Cytokine production and cytolytic mechanism of CD4+cytotoxic T lymphocytes in ex vivo expanded therapeutic Epstein-Barr virus–specific T-cell cultures

Blood ◽  
2002 ◽  
Vol 99 (9) ◽  
pp. 3302-3309 ◽  
Author(s):  
Qi Sun ◽  
Robert L. Burton ◽  
Kenneth G. Lucas
Keyword(s):  
Cell Culture ◽  
T Cells ◽  
T Cell ◽  
Cytotoxic T Lymphocytes ◽  
Epstein Barr Virus ◽  
Ex Vivo ◽  
Cd4 Cells ◽  
Barr Virus ◽  
Ifn Γ ◽  
Epstein Barr

Abstract Ex vivo expanded Epstein-Barr virus (EBV)–specific T cells have been successfully applied clinically for adoptive immunotherapy. However, the role of CD4+ T cells in the therapeutic T-cell culture has not been established for the reconstitution of EBV-specific immunity. We isolated and characterized CD4+ T-cell lines from the ex vivo T-cell cultures. Monoclonal line PD-F4 and oligoclonal lines ND-R4 and TD-B4 were CD3+CD4+CD8−. Cytolytic tests with targets of mismatched major histocompatibility complex (MHC) and anti-MHC antibodies confirmed that the cytotoxicity of these CD4+ cells was restricted by MHC class II. Single cells of ND-R4 expressed interferon-γ (IFN-γ, or interleukin 4 (IL-4), but rarely coexpressed these 2 cytokines. In contrast, PD-F4 coexpressed IFN-γ, IL-2, and IL-4. Kinetic studies with PD-F4 showed that expression of the 3 cytokines plateaued 5 hours upon stimulation and was then drastically reduced, with a pattern consistent with independent modulation and differential off-cycle signal requirements. The cytotoxicity of these CD4+ cells was largely resistant to brefeldin A, an inhibitor for cytolytic pathways by Fas-ligand family molecules. Although sensitive to concanamycin A and ethyleneglycotetraacetic acid, which inhibit cytotoxicity by granule exocytosis, the CD4+ cytotoxic T lymphocytes (CTLs) did not express perforin, suggesting a cytotoxic mechanism independent of perforin although involving exocytosis. Flow cytometric analysis showed that the CD4+ CTLs expressed granulysin, a recently identified cytolytic molecule associated with exocytotic cytolytic granules. These data suggested that CD4+ T cells in the therapeutic B-lymphoblastoid cell lines–primed T-cell culture are diverse in producing TH1 and TH2 cytokines, and may exert specific cytotoxicity via exocytosis of granulysin.

scholarly journals Repertoire and frequency of immune cells reactive to Epstein-Barr virus–derived autologous lymphoblastoid cell lines

Blood ◽  
2008 ◽  
Vol 111 (3) ◽  
pp. 1334-1343 ◽  
Author(s):  
Sumita Bhaduri-McIntosh ◽  
Marisa J. Rotenberg ◽  
Benjamin Gardner ◽  
Marie Robert ◽  
George Miller
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Lines ◽  
Immune Cells ◽  
Epstein Barr Virus ◽  
Ex Vivo ◽  
Lymphoblastoid Cell Lines ◽  
Barr Virus ◽  
Different Types ◽  
Epstein Barr

AbstractAnswers to questions about frequency and repertoire of immune cells, relative contributions made by different types of immune cells toward the total Epstein-Barr virus (EBV)–directed response and the variation of such responses in healthy persons have been elusive because of disparities in assays, antigen presenting cells, and antigenic sources used in previous experiments. In this study, we addressed these questions using an assay that allowed direct comparison of responses generated by different types of cells of the immune system. This short-term (20-hour) ex vivo assay measured interferon-γ production by blood cells in response to autologous EBV-transformed lymphoblastoid cell lines (LCLs). Our experiments defined the variation in responses among persons and clearly distinguished 10 healthy EBV-immune from 10 healthy EBV-naive persons. In EBV-immune persons, 33% of responding cells were CD4+, 43.3% were CD8+, and 12.9% were γ-δ T cells. LCL-reactive CD8+ T cells were only 1.7-fold more frequent than similarly reactive CD4+T cells. Responses by γ-δ T cells were 6-fold higher in seropositive than in seronegative persons. Our findings emphasize the importance of CD4+ and γ-δ T-cell responses and have implications for immunotherapy and for identifying defects in T-cell populations that might predispose to development of EBV-associated lymphomas.

scholarly journals Acquisition of Polyfunctionality by Epstein-Barr Virus-Specific CD8+ T Cells Correlates with Increased Resistance to Galectin-1-Mediated Suppression

2009 ◽  
Vol 83 (12) ◽  
pp. 6192-6198 ◽  
Author(s):  
Corey Smith ◽  
Leone Beagley ◽  
Rajiv Khanna
Keyword(s):  
T Cells ◽  
T Cell ◽  
Epstein Barr Virus ◽  
Ex Vivo ◽  
Interleukin 2 ◽  
Nuclear Proteins ◽  
Proliferative Potential ◽  
Peptide Epitopes ◽  
Barr Virus ◽  
Epstein Barr

ABSTRACT Latent membrane antigen 1 and -2 (LMP-1/2)-specific CD8+ T cells from newly diagnosed and relapsed Hodgkin's lymphoma (HL) patients display a selective functional impairment. In contrast, CD8+ T cells specific for Epstein-Barr virus (EBV) nuclear proteins and lytic antigens retain normal T-cell function. Reversion to a dysfunctional phenotype of LMP-1/2-specific T cells is coincident with the regression of HL. To delineate the potential basis for this differential susceptibility for the loss of function, we have carried out a comprehensive functional analysis of EBV-specific T cells using ex vivo multiparametric flow cytometry in combination with assessment of antigen-driven proliferative potential. This analysis revealed that LMP-1/2-specific T cells from healthy virus carriers display a deficient polyfunctional profile compared to that of T cells specific for epitopes derived from EBV nuclear proteins and lytic antigens. Furthermore, LMP-specific T-cells are highly susceptible to galectin-1-mediated immunosuppression and are less likely to degranulate following exposure to cognate peptide epitopes and poorly recognized endogenously processed epitopes from virus-infected B cells. More importantly, ex vivo stimulation of these T cells with an adenoviral vector encoding multiple minimal CD8+ T-cell epitopes as a polyepitope, in combination with a γC cytokine, interleukin-2, restored polyfunctionality and shielded these cells from the inhibitory effects of galectin-1.

scholarly journals Signaling by the Epstein–Barr virus LMP1 protein induces potent cytotoxic CD4+ and CD8+ T cell responses

2018 ◽  
Vol 115 (4) ◽  
pp. E686-E695 ◽  
Author(s):  
Il-Kyu Choi ◽  
Zhe Wang ◽  
Qiang Ke ◽  
Min Hong ◽  
Yu Qian ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Epstein Barr Virus ◽  
T Cell Responses ◽  
Cd8 T Cell ◽  
Cd4 Cells ◽  
Barr Virus ◽  
Cell Responses ◽  
Epstein Barr

The B-lymphotropic Epstein–Barr virus (EBV), pandemic in humans, is rapidly controlled on initial infection by T cell surveillance; thereafter, the virus establishes a lifelong latent infection in the host. If surveillance fails, fatal lymphoproliferation and lymphomagenesis ensue. The initial T cell response consists of predominantly CD8+ cytotoxic T cells and a smaller expansion of CD4+ cells. A major approach to treating EBV-associated lymphomas is adoptive transfer of autologous or allogeneic T cells that are stimulated/expanded on EBV-transformed B cells. Strikingly, the clinical response correlates with the frequency of CD4 cells in the infused T cells. Although in vitro studies suggested that EBV-specific CD4 cells develop cytotoxicity, they have not been comprehensively characterized and the molecular mechanism underlying their formation remains unknown. Our recent work, using a transgenic approach in mice, has revealed a central role for the EBV signaling molecule LMP1 in immune surveillance and transformation of EBV-infected B cells. The mouse model offers a unique tool for uncovering basic features of EBV immunity. Here, we show that LMP1 expression in B cells induces potent cytotoxic CD4 and CD8 T cell responses, by enhancing antigen presentation and costimulation by CD70, OX40 ligand, and 4-1BB ligand. Our data further suggest that cytotoxic CD4 cells hold superior therapeutic value for LMP1 (EBV)-driven lymphomas. These findings provide insights into EBV immunity, demonstrating that LMP1 signaling alone is sufficient to induce a prominent cytotoxic CD4 response, and suggest strategies for immunotherapy in EBV-related and other cancers.

Quantitation, selection, and functional characterization of Epstein-Barr virus–specific and alloreactive T cells detected by intracellular interferon-γ production and growth of cytotoxic precursors

Blood ◽  
2002 ◽  
Vol 99 (5) ◽  
pp. 1730-1740 ◽  
Author(s):  
Guenther Koehne ◽  
Katherine M. Smith ◽  
Teresa L. Ferguson ◽  
Roxanne Y. Williams ◽  
Glenn Heller ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Epstein Barr Virus ◽  
Flow Cytometric Analysis ◽  
Interferon Γ ◽  
Cytotoxic T Cell ◽  
Barr Virus ◽  
Alloreactive T Cells ◽  
Ifn Γ ◽  
Epstein Barr

Techniques for the quantitation of virus-specific and alloantigen-reactive T cells vary in their measurement of clinically relevant T-cell effector populations, their sensitivity and quantitative accuracy, and the time required to obtain measurable results. We compared frequencies of Epstein-Barr virus (EBV)–specific and major alloantigen-reactive T cells as measured by flow cytometric analysis of responding T cells producing intracellular interferon-γ (IFN-γ) and by limiting-dilution analysis (LDA) of cytotoxic T-cell precursors (CTLp) at sequential time points during the generation of EBV-specific T-cell lines. The expansion of EBV-specific T lymphocytes and the depletion of alloreactive T cells in cultures of T cells sensitized with autologous EBV-transformed targets followed similar kinetics when measured by either method. Frequencies of EBV- specific T cells generating intracellular IFN-γ exceeded by 25- to 90-fold the frequencies of responding CTLp at each stage of expansion, whereas the frequencies of alloreactive T cells generating intracellular IFN-γ exceeded by 30- to 220-fold those detected by LDA. The assay that quantitated T cells producing IFN-γ yielded more reproducible and precise results than LDA. Furthermore, frequencies detected by the enumeration of T cells responding to immunodominant EBNA 3a and EBNA 3c peptides by IFN-γ production or their capacity to bind peptide-HLA tetramers were strikingly similar and represented significant fractions of T cells generating IFN-γ in response to autologous EBV B lymphoblastoid cell line. Functional analysis of responding viable T cells, fractionated on the basis of their secretion of IFN-γ, demonstrated that EBV-specific and alloantigen cytotoxic T cells were predominately or exclusively detected in the CD8+IFN-γ+ fraction of T cells. Strikingly, the CD4+IFN-γ+ cell fractions were not cytotoxic against EBV-transformed or allogeneic targets.

scholarly journals Determination of Optimal Target Antigen and Immune Modulatory Approaches to Improve and Standardize Epstein-Barr Virus-Specific Adoptive T Cell Therapy

Blood ◽  
2014 ◽  
Vol 124 (21) ◽  
pp. 5817-5817
Author(s):  
Monica L Mitchell ◽  
John T. Patton ◽  
Lynn O'Donnell ◽  
Robert A. Baiocchi
Keyword(s):  
T Cells ◽  
T Cell ◽  
Epstein Barr Virus ◽  
Ex Vivo ◽  
Viral Proteins ◽  
Adoptive T Cell Therapy ◽  
T Cell Therapy ◽  
Barr Virus ◽  
Epstein Barr ◽  
Amino Acid Depletion

Abstract Epstein-Barr Virus (EBV) is a ubiquitous gamma herpesvirus that infects lymphocytes and epithelial cells, establishing lifelong latency. Immunosuppression is associated with a transition from a quiescent, latent EBV infection to either a lytic or active latent life cycle, which involves transformation of memory B cells to rapidly growing lymphoblastoid cell lines (LCLs). Due to the immunogenicity of EBV viral proteins, cells expressing most latent or lytic peptides are eradicated through immune surveillance mechanisms. EBV-associated lymphoproliferative disorders (LPDs) arise in immunocompromised individuals and are facilitated by the expression of the same viral proteins that are expressed in the immune tolerant environment. Furthermore, EBV-LPDs acquire various mechanisms to suppress immune function, such as expression of the programmed death ligand (PD-L1), CTLA-4, amino acid depletion, and propagation of suppressive immune cell subsets, including tumor-associated macrophages (TAMs) that attenuate T cell receptor signaling and cytokine production. These immunosuppressive mechanisms increase the risk of acquiring EBV viremia and can facilitate the development of LPDs. In other work reported by our group (Patton et al, ASH abstract 2014), a TAM-like population has recently been discovered to spontaneously expand in peripheral blood mononuclear cell (PBMC)/LCL co-cultures. These TAM-like macrophages are capable of potent cytotoxicity against EBV-specific T cells, thus novel strategies to counteract this negative regulatory network are needed. One effective strategy developed to reestablish EBV-specific immunity is adoptive T cell therapy. However, the optimal combination of target antigens required to protect against EBV-LPD is not well characterized. It remains plausible that expansion of cytotoxic and/or helper T cell populations specific for multiple EBV antigens will improve the efficiency of the T cell response. The suppression of negative immune regulators may further enhance the effectiveness of the adoptive therapies to provide optimal control of viremia and effectively treat EBV-driven malignancies. Here, we generated EBV-specific T cell preparations by culturing PBMCs supplemented with interleukins 4 and 7 (IL4, IL7) in the presence of latent (EBNA1, EBNA3, LMP1, LMP2) or lytic (BZLF1, BRLF, BMLF1, BMRF1) PepMixes (complete protein-spanning pools of overlapping peptides). Cells were separated into two groups, a test and expansion group, to determine optimal ex vivo expansion. Activity of T cells was measured by direct flow-based cytotoxicity against LCL targets and interferon gamma (IFNγ) production via Enzyme-Linked Immuno Spot (ELISpot). Preliminary ELISpot assays from two donors in the test group showed that BZLF1-specific T cells produced as much as 34-fold more IFNγ in comparison to other EBV antigens (standardized to a negative control). T cells produced in culture with a peptivator PepMix, a collection of 13 latent and lytic peptide pools, on average exhibited the second highest levels of IFNγ release, expressing as much as 1.4-fold more IFNγ than the next highest antigen-specific response, depending on the donor. Additionally, LMP1 produced as much as 24-fold higher levels of IFNγ. Therefore, given the heterogeneous EBV viral gene expression profiles and variation of immune dominance across human leukocyte antigen (HLA) types, use of polyclonal immunogen-specific T cells may be more effective in improving survival and long-term protection from EBV-driven complications via improved memory T cell surveillance. Additional ex vivo work is currently underway examining combinations of lytic and latent antigens based on immunodominance analysis across HLA types. Finally, use of immune modulatory mechanisms to address newly discovered TAM-like populations contributing to reduced immune effector function against EBV-LPD are currently being evaluated. We are testing strategies to promote PD-L1 and CTLA-4 blockade, inhibition of amino acid depletion, and down regulation of regulatory T cells, to allow for enhanced expansion of antigen-specific T cell populations. Confirmation of these hypotheses will be directed toward developing streamlined methods for efficient, rapid, and personalized T cell preparation that can be standardized for translation into clinical trials. Disclosures No relevant conflicts of interest to declare.

Peripheral T-cell Lymphoma, NOS With Epstein-Barr Virus Positivity in an Elderly Patient With Myelodysplastic Syndrome: An Autopsy Case Report

2020 ◽  
Vol 154 (Supplement_1) ◽  
pp. S81-S81
Author(s):  
J Lanceta ◽  
W Xue ◽  
M Hurford ◽  
H Wu
Keyword(s):  
Bone Marrow ◽  
T Cells ◽  
Lymph Node ◽  
T Cell ◽  
Epstein Barr Virus ◽  
Cell Lymphoma ◽  
T Cell Lymphoma ◽  
Peripheral T Cell Lymphoma ◽  
Barr Virus ◽  
Epstein Barr

Abstract Casestudy Epstein-Barr virus (EBV)-associated peripheral T-cell lymphomas are a group of aggressive neoplasms with a geographic predilection for South America and Asia, but are very rare in Western populations. Results We report a case of a 74-year-old Caucasian female who presented with pancytopenia and B symptoms with EBV-IgG detected on admission. Past medical history included: ITP, chronic urticaria, and recently diagnosed myelodysplastic syndrome (MDS) on bone marrow biopsy one month prior to admission. Excisional biopsies of an enlarged right neck lymph node (repeated within 6 months) and right axillary lymph node five years ago were negative for a lymphoproliferative disorder at the time. Repeated bone marrow biopsy, performed during the current admission, confirmed the diagnosis of MDS, with scattered T-cells without aberrant immunophenotype. Despite aggressive treatment from multiple specialties, the patient deteriorated and expired four weeks later from complications of MDS. At autopsy, there was diffuse lymphadenopathy involving the mediastinum, axilla, pelvis and peripancreatic fat. Lymph node sections demonstrated nodal architecture effacement by diffuse, vaguely nodular lymphoid infiltrates. Histologically, the infiltrates were composed of medium to large lymphocytes with round to slight irregular nuclei, rare Reed-Sternberg-like multinucleated cells, clumped chromatin, and indistinct nucleoli. Individual cell necrosis was abundant with mitotic figures readily identifiable. Immunohistochemistry revealed CD2+ CD3+ neoplastic T-cells that co-express MUM1 and a subset of CD30, while negative for CD4, CD5, CD8, CD56, ALK1, and TDT. EBV-encoded RNA in-situ hybridization was focally positive. The final postmortem diagnosis was peripheral T-cell lymphoma, not otherwise specified (NOS), with focal EBV positivity. Conclusion Co-existence of a de-novo MDS and non-Hodgkin lymphoma without any prior chemotherapeutic exposure is a highly unusual finding, although MDS-like presentations can occur with EBV-associated lymphomas. Peripheral T-cell lymphoma, NOS is an aggressive lymphoma and EBV positivity has been found correlated with a poor prognosis. This case demonstrates how postmortem examination remains an important tool in clinical- pathological correlation and highlights the potential pathogenetic role EBV plays in MDS and T-cell lymphoma.

Infusion of Cytotoxic T Cells for the Prevention and Treatment of Epstein-Barr Virus–Induced Lymphoma in Allogeneic Transplant Recipients

Blood ◽  
1998 ◽  
Vol 92 (5) ◽  
pp. 1549-1555 ◽  
Author(s):  
Cliona M. Rooney ◽  
Colton A. Smith ◽  
Catherine Y.C. Ng ◽  
Susan K. Loftin ◽  
John W. Sixbey ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Epstein Barr Virus ◽  
Mononuclear Cells ◽  
Cytotoxic T Cells ◽  
Marker Gene ◽  
Barr Virus ◽  
Immunoblastic Lymphoma ◽  
Epstein Barr ◽  
T Cell Lines

Abstract Epstein-Barr virus (EBV) causes potentially lethal immunoblastic lymphoma in up to 25% of children receiving bone marrow transplants from unrelated or HLA-mismatched donors. Because this complication appears to stem from a deficiency of EBV-specific cytotoxic T cells, we assessed the safety and efficacy of donor-derived polyclonal (CD4+ and CD8+) T-cell lines as immunoprophylaxis and treatment for EBV-related lymphoma. Thirty-nine patients considered to be at high risk for EBV-induced lymphoma each received 2 to 4 intravenous infusions of donor-derived EBV-specific T lymphocytes, after they had received T-cell–depleted bone marrow from HLA-matched unrelated donors (n = 33) or mismatched family members (n = 6). The immunologic effects of this therapy were monitored during and after the infusions. Infused cells were identified by detection of the neo marker gene. EBV-specific T cells bearing theneo marker were identified in all but 1 of the patients. Serial analysis of DNA detected the marker gene for as long as 18 weeks in unmanipulated peripheral blood mononuclear cells and for as long as 38 months in regenerated lines of EBV-specific cytotoxic T cells. Six patients (15.5%) had greatly increased amounts of EBV-DNA on study entry (>2,000 genome copies/106 mononuclear cells), indicating uncontrolled EBV replication, a complication that has had a high correlation with subsequent development of overt lymphoma. All of these patients showed 2 to 4 log decreases in viral DNA levels within 2 to 3 weeks after infusion and none developed lymphoma, confirming the antiviral activity of the donor-derived cells. There were no toxic effects that could be attributed to prophylactic T-cell therapy. Two additional patients who did not receive prophylaxis and developed overt immunoblastic lymphoma responded fully to T-cell infusion. Polyclonal donor-derived T-cell lines specific for EBV proteins can thus be used safely to prevent EBV-related immunoblastic lymphoma after allogeneic marrow transplantation and may also be effective in the treatment of established disease. © 1998 by The American Society of Hematology.

scholarly journals Analysis of the defects responsible for the impaired regulation of Epstein-Barr virus-induced B cell proliferation by rheumatoid arthritis lymphocytes. I. Diminished gamma interferon production in response to autologous stimulation.

1983 ◽  
Vol 157 (1) ◽  
pp. 173-188 ◽  
Author(s):  
F Hasler ◽  
H G Bluestein ◽  
N J Zvaifler ◽  
L B Epstein
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
T Cell ◽  
B Cell ◽  
Epstein Barr Virus ◽  
Gamma Interferon ◽  
Barr Virus ◽  
T Cell Regulation ◽  
Epstein Barr

T cells of patients with rheumatoid arthritis (RA) do not control the rate of B lymphoblast transformation induced by Epstein-Barr virus (EBV) as efficiently as T cells from healthy individuals; thus, lymphoblast cell lines are established more readily in RA lymphocytes in vitro after EBV infection. In the present experiments, we have asked whether this T cell regulation can be reproduced by lymphocytes. We found that normal T cells, activated in allogeneic or autologous mixed leukocyte reactions (MLR), produce lymphokines that inhibit in vitro EBV-induced B cell proliferation. Allogeneic MLR supernatants inhibited EBV-induced DNA synthesis 62 +/- 4% (mean +/- SE) at 10 d post-infection, whereas autologous MLR supernatants suppressed it 50 +/- 3%. RA T cell supernatants produced in an allogeneic MLR suppressed as well as normal T cell supernatants (64 +/- 5% inhibition). In contrast, supernatants from RA autologous MLR had little inhibitory activity. EBV-induced DNA synthesis at 10 d was reduced only 8 +/- 3%, compared with the 50 +/- 3% suppressive activity of normal autologous MLR supernatants. The magnitude of the proliferative responses in the autologous MLR regenerating the lymphokines was similar in the normal and RA populations. After depletion of adherent cells from the RA auto-MLR stimulators, supernatant inhibitory activities increased to normal levels (from 11 +/- 6 [SE] to 52 +/- 6% [SE]). The inhibitory factor involved in the regulation of in vitro EBV infection is a protein with a molecular weight of approximately 50,000. Its activity is eliminated by hearing at 56 degrees C and by exposure to acid at pH 2. The inhibitory activity is blocked by mixing the MLR supernatants with a polyvalent antisera or monoclonal antibodies specific for human gamma interferon. Gamma interferon produced by activating T cells in allo- or auto-MLR can reproduce T cell-mediated regulation of EBV-induced B cell proliferation, and the failure of RA auto-MLR to generate that lymphokine parallels the defective T cell regulation of EBV-induced B cell proliferation characteristic of RA lymphoid cells.

Infusion of Cytotoxic T Cells for the Prevention and Treatment of Epstein-Barr Virus–Induced Lymphoma in Allogeneic Transplant Recipients

Blood ◽  
1998 ◽  
Vol 92 (5) ◽  
pp. 1549-1555 ◽  
Author(s):  
Cliona M. Rooney ◽  
Colton A. Smith ◽  
Catherine Y.C. Ng ◽  
Susan K. Loftin ◽  
John W. Sixbey ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Epstein Barr Virus ◽  
Mononuclear Cells ◽  
Cytotoxic T Cells ◽  
Marker Gene ◽  
Barr Virus ◽  
Immunoblastic Lymphoma ◽  
Epstein Barr ◽  
T Cell Lines

Epstein-Barr virus (EBV) causes potentially lethal immunoblastic lymphoma in up to 25% of children receiving bone marrow transplants from unrelated or HLA-mismatched donors. Because this complication appears to stem from a deficiency of EBV-specific cytotoxic T cells, we assessed the safety and efficacy of donor-derived polyclonal (CD4+ and CD8+) T-cell lines as immunoprophylaxis and treatment for EBV-related lymphoma. Thirty-nine patients considered to be at high risk for EBV-induced lymphoma each received 2 to 4 intravenous infusions of donor-derived EBV-specific T lymphocytes, after they had received T-cell–depleted bone marrow from HLA-matched unrelated donors (n = 33) or mismatched family members (n = 6). The immunologic effects of this therapy were monitored during and after the infusions. Infused cells were identified by detection of the neo marker gene. EBV-specific T cells bearing theneo marker were identified in all but 1 of the patients. Serial analysis of DNA detected the marker gene for as long as 18 weeks in unmanipulated peripheral blood mononuclear cells and for as long as 38 months in regenerated lines of EBV-specific cytotoxic T cells. Six patients (15.5%) had greatly increased amounts of EBV-DNA on study entry (>2,000 genome copies/106 mononuclear cells), indicating uncontrolled EBV replication, a complication that has had a high correlation with subsequent development of overt lymphoma. All of these patients showed 2 to 4 log decreases in viral DNA levels within 2 to 3 weeks after infusion and none developed lymphoma, confirming the antiviral activity of the donor-derived cells. There were no toxic effects that could be attributed to prophylactic T-cell therapy. Two additional patients who did not receive prophylaxis and developed overt immunoblastic lymphoma responded fully to T-cell infusion. Polyclonal donor-derived T-cell lines specific for EBV proteins can thus be used safely to prevent EBV-related immunoblastic lymphoma after allogeneic marrow transplantation and may also be effective in the treatment of established disease. © 1998 by The American Society of Hematology.

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