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In vitro expansion of tumor-specific, HLA-restricted human CD8+ cytolytic T lymphocytes.

The effect of cytokines with human recombinant interleukin-2 (rIL-2) on cytolytic T cell (CTL) generation was studied. Lymphocytes were isolated from involved lymph nodes of melanoma patients and expanded in medium containing rIL-2 alone or in combination with other human cytokines (rIL-1, rIL-4, rIL-6, and recombinant human tumor necrosis factor-alpha (rTNF alpha)). Lymphocytes incubated with rIL-2 alone did not grow, whereas addition of the other cytokines augmented IL-2-mediated lymphocyte proliferation. In all cultures, the majority of expanded lymphocytes were CD3+, CD56- T cells. Lymphocytes cultured with rIL-1, rIL-2, rIL-4, and rIL-6 exhibited cytolytic activity specific for autologous melanoma, which increased during the culture period (24.08 and 58.18% at 16 and 30 days in culture, respectively) without detectable changes in cell surface phenotype and remained high even after 100 days in culture. Moreover, the cytolytic activity was inhibited by monoclonal antibodies (mAbs) against HLA-class I, CD3, and CD8 molecules but not by mAbs against HLA-class II or CD4 molecules. Lymphokine-activated killer (LAK) activity was detected in lymphocytes cultured with rIL-1, rIL-2, and rIL-6 in the presence or absence of rTNF alpha. These data indicate that lymphocytes derived from melanoma-invaded lymph nodes and cultured in the presence of rIL-1, rIL-2, rIL-4, and rIL-6 offers an efficient system to expand CD8+ CTLs with HLA-restricted cytolytic specificity against autologous tumor cells.