Induction of Melanoma Antigen-Specific Cytotoxic T Lymphocytes In Vitro by Stimulation with B7-Expressing Human Melanoma Cell Lines

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Crosslinking of CD28 receptors on resting T lymphocytes by B7 cost-imulatory molecules expressed by antigen-presenting cells (APCs) plays a critical role in T-cell activation. Human melanomas express major histocompatibility complex (MHC)-restricted tumor-associated antigens that can be recognized by cytotoxic T lymphocytes (CTL), yet they remain poorly immunogenic. One mechanism for the failure of T-cell response is the lack of expression of costimulatory molecules by human melanoma cells. We have transfected the B7–1 gene into three HLA-A2-expressing human melanoma cell lines, and studied their capacity to stimulate primary human T cells. B7-expressing melanoma cells were excellent inducers of T-cell proliferation, cytokine production, and cytolytic activity in allogeneic mixed lymphocyte cultures through a process dependent on the function of the T-cell receptor as well as interactions between B7:CD28, CD2:LFA-3, and LFA-1:ICAM-1. Subset analysis demonstrated that CD4* T cells or addition of exogenous interleukin-2 was required for the induction of CD8+ CTL. Untransfected parental melanoma cells were inert as APCs in these cultures. Rotating stimulation of T cells with the three B7-expressing cell lines led to the generation of T-cell lines that were cytolytic for HLA-A2 melanoma cells and other HLA-A2+ targets that were pulsed with HLA-A2-restricted MART-1 peptides: These data demonstrate that expression of B7–1 by human mela-noma cells converts them into effective APCs for the in vitro induction of MHC-restricted, melanoma-specific CTL.

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