Abstract 2571: 1,25-dihydroxyvitamin D regulation of pyruvate carboxylase in MCF10A-ras cells decreases viability in extracellular matrix detached conditions

Abstract

Abstract Metastasis is the main cause of most breast cancer related deaths. Throughout the process of metastasis, cancer cells strive to survive in conditions where they are detached from the extracellular matrix (ECM). Vitamin D may play a role in preventing breast cancer metastasis by modulating metabolism in breast cancer cells detached from ECM. The anaplerotic enzyme pyruvate carboxylase (PC) is essential for breast to lung metastasis and the active metabolite of vitamin D, 1,25-dihydroxyvitamin D3 (1,25D), downregulates PC expression through a vitamin D response element. PC replenishes the TCA cycle with the intermediate oxaloacetate (OAA), and lower expression of PC following 1,25D treatment may cause impaired energy metabolism via the TCA cycle. We hypothesize that the 1,25D mediated decrease in PC expression decreases breast cancer cell viability in ECM detached conditions. The Harvey-ras transformed MCF10A human breast epithelial cell line (MCF10A-ras) treated with vehicle (ethanol) or 10 nM 1,25D and maintained in varying nutrient concentrations were utilized for these studies. Poly-HEMA coated plates were used to model ECM detachment paired with MTT assays to measure cell viability. mRNA expression of metabolic genes is quantified using qPCR. Results show that when compared to attached cells, detached cells have a 94% increase in PC mRNA expression, while 1,25D treatment decreases PC mRNA expression in both attached by 75% and detached conditions by 63%. In addition 1,25D treatment and PC knockdown decreases viability in ECM detached conditions significantly by 16% and 11%, with no additional effect when combined. Glutamine deprivation also decreases viability by 40% in detached conditions, however, the combination of 1,25D treatment and glutamine deprivation results in a 71% decrease in viability. These results suggest that while glutamine is required for ECM survival, regulation of glutamine metabolism is not the mechanism by which 1,25D affects ECM survival. Replenishing the TCA cycle with OAA rescues glutamine depleted cells by 84% as well as PC knockdown cells by 14%. Further, when stressed with low glutamine conditions, cells in attached conditions increase mRNA expression of PC by 311%, suggesting that changes in PC expression may be a mechanism that cancer cells use to adapt to energy related stress. These results suggest that 1,25D regulation of metabolism may sensitize breast cancer cells to ECM detachment death via downregulation of PC, reducing the required flux of substrates into the TCA cycle which are needed to survive ECM detached conditions during metastasis. Citation Format: Madeline Sheeley, Dorothy Teegarden, Katie Wong. 1,25-dihydroxyvitamin D regulation of pyruvate carboxylase in MCF10A-ras cells decreases viability in extracellular matrix detached conditions [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2571.