Author ORCID Identifier

https://orcid.org/0000-0002-7567-1891

Date of Graduation

12-2021

Document Type

Dissertation (PhD)

Program Affiliation

Genetics and Epigenetics

Degree Name

Doctor of Philosophy (PhD)

Advisor/Committee Chair

Boyi Gan

Committee Member

Junjie Chen

Committee Member

Pierre McCrea

Committee Member

Bingliang Fang

Committee Member

Haoqiang Ying

Abstract

The metabolic vulnerability of cancers has long been envisaged as an attractive window to develop novel therapeutic strategies. Metabolic flexibility at the cellular level encompasses the efficient rerouting of anabolic and catabolic pathways in response to varying environmental stimuli to maintain cellular homeostasis and sustain proliferation. The primary objective of this study is to identify metabolic vulnerabilities bestowed by KEAP1/NRF2 signaling axis through SLC7A11. SLC7A11 is a transcriptional target of NRF2, an essential regulator of cellular anti-oxidant response. Under unstressed basal conditions, NRF2 interacts with KEAP1, a tumor suppressor gene and a substrate adaptor protein of the Cullin3-dependent ubiquitin ligase complex, which targets it for polyubiquitination and proteasomal degradation. KEAP1-NRF2 pathway is altered in upto 34% of lung squamous cell carcinoma (SqCC), with mutations/amplifications of NRF2 in 19% cases; mutations/deletions of KEAP1 in 12% cases of SqCC. KEAP1 is mutated in up to 20% of lung adenocarcinoma. However, currently, there is no effective therapy to target KEAP1/NRF2-mutant lung cancer. The interesting role of SLC7A11 underlies its double-edged role in regulating redox balance and nutrient dependency through the exchange of two critical cellular metabolites. The cystine imported by SLC7A11 is quickly reduced to cysteine, the rate-limiting precursor in the synthesis of the anti-oxidant glutathione (GSH). Thus, SLC7A11 plays an essential role in maintaining intracellular GSH levels to protect cells from oxidative stress. In light of the established pro-survival function of SLC7A11, several surprising recent findings reveal a pro-death function of SLC7A11 upon glucose starvation. This project aims to determine the roles of the KEAP1-NRF2 signaling axis in regulating glucose dependency in lung cancer.

In this study, I provide evidence that, contrasting to its pro-survival roles, high expression of SLC7A11 renders cancer cells more dependent on glucose for survival. I further demonstrate that this SLC7A11 regulated metabolic dependency is mediated through its transcription factors NRF2 and ATF. I extend the observations of NRF2/SLC7A11 mediated glucose dependency to NSCLC cells carrying mutated KEAP1. I show that KEAP1 mutation/deficiency leads to constitutive activation of NRF2 and aberrant expression of NRF2 target cystine/glutamate antiporter SLC7A11. I further show a toxic accumulation of disulfide molecules and NADPH depletion by high cystine uptake in KEAP1- deficient lung cancer cells. Lastly, I also show that KEAP1 expression status drives sensitivity to glucose transporter inhibitor as tested in cell lines and mouse xenograft studies. The Pro-cell death function of KEAP1-NRF2-SLC7A11 signaling in the context of glucose starvation revealed in this work is unexpected and unique, representing a significant shift from the current understanding of KEAP1-NRF2-SLC7A11 function in stress response and proposes a novel therapy development to target such tumors.

Keywords

Lung Cancer, Keap1, Cancer Metabolism, Nutrient Dependency, Metabolic Vulnerability, disulfide stress

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