The HEXB Knockout HCT 116 Polyclonal Cells are a CRISPR/Cas9-edited polyclonal knockout cell population derived from the human colorectal carcinoma cell line HCT 116. This product features targeted disruption of the HEXB gene, which encodes the ??-subunit of ??-hexosaminidase, using CRISPR/Cas9-mediated genome editing. The resulting polyclonal population contains a heterogeneous mix of edited alleles, providing a loss-of-function model without isolation of individual clones. This polyclonal format is suitable for experiments where gene knockout effects are analyzed at the population level, offering a robust tool for investigating HEXB-dependent processes in a cancer cell background. Researchers can use these cells to dissect lysosomal sphingolipid degradation pathways and evaluate the consequences of ??-hexosaminidase deficiency in a genetically defined host.
The host cell line HCT 116 is a widely used model of human colorectal carcinoma, characterized by activating mutations in KRAS (G13D), CTNNB1 (??45), and PIK3CA (H1047R), and displays microsatellite instability (MSI) due to MLH1 promoter methylation. This genetic profile makes HCT 116 cells highly relevant for studying epithelial tumorigenesis, Wnt/??-catenin signaling, and MAPK pathway activation. The MSI status further links this line to defective DNA mismatch repair, a hallmark of certain colorectal cancers. In this context, HEXB knockout enables exploration of how lysosomal dysfunction intersects with oncogenic signaling pathways, providing insights into the role of sphingolipid metabolism in tumor progression and therapy response.
HEXB encodes the ??-hexosaminidase subunit that dimerizes with the ??-subunit (HEXA) to form the heterodimeric Hex A enzyme, or homodimerizes to produce Hex B. Both enzymes are essential for the lysosomal degradation of GM2 gangliosides, a step requiring the GM2 activator protein (GM2A) for substrate presentation. The HEXB gene is regulated by transcription factors TFEB and MITF, which are downstream effectors of mTORC1 signaling that control lysosomal biogenesis and autophagy. In the glycosphingolipid metabolic pathway, HEXB functions in concert with HEXA, GM2A, GLB1, and NEU1 to process complex sphingolipids. Disruption of HEXB leads to accumulation of GM2 ganglioside and related substrates, resulting in lysosomal storage pathology reminiscent of Sandhoff disease and triggering secondary effects on lysosomal homeostasis.
In the HCT 116 cellular environment, HEXB knockout creates a unique model to study the interplay between lysosomal storage disorders and colorectal cancer biology. Loss of ??-hexosaminidase activity impairs GM2 ganglioside catabolism, causing substrate accumulation that can disrupt lysosomal function, alter autophagic flux, and affect cellular proliferation and apoptosis. The oncogenic mutations in HCT 116, particularly KRAS G13D and CTNNB1 ??45, provide an instructive background to investigate how lysosomal stress modulates tumorigenic signaling networks. This model facilitates examination of whether HEXB deficiency sensitizes cancer cells to lysosomotropic agents or chemotherapeutics, and allows assessment of potential compensatory mechanisms such as upregulation of alternative glycosphingolipid degradation pathways.
Typical research applications for these polyclonal HEXB knockout cells include characterization of GM2 ganglioside accumulation by immunofluorescence or lipidomic analysis, assessment of lysosomal morphology and pH using LysoTracker staining, and evaluation of lysosomal enzyme activities. Functional assays such as Western blotting for HEXB and downstream markers, and RT-qPCR for transcriptional targets of TFEB and MITF, can be used to confirm knockout and pathway perturbations. Cell proliferation and apoptosis assays offer insights into the phenotypic consequences of HEXB loss in a colorectal cancer context. These cells are also valuable for screening small molecules that modulate lysosomal function or rescue the Sandhoff disease phenotype. For further technical details and ordering information, please contact Ascent Research.
