Abstract
BackgroundIn 2014 the European Medicines Agency included exon 2, 3 and 4 KRAS and NRAS testing for the selection of metastatic colorectal cancer (mCRC) patients eligible for the therapy with anti-EGFR monoclonal antibodies. The Italian Association of Medical Oncology (AIOM) and the Italian Society of Pathology and Cytology (SIAPEC) organized an external quality assessment (EQA) scheme for CRC to evaluate inter-laboratory consistency and to ensure standardization of the results in the transition from KRAS to all-RAS testing.MethodsTen formalin fixed paraffin embedded specimens including KRAS/NRAS (exons 2, 3, 4) and BRAF (codon 600) mutations were validated by three referral laboratories and sent to 88 participant centers. Molecular pathology sample reports were also requested to each laboratory. A board of assessors from AIOM and SIAPEC evaluated the results according to a predefined scoring system. The scheme was composed of two rounds.ResultsIn the first round 36 % of the 88 participants failed, with 23 centers having at least one false positive or false negative while 9 centers did not meet the deadline. The genotyping error rate was higher when Sanger sequencing was employed for testing as compared with pyrosequencing (3 vs 1.3 %; p = 0.01; Pearson Chi Square test). In the second round, the laboratories improved their performance, with 23/32 laboratories passing the round. Overall, 79/88 participants passed the RAS EQA scheme. Standardized Human Genome Variation Society nomenclature was incorrectly used to describe the mutations identified and relevant variations were noticed in the genotype specification.ConclusionThe results of the Italian RAS EQA scheme indicate that the mutational analyses are performed with good quality in many Italian centers, although significant differences in the methods used were highlighted. The relatively high number of centers failing the first round underlines the fundamental role in continued education covered by EQA schemes.
Highlights
In 2014 the European Medicines Agency included exon 2, 3 and 4 KRAS and NRAS testing for the selection of metastatic colorectal cancer patients eligible for the therapy with anti-EGFR monoclonal antibodies
Cetuximab and panitumumab are two monoclonal antibodies that bind the extracellular domain of the EGFR, block its interaction with ligands and inhibit its downstream signalling [1, 2]
One 10-μm-thick slide derived from samples with an adequate content of tumor cells (≥50 %) was selected and analysed by three referral centers that were selected based on their experience in molecular pathology, their track of scientific publications and their expertise in external quality assessment (EQA) organization: the Center of Predictive Molecular Medicine at CeSI Biotech in Chieti, the Department of Human Pathology and Oncology at the University of Florence and the Laboratory of Pharmacogenomics, at Centro di Ricerche Oncologiche di Mercogliano (CROM)—INT “Fondazione Giovanni Pascale” in Mercogliano
Summary
In 2014 the European Medicines Agency included exon 2, 3 and 4 KRAS and NRAS testing for the selection of metastatic colorectal cancer (mCRC) patients eligible for the therapy with anti-EGFR monoclonal antibodies. On the basis of these studies, in 2014 the EMA restricted the use of cetuximab and panitumumab to patients with RAS (exon 2, 3 and 4 of KRAS and NRAS genes) wild-type status. Other genes involved in EGFR downstream pathways, such as BRAF, PIK3CA and PTEN, might be involved in the development of resistance mechanisms to cetuximab and panitumumab in CRC, their role is not completely clarified [10,11,12] Despite this controversial position, the analysis of the mutational status of BRAF is often required by the oncologists, since it is a strong negative prognostic biomarker for mCRC patients and BRAF mutant patients are often treated with aggressive therapeutic regimens
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