Isocitrate Dehydrogenase Protein Research Articles

FOXM1 requires IDH1 for late genes expression in mitotic cells.

Isocitrate dehydrogenase 1 (IDH1) is a metabolic enzyme that converts isocitrate to α-ketoglutarate in cells. However, research on IDH1 is more focused on the metabolite D-2-hydroxyglutarate than the cellular roles of the IDH1 protein. Metabolic enzymes can moonlight by participating in diverse cellular processes in cancer cells. This moonlighting function of the metabolic enzymes can contribute to changes in gene expression. It is unknown whether IDH1 associates with any transcription factor. We asked whether IDH1 coordinates with forkhead box protein M1 (FOXM1) in mitotic cells to regulate late genes expression. We found that depletion of IDH1 reduces canonical FOXM1-target expression in mitotic cells. Also, IDH1 binds to FOXM1 and a subset of MuvB proteins, Lin-9 and Lin-54, in mitotic cells. Based on these observations, we suggest that IDH1 coordinates with FOXM1 in mitotic cells to regulate late genes expression.

Abstract 4333: Identification of glucose-6-phosphate dehydrogenase dependency in IDH1 mutant glioma cells using functional genomics

Abstract Astrocytoma and oligodendroglioma are most commonly initiated by missense mutations at the arginine 132 codon of isocitrate dehydrogenase 1 (IDH1). Biochemically the mutant IDH1 protein acquires neomorphic activity: reductive NADPH-dependent catalysis of α-ketoglutarate (αKG) to (R)-2-hydroxyglutarate [(R)-2-HG]. (R)-2-HG interferes with the function of αKG-dependent epigenetic modifiers, resulting in extensive epigenetic remodeling that impairs cellular differentiation and promotes gliomagenesis. Strategies for selectively targeting IDH1-mutant cells offer promise for improved treatment over the standard modalities of surgery, chemotherapy and radiotherapy. Whole-genome CRISPR/Cas9 knockout screens were conducted to identify gene dependencies in isogenic IDH1-mutant and wild-type (WT) U-87 MG cells. Cells were transduced with the Brunello single guide RNA (sgRNA) library, grown for 21 days, then sgRNA distributions were assessed by amplicon sequencing. Gene dependencies were identified using CRISPRcleanR and BAGELR pipelines. We discovered that glucose-6-phosphate dehydrogenase (G6PD), a pentose phosphate pathway (PPP) enzyme, was needed for viability of IDH1-mutant cells, but was dispensable for U-87 MG WT cells. The PPP is the main producer of cytosolic NADPH that is required for de novo lipogenesis, maintenance of antioxidants and production of (R)-2-HG by mutant IDH1. Previous reports demonstrated that IDH1-mutant cells increase PPP flux to support these processes. We have validated this dependency using G6PDi-1, a cell-active inhibitor of G6PD. Growth inhibition (IC50) assays demonstrated that IDH1-mutant U-87 MG cells were threefold more sensitive to G6PDi-1 with IC50 of 23 μM and enhanced cell death (increased propidium iodide uptake), compared with WT U-87 MG cells with IC50 of 67 μM. G6PD knockout was then conducted to further validate this dependency. Nucleofection of IDH1 WT U-87 MG cells with G6PD multi guide RNA-Cas9 ribonucleoproteins resulted in highly efficient knockout of G6PD at day 5 (99-100%), which was maintained at day 21 (96-98%). In contrast, efficient G6PD knockout in IDH1-mutant U-87 MG cells at day 5 (98-100%) was lost following 21 days culture (0-5%), indicating death of almost all edited cells within the culture. Finally, Kaplan-Meier analysis of IDH1-mutant patients in the TCGA lower grade glioma dataset demonstrated that higher G6PD expression correlated with worse median overall survival (7.3 versus 9.5 years; log-rank p=0.01), but was not prognostic in the IDH1 WT TCGA glioblastoma multiforme dataset. These findings demonstrate that a cell line model of IDH1-mutant glioma is highly dependent on G6PD for survival. This observation builds on the recognized importance of the PPP in the metabolic biology of IDH1-mutant malignancy by establishing G6PD as a targetable dependency in this disease. Citation Format: Kim M. Paras, Pooja Patel, Paul Choi, William R. Wilson, Tet-Woo Lee, Stephen M. Jamieson, Dean C. Singleton. Identification of glucose-6-phosphate dehydrogenase dependency in IDH1 mutant glioma cells using functional genomics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4333.

Cytosolic isocitrate dehydrogenase regulates plant stem cell maintenance in response to nutrient deficiency.

WUSCHEL (WUS) and WUSCHEL-RELATED HOMEOBOX (WOX) proteins determine stem cell maintenance for continual plant growth and development under changing environmental conditions. Nutrient availability is an environmental factor that substantially controls plant growth and development. However, how plant stem cell homeostasis is regulated under nutrient deficiency remains to be elucidated. Here, we showed that cytosolic isocitrate dehydrogenase (ICDH) plays an important role in nutrient sensing of stem cells in Arabidopsis (Arabidopsis thaliana). Nutrient deficiency induced the cytoplasmic-to-nuclear translocation of cytosolic ICDH protein. ICDH can interact with WUS/WOX protein as a complex that further promotes WUS/WOX expression by binding to its promoter. WUS/WOX expression in the icdh-2 mutant was lower than that of wild-type plants under nutrient deficiency. Consistently, loss of ICDH function caused a more serious growth repression under nutrient deficiency that was independent of ICDH's catalytic activity. Therefore, cytosolic ICDH regulates stem cell homeostasis of plants in response to nutrient deficiency.

Isocitrate dehydrogenase mutations in gliomas: A review of current understanding and trials.

Isocitrate dehydrogenase (IDH) is a key enzyme in normal metabolism and homeostasis. However, mutant forms of IDH are also defining features of a subset of diffuse gliomas. In this review, we highlight current techniques targeting IDH-mutated gliomas and summarize current and completed clinical trials exploring these strategies. We discuss clinical data from peptide vaccines, mutant IDH (mIDH) inhibitors, and PARP inhibitors. Peptide vaccines have the unique advantage of targeting the specific epitope of a patient's tumor, inducing a highly tumor-specific CD4+ T-cell response. mIDH-inhibitors, on the other hand, specifically target mutant IDH proteins in cancer cell metabolism and thus help halt gliomagenesis. We also explore PARP inhibitors and their role in treating diffuse gliomas, which exploit IDH-mutant diffuse gliomas by allowing the persistence of unrepaired DNA complexes. We summarize various completed and current trials targeting IDH1 and IDH2 mutations in diffuse gliomas. Therapies targeting mutant IDH have significant promise in treating progressive or recurrent IDH-mutant gliomas and may significantly change treatment paradigms in the next decade.

Anaplastic ganglioglioma-A diagnosis comprising several distinct tumour types.

Anaplastic ganglioglioma is a rare tumour, and diagnosis has been based on histological criteria. The 5th edition of the World Health Organization Classification of Tumours of the Central Nervous System (CNS WHO) does not list anaplastic ganglioglioma as a distinct diagnosis due to lack of molecular data in previous publications. We retrospectively compiled a cohort of 54 histologically diagnosed anaplastic gangliogliomas to explore whether the molecular profiles of these tumours represent a separate type or resolve into other entities. Samples were subjected to histological review, desoxyribonucleic acid (DNA) methylation profiling and next-generation sequencing. Morphological and molecular data were summarised to an integrated diagnosis. The majority of tumours designated asanaplastic gangliogliomas resolved into other CNS WHO diagnoses, most commonly pleomorphic xanthoastrocytoma (16/54), glioblastoma, isocitrate dehydrogenase protein (IDH) wild type and diffuse paediatric-type high-grade glioma, H3 wild type and IDH wild type (11 and 2/54), followed by low-grade glial or glioneuronal tumours including pilocytic astrocytoma, dysembryoplastic neuroepithelial tumour and diffuse leptomeningeal glioneuronal tumour (5/54), IDH mutant astrocytoma (4/54) and others (6/54). A subset of tumours (10/54) was not assignable to a CNS WHO diagnosis, and common molecular profiles pointing to a separate entity were not evident. In summary, we show that tumours histologically diagnosed as anaplastic ganglioglioma comprise a wide spectrum of CNS WHO tumour types with different prognostic and therapeutic implications. We therefore suggest assigning this designation with caution and recommend comprehensive molecular workup.

Determination of intracellular 2-hydroxyglutarate enantiomers using two-dimensional liquid chromatography

In this study, a heart-cutting two-dimensional liquid chromatography (2D-LC) method for the determination of intracellular 2-hydroxyglutarate (2-HG) enantiomers is proposed. D-2-HG is an oncometabolite produced by tumors harboring isocitrate dehydrogenase (IDH) mutations. Hence, achieving accurate quantification of the 2-HG enantiomers in cancer cells is important. In this study, a fluorescence derivatization reagent, 4-nitro-7-piperazino-2,1,3-benzoxadiazole (NBD-PZ), was used. We found that 2-HG was successfully derivatized with NBD-PZ after lactonization. The enantiomers of 2-HG derivatized with NBD-PZ were quantified using the heart-cutting 2D-LC method. NBD-PZ-2-HG was stable for 1 week and was separated from endogenous compounds using an octadecyl silica column before enantiomeric separation on a chiral column. The developed heat-cutting 2D-LC method was validated and successfully applied to cell samples. The amount of D-2-HG and L-2-HG in U937 cells were 9.8 and 8.3 pmol/5.0 × 105 cells, respectively. Furthermore, high amounts of D-2-HG were measured in U937 cells expressing mutant IDH proteins as expected. This method can therefore be applied to further elucidate the role of 2-HG in cancer cells, particularly those harboring isocitrate dehydrogenase mutations.

Genetic Profiling and Deep IDH1 Mutation Clearance to ≤0.04% in Ivosidenib (AG-120)-Treated Patients with Mutant IDH1 Relapsed or Refractory and Untreated AML

Genetic Profiling and Deep IDH1 Mutation Clearance to ≤0.04% in Ivosidenib (AG-120)-Treated Patients with Mutant IDH1 Relapsed or Refractory and Untreated AML

Identification and characterization of a novel mutant isocitrate dehydrogenase 1 inhibitor for glioma treatment

Isocitrate dehydrogenase 1 (IDH1) mutant R132H, promoting the oncometabolite D-2-hydroxyglutarate (D2HG), is a driver mutation and an emerging therapeutic target in glioma. This study identified a novel mutant IDH1 inhibitor, WM17, by virtual screening and enzymatic confirmation. It could bind to and increase mutant IDH1 protein’s thermostability in both endogenous heterozygous cells and exogenous overexpressed cells. Consequently, WM17 reversed the accumulation of D2HG and histone hypermethylation in IDH1 mutated cells. Finally, we concluded that WM17 significantly inhibited cell migration in IDH1 mutated glioma cells, although it has no apparent effect on cell proliferation. Further studies are guaranteed toward the development of WM17 as a therapeutic agent for IDH1 mutated glioma.

AHA1 upregulates IDH1 and metabolic activity to promote growth and metastasis and predicts prognosis in osteosarcoma

Osteosarcoma (OS) is the most common primary malignant bone tumor in children and adolescents. Although activator of HSP90 ATPase activity 1 (AHA1) is reported to be a potential oncogene, its role in osteosarcoma progression remains largely unclear. Since metabolism reprogramming is involved in tumorigenesis and cancer metastasis, the relationship between AHA1 and cancer metabolism is unknown. In this study, we found that AHA1 is significantly overexpressed in osteosarcoma and related to the prognosis of osteosarcoma patients. AHA1 promotes the growth and metastasis of osteosarcoma both in vitro and in vivo. Mechanistically, AHA1 upregulates the metabolic activity to meet cellular bioenergetic needs in osteosarcoma. Notably, we identified that isocitrate dehydrogenase 1 (IDH1) is a novel client protein of Hsp90-AHA1. Furthermore, the IDH1 protein level was positively correlated with AHA1 in osteosarcoma. And IDH1 overexpression could partially reverse the effect of AHA1 knockdown on cell growth and migration of osteosarcoma. Moreover, high IDH1 level was also associated with poor prognosis of osteosarcoma patients. This study demonstrates that AHA1 positively regulates IDH1 and metabolic activity to promote osteosarcoma growth and metastasis, which provides novel prognostic biomarkers and promising therapeutic targets for osteosarcoma patients.

Exosomal IDH1 increases the resistance of colorectal cancer cells to 5-Fluorouracil.

Chemoresistance challenges the clinical treatment of colorectal cancer and requires an urgent solution. Isocitrate dehydrogenase 1 (IDH1) is a key enzyme involved in glucose metabolism that mediates the malignant transformation of tumors. However, the mechanisms by which IDH1 is involved in colorectal cancer cell proliferation and drug resistance induction remain unclear. In this study, we found that IDH1 was highly expressed in human colorectal cancer tissues and could be used to indicate a high-grade tumor. In vitro gene overexpression and knockdown were used to determine whether IDH1 promoted the proliferation of the colorectal cancer cell line HCT8 and resistance to 5-Fluorouracil (5FU). Further studies have shown that the 5FU-resistant cell line, HCT8FU, secreted exosomes that contained a high level of IDH1 protein. The exosomal IDH1 derived from 5FU-resistant cells enhanced the resistance of 5FU-sensitive cells. Metabolic assays revealed that exosomes derived from 5FU-resistant cells promoted a decrease in the level of IDH1-mediated NADPH, which is associated with the development of 5FU resistance in colorectal cancer cells. Therefore, exosomal IDH1 may be the transmitter and driver of chemoresistance in colorectal cancer and a potential chemotherapy target.

LncRNA IDH1-AS1 suppresses cell proliferation and tumor growth in glioma.

Glioma is a type of brain tumor that is common globally, and is associated with a variety of genetic changes. It has been reported that isocitrate dehydrogenase 1 (IDH1) is overexpressed in glioma and in HeLa cells. The lncRNA IDH1-AS1 is believed to interact with IDH1, and when IDH1-AS1 is overexpressed, HeLa cell proliferation is inhibited. However, the effects of IDH1-AS1 on glioma were relatively unknown. The results from this work show that IDH1-AS1 is downregulated in the glioma tissues. We used primary glioblastoma cell lines U251 and U87-MG to study the effects of IDH1-AS1 on glioma cell growth, in vitro and in vivo. We found that when IDH1-AS1 is overexpressed cell proliferation is inhibited, cell cycle is arrested at the G1 phase, and the protein expression levels of cyclinD1, cyclinA, cyclinE, CDK2, and CDK4 are decreased. We found that cell apoptosis was increased when IDH1-AS1 was overexpressed, as evidenced by increases in the levels of cleaved caspase-9 and -3. Conversely, knockdown of IDH1-AS1 promoted cell proliferation. Moreover, we proved that overexpression of IDH1-AS1 inhibits the tumorigenesis of U251 cells, in vivo. Furthermore, IDH1-AS1 did not affect IDH1 protein expression, but altered its enzymatic activities in glioma cells. Silencing of IDH1 reversed the effects of IDH1-AS1 upregulation on cell viability. Hence, our study provides first-hand evidence for the effects of lncRNA IDH1-AS1 on gliomas. Because overexpressing IDH1-AS1 inhibited cell growth, IDH1-AS1 could also be considered as a potential target for glioma treatment.

Abstract 2046: Inhibiting IDH mutations in low-grade glioma alters cellular function and the immune environment

Abstract Somatic mutations in the genes encoding the metabolic enzymes isocitrate dehydrogenase (IDH) 1 and 2 occur in many cancers, including glioma, cholangiocarcinoma, and acute myeloid leukemia. Mutant IDH (mIDH) proteins have a gain-of-function enzyme activity, catalyzing the reduction of alpha-ketoglutarate (α-KG) to the oncometabolite D-2-hydroxyglutarate (2-HG). 2-HG competitively inhibits α-KG-dependent enzymes, leading to epigenetic dysregulation and tumorigenesis. TET2 is an α-KG-dependent enzyme mediating the conversion of 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC), which leads to DNA demethylation. In mIDH tumors, TET2 activity is inhibited by 2-HG, leading to DNA hypermethylation. In addition, IDH mutations are associated with immune evasion in gliomas. IDH mutations are common in low-grade gliomas (LGG; WHO grade 2/3) and include mutations in IDH1 (~80%) and IDH2 (~4%). Ivosidenib (AG-120, IVO) is a first-in-class, oral inhibitor of mIDH1, and vorasidenib (AG-881, VOR) is an oral, potent, brain-penetrant inhibitor of both mIDH1 and mIDH2. In an ongoing perioperative study (NCT03343197), IVO and VOR demonstrated brain penetrance and >90% suppression of 2-HG in resected mIDH1 gliomas after pre-operative treatment for approximately 4 weeks. In this analysis, we examined the cellular mechanisms underlying mIDH inhibition in LGG using resected tumor tissues collected following treatment with IVO and VOR. Optimal 2-HG suppression (defined by post-treatment 2-HG levels equal to or lower than the upper range of IDH wild-type tumor 2-HG levels) in IVO- or VOR-treated samples (n=17) led to a ~2-fold increase in 5hmC compared with untreated controls and banked reference samples (n=35; P=0.04). Furthermore, immunohistochemistry (IHC) analysis of the proliferation marker Ki-67 showed a ~3-fold decrease in the percentage of Ki-67-positive cells in IVO- or VOR-treated samples with optimal 2-HG suppression (n=21) compared with untreated controls (n=5; P=0.02). Tumor immune microenvironment analyses were also conducted. Comparing IVO- or VOR-treated samples (n=21) with untreated controls and banked reference samples (n=33), RNAseq analysis showed a ~2-fold decrease in the expression of tumor-associated macrophage markers CD68 (P=0.01) and CD163 (P=0.03) in mIDH oligodendrogliomas. In addition, IHC analysis from paired pre-treatment (archival) and post-treatment samples showed an increase of CD3+ T-cell infiltration (P=0.03) and CD8+ T-cell infiltration (P=0.01) in IVO- or VOR-treated samples with optimal 2-HG suppression (n=21). Single-cell RNAseq was conducted for a subset of samples, and the results will be presented. Overall, these data suggest that mIDH inhibition decreases DNA hypermethylation and tumor cell proliferation, and activates the immune microenvironment in mIDH LGG. In conclusion, these data elucidate the mechanisms underlying mIDH inhibition and support further development of mIDH inhibitors in mIDH LGG. Page 1 of 1 Citation Format: Min Lu, Ingo K. Mellinghoff, Aaron Diaz, Jennie W. Taylor, Sung Choe, Ania Tassinari, Dongwei Zhu, Katie Sellers, Kha Le, Feng Tai, Islam Hassan, Shuchi S. Pandya, Lori Steelman, Bin Wu. Inhibiting IDH mutations in low-grade glioma alters cellular function and the immune environment [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 2046.

Abstract 3731: Targeting the isocitrate dehydrogenase 1 (IDH1) metabolic enzyme in prostate cancer

Abstract Prostate cancer cells (PCa) are dependent on the androgen receptor (AR) for their aberrant proliferation and survival. We have recently discovered that AR induces a reprogramming of PCa cell metabolism by controlling the cytoplasmic wild-type enzyme isocitrate dehydrogenase 1 (IDH1), which results in an enhanced proliferation of tumour cells. However, the specific metabolic functions of IDH1 in PCa or how to use the reliance of tumour cells on this enzyme as a therapeutic avenue, is elusive. In in vitro human PCa models, we showed that IDH1 protein levels and activity are increased in an AR-dependent manner. Using pharmacological and genetic tools, we showed that IDH1 is a major contributor to the replenishment of NADPH levels in PCa. This cofactor plays a key role in the synthesis of biomaterials required for cellular division, and our results indicate that IDH1 contribute to 30-40% of total cellular NADPH levels. In that context, blockade of IDH1 was shown to alter the mTOR signaling, a central regulator of cellular anabolism, which is linked to decreased cellular proliferation rates. FDA-approved pharmacological inhibitors of mutant IDH1 significantly inhibited IDH activity and proliferation in PCa cells, suggesting that such inhibitors could be used to treat PCa patients even in absence of IDH1 mutation. Globally, our results demonstrate that IDH1 is a key player in proliferative anabolic pathways in PCa. Importantly, they also support the hypothesis that inhibition of IDH1 using already-approved molecules represents one viable therapeutic solution. Citation Format: Kevin Gonthier, Cindy Weidmann, Lilianne Frégeau-Proulx, Étienne Audet-Walsh. Targeting the isocitrate dehydrogenase 1 (IDH1) metabolic enzyme in prostate cancer [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3731.

Abstract 772: Performance characteristics of the Oncomine™ Dx Target Test for the detection of IDH1 R132 variants from formalin-fixed, paraffin-embedded (FFPE) cholangiocarcinoma samples

Abstract Background: Cholangiocarcinoma (CC) is an aggressive bile-duct cancer that affects 8000 people per year in the US. This silent disease manifests symptoms only in advanced stages, resulting in a 5-year overall survival rate of < 10%. Mutations in the metabolic enzyme isocitrate dehydrogenase 1 (IDH1) have been observed in ~20% of CC. These mutations are implicated in promoting tumorigenesis potentially by generation of an ‘oncometabolite' resulting in changes in DNA methylation. There are currently no approved targeted therapies for CC. Ivosidenib, developed by Agios Pharmaceuticals, is a first-in-class, oral, targeted, small-molecule inhibitor of the mutant IDH1 protein currently being evaluated in patients with advanced CC with an IDH1 mutation (ClarIDHy). The Oncomine Dx Target Test (ODxTT) is an FDA approved in vitro diagnostic test for the detection of somatic alterations in human DNA and RNA isolated from FFPE non-small cell lung cancer samples. Thermo Fisher Scientific has partnered with Agios Pharmaceuticals to validate the ODxTT for the detection of IDH1 R132 mutations in CC. Here we present the results from our analytical studies which establish sensitivity, reproducibility and accuracy of the IUO ODxTT to detect 5 IDH1 R132 variants from FFPE CC specimens. Methods: Analytical studies were performed to determine the limit of detection (LOD) and reproducibility of the assay to detect the variants of interest. LOD was determined for each of the 5 variants of interest using either FFPE clinical sample or FFPE cell line blends. Reproducibility of the assay was evaluated by testing clinical samples blends for 3 of 5 variants at ~2x LOD across 3 sites, 6 operators and 3 lots of reagents. Analytical accuracy of the ODxTT was determined by testing over 340 clinical samples with both the ODxTT and a validated orthogonal method. The orthogonal method was Sanger sequencing coupled with Applied Biosystems™ Minor Variant Finder Software to allow calling minor variants down to 5%. Results: 1. LOD of the ODxTT for all 5 variants of interest (R132C, R132G, R132S, R132L and R132H) was determined to be at an allelic frequency (AF) between 3.7% - 5.5%. 2. Reproducibility of the ODxTT to detect R132C, R132G and R132L variants from clinical sample blends was 100% when each variant was tested at an AF of ~2x LOD. 3. Analytical accuracy: The positive percent agreement (PPA) of the ODxTT to Sanger sequencing was 99.4% (163/164) and the negative percent agreement (NPA) was 96.5% (164/170) when excluding unknowns (invalids and no calls). Including unknowns, the PPA of the ODxTT to Sanger sequencing was 97% (163/168) and the NPA was 90.6% (164/181). Conclusion: The data presented here demonstrated that the ODxTT is a sensitive, reproducible and accurate assay for the detection of IDH1 R132 variants from 10 ng of DNA extracted from FFPE CC specimens. Citation Format: Aparna R. Aiyer, Tristan Nater, Matthew Brentnall, Foram Rathi, Bharath Bharadwaj, Mini Manchanda, Viet Nhan, Lauren Abutin, Peter LoCoco, Bin Wu, Jay Shaw. Performance characteristics of the Oncomine™ Dx Target Test for the detection of IDH1 R132 variants from formalin-fixed, paraffin-embedded (FFPE) cholangiocarcinoma samples [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 772.

The Long Non-Coding RNA IDH1-AS1 Promotes Prostate Cancer Progression by Enhancing IDH1 Enzyme Activity.

PurposeLong non-coding RNAs (lncRNAs) are involved in the development of various tumors including prostate cancer. The purpose of this study was to explore the function of a natural antisense RNA, IDH1-AS1, exerting potential carcinogenic effects in prostate cancer through a novel molecular mechanism.Materials and MethodsGEPIA and CCLE databases were searched to identify alterations in the expression of IDH1-AS1, which were then verified by RT-qPCR in 20 pairs of matched tumor and normal tissue samples. Subsequently, CCK-8, EdU, and transwell assays were conducted to investigate the carcinogenic effect of IDH1-AS1. RT-qPCR, Western blot, and isocitrate dehydrogenase 1 (IDH1) enzyme activity assays were used to explore the functional relationship between IDH1-AS1 and its sense gene IDH1.ResultsIDH1-AS1 expression was found to be significantly increased in prostate cancer tissues and cell lines. IDH1-AS1 knockdown significantly inhibited the proliferation and migration of prostate cancer cells. Interestingly, RT-qPCR and Western blot analyses revealed that IDH1-AS1 did not significantly affect the expression of IDH1 mRNA or protein but was involved in the regulation of IDH1 enzyme activity in prostate cancer cells.ConclusionOur experiments revealed that the carcinogenic effects of IDH1-AS1 in prostate cancer may depend on a new molecular mechanism, which directly alters IDH1 enzyme activity. Our findings indicate that IDH1-AS1 is a novel candidate target for prostate cancer treatment.

IDH1 mutation detection in plasma circulating tumor DNA (ctDNA) and association with clinical response in patients with advanced intrahepatic cholangiocarcinoma (IHC) from the phase III ClarIDHy study.

4576 Background: Mutations in isocitrate dehydrogenase 1 ( IDH1) are detected in ~13% of IHCs. Ivosidenib (IVO) is a first-in-class, oral inhibitor of the mutant IDH1 (mIDH1) protein. In ClarIDHy, a global, phase 3, double-blind study in previously treated patients with advanced m IDH1 IHC (N = 186), IVO demonstrated an improvement in progression-free survival (PFS) vs placebo (hazard ratio 0.37, p < 0.001) (Abou-Alfa et al., Ann Oncol 2019; NCT02989857). Feasibility of m IDH1 detection in plasma ctDNA from patients with IHC was demonstrated and was highly concordant with mutation status in tumor tissue (Aguado-Fraile et al., Cancer Res 2019). This analysis was extended to the larger patient cohort from ClarIDHy, and longitudinal m IDH1 detection from ctDNA was assessed and correlated with clinical response. Methods: Baseline plasma and tumor tissue samples were obtained before randomization; longitudinal plasma samples were collected on day 1 of each treatment cycle. m IDH1 status in tissue was prospectively and centrally confirmed using Oncomine Focus Assay. A BEAMing digital PCR test was used for quantification of m IDH1 in plasma. IDH1 mutation clearance ( IDH1-MC) was achieved when plasma m IDH1 variant allele frequency was below the assay’s sensitivity (0.02% for R132C/L/S/G; 0.04% for R132H). Results: m IDH1 detection in plasma ctDNA and tissue was concordant in 92% (193/210) of samples screened. As of 31 Jan 2019, median PFS was 2.7 months for IVO vs 1.4 months for placebo. Longitudinal analysis with biomarker data available as of Jan 2020 demonstrated IDH1-MC in plasma from 10 IVO-treated patients with PFS ≥2.7 months (n = 36) vs 0 patients with PFS < 2.7 months (n = 40). No IDH1-MC was observed in patients treated with placebo, irrespective of response (n = 49). Conclusions: These results reinforce the feasibility of IDH1-R132 detection in plasma from patients with IHC, with a 92% concordance with detection in tumor tissue, supporting m IDH1 detection in liquid biopsy as a viable patient selection strategy where tissue exhaustion can limit conventional methods. Plasma IDH1-MC was also observed in a subset of IVO-treated patients who achieved disease control. Clinical trial information: NCT02989857 .

Gray Areas in the Gray Matter: IDH1/2 Mutations in Glioma.

Since the first discovery of isocitrate dehydrogenase (IDH) mutations in cancer, considerable progress has been made in our understanding of their contribution to cancer development. For glioma, this has helped to identify two diagnostic groups of tumors (oligodendroglioma and astrocytoma IDHmt) with distinct clinical characteristics and that are now diagnosed by the presence of the IDH mutations. The metabolic changes occurring as the consequence of the altered substrate affinity of the mutant IDH protein results in a cascade of intracellular changes, also inducing a relative sensitivity to chemotherapy and radiotherapy compared with IDHwt tumors. Pharmacologic blockade of the mutant enzyme with first-in-class inhibitors has been efficacious for the treatment of IDH-mutant acute myeloid leukemia (AML) and is currently being evaluated in phase III trials for IDH-mutant glioma (INDIGO) and cholangiocarcinoma (ClarIDHy). It seems likely that acquired resistance to mutant IDH inhibitors will eventually emerge, and combination therapies to augment the antitumor activity of mutant IDH inhibitors have already been initiated. Approaches to exploit, rather than inhibit, the unique metabolism of IDH-mutant cancer cells have emerged from laboratory studies and are now also being tested in the clinic. Results of these clinical trials are eagerly awaited and will likely provide new key insights and direction of the treatment of IDH-mutant human cancer.

Biochemical and phylogenetic characterization of a monomeric isocitrate dehydrogenase from a marine methanogenic archaeon Methanococcoides methylutens.

Monomeric isocitrate dehydrogenase (IDH) stands for a separated subgroup among IDH protein family. Up to now, all reported monomeric IDHs are from prokaryotes. Here, a monomeric IDH from a marine methanogenic archaeon Methanococcoides methylutens (MmIDH) was reported for the first time. BLAST search demonstrated that only a few marine archaea encode the monomeric IDH and all these organisms are methylotrophic. MmIDH shows the highest homology (~ 70%) to the monomeric IDHs from some marine bacteria, suggesting a lateral gene transfer event between marine bacteria and archaea. The monomeric state of MmIDH was determined by size exclusion chromatography. MmIDH is divalent cation-dependent and Mn2+ is the most favored. Kinetic analysis showed that MmIDH is highly specific to NADP+ and cannot utilize the NAD+. The optimal temperature for MmIDH activity is 50°C and the optimal pH is 8.2. Heat inactivation assay revealed that MmIDH is a mesophilic enzyme. It sustained 50% activity after incubation at 39°C for 20min. Moreover, the putative coenzyme binding residues (His590, Arg601, and Arg650) of MmIDH were explored by mutagenesis. The triple mutant H590L/R601D/R650S displayed a 5.93-fold preference for NAD+ over NADP+, indicating that the coenzyme specificity of MmIDH was significantly switched from NADP+ to NAD+ by three key mutations.

LBA10_PR - ClarIDHy: A global, phase III, randomized, double-blind study of ivosidenib (IVO) vs placebo in patients with advanced cholangiocarcinoma (CC) with an isocitrate dehydrogenase 1 (IDH1) mutation

LBA10_PR - ClarIDHy: A global, phase III, randomized, double-blind study of ivosidenib (IVO) vs placebo in patients with advanced cholangiocarcinoma (CC) with an isocitrate dehydrogenase 1 (IDH1) mutation

2031 - DELINEATING MECHANISMS OF ACQUIRED RESISTANCE TO IDH INHIBITORS IN AML

Approximately 20% of acute myeloid leukaemia (AML) patients carry mutations in genes that encode the metabolic enzymes isocitrate dehydrogenase (IDH) 1 and -2. Mutant IDH proteins possess a neomorphic enzymatic activity and produce the oncometabolite D-2-hydroxyglutarate (2-HG) that modulates numerous pathways implicated in cell fate decisions and cancer. We and others have shown that IDH mutations can contribute to leukaemia initiation and maintenance both in vitro and in vivo. Small molecule inhibitors that block production of 2-HG are starting to enter the clinic, however, the biomarkers of response and mechanisms of de novo and acquired resistance to mutant IDH inhibition remain poorly understood. We have engineered a series of murine AML models driven by inducible expression of mutant IDH proteins. To uncover the molecular mechanisms that underpin response and adaptation to mutant IDH inhibition we integrated bulk and single-cell RNA sequencing with chromatin immuno-precipitation sequencing. We demonstrate that genetic depletion and pharmacological inhibition of mutant IDH has striking effects, altering self-renewal and differentiation programs of leukaemic cells. Importantly, we identify a population of cells that persist during treatment and characterise genetic programs that circumvent effective IDH inhibition, ultimately leading to therapy resistance and disease relapse. Our studies provide detailed mechanistic insight into the oncogenic properties of mutant IDH proteins in maintaining the transformed state of AML cells.