Differences in the frequencies of K‐ras c12–13 genotypes by gender and pathologic phenotypes in colorectal tumors measured using the allele discrimination method

Abstract

The frequencies of different genotypes of the K-ras oncogene in colorectal cancer (CRC) reveal complex relationships among gender, age, and tumor aggression, however, differences among these studies could also be attributed to a lack of standardization of the detection methods used. We developed the allele discrimination assay, which uses dual-color real-time polymerase chain reaction (qPCR) as a fast K-ras genotyping method, and demonstrated higher sensitivity and specificity than DNA sequencing with formalin-fixed paraffin tissues. The assay detected K-ras mutations among 83 of 204 patients with CRC (40.7%); 20.6% of these mutations were G12D (GAT) mutations, 7.4% were G13D (GAC) and G12V (GTT), and 5.3% were other types. A higher proportion of females was observed overall in tumors with K-ras mutations (60.2%, P = 0.01), codon 12 mutations (63.2%, P = 0.005), and transversions (69.6%, P = 0.02), which reflected the higher prevalence of females among the well- to moderately differentiated tumors (29% in males vs. 53% in females; interaction P = 0.03). The opposite was observed for poorly differentiated tumors (47% in males vs. 35% in females). No significant influence of age was found on the prevalence of K-ras mutation. Males with pathological changes and females with poorly differentiated tumors displayed GAT as a less common genotype compared with most other prevalence studies. In conclusion, allele discrimination, with no additional amplification step, is a fast and reliable genotyping method for detecting K-ras c12-13 mutations. Using this method, we demonstrate differences in the frequencies of K-ras genotypes by gender and pathologic phenotypes of CRC.

Oncogene Mutations in Colorectal Polyps Identified in the Norwegian Colorectal Cancer Prevention (NORCCAP) Screening Study

“…39 This complies with observations of higher KRAS mutation frequencies in CRC from female than in male patients. 13,43 Some studies, however, find the higher KRAS frequency in females only relevant in younger individuals or limited to CRC located in the distal colon or rectum. 39,[44][45][46] These factors are consistent with our data as both young age in terms of CRC risk and dominance of distally located polyps are present.…”

Molecular Biomarkers for the Evaluation of Colorectal Cancer

“…This recommendation is supported by 15 studies, [128][129][130][131][132][133][134][135][136][137][138][139][140][141][142] comprising eight prospective cohort studies [130][131][132][133][136][137][138]142 and seven retrospective cohort studies. 128,129,134,135,[139][140][141] For the eight prospective cohort studies, [130][131][132][133][136][137][138]142 all reported balance between the treatment and assessment groups, as all but one 132 used a single cohort design allowing for within-group comparisons. Only this single study, reported by Tol et al, 132 would have required making adjustments for imbalances between the treatment and assessment groups, but none were needed.…”

“…Only this single study, reported by Tol et al, 132 would have required making adjustments for imbalances between the treatment and assessment groups, but none were needed. Five studies 130,133,[136][137][138] reported nonindustry funding, one 132 reported at least partial industry funding, one 142 reported industry funding, and one 131 did not disclose the source of funding, if any. Seven 130,131,133,[136][137][138]142 were deemed to have a low risk of bias, and one 132 was deemed to have a low to moderate risk of bias.…”

“…Five studies 130,133,[136][137][138] reported nonindustry funding, one 132 reported at least partial industry funding, one 142 reported industry funding, and one 131 did not disclose the source of funding, if any. Seven 130,131,133,[136][137][138]142 were deemed to have a low risk of bias, and one 132 was deemed to have a low to moderate risk of bias.…”

“…Due to intratumoral heterogeneity, subclones, and the nature of tissue sampling, clinical trials have used 5% as the lower LOD, and for clinical purposes, it is recommended that the lower LOD for a mutant allele be at least 5%. 12 Therefore, the utilization of methods such as PCR, HRM, single-strand conformation polymorphism, pyrosequencing, or commercially available kits that achieve this level of sensitivity is recommended 130,137,138,142 (Table 13).…”