Alterations in key-regulator genes of disease pathogenesis (BRAF, cKIT, CyclinD1) have been evaluated in patients with multiple primary melanoma (MPM).,One hundred twelve MPM patients (96 cases with two primary melanomas, 15 with three, and 1 with four) were included into the study.,Paired synchronous/asynchronous MPM tissues (N = 229) were analyzed for BRAF mutations and cKIT/CyclynD1 gene amplifications.,BRAF mutations were identified in 109/229 (48%) primary melanomas, whereas cKIT and CyclinD1 amplifications were observed in 10/216 (5%) and 29/214 (14%) tumor tissues, respectively.,While frequency rates of BRAF mutations were quite identical across the different MPM lesions, a significant increase of cKIT (p < 0.001) and CyclinD1 (p = 0.002) amplification rates was observed between first and subsequent primary melanomas.,Among the 107 patients with paired melanoma samples, 53 (49.5%) presented consistent alteration patterns between first and subsequent primary tumors.,About one third (40/122; 32.8%) of subsequent melanomas presented a discrepant pattern of BRAF mutations as compared to incident primary tumors.,The low consistency in somatic mutation patterns among MPM lesions from same patients provides further evidence that melanomagenesis is heterogeneous and different cell types may be involved.,This may have implications in clinical practice due to the difficulties in molecularly classifying patients with discrepant primary melanomas.
Oncogenic BRAF mutation had been considered to be a founder event in the formation of melanocytic tumours; however, we recently argued against this notion by showing marked polyclonality of BRAF mutations in acquired melanocytic nevi (Lin et al, J Natl Cancer Inst., 2009; 101:1423-7).,Here, we tested whether similar heterogeneity of BRAF mutations exists in primary melanomas.,We isolated and sequenced single melanoma cells from five primary melanoma tissues using antibodies against human high-molecular-weight melanoma-associated antigen.,We also examined 10 primary melanomas by the sensitive Mutector assay detecting the BRAFV600E mutation, as well as by cloning and sequencing of separated alleles.,Furthermore, we estimated the frequency of BRAF mutant alleles in paired samples of primary tumour and recurrence or metastasis in three patients.,Single-cell mutation analyses revealed that four of five primary melanomas contained both BRAF-wild-type and BRAF-mutant tumour cells.,Tumour heterogeneity in terms of BRAF mutations was also shown in 8 of 10 primary melanomas.,Selection of BRAF mutant alleles during progression was demonstrated in all the three patients.,Acquisition of a BRAF mutation is not a founder event, but may be one of the multiple clonal events in melanoma development, which is selected for during the progression.