scholarly journals CAR-T cells and TRUCKs that recognize an EBNA-3C-derived epitope presented on HLA-B*35 control Epstein-Barr virus-associated lymphoproliferation

2020 ◽  
Vol 8 (2) ◽  
pp. e000736
Author(s):  
Anna Christina Dragon ◽  
Katharina Zimmermann ◽  
Thomas Nerreter ◽  
Deborah Sandfort ◽  
Julia Lahrberg ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Immune Cells ◽  
Epstein Barr Virus ◽  
Target Cells ◽  
Barr Virus ◽  
Car T Cells ◽  
Car T ◽  
Epstein Barr

BackgroundImmunosuppressive therapy or T-cell depletion in transplant patients can cause uncontrolled growth of Epstein-Barr virus (EBV)-infected B cells resulting in post-transplant lymphoproliferative disease (PTLD). Current treatment options do not distinguish between healthy and malignant B cells and are thereby often limited by severe side effects in the already immunocompromised patients. To specifically target EBV-infected B cells, we developed a novel peptide-selective chimeric antigen receptor (CAR) based on the monoclonal antibody TÜ165 which recognizes an Epstein-Barr nuclear antigen (EBNA)−3C-derived peptide in HLA-B*35 context in a T-cell receptor (TCR)-like manner. In order to attract additional immune cells to proximity of PTLD cells, based on the TÜ165 CAR, we moreover generated T cells redirected for universal cytokine-mediated killing (TRUCKs), which induce interleukin (IL)-12 release on target contact.MethodsTÜ165-based CAR-T cells (CAR-Ts) and TRUCKs with inducible IL-12 expression in an all-in-one construct were generated. Functionality of the engineered cells was assessed in co-cultures with EBNA-3C-peptide-loaded, HLA-B*35-expressing K562 cells and EBV-infected B cells as PTLD model. IL-12, secreted by TRUCKs on target contact, was further tested for its chemoattractive and activating potential towards monocytes and natural killer (NK) cells.ResultsAfter co-cultivation with EBV target cells, TÜ165 CAR-Ts and TRUCKs showed an increased activation marker expression (CD137, CD25) and release of proinflammatory cytokines (interferon-γ and tumor necrosis factor-α). Moreover, TÜ165 CAR-Ts and TRUCKs released apoptosis-inducing mediators (granzyme B and perforin) and were capable to specifically lyse EBV-positive target cells. Live cell imaging revealed a specific attraction of TÜ165 CAR-Ts around EBNA-3C-peptide-loaded target cells. Of note, TÜ165 TRUCKs with inducible IL-12 showed highly improved effector functions and additionally led to recruitment of monocyte and NK cell lines.ConclusionsOur results demonstrate that TÜ165 CAR-Ts recognize EBV peptide/HLA complexes in a TCR-like manner and thereby allow for recognizing an intracellular EBV target. TÜ165 TRUCKs equipped with inducible IL-12 expression responded even more effectively and released IL-12 recruited additional immune cells which are generally missing in proximity of lymphoproliferation in immunocompromised PTLD patients. This suggests a new and promising strategy to specifically target EBV-infected cells while sparing and mobilizing healthy immune cells and thereby enable control of EBV-associated lymphoproliferation.

scholarly journals Targeting CAR to the Peptide-MHC Complex Reveals Distinct Signaling Compared to That of TCR in a Jurkat T Cell Model

Cancers ◽  
2021 ◽  
Vol 13 (4) ◽  
pp. 867
Author(s):  
Ling Wu ◽  
Joanna Brzostek ◽  
Shvetha Sankaran ◽  
Qianru Wei ◽  
Jiawei Yap ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cell Model ◽  
Cell Receptor ◽  
Target Cells ◽  
Tcr Signaling ◽  
Barr Virus ◽  
Car T ◽  
Epstein Barr ◽  
Lower Magnitude

Chimeric antigen receptor T cells (CAR-T) utilize T cell receptor (TCR) signaling cascades and the recognition functions of antibodies. This allows T cells, normally restricted by the major histocompatibility complex (MHC), to be redirected to target cells by their surface antigens, such as tumor associated antigens (TAAs). CAR-T technology has achieved significant successes in treatment of certain cancers, primarily liquid cancers. Nonetheless, many challenges hinder development of this therapy, such as cytokine release syndrome (CRS) and the efficacy of CAR-T treatments for solid tumors. These challenges show our inadequate understanding of this technology, particularly regarding CAR signaling, which has been less studied. To dissect CAR signaling, we designed a CAR that targets an epitope from latent membrane protein 2 A (LMP2 A) of the Epstein–Barr virus (EBV) presented on HLA*A02:01. Because of this, CAR and TCR signaling can be compared directly, allowing us to study the involvement of other signaling molecules, such as coreceptors. This comparison revealed that CAR was sufficient to bind monomeric antigens due to its high affinity but required oligomeric antigens for its activation. CAR sustained the transduced signal significantly longer, but at a lower magnitude, than did TCR. CD8 coreceptor was recruited to the CAR synapse but played a negligible role in signaling, unlike for TCR signaling. The distinct CAR signaling processes could provide explanations for clinical behavior of CAR-T therapy and suggest ways to improve the technology.

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 CAR-T Cells Targeting Epstein-Barr Virus gp350 Validated in a Humanized Mouse Model of EBV Infection and Lymphoproliferative Disease

2020 ◽  
Vol 18 ◽  
pp. 504-524
Author(s):  
Constanze Slabik ◽  
Maja Kalbarczyk ◽  
Simon Danisch ◽  
Reinhard Zeidler ◽  
Frank Klawonn ◽  
...  
Keyword(s):  
T Cells ◽  
Epstein Barr Virus ◽  
Lymphoproliferative Disease ◽  
Humanized Mouse ◽  
Humanized Mouse Model ◽  
Barr Virus ◽  
Ebv Infection ◽  
Car T Cells ◽  
Car T ◽  
Epstein Barr

scholarly journals Preferential Localization of the Epstein-Barr Virus (EBV) Oncoprotein LMP-1 to Nuclei in Human T Cells: Implications for Its Role in the Development of EBV Genome-Positive T-Cell Lymphomas

2002 ◽  
Vol 76 (8) ◽  
pp. 4080-4086 ◽  
Author(s):  
Jingwu Xu ◽  
Ali Ahmad ◽  
José Menezes
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Epstein Barr Virus ◽  
Phenotypic Changes ◽  
Barr Virus ◽  
T Cell Lymphomas ◽  
Cell Clones ◽  
Epstein Barr

ABSTRACT The Epstein-Barr virus (EBV)-encoded latent membrane protein-1 (LMP-1) is thought to play a role in the EBV-induced B-cell transformation and immortalization. EBV has also been implicated in certain human T-cell lymphomas; however, the phenotypic effects of the expression of this oncoprotein in T cells are not known. To learn whether LMP-1 also induces phenotypic changes in T cells, we stably expressed it in human cell lines of T and B lineages and 25 LMP-1-expressing T-cell clones and 7 B-cell clones were examined. Our results show for the first time that, in sharp contrast to B cells, LMP-1 preferentially localizes to nuclei in T cells and does not induce the phenotypic changes in these cells that it induces in B cells, does not associate with TRAF proteins, and does not arrest the cell cycle in the G2/M phase. A computer-assisted analysis revealed that LMP-1 lacks the canonical nuclear localization signal. Our results suggest that this oncoprotein may not play the same role in the lymphomagenesis of T cells as it does in B cells.

scholarly journals Heterogeneous Epstein-Barr virus infection patterns in peripheral T- cell lymphoma of angioimmunoblastic lymphadenopathy type

Blood ◽  
1992 ◽  
Vol 80 (7) ◽  
pp. 1804-1812 ◽  
Author(s):  
I Anagnostopoulos ◽  
M Hummel ◽  
T Finn ◽  
M Tiemann ◽  
P Korbjuhn ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
Epstein Barr Virus ◽  
Cell Lymphoma ◽  
Angioimmunoblastic Lymphadenopathy ◽  
Barr Virus ◽  
Ebv Infection ◽  
Infected Cells ◽  
Epstein Barr

Abstract In this study, 32 cases of T-cell lymphoma of angioimmunoblastic lymphadenopathy type (AILD-TCL) were investigated for their association with Epstein-Barr virus (EBV). For this purpose, three different approaches were applied: polymerase chain reaction (PCR) for the presence of EBV-DNA, in situ hybridization (ISH) for EBV-encoded small nuclear RNAs (EBER), and immunohistology for EBV-encoded latent membrane protein (LMP). PCR and EBER-ISH produced almost identical results, showing that all but one case of AILD-TCL contained EBV genomes. Three distinctive patterns of EBV infection were observed after immunophenotypical characterization of EBER-positive cells: (1) in 26% of the cases, B and T cells were infected, the majority of which were B cells of immunoblastic morphology located in the remnants of lymphoid follicles; (2) in 42% of the cases, the vast majority of infected cells were neoplastic T cells diffusely distributed in the lymph nodes, but infected B cells were also present; and (3) in 32% of the cases, there were only a few infected small lymphoid cells. Detectable LMP was frequent in cases exhibiting patterns 1 and 2. These findings suggest that in AILD-TCL patients, B cells and especially T cells are highly susceptible to a persistent EBV infection, which often leads to a growth advantage of the infected cells. Thus EBV, in conjunction with genetic abnormalities and selective defects of the immune system, might be involved in the pathogenesis of AILD-TCL.

scholarly journals CD4+ T-Cell Effectors Inhibit Epstein-Barr Virus-Induced B-Cell Proliferation

2001 ◽  
Vol 75 (8) ◽  
pp. 3740-3752 ◽  
Author(s):  
Sarah Nikiforow ◽  
Kim Bottomly ◽  
George Miller
Keyword(s):  
Cell Proliferation ◽  
T Cells ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Cell Lines ◽  
Epstein Barr Virus ◽  
Barr Virus ◽  
Epstein Barr

ABSTRACT In immunodeficient hosts, Epstein-Barr virus (EBV) often induces extensive B-cell lymphoproliferative disease and lymphoma. Without effective in vitro immune surveillance, B cells infected by the virus readily form immortalized cell lines. In the regression assay, memory T cells inhibit the formation of foci of EBV-transformed B cells that follows recent in vitro infection by EBV. No one has yet addressed which T cell regulates the early proliferative phase of B cells newly infected by EBV. Using new quantitative methods, we analyzed T-cell surveillance of EBV-mediated B-cell proliferation. We found that CD4+ T cells play a significant role in limiting proliferation of newly infected, activated CD23+ B cells. In the absence of T cells, EBV-infected CD23+ B cells divided rapidly during the first 3 weeks after infection. Removal of CD4+ but not CD8+ T cells also abrogated immune control. Purified CD4+ T cells eliminated outgrowth when added to EBV-infected B cells. Thus, unlike the killing of EBV-infected lymphoblastoid cell lines, in which CD8+ cytolytic T cells play an essential role, prevention of early-phase EBV-induced B-cell proliferation requires CD4+ effector T cells.

scholarly journals Functional Analysis of the CD4+ T-Cell Response to Epstein-Barr Virus: T-Cell-Mediated Activation of Resting B Cells and Induction of Viral BZLF1 Expression

2000 ◽  
Vol 74 (14) ◽  
pp. 6675-6679 ◽  
Author(s):  
Zheng Fu ◽  
Martin J. Cannon
Keyword(s):  
Functional Analysis ◽  
T Cells ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Epstein Barr Virus ◽  
Lymphoproliferative Disorders ◽  
T Cell Response ◽  
Barr Virus ◽  
Epstein Barr

ABSTRACT In contrast to the major role played by Epstein-Barr virus (EBV)-specific CD8+ cytotoxic T-cell responses in immunosurveillance, recent studies have offered the apparently paradoxical suggestion that development of EBV-driven human B-cell lymphoproliferative disorders and tumors in SCID/hu mice is dependent on the presence of T cells, in particular CD4+ T cells. This study presents a functional analysis of the CD4+T-cell response to EBV and shows that while CD4+ T cells may be cytotoxic, they also express Th2 cytokines and CD40 ligand (gp39) and possess B-cell helper function. We show that EBV-specific CD4+ T cells can provide non-HLA-restricted help for activation of resting B cells via a gp39-CD40-dependent pathway and are able to induce expression of BZLF1, a viral lytic cycle transactivator in latently infected resting B cells, ultimately resulting in rapid outgrowth of transformed B-cell colonies. These results support the proposal that CD4+ T cells may play a key role in reactivation of latent EBV infection and may thus contribute to the pathogenesis of EBV-driven lymphoproliferative disorders.

scholarly journals Combined immunodeficiency and Epstein-Barr virus–induced B cell malignancy in humans with inherited CD70 deficiency

2016 ◽  
Vol 214 (1) ◽  
pp. 91-106 ◽  
Author(s):  
Hassan Abolhassani ◽  
Emily S.J. Edwards ◽  
Aydan Ikinciogullari ◽  
Huie Jing ◽  
Stephan Borte ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
B Cells ◽  
B Cell ◽  
Cd8 T Cells ◽  
Epstein Barr Virus ◽  
Combined Immunodeficiency ◽  
Memory Cd8 T Cells ◽  
Barr Virus ◽  
Epstein Barr

In this study, we describe four patients from two unrelated families of different ethnicities with a primary immunodeficiency, predominantly manifesting as susceptibility to Epstein-Barr virus (EBV)–related diseases. Three patients presented with EBV-associated Hodgkin’s lymphoma and hypogammaglobulinemia; one also had severe varicella infection. The fourth had viral encephalitis during infancy. Homozygous frameshift or in-frame deletions in CD70 in these patients abolished either CD70 surface expression or binding to its cognate receptor CD27. Blood lymphocyte numbers were normal, but the proportions of memory B cells and EBV-specific effector memory CD8+ T cells were reduced. Furthermore, although T cell proliferation was normal, in vitro–generated EBV-specific cytotoxic T cell activity was reduced because of CD70 deficiency. This reflected impaired activation by, rather than effects during killing of, EBV-transformed B cells. Notably, expression of 2B4 and NKG2D, receptors implicated in controlling EBV infection, on memory CD8+ T cells from CD70-deficient individuals was reduced, consistent with their impaired killing of EBV-infected cells. Thus, autosomal recessive CD70 deficiency is a novel cause of combined immunodeficiency and EBV-associated diseases, reminiscent of inherited CD27 deficiency. Overall, human CD70–CD27 interactions therefore play a nonredundant role in T and B cell–mediated immunity, especially for protection against EBV and humoral immunity.

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