Understanding the Structure‐Function Relationship of IDH1 R132 mutants and the Effect of Allosteric Inhibitors

Alterations in tumor cell metabolism were first described nearly a century ago by Otto Warburg. The Warburg effect consists of a shift from oxidative phosphorylation to glycolysis as the main source of energy for tumor cells. Isocitrate dehydrogenase 1 (IDH1), an important dimeric metabolic enzyme, is responsible for the NADP + ‐dependent oxidative decarboxylation of isocitrate to α‐ketoglutarate (αKG) in the cytosol and peroxisomes of the cell. There are two other isoforms, IDH2 and IDH3, that reside in the mitochondria. Mutations in the IDH1 gene, most commonly at residue 132, drive ~80% of lower grade gliomas and secondary glioblastomas. The most common IDH1 mutant observed in cancer is R132H, but other less frequent mutations have been identified such as R132C, R132G, and R132S. The R132 mutant IDH1 enzymes lose their activity for the native reaction but they gain a neomorphic function consisting of the NADPH‐dependent reduction of αKG to D‐2‐hydroxyglutarate (D2HG). Consequently, cells that contain IDH1 mutations associated with cancer show accumulation of D2HG. D2HG is a proposed oncometabolite since it acts as a competitive inhibitor of many αKG‐dependent enzymes such as DNA and histone demethylases, resulting in genome hypermethylation to alter gene regulation. With the help of steady‐state kinetic analysis, we observed a tumor‐relevant R132Q IDH1 mutant that has conserved the native reaction as well as gained the neomorphic one. Due to its high frequency of mutations in gliomas, their critical downstream effects, and their inhibition specificity, IDH1 has become an attractive therapeutic target. Interestingly, IDH1 inhibitors are very selective and potent against mutant forms and have low affinity for wild type IDH1. Although many of the downstream effects of IDH1 mutations have been studied, a full structure‐function characterization of mutant IDH1 catalysis and the features driving selectivity of allosteric inhibitors are currently limited. We hypothesize that IDH1 mutants that conserve wildtype activity will have a more stable regulatory domain, thus, decreasing the potency of the inhibitor on the enzyme. We show that ML309 and AGI‐5198 have over a 100‐fold increase in IC 50 for R132Q compared to R132H. These results suggest a possible stabilization of the regulatory domain in IDH1 R132Q, causing loss of selectivity and potency of the inhibitor. Since the structure of R132Q has not been determined, we will create models containing that mutation as well as molecular dynamics simulations to determine key conformational changes that will explain the lack of inhibition for R132Q as also seen in wild type. In the future, we aim to perform structural analysis of IDH1 R132Q. This work will clarify which IDH1 mutations in patients will likely respond to targeted mutant IDH1 therapy.AGI‐5198 inhibition on different R132 IDH1 mutants. R132Q has the highest IC 50 (>100 fold) compare to the other IDH1 mutants.ML309 inhibition on different R132 IDH1 mutants. R132Q has the highest IC50 (>100) compare to the other IDH1 mutants.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal .