The EHHADH Knockout NCI-H1299 Polyclonal Cells comprise a CRISPR/Cas9-edited polyclonal knockout cell population designed to enable loss-of-function studies of the EHHADH gene within a human non-small cell lung cancer (NSCLC) context. This product consists of a heterogeneous pool of NCI-H1299 cells harboring CRISPR/Cas9-mediated disruptions at the EHHADH locus, providing a versatile model system for dissecting peroxisomal beta-oxidation and its interplay with cancer cell metabolism. The polyclonal format preserves population-level genetic diversity while allowing robust interrogation of EHHADH-dependent phenotypes, making it suitable for both pooled screening approaches and bulk biochemical analyses.
The host cell line, NCI-H1299, is an adherent epithelial cell line originally derived from a lymph node metastasis of a lung adenocarcinoma. These cells are widely employed as a model system for NSCLC research, particularly due to their defined genetic background: they are TP53 null and carry wild-type EGFR and KRAS alleles. The absence of common driver mutations in EGFR and KRAS makes NCI-H1299 an informative platform for studying alternative oncogenic pathways and metabolic adaptations in lung cancer, including those potentially linked to peroxisomal function.
EHHADH encodes the L-bifunctional protein, a central peroxisomal enzyme that catalyzes the second (enoyl-CoA hydratase) and third (3-hydroxyacyl-CoA dehydrogenase) steps of the fatty acid beta-oxidation spiral. This enzyme acts on medium- and long-chain fatty acyl-CoA substrates, converting trans-2-enoyl-CoA to L-3-hydroxyacyl-CoA and subsequently to 3-ketoacyl-CoA in an NAD+-dependent manner. EHHADH function is regulated upstream by the peroxisome proliferator-activated receptor alpha (PPARA) and its fatty acid ligands, and it works within a multi-enzyme complex that includes ACOX1 and ACAA1, with peroxisomal import mediated by PEX5 and PEX7 receptors. The 3-ketoacyl-CoA product is further processed by the thiolase ACAA1, highlighting EHHADH’s pivotal role in peroxisomal fatty acid degradation.
In the context of NCI-H1299 lung adenocarcinoma cells, EHHADH disruption provides a valuable tool for investigating the contribution of peroxisomal fatty acid oxidation to NSCLC metabolism. Recent evidence suggests that peroxisomal beta-oxidation may support cancer cell proliferation and survival under nutrient stress, with potential links to drug sensitivity and resistance. The TP53-null background of NCI-H1299 further allows exploration of p53-independent metabolic vulnerabilities that may be unmasked upon EHHADH loss. By eliminating EHHADH function, researchers can assess the impact on cellular energetics, very long-chain fatty acid accumulation, and peroxisome-mitochondria crosstalk in a well-characterized lung cancer model.
This polyclonal knockout population supports diverse experimental workflows, such as fatty acid oxidation assays with radiolabeled palmitate, very long-chain fatty acid metabolomics, and peroxisomal enzyme activity measurements. Immunoblotting and RT-qPCR confirm EHHADH disruption, while cell proliferation and drug sensitivity assays enable functional phenotyping. Co-culture and xenograft studies can further probe peroxisomal metabolism in tumor growth and therapy response. For additional details, please contact Ascent Research.
