Abstract 747: Glutamine, glutaminase and γ-glutamyl-transferase activities are essential for lung tumorigenesis

Increased glutamine uptake has been found to drive cancer cell proliferation, making tumor cells glutamine-dependent or even addicted. Glutamine is believed to provide additional carbon and nitrogen sources for cell growth. The first step in glutamine utilization is its conversion to glutamate by glutaminase (GLS). Glutamate is a precursor for glutathione synthesis, a pathway known to be activated in tumor cells. Additionally, glutathione concentrations are known to be elevated in non-small cell lung cancer (NSCLC) and is believed to protect against oxidative stress. In contrast, glutathione9s participation in the γ-glutamyl-cycle is less well studied. Here we investigated the hypothesis that glutamine drives glutathione synthesis and export, drives the γ-glutamyl-cycle, and thereby contributes to NSCLC tumor cell proliferation. Glutamine consumption and glutathione excretion were measured in H460, A549 and MRC-5 cell lines. Treatment with BPTES, a known GLS inhibitor, led to reduced cell proliferation. This was also demonstrated when γ-glutamyl-transferase (GGT) was inhibited with acivicin. GGT transfers glutamyl groups from glutathione to amino acids to facilitate the amino acid transport into the cell. These results suggest active utilization of the γ-glutamyl cycle in the tumor cells. The formation of metabolites downstream of GLS and GGT were essentially abolished at the lowest inhibitor concentrations studied. In contrast, biological endpoints, such as MTT assay response, required up to 30-fold higher inhibitor concentrations. Culturing in stable isotope-labeled glutamine demonstrated that >50% of excreted glutathione and >80% of γ-glutamyl amino acids were derived from glutamine. Together these results demonstrate that glutamine is actively utilized for synthesis and excretion of glutathione which, in turn, is utilized to promote amino acid uptake by the formation of γ-glutamyl amino acids. These findings add yet another important function to the glutamine dependence of tumor cells. In addition to providing TCA cycle intermediates, we show that glutamine promotes glutathione synthesis to increase cellular defense mechanisms, and drives the γ-glutamyl cycle to enhance amino acid uptake. Citation Format: Daniel R. Sappington, Eric R. Siegel, Rosalind B. Penney, Gunnar Boysen. Glutamine, glutaminase and γ-glutamyl-transferase activities are essential for lung tumorigenesis. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 747. doi:10.1158/1538-7445.AM2015-747