Browsing by Subject "Glutamine"
Now showing 1 - 3 of 3
- Results Per Page
- Sort Options
Item Glutamine Antagonism and Its Utlity [sic] as a Therapeutic Modality in Cancer(2022-05) Rosales, Tracy Ibarra; Kim, James; DeBerardinis, Ralph J.; Whitehurst, Angelique Wright; Conacci-Sorrell, MaraliceGlutamine metabolism is important in cancer as it fuels the TCA cycle, plays a role in redox homeostasis, and contributes to the production of nucleotides, amino acids, and lipids for survival, making glutamine metabolism a promising target in cancer therapy. The work outlined in this dissertation focuses on understanding the mechanism of the broad glutamine metabolism inhibitor, 6-diazo-5-oxo-L-norleucine (DON) and its prodrug, JHU-083, while comparing them to the effects of CB-839, a specific glutaminase inhibitor. DON is one of the oldest and well-known glutamine antagonists and can effectively limit tumor growth in a preclinical setting. Unfortunately, DON was removed from early phase clinical trials due to unacceptable toxicity in the gastrointestinal tract (GI). Thus, DON prodrugs were recently developed to be inactive until cleaved by cathepsins enriched in the tumors or by plasma esterases, bypassing toxicity in the GI tract. Using isotope tracer studies in cancer cells and mouse xenograft models, I found that DON and JHU-083 mainly inhibit glutamine-derived nitrogen labeling in purines but unexpectedly does not limit the contribution of glutamine-derived carbon labeling of tricarboxylic acid (TCA) cycle metabolites. Additionally, I found that DON and JHU-083 can limit the levels of purines but not the levels of most TCA cycle metabolites. These findings suggest that these drugs are poor inhibitors of glutaminase in the cancer cell lines tested and that DON and JHU-083 mainly target purine metabolism. Recognizing DON and JHU-083 as effective purine metabolism inhibitors can offer insight into which cancer patients could benefit from these drugs. Relapsed small-cell lung cancer (SCLC) is characterized by an upregulation of de novo purine biosynthesis and have few durable therapies. Using metabolic tracing and untargeted metabolomics, I found that DON can inhibit purine metabolism in treatment-naïve and chemoresistant pairs of SCLC. In a mouse xenograft model of relapsed SCLC, JHU-083 induces a delay in tumor growth without overt side effects. My work provides an opportunity to explore JHU-083 as an anti-cancer therapy for diseases that depend on purine biosynthesis.Item Reductive Carboxylation Is a Novel Pathway of Glutamine Metabolism That Supports the Growth of Tumor Cells with Metabolic Defects(2013-10-23) Mullen, Andrew Robbins; Burgess, Shawn C.; Abrams, John M.; Pearson, Gray W.; DeBerardinis, Ralph J.In growing cancer cells, oxidative metabolism of glucose and glutamine in the mitochondria provide precursors needed for de novo synthesis of proteins, nucleic acids and lipids. Yet, a subset tumors harbor genetic mutations in the electron transport chain or tricarboxylic acid cycle that disable normal oxidative mitochondrial function. Importantly, it has been unknown how these cells generate the biosynthetic precursors required for growth. To address this, I used models of mitochondrial dysfunction in isogenic cancer cell lines and studied their metabolism using a combination of Gas Chromatography- Mass Spectrometry and Nuclear Magnetic Resonance spectroscopy. In all cases, mitochondrial dysfunction stimulated a novel pathway of glutamine metabolism, characterized by reversal of the canonical tricarboxylic acid cycle, termed reductive carboxylation; providing a plausible mechanism for how cancer cells with mitochondrial defects generate biosynthetic precursors required for growth. To gain mechanistic insight into how this unusual pathway was regulated I carried out a targeted metabolomics analysis in our isogenic tumor cell models. This led to the striking discovery that cells engaged in the reductive carboxylation pathway also operate an additional metabolic pathway that, at first glance, would appear to be superfluous and inefficient. Functional characterization of this second pathway revealed, however, that its activity was necessary for the optimal function of the reductive carboxylation pathway. In summary, this work has given us insights into how cancer cells are able to grow in the context of defective mitochondria. Additionally, this has exposed a potential Achilles ’ heel that might be used to selectively kill tumors which rely on this pathway for growth.Item Targeting Glutamine Metabolism in Kidney Development and Polycystic Kidney Disease(2018-07-30) Flowers, Ebony Michelle; Zhu, Hao; Abrams, John M.; Cobb, Melanie H.; DeBerardinis, Ralph J.; Carroll, Thomas J.Polycystic kidney disease is a hereditary disorder characterized by the progressive manifestation of numerous fluid-filled sacs, known as cysts, within the renal epithelia. The enlargement of the cysts causes the gradual replacement of normal kidney parenchyma which leads to impairment of renal function, and ultimately, renal failure. While the primary causes of PKD are genetic mutations in one of the polycystins that encode the PKD1 and PKD2 proteins, the age of onset and severity of PKD cases greatly varies, suggesting other genes/processes are involved. Lkb1 is a serine-threonine kinase involved in the regulation of several molecular processes including cellular polarity, autophagy, mTOR signaling, and energetic stress response, all of which are dysregulated in PKD. We generated mice lacking Lkb1 in developing kidney epithelia to establish which of these processes contributed to cyst formation and progression in the absence of Lkb1. Surprisingly, Lkb1 mutant mice showed no defects in renal tubule development or maintenance. However, later studies revealed the co-ablation Lkb1 along with Tsc1, a gene known to play a role in human PKD, within the developing renal epithelia prompted a drastic acceleration in the timing, number, and size of cyst formation. We utilized in vitro cell culture coupled with ex vivo culture of embryonic kidneys with defined media allowed us to determine which metabolic pathways were affected by the deficit of Lkb1. Our results revealed that Lkb1 mutant cells and embryonic kidneys require glutamine for growth while wild-type cells and kidneys did not. Subsequent studies demonstrated that Pkd1 embryonic kidneys phenocopied Lkb1 mutant kidneys in respect to their reliance on glutamine for growth. Further investigation into defining which metabolic enzymes/pathways are regulated in normal kidney development and how the absence of Lkb1 or Pkd1 alters these metabolic processes will allow us to gain a greater understanding of the role of metabolism in PKD and potentially lead to t the development of therapeutics to reduce cyst number and/or size.