title: KRAS, NRAS and BRAF mutations in Greek and Romanian patients with colorectal cancer: a cohort study.
abstract: OBJECTIVE
  Treatment decision-making in colorectal cancer is often guided by tumour tissue molecular analysis. The aim of this study was the development and validation of a high-resolution melting (HRM) method for the detection of KRAS, NRAS and BRAF mutations in Greek and Romanian patients with colorectal cancer and determination of the frequency of these mutations in the respective populations.
  
  
  METHODS
  Diagnostic molecular laboratory located in Athens, Greece.
  
  
  METHODS
  2425 patients with colorectal cancer participated in the study.
  
  
  METHODS
  2071 patients with colorectal cancer (1699 of Greek and 372 of Romanian origin) were analysed for KRAS exon 2 mutations. In addition, 354 tumours from consecutive patients (196 Greek and 161 Romanian) were subjected to full KRAS (exons 2, 3 and 4), NRAS (exons 2, 3 and 4) and BRAF (exon 15) analysis. KRAS, NRAS and BRAF mutation detection was performed by a newly designed HRM analysis protocol, followed by Sanger sequencing.
  
  
  RESULTS
  KRAS exon 2 mutations (codons 12/13) were detected in 702 of the 1699 Greek patients with colorectal carcinoma analysed (41.3%) and in 39.2% (146/372) of the Romanian patients. Among the 354 patients who were subjected to full KRAS, NRAS and BRAF analysis, 40.96% had KRAS exon 2 mutations (codons 12/13). Among the KRAS exon 2 wild-type patients 15.31% harboured additional RAS mutations and 12.44% BRAF mutations. The newly designed HRM method used showed a higher sensitivity compared with the sequencing method.
  
  
  CONCLUSIONS
  The HRM method developed was shown to be a reliable method for KRAS, NRAS and BRAF mutation detection. Furthermore, no difference in the mutation frequency of KRAS, NRAS and BRAF was observed between Greek and Romanian patients with colorectal cancer.
authors.affiliation: University of Medicine and Pharmacy of Timisoara, Timisoara, Romania.
authors.forename: Serban
authors.initials: S
authors.lastname: Negru
authors.affiliation: Department of Molecular Biology, GENEKOR, Athens, Greece.
authors.forename: Eirini
authors.initials: E
authors.lastname: Papadopoulou
authors.affiliation: Department of Molecular Biology, GENEKOR, Athens, Greece.
authors.forename: Angela
authors.initials: A
authors.lastname: Apessos
authors.affiliation: Institute of Oncology, Bucharest, Romania.
authors.forename: Dana Lucia
authors.initials: DL
authors.lastname: Stanculeanu
authors.affiliation: Institute of Oncology Ion Chiricuta, Cluj-Napoca, Romania.
authors.forename: Eliade
authors.initials: E
authors.lastname: Ciuleanu
authors.affiliation: Centrul de Oncologie Medicala, Iasi, Romania.
authors.forename: Constantin
authors.initials: C
authors.lastname: Volovat
authors.affiliation: Department of Medical Oncology, Fundeni Clinical Institute Bucharest, Bucharest, Romania.
authors.forename: Adina
authors.initials: A
authors.lastname: Croitoru
authors.affiliation: Department of Medical Oncology, University General Hospital of Alexandroupolis, Thrace, Greece.
authors.forename: Stylianos
authors.initials: S
authors.lastname: Kakolyris
authors.affiliation: Second Department of Medical Oncology, "Agii Anargiri" Cancer Hospital, Athens, Greece.
authors.forename: Gerasimos
authors.initials: G
authors.lastname: Aravantinos
authors.affiliation: First Department of Medical Oncology, 'METAXA' Anticancer Hospital of Athens, Athens, Greece.
authors.forename: Nikolaos
authors.initials: N
authors.lastname: Ziras
authors.affiliation: Department of Medical Oncology, Mitera Hospital, Athens, Greece.
authors.forename: Elias
authors.initials: E
authors.lastname: Athanasiadis
authors.affiliation: Department of Medical Oncology, Interbalkan Medical Center, Thessaloniki, Greece.
authors.forename: Nikolaos
authors.initials: N
authors.lastname: Touroutoglou
authors.affiliation: Department of Medical Oncology, University of Ioannina School of Medicine, Ioannina, Greece.
authors.forename: Nikolaos
authors.initials: N
authors.lastname: Pavlidis
authors.affiliation: Clinical Oncology Laboratory, Division of Oncology, Department of Medicine, University of Patras, Patras Medical School, Rio, Greece.
authors.forename: Haralabos P
authors.initials: HP
authors.lastname: Kalofonos
authors.affiliation: Department of Molecular Biology, GENEKOR, Athens, Greece.
authors.forename: George
authors.initials: G
authors.lastname: Nasioulas
mesh_terms: D015331:Cohort Studies; D015179:Colorectal Neoplasms / Q000235:genetics*; D020558:GTP Phosphohydrolases / Q000235:genetics*; D006115:Greece; D006801:Humans; D008565:Membrane Proteins / Q000235:genetics*; D025202:Molecular Diagnostic Techniques / Q000379:methods; D009154:Mutation; D009691:Nucleic Acid Denaturation; D048493:Proto-Oncogene Proteins B-raf / Q000235:genetics*; D016283:Proto-Oncogene Proteins p21(ras) / Q000235:genetics*; D012383:Romania
chemical_list: C117307:KRAS protein, human; D008565:Membrane Proteins; C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf; D020558:GTP Phosphohydrolases; C579846:NRAS protein, human; D016283:Proto-Oncogene Proteins p21(ras)
keywords: Molecular Biology