Implementation of novel pyrosequencing assays to screen for common mutations of BRAF and KRAS in a cohort of sporadic colorectal cancers.

摘要: Activating mutations of the BRAF and KRAS genes cause constitutive stimulation an important cell-signaling pathway promoting tumorigenesis, are increasingly recognized as determinants response to targeted cancer therapies. The V600E mutation accounts for most in cancer, predominantly encoded by nucleotide substitutions within codons 12 13. We designed novel pyrosequencing assays detection common "hotspot" these genes, which demonstrated analytical sensitivities ≤10% titrations mutant cell lines. assay has ability simultaneously identify all potential changes cluster at 13, with a sequence output sense direction facilitate results interpretation. These were used determine status prospective series 1198 sporadic colorectal cancers. was detected 13.2% frequency our cohort 32.4%, G>A transitions position 2 13 being prevalent. Both proved highly sensitive specific when applied clinical specimens, applicable both fresh-frozen formalin-fixed paraffin-embedded archival tissues. would serve suitable platform large-scale specimens where facility is available.