Duplex Reverse-Hybridization Assay for The Simultaneous Detection of KRAS/BRAF Mutations in FFPE-extracted Genomic DNA from Colorectal Cancer Specimens
نویسندگان
چکیده
We report the performance evaluation of a non-quantitative reverse-hybridization assay (KRAS-BRAF StripAssay) designed for the simultaneous detection of 10 mutations in codons 12 and 13 of the KRAS gene and BRAF mutation V600E. Dilution experiments using DNA from tumor cell lines or from formalin-fixed paraffin-embedded (FFPE) colorectal cancer (CRC) tissue were performed to assess assay sensitivity. Using 50 ng of total DNA (mutant and wild-type), the KRAS-BRAF StripAssay demonstrated a detection limit of 1% mutant sequence in a background of wild-type DNA. With respect to BRAF V600E, the KRAS-BRAF StripAssay was evaluated using 60 FFPE CRC samples previously analyzed by high resolution melting (HRM). Test strip hybridization identified 2/60 (3%) samples to carry the BRAF V600E mutation, and results were in agreement with those obtained by HRM analysis. This work demonstrates the KRAS-BRAF StripAssay to be a robust and sensitive method for the detection of common KRAS/BRAF mutations in genomic DNA isolated from FFPE tissue samples.
منابع مشابه
KRAS and BRAF mutations in Iranian colorectal cancer patients: A systematic review and meta-analysis
Background: Mutations in the EGFR signaling pathway play an important role in the development of colorectal cancer (CRC). Mutations in these genes, like KRAS and BRAF, affect the treatment strategies and associated with poor prognosis and relative resistance to anti-EGFR therapies. Our aim was to conduct a systematic and meta-analysis on all studies that have been conducted on the prevalence of...
متن کاملارزیابی فراوانی جهشهای ژن KRAS در بیماران ایرانی مبتلا به سرطان کولورکتال
Background: Kirsten rat sarcoma (KRAS) gene is a target of genetic alterations which are diagnostic and prognostic biomarkers in patients with metastatic colorectal cancer who are treated with monoclonal anti-EGFR antibodies such as cetuximab and panitumumab. KRAS mutations are seen in 35-42% of patients with colorectal cancer. The high frequency of these mutations in colorectal cancer represen...
متن کاملA multiplex technology platform for the rapid analysis of clinically actionable genetic alterations and validation for BRAF p.V600E detection in 1549 cytologic and histologic specimens.
CONTEXT Current clinicopathologic assessment of malignant neoplastic diseases entails the analysis of specific genetic alterations that provide diagnostic, prognostic, or therapy-determining information. OBJECTIVE To develop and validate a robust molecular method to detect clinically relevant mutations in various tissue types and anatomic pathology specimens. DESIGN Genes of interest were a...
متن کاملSensitive multiplex detection of KRAS codons 12 and 13 mutations in paraffin-embedded tissue specimens.
BACKGROUND Colorectal cancer patients harbouring KRAS mutations in codon 12 or 13 do not benefit from current anti-epidermal growth factor receptor (EGFR) monoclonal antibody therapies. Efficient and robust methods are therefore required for routine clinical testing of KRAS mutation status. AIMS To evaluate a novel multiplex assay for the rapid detection of common KRAS mutations in formalin-f...
متن کاملA Novel Fully Automated Molecular Diagnostic System (AMDS) for Colorectal Cancer Mutation Detection
BACKGROUND KRAS, BRAF and PIK3CA mutations are frequently observed in colorectal cancer (CRC). In particular, KRAS mutations are strong predictors for clinical outcomes of EGFR-targeted treatments such as cetuximab and panitumumab in metastatic colorectal cancer (mCRC). For mutation analysis, the current methods are time-consuming, and not readily available to all oncologists and pathologists. ...
متن کامل