Melanoma is a deadly variety of skin cancer accounting for three-fourths of skin cancer-related deaths, making it critical to investigate the pathogenesis involved in the advanced stages of melanoma, and design effective therapeutics for its treatment. Drugs targeting epigenetic modifiers such as EZH2 are becoming increasingly popular in cancer treatment. However, the effect of EZH2 inhibitors have not been investigated in melanoma. The goal of this work was to investigate the role of EZH2 in the pathogenesis involved in the advanced stages of melanoma, to enable the design of effective therapeutic strategies for the treatment of melanoma. In this study, the relationship between EZH2 and tumor suppressors E-cadherin and RUNX3 in melanoma has been investigated using matched melanoma cell lines (WM115 and WM266-4), formalin-fixed-paraffin-embedded (FFPE) patient tissue samples and an in vivo xenograft model. The studies performed in cell lines and FFPE tissue samples suggested EZH2 mediated silencing of E-cadherin and RUNX3 by promoting H3K27-trimethylation of the E-CADHERIN and RUNX3 promoters in WM266-4 cells (metastatic; high EZH2; low E-cadherin) relative to WM115 cells (primary; low EZH2; high E-cadherin). Furthermore, it was observed that EZH2 inhibition restored E-cadherin and RUNX3 expression in metastatic melanoma cells by reducing EZH2 and H3K27me3 occupancy at E-CADHERIN and RUNX3 promoters respectively. In addition, EZH2 inhibition induced apoptosis, inhibited cellular proliferation, and adversely affected invasion of metastatic melanoma cells. In vivo, treatment of WM266-4 xenografts with EZH2 inhibitor resulted in significant reduction of tumor growth. In combination with the BRAF inhibitor Dabrafenib, this effect was highly synergistic. Analysis of harvested tumors indicated increased E-cadherin and RUNX3 levels in EZH2 inhibited group compared to vehicle, due to reduced H3K27me3 enrichment by EZH2 at E-CADHERIN and RUNX3 promoters, validating our in vitro finding. Tumor regression by combination therapy was also accompanied by a significant increase in pro-apoptotic proteins Bak and Bax and decreases in anti-apoptotic proteins Bcl-xL and Bcl-2. In the later part of the dissertation, epigenetic mechanisms operating in another type of skin cancer - Merkel cell carcinoma has been investigated. Overall, the results of this dissertation provide strong experimental support for the use of EZH2 inhibitors as promising melanoma drugs as monotherapy, or in conjunction with other chemotherapeutics in combination therapies.
|Advisor:||Tackett, Alan J.|
|Commitee:||Baldini, Giulia, Chambers, Timothy, Davidson, Mari K., Gao, Ling|
|School:||University of Arkansas for Medical Sciences|
|Department:||Biochemistry and Molecular Biology|
|School Location:||United States -- Arkansas|
|Source:||DAI-B 78/12(E), Dissertation Abstracts International|
|Keywords:||BRAF, EPZ-6438, EZH2, Melanoma, Merkel cell carcinoma, Proteomics|
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