High sensitivity of reverse-hybridization methodology in the detection of KRAS mutations from formalin-fixed paraffin-embedded colorectal cancer samples

Abstract

Colorectal cancer is ranked the third most common cancer worldwide in terms of incidence and the second in terms of mortality. Recent advances in therapeutic approaches to colorectal cancer have identified a potential role of anti-epidermal growth factor receptor (EGFR) targeted therapies as adjuvant treatment in advanced disease. New evidences showed that patients harboring KRAS mutations on codons 12 and 13 are not responsive to anti-EGFR monoclonal antibodies. Therefore, new mutational screening tools have been proposed to select patients who will benefit from anti-EGFR targeted therapy, reducing inappropriate, expensive treatments and unwarranted side effects. We evaluated the performance of a reverse-hybridization-based assay in the identification of the most frequent KRAS mutations on a series of 50 formalin-fixed, paraffin-embedded, advanced colorectal cancer specimens, in comparison with the direct gene sequencing technique. Thirty-two of the 50 cases (64%) showed KRAS single point mutations by reverse-hybridization technique. In particular, 93.8% of the mutations were reported on codon 12, whereas 6.2% of the mutations were reported on codon 13. Direct gene sequencing showed KRAS mutations on 28 of the 50 cases (56%) with 96.4% of the mutations on codon 12 and 3.6% on codon 13. Concordance between the assays was observed in 92% of the cases. Both reverse hybridization and gene sequencing methods have been shown to be suitable tests in detecting KRAS mutations from formalin-fixed, paraffin-embedded tumor specimens. In our experience, reverse-hybridization technique has been shown to be an effective and more sensitive assay for the identification of the most common KRAS mutations.