IDH2 mutation-induced histone and DNA hypermethylation is progressively reversed by small-molecule inhibition

Abstract

Key Points• IDH2 R140Q expression in TF-1 cells can induce DNA and histone hypermethylation that mirrors human IDH2 mutant acute myeloid leukemia.• The hypermethylation can be reversed on treatment with AGI-6780, an IDH2 mutantspecific small-molecule inhibitor.Mutations of IDH1 and IDH2, which produce the oncometabolite 2-hydroxyglutarate (2HG), have been identified in several tumors, including acute myeloid leukemia. Recent studies have shown that expression of the IDH mutant enzymes results in high levels of 2HG and a block in cellular differentiation that can be reversed with IDH mutant-specific smallmolecule inhibitors. To further understand the role of IDH mutations in cancer, we conducted mechanistic studies in the TF-1 IDH2 R140Q erythroleukemia model system and found that IDH2 mutant expression caused both histone and genomic DNA methylation changes that can be reversed when IDH2 mutant activity is inhibited. Specifically, histone hypermethylation is rapidly reversed within days, whereas reversal of DNA hypermethylation proceeds in a progressive manner over the course of weeks. We identified several gene signatures implicated in tumorigenesis of leukemia and lymphoma, indicating a selective modulation of relevant cancer genes by IDH mutations. As methylation of DNA and histones is closely linked to mRNA expression and differentiation, these results indicate that IDH2 mutant inhibition may function as a cancer therapy via histone and DNA demethylation at genes involved in differentiation and tumorigenesis. (Blood. 2015;125(2):296-303) IntroductionActive site mutations in IDH1 (R132) and IDH2 (R172 and R140) that produce high levels of 2-hydroxyglutarate (2HG) have been identified in several human cancers.1-3 IDH mutations have been shown to cause DNA hypermethylation in both gliomas and leukemias via inhibition of methylcytosine dioxygenase TET2.4,5 Mutant IDH can also promote histone hypermethylation through competitive inhibition of a-ketoglutarate (aKG)-dependent Jumonji-C histone demethylases, thereby activating or deactivating expression of associated genes. 4,6,7 We have shown that mutant IDH1 and IDH2 can affect cell differentiation in solid and liquid tumors. [8][9][10] An IDH1 R132H inhibitor, AGI-5198, delayed growth and promoted differentiation of glioma cells while reducing histone H3K9 trimethylation. 8 Leukemic cell differentiation was also induced in primary human patient samples harboring an IDH2 R140Q mutation when they were treated ex vivo with AGI-6780, an IDH2 R140Q allosteric inhibitor.9 However, the mechanism by which IDH2 mutant activity and 2HG levels contribute to cellular differentiation and tumorigenesis is not fully understood. High levels of 2HG have been shown to competitively inhibit aKGdependent dioxygenases, leading to broad epigenetic changes. Therefore, we sought to investigate the global and gene-specific effects of mutant IDH2 inhibition in TF-1 cells expressing IDH2 R140Q. Probing the effects of IDH2 R140Q expression on histone and DNA methylation and gene expres...

AG-221, a First-in-Class Therapy Targeting Acute Myeloid Leukemia Harboring Oncogenic IDH2 Mutations

“…IDH1/2 mutations can be identified in approximately 20% of patients with AML and approximately 5% of patients with MDS (37,38), and can contribute to leukemia via a block in hematopoietic cell differentiation (16,17,23,24,39). We have discovered AG-221, an oral, selective, first-in-class inhibitor of the mutant IDH2 enzyme.…”

Mutant IDH : A Targetable Driver of Leukemic Phenotypes Linking Metabolism, Epigenetics and Transcriptional Regulation

“…Expression of IDH2 R140Q in primary murine bone marrow cells resulted in inhibition of myeloid differentiation and an increase in the proportion of immature myeloid progenitor cells [25]. Concordantly, expression of IDH2 R140Q or IDH1 R132H in the cytokinedependent TF-1 human erythroleukemia cell line induced cytokine independent growth and a block in cell differentiation [45,62,66]. In the IDH1 R132H model the phenotype required several passages to manifest, was inducible upon exposure to a cell permeable 2-HG and was reversible upon 2-HG withdrawal.…”

“…These were shown to reverse putative IDHm transforming effects, and promote cellular differentiation in leukemia [45,66] and glioma [73] model systems. The IDH1m inhibitor AGI-5198 was able to significantly decrease intracellular 2-HG in glioblastoma cells [73] and intra-and extra-cellular 2-HG levels in in vitro models of chondrosarcoma [74].…”

“…In vitro enzymatic and cell-based assays established that 2-HG inhibits the function of KDM7A (Histone [H] 3 lysine [K] 9 and H3K27 demethylase), KDM2A (H3K36 demethylase) and KDM5B (H3K4 demethylase) (Figure 1) [40,44]. While AML patient samples have not been interrogated extensively for the effects of IDH1 and IDH2 mutations on histone methylation patterning, IDH2m expression in the cytokine-dependent human erythroleukemia TF-1 cell line, was associated with an increase in H3K4, H3K9, H3K27 and H3K36 methylation [45]. This is in agreement with reports of IDH1m and IDH2m expression in other cell based systems [46,47] and suggests that histone methylation patterning may also be contributing to gene expression aberrations in IDH1/2 mutant AMLs.…”

Oncometabolite Tinkers with Genome Folding, Boosting Oncogene Expression

“…Alternatively, pharmacological inhibition of IDH, or selective inhibition of the abnormal enzymatic activity could be used to specifically target tumour cells [1]. IDH inhibitors can revert the impact of IDH mutations on chromatin state [6]. Whether this includes correction of CTCF binding, genome folding and gene expression is an interesting question that remains to be explored.…”