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Detecting BRAF mutations in formalin-fixed melanoma: experiences with two state-of-the-art techniques


Schoenewolf, Nicola L; Dummer, Reinhard; Mihic-Probst, Daniela; Moch, Holger; Simcock, Mathew; Ochsenbein, Adrian; Gillessen, Silke; Schraml, Peter; von Moos, Roger (2012). Detecting BRAF mutations in formalin-fixed melanoma: experiences with two state-of-the-art techniques. Case Reports in Oncology, 5(2):280-289.

Abstract

BACKGROUND: Melanoma is characterized by a high frequency of BRAF mutations. It is unknown if the BRAF mutation status has any predictive value for therapeutic approaches such as angiogenesis inhibition. PATIENTS AND METHODS: We used 2 methods to analyze the BRAF mutation status in 52 of 62 melanoma patients. Method 1 (mutation-specific real-time PCR) specifically detects the most frequent BRAF mutations, V600E and V600K. Method 2 (denaturing gel gradient electrophoresis and direct sequencing) identifies any mutations affecting exons 11 and 15. RESULTS: Eighteen BRAF mutations and 15 wild-type mutations were identified with both methods. One tumor had a double mutation (GAA) in codon 600. Results of 3 samples were discrepant. Additional mutations (V600M, K601E) were detected using method 2. Sixteen DNA samples were analyzable with either method 1 or method 2. There was a significant association between BRAF V600E mutation and survival. CONCLUSION: Standardized tissue fixation protocols are needed to optimize BRAF mutation analysis in melanoma. For melanoma treatment decisions, the availability of a fast and reliable BRAF V600E screening method may be sufficient. If other BRAF mutations in exons 11 and 15 are found to be of predictive value, a combination of the 2 methods would be useful.

Abstract

BACKGROUND: Melanoma is characterized by a high frequency of BRAF mutations. It is unknown if the BRAF mutation status has any predictive value for therapeutic approaches such as angiogenesis inhibition. PATIENTS AND METHODS: We used 2 methods to analyze the BRAF mutation status in 52 of 62 melanoma patients. Method 1 (mutation-specific real-time PCR) specifically detects the most frequent BRAF mutations, V600E and V600K. Method 2 (denaturing gel gradient electrophoresis and direct sequencing) identifies any mutations affecting exons 11 and 15. RESULTS: Eighteen BRAF mutations and 15 wild-type mutations were identified with both methods. One tumor had a double mutation (GAA) in codon 600. Results of 3 samples were discrepant. Additional mutations (V600M, K601E) were detected using method 2. Sixteen DNA samples were analyzable with either method 1 or method 2. There was a significant association between BRAF V600E mutation and survival. CONCLUSION: Standardized tissue fixation protocols are needed to optimize BRAF mutation analysis in melanoma. For melanoma treatment decisions, the availability of a fast and reliable BRAF V600E screening method may be sufficient. If other BRAF mutations in exons 11 and 15 are found to be of predictive value, a combination of the 2 methods would be useful.

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Additional indexing

Item Type:Journal Article, refereed, original work
Communities & Collections:04 Faculty of Medicine > University Hospital Zurich > Institute of Pathology and Molecular Pathology
04 Faculty of Medicine > University Hospital Zurich > Dermatology Clinic
Dewey Decimal Classification:610 Medicine & health
Uncontrolled Keywords:BRAF mutations; Melanoma; Mutation detection methods; V600E
Language:English
Date:2012
Deposited On:02 Jul 2012 15:40
Last Modified:25 Aug 2017 16:14
Publisher:Karger
ISSN:1662-6575
Free access at:PubMed ID. An embargo period may apply.
Publisher DOI:https://doi.org/10.1159/000339300
PubMed ID:22740817

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