Investigating Mechanisms of Cancer Redox Homeostasis with Metabolic CRISPR-Based Screens

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Project Summary and Abstract The broad objectives of this NRSA Individual Fellowship are two-fold: 1) to facilitate the development of essential skills that will allow the candidate to become an effective physician-scientist, and 2) to investigate mechanisms of cancer redox homeostasis uncovered by CRISPR-based screens. The candidate and his mentor have designed a specific training plan to achieve these objectives. The plan includes a rigorous research component that lays the foundation for a successful career. This proposal concerns lung adenocarcinoma and regulation of oxidative stress. Lung cancer remains the leading cause of cancer death among both men and women. The recent success of metabolic drugs, such as complex I inhibitors, have revitalized interest in targeting cancer metabolism. However, many of these drugs have unacceptable side effects. A more thorough understanding of cancer metabolic reprogramming is necessary for the development of effective and safer, metabolically targeted cancer therapies. Recent studies have uncovered that the dietary consumption of cruciferous vegetables, such as broccoli, is associated with a decreased risk of cancer. Their chemoprotective properties are attributed to isothiocyanates such as phenethyl isothiocyanate (PEITC). Although it known that PEITC disrupts the delicate cancer redox balance, the mechanism by which this occurs is unknown. The candidate has preliminary data utilizing a metabolism focused CRISPR-based screen revealing KDM5C as required for PEITC-induced lung adenocarcinoma cell death and SCD1 as synthetic lethal with PEITC. The focus of this proposal is to validate these gene hits and examine the mechanisms by which KDM5C and SCD1 regulate the cellular antioxidant response. The long-term goal of this proposal is to identify therapeutic metabolic targets that may lead to more effective and safer therapies for patients with lung cancer. In Specific Aim 1, the candidate will determine whether KDM5C is required for PEITC-induced lung adenocarcinoma cell death by repressing antioxidant genes. Analysis of KDM5C regulation of histone modifications and their downstream effects on gene expression will be used to determine the extent to which KDM5C is a negative regulator of cellular antioxidant response. In Specific Aim 2, the candidate will determine the molecular mechanism by which inhibition of SCD1 is synthetic lethal with PEITC to induce lung adenocarcinoma cell death. Lipidomics will quantify the extent to which SCD1 regulates cellular lipid levels. Measurements of ER and mitochondrial stress will provide insight into the mechanism by which SCD1 is synthetic lethal with PEITC. The proposed studies will identify and causally validate previously unknown mechanisms of cellular redox homeostasis in cancer and provide evidence to devise effective combinatorial therapies to target this balance.