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Mutations in isocitrate dehydrogenase 1 (IDH1) are found in 6% of AML patients. Mutant IDH produces R-2-hydroxyglutarate (R-2HG), which induces histone- and DNA-hypermethylation through the inhibition of epigenetic regulators, thus linking metabolism to tumorigenesis. Here we report the biochemical characterization, in vivo antileukemic effects, structural binding, and molecular mechanism of the inhibitor HMS-101, which inhibits the enzymatic activity of mutant IDH1 (IDH1mut). Treatment of IDH1mut primary AML cells reduced 2-hydroxyglutarate levels (2HG) and induced myeloid differentiation in vitro. Co-crystallization of HMS-101 and mutant IDH1 revealed that HMS-101 binds to the active site of IDH1mut in close proximity to the regulatory segment of the enzyme in contrast to other IDH1 inhibitors. HMS-101 also suppressed 2HG production, induced cellular differentiation and prolonged survival in a syngeneic mutant IDH1 mouse model and a patient-derived human AML xenograft model in vivo. Cells treated with HMS-101 showed a marked upregulation of the differentiation-associated transcription factors CEBPA and PU.1, and a decrease in cell cycle regulator cyclin A2. In addition, the compound attenuated histone hypermethylation. Together, HMS-101 is a unique inhibitor that binds to the active site of IDH1mut directly and is active in IDH1mut preclinical models.

D-2-hydroxyglutaric aciduria (D2HGA) type II is a rare neurometabolic disorder caused by germline gain-of-function mutations in isocitrate dehydrogenase 2 (IDH2), resulting in accumulation of D-2-hydroxyglutarate (D2HG). Patients exhibit a wide spectrum of symptoms including cardiomyopathy, epilepsy, developmental delay and limited life span. Currently, there are no effective therapeutic interventions. We generated a D2HGA type II mouse model by introducing the Idh2R140Q mutation at the native chromosomal locus. Idh2R140Q mice displayed significantly elevated 2HG levels and recapitulated multiple defects seen in patients. AGI-026, a potent, selective inhibitor of the human IDH2R140Q-mutant enzyme, suppressed 2HG production, rescued cardiomyopathy, and provided a survival benefit in Idh2R140Q mice; treatment withdrawal resulted in deterioration of cardiac function. We observed differential expression of multiple genes and metabolites that are associated with cardiomyopathy, which were largely reversed by AGI-026. These findings demonstrate the potential therapeutic benefit of an IDH2R140Q inhibitor in patients with D2HGA type II., Competing Interests: None. Animal rights All institutional and national guidelines for the care and use of laboratory animals were followed. This article does not contain studies with human subjects performed by any of the authors. Funding This work was funded by Agios. Writing assistance was provided by Helen Varley, PhD, CMPP, Excel Scientific Solutions, Horsham, UK and funded by Agios.

Mutations in the metabolic enzymes isocitrate dehydrogenase 1 (IDH1) and 2 (IDH2) are frequently found in glioma, acute myeloid leukemia (AML), melanoma, thyroid cancer, and chondrosarcoma patients. Mutant IDH produces 2-hydroxyglutarate (2HG), which induces histone- and DNA-hypermethylation through inhibition of epigenetic regulators. We investigated the role of mutant IDH1 using the mouse transplantation assay. Mutant IDH1 alone did not transform hematopoietic cells during 5 months of observation. However, mutant IDH1 greatly accelerated onset of myeloproliferative disease-like myeloid leukemia in mice in cooperation with HoxA9 with a mean latency of 83 days compared with cells expressing HoxA9 and wild-type IDH1 or a control vector (167 and 210 days, respectively, P = .001). Mutant IDH1 accelerated cell-cycle transition through repression of cyclin-dependent kinase inhibitors Cdkn2a and Cdkn2b, and activated mitogen-activated protein kinase signaling. By computational screening, we identified an inhibitor of mutant IDH1, which inhibited mutant IDH1 cells and lowered 2HG levels in vitro, and efficiently blocked colony formation of AML cells from IDH1-mutated patients but not of normal CD34(+) bone marrow cells. These data demonstrate that mutant IDH1 has oncogenic activity in vivo and suggest that it is a promising therapeutic target in human AML cells.

Pyridoxine-dependent epilepsy (PDE) is an inborn error of metabolism resulting from antiquitin deficiency. There is marked elevation of α-amino adipic semi-aldehyde (αAASA), piperidine-6-carboxylate (P6C) and pipecolic acid. The diagnosis can be confirmed by identifying the mutation in the ALDH7A1 gene in chromosome 5q3l. An 8-year-old Indian girl presented with severe developmental delay and seizures and was found to have pyridoxine-dependent epilepsy owing to an antiquitin mutation. Genetic evaluation of the parents allowed antenatal diagnosis to be made during the next pregnancy.

The Krebs cycle is of fundamental importance for the generation of the energetic and molecular needs of both prokaryotic and eukaryotic cells. Both enantiomers of metabolite 2-hydroxyglutarate are directly linked to this pivotal biochemical pathway and are found elevated not only in several cancers, but also in different variants of the neurometabolic disease 2-hydroxyglutaric aciduria. Recently we showed that cancer-associated IDH2 germline mutations cause one variant of 2-hydroxyglutaric aciduria. Complementary to these findings, we now report recessive mutations in SLC25A1, the mitochondrial citrate carrier, in 12 out of 12 individuals with combined D-2- and L-2-hydroxyglutaric aciduria. Impaired mitochondrial citrate efflux, demonstrated by stable isotope labeling experiments and the absence of SLC25A1 in fibroblasts harboring certain mutations, suggest that SLC25A1 deficiency is pathogenic. Our results identify defects in SLC25A1 as a cause of combined D-2- and L-2-hydroxyglutaric aciduria., (Copyright © 2013 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.)

We report two siblings with atypical pyridoxine-dependant epilepsy, modest elevation of biomarkers, in which the open reading frame and the splice sites of ALDH7A1 did not show any mutations. Subsequent genetic analysis revealed a deep homozygous intronic mutation in ALDH7A1 resulting in two types of transcripts: the major transcript containing a pseudoexon, and the minor transcript representing the authentic spliced transcript. In future, this mutation may be targeted with antisense-therapy aiming at exclusion of the pseudoexon., (Copyright © 2012 Elsevier Inc. All rights reserved.)

Human induced pluripotent stem cells (iPSCs) have been recently found to harbor genomic alterations. However, the integrity of mitochondrial DNA (mtDNA) within reprogrammed cells has yet to be investigated. mtDNA mutations occur at a high rate and contribute to the pathology of a number of human disorders. Furthermore, the lack of mtDNA integrity may alter cellular bioenergetics and limit efficient differentiation. We demonstrated previously that the derivation of iPSCs is associated with mitochondrial remodeling and a metabolic switch towards glycolysis. Here, we have discovered that alterations of mtDNA can occur upon the induction of pluripotency. Massively parallel pyrosequencing of mtDNA revealed that human iPSCs derived from young healthy donors harbored single base mtDNA mutations (substitutions, insertions, and deletions), both homoplasmic (in all mtDNA molecules) and heteroplasmic (in a fraction of mtDNAs), not present in the parental cells. mtDNA modifications were mostly common variants and not disease related. Moreover, iPSC lines bearing different mtDNA mutational loads maintained a consistent human embryonic stem cell-like reprogramming of energy metabolism. This involved the upregulation of glycolytic enzymes, increased glucose-6-phosphate levels, and the over-expression of pyruvate dehydrogenase kinase 1 protein, which reroutes the bioenergetic flux toward glycolysis. Hence, mtDNA mutations within iPSCs may not necessarily impair the correct establishment of pluripotency and the associated metabolic reprogramming. Nonetheless, the occurrence of pathogenic mtDNA modifications might be an important aspect to monitor when characterizing iPSC lines. Finally, we speculate that this random rearrangement of mtDNA molecules might prove beneficial for the derivation of mutation-free iPSCs from patients with mtDNA disorders., (Copyright © 2011 AlphaMed Press.)

Pyridoxine-dependent seizures (PDS) is a rare autosomal recessive disorder causing intractable seizures in neonates and infants. Patients are typically resistant to conventional anticonvulsants but respond well to the administration of pyridoxine. We report two unrelated patients affected with PDS as a result of alpha-aminoadipic semialdehyde (alpha-AASA) dehydrogenase deficiency caused by pathogenic ALDH7A1/antiquitin mutations. Two of the three reported mutations are novel and result in erroneous splicing, as showed by messenger RNA (mRNA) studies. So far, the vast majority of the patients clinically diagnosed as PDS show alpha-AASA dehydrogenase deficiency, caused by mutations in the ALDH7A1 gene. However, despite the availability of reliable biomarkers, early consideration of a pyridoxine trial is still the most important issue in a child with therapy-resistant seizures.

The recent discovery of two defects (ribose-5-phosphate isomerase deficiency and transaldolase deficiency) in the reversible part of the pentose phosphate pathway (PPP) has stimulated interest in this pathway. In this review we describe the functions of the PPP, its relation to other pathways of carbohydrate metabolism and an overview of the metabolic defects in the reversible part of the PPP.

D-2-hydroxyglutaric aciduria is a neurometabolic disorder with mild and severe phenotypes. Recently, we reported pathogenic mutations in the D-2-hydroxyglutarate dehydrogenase gene as the cause of the severe phenotype of D-2-hydroxyglutaric aciduria in two patients. Here, we report two novel pathogenic mutations in this gene in one patient with a mild presentation and two asymptomatic siblings with D-2-hydroxyglutaric aciduria from two unrelated consanguineous Palestinian families: a splice error (IVS4-2A-->G) and a missense mutation (c.1315A-->G;p.Asn439Asp). Overexpression of this mutant protein showed marked reduction of the enzyme activity.