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Tracing Compartmentalized Metabolism in Mutant IDH Cancer Cells

  • Author(s): Parker, Seth Jameson
  • Advisor(s): Metallo, Christian M
  • et al.
Abstract

Cellular metabolism comprises of a network of reactions responsible for converting nutrients into energy and building blocks necessary for cells to carry out specific functions and to proliferate. In many diseases, including cancer, metabolism is dysfunctional and distinct from normal, non-diseased cells largely due to the presence of mutations impacting the regulation of cellular signaling and metabolic pathways. The chapters of this dissertation are independent bodies of work that explore how specific mutations can influence the flow of nutrients through the metabolic network. Chapter 1, titled "Metabolic Consequences of Oncogenic IDH Mutations", is a review on a set of mutations in a specific enzyme involved in metabolism observed in several different types of human cancers and how the this mutation affects the flow of major nutrients through the metabolic network. The chapter also discusses possible therapeutic targets present in metabolism and current efforts to treat cancers that harbor these mutations. Chapter 2, titled "IDH1 Mutations Alter Citric Acid Cycle Metabolism and Increase Dependence on Oxidative Mitochondrial Metabolism", explores how mutations in the cytosolic enzyme isocitrate dehydrogenase 1 (IDH1) affects central carbon metabolism in response to the low oxygen tensions commonly experienced by tumors. Cancer cells harboring mutations in IDH1 exhibited a greater reliance on oxidative mitochondrial metabolism under low oxygen tensions and were significantly more sensitive to mitochondrial inhibitors, such as the biguanide phenformin. Chapter 3, titled "Tracing Compartmentalized NADPH Metabolism in the Cysotol and Mitochondria of Mammalian Cells", describes the development of a tracing technique utilizing deuterium-labeled substrates and mass spectrometry to understand how NADPH and NADH, two cofactors critical for a variety of cellular processes, are metabolized in intact cells. In addition, Chapter 3 discusses the development of a reporter system to detect metabolism of NADPH in distinct organelles, such as the cytosol and mitochondria, in order to elucidate the activity of NADPH-dependent isozymes that exist in both compartments. Lastly, Chapter 3 explores the directionality of folate-mediated one carbon metabolism, a pathway critical for synthesizing nucleotides and supplying NAD(P)H, utilizing the reporter system. Chapter 4, titled "Hypoxic Reprogramming of Redox Metabolism in Mutant IDH2 Cells", applies the deuterium tracing technique discussed in Chapter 3 to understand how redox metabolism, specifically NADH and NADPH metabolism, is reprogrammed in response to hypoxia. Furthermore, the reporter system is applied to understand how NADPH pools in the mitochondria are affected by mutations in the mitochondrial enzyme IDH2 in cancer. Chapter 4 highlights an interesting, often overlooked, aspect of metabolism---the compartmentalization of metabolic pathways and how mutations affecting enzymes localized to distinct compartments reprogram metabolism differently.

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