Molecular Mechanisms of Isocitrate Dehydrogenase 1 Mutants in Driving the Oncogenic Neomorphic Reaction

Abstract

Brain tumors result in a mortality rate > 60% within 5 years of diagnosis. Mutations in cytosolic isocitrate dehydrogenase 1 (IDH1) drive over 70% of low grade gliomas and secondary glioblastomas. IDH1 catalyzes the NADP+‐dependent formation of α‐ketoglutarate (α‐KG) from isocitrate via oxidative decarboxylation. Mutation of IDH1 results in a neomorphic activity, the formation of the oncometabolite, D‐2‐hydroxyglutarate (D2HG) from α‐KG in the presence of NADPH, via reduction. The most common mutations occur at residue 132, where arginine (R132) is typically replaced by histidine (R132H). We have shown previously that replacing arginine with other amino acids resulted in a trend in catalytic efficiency, with larger, more polar residues at the 132‐position promoting the normal reaction, and smaller, more hydrophobic residues driving the neomorphic reaction. Based on previous data with R132H and R132L, we hypothesize that R132Y and R132I, having similar structures respectively, will promote similar rates of catalytic efficiency in the neomorphic reaction, and R132P will have little to no catalytic activity due to likely disruption of local secondary structure. To test this hypothesis, we generated cDNA constructs and heterologously expressed and purified R132I, R132P, and R132Y‐IDH1 from E. coli to determine how these residues affect to the normal or neomorphic reaction. Kinetic assays were performed to determine the rates of the neomorphic reaction of the IDH1 mutant in the presence of NADPH and αKG. We found that R132Y‐IDH1 was the most efficient in catalyzing the neomorphic reaction followed by R132I, and R132P‐IDH1, having the lowest catalytic activity for the neomorphic reaction. By replacing the R132 residue with a wide range of amino acids, and studying catalytic efficiency of each mutation, we will be able to discover the molecular mechanisms that facilitate this neomorphic activity and help inform drug design for selective mutant IDH1 inhibitors that are effective against a wide range of tumor‐relevant mutants.Support or Funding InformationThis work was funded by the National Institutes of Health R00 CA187594 (C.D.S.), U54CA132384 (SDSU) & U54CA132379 (UC San Diego), MARC 5T34GM008303 (SDSU), and IMSD 5R25GM058906 (SDSU), as well as the California Metabolic Research Foundation (SDSU) and SDSU startup funds (C.D.S). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.”This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.