SIRT3 Suppresses Rewiring of Glutamine Metabolism in Cancer

Abstract

In the process of transformation, a cancer cell selects for a strategy to enhance its survival, growth, and proliferation. Metabolic rewiring is a hallmark of cancer cells and is at the heart of such a mechanism. Although oncogenes and tumor suppressors contribute to metabolic reprogramming in cancer, the exact mechanisms underlying deregulated nutrient metabolism continue to be elucidated. The mitochondrial sirtuin, SIRT3 has tumor suppressive function in various types of human cancer and represses the Warburg effect in breast cancer cells. Despite what is known about SIRT3 in the regulation of metabolism, the challenge in translating our findings into development of therapies for treatment of cancer remains. Thus, the goal of this dissertation was to develop a better understanding of the role of SIRT3 in tumorigenesis and to identify metabolic vulnerabilities associated with loss of SIRT3 in cancer.

In this dissertation, I identify deregulated glutamine metabolism as a vulnerability in tumor-like cells with SIRT3 deletion. First, I utilized immortalized WT and SIRT3 null (KO) cells and performed a small molecule screen to identify compounds that selectively decreased proliferation of cells lacking SIRT3. The top hit from the screen was a glutamine analog, azaserine. Like glucose, glutamine is crucial for synthesis of building blocks to support tumorigenesis. In combination with metabolomics analysis, RNA sequencing (RNA Seq), and bioinformatics analysis, I revealed that azaserine significantly represses a dependency on elevated de novo nucleotide metabolism, driven in part by increased mTORC1 signaling, to suppress growth of cells with decreased SIRT3 levels. Additionally, in human breast tumors, SIRT3 expression is lower in the more aggressive basal-like tumors compared to luminal breast tumors, and is associated with increased mTORC1 signaling. Moreover, I demonstrate that SIRT3 represses mTORC1 signaling to suppress tumorigenesis.

Given that azaserine is a glutamine analog and that SIRT3 loss results in enhanced glutamine consumption, I further examined the effect of SIRT3 loss on glutamine metabolism. Using a combination of glutamine tracing and RNA Seq, I uncover alterations in various nodes of glutamine metabolism in the absence of SIRT3. More specifically, I identify increased distribution of glutamine to glutathione and the tricarboxylic acid (TCA) cycle, which is coupled with production of nonessential amino acids (NEAAs) in SIRT3 KO cells. Lastly, I find that SIRT3 represses glutaminolysis in breast cancer cells.

Taken together, my findings reveal a pathway through which glutamine contributes to tumor growth and identifies previously undescribed nodes of tumor metabolism deregulated in the absence of SIRT3. My findings shed light on potential metabolic pathways that may be targeted in cancers with decreased SIRT3 expression to suppress tumorigenesis.