Abstract 5482: L-2HG/ L2HGDH axis as therapeutic target for kidney cancer

Abstract

Background: The D-enantiomer of 2-hydroxygultarate (D-2HG), along with fumarate and succinate, are considered oncometabolites that promote tumorigenesis. Our lab has reported elevated levels of the L-enantiomer of 2-hydroxyglutare (L-2HG) in clear cell renal cell carcinoma (ccRCC), in part due to reduced expression of the enzyme L-2HG dehydrogenase (L2HGDH). Objectives: Here we evaluated the underlying biochemical mechanisms of L-2HG accumulation and characterized the contribution of the L-2HG/L2HGDH axis to tumorigenesis. Experimental strategy: We assessed the role of raised L-2HG in renal carcinogenesis via both in vitro and in vivo means. Mutagenesis and knock down approaches were applied to study involvement of loss of L2HGDH in accumulation of high L-2HG levels and associated tumor phenotypes. Metabolomics coupled with 13C tracer labeling studies were utilized to dissect the biochemical axis that promotes L-2HG accumulation in RCC cells. Results: Translational based studies demonstrate that loss of L2HGDH expression is associated with both cancer progression and worsened outcomes. In connection with our previous findings, we demonstrate that raising cellular L-2HG levels by treatment with cell permeable octyl ester of L-2HG as well as by shRNA-mediated knock down of L2HGDH in renal epithelial cells (HK-2) promotes in vitro tumor phenotypes. Concurrently, the epigenetic mark 5hmc was significantly decreased under high L-2HG levels. Further investigation with restoration of L2HGDH in RCC cells (RXF393 and A498) shows decreased L-2HG levels and suppression of in vivo tumor growth in nude mice (NU/NU). Interestingly, expression of loss-of-function mutant of L2HGDH was unable to decrease L-2HG levels and failed to suppress in vitro and in vivo tumor phenotypes. In addition, high L-2HG levels were found to upregulate epithelial-mesenchymal-transition (EMT) marker SNAIL1 and correspondingly downregulate E-cadherin. Biochemical studies demonstrate that the predominant carbon source for L-2HG in RCC is glutamine through the activity of glutaminase and malate dehydrogenase (MDH). Pharmacological inhibition of the glutamine/MDH axis by treatment of glutaminase inhibitor (CB-839) and MDH inhibitor (4k) in RCC cells reduced L-2HG levels and mitigates in vitro tumor phenotypes. Furthermore, suppression of in vitro phenotypes by CB-839 and shRNA-mediated MDH2 knockdown was rescued by concurrent treatment with octyl ester of L-2HG. Finally, restoration of L2HGDH promoted the expression of genes targeted by the polycomb repressor complex 2 (PRC2), whereas inhibition of PRC2 in high L-2HG cells suppressed tumor phenotypes. Conclusion: Collectively, our data demonstrate the biologic relevance of high L-2HG to renal carcinogenesis and reveal novel therapeutic opportunities for L-2HG driven kidney tumors. Citation Format: Sandeep Balu Shelar, Eun-hee Shim, Garrett Brinkley, Anirban Kundu, Hyeyoung Nam, Francesca Carobbio, Tyler Poston, Jubilee Tan, Daniel Benson, Dinesh Rakheja, Richard Kirkman, Yusuke Sato, Seishi Ogawa, Shilpa Dutta, Sadanandan E. Velu, David Crossman, Anja Becker, Conrad Kunick, Sunil Sudarshan. L-2HG/ L2HGDH axis as therapeutic target for kidney cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 5482.

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“…2-HG is considered as an oncometabolite and strongly contributes to tumorigenesis in many cancers (Parsons et al, 2008) (Mardis et al, 2009) (Shelar et al, 2018). Mutated forms of isocitrate dehydrogenase 1/2 (IDHmt1/2) that catalyze the conversion from α-ketoglutarate to the D enantiomer of 2-HG (D-2-HG) were famously identified as potential targets with the development of the first-in-class metabolic inhibitors (Parsons et al, 2008) (Dang et al, 2009).…”