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Low pH Facilitates Heterodimerization of Mutant Isocitrate Dehydrogenase IDH1-R132H and Promotes Production of 2-Hydroxyglutarate.

ABSTRACT: Isocitrate dehydrogenase 1 (IDH1) is a key metabolic enzyme for maintaining cytosolic levels of α-ketoglutarate (AKG) and preserving the redox environment of the cytosol. Wild-type (WT) IDH1 converts isocitrate to AKG; however, mutant IDH1-R132H that is recurrent in human cancers catalyzes the neomorphic production of the oncometabolite d-2-hydroxyglutrate (D-2HG) from AKG. Recent work suggests that production of l-2-hydroxyglutarte in cancer cells can be regulated by environmental changes, including hypoxia and intracellular pH (pHi). However, it is unknown whether and how pHi affects the activity of IDH1-R132H. Here, we show that in cells IDH1-R132H can produce D-2HG in a pH-dependent manner with increased production at lower pHi. We also identify a molecular mechanism by which this pH sensitivity is achieved. We show that pH-dependent production of D-2HG is mediated by pH-dependent heterodimer formation between IDH1-WT and IDH1-R132H. In contrast, neither IDH1-WT nor IDH1-R132H homodimer formation is affected by pH. Our results demonstrate that robust production of D-2HG by IDH1-R132H relies on the coincidence of (1) the ability to form heterodimers with IDH1-WT and (2) low pHi or highly abundant AKG substrate. These data suggest cancer-associated IDH1-R132H may be sensitive to physiological or microenvironmental cues that lower pH, such as hypoxia or metabolic reprogramming. This work reveals new molecular considerations for targeted therapeutics and suggests potential synergistic effects of using catalytic IDH1 inhibitors targeting D-2HG production in combination with drugs targeting the tumor microenvironment.

SUBMITTER: Sesanto R

PROVIDER: S-EPMC8246651 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundMultiple studies concluded that oncometabolites (e.g. D-2-hydroxyglutarate (2-HG) related to mutant isocitrate dehydrogenase 1/2 (IDH1/2) and lactate) have tumour promoting potential. Regulatory mechanisms implicated in the maintenance of oncometabolite production have great interest. mTOR (mammalian target of rapamycin) orchestrates different pathways, influences cellular growth and metabolism. Considering hyperactivation of mTOR in several malignancies, the question has been addressed whether mTOR operates through controlling of oncometabolite accumulation in metabolic reprogramming.MethodsHT-1080 cells - carrying originally endogenous IDH1 mutation - were used in vitro and in vivo. Anti-tumour effects of rapamycin were studied using different assays. The main sources and productions of the oncometabolites (2-HG and lactate) were analysed by 13C-labeled substrates. Alterations at protein and metabolite levels were followed by Western blot, flow cytometry, immunohistochemistry and liquid chromatography mass spectrometry using rapamycin, PP242 and different glutaminase inhibitors, as well.ResultsRapamycin (mTORC1 inhibitor) inhibited proliferation, migration and altered the metabolic activity of IDH1 mutant HT-1080 cells. Rapamycin reduced the level of 2-HG sourced mainly from glutamine and glucose derived lactate which correlated to the decreased incorporation of 13C atoms from 13C-substrates. Additionally, decreased expressions of lactate dehydrogenase A and glutaminase were also observed both in vitro and in vivo.ConclusionsConsidering the role of lactate and 2-HG in regulatory network and in metabolic symbiosis it could be assumed that mTOR inhibitors have additional effects besides their anti-proliferative effects in tumours with glycolytic phenotype, especially in case of IDH1 mutation (e.g. acute myeloid leukemias, gliomas, chondrosarcomas). Based on our new results, we suggest targeting mTOR activity depending on the metabolic and besides molecular genetic phenotype of tumours to increase the success of therapies.

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Low pH Facilitates Heterodimerization of Mutant Isocitrate Dehydrogenase IDH1-R132H and Promotes Production of 2-Hydroxyglutarate.

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