Functional Genomics of Metastatic Determinants of Melanoma

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DESCRIPTION (provided by applicant): Metastasis is a complex, multi-step process and while much has been investigated about the genetic and genomic alterations driving primary tumor formation, little has been elucidated about the genetic basis for metastasis. Evidence shows that genomic profiles of metastatic tumors are distinct from primary tumors implying that novel, distinct genes drive metastasis. While genomic data provides insight into alterations that are critical in the etiology of tumors, true genetic determinants must be identified through functional in vitro and in vivo studies. To this end, a cross-species oncogenomic screen has identified a stringently filtered list of genes up-regulated in metastatic melanoma. This research proposes to employ these candidates in a low-complexity in vitro screen for resistance to anoikis, or apoptosis due to loss of surface attachment. Additionally, an in vivo metastasis screen will be conducted to identify genes driving various steps of metastasis including intravasation, survival while in circulation, and secondary tumor formation. Finally, all pro-anoikis resistance and pro-metastatic genes will be enlisted into in-depth functional and clinicopathological studies. Using low-adherent plates, a high-throughput 96-well format in vitro anoikis screen to identify genes that confer anoikis resistance to anoikis sensitive cells will be conducted. Since anoikis resistance is but one aspect of metastasis, this research will also screen in vivo for the combination of genes capable of driving multiple steps of metastasis. Human cells expressing candidate genes will be sub-cutaneously injected into mice and mice will be monitored for primary and secondary tumor formation and survival of tumor cells in circulation. Candidates conferring metastatic potential to non-metastatic cells will be enlisted into in-depth biochemical studies through gain-of-function and loss-of-function assays. In parallel, using immunohistochemistry, metastatic determinants will be examined for expression on staged human metastatic melanoma tissues to establish clinical relevancy. PUBLIC HEALTH RELEVANCE: Unlike many cancers, the incidence of melanoma has steadily increased world-wide. Whereas primary localized melanoma is surgically excisable, therapies for metastatic melanoma have poor clinical efficacy, resulting in only 10% of patients having greater than five year survival. Therefore, it is of utmost importance to identify the genetic determinants of metastatic melanoma and thereby generate a list of genes that can serve not only as therapeutic targets but also as prognostic biomarkers with high translational potential.