Biochemical Background in Mitochondria Affects 2HG Production by IDH2 and ADHFE1 in Breast Carcinoma.

Jitka Špačková,Katarína Smolková,Aleš Dvořák,Petra Tesařová,Kateřina Pospíšilová,David Větvička,Alberto Leguina-Ruzzi,Petr Ježek,Klára Gotvaldová,Alexandra Urbančoková,Libor Vítek

doi:10.3390/cancers13071709

Abstract

Mitochondrial production of 2-hydroxyglutarate (2HG) can be catalyzed by wild-type isocitrate dehydrogenase 2 (IDH2) and alcohol dehydrogenase, iron-containing 1 (ADHFE1). We investigated whether biochemical background and substrate concentration in breast cancer cells promote 2HG production. To estimate its role in 2HG production, we quantified 2HG levels and its enantiomers in breast cancer cells using analytical approaches for metabolomics. By manipulation of mitochondrial substrate fluxes using genetic and pharmacological approaches, we demonstrated the existence of active competition between 2HG producing enzymes, i.e., IDH2 and ADHFE1. Moreover, we showed that distinct fractions of IDH2 enzyme molecules operate in distinct oxido-reductive modes, providing NADPH and producing 2HG simultaneously. We have also detected 2HG release in the urine of breast cancer patients undergoing adjuvant therapy and detected a correlation with stages of breast carcinoma development. In summary, we provide a background for vital mitochondrial production of 2HG in breast cancer cells with outcomes towards cancer biology and possible future diagnosis of breast carcinoma.

Highlights

Metabolic reprogramming is a general feature of cancer phenotype
We evaluated 2HG levels in several breast carcinoma cultured cell lines, derived from primary and secondary tumors and we demonstrated that 2HG is actively formed by isocitrate dehydrogenase 2 (IDH2) and ADHFE1 and that there is a competition between these two enzymes, which regulate substrate flux in mitochondria
1- C-glutamine (IDH2-related flux) and we have demonstrated that 2HG production in According to our previous work [8], 13C-labeling of 2HG has been observed when we used

Summary

Introduction

Metabolic reprogramming is a general feature of cancer phenotype. Substantial attention concerning research on mitochondrial isocitrate dehydrogenase 2 (IDH2) and cytosolic IDH1 in cancer cells has been devoted to mutant forms of IDHs, representing oncogenes, producing 2-hydroxyglutarate (2HG) [2]. We have demonstrated that 2HG can be formed by mitochondrial wild-type IDH2 despite the absence of any IDH2/1 mutations in breast cancer cells [8,9,10]. R-2HG is a product or bi-product of several metabolic enzymes, including mitochondrial ones. We investigated whether production of mitochondrial 2HG is elevated in breast cancer cell lines and identified active competition for initial substrate, 2OG, between enzymes isocitrate dehydrogenase IDH2 and alcohol dehydrogenase ADHFE1. We have investigated possible substrate and cofactor NADPH channeling between the two IDH2 molecules within mitochondria

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