The AAK1 Knockout HCT 116 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population with targeted disruption of the AAK1 gene in HCT 116 colorectal carcinoma cells. This heterogeneous pool maintains polyclonal variation, avoiding clonal selection artifacts, and serves as a versatile loss-of-function model for investigating AAK1-dependent processes. No specific editing mechanism or clonal genotype is claimed. The product is supplied as a live cell population, ready for immediate experimental use.
The host cell line, HCT 116, is an adherent human colorectal carcinoma epithelial cell line derived from colorectal adenocarcinoma. It exhibits microsatellite instability (MSI-H), harbors an activating KRAS G13D mutation, and expresses wild-type p53. These molecular features establish it as a clinically relevant model for colorectal cancer studies, enabling detailed analysis of signaling pathways and therapeutic responses.
AAK1 encodes a serine/threonine kinase that phosphorylates the AP2M1 subunit of the AP2 complex, a critical event in clathrin-mediated endocytosis. It is activated by upstream signals including EGF receptor stimulation, Notch ligands (DLL4, JAG1), and Wnt3a, promoting internalization of cargo such as EGFR and components of Notch and Wnt receptor complexes. This kinase thereby modulates Notch intracellular domain (NICD) levels and beta-catenin stabilization, with downstream effects on TCF/LEF-dependent transcription. AAK1 interacts with all AP2 complex subunits, clathrin heavy chain, and the adaptor NUMB, forming a regulatory hub that couples extracellular cues to endocytosis and subsequent signal transduction.
Disrupting AAK1 in the HCT 116 background is particularly instructive for studying endocytic regulation in KRAS-mutant, MSI-H colorectal cancer. KRAS-driven tumors frequently show altered endocytosis, which can sustain EGFR pathway activity and contribute to therapeutic resistance. The polyclonal knockout format mimics tumor heterogeneity, providing a more physiologically relevant system to evaluate how AAK1 loss impacts oncogenic signaling and drug sensitivity.
This model supports a variety of experimental applications: assessment of AP2M1 phosphorylation by western blotting, visualization of clathrin-coated pits via immunofluorescence, quantitative EGFR internalization assays, and Notch pathway luciferase reporter assays. It is also suitable for colony formation assays and drug sensitivity testing, notably with EGFR inhibitors such as cetuximab. By enabling precise manipulation of the endocytic machinery, these cells provide a powerful tool for colorectal cancer research and beyond. For further information, please contact Ascent Research.
