etc
Can be used to prevent viral induced skin diseases or cancers.
Can be used to treat malignancies other than PTLD. Example glioblastoma and hepatic cancers.
Can be used in treating vasculitis and connective tissue disease.
What are the side effects of adoptive immunotherapy?
Graft versus host disease.
Allergic reaction, flue like illness, fever, palpitation and shortness of breath.
Neurotoxicity.
Cytokine release syndrome.
That is an excellent summary and very well structured reply.
What is your analysis of level of evidence of this study?
level of evidence V – erratic review
1- Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD:-
====================================================================
Introduction
====================================================================
Manufacturing of EBVSTs
====================================================================
Ex vivo expansion of EBVSTs
=================================================================
Antigen-specific T cell selection
====================================================================
Post-HSCT donor-derived T cell therapy
Donor lymphocyte infusions
Donor-derived EBVSTs
Autologous EBVSTs
Third-party T cells
====================================================================
Modification of EBVSTs to enhance activ
Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo
Calcineurin resistance to enhance EBVSTs
Chimeric antigen receptor T cells
====================================================================
Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance EBVST activity in vivo
Demethylating agents
Checkpoint inhibitors
BCL-2 inhibitors
====================================================================
Conclusions
====================================================================
2- What are the indications of adoptive immunotherapy?
1- If conventional tratment fails.
2- It uses EBV-specific cytotoxic T lymphocytes(EBV-CTLs)or donor lymphocyte infusion(DLI)in attempt to about the EBV-driven proliferation of B cells in EBV -association PTLD.
3- Prevention and remmision of EBV-induced PTLD have been achieved in 52 to 75% of pateint .
====================================================================
3- an adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
1- Adoptive cellular treatment using EBVSTs has produced outstanding outcomes in clinical trials for immunocompromised patients with EBV-LD, and there is potential for wider application.
2- The focus of current research is on genetic engineering and combination therapy to increase EBVST potency in vivo.
====================================================================
4- What are the side effects of adoptive immunotherapy?
1- Acute and chronic graft -versus-host disease(GVHD).
2- DLI may produce GVHD,which dose not appear to be problem with EBV-CTLs.
1. Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
EBV maintains a lifelong latency in B cells and oral epithelial cells.
During primary infection, EBV enters the oropharynx replicating within the epithelial cells and infect B-lymphocytes subsequently it undergoes several steps from lytic, latency III to latency 0.
each step can result in different types of malignancies
· lytic none
· latency III PTLD, HIV lymphoma
· latency II hodgkins lymphoma, DLBCL, diffuse large B-cell lymphoma; NK, T cell lymphoma, nasopharangeal carcinoma
· latency I Burkitt lymphoma, Gastric carcinoma
· latency 0 none
EBV-infected B-lymphocytes in latency III express proteins EBNA1, EBNA2, EBNA3, and EBNALP, LMP1, and LMP2, BARF1. then it enters the lymphoid follicles and downregulate the immunogenic proteins to express less immunogenic proteins and save them into the memory compartment then it continue to further downregulating expression of viral proteins so as to become invisible to EBV-specific T-lymphocytes (EBVSTs).
In individuals with weakened immune systems, such as
(I) patients with primary immunodeficiency (PID) or
(II) infection with human immunodeficiency virus (HIV),
(III) recipients of hematopoietic stem cell transplantation (HSCT) or
(IV) solid organ transplant (SOT),
the lack of a robust EBVST response can lead to uncontrolled proliferation of type 3 latency EBV-infected B cells resulting in
· EBV-associated LPD and
· Malignancies
The scientific rationale for adoptive transfer of EBVSTs is based on harnessing the immunogenicity of type III latency malignancies to control the proliferation of the latently infected B cells.
2. What are the indications of adoptive immunotherapy?
Either prophylaxis or treatment of EBV-associated LPD and malignancies in the following populations:
· patients with chronic immunosuppression
· against EBV latency proteins
· as a multi-virus specific product with activity against multiple viruses
· patients with primary immunodeficiency (PID)
· infection with human immunodeficiency virus (HIV)
· recipients of hematopoietic stem cell transplantation (HSCT)
· solid organ transplant (SOT)
· post-HSCT (33) or SOT (13) and PTLD
3. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients.
refractory PTLD
treatment of multi-VSTs [EBV, cytomegalovirus (CMV), adenovirus, +/− BK virus (BKV) and human herpes virus 6 (HHV6)] products
human B-cell lymphoma
acute B-lymphoblastic leukemia
4. What are the side effects of adoptive immunotherapy?
skin GVHD
TMA
GI hemorrhage
Well done.
Adoptive cellular immunotherapy for EBV-associated PTLD
Introduction:
· EBV associated PTLD is a well described complication after both hematopoietic stem cell (HPSC) and solid organ transplantation (SOT) in about 1-25% of cases.
· The risk of PTLD is dependent on the EBV serostatus of the donor (higher risk with positive donor and negative recipient) and the degree of immunosuppression (higher risk with ATG induction and TAC based maintenance therapy).
· Risk of EBV-PTLD is higher in pediatric SOT (as they are transplanted early before acquiring infection) while > 90 % of adult donors are seropositive.
· EBV is one of herpes virus family and present in >90 % of adult population. It is either asymptomatic if acquired in early childhood or manifests as IMN when acquired in adolescents.
· The degree of viral replication and antigen presentation is maximal during the lytic stage then decrease with time in latency stage as occurs in healthy individuals with intact immune surveillance.
· Usually lytic stage is asymptomatic or causing IMN, while latency phase is associated with risk of LPD.
· After the initial infection and viral replications, it remains dormant in B cell and oral epithelial cells.
· In case of immunodeficient individuals as primary immune deficiency (PID), HIV or SOT: the absence of EBV specific T lymphocytes (EBVSTs) allows the uncontrolled expansion and proliferation of EBV infected B lymphocytes that leads to EBV- lymphoproliferative diseases.
· EBV infection is then associated with many diseases as PTLD, Hodgkin lymphoma, NK and T cell lymphoma, Burkitt’s lymphoma, nasopharyngeal and gastric carcinoma.
· Most of EBV-PTLD (90%) are of B cell origin.
· Rituximab is an effective monotherapy in case of HSCT (50-100 % of cases) and effective in 50 % of SOT cases. In children, combination of rituximab, steroids and cyclophosphamide is effective in treatment but it increases the risk of infections,
1. Summarize principles of adaptive immunotherapy
· Immunotherapy includes:
1. Use of Donor lymphocytes infusion (derived from EBV seropositive donors) was utilized in treatment of PTLD after HSCT. However, it was associated with higher risk of graft versus host disease (GVHD).
2. EBVSTs which are prepared from seropositive donors and cro-preserved for future use.
· Genetic engineering and new methods targeting enhanced capacity of EBVST are investigated to improve outcome of patients with PTLD and EBV related lymphoproliferative disease (EBVLD).
· Methods of preparation of EBVSTs are either Ex-vivo expansion or use of antigen specific T cell selection:
· Ex-vivo expansion of EBVSTs:
o It depends on many steps for good manufacturing-practice GMP-grade EBV-STs.
o Mononuclear cells are isolated from the peripheral blood of donor.
o Then the B-lymphocytes are infected with laboratory strain EBV and transduced with an adenoviral vector expressing latent membrane protein LMP1 and LMP2 and then irradiated.
o Monocytes are separately transduced with the same adenoviral vector and cocultured with T cells followed by a second stimulation by the transduced lymphoblastoid cell lines (LCL) to create and further expand LMP-specific T-cell product.
· Use of antigen specific T cell selection in EBVSTs:
o Using IFN-γ capture approaches where mononuclear cells are pulsed with antigen to isolate CD8+ and CD4+ T cells which secrete IFN-γ in response to the viral antigens.
o Then use of the HLA-restriction requirement to purely select CD8+ T cell product that lacks CD4+T cell (which is necessary for a sustained immune response) and limits the overall applicability of the multimer-selection approach.
· EBVSTs can be either allogenic (T cell are derived from the donor as in case of available living donor) or autologous (used in case of unavailable donor as in deceased organ transplantation or 3rd party (off the shelf) T cell therapy.
· Autologous is more difficult due to immunosuppressed state of transplant recipients. However, it seems more beneficial as mostly PTLD is of recipient origin.
· As regard the outcome of allogenic EBVST immunotherapy in HSCT in either prophylaxis or treatment of EBV-PTLD; it helps in prophylaxis against PTLD and decrease viral load. However, GVHD was a well-known adverse effect. It was also used in SOT and was effective. (mainly for prophylaxis, few cases for treatment of PTLD).
· While autologous EBVST immunotherapy was effective in prophylaxis against PTLD after SOT.
· Type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC required autologous EBVST immunotherapy as an adjunctive therapy to chemotherapy and/or in relapsed patients as they have more restrictive antigen presentation than in type III latency tumors as Burkitt Lymphoma and Gastric Carcinoma.
· 3rd party cell bank (off the shelf T cell therapy):
o Available banked EBVSTs with use of HLA matched (2-5 out of 6) to those with refractory PTLD with reasonable success rate (50-60%.
o The response was better in pediatric population (negative EBV at time of transplantation and has early onset disease).
o It shows reasonable results in BMT and HSCT in children with refractory viral infections (EBV, CMV) and PTLD.
o Its advantage is time saving and early availability without waiting time to access the donor or recipient derived cells.
· The response to EBVSTs can be assessed through measuring the decrease in EBV viral load. However, the PCR is not standardized in all labs (measured in plasma vs whole blood).
· Methods to enhance In-Vivo activity of EBVSTs especially in less antigenic cells of type I and II latency stages:
o Producing TGF-β receptor negative cells to avoid the inhibitory effect of TGF-β secreted by tumor cells.
o Genetic engineering to modify EBVSTs to be CNI and steroid resistant to allow its persistent activity in spite of use of immunosuppressive therapy in SOT.
o Chimeric antigen receptor T cells (CD19) is being investigated in SOT, however nephrotoxicity and neurotoxicity were main side effects.
o Demethylating agents as Azacytidine and decitabine which prevent downregulation of viral antigen expression by EBV-infected B lymphocytes.
2. Indications of its use
o Treatment of PTLD in case of:
§ Refractory cases to standard immunosuppressive medication reduction and chemotherapy as steroids and cyclophosphamide.
§ Advanced and late onset cases.
§ Recurrent cases.
§ EBV +ve cases.
§ Still just tried in case reports and series (few number of cases) and mainly in HSCT not in SOT.
o Treatment of glioblatoma and GIT malignancies.
o Treatment of chronic HIV infection.
o Treatment of autoimmune diseases as type 1 DM and SLE.
3. Can it be used in ttt of other conditions related to SOT
· Ttt of resistant viral infections as CMV and BK.
· Immunomodulation through T reg in treatment of acute rejection.
4. Side effects of adaptive immunotherapy
· DLI is associated with GVHD. However, EBVSTs therapy is not associated with this side effets.
· Generally, may be associated with allergic reactions, GIT upset and flu like symptoms.
Exellent
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
PTLD is a serious complication occurring after solid organ transplantation, around 95% occur due to activation of EBV viruses acquired in childhood, and 95% of cases are B cell type, T cell lymphoma accounts for 5%
After the primary infection EBV, the virus infect B cells and passes into 2 main stages :
Latency III is a stage in which EBV-infected B-lymphocytes express some protein that comprise the EBV genome such as EBV nuclear antigens EBNA1, 2, 3, EBNALP, LMP1, 2 and BARF1. this high antigenic expression is associated with potent T cell response that limit proliferation of B cells to < 2%. Latency III stage is associated with lymphoproliferative disease and malignancy once the patient is immunosuppressed so the B cell start to develop uncontrolled, unchecked proliferation, the most common form of malignancy associated with this stage is PTLD.
On the other hand, Latency II and I stages are less immunogenic , in which there is down regulation of the immunodominant antigens such as EBNA3 and EBNA2. Latency I and II stages are associated with the development of lymphoproliferative diseases and malignancies in both immune-competent and immune-suppressed patients such Hodgkin’s lymphoma, natural killer /T-cell lymphoma, nasopharyngeal carcinoma (Latency II) and Burkitt lymphoma or gastric carcinoma (Latency I)
The principle of adoptive immunotherapy in treatment of PTLD is using EBV specific cytotoxic T lymphocytes or donor lymphocyte infusion to kill dividing B cells containing EBV.
What are the indications of adoptive immunotherapy?
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
What are the side effects of adoptive immunotherapy?
Exellent and clear .
That is an excellent summary and very well structured reply.
What is your analysis of level of evidence of this study?
Introduction:
EBV is the main cause of PTLD.
When the virus enter the oropharynx in primary infection ,it replicate in B-lymphocyte ,which produce different protein comprises wide range of viral genome , EBV nuclear antigens EBNA1, EBNA2, EBNA3, and EBNALP, membrane proteins LMP1, and LMP2 as well as BARF1 and two small non-translated ribonucleic acids (RNA) (Type III latency).
EBV infected B-lymphocytes then downregulate the immunogenic proteins to express less immunogenic type II latency proteins (EBNA1, LMP1 and LMP2) and thus rescue them into the memory compartment where the virus persists in latently infected B-lymphocytes by further downregulating expression of viral proteins so as to become invisible to EBV-specific T-lymphocytes (EBVSTs).
The frequency of EBV-infected B cells in a healthy person remains stable [approximately 0.1–50 EBV infected B lymphocytes per 1,000,000 peripheral blood mononuclear cells (PBMCs)].
EBV-infected B cells, controlled by a potent EBVST response.
Individuals with weakened immune systems like solid organ transplant, the lack of a robust EBVST response can lead to uncontrolled proliferation of type 3 latency EBV-infected B cells resulting in EBV-associated LPD and malignancies.
The transformed B-cells in EBV-associated lymphoproliferative disease associated with latency type III presents several antigenic viral proteins including EBNA 1-3, LMP1 and LMP2 that induce potent EBVST responses. Such potent T cell immunity maintains the infected B cell pool at less than 2%
B cells in immunocompetent individuals but is lacking in immunosuppressed individuals leading to uncontrolled lymph proliferation of the infected B cells.
The scientific rationale:
Adoptive transfer of EBVSTs is based on harnessing the immunogenicity of type III latency malignancies to control the proliferation of the latently infected B cells.
Manufacturing of EBVSTs:
EBVSTs can be readily produced from EBV-positive donors.
Methods:
Ex vivo expansion of EBVSTs:
Where EBVSTs were selectively expanded utilizing irradiated EBV transformed lymphoblastoid cell lines (LCL) as antigen presenting cells (APCs) to selectively expand EBVSTs.
Adenovirus vectors also used to enhance the specificities.
Disadvantage:
Complex.
Time consuming.
APC such as dendritic cell (DC) pulsed with overlapping peptide libraries spanning whole antigen coupled with the use of artificial APCs [e.g., the novel antigen-presenting complex (KATpx)] can facilitate the rapid (10–21 days) expansion.
Antigen-specific T cell selection:
Products can be rapidly produced.
(I) major histocompatibility complex (MHC) multimer selection where oligomeric forms of MHC molecules are designed and conjugated to magnetic beads to isolate typically CD8+ T cells with high affinity to a specific viral epitope/peptide in an HLA restricted manner .
(II) IFN-γ capture approaches where mononuclear cells are pulsed with antigen to isolate CD8+ and CD4+ T cells which secrete IFN-γ in response to the viral antigens.
Post-HSCT donor-derived T cell therapy:
Donor lymphocyte infusions:
Unmodified DLI derived from the patient’s EBV seropositive HSCT donor which contained effector cells with activity against EBV.
Donor-derived EBVSTs:
Donor derived EBVSTs for the prevention and treatment of EBV+ PTLD in the post-HSCT setting when the donor is available.
Autologous EBVSTs
Autologous EBVSTs has been used for patients with EBV-associated malignancies in SOT recipients with PTLD where donors usually are not available because of the use of cadaveric grafts.
Autologous EBVSTs have also been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patient
Third-party T cells:
Off the shelf” T cell therapy products. Many bank have being created to reduce need for donors for patient-specific EBVST product (autologous or allogeneic).
Modification of EBVSTs to enhance activity:
Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo:
EBV/LMP-specific T-cell (LST) product expressing a dominant negative TGF-β receptor type II (DNRII) .
Calcineurin resistance to enhance EBVSTs:
EBVSTs were genetically engineered to express a mutant form of calcineurin thus rendering them calcineurin inhibitor (CNI) resistant.
Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance EBVST activity in vivo Demethylating agent:
Pretreatment with decitabine induced expression of LMP1 and ENBA3 associated with latency type 3 which sensitized tumor cells to subsequent therapy with EBVSTs.
Conclusions:
The use of adoptive immunotherapy, particularly using third-party “off the shelf” EBVSTs for the treatment of EBV-LD has shown promise in several studies conducted at specialized center.
1. What are the indications of adoptive immunotherapy?
Prophylaxis; treatment of EBV virus-associated lymphoproliferative disease.
1. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Natural killer /T-cell lymphoma, nasopharyngeal carcinoma or type I latency tumors (e.g., Burkitt lymphoma or gastric carcinoma)
1. What are the side effects of adoptive immunotherapy?
GVHD.
Anaphylactic.
Other side effects:
Fever ,chills, edema ,hypotension .
Autoimmune disease, uveitis’, vitiligo.
Cytokines release syndrome.
Other indication of EBV ST therapy:
metastatic melanoma.
Breast cancers.
Cholangiocarcinoma.
RCC.
That is an excellent summary and very well structured reply. Under the heading of Introduction, I wish you could type sub-headings in bold or underline or in italics. That would make it easier to read.
What is your analysis of level of evidence of this study?
The major complication of EBV positive post transplant is PTLD in HSCT and solid organ transplantation. Less than 25% of HSCT recipient have PTLD and is associated with EBV sero status, degree of T-cell depletion, and immunosupression.
Principals;
PTLD is highly immunogenic and responded to immunotherapy with EBVSTs.
EBVSTs can be produced from EBV-positive donors using good manufacturing grade compliant methodology.
The commonly used methods consist of ex vivo expansion of VSTs versus antigen specific.T-cell selection
The major risk factor for developing EBV-associated PTLD post-SOT(solid organ transplant) is EBV
seronegativity at the time of transplant. Since most children are transplanted at a young age while still being EBV seronegative and convert to EBV seropositivity within 2 years of transplant, EBV driven PTLD is much more common in paediatric SOT recipient
Adoptive cellular therapy using EBVSTs (EBV specific T-lymphocyte) has shown impressive results in immunocompromised patients with
Adoptive immunotherapy of autologous or allogeneic EBV-specific CTLs has and will have an important role in establishing scientific principles and might find clinical application in conjunction with conventional therapies
Recent research is focused on strategies to enhance EBVST potency in vivo through genetic engineering as well as combination therapy
. What are the indications for adoptive immunotherapy?
-EBSTs and multi-VSTs presumed by third parties in PTLD
-recipient with immunosuppression and latent EBV infection
To decrease in patients EBV load and prevention of treatment
In refractory and treatment resistant cases of PTLD
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Autoimmune disease
Anti rejection treatment
It can be used as a anti fibrotic therapy
Side effect of adoptive therapy
Fever, chills, flue
Kidney injury
Neurological side effect
GVHD
That is an excellent summary and very well structured reply.
What is your analysis of level of evidence of this study?
1. Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
rituximab:
– anti cd20
– response rate 55-100% in HSCT& around 50% in SOT
– in pediatric SOT recipients, a combination regimen using
– low dose cyclophosphamide with prednisone and rituximab
– has shown event-free survival (EFS) rates of 72%.
Adoptively transferred virus-specific T cells (VSTs):
– ranging from the use of donor lymphocyte infusions (DLI) to donor-derived multi-antigen specific VSTs
– can be readily produced from EBV-positive donors using good-manufacturing-grade (GMP) compliant methodologies
– Ex vivo expansion of EBVSTs
– Antigen-specific T-cell selection
– Post-HSCT donor-derived T cell therapy ( Donor lymphocyte infusions, Donor-derived EBVSTs )
– Autologous EBVSTs
– Third-party T cells
modification of EBVSTs to enhance the activity:
– Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo
– Calcineurin resistance to enhance EBVSTs
– Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance EBVST activity in vivo (Demethylating agents, Checkpoint inhibitors, BCL-2 inhibitors)
2. What are the indications of adoptive immunotherapy?
– EBV-associated PTLD in HSCT & SOT as prophylaxis to reduce the viral load, induction of remission & decrease relapses
3. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
Trials showing promising efficacy in:
– Auto-immune triggered skin disease(pemphigus)
– autoimmune diseases: Lupus, Multiple sclerosis.
4. What are the side effects of adoptive immunotherapy?
– Flu-like symptoms: chills, fever
– Neurological toxicity: delirium, headache, confusion.
– GVHD
– AKI
– Anaphylactic reaction, palpitation
That is an excellent summary and very well structured reply.
What is your analysis of level of evidence of this study?
1. Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD?
EBV infection has different phases from lytic phase (when the virus express all the antigens) to different phases of latency periods which allows virus escaping from immune surveillance.
There is associated malignancies with every virus phase, PTLD is associated with latency phase III (LMP1, 2A, 2B virus antigens).
Immunosuppressed population lack of a robust EBV specific T cell (EBVST) response can lead to uncontrolled proliferation of type 3 latency EBV-infected B cells resulting in EBV-associated LPD and malignancies. The scientific rationale for adoptive transfer of EBVSTs is based on harnessing the immunogenicity of type III latency malignancies to control the proliferation of the latently infected B cells.
2. What are the indications of adoptive immunotherapy?
EBV-associated PTLD that persists following initial therapy
3. Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
malignancies includes PTLD, melanoma, cervical squamous cell carcinoma and cholangiocarcinoma.
4- What are the side effects of adoptive immunotherapy?
It varies according to the type and target, it includes:
acute and chronic graft-versus host disease (GVHD)
cytokine storm
Neurotoxicity
Others: AKI, Arrhythmia, GI upset, neutropenia, infections and constitutional symptoms
Resources:
That is an excellent summary and very well structured reply.
What is your analysis of level of evidence of this study?
This is a systematic review is based on a review of published literature spanning 20 years from 2000 to 2020 in addition to personal experience and data from the authors
EBV is prevalent in adult population by more than 90% infection acquired during childhood where it get attach to B lymphocyte through specific site ( CD21 ) and stimulate the cell protein factory to produce its antigens (3 nuclear antigen and three surface membrane protein) that stimulate Cytotoxic T lymphocyte to control the infection . hence the virus within the infected slave B lymphocyte enter in latent phase inside lymphoid tissue so long the immune system is potent . under any weakness of immune system ( HIV infection or solid organ transplantation SOT) latent antigenic machine inside B lymphocyte will be active and can cause lymphoproliferative disease
So the rule of r adoptive transfer of EBV specific T lymphocytes (EBVSTs) is to control the proliferation of the latently infected B cells
In solid organ recipients, incidence of PTLD ranges from 2% to 25% depending on
– The organ transplanted,
– Passenger lymphocytes in the transplanted organ
– Type of immunosuppression
– EBV seronegativity at the time of transplant
Manufacturing of EBVSTs
1-Ex vivo expansion of EBVST:
Disadvantage: long production time
2- Antigen-specific T cell selection
– Advantages: less time consuming
– Disadvantages: difficulties in finding donor with specific cells
3- Autologous EBVSTs
indication of adoptive immunotherapy:
prophylactic to reduce EBV viral load
therapeutic : immunosuppressed recipient with EBV- latent disease causing PTLD
Uses of adoptive immunotherapy other than cancer treatment:
Autoimmune disease
side effects
GVHD
acute kidney injury,
Heart : heart arrhythmias, tachycardia, hypotension
chills, fever
GIT: constipation, diarrhea nausea, vomiting decreased appetite,
Chest : cough, cytokine release syndrome (cytokine storm),
Neurological : delirium, , dizziness, headache, encephalopathy, fatigue, tremors,
Hematology: febrile neutropenia, , hypogammaglobulinemia, bleeding episodes,
That is an excellent summary and very well structured reply. I wish you could type sub-headings in bold or underline or in italics.
You have used too much of bold type in one heading. Typing in bold or capitals amounts to shouting!
What is your analysis of level of evidence of this study?
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
————————————————————————————————————
PTLD is highly immunogenic and amenable to immunotherapy with EBVST . Adoptively transferred virus-specific T cells (VSTs) have been evaluated for more than two decades, ranging from the use of donor lymphocyte infusions (DLI) to donor derived multi-antigen specific VSTs in the HSCT setting to autologous as well as allogeneic VSTs in the SOT setting .
Donor lymphocyte infusions;
———————————————————
The earliest reported experience of cellular therapy for the treatment of PTLD utilized unmodified DLI derived from the patient’s EBV seropositive HSCT donor which contained effector cells with activity against EBV . Although effective in inducing remissions, this therapy carries a significant risk of graft-versus-host disease (GVHD) .
Donor-derived EBVSTs;
———————————————–
There is extensive reported experience using donor derived EBVSTs for the prevention and treatment of EBV+ PTLD in the post-HSCT setting when the donor is available . This therapy was well tolerated without significant complications with remarkable reductionin EBV viral copy numbers within 4 weeks, including in a patient with immunoblastic lymphoma .
Autologous EBVST;
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Autologous EBVSTs are preferrable to donor EBVSTs even if available in the post-SOT setting because PTLD is usually of recipient origin and solid organ grafts are not routinely HLA matched to the recipient. However, the challenges in production and the production time leading to delays in initiation of therapy in patients with a rapidly progressive disease impede the routine use of these products in this setting .
Several strategies have been developed to manufacture EBVST products with minimal alloreactivity and broad specificity against EBV latency proteins or as a multi-virus specific product with activity against multiple viruses . The most commonly utilized methods consist of ex vivo expansion of VSTs versus antigen-specific T cell selection [e.g., interferon γ (IFN-γ) capture.
1-Ex vivo expansion of EBVSTs;
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Ex vivo expansion of T cells targeting viral antigens via native T cell receptor .
2-Antigen-specific T cell selection;
—————————————————–
1-major histococomplex (MHC) multimer selection; where oligomeric forms of MHC molecules are designed and conjugated to magnetic beads to isolate typically CD8 + T cells with high affinity to a specific viral epitope/peptide in an HLA restricted manner .
2- IFN- γ capture approaches; where mononuclear cells are pulsed with antigen to isolate CD8 + and CD4 + T cells which secrete IFN- γ in response to the viral antigens.
Third-party T cells ;
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It is a readily available “off the shelf” T cell therapy product .The feasibility of third-party cell banks able to cover a majority of the referred patient population had been demonstrated in different reports with impressive response rates .
Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo;
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TGF-β can be released into the tumor microenvironment by tumor cells, fibroblasts and immune cells and creates an immunosuppressive environment by impeding T-cell activation, proliferation and migration. In addition, it affects DC and macrophage antigen presentation and chemo taxis .
Calcineurin resistance to enhance;
——————————————————
Given the concerns regarding EBVST persistence in patients who require ongoing immune suppression , several groups have explored gene engineering of virus specific T cells to render them resistant to immune suppressive agents including steroids and calcineurin inhibitor .
Chimeric antigen receptor T cells ;
——————————————————-
CD19-chimeric antigen T cells (CD19-CART) have shown impressive efficacy in acute B-lymphoblastic leukemia and in B-cell lymphomas . In addition to viral antigens, EBV-LD expresses a variety of B-cell antigens targetable by CARTs including CD19, CD20 and CD30. However, their use in EBV-lymphoproliferation is limited by the patient’s immunosuppressive state impeding T cell manufacture and the length of production time.
.
Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance EBVST activity in vivo;
————————————————————————
1-Demethylating agent;
Newly EBV-infected B-cells express up to 90 viral genes; however rapid CpG methylation of viral antigens leads to downregulation of viral protein expression and the latency . Azacytidine and decitabine are potent CpG demethylating agent .
2-Checkpoint inhibitors ;
LMP1 has been shown to induce expression of the checkpoint protein PD-L1 in classic HL (CHL) without 9p24.1 alteration . PD-L1 is expressed in 73% of EBV-positive PTLD . . There have been several trials using checkpoint blockade in DLBCL hinting at single agent activity . A phase I study at BCM is exploring combination therapy of checkpoint inhibitors with EBV directed T cells for EBV + HL and NHL .
3-BCL-2 inhibitotors;
In preclinical, in vitro studies, pretreatment of malignant B-cell lines with the BCL-2 inhibitor venetoclax increased proapoptotic proteins and sensitivity to CD19.
2-What are the indications of adoptive immunotherapy?
——————————————————————————————–
1- Inducing remission of PTLD .
2- Prevention and treatment of EBV+ PTLD in the post-HSC.
3-Treatment of type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients .
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
—————————————————————————
Autologous EBVSTs have also been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients .
What are the side effects of adoptive immunotherapy?
———————————————————————————
1-The risk of graft-versus-host disease (GVHD)
2-The risk of graft rejection and autoimmunity .
That is an excellent summary and very well structured reply.
What is your analysis of level of evidence of this study?
Thanks professor
Level V
Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD
The major risk factor for developing EBV-associated PTLD post-SOT is EBV seronegativity at the time of transplant
More than 90% of EBV-associated PTLD is of B-cell origin with cell surface expression of CD20
PTLD is highly immunogenic and adjustable to immunotherapy with EBVSTs
Donor lymphocyte infusion or EBV-specific T-lymphocytes (EBVSTs) in specialized centers
The aims of this study are to define current best manufacturing strategies for EBV-specific VSTs, clinical experience of their use, discuss opportunities to broaden the applicability of this approach and to explore future strategies to enhance their efficacy
Manufacturing of EBVSTs
Produced from EBV-positive donors using good-manufacturing-grade (GMP) compliant methodologies
Donor type source: autologous and allogeneic (including third party)
Two methods: ex vivo expansion of VSTs and antigen-specific T cell selection
Ex vivo expansion of EBVSTs: complex and time consuming. Can be manufactured from individuals irrespective of their HLA type. Can be produced from EBV seronegative donors
Antigen-specific T cell selection [interferon γ (IFN-γ)]: EBVST products can be rapidly produced (in 48 hours) from EBV seropositive autologous and allogeneic donors
Donor-derived T cell therapy (Post-HSCT)
Donor lymphocyte infusions: produced from EBV seropositive donor. Effective but high risk of GVHD
Donor-derived EBVSTs: safe and rare GVHD
Autologous EBVSTs
Technically challenging (ongoing immunosuppression)
Used in case of no available donor (cadaveric grafts). Effective and rare PTLD
Third-party T cells
“Off the shelf” T cell therapy product. Mostly used in the post-transplant setting
Modification of EBVSTs to enhance activity
1. Overcoming the immune suppressive effects of TGF-β
2. Calcineurin resistance
3. Chimeric antigen receptor T cells
4. combination strategies administering EBVSTs with other therapeutic modalities
(a) Demethylating agents
(b) Checkpoint inhibitors
(c) BCL-2 inhibitors
Conclusions
Use of adoptive immunotherapy for the treatment of EBV-LD is promising in several studies (EBVSTs
Is now widely available)
What are the indications of adoptive immunotherapy?
Patients with EBV-associated PTLD who fail conventional treatments or cannot tolerate it (RI, rituximab, and chemotherapy)
Treatment of PTLD with EBV-specific CTLs should be considered where available with refractory or relapsed (R/R) EBV-positive PTLD (1C)]
Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
CMV, BK virus, and rejection
What are the side effects of adoptive immunotherapy?
Vary according to the type of adoptive immunotherapy, location of cancer, type of cancer, and patient overall health
In most cases side effects can be managed safely when recognized early
Overactive immune response (cytokine release syndrome), and neurotoxicity, AKI, bleeding disorders, arrhythmias, chills, GIT symptoms, febrile neutropenia, and hypotension
That is an excellent summary and very well structured reply.
What is your analysis of level of evidence of this study?
Thank yoy Prof. Ajay
Level 5
1. The principle of adoptive immunotherapy in PTLD treatment:
It involves generating EBV-specific T lymphocytes (EBVSTs) from EBV-positive autologous or allogenic donors with limited alloreactivity and wide virus-fighting activity.
It is necessary to use ex vivo virus-specific T cell growth or antigen-specific T cell selection.
MHC selection involves isolating CD8+ T cells with high affinity to a specific viral epitope or peptide in an HLA-restricted manner; IFN- capturing involves isolating CD8+ and CD4+ T cells that release IFN- in response to viral antigens.
Technically challenging approaches.
Post-HSCT donor-derived T cells
Donor-lymphocyte infusions
Unmodified DLI from PTLD patients’ EBV-positive HSCT donors induces remission.
Donor-derived EBVSTs may prevent and treat EBV+ PTLD in post-HSCT patients without reactivation or GVHD by lowering EBV burden.
EBV-transformed lymphoblastoid cell lines (LCLs) are used to selectively grow EBV-specific T lymphocytes as antigen-presenting cells.
DC and LCLs were transduced with adenovirus vectors to overexpress LMP1 or LMP2 to increase specificity for less immunogenic EBV antigens, but it is difficult and wastes time.
2. What are the indications for adoptive immunotherapy?
-EBSTs and multi-VSTs provided by third parties in PTLD
-recipient with immunosuppression and latent EBV infection
-Treatment for prevention and a decrease in the patient’s EBV load
-Treatment resistant and refractory cases of PTLD
3-Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
T-cell adoptive immunotherapy for BK nephropathy in renal transplantation
The treatment of lymphoproliferative diseases connected to viral (EBV) loads
4—What are the side effects of adoptive immunotherapy?
GVHD, acute kidney injury, bleeding episodes, heart arrhythmias, chills, constipation, cough, cytokine release syndrome (cytokine storm), decreased appetite, delirium, diarrhea, dizziness, edema, and encephalopathy.
1. Briefly summarise the principle of adoptive immunotherapy in the treatment of PTLD.
-EBV-positive post-transplant lymphoproliferative disease (PTLD) is a major complication of hematopoietic stem cell and solid organ transplantation,
– The biggest risk factor for developing EBV-associated PTLD post-SOT is EBV seronegativity at the time of transplant. Since most children are transplanted at a young age while still being EBV seronegative and convert to EBV seropositivity within 2 years of transplant, EBV driven PTLD is much more common in pediatric SOT recipients,
-Adoptive cellular therapy using EBVSTs has shown impressive results in immunocompromised patients with EBV-LD and there is the potential for wider use.
– Adoptive immunotherapy uses EBV-specific cytotoxic T lymphocytes (EBV-CTLs) (autologous or allogeneic) or donor lymphocyte infusion (DLI) in an attempt to kill dividing B cells in EBV-associated PTLD,
–“off the shelf” T cell therapy product is desirable. Because donor cells may not be available (e.g., recipients of umbilical cord blood transplants or cadaveric organ transplants). Hence, manufacture of patient-specific products may not be possible or may be so delayed that patients with rapidly progressive disease are not able to access these therapies,
-Potential for combination strategies administering EBVSTs with other therapeutic modalities to enhance EBVST activity in vivo,
-Demethylating agents
-Checkpoint inhibitors
-BCL-2 inhibitors
2- What are the indications of adoptive immunotherapy?
–Allogenic virus-specific T cells in PTLD in HSCT recipients,
-Autologous EBVSTs for EBV PTLD in SOT recipients,
-Third-party EBSTs and multi-VSTs in PTLD,
-Type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients. Such tumors are more challenging targets because of the reduced expression of immunogenic viral antigens in these type II latency tumors which express a more restricted array of antigens compared to type III.
3- Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation?
-Autologous EBVSTs have also been used to treat type II latency EBV-associated malignancies including HL, T/NK lymphoma and NPC as adjunctive therapy to chemotherapy and/or in relapsed patients.
4- What are the side effects of adoptive immunotherapy?
-The major complication of adoptive immunotherapy is acute and chronic graft-versus-host disease (GVHD) . DLI may produce GVHD, which does not appear to be a problem with EBV-CTLs.
-Impaired Graft Function
-CRS
Thank you for your reply. What is the level of evidence provided by this article?
Please explain the abbreviation.
THANK YOU MY DEAR PROF;-
-THIS REVIEW ;. LEVEL III OF EVIDENCE
-THESE MY ABBREVATIONS;.
(PTLD);. Post-transplant lymphoproliferative disorder(EBV);.Epstein-Barr virus
(SOT);.Solid Organ Transplant
(EBV-CTLs);.EBV Epstein-Barr virus (EBV) specific cytotoxic T lymphocytes
(EBVSTs) ;.(EBV)-specific T lymphocytes
(DLI);. donor lymphocyte infusion
(HSCT);.(haematopoietic stem cell transplant)
(GVHD);.chronic graft-versus-host disease
(CRS);. Cytokine release syndrome
sorry; Level V
1- Principles of adoptive immunotherapy:
– EBV-infected B-lymphocytes pass through out many phases: lytic phase with high antigen expression, Latency III with less expression, Latency II, Latency I and Latency 0 with no anitigen expression (dormant infection) and become invisible to specific T cells.
-Impaired immunity—replication and proliferation of B cells and cause
EBV- malignancies which are:
1- type III latency: PTLD
2- Type II latency : HL, natural killer (NK)/T-cell lymphoma, nasopharyngeal
carcinoma (NPC)
3- type I latency: Burkitt lymphoma or gastric carcinoma
– So, he rational for adoptive immunotherapy is to attack the proliferating B-cells with EBV-specific T cells.
2- Indications of adoptive immunotherapy:
EBV-lymph proliferative diseases either post transplantation (HSCT or SOT) or not.
3- Can adoptive immunotherapy be used in the treatment of other conditions related to organ transplantation:
prophylaxis against EBV-related PTLD
4- side effects:
Graft versus host disease
impaired graft function
non-response
Thank you for your reply. What is the level of evidence provided by this article?
level of evidence: is V
It’s mainly for eradication of Epstein-Barr virus (EBV), to prevent risk of lymphoma and lymph proliferative disorders in solid organ transplant.
The incidence of PTLD is 1-25% and mainly express CD20 on the surface of B cell. Only 10% is T cell and responded well to rituximab in case of HSCT but it’s limited in use because risk of infection and response low in solid organ transplant.
Post-HSCT donor-derived T cell therapy
Donor lymphocyte infusions:
Enhance remission by eradicate of EBV but carrying high risk of GVHD, However it’s limited by depletion of T cell regulators before infusion.
Donor-derived EBVSTs:
prevent and treat of EBV related PTLD; with no risk of GVHD.
Autologous EBVSTs:
It’s used to treat type II latency EBV-associated malignancies including Hodgkin’s lymphoma and natural killer lymphoma, with complete remission but not reduce viral load of EBV.
Third-party T cells:
This therapy use to treatment of lymphoproliferative disorder related to refractory infection like EBV and CMV and adenovirus by reducing viral load.
It’s has good effects.
Modification of EBVSTs to enhance activity
Overcoming the immune suppressive effects of TGF-β to enhance EBVST activity in vivo:
Calcineurin resistance to enhance EBVSTs
Chimeric antigen receptor T cells
Combination therapy of EBVSTs with other therapy like
Demethylating agents
Checkpoint inhibitors
BCL-2 inhibitors
Treatment of lymph proliferation disease related to viral load especially EBV
Prevention and reduce virama of EBV
Prophylactic and treat viral load
Autoimmune disease
Graft -versus-host disease
Flu like illness
Thank you for your reply. What is the level of evidence provided by this article?
There are more side effects than what you have written.
Adoptive cellular immunotherapy for Epstein-Barr virus-associated lymphoproliferative disease.
Summary of the article:
Introduction:
EBV, the worldwide common viral infection in childhood, affects 90% of the adult population.
In healthy individuals, the immune system analyses the virus and protects against spreading infection, although rare cases can go into fulminant disease.
EBV goes into a dormant phase latent in the B cells.
In contrast to healthy individuals, the patients with the defective immune system, include:
can lead to uncontrolled infection and proliferation which can lead to EBV- associated LPD or malignancies.
The transformed B-cells in EBV-associated LD, associated with latency type III, present Viral proteins:
such antigens in intact immune systems, T-cells can control B cells and maintain infected B cells < 2%.
Type II latency associated with malignancies:
Type I latency tumors:
The occurrence of PTLD in HSCT ranges from 1-25%, depending on the following:
Sero negativity is the top determinant factor of the development of post-transplant LPD.
Since pediatric patients are being still seronegative as they transplant earlier at a younger age, PTLDs are more common in pediatric patients.
Monotherapy with Rituximab anti-CD20 is more effective in HSCT with a response rate of 55 -100%, while combination therapy (prednisolone/cyclophosphamide/rituximab) is shown to better survival up to 725.
#EBVSTs manufacturing:
#Post-HSCT donor-derived T-cell therapy:
# Autologous EBVSTs.
# Third-party T cells.
Modification of EBVSTs to enhance activity:
Combination strategy, with EBVSTs:
Conclusion:
Indication of adoptive immunotherapy:
Uses of adoptive immunotherapy other than cancer treatment:
Side effects:
Excellent
Thank you prof.
1- The principal of adoptive immunotherapy in PTLD treatment
EBVSTs Manufacture
It includes producing EBV specific T lymphocytes (EBVSTs) from EBV positive donors either autologous or allogenic donors with minimal alloreactivity and broad spectrum activity against multiple viruses.
The methodology involve either ex vivo expansion of virus specific T cells or antigen-specific
T cell selection.
Ex vivo expansion of EBVST
By using irradiated EBV transformed lymphoblastoid cell lines(LCLs) expressing high levels of class I and class II HLA and co-stimulatory molecules acting as antigen presenting cells to selectively expand EBV specific T lymphocytes.
DC and LCLs were transduced with adenovirus vectors to overexpress LMP1 or LMP2 to strengthen the specificity to less immunogenic EBV antigens but in fact it is difficult and wastes time on the other hand it enhances rapid increase of T cells targeting EBNA1, LMP1 and LMP2 from healthy donors and from cases with type 2 latency EBV-associated malignancies, also EBVTs can be derived from EBV seronegative donors.
Antigen specific T cell selection
-MHC selection by isolating CD8+T cells having high affinity to a specific viral epitope/peptide in an HLA restricted manner
– IFN-γ capturing by isolating CD8+and CD4+ T cells that secrete IFN-γ in response to the viral antigens
Those methods are difficult technically.
Post-HSCT donor-derived T cell therapy
Donor lymphocyte infusions
Used in the treatment of PTLD by unmodified DLI taken from patient’s EBV seropositive HSCT donor having activity against EBV, inducing remission.
Donor-derived EBVSTs
Using ex vivo expanded EBVSTs derived from seropositive donors for the prevention and treatment of EBV+ PTLD in the post-HSCT efficiently without reactivation nor occurrence of GVHD , through decreasing EBV load in cases with reactivation or high risk for reactivation.
Autologous EBVSTs
It was used for treatment of PLTD in SOT patients from cadavers ,which is challenging due to the immunosuppression and that can delay starting therapy.
EBVST produced by LCL stimulation consist of T cells specific to early lytic antigens and the immunodominant EBNA3 antigens but less active towards viral antigens expressed
in latency type II.
Meanwhile in a trial latent membrane protein (LMP)-specific T cells were used as adjuvant therapy with acceptable outcomes with type II latency malignancies
Third party T cells
Donor cells may not be available therefore off the shelf T cells will be needed.
33 transplant recipients of different organs ,having refractory PTLD were given partially HLA matched EBVSTs and the cases with the higher HLA match had a better response with less rejection and GVHD risk.
Third-party multi-VSTs use for EBV-associated PTLD had acceptable outcomes.
Third party EBV gave promising results for CNS PLTD cases
EBVSTs modification to increase efficiency
-by inhibiting TGFB immunosuppressive activity
In EBV positive HL ,EBV/LMP-specific T-cell (LST) product expressing a dominant negative TGF-β receptor type II (DNRII) were resistant to inhibitory concentrations of TGF-β produced by tumor cells with favourable response
– Calcineurin resistance to enhance EBVSTs
Gene engineering of virus specific T cells to resist immune suppressives
– Chimeric antigen receptor T cells
– Combining EBVSTs with other therapies as
· Demethylating agents that can sensitize tumor cells to EBVST
· EBV-positive diffuse large B-cell lymphoma (DLBCL) and 73% of EBV-positive PTLD express PD-L1 but combining checkpoint inhibitors with EBV directed T cells for EBV+ PTLD had side effects
· BCL-2 inhibitors ,as BCL-2 is an antiapoptotic protein for latent EBV infected cells upregulated by LMP1 protein.
Combing BCL-2 inhibitors with CD 19 CART for treatment of EBV lymphoproliferative is under trial
2-Adoptive therapy indications are EBV positive PLTD in HSCT and in SOT recipients, EBV-associated malignancies as HL, T/NK lymphoma and NPC and in BMT recipeints infected with EBV,CMV ,HHV6,and BKV
3- Autologous EBVSTs was involved in treat of type II latency EBV-associated malignancies as HL, T/NK lymphoma and NPC with chemotherapy and/or in relapsed patients.
–Multi-VSTs [EBV, cytomegalovirus (CMV), adenovirus, +/− BK virus (BKV) and human herpes virus 6 (HHV6)] products were given to HSCT recipients having refractory viral disease after BMT targeting EBV, CMV and/or adenovirus
Third party EBV was given to post-HSCT or SOT cases ,no published data on it’s use in HIV-associated lymphoma
EBV/LMP-specific T-cell (LST) use in EBV positive HL
CNI-resistant EBVST for human B cell lymphoma bared by a mouse (under trial)
CD19-chimeric antigen T cells in acute B-lymphoblastic leukemia and in B-cell lymphomas
EBVSTs with demethylating agent in Burkitt lymphoma
Combining checkpoint inhibitors with EBV directed T cells for EBV+ HL and NHL under trial
4- Side effects of adoptive immunotherapy
-DLI can lead to GVHD through selective depletion of T reg, while depleting
naïve T cells can decrease the GVHD risk
-Multi VST can lead to TMA and GI hemorrhage
– CD19-chimeric antigen T cells (CD19-CART) caused cytokine release syndrome (CRS), neurotoxicity and acute kidney injury.
– Combining checkpoint inhibitors with EBV directed T cells for EBV+PTLD lead to graft rejection and autoimmune complications.
Thank you for your Doaa for your reply. What is the level of evidence provided by this article?
level of evidence is V
2.Indications of adoptive immunotherapy: Prophylaxis and treatment
3.Can adoptive immunotherapy be used in the treatment of other conditions?
4.What are the side effects of adoptive immunotherapy?
Thank you Ben for your reply. What is the level of evidence provided by this article?
Please explain the abbreviation.
Welcome prof