Metabolic Stress in Non-Metastatic and Metastatic Murine Mammary Cancer Cells

Breast cancer is a major public health concern, with one in eight women in the United States expected to be diagnosed throughout the course of her lifetime. Metastasis of cancer to secondary sites in the body is the primary cause of death among breast cancer patients, highlighting the critical need to understand mechanisms that contribute to metastatic progression. Throughout metastatic progression, cancer cells are exposed to cell stresses, including metabolic, oxidative, and hypoxic cell stress, which cells must overcome in order to survive and progress. In the present studies, we determined the effects of metabolic cell stresses in non-metastatic M-Wnt and metastatic metM-Wnt^lung murine mammary cancer cell lines. Culturing both cell lines in high (4 mM) compared to low (2 mM) glutamine conditions suppressed viability of metM-Wnt^lung cells. M-Wnt cells had no change in viability in response to glutamine concentration, and high glutamine concentrations decreased mRNA levels of genes involved in glutamine catabolism in M-Wnt cells only. In accordance with the differences in glutamine metabolism, metM-Wnt^lung cell demonstrated an increase in glutamine flux into the TCA cycle in high glutamine, whereas M-Wnt cells had no change in glutamine flux in response to glutamine concentration. metM-Wnt^lung cells were significantly more sensitive to treatment with ammonium, a byproduct of glutamine catabolism, suggesting that a high rate of metabolism and ammonium production may decrease cell viability in high glutamine conditions. These data suggest that glutamine utilization and metabolism change in cancer cells at different stages of metastatic progression. In addition to metabolic stress from variable nutrient availability, changes in oxygen availability are a source of metabolic stress for cancer cells. Hypoxia, or low oxygen tension, is associated with metastasis and reduced survival, making it an important biological process to study in the context of cancer. Culturing non-metastatic and metastatic cells in hypoxia increased mRNA levels of genes related to antioxidant defense only in metM-Wnt^lung cells. Hypoxia also induced expression of the integrated stress response effector protein activating transcription factor 4 (ATF4) and its target gene glutamic pyruvic transaminase (Gpt2) in metM-Wnt^lung cells. Furthermore, genetic depletion of ATF4 reduced survival of hypoxic metM-Wnt^lung cells in detached conditions. These results suggest that cancer cells accumulate cell stress throughout the course of progression and must adapt their gene expression for continued survival throughout metastatic processes. The results of these two studies highlight metabolic adaptations and vulnerabilities of cancer cells at different stages of progression. These data will contribute to improving our understanding of therapeutic targets to prevent or delay metastasis in cancer patients, thereby reducing cancer mortality.