Robey, R.C.; (2010) Characterising the adaptive t-cell immune response against Kaposi’s sarcoma-associated herpesvirus. Doctoral thesis , UCL (University College London).

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Abstract

Kaposi’s sarcoma-associated herpesvirus (KSHV) is causally related to Kaposi’s sarcoma (KS), the most common malignancy in individuals with untreated HIV/AIDS. Several lines of evidence indicate that KS oncogenesis is associated with loss of T cell-mediated control of KSHV-infected cells. However, the adaptive CD8 and CD4 T-cell responses against KSHV have not been fully characterised. Neither the antigenic repertoire nor the immunodominant targets of CD8 and CD4 KSHV-specific T cells are fully understood, and the phenotypes and functions of these cells remain largely unexplored. To investigate the targets of the CD8 and CD4 T-cell responses against KSHV, a novel approach for a large-scale screen of KSHV antigens was proposed that used lentiviral-transduced monocyte-derived dendritic cells (moDCs) expressing a panel of KSHV open reading frames (ORFs). Transduced moDCs naturally process the KSHV gene products and present the resulting antigenic peptides in the context of MHC class I and II. Transduced moDCs were cultured with autologous T cells and the CD8 and CD4 proliferative responses to each KSHV ORF (or pool of ORFs) were assessed. CD8 and CD4 KSHV-specific responses were investigated in 14 KSHV-seropositive individuals. Unexpectedly, both the CD8 and CD4 T-cell responses against KSHV were found to be skewed towards ORFs expressed in the early and late phases of the viral lytic cycle. The most frequently recognised CD8 target was a pool of late lytic KSHV ORFs, [ORF28/ORF36/ORF37]. Identification of novel KSHV CD8 epitopes from within the late lytic ORF pool was attempted. Peptide-MHC binding and denaturation assays identified peptides that had the highest affinity for HLA-A*0201. Recognition of these potential epitopes was tested in clinical samples by IFNγ ELISpot, and compared with recognition of nine previously published HLA-A*0201-restricted KSHV epitopes. Finally, the use of pentamers as tools to investigate the memory phenotypes and functions of virus-specific T cells was explored.

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