Sensitive High-Resolution Melting Analysis for Screening of KRAS and BRAF Mutations in Iranian Human Metastatic Colorectal Cancers

Abstract

<br /> <strong><span style="font-size: small;">Background: </span></strong><span style="font-family: Times New Roman,Times New Roman; font-size: small;"><span style="font-family: Times New Roman,Times New Roman; font-size: small;">Investigations of methods for detection of mutations have uncovered major weaknesses of direct sequencing and pyrosequencing, with their high costs and low sensitivity in screening for both known and unknown </span></span><span style="font-family: Times New Roman,Times New Roman; font-size: small;"><span style="font-family: Times New Roman,Times New Roman; font-size: small;">mutations. High resolution melting (HRM) analysis is an alternative tool for the rapid detection of mutations. Here we </span></span><span style="font-family: Times New Roman,Times New Roman; font-size: small;"><span style="font-family: Times New Roman,Times New Roman; font-size: small;">describe the accuracy of HRM in screening for KRAS and BRAF mutations in metastatic colorectal cancer (mCRCs) </span></span><span style="font-family: Times New Roman,Times New Roman; font-size: small;"><span style="font-family: Times New Roman,Times New Roman; font-size: small;">samples. </span></span><strong><span style="font-size: small;">Materials and Methods: </span></strong><span style="font-family: Times New Roman,Times New Roman; font-size: small;"><span style="font-family: Times New Roman,Times New Roman; font-size: small;">A total of 1000 mCRC patients in Mehr Hospital, Tehran, Iran, from Feb 2008 to </span></span><span style="font-family: Times New Roman,Times New Roman; font-size: small;"><span style="font-family: Times New Roman,Times New Roman; font-size: small;">May 2012 were examined for KRAS mutations and 242 of them were selected for further assessment of BRAF mutations </span></span><span style="font-family: Times New Roman,Times New Roman; font-size: small;"><span style="font-family: Times New Roman,Times New Roman; font-size: small;">by HRM analysis. In order to calculate the sensitivity and specificity, HRM results were checked by pyrosequencing </span></span><span style="font-family: Times New Roman,Times New Roman; font-size: small;"><span style="font-family: Times New Roman,Times New Roman; font-size: small;">as the golden standard and Dxs Therascreen as a further method. </span></span><strong><span style="font-size: small;">Results: </span></strong><span style="font-family: Times New Roman,Times New Roman; font-size: small;"><span style="font-family: Times New Roman,Times New Roman; font-size: small;">In the total of 1,000 participants, there were 664 (66.4%) with wild type and 336 (33.6%) with mutant codons 12 and/or 13 of the KRAS gene. Among 242 </span></span><span style="font-family: Times New Roman,Times New Roman; font-size: small;"><span style="font-family: Times New Roman,Times New Roman; font-size: small;">samples randomly checked for the BRAF gene, all were wild type by HRM. Pyrosequencing and Dxs Therascreen results were in line with those of the HRM. In this regard, the sensitivity and specificity of HRM were evaluated as </span></span><span style="font-family: Times New Roman,Times New Roman; font-size: small;"><span style="font-family: Times New Roman,Times New Roman; font-size: small;">100%. </span></span><strong><span style="font-size: small;">Conclusion: </span></strong><span style="font-family: Times New Roman,Times New Roman; font-size: small;"><span style="font-family: Times New Roman,Times New Roman; font-size: small;">The findings suggest that the HRM, in comparison with DNA sequencing, is a more appropriate </span></span><span style="font-family: Times New Roman,Times New Roman; font-size: small;"><span style="font-family: Times New Roman,Times New Roman; font-size: small;">method for precise scanning of KRAS and BRAF mutations. It is also possible to state that HRM may be an attractive </span></span><span style="font-family: Times New Roman,Times New Roman; font-size: small;"><span style="font-family: Times New Roman,Times New Roman; font-size: small;">technique for the detection of known or unknown somatic mutations in other genes. </span></span>