Cancer cells from a variety of tumor types rely on the metabolic process of glycolysis even in the presence of oxygen. The cancer cell needs glucose for the production of energy and the biosynthesis of nucleic acids, amino acids and lipids needed to sustain high levels of cellular proliferation. The rate of aerobic glycolysis in cancer cells is elevated to a much higher degree compared to normal tissue, such that whole-body imaging of tumors is routinely done in the clinic using the radioactive glucose analogue, 2-deoxy-2-(18F)fluoro-D-glucose and positron emission tomography (FDG-PET). A major consequence of this sustained elevation of glycolysis is the generation of toxic by-products such as the highly reactive alpha-oxoaldehyde, methylglyoxal (MG). MG reacts with amino acids, nucleic acids and lipids to generate advanced glycation end-products (AGEs). The predominant DNA AGE formed is the mutagenic DNA adduct, CEdG. To combat the accumulation of MG, cells deploy the highly-conserved, two enzyme glyoxalase system that detoxifies MG in a glutathione-dependent fashion to form D-lactate. This system is important to prevent CEdG and other AGE formation in healthy cells, but in highly glycolytic cancer cells, it may represent an Achille?s heel that can be exploited for treatment. In cancer cells with elevated Glyoxalase 1 (Glo1) activity, Glo1 inhibition using glutathione analogues leads to programmed cell death. In breast cancer patients, Glo1 is over-expressed in the tumor relative to pair-matched, adjacent tissue. In prostate cancer patients, high levels of Glo1 predict poor prognosis. Thus, Glo1 has high potential to be a useful therapeutic target in these and other glycolytic tumors. We hypothesize that Glo1 inhibition will lead to intracellular accumulation of MG, which will lead to the formation of the mutagenic CEdG and cell death. The proposed research will thus interrogate the molecular intersection of metabolism and DNA repair.