1,25-Dihydroxyvitamin D inhibits glutamine metabolism in Harvey-ras transformed MCF10A human breast epithelial cell.

Abstract Breast cancer is the second most common cancer among women in the US. The active form of vitamin D, 1,25-dihydroxyvitamin D (1,25(OH) 2 D), is proposed to inhibit cellular processes and to prevent breast cancer. The current studies investigated the effect of 1,25(OH) 2 D on glutamine metabolism during cancer progression employing Harvey -ras oncogene transformed MCF10A human breast epithelial cells (MCF10A -ras ). Treatment with 1,25(OH) 2 D significantly reduced intracellular glutamine and glutamate levels measured by nuclear magnetic resonance (NMR) by 23 ± 2% each. Further, 1,25(OH) 2 D treatment reduced glutamine and glutamate flux, determined by [U- 13 C 5 ] glutamine tracer kinetics, into the TCA cycle by 31 ± 0.2% and 17 ± 0.4%, respectively. The relative levels of mRNA and protein abundance of the major glutamine transporter, solute linked carrier family 1 member A5 (SLC1A5), was significantly decreased by 1,25(OH) 2 D treatment in both MCF10A- ras cells and MCF10A which overexpress ErbB2 ( HER-2/neu ). Consistent with these results, glutamine uptake was reduced by 1,25(OH) 2 D treatment and the impact was eliminated with the SLC1A5 inhibitor L-γ-Glutamyl- p -nitroanilide (GPNA). A consensus sequence to the vitamin D responsive element (VDRE) was identified in silico in the SLC1A5 gene promoter, and site-directed mutagenesis analyses with reporter gene studies demonstrate a functional negative VDRE in the promoter of the SLC1A5 gene. siRNA-SLC1A5 transfection in MCF10A- ras cells significantly reduced SLC1A5 mRNA expression as well as decreased viable cell number similar to 1,25(OH) 2 D treatment. SLC1A5 knockdown also induced an increase in apoptotic cells in MCF10A -ras cells. These results suggest 1,25(OH) 2 D alters glutamine metabolism in MCF10A- ras cells by inhibiting glutamine uptake and utilization, in part through down-regulation of SLC1A5 transcript abundance. Thus, 1,25(OH) 2 D down-regulation of the glutamine transporter, SLC1A5, may facilitate vitamin D prevention of breast cancer.