951. Antitumor Activity of AAV-2 Vector Encoding Modified TRAIL Gene

Abstract

Prostate cancer is the second most prevalent cancer in men, with limited effective treatment options, especially for metastatic disease. We propose to address this problem by developing enhanced Fas-mediated apoptosis as a treatment modality for prostate cancer. It has been shown that Fas expression on some prostate tumor lines, including the murine RM-1 line, can be up-regulated by a variety of treatment regimens. In this study, primary T cells and functionally-defined subsets of these T cells will be engineered to over-express Fas and will be introduced into tumor-bearing mice in order to interact with normal or up-regulated Fas receptor on prostate tumors. To do this, three recombinant onco-retroviruses have been produced which engineer over-expression of FasL. The first carries only the FasL gene, while the second contains cDNA for a modified, non-cleavable form of FasL (ncFasL) with potentially reduced liver toxicity. The third, a bicistronic construct, includes the cDNA for c-FLIP, an inhibitor of the Fas-mediated apoptotic pathway, in addition to ncFasL. c-FLIP is included to prevent self-killing of transduced, FasL-expressing T cells. The constructs have been packaged in ecotropic E86 and amphotropic FLYRD18 cells and shown to direct production of FasL as well as produce infectious virions, as measured by staining with anti-FasL antibody and flow cytometric analysis. Assays have confirmed that the FasL and ncFasL produced by infected cells is able to mediate killing of Fas-expressing Jurkat cells, and is thus functional. Transduction of primary murine C57BL/6 pan-T cells has been optimized. Irradiation of several tumor cell lines, both murine and human, has shown this to be a plausible method to increase Fas expression. The ability of interferons and chemotherapeutic drugs, including mitoxantrone, to increase tumor Fas expression will also be examined. Experiments are underway in tumor-bearing mice to assess the efficiency of killing by these modified T cells, and to compare the effectiveness of the three viral constructs. We expect to see effective activation of death pathways, leading to tumor destruction. Finally, the relative abilities of the type 1 versus type 2 T lymphocytes as delivery vehicles for this treatment will be compared. This approach may provide a non-invasive treatment for prostate cancer and may also be extended to other malignancies such as breast cancer.