Angiogenic and Metastatic Determinants of Malignant Melanoma

Cutaneous melanoma or malignant melanoma of the skin is a highly metastatic disease, with an increasing rate of incidence, poor prognosis and high resistance to therapeutic intervention. Although early diagnosis and surgical resection of the primary lesion could significantly improve survival, the high propensity of melanomas to disseminate through intradermal, haematogenous and lymphatic routes to regional and visceral sites leads to poor prognosis and high mortality rates. While the clinical staging and progression of melanoma is well defined, the molecular etiology of the disease is less well characterized. The principal goal of this thesis was to obtain novel mechanistic insights into melanoma biology and identify molecular determinants of this disease.

We identified the capacity of melanoma-derived small molecules to promote long-term survival of endothelial cells under severe hypoxic conditions. This observation excludes known proangiogenic molecules which have been the focus of anti-angiogenic therapies and sets the stage for the identification of novel regulators with therapeutic potential. Moreover, TIMP3 was identified as a dominant negative regulator of melanoma development. The inhibitory role of TIMP3 in melanoma angiogenesis was validated and further extended to clinical samples from melanoma patients. We showed that promoter methylation mediated TIMP3 silencing impacts clinical outcome in melanoma and also evaluated the implications of the tumor suppressor role of TIMP3 in melanoma metastasis, using matched samples from melanoma patients. A decrease in TIMP3 expression with observed with disease progression and further TIMP3 inhibited melanoma cell migration and invasion. To characterize core mediators of the metastatic cascade of melanomas, we determined the migratory profile of melanoma cell lines and performed correlation analysis to identify potential genetic modulators. WNT5A was identified as a dominant regulator of the metastatic cascade in melanoma and further we show that WNT5A inhibition decreases the migratory and metastatic potential of melanoma cells.

Additionally, this thesis describes novel tools to quantitatively characterize several biological processes. The ring barrier-based migration assay enables the quantitative assessment of several parameters of cell migration. The eNOS-Tag-GFP mouse model provides a platform for the in vivo and ex vivo study of early angiogenic events in physiological and pathological conditions. Collectively, the results presented in this thesis identify crucial pathophysiological determinants of melanoma. These insights and tools may further guide the discovery of novel regulators of melanoma biology and result in the implementation of new treatment rationales for therapeutic benefit.