Tumor necrosis factor mutants with selective cytotoxic activity

Abstract

Tumor necrosis factor (TNF-alpha) has a cytotoxic or cytostatic effect when tested with various malignant cell lines. Clinical trials in cancer patients, however, revealed high systemic toxicity of TNF-alpha. The existence of two types of receptor may partially explain the pleiotropic activity of TNF-alpha. The purpose of this study was to characterize the relative cytotoxic activity of TNF-alpha and TNF mutants on the mouse fibrosarcoma L929 cells in a standard cytotoxicity test, on human larynx carcinoma HEp-2 cells, and on human monoblastoid leukemic cells U937. TNF mutants were obtained by site-directed mutagenesis. The purity of TNF-alpha was established by capillary electrophoresis. TNF-alpha and TNF mutants were analysed by Western blot analysis using monoclonal antibodies against TNF-alpha. The results show that TNF mutants can recognize the different TNF-receptors (TNF-R) selectivity. It is generally believed that activation of TNF-R75 is responsible for the systemic toxicity of TNF-alpha. Hence, the development of TNF mutants, binding selectively to TNF-R55, could lead to new option for an anticancer treatment that would be devoid of the deleterious effect of TNF-alpha.