Mechanisms of immune escape in EBV associated malignancies : Hodgkin's disease and Burkitt's lymphoma

The work presented in this thesis is focused on the analysis of the mechanisms operating in the immune escape of two Epstein-Barr virus (EBV) associated tumors: Hodgkin's Disease (HD) and Burkitt's lymphoma (BL).

EBV positive HD tumor cells express at least 3 viral proteins: the EBV nuclear antigen (EBNA)1, the latent membrane proteins (LMP)1 and LMP2A. LMP1 and LMP2A are targets for cytotoxic T lymphocyte (CTL) mediated responses. To examine whether EBV triggered reactivities can be detected within tumor, infiltrating lymphocytes (TILs), cytokine mRNA expression, cell phenotype and EBV specific Cytotoxic activity were analysed in biopsies from 8 EBV carrying and 6 EBV negative HD lesions. Neither the pattern of lymphokine production, nor the cell phenotype of the in vivo activated IL-2 responding T cell population provided a clear discrimination between EBV+ and EBV- cases. HLA class I restricted EBV specific cytotoxicity was demonstrated in IL-2 dependent cultures of TILs from 3 out of 3 EBV- tumors, whereas cultures from 6 out of 6 EBV+ tumors were either noncytotoxic, or exerted LAK-type cytotoxicity. EBV specific CTL precursors were present in the blood of one patient, carrying an EBV positive HD. These results suggest that suppression of EBV specific T cell responses at the tumor site may be an important component in the pathogenesis of EBV+ HD.

EBV positive BL derived cell lines do not present endogenous antigens to CD8+ CTLs. Impaired presentation of the HLA A11-restricted epitope E4416-424 derived from the viral protein EBNA4 was associated with downregulation of the peptide transporters associated with antigen processing (TAP)1 and TAP2 and with altered expression of certain enzymatic and regulatory subunits of the proteasome. This may result in the production of different sets of endogenous peptides. Stimulation of BL cells via CD40 cross-linking induced upregulation of adhesion and costimulatory molecules, MHC class I and increased TAP1 and TAP2 expression, without affecting the subunit composition of the proteasome. These phenotypic changes were associated with increased stimulatory capacity in mixed lymphocyte culture (MLC) and enhanced sensitivity to natural killer (NK) cells. However, BL cells remained unable to present endogenously expressed viral antigens to EBV specific CTLs.

The results highlight two possible routes of immune escape: a) local inhibition of T cell responses that could potentially recognise and destroy tumor cells expressing antigenic proteins; b) defective antigen presentation which would render the malignant cells invisible to CTLs specific for tumor antigens.