Strategies to Overcome Drug Resistance in Melanoma
Open Access
- Author:
- Chung, Chin-ying
- Graduate Program:
- Molecular Toxicology
- Degree:
- Doctor of Philosophy
- Document Type:
- Dissertation
- Date of Defense:
- June 24, 2013
- Committee Members:
- Gavin Peter Robertson, Dissertation Advisor/Co-Advisor
Gavin Peter Robertson, Committee Chair/Co-Chair
Gary Alan Clawson, Committee Member
Rogerio I Neves, Special Member
Jiyue Zhu, Committee Member - Keywords:
- Melanoma
B-Raf(V600E)
Vemurafenib
Epigenetics
Cell invasion
Integrin - Abstract:
- B-Raf is the most mutated gene in melanoma. The most common V600E mutation constitutively activates the MAP kinase-signaling cascade. The recently FDA approved selective V600EB-Raf inhibitor, Vemurafenib, gave great promise for patients with metastatic disease and exhibited manageable toxicity; however, most of the patients eventually developed more aggressive resistant tumors. The focus of this dissertation is to identify molecular mechanisms leading to resistance for B-Raf targeting agents, such as Vemurafenib and B-Raf siRNA, in order to develop therapeutic strategies to overcome drug resistance in melanoma. We found that agents targeting V600EB-Raf decrease expression levels of the tetraspanin CD82 (a tumor metastasis suppressor) leading to more aggressive cell invasion for resistance development. Here, B-Raf targeting agents are shown to increase expression of DNA methyltransferases leading to CD82 promoter methylation, decreasing CD82 expression, and promoting integrin expression for cell attachment on extracellular matrix (ECM) under flow conditions. Decreased CD82 increases the invasive migratory potential of melanoma cells through vessel walls to enhance lung metastases development. CD82 promoter methylation could be reversed using the demethylating agent 5-aza-2'-deoxycytidine (Decitabine), suggesting that the combination of agents might overcome melanoma spread in patients. Deregulated PI3 and MAP kinase pathways promote early melanocytic lesion development and confer drug resistance. Few treatment options are available to target these deregulated pathways to prevent cutaneous non-invasive melanocytic cells or invasive melanomas from developing into more aggressive, widely-disseminated metastatic disease. In this dissertation study, we examined the ability of a selenium containing compound called PBISe to moderate these two major signaling pathways to prevent cutaneous melanocytic lesion or metastatic melanoma development. Topical PBISe treatment inhibited melanocytic lesion development in laboratory-generated skin by 70-80% and in animal skin by ~50% through targeting both the Akt3 and MAPK pathways. Thus, PBISe treatment has the potential to treat non-invasive melanocytic lesions and invasive metastatic melanoma development in skin. The Vemurafenib resistance mechanisms are complex. We also found that the Vemurafenib-mediated ERK1/2 negative feedback loop inhibits tumor suppressor Sprouty4 and increases Akt3 activity, ultimately leading to the development of drug-resistant melanomas. In this study, combining Vemurafenib with sigma receptor-targeted LPD (Liposomal-Polycation-DNA) nanoparticles containing Akt3 siRNA and suppressor Sprouty4 plasmid simultaneously inhibits the MAPK and Akt3 signaling cascades. The combination-mediated inhibition prevents the development of resistant tumors in xenografted mice. Further, the novel melanoma cell-targeting nanoparticle LPD can also carry potential siRNA to silence genes that lead to drug resistance. Therefore, this nanoparticle based agent has important potential for development as an effective agent for combating recurrent resistant disease.