SW480 (CCL-288, ATCC), SW620 (CCL-227, ATCC) and LIM1863 cells175 (Ludwig Institute for Cancer Research, Melbourne) cells were cultured in RPMI-1640 (Life Technologies). MDA MB 231 (HTB-26, ATCC) was cultured in Dulbecco’s modified Eagle medium (Life Technologies). Primary human cell sources...

Objective(s): Colorectal cancer (CRC) is a major cause of mortality, and KRAS mutations drive tumorigenesis and therapy resistance. Although CRISPR-Cas9 enables precise genome editing, intracellular delivery of Cas9 ribonucleoproteins (RNPs) remains challenging. We evaluated direct electroporation of Cas9-sgRNA RNPs targeting KRAS p.G12V in SW480 CRC cells to assess delivery efficiency, on-target editing, off-target activity, and viability effects. Materials and Methods: An sgRNA targeting KRAS p.G12V was designed computationally. Cas9-sgRNA RNPs were assembled and validated by in vitro digestion. RNPs were electroporated into SW480 cells using a square-wave electroporation protocol; pmaxGFP served as the delivery control. On-target editing and five predicted off-target loci were assessed by Sanger sequencing and TIDE analysis. Cell viability was measured by the MTT assay.Results: Electroporation resulted in high GFP expression (~97%). TIDE analysis at the KRAS locus estimated ~27% tot

Journal of Proteome ResearchDOI: 10.1021/acs.jproteome.3c00833

Colorectal cancer remains one of the leading causes of cancer-related mortality worldwide, highlighting the need for novel therapeutic agents with multitarget anticancer activity. Probucol is a pheno...