The DNPH1 Knockout HCT 116 Polyclonal Cells product provides a CRISPR/Cas9?edited polyclonal knockout cell population derived from the HCT 116 human colorectal carcinoma cell line. These cells carry a targeted disruption of the DNPH1 gene, resulting in loss of 5?formyluracil/5?hydroxymethyluracil DNA glycosylase function. This heterogeneous polyclonal pool offers a robust loss?of?function model for investigating DNPH1 biology while avoiding clonal selection artifacts.
The parental HCT 116 line is a well?characterized colorectal adenocarcinoma model featuring a KRAS G13D mutation and homozygous MSH2 inactivation, leading to mismatch?repair deficiency and microsatellite instability (MSI?H). These adherent epithelial cells recapitulate key features of aggressive colorectal cancer and are routinely used to study DNA damage responses, genomic instability, and tumor?suppressor pathways in a mismatch?repair?compromised background.
DNPH1 encodes a monofunctional DNA glycosylase that initiates base excision repair (BER) by removing oxidized pyrimidines, specifically 5?formyluracil and 5?hydroxymethyluracil. The enzyme physically interacts with the BER scaffold protein XRCC1 and with PCNA, and functionally cooperates with APE1 and DNA polymerase ?? (POLB). DNPH1 expression is activated by MYC, E2F1, and TP53 under oxidative stress. In the knockout, accumulation of unrepaired oxidative lesions triggers p53?dependent signaling, leading to transcriptional upregulation of CDKN1A (p21), BAX, and GADD45A, resulting in cell cycle arrest and apoptosis.
Within the HCT 116 genetic landscape, DNPH1 knockout exacerbates pre?existing mismatch?repair deficiency and KRAS?driven proliferation, elevating spontaneous and induced genomic instability. This combination sensitizes cells to oxidative DNA?damaging agents and modulates p53?mediated damage responses. The model is particularly suited for dissecting how BER?deficient colorectal cancer cells manage replication stress and for identifying synthetic vulnerabilities exploitable in MSI?H/KRAS?mutant tumors.
These polyclonal DNPH1?knockout cells support diverse experimental applications: comet assays for DNA strand breaks, ??H2AX immunofluorescence for damage foci, western blotting for p53, p21, and BAX, cell viability and annexin V apoptosis assays under oxidative challenge, RT?qPCR profiling of BER genes, BER enzymatic activity measurements, H2O2 sensitivity screening, and RNA?seq transcriptomics. The model is thus valuable for mechanistic studies of DNA repair, functional genomics of base excision repair, and preclinical validation of DNA?repair?targeted therapeutics. For further information, please contact Ascent Research.
