title: Preponderance of the oncogenic V599E and V599K mutations in the B-raf kinase domain is enhanced in melanoma lymph node metastases.
abstract: Downstream of Ras, the serine/threonine kinase B-raf has been reported to be mutated, amongst other carcinomas, in a substantial subset of primary melanomas, with a preponderance of mutations within the kinase domain, including the activating V599E and V599K transitions. We investigated a representative series of 54 resection specimens of melanoma lymph node metastases for the presence of mutations within the activation segment (exon 15) of the B-raf kinase domain by polymerase chain reaction (PCR) and single-strand conformational polymorphism (SSCP) gel electrophoresis. Sequencing of cloned PCR-SSCP amplicons resulted in 24 (44%) samples harbouring somatic mutations, which is not significantly different from the mutation frequency found in recently investigated primary cutaneous melanomas (Deichmann M, Thome M, Benner A, Näher H. B-raf exon 15 mutations are common in primary melanoma resection specimens but not associated with clinical outcome. Oncology 2004; 66:411-419). The activating mutation T1796A was present in 20 (83%) of these resection specimens, followed in frequency by the oncogenic g1795A mutation in five (21%) cases. With regard to the B-raf protein sequence, the acidic amino acid transitions V599E and V599K were predicted in 15 (62%) and five (21%) of the 24 positive metastases, respectively. The detection of mutations at this hot spot codon was significantly associated with subsequent visceral metastasis (P=0.03; Fisher's exact test). During the transition from primary melanomas (see reference above) to lymph node metastases, the spectrum of B-raf mutations narrows significantly (P=0.00047). The oncogenic B-raf mutations V599E and V599K, as early events in melanocyte transformation, persist throughout metastasis with important prognostic implications.
authors.affiliation: Department of Dermatology, Heidelberg University Clinics, Arnsburger Weg 1, 63571 Gelnhausen, Germany.
authors.forename: Martin
authors.initials: M
authors.lastname: Kirschner
authors.forename: Burkhard
authors.initials: B
authors.lastname: Helmke
authors.forename: Hans
authors.initials: H
authors.lastname: Starz
authors.forename: Axel
authors.initials: A
authors.lastname: Benner
authors.forename: Marianne
authors.initials: M
authors.lastname: Thome
authors.forename: Martin
authors.initials: M
authors.lastname: Deichmann
mesh_terms: D005091:Exons; D006801:Humans; D008198:Lymph Nodes / Q000473:pathology; D008207:Lymphatic Metastasis; D008545:Melanoma / Q000201:enzymology / Q000235:genetics* / Q000473:pathology*; D009154:Mutation; D016612:Paraffin Embedding; D016133:Polymerase Chain Reaction; D018807:Polymorphism, Single-Stranded Conformational; D048493:Proto-Oncogene Proteins B-raf / Q000235:genetics*
chemical_list: C482119:BRAF protein, human; D048493:Proto-Oncogene Proteins B-raf