Giovanna De Maglio et al., The pyrosequencing experience: testing colorectal cancer patients for KRAS status in the clinical practice

Testing KRAS status in advanced colorectal cancers has gained a practice-changing level of evidence, being predictive of resistance to the point of rendering the use of EGFR-inhibitors useless in patients carrying KRAS mutations. Since there is noagreement on the best testing method, different assays are commonly used in the clinical practice to determinate KRAS status. Among others, real-time PCR, direct sequencing analysis, and PCR-RFLP are the most widely used in Italy, while <10% of laboratories use pyrosequencing technology, a nucleotide extension based sequencing technique. Most of KRAS mutation tests are laboratory based. Limited information has been published on the application of CE-marked commercial kits with pyrosequencing.
Aims of our study were to verify the concordance of pyrosequencing technique with a CE-marked kit for clinical use with literature data and to validate the efficiency of this methodology in a consecutive series of 122 advanced CRC patients.

Pyrosequencing is a promising, highly sensitive method, and the widespread availability of CE-marked kits to test KRAS and BRAF status would improve their implementation in routine practice. Data obtained is robust and confirms frequencies and pattern of mutations reported in the literature. Notably, the possibility to accurately determine KRAS status even in limited tissue samples obtained from small biopsies is key in the clinical practice. The slightly higher reported rate of KRAS mutations may reflect the high sensibility of the method.

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