The prognostic IDH1( R132 ) mutation is associated with reduced NADP+-dependent IDH activity in glioblastoma

Abstract

Somatic mutations in the isocitrate dehydrogenase 1 gene (IDH1) occur at high frequency in gliomas and seem to be a prognostic factor for survival in glioblastoma patients. In our set of 98 glioblastoma patients, IDH1 ( R132 ) mutations were associated with improved survival of 1 year on average, after correcting for age and other variables with Cox proportional hazards models. Patients with IDH1 mutations were on average 17 years younger than patients without mutation. Mutated IDH1 has a gain of function to produce 2-hydroxyglutarate by NADPH-dependent reduction of alpha-ketoglutarate, but it is unknown whether NADPH production in gliomas is affected by IDH1 mutations. We assessed the effect of IDH1 (R132 ) mutations on IDH-mediated NADPH production in glioblastomas in situ. Metabolic mapping and image analysis was applied to 51 glioblastoma samples of which 16 carried an IDH1 (R132 ) mutation. NADP+-dependent IDH activity was determined in comparison with activity of NAD+-dependent IDH and all other NADPH-producing dehydrogenases, glucose-6-phosphate dehydrogenase, 6-phosphogluconate dehydrogenase, malate dehydrogenase, and hexose-6-phosphate dehydrogenase. The occurrence of IDH1 mutations correlated with approx. twofold diminished NADP+-dependent IDH activity, whereas activity of NAD+-dependent IDH and the other NADP+-dependent dehydrogenases was not affected in situ in glioblastoma. The total NADPH production capacity in glioblastoma was provided for 65% by IDH activity and the occurrence of IDH1 (R132 ) mutation reduced this capacity by 38%. It is concluded that NADPH production is hampered in glioblastoma with IDH1 (R132 ) mutation. Moreover, mutated IDH1 consumes rather than produces NADPH, thus likely lowering NADPH levels even further. The low NADPH levels may sensitize glioblastoma to irradiation and chemotherapy, thus explaining the prolonged survival of patients with mutated glioblastoma.

Fig. 1

Adjusted survival curves comparing IDH1 ^R132 mutated and non-mutated glioblastoma patients. Constructed with the use of multivariable Cox proportional hazards regression, and adjusted for age, Karnofsky performance score, irradiation and additional treatments

Fig. 2

NADP⁺-dependent IDH activity in IDH1 ^R132 non-mutated (n = 36) and mutated (n = 15) tumors. NADP⁺-dependent IDH activity staining of IDH1 ^R132 non-mutated a and mutated c glioblastoma cryostat sections. The amount of blue color (nitro BT-formazan) directly reflects NADP⁺-dependent IDH activity (production of NADPH). H&E staining of serial sections of non-mutated (b) and mutated (d) samples

Fig. 3

Capacity (maximum velocity; V _max) of dehydrogenases in IDH^R132 non-mutated and mutated tumors expressed as micromoles NADPH or NADH produced/ml of tissue/min, ±SD. NADP⁺-dependent IDH activity (a); individual values of NADP⁺-dependent IDH activity (b); NADP⁺-dependent G6PD activity (c); NAD⁺-dependent IDH activity (d)