The L2HGDH Knockout NCI-H1975 Polyclonal Cells are a pool of CRISPR/Cas9-edited cells derived from the NCI-H1975 human lung adenocarcinoma cell line, featuring a heterogeneous loss-of-function disruption of the L2HGDH gene. This polyclonal knockout population provides a biologically relevant model for studying the consequences of L2HGDH deficiency without single-cell clonal selection, preserving the genetic heterogeneity inherent to cancer cell populations.
The parental NCI-H1975 cell line was established from a pleural effusion of a female nonsmoker with non-small cell lung adenocarcinoma and harbors two canonical epidermal growth factor receptor (EGFR) mutations: L858R and T790M. These alterations render the cells dependent on EGFR signaling and sensitive to first- and second-generation tyrosine kinase inhibitors, making NCI-H1975 a widely employed model for studying EGFR-mutant lung cancer biology and therapeutic responses.
L2HGDH encodes a mitochondrial flavoprotein that catalyzes the flavin adenine dinucleotide (FAD)-dependent oxidation of the oncometabolite L-2-hydroxyglutarate (L-2HG) to ??-ketoglutarate (??-KG), a critical TCA cycle intermediate. This reaction is essential for preventing the accumulation of L-2HG, which competitively inhibits ??-KG-dependent dioxygenases, including the ten-eleven translocation (TET) family of DNA demethylases (TET1, TET2, TET3) and Jumonji C-domain-containing histone demethylases (e.g., KDM4A, KDM5). The enzyme is transcriptionally upregulated under hypoxic conditions by hypoxia-inducible factor 1-alpha (HIF1A) and at acidic pH, linking its activity to the tumor microenvironment. L2HGDH interacts with electron transfer flavoprotein (ETF) and mitochondrial membrane proteins, positioning it at the interface of cellular metabolism and epigenetic regulation.
In the context of NCI-H1975 cells, disruption of L2HGDH is expected to elevate intracellular L-2HG levels, thereby perturbing the epigenetic landscape through TET-mediated DNA demethylation and JmjC-dependent histone demethylation pathways. Given that EGFR-mutant lung adenocarcinomas exhibit distinct epigenetic dependencies and hypoxia-driven molecular adaptations, this knockout model permits investigation into how oncometabolite-induced epigenetic reprogramming intersects with oncogenic EGFR signaling. The L858R/T790M background also allows assessment of whether L2HGDH loss modulates sensitivity to EGFR tyrosine kinase inhibitors, potentially informing clinical observations of metabolic resistance mechanisms.
Researchers can employ these polyclonal knockout cells to dissect the role of L-2-hydroxyglutarate in DNA and histone methylation dynamics using liquid chromatography-mass spectrometry (LC-MS) for L-2HG quantification, global DNA methylation ELISAs, bisulfite sequencing, and chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR) targeting histone marks like H3K9me3 and H3K27me3. Combined with cell proliferation assays and drug sensitivity profiling with EGFR inhibitors under normoxic and hypoxic conditions, the model supports comprehensive studies of metabolic-epigenetic crosstalk in lung adenocarcinoma. Transcriptomic profiling by RNA-seq further enables discovery of L2HGDH-dependent gene signatures. For technical inquiries and ordering, please contact Ascent Research.
