title: The usability of allele-specific PCR and reverse-hybridization assays for KRAS genotyping in Serbian colorectal cancer patients.
abstract: BACKGROUND
  Colorectal cancers (CRCs) with wild-type KRAS respond to EGFR-targeted antibody treatment. Analysis of the hotspot clustered mutations in codons 12 and 13 is compulsory before therapy and no standardized methodology for that purpose has been established so far. Since these mutations may have different biological effects and clinical outcome, reliable frequency and types of KRAS mutations need to be determined for individual therapy.
  
  
  OBJECTIVE
  The purpose of this study was to describe the KRAS mutation spectrum in a group of 481 Serbian mCRC patients and to compare the general performances of allele-specific PCR and reverse-hybridization assays.
  
  
  METHODS
  KRAS testing was performed with two diagnostic analyses, DxS TheraScreen K-RAS PCR Kit and KRAS StripAssay™.
  
  
  RESULTS
  KRAS mutations in codons 12 and 13 were present in 37.6 % of analyzed formalin-fixed paraffin-embedded (FFPE) DNA samples. The seven most frequent mutation types were observed with both assays: p.G12D 34.6 %, p.G12V 24.9 %, p.G12A 10.3 %, p.G12C 8.1 %, p.G12S 5.4 %, p.G12R 1.6 %, and p.G13D 15.1 %. Regarding double mutants, 0.8 % of them were present among all tested samples and 2.2 % among KRAS mutated ones.
  
  
  CONCLUSIONS
  Two screening approaches that were used in this study have been shown as suitable tests for detecting KRAS mutations in diagnostic settings. In addition, they appear to be good alternatives to methods presently in use. In our experience, both methods showed capacity to detect and identify double mutations which may be important for potential further subgrouping of CRC patients.
authors.affiliation: Department of Experimental Oncology, Institute for Oncology and Radiology of Serbia, Pasterova 14, 11000 Belgrade, Serbia. jakovljevicksenija@yahoo.com
authors.forename: Ksenija
authors.initials: K
authors.lastname: Brotto
authors.forename: Emina
authors.initials: E
authors.lastname: Malisic
authors.forename: Milena
authors.initials: M
authors.lastname: Cavic
authors.forename: Ana
authors.initials: A
authors.lastname: Krivokuca
authors.forename: Radmila
authors.initials: R
authors.lastname: Jankovic
mesh_terms: D000483:Alleles; D002277:Carcinoma / Q000235:genetics*; D015179:Colorectal Neoplasms / Q000235:genetics*; D004252:DNA Mutational Analysis; D060005:Genotyping Techniques; D006801:Humans; D016133:Polymerase Chain Reaction; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D016283:Proto-Oncogene Proteins p21(ras); D055771:Serbia; D018631:ras Proteins / Q000235:genetics*
chemical_list: C117307:KRAS protein, human; D011518:Proto-Oncogene Proteins; D016283:Proto-Oncogene Proteins p21(ras); D018631:ras Proteins