291. Comparison of Anti-Tumor Effects between Transmembrane and Secretory Tumor Necrosis Factor-alpha In Vitro and In Vivo

Abstract

Tumor necrosis factor-alpha (TNF-a) exists naturally in two forms, transmembrane and secretory form. In the present study, three recombinant retroviral vectors containing human wild type TNF-a (Wt-TNF), secretory TNF-a (S-TNF, a mutant replaced its signal peptide by that of IL-2), and transmembrane TNF-a (TM-TNF, a mutant deleted a site for cleavage of TNF into the secretory form) were constructed respectively and transfected stably into murine hepatic carcinoma cell line (H22). TNF-a either secreted in supernatants by S-TNF and Wt-TNF producing tumor cells or expressed on cell surface of TM-TNF and Wt-TNF synthesizing tumor cells was demonstrated to be cytotoxic against TNF sensitive L929 cells. Also, the H22 cells transfected with three types of TNF-a can kill parental H22 in effect/target (E/T) dependent manner in vitro and their maximal killing rates were about 38|[sim]|43% at E/T ratio of 5:1. Injection of total 2.5|[prime]|105 cells mixed with transfected and parental H22 tumor cells at different ratios into syngeneic mice resulted in the inhibition of tumor growth and their maximal inhibition rates were about 57|[sim]|72% at E/T 5:1. A transient weight loss was found in mice bearing S-TNF producing tumors, whereas no obvious side effects were seen in mice bearing TM-TNF or Wt-TNF expressing tumors. It has been shown that S-TNF secreting tumor promoted significantly lymphocytes infiltration and attracted more CD4+ T cells than CD8+ T cells to the tumor site, while TM-TNF expressing tumor stimulated Fas expression and inhibited a tumor metastasis associated molecule CD44v3 expression on tumor cells. These results suggest that S-TNF and TM-TNF kill cancer cells in vivo through different mechanisms and that transmembrane form of TNF-a might be a safer candidate for tumor gene therapy due to its less side effects and retainable antitumor activity.