CD4+ T-Cell Responses to Epstein-Barr Virus (EBV) Latent-Cycle Antigens and the Recognition of EBV-Transformed Lymphoblastoid Cell Lines

CRUK Institute for Cancer Studies, The University of Birmingham, Vincent Dr., Edgbaston, Birmingham B15 2TT, United Kingdom.
Journal of Virology (Impact Factor: 4.44). 05/2005; 79(8):4896-907. DOI: 10.1128/JVI.79.8.4896-4907.2005
Source: PubMed


There is considerable interest in the potential of Epstein-Barr virus (EBV) latent antigen-specific CD4+ T cells to act as direct effectors controlling EBV-induced B lymphoproliferations. Such activity would require direct CD4+ T-cell recognition of latently infected cells through epitopes derived from endogenously expressed viral proteins and presented
on the target cell surface in association with HLA class II molecules. It is therefore important to know how often these conditions
are met. Here we provide CD4+ epitope maps for four EBV nuclear antigens, EBNA1, -2, -3A, and -3C, and establish CD4+ T-cell clones against 12 representative epitopes. For each epitope we identify the relevant HLA class II restricting allele
and determine the efficiency with which epitope-specific effectors recognize the autologous EBV-transformed B-lymphoblastoid
cell line (LCL). The level of recognition measured by gamma interferon release was consistent among clones to the same epitope
but varied between epitopes, with values ranging from 0 to 35% of the maximum seen against the epitope peptide-loaded LCL.
These epitope-specific differences, also apparent in short-term cytotoxicity and longer-term outgrowth assays on LCL targets,
did not relate to the identity of the source antigen and could not be explained by the different functional avidities of the
CD4+ clones; rather, they appeared to reflect different levels of epitope display at the LCL surface. Thus, while CD4+ T-cell responses are detectable against many epitopes in EBV latent proteins, only a minority of these responses are likely
to have therapeutic potential as effectors directly recognizing latently infected target cells.

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Available from: Elise Landais, Jan 08, 2014
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    • "One of the most extensively studied ThCTL subsets in humans are those generated against Epstein Barr Virus (EBV), a herpes virus typically harbored in latent form by B cells. ThCTL have been found to recognize both lytic and latent EBV class II antigens presented by conventional and transformed B cells [16–19]. ThCTL have also been identified in HIV-1 seropositive individuals [7, 12], a lentivirus that infects professional APCs and CD4 T cells that can also express class II upon activation in humans but not in mice [20–22]. "
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    ABSTRACT: CD4 T cells that acquire cytotoxic phenotype and function have been repeatedly identified in humans, mice, and other species in response to many diverse pathogens. Since CD4 cytotoxic T cells are able to recognize antigenic determinants unique from those recognized by the parallel CD8 cytotoxic T cells, they can potentially contribute additional immune surveillance and direct effector function by lysing infected or malignant cells. Here, we briefly review much of what is known about the generation of cytotoxic CD4 T cells and describe our current understanding of their role in antiviral immunity. Furthering our understanding of the many roles of CD4 T cells during an anti-viral response is important for developing effective vaccine strategies that promote long-lasting protective immunity.
    BioMed Research International 11/2011; 2011(4):954602. DOI:10.1155/2011/954602 · 2.71 Impact Factor
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    • "Outgrowth assay was carried out as previously described [12]. Briefly, target LCL were seeded as replicates in U-bottom 96-well plates at doubling dilution, starting from 104 cells/well to 78 cells/well. "
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    ABSTRACT: Recent preclinical adoptive immunotherapy studies in murine models prompt to employ "proper" rather than "as many as possible" antigen-specific T cells to gain better therapeutic results. Ideally, "proper" T cells are poorly differentiated in vitro, but retain the capacity to fully differentiate into effector cells in vivo, where they can undergo long-term survival and strong proliferation. Such requirements can be achieved by modifying culture conditions, namely using less "differentiating" cytokines than IL-2. To evaluate this issue in human T cell cultures, we exploited a well characterized and clinical-grade protocol finalized at generating EBV-specific CTL for adoptive immunotherapy. In particular, we studied the impact of IL-7, IL-15 and IL-21 compared to IL-2 on different aspects of T cell functionality, namely growth kinetics, differentiation/activation marker expression, cytokine production, and short-term and long-term cytotoxicity. Results disclosed that the culture modifications we introduced in the standard protocol did not improve activity nor induce substantial changes in differentiation marker expression of EBV-specific CTL. Our data indicated that the addition of γ-chain cytokines other than IL-2 for the generation of EBV-specific T cell cultures did not produce the improvements expected on the basis of recent published literature. This fact was likely due to the intrinsic differences between murine and human models and highlights the need to design ad hoc protocols rather than simply modify the cytokines added in culture.
    Journal of Translational Medicine 11/2010; 8(1):121. DOI:10.1186/1479-5876-8-121 · 3.93 Impact Factor
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    • "The results in Fig. 2 suggest that this recognition is impaired by the V-protein through its multiple effects on the MHC class I antigen presentation pathway and on the expression of adhesion molecules necessary for efficient CTL–target binding. To test this hypothesis, we compared the recognition of IARC-171 pBabe and V-protein LCLs by EBV-specific CD8 + CTLs using an ELISA for IFN-c released by T cells that gives a quantitative measurement of the T cell recognition of the EBV + target cells (Long et al., 2005). This assay was only performed on IARC-171 pBabe and V-protein LCLs as they demonstrated the biggest difference in MHC class I cell surface expression (Fig. 2) and the HLA-type was matched to two available T cell clones: HPV (Blake et al., 1997) and AVF (Gavioli et al., 1993). "
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    ABSTRACT: The transformation of B cells by Epstein-Barr virus (EBV), into lymphoblastoid cell lines (LCLs) results in the upregulation of STAT1, a key transcription factor in the interferon signalling pathway. Although the mechanism of EBV induction of STAT1 protein expression has been intensively studied, there has been little investigation into the function of STAT1 in EBV-transformed LCLs. In this study, we have implemented a novel strategy to investigate the functional role of STAT1 through the introduction of the simian virus 5 (SV5) V-protein into LCLs by retroviral gene transfer. The V-protein is a virally evolved STAT1 inhibitor that specifically targets STAT1 for proteasomal degradation. Using this in vitro model, we have shown that major histocompatibility complex (MHC) class I and class II molecules are downregulated at the cell surface following a reduction in STAT1 protein expression. With regards to MHC class I, the impairment of the antigen processing machinery renders the cells less recognized by the host EBV-specific immunosurveillance. In addition, downregulation of STAT1 increases the expression of LMP2A and lytic cycle antigens and results in a higher proportion of cells entering the lytic cycle. These results suggest that STAT1 is involved in maintaining the latency III viral program observed in transformed B cells and regulating immunorecognition by EBV-specific T cells.
    Journal of General Virology 08/2009; 90(9-Pt 9):2239-50. DOI:10.1099/vir.0.011627-0 · 3.18 Impact Factor
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