Author ORCID Identifier
0000-0002-5200-7488
Date of Graduation
5-2018
Document Type
Dissertation (PhD)
Program Affiliation
Genes and Development
Degree Name
Doctor of Philosophy (PhD)
Advisor/Committee Chair
Sendurai A. Mani
Committee Member
Richard Behringer
Committee Member
Mikhail Kolonin
Committee Member
Suzanne Fuqua
Committee Member
Arun Sreekumar
Abstract
Metastasis is currently incurable. Recent studies suggest that metabolic reprogramming significantly impacts metastatic progression. Understanding the underlying biology of how metabolic reprogramming influences metastasis will shed light on developing novel ways to treat metastasis and is of urgent clinical need. A potent mediator of metastatic potential in breast cancer is the winged helix/Forkhead domain transcription factor FOXC2. We have previously shown that FOXC2 is necessary and sufficient to promote metastasis through induction of the epithelial to mesenchymal transition (EMT). EMT is a latent embryonic program that is aberrantly induced in breast cancer cells conferring migratory and invasive capabilities in addition to cancer stem cell (CSC) properties. In normal adipose tissue, FOXC2 regulates adipocyte metabolism by promoting the expression of genes associated with energy turnover, insulin sensitivity, differentiation, and mitochondrial biogenesis. I hypothesized that similar to its regulatory role of adipocyte metabolism, FOXC2 dictates metabolic functions during EMT.
Here we report that distinct metabolic alterations occur with the onset of EMT. We identified an EMT metabolite signature (EMS) composed of four elevated metabolites (glutamine, glutamate, beta-alanine, and glycyleucine) and found that the EMS is predictive of poor clinical outcome in breast cancer patients. In efforts to characterize the functional relevance of the metabolic alterations defined by the EMS, we examined genes associated with metabolism that change with the onset of EMT. From this analysis, we identified genes associated with purine nucleotide and glutamine metabolism that became downregulated with the onset of EMT. We measured the expression of these genes in EMT-induced cells with FOXC2 knockdown. We observed that depletion of FOXC2 resulted in upregulation of these purine and glutamine metabolism genes and a restoration of their associated metabolism. Moreover, we found that the induction of EMT results in reduced mitochondrial metabolism and glutamine independence. This metabolic program is maintained by the downregulation of Glutaminase 2 (GLS2) expression, an enzyme that catalyzes the conversion of glutamine to glutamate. We also observed that GLS2 expression is FOXC2 dependent. Inhibition of FOXC2 signaling by treatment of cells with a small moleculae inhibitor of its upstream regulator p38MAPK with a chemical inhibitor enhanced GLS2 mRNA expression and mitochondrial metabolism. Moreover, we demonstrate that GLS2 over-expression alone increases mitochondrial metabolism, reduces the glycolytic metabolic program, and CSC mammosphere formation. Collectively these data are in line with an analysis of patient data that demonstrated that higher GLS2 expression is associated with better over-all survival. Lastly, we tested a novel therapeutic strategy that combined a p38MAPK inhibitor, which inhibits the FOXC2 signaling pathway, with a standard-of-care chemotherapeutic to reduce the tumor bulk. We observed both a decrease in lung metastasis and tumor size in mice treated with the combination strategy compared to mice receiving monotherapy.
Keywords
Epithelial to Mesenchymal Transition, Breast Cancer, Glutamine Metabolism, Cancer Stem Cells, Metabolic Flexibility, FOXC2, Metastasis