Active site remodeling in tumor-relevant IDH1 mutants drives distinct kinetic features and potential resistance mechanisms

Abstract

Human isocitrate dehydrogenase 1 (IDH1) is a highly conserved enzyme that converts isocitrate to α-ketoglutarate (αKG). IDH1 mutations affecting residue R132 drive tumor formation, and several FDA-approved mutant IDH1 inhibitors are available. Most mutants cannot catalyze the conventional reaction, but gain a neomorphic activity that produces D-2-hydroxyglutarate (D2HG). We previously reported that IDH1 R132Q uniquely preserves conventional activity while catalyzing robust D2HG production, allowing an opportunity to compare these reaction mechanisms within a single active site. Here, we employed static and dynamic structural methods and found that, compared to R132H, the R132Q active site adopted a conformation primed for catalysis with optimized substrate binding, hydride transfer, and prevention of inhibitor binding. When binding isocitrate, IDH1 R132Q exhibited a closed conformation with features consistent with decreased catalysis versus WT. Binding neomorphic reaction substrates resulted in semi-closure of IDH1 R132Q with profoundly limited active site solvent accessibility, and featured a distinct αKG binding pocket compared to R132H. This work enhances our understanding of fundamental IDH1 mechanisms while pinpointing regions that can improve inhibitor selectivity and drive resistance.

Catalytically distinct IDH1 mutants tune phenotype severity in tumor models

“…We sought to determine how the degree of D2HG production efficiency tunes phenotype severity in models of mutant IDH1-driven tumors. We have shown previously that R132Q mutant homodimers produce D2HG more efficiently than R132H mutant homodimers 20–22 , though WT:mutant heterodimers may also be found as IDH1 mutations are found heterozygously in patients. Thus, we first generated 1:1 mixtures of purified WT and mutant IDH1 enzymes to allow heterodimers to form.…”

“…Others have reported that glioma tissue (expressing IDH1 R132H/C/G mutations) and cell line models (expressing IDH1 R132H/C/G/S/L mutations) had varying concentrations of D2HG depending on the IDH1 mutation present, with R132H associated with the lowest levels of D2HG 18 , though no changes in overall survival were observed with patients with IDH1 R132C/G/S/L/K mutations versus R132H 19 . We previously reported that kinetic features may play some role in these observed differences, noting that purified IDH1 R132Q was far more catalytically efficient at D2HG production than R132H 20,21 due to activating structural features 22 . An extensive study on astrocytomas binned as R132H mutated and non-R132H IDH1/2- mutated (which included R132C/L/G/S and IDH2 R140Q/W/G/L and R172W/K/M/G/S/T) showed that non- R132H IDH1/2-mutated tumors had overall increased DNA methylation, decreased gene expression, and better prognosis, suggesting an intriguing possibility of differences in methylation phenotype intensity within IDH1 mutants and/or among IDH1 and IDH2 mutants 23 .…”