Molecular Targeting of 15-Lipoxygenase-1 in Colon Cancer

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DESCRIPTION (provided by applicant): Apoptosis is being targeted to develop better therapies for colorectal cancer. We have found that (a) linoleic acid's product through 15-1ipoxygenase-1 (15-LOX-1), 13-S-hydroxyoctadecadienoic acid (13-S-HODE), restores apoptosis in colorectal cancer cells, (b) 13-S-HODE and 15-LOX-1 are downregulated in human colorectal cancer, and (c) NSAIDs restore 15-LOX-1 expression in human colorectal cancer cells to induce apoptosis and inhibit tumorigenesis. Human telomerase reverse transcriptase (hTERT) promoter can selectively express genes in cancer cells. The proposed research tests the following hypothesis: Selective restoration of 15-LOX-1 expression under the control of hTERT promoter in colorectal cancer cells via an adenoviral delivery system will reestablish apoptosis and inhibit tumorigenesis in colorectal cancer cells. Specific Aims: Aim 1: To determine whether hTERT-driven 15-LOX-1 expression via an adenoviral vector reestablishes apoptosis through the expression of 15-LOX-1 in colorectal cancer cells, we will transfect colorectal cancer cell lines with an adenoviral vector (Ad-15-LOX-1) that expresses 15-LOX-1 under the control of the hTERT promoter and will measure 15-LOX-1 expression, 13-S-HODE levels, and apoptosis rates in vitro. Aim 2: To assess whether 15-LOX-1 expression driven by hTERT promoter through an adenoviral transfection system suppresses colorectal tumorigenesis in vivo, we will transfect Ad-15-LOX-1 by intratumoral injections into subcutaneous xenografts of human colorectal cancer cells in nude mice and assess the effects of 15-LOX-1 expression on tumorigenesis and apoptosis rates. Aim 3: To determine whether the systemic administration of hTERT-15-LOX-1 adenovirus selectively expresses 15-LOX-1 in tumor cells suppressing tumor growth, we will administer Ad-15-LOX-1 intravenously to nude mice carrying human colon cancer cell liver metastases and evaluate the 15-LOX-1 expression effects on tumor growth. Aim 4: To determine whether 15-LOX-1 downregulates Bcl-2 to induce apoptosis in colorectal cancer cells, we will examine (a) the effects of transfecting Ad-15-LOX-1 into colorectal cancer cells on Bcl-2 expression and (b) if overexpressing Bcl-2 inhibits 15-LOX-1 induced apoptosis. Confirming the feasibility of tumor selective targeting of 15-LOX-1 to inhibit tumorigenesis will pave the way for future preclinical and clinical development of therapeutic strategies based on molecularly targeting 15-LOX-1