The ACADM Knockout HT29 Polyclonal Cells constitute a CRISPR/Cas9-edited polyclonal knockout cell population derived from the HT29 colorectal adenocarcinoma line, engineered to disrupt the ACADM gene and ablate medium-chain acyl-CoA dehydrogenase (MCAD) function. This heterogeneous pool retains genetic diversity characteristic of polyclonal editing, avoiding clonal selection bias and providing a faithful loss-of-function model system. The product is supplied as a viable, proliferating culture suitable for immediate expansion, enabling robust interrogation of mitochondrial fatty acid beta-oxidation and its roles in intestinal epithelial biology and cancer metabolism. Researchers can employ these cells to dissect metabolic vulnerabilities without the confounding effects of single-cell adaptation.
The host HT29 cell line originates from a 44-year-old Caucasian female with colorectal adenocarcinoma, exhibiting adherent epithelial morphology and mucin production. As a widely utilized model in colon cancer research, HT29 cells recapitulate key features of intestinal epithelium, including barrier function, secretory activity, and drug absorption/metabolism. Their well-documented growth characteristics and responsiveness to metabolic perturbations make them an ideal platform for studying how ACADM dysfunction affects colorectal cancer cell energetics, stress adaptation, and pharmacological responses.
ACADM encodes the mitochondrial enzyme catalyzing the initial, rate-limiting step of medium-chain fatty acid beta-oxidation: FAD-dependent dehydrogenation of medium-chain fatty acyl-CoAs to trans-2-enoyl-CoAs. Electrons are then shuttled via electron transfer flavoprotein (ETF) and ETF dehydrogenase to the respiratory chain, ultimately generating acetyl-CoA for the TCA cycle and ATP. The enzyme is transcriptionally activated by PPARA and AMPK signaling downstream of glucagon and fasting stimuli, while insulin suppresses expression. Its activity is critical for ketone body production during catabolic stress, and it interacts directly with ETF, enoyl-CoA hydratase, 3-hydroxyacyl-CoA dehydrogenase, and 3-ketoacyl-CoA thiolase within the beta-oxidation spiral. Knockout disrupts this cascade, leading to accumulation of medium-chain acyl-carnitines, diminished acetyl-CoA and ketone body output, and reduced ATP synthesis under glucose-limiting conditions, paralleling the metabolic crisis observed in medium-chain acyl-CoA dehydrogenase deficiency (MCADD).
In the HT29 colorectal adenocarcinoma context, ACADM knockout creates a powerful tool for exploring the intersection of fatty acid oxidation and cancer metabolism. Colorectal tumors frequently rewire metabolic pathways, and reliance on fatty acid oxidation can represent a therapeutic vulnerability. This polyclonal model enables the study of how loss of MCAD affects cell viability, clonogenicity, and mitochondrial respiration in an intestinal epithelial background. Additionally, HT29 cells endogenously express key regulators like AMPK and PPAR signaling components, allowing dissection of compensatory responses to impaired beta-oxidation under various nutrient regimes. The model thus links metabolic disorder research (MCADD) with cancer biology, providing insights into hypoketotic hypoglycemia mechanisms and potential metabolic targets in colon cancer.
Investigators can deploy these ACADM knockout polyclonal cells in a wide range of experimental workflows. Common applications include phenotypic rescue experiments to validate MCAD function, mitochondrial respiration profiling via Seahorse analysis, and fatty acid oxidation flux measurements using labeled palmitate or octanoate. The cells are suited for viability assays under glucose/glutamine restriction to mimic fasting stress, as well as for acyl-carnitine profiling by mass spectrometry to quantify lipid intermediates. Immunofluorescence, western blotting, and RT-qPCR can confirm knockout at the protein and transcript levels, while flow cytometry enables cell cycle and apoptosis studies. The system also supports drug metabolism screening where fatty acid oxidation influences pharmacological responses, and it serves as a platform for identifying synthetic lethal interactions in colorectal cancer. For additional technical guidance or bulk orders, please contact Ascent Research.
