PuSH - Publication Server of Helmholtz Zentrum München: B cells immortalized by a mini-Epstein-Barr virus encoding a foreign antigen efficiently reactive specific cytotoxic T cells.

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Moosmann, A.* ; Khan, N.* ; Cobbold, M.* ; Zentz, C. ; Delecluse, H.-J.* ; Hollweck, G. ; Hislop, A.D.* ; Blake, N.W.* ; Croom-Carter, D.* ; Wollenberg, B. ; Moss, P.A.H.* ; Zeidler, R.* ; Rickinson, A.B.* ; Hammerschmidt, W.

B cells immortalized by a mini-Epstein-Barr virus encoding a foreign antigen efficiently reactive specific cytotoxic T cells.

Blood 100, 1755-1764 (2002)

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Lymphoblastoid cell lines (LCLs) are human B cells latently infected and immortalized by Epstein-Barr virus (EBV). Presenting viral antigens, they efficiently induce EBV-specific T-cell responses in vitro. Analogous ways to generate T-cell cultures specific for other antigens of interest are highly desirable. Previously, we constructed a mini-EBV plasmid that consists of less than half the EBV genome, is unable to cause virus production, but still immortalizes B cells in vitro. Mini-EBV–immortalized B-cell lines (mini-LCLs) are efficiently produced by infection of B cells with viruslike particles carrying only mini-EBV DNA. Mini-EBV plasmids can be engineered to express an additional gene in immortalized B cells. Here we present a mini-EBV coding for a potent CD8⁺ T-cell antigen, the matrix phosphoprotein pp65 of human cytomegalovirus (CMV). By means of this pp65 mini-EBV, pp65-expressing mini-LCLs could be readily established from healthy donors in a one-step procedure. We used these pp65 mini-LCLs to reactivate and expand effector T cells from autologous peripheral blood cells in vitro. When generated from cytomegalovirus (CMV)–seropositive donors, these effector T-cell cultures displayed strong pp65-specific HLA-restricted cytotoxicity. A large fraction of CD8⁺ T cells with pp65 epitope specificity was present in such cultures, as demonstrated by direct staining with HLA/peptide tetramers. We conclude that the pp65 mini-EBV is an attractive tool for CMV-specific adoptive immunotherapy. Mini-EBVs could also facilitate the generation of T cells specific for various other antigens of interest.

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