IDH2 Knockout HAP1 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal population targeting the mitochondrial isocitrate dehydrogenase 2 (IDH2) gene in the near-haploid HAP1 human cell line. This loss-of-function model enables dissection of IDH2-dependent metabolic and redox pathways, providing a robust tool for functional genomics and drug discovery without selection for single clones. The polyclonal nature ensures a representative assessment of gene disruption effects.
The HAP1 cell line, derived from KBM-7 chronic myeloid leukemia cells, maintains a near-haploid karyotype with disomy for chromosome 8. This genetic simplicity reduces functional redundancy, facilitating clean knockout phenotypes. Its leukemic background provides a relevant cancer model for studying hematological malignancies and metabolic adaptations.
IDH2 catalyzes the oxidative decarboxylation of isocitrate to ??-ketoglutarate (??KG), a reaction that generates NADPH. Its activity is allosterically enhanced by citrate and reduced by NADH and ATP; deacetylation by SIRT3 increases enzymatic function. IDH2-generated ??KG is a limiting co-substrate for TET DNA dioxygenases and histone demethylases, linking mitochondrial metabolism to epigenetic regulation. Meanwhile, NADPH serves as a cofactor for glutathione reductase and prolyl hydroxylases, crucial for redox balance and HIF-1?? stabilization. IDH2 forms a homodimer and interacts with mitochondrial chaperones and TCA cycle enzyme complexes. Knockout of IDH2 therefore depletes both ??KG and NADPH pools, impairing antioxidant defenses and ??KG-dependent dioxygenases, with consequences for gene expression and cell survival.
In HAP1 leukemic cells, IDH2 ablation models the metabolic dependencies of IDH2-mutant cancers, where loss of normal IDH2 function can be synthetically lethal or reveal therapeutic targets. The polyclonal knockout population captures heterogeneous editing events, enabling studies of redox vulnerability, metabolic reprogramming, and epigenetic alterations in a near-haploid background that simplifies genotype?Cphenotype correlations.
Key experimental applications include NADPH/NADP+ ratio quantification, LC-MS-based metabolic profiling, Seahorse mitochondrial stress analysis, and ROS detection to evaluate oxidative stress. Epigenetic analyses via ChIP-qPCR for histone methylation marks can delineate ??KG-dependent changes. Apoptosis, proliferation, and drug sensitivity assays support target validation and compound screening workflows. For further technical details, please contact Ascent Research.
