Date of Award

Spring 2021

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Pharmacology

First Advisor

Turk, Benjamin

Abstract

Melanoma, the deadliest form of skin cancer, is characterized by aberrant hyperactivation of the ERK mitogen-activated protein kinase signaling pathway. Genetic lesions in the core components of the RAS-RAF-MEK-ERK protein kinase cascade as well as its upstream regulators are key features of melanoma progression and drug resistance. MEK, the central kinase within the cascade, is constitutively activated by many upstream oncogenic events and is an important drug target. MEK inhibition in combination with BRAF inhibition is the standard of care for treating BRAFV600E melanoma. However, not all BRAFV600E melanomas respond to these inhibitors, and those that do respond eventually acquire resistance. To better understand mechanisms of MEK inhibitor susceptibility and MEK regulation in BRAFV600E melanoma, I performed a loss-of-function screen to identify kinases and phosphatases that modulate sensitivity to two clinical MEK1/2 inhibitors. In this screen, I identified PPP6C, the catalytic subunit of protein phosphatase 6 (PP6), as a factor promoting sensitivity to MEK inhibition. I established PPP6C as a major MEK phosphatase in cells exhibiting oncogenic ERK pathway activation. Recruitment of MEK to PPP6C occurs through an interaction with its associated regulatory subunits. Loss of PPP6C causes hyperphosphorylation of MEK at both activating and crosstalk phosphorylation sites, promoting signaling through the ERK pathway and decreasing sensitivity to the growth inhibitory effects of MEK inhibitors. Consistent with its role in regulating ERK signaling, PPP6C is frequently mutated in melanoma, as is MEK1. I found that recurrent melanoma-associated PPP6C mutations cause MEK hyperphosphorylation and ERK signaling hyperactivation when expressed in cells. Recurrent MEK1 mutations all promote MEK1 kinase activity but are activated by different mechanisms of action. The elevated MEK activity associated with PPP6C mutations or MEK1 mutations suggests that they promote disease by a common mechanism: activating the core oncogenic pathway driving melanoma. Collectively, our studies identify novel modulators of susceptibility to ERK pathway targeted cancer therapies, including PPP6C, a key negative regulator of ERK signaling, and cancer-associated mutations that influence ERK signaling activation.

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