Because of their ability to elicit cell death in a variety of tumor cells but not in normal cells, recombinant proteins of the TNFalpha-related-apoptosis-inducing ligand (TRAIL) appear to be promising medicine for cancer patients. However, a recent finding of apoptosis induction by recombinant TRAIL protein in primary normal human hepatocytes raised the alarm about the possibility of serious toxicity when the TRAIL proteins are administered systemically. Our preliminary study also showed that expression of the full-length TRAIL protein in primary human hepatocytes caused massive apoptosis in these cells. Consequently, localized expression of the gene in tumors may be a good way to reduce systemic toxic effects while providing a constant therapeutic effect at the cancer site. The goal of this proposal is to develop technology that can target pharmaceutical effects of the TRAIL gene to cancer cells. We hypothesize that the promoter for human telomerase reverse transcriptase (hTERT), active in 85 percent of primary human cancers, will be able to elicit tumor-specific expression of the TRAIL gene and thus can be used to avoid toxicity in susceptible normal cells. In a preliminary study, we found that in vitro transfer of the TRAIL gene elicited apoptosis and bystander effects in colon cancer DLD-1 cells but not in normal fibroblasts. Intratumoral delivery of the TRAIL gene effectively suppressed tumor growth of DLD-1 xenograft. We also demonstrated that the hTERT promoter could induce high levels of transgene expression in cancer cells but not in normal cells in vitro and in vivo. Our preliminary results showed that the hTERT promoter can be used to prevent GFP-TRAIL fusion gene expression in liver cells in vitro and in vivo. To achieve our goals, we will assess therapeutic value of the TRAIL gene expressed from the hTERT promoter. In vitro and in vivo studies are proposed to evaluate the efficiency of gene transfer, levels of transgene expression, apoptosis induction, the bystander effect and its possible mechanism, antitumor activity, anti-metastasis activity and possible toxicity of the GFP/TRAIL gene expressed from the hTERT promoter and delivered by adenoviral vector. Completion of the proposed studies will allow us to develop technologies for targeting the pharmaceutical effects of the TRAIL gene to cancers so that the treatment will have maximum therapeutic effects but minimum toxicity. The preclinical documentation we obtain on tumor-specific expression driven by the hTERT promoter and on the therapeutic/toxic profiles of hTERT-TRAIL constructs may provide a scientific basis for future clinical application of this approach in humans.
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