Investigating Metabolic Adaptations of Cancer Cells to Nutrient Stress

Metabolic reprogramming is a classic hallmark of cancer. In addition to increased glucose uptake and aerobic glycolysis, many cancer cells exhibit extreme dependency on glutamine in order to maintain anaplerotic, anabolic, and antioxidant needs to support rapid proliferation and growth. In particular, cancers exhibiting amplification of the oncogene Myc are extremely glutamine dependent and undergo apoptosis when glutamine is removed in culture, suggesting glutamine is an essential metabolite for survival and growth. How these cells adapt to glutamine limitation and maintain cell proliferation is poorly understood. Furthermore, cancer therapies targeting glutamine metabolism are currently undergoing clinical trials, making it imperative to investigate how treatment with these drugs will select for cells that are resistant to glutamine limitation. For my thesis, I utilized an unbiased screening strategy in order to identify Myc-amplified clones that resist glutamine deprivation. The top validated hit in this screen was Aldh18a1, a gene that encodes the mitochondrial protein pyrroline-5-carboxylate synthase (P5CS), which catalyzes the rate-limiting step in proline biosynthesis. In this reaction, intracellular glutamate is reduced to the metabolic intermediate P5C, which is subsequently catalyzed to proline. Importantly, the role this gene plays in cancer remains unknown. I hypothesized that by negatively regulating P5CS, cells compensate for the lack of exogenous glutamine by accumulating intracellular glutamate, which can then be used for a variety of metabolic processes. Indeed, through use of genetic knockdown strategies, LC-MS metabolomic profiling and 15NH4 labeling, and nutrient challenges/supplementations, I found that these cells were able to repurpose carbon and nitrogen towards alpha-ketoglutarate, asparagine, and nucleotides; all of which support cell survival and proliferation in the absence of glutamine. Furthermore, Aldh18a1 deficient cells become exquisitely sensitive to glutamine synthetase inhibition when glutamine is limiting. Lastly, I found that certain human cancer cells downregulate P5CS in response to glutamine deprivation, and knockdown of this enzyme offered a growth advantage under glutamine restricted conditions. My findings reveal a novel adaptive pathway that cancer cells acquire to survive conditions of nutrient stress, providing insight that could inform new metabolically-driven therapies that would reduce morbidity and recurrence in patients suffering from highly aggressive cancers.
Medical Sciences