KRAS and BRAF mutation analysis in routine molecular diagnostics: comparison of three testing methods on formalin-fixed, paraffin-embedded tumor-derived DNA.
J Mol Diagn
; 14(3): 247-55, 2012.
Article
em En
| MEDLINE
| ID: mdl-22425762
ABSTRACT
Accurate mutation detection assays are strongly needed for use in routine molecular pathology analyses to aid in the selection of patients with cancer for targeted therapy. The high-resolution melting (HRM) assay is an ideal prescreening tool, and SNaPshot analysis offers a straightforward genotyping system. Our present study was determined to compare these mutation testing methods on formalin-fixed, paraffin-embedded (FFPE) tumor-derived DNA. We compared the performance of HRM, followed by cycle sequencing (HRM-sequencing); multiplex PCR assay, followed by SNaPshot analysis (multiplex mutation assay); and a successor assay using HRM, followed by SNaPshot (HRM-SNaPshot) for mutation analysis of both KRAS (codon 12/13/61) and BRAF (codon 600/601). In a series of 195 FFPE-derived DNA specimens, a high genotypic concordance between HRM-sequencing and multiplex mutation assay was found (κ, 0.98; 95% CI, 0.94 to 1), underlining the potential of a combined HRM-SNaPshot approach. In reconstruction experiments, the analytical sensitivity of HRM-SNaPshot was twofold to fourfold higher than HRM-sequencing and multiplex mutation assay, respectively. In addition, HRM-SNaPshot had a good performance rate (99%) on FFPE tumor-derived DNA, and mutation detection was highly concordant with the predecessor assays (κ for both, 0.98). The occurrence of BRAF and KRAS mutations is mutually exclusive. HRM-SNaPshot is an attractive method for mutation analysis in pathology, given its good performance rate on FFPE-derived DNA, high analytical sensitivity, and prescreening approach.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Análise Mutacional de DNA
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Neoplasias Colorretais
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Genes ras
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Proteínas Proto-Oncogênicas
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Carcinoma Pulmonar de Células não Pequenas
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Proteínas ras
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Técnicas de Diagnóstico Molecular
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Proteínas Proto-Oncogênicas B-raf
Tipo de estudo:
Diagnostic_studies
/
Evaluation_studies
Limite:
Humans
Idioma:
En
Revista:
J Mol Diagn
Assunto da revista:
BIOLOGIA MOLECULAR
Ano de publicação:
2012
Tipo de documento:
Article
País de afiliação:
Holanda