Discovery of selective inhibitors of Glutaminase-2, which inhibit mTORC1, activate autophagy and inhibit proliferation in cancer cells

Abstract

Glutaminase, which converts glutamine to glutamate, is involved in Warburg effect in cancer cells. Two human glutaminase genes have been identified, GLS (GLS1) and GLS2. Two alternative transcripts arise from each glutaminase gene: first, the kidney isoform (KGA) and glutaminase C (GAC) for GLS; and, second, the liver isoform (LGA) and glutaminase B (GAB) for GLS2. While GLS1 is considered as a cancer therapeutic target, the potential role of GLS2 in cancer remains unclear. Here, we discovered a series of alkyl benzoquinones that preferentially inhibit glutaminase B isoform (GAB, GLS2) rather than the kidney isoform of glutaminase (KGA, GLS1). We identified amino acid residues in an allosteric binding pocket responsible for the selectivity. Treatment with the alkyl benzoquinones decreased intracellular glutaminase activity and glutamate levels. GLS2 inhibition by either alkyl benzoquinones or GLS2 siRNA reduced carcinoma cell proliferation and anchorage-independent colony formation, and induced autophagy via AMPK mediated mTORC1 inhibition. Our findings demonstrate amino acid sequences for selective inhibition of glutaminase isozymes and validate GLS2 as a potential anti-cancer target.

A Tale of Two Glutaminases: Homologous Enzymes with Distinct Roles in Tumorigenesis

“…While this report is at odds with previous literature and the predicted localization of either LGA isoform, no study has yet been published which either disputes or supports these findings. The N-terminal residues of the longer LGA isoform are truncated following mitochondrial localization, but it is currently unclear if a similar tr uncation occurs for the shorter LGA variant [51,58].…”

“…The same group later demonstrated that transfection of the same cells with LGA increases their susceptibility to alkylating agents, by downregulating the DNA repair gene MGMT [116]. However, Lee et al showed that LGA inhibition or genetic silencing inhibited the growth of both A549 lung cancer cells and HepG2 hepatoma cells [58].…”

Glutamine Addiction in Kidney Cancer Suppresses Oxidative Stress and Can Be Exploited for Real-Time Imaging

“…There exist two glutaminases, GLS (kidney type KGA and GAC isoforms) and GLS2 (liver isoform (LGA) and glutaminase B (GAB)), yet GLS is the isoform upregulated in cancer, as well as the main glutaminase within the kidney(25,26). Since GSH synthesis requires glutamate in addition to glycine and cysteine, we tested the hypothesis that blocking glutamate production from glutamine by GLS inhibition will down-regulate this important antioxidant pathway, resulting in higher ROS levels, which will be selectively toxic to cancer cells due to their increased local ROS levels.…”

Divergent in vitro/in vivo responses to drug treatments of highly aggressive NIH-Ras cancer cells: a PET imaging and metabolomics-mass-spectrometry study

“…Specifically, the metabolic rewiring shown by cancer cells involves not only the Warburg effect with enhanced glycolysis and high glutamine consumption, but also elevated rates of lipid biosynthesis, maintenance of redox homeostasis and limited levels of autophagy in the first steps of oncogenesis [10–12]. Recent evidence has shown that cancer cells can engage glutamine metabolism enzymes, such as PEP-carboxykinase (PCK2) [13] or pyruvate carboxylase (PC) [14], to develop metabolic adaptation and promote enhanced cell growth when even glucose is limited. In addition, they can activate autophagy when glutamine is inhibited [15].…”

“…Recent evidence has shown that cancer cells can engage glutamine metabolism enzymes, such as PEP-carboxykinase (PCK2) [13] or pyruvate carboxylase (PC) [14], to develop metabolic adaptation and promote enhanced cell growth when even glucose is limited. In addition, they can activate autophagy when glutamine is inhibited [15]. Furthermore, metabolites, such as ATP, acetyl-CoA (AcCoA), reactive oxygen species (ROS) and enzymes with key role in metabolic regulation may act on different signaling pathways controlling tumor growth [9, 16–19].…”