Abstract
D-2-Hydroxyglutarate (D-2-HG) is regarded as an oncometabolite. It is found at elevated levels in certain malignancies such as acute myeloid leukaemia and glioma. It is produced by a mutated isocitrate dehydrogenase IDH1/2, a low-affinity/high-capacity enzyme. Its degradation, in contrast, is catalysed by the high-affinity/low-capacity enzyme D-2-hydroxyglutarate dehydrogenase (D2HDH). So far, it has not been proven experimentally that the accumulation of D-2-HG in IDH mutant cells is the result of its insufficient degradation by D2HDH. Therefore, we developed an LC-MS/MS-based enzyme activity assay that measures the temporal drop in substrate and compared this to the expression of D2HDH protein as measured by Western blot. Our data clearly indicate, that the maximum D-2-HG degradation rate by D2HDH is reached in vivo, as vmax is low in comparison to production of D-2-HG by mutant IDH1/2. The latter seems to be limited only by substrate availability. Further, incubation of IDH wild type cells for up to 48 hours with 5 mM D-2-HG did not result in a significant increase in either D2HDH protein abundance or enzyme activity.
Highlights
Among the oncogenic mutations reported to alter cancer cell metabolism, mutations in isocitrate dehydrogenase (IDH), which normally catalyses the oxidative decarboxylation of isocitrate to α-ketoglutarate, have attracted particular attention[1,2]
Similar to cancer, IDH2 mutation leads to the production and accumulation of D-2-HG in the presence of functional D-2-hydroxyglutarate dehydrogenase (D2HDH), which results in an even higher urinary excretion of D-2-HG compared to patients with D-2-HGA type I20
Elevated D-2-HG levels impact prognosis and therapy of patients with mutated IDH1/2 tumours like glioma or acute myeloid leukaemia (AML). They are problematical for HG-aciduria patients. 2-HG is involved in the development and progression of these diseases and antibodies targeting mutated IDH1/2 in cancer can prolong survival in treated patients[29,30]
Summary
Among the oncogenic mutations reported to alter cancer cell metabolism, mutations in isocitrate dehydrogenase (IDH), which normally catalyses the oxidative decarboxylation of isocitrate to α-ketoglutarate (αKG), have attracted particular attention[1,2]. In these patients, similar to cancer, IDH2 mutation leads to the production and accumulation of D-2-HG in the presence of functional D2HDH, which results in an even higher urinary excretion of D-2-HG compared to patients with D-2-HGA type I20. Similar to cancer, IDH2 mutation leads to the production and accumulation of D-2-HG in the presence of functional D2HDH, which results in an even higher urinary excretion of D-2-HG compared to patients with D-2-HGA type I20 This suggests that D-2-HG production exceeds its degradation and excretion. The results contribute to a better understanding of the mechanism of D-2-HG accumulation in tumours with IDH1/2 mutations
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