Bone-induced c-kit in Prostate Cancer Cells: Implications for Bone Metastasis

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DESCRIPTION (provided by applicant): Bone metastasis is the main cause of morbidity and mortality in prostate cancer (PCa) patients. Currently, PCa patients with bone metastatic disease respond poorly to chemotherapy. In cancer, deregulation of the receptor tyrosine kinase c-kit is a frequent event. However, its precise role in PCa and, more specifically in PCa bone metastasis, remains largely undetermined. In studies using clinical samples, we have shown that c-kit expression by prostate cells is gradually increased during malignant progression, with the highest expression seen in PCa bone metastases. We found that c-kit is neither expressed by PCa cell lines in vitro, nor by subcutaneous tumors formed by them. However, when these same cells are either co-cultured with bone marrow-derived cells or injected intraosseously using different experimental models, we found a de novo expression of c-kit in all the cases. Importantly, a correlation between c-kit expression and chemoresistance has been established in tumor systems other than prostate, and our preliminary data indicate that this might be also true for PCa, suggesting a role for c-kit in the poor response of patients with PCa bone metastasis to chemotherapy. Taken together, we hypothesize that de novo expression of c-kit induced by bone-derived factors is a prerequisite for the expansion of PCa cells within the bone, and is responsible for the acquisition of chemotherapy-resistance. To test our hypothesis, the specific aims are: (1) Determine the functional significance of bone-induced c-kit expression in PCa cells in intraosseous tumor growth and bone response, and the effect of de novo c-kit expression by PCa cells on the acquisition of chemoresistance, and (2) Identify causative bone-derived factors involved in the induction of c-kit in PCa cells. To provide causal evidence for the role of c-kit in intraosseous PCa growth, lentiviral-mediated RNA interference or transfect ion with a plasmid containing a dominant-negative (kinase-dead) c-kit mutant construct will be used to impede de novo c-kit induction in PCa cells within the bone microenvironment. The contribution of PCa-associated c-kit to intraosseous expansion of PCa cells will be studied using the SCID-hu model of bone metastasis developed in our laboratory, as well as its effect on osteoblast and osteoclast differentiations. The effect of de novo c-kit expression by PCa cells and its consequence to response to docetaxel will be studied in vivo and in vitro with c-kit-inducible and non-inducible c-kit PCa cells, and molecular mechanisms involved will be investigated. Using laser capture microdisection we will extract RNA from experimental PCa bone tumors and then use a) functional cloning from a retroviral cDNA expression library and b) differential expression analysis by gene microarray, to identify bone-derived factors inducing c-kit expression in PCa cells (Aim 2). It is expected that this high-risk, high-gain proposal will culminate in the design of new tailored therapies aimed at filling the gap in the treatment of PCa patients with bone metastasis.