The HADH Knockout A-549 Polyclonal Cells provide a CRISPR/Cas9-mediated loss-of-function model, targeting the HADH gene in the human A-549 lung adenocarcinoma cell line. Supplied as a polyclonal knockout population, this product consists of a heterogeneous pool of edited cells, capturing a range of knockout efficiencies without single-cell cloning. The use of CRISPR/Cas9 ensures stable gene disruption, and the polyclonal nature preserves genetic diversity, which is advantageous for studying phenotypic heterogeneity and population-level effects in cancer metabolism.
A-549 cells are a human lung adenocarcinoma line with alveolar basal epithelial type II pneumocyte characteristics, originally derived from an explanted tumor. They carry a KRAS G12S activating mutation, one of the most frequent oncogenic drivers in lung adenocarcinoma, resulting in constitutive signaling that promotes proliferation and metabolic reprogramming, including altered glucose and lipid handling. Their adherent growth and well-characterized biology facilitate routine culture, genetic manipulation, and functional assays, making them an ideal host for studying metabolic gene knockouts in an oncogene-driven context.
HADH encodes mitochondrial short-chain 3-hydroxyacyl-CoA dehydrogenase (SCHAD), which catalyzes the NAD+-dependent oxidation of C4-C10 3-hydroxyacyl-CoAs in the third step of mitochondrial fatty acid ??-oxidation. This produces NADH and 3-ketoacyl-CoA, subsequently converted to acetyl-CoA for the TCA cycle and oxidative phosphorylation. HADH expression is controlled by PPAR??, and its activity is influenced by the NAD+/NADH ratio, short-chain acyl-CoA levels, and insulin signaling. HADH directly binds NAD+ and short-chain 3-hydroxyacyl-CoAs, and cooperates with ??-oxidation enzymes CPT1A, ACADS, ECHS1, and ACAA2. The generated NADH and acetyl-CoA sustain respiratory chain activity and anaplerotic reactions.
In A-549 cells, HADH knockout disrupts mitochondrial short-chain ??-oxidation, reducing acetyl-CoA and NADH output. This metabolic impairment likely induces compensatory glycolysis, reminiscent of the Warburg effect, altering energy charge and redox balance, which can impact proliferation, survival, and drug sensitivity. Additionally, HADH??s role in insulin secretion links this model to hyperinsulinemic hypoglycemia, enabling studies of oncogenic and metabolic pathway cross-talk. The HADH-null A-549 line may reveal synthetic lethal vulnerabilities in KRAS-mutant lung adenocarcinoma.
This polyclonal HADH knockout product supports applications in cancer metabolism research, metabolic drug screening, and disease modeling of fatty acid oxidation disorders and hyperinsulinemic hypoglycemia. Validation and functional characterization can be performed using Western blotting, RT-qPCR, Seahorse extracellular flux analysis, fatty acid oxidation assays, ATP luminescence, and LC-MS metabolomics. The polyclonal format captures population-level metabolic responses, making it suitable for both bulk assays and single-cell analysis. For technical support and ordering information, please contact Ascent Research.
