Abstract 5402: A new anticancer strategy based on inhibiting mitochondrial proline dehydrogenase (PRODH) and exploiting synthetic lethal interactions with p53 restoration and/or glutaminase (GLS1) inhibition
Authors: Scott Gary K. , Rutter Justine , Frazier Katya , Rothschild Daniel , Yau Christina , Benz Christopher 
Published: August 2015

From prokaryotes to the highest eukaryotes, proline is catabolized by a unique and structurally conserved flavoprotein, proline dehydrogenase (PRODH). In eukaryotes PRODH associates with the inner mitochondrial membrane and catalyzes the first and rate limiting catabolic step, transferring two electrons to the electron transport chain where they can produce ATP and/or reactive oxygen species (ROS). Following recognition that PRODH is one of the most strongly upregulated genes by the tumor suppressing protein, p53, its capacity to generate reactive oxygen species (ROS) and induce apoptosis initially qualified it as a tumor suppressing response, mediating the effects of p53 upregulation). However, we have definitively shown that while PRODH can induce mitochondrial ROS production it does so almost exclusively at other mitochondrial sites (e.g. complex I) via its anaplerotic glutamate production and not directly by itself. Furthermore, our recent studies indicate that PRODH critically supports breast cancer cell growth and survival by consuming proline for anaplerotic glutamate production, bypassing glutaminase (GLS1) to fuel oxidative phosphorylation and sustain ATP levels. We now show that PRODH knockdown, as well as its enzymatic inhibition by proline competitive inhibitors like L-tetrahydro-2-furoic acid (L-THFA) or (S)-5-oxo-2-tetrahydrofurancarboxylic acid (5-oxo), induce breast cancer cell apoptosis (cleaved PARP)and reduce viable cell growth within 48 h. Given that PRODH can provide an alternate source of mitochondrial glutamate for glutamine addicted and GLS1-dependent cancer cells, we evaluated the expression microarray profiles of 51 different human breast cancer lines and found a significant breast cancer subtype association with PRODH expression (luminal and HER2+ &gt; basal-like), and also a strong inverse correlation between PRODH and GLS1 expression, suggesting that one or the other of these mitochondrial pathways is needed to feed breast cancer's anaplerotic addiction to glutamate. Consistent with our hypothesis of synthetic lethal interactions between PRODH, p53wt upregulation and glutamine addiction, we combined PRODH knockdown or enzymatic inhibition with either a p53wt restoring drug (e.g. MI-63, nutlin-3a) or a clinical glutaminase (GLS1) inhibitor (CB-839, Calithera) and observed synergistic induction of apoptosis and growth inhibition against malignant (e.g. MCF7, ZR-75-1, DU4475) but not normal (MCF10A) breast epithelial cells. Using an in vitro mitochondrial PRODH bioassay and a computationally based structural model of human PRODH's catalytic site, we are now designing new competitive and mechanism-based irreversible PRODH inhibitors capable of exploiting these synthetic lethal interactions to develop more effective and less toxic cancer therapy.

Citation Format: Gary K. Scott, Justine Rutter, Katya Frazier, Daniel Rothschild, Christina Yau, Christopher Benz. A new anticancer strategy based on inhibiting mitochondrial proline dehydrogenase (PRODH) and exploiting synthetic lethal interactions with p53 restoration and/or glutaminase (GLS1) inhibition. [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 5402. doi:10.1158/1538-7445.AM2015-5402