DNA Repair and Immune Response Pathways Are Deregulated in Melanocyte-Keratinocyte Co-cultures Derived From the Healthy Skin of Familial Melanoma Patients.

Familial melanoma accounts for 10% of cases, being CDKN2A the main high-risk gene. However, the mechanisms underlying melanomagenesis in these cases remain poorly understood. Our aim was to analyze the transcriptome of melanocyte-keratinocyte co-cultures derived from healthy skin from familial melanoma patients vs. controls, to unveil pathways involved in melanoma development in at-risk individuals. Accordingly, primary melanocyte-keratinocyte co-cultures were established from the healthy skin biopsies of 16 unrelated familial melanoma patients (8 CDKN2A mutant, 8 CDKN2A wild-type) and 7 healthy controls. Whole transcriptome was captured using the SurePrint G3 Human Microarray. Transcriptome analyses included: differential gene expression, functional enrichment, and protein-protein interaction (PPI) networks. We identified a gene profile associated with familial melanoma independently of CDKN2A germline status. Functional enrichment analysis of this profile showed a downregulation of pathways related to DNA repair and immune response in familial melanoma (P < 0.05). In addition, the PPI network analysis revealed a network that consisted of double-stranded DNA repair genes (including BRCA1, BRCA2, BRIP1, and FANCA), immune response genes, and regulation of chromosome segregation. The hub gene was BRCA1. In conclusion, the constitutive deregulation of BRCA1 pathway genes and the immune response in healthy skin could be a mechanism related to melanoma risk.

Patterns of distribution of giant congenital melanocytic nevi (GCMN): The 6B rule.

BACKGROUND: Garment-related terms have been used to describe the pattern of distribution of giant congenital melanocytic nevi (GCMN). OBJECTIVE: We sought to describe patterns of distribution of GCMN and propose a classification scheme. METHODS: Photographic records of patients with GCMN from the Hospital Clinic of Barcelona were analyzed and a classification based on observed GCMN distribution patterns was created. The classification was independently applied by 8 observers to cases found in the literature. The interobserver agreement was assessed. RESULTS: Among 22 patients we observed 6 repeatable patterns of distribution of GCMN, which we termed the "6B": bolero (involving the upper aspect of the back, including the neck), back (on the back, without involvement of the buttocks or shoulders), bathing trunk (involving the genital region and buttocks), breast/belly (isolated to the chest or abdomen without involvement of bolero or bathing trunk distributions), body extremity (isolated to extremity), and body (both bolero and bathing trunk involvement). Our literature search found 113 cases of GCMN, which we were able to classify into 1 of the 6B patterns with an overall kappa of 0.89. LIMITATIONS: Some patterns occur infrequently with a dearth of images available for analysis. CONCLUSIONS: The anatomic distribution of GCMN occurs in 6 recognizable and repeatable patterns.

Prognostic role of the histological subtype of melanoma on the hands and feet in Caucasians.

Acral melanoma (AM) is associated with a poor prognosis in part because of delayed diagnosis, but probably also because of other intrinsic characteristics of location. The aim of this study was to review the specific characteristics and outcome of AM in Caucasians. This was a cross-sectional retrospective clinical-pathological study of 274 patients identified with AM in the database of a referral unit in Europe from 1986 to 2010. The mean age of the patients was 56.6 (SD 17.7) years. 269 cases could be histologically classified and included in the study. In all, 222 (82.5%) were located on feet. According to melanoma subtype, 165 (61.3%) were acral lentiginous melanoma (ALM), 84 (31.2%) were superficial spreading melanoma (SSM), and 20 (7.5%) were nodular melanoma (NM). SSM patients were characterized by female predominance (77.4%), younger age, and classic melanoma-risk phenotype (fair skin and multiple nevi). Among the 198 invasive cases with a mean follow-up of 56.2 months, the mean (SD) Breslow's thickness was 3.1 (3.6) mm, being 1.4 (1.4) mm in SSM, 3.5 (4.1) mm in ALM and 4.9 (2.9) mm in NM (P<0.001). Ulceration was present in 33.3%, 2.9% in SSM, 38.6% in ALM, and 76.9% in NM (P<0.001). A total of 29.3% relapsed (7.3% of SSM, 35% of ALM and 55% of NM) and 24.2% died because of AM. In multivariate analysis, age at diagnosis, Breslow, and histopathological subtype were independent prognostic factors for both disease-free and AM-specific survival. The ALM and NM subtypes presented poorer outcome after weighting Breslow and age (P=0.02). Histological subtype of AM could have an impact on biological behavior, ALM and NM subtypes presenting a poorer prognosis after adjusting for age and Breslow's thickness.

Genomic analysis and clinical management of adolescent cutaneous melanoma.

Melanoma in young children is rare; however, its incidence in adolescents and young adults is rising. We describe the clinical course of a 15-year-old female diagnosed with AJCC stage IB non-ulcerated primary melanoma, who died from metastatic disease 4 years after diagnosis despite three lines of modern systemic therapy. We also present the complete genomic profile of her tumour and compare this to a further series of 13 adolescent melanomas and 275 adult cutaneous melanomas. A somatic BRAFV600E mutation and a high mutational load equivalent to that found in adult melanoma and composed primarily of C>T mutations were observed. A germline genomic analysis alongside a series of 23 children and adolescents with melanoma revealed no mutations in known germline melanoma-predisposing genes. Adolescent melanomas appear to have genomes that are as complex as those arising in adulthood and their clinical course can, as with adults, be unpredictable.

Multiple BRAF Wild-Type Melanomas During Dabrafenib Treatment for Metastatic BRAF-Mutant Melanoma.

IMPORTANCE: BRAF inhibitors have become the standard of care in metastatic BRAF-mutant melanomas. Compared with chemotherapies, BRAF inhibitors improve overall and disease-free survival and speed the recovery of symptomatic patients with metastatic disease. The most worrisome finding is the possible development of resistance to new malignant tumors. OBSERVATIONS: A patient in her 30s developed massive BRAFV600E melanoma metastasis during her 30th week of pregnancy. After emergency cesarean delivery, oral dabrafenib treatment was initiated, and a partial radiologic response was confirmed within 1 month. At dermatologic digital follow-up aided by confocal microscopy 8 weeks after initiation of dabrafenib treatment, 4 melanomas were detected. Unfortunately, within the next month, the melanoma rapidly progressed. The 4 new melanomas were wild-type BRAFmelanomas, whereas the new metastasis carried a different BRAF mutation (S467L). CONCLUSIONS AND RELEVANCE: Cutaneous malignant tumors are the most frequent adverse events of BRAF inhibitors; therefore, strict dermatologic surveillance in a referral center aided by digital follow-up is mandatory, especially when multiple nevi are present and these drugs are used in an adjuvant setting. In view of our findings, the pathogenesis of the development of new melanomas seems to be different from therapy resistance. Whether paradoxical RAF activation could explain these BRAF wild-type secondary malignant tumors is still unknown.

Evaluation of PAX3 genetic variants and nevus number.

The presence of a high nevus number is the strongest phenotypic predictor of melanoma risk. Here, we describe the results of a three-stage study directed at identifying risk variants for the high nevus phenotype. At the first stage, 263 melanoma cases from Barcelona were genotyped for 223 single-nucleotide polymorphisms (SNPs) in 39 candidate genes. Seven SNPs in the PAX3 gene were found to be significantly associated with nevus number under the additive model. Next, the associations for seven PAX3 variants were evaluated in 1217 melanoma cases and 475 controls from Leeds; and in 3054 healthy twins from TwinsUK. Associations with high nevus number were detected for rs6754024 (P values < 0.01) in the Barcelona and Leeds datasets and for rs2855268 (P values < 0.01) in the Barcelona and the TwinsUK sets. Associations (P values < 0.001) in the opposite direction were detected for rs7600206 and rs12995399 in the Barcelona and TwinsUK sets. This study suggests that SNPs in PAX3 are associated with nevus number, providing support for PAX3 as a candidate nevus gene. Further studies are needed to examine the role of PAX3 in melanoma susceptibility.

Impact of sunscreens on preventing UVR-induced effects in nevi: in vivo study comparing protection using a physical barrier vs sunscreen.

IMPORTANCE: Sun damage is the most important environmental factor associated with malignant melanoma. To address the health threat, as well as the economic burden, primary prevention and early detection are crucial. OBJECTIVE: To test the efficacy of a topical sunscreen in the prevention of UV-induced effects in nevi. DESIGN: Prospective study of nevi protected by sunscreen vs a physical barrier. SETTING AND PATIENTS: Twenty-three nevi from 20 patients attending a referral hospital. INTERVENTION: Half of each nevus was protected by either a physical barrier or a sunscreen. Lesions were completely irradiated by a single dose of UV-B. MAIN OUTCOMES AND MEASURES: In vivo examination before and 7 days after irradiation and histopathologic-immunopathologic evaluation after excision on the seventh day. RESULTS: The most frequent clinical changes after UV radiation were pigmentation, scaling, and erythema; the most frequent dermoscopic changes were increased globules/dots, blurred network, regression, and dotted vessels. Both physical barrier- and sunscreen-protected areas showed some degree of these changes. More than 30% (7) of nevi did not show any change on clinical examination, and 18% (4) had no dermoscopic change. Immunohistopathologic differences between the halves of each nevus were demonstrable even when in vivo examination detected nothing. Parakeratotic scale, increased number and activation of superficial melanocytes, and keratinocyte proliferation were the most remarkable features. The only difference between both barriers was more enhanced melanocytic activation and regression features in the sunscreen group. No phenotypic features were found to predict a specific UV-B response. CONCLUSIONS AND RELEVANCE: Both physical barriers and sunscreens can partially prevent UV-B effects on nevi. Subclinical UV radiation effects, not always associated with visible changes, can develop even after protection. Sunscreens are not quite as effective as physical barriers in the prevention of inflammatory UV-B-induced effects.

Genetic alterations in RAS-regulated pathway in acral lentiginous melanoma.

Studies integrating clinicopathological and genetic features have revealed distinct patterns of genomic aberrations in Melanoma. Distributions of BRAF or NRAS mutations and gains of several oncogenes differ among melanoma subgroups, while 9p21 deletions are found in all melanoma subtypes. In the study, status of genes involved in cell cycle progression and apoptosis was evaluated in a panel of 17 frozen primary acral melanomas. NRAS mutations were found in 17% of the tumors. In contrast, BRAF mutations were not found. Gains of AURKA gene (20q13.3) were detected in 37.5% of samples, gains of CCND1 gene (11q13) or TERT gene (5p15.33) in 31.2% and gains of NRAS gene (1p13.2) in 25%. Alterations in 9p21 were identified in 69% of tumors. Gains of 11q13 and 20q13 were mutually exclusive, and 1p13.2 gain was associated with 5p15.33. Our findings showed that alterations in RAS-related pathways are present in 87.5% of acral lentiginous melanomas.

Genetic counseling in melanoma.

Genetic counseling may be offered to families with melanoma and to individuals with multiple melanomas to better understand the genetic susceptibility of the disease, the influence of environmental factors, the inheritance of the risk, and behavior that decreases the risk of dying from melanoma, including specific dermatological follow-up such as total body photography and digital dermoscopy. Genetic testing may be offered to those individuals with more than a 10% chance of being a carrier of a mutation. This risk varies according to the incidence of melanoma in the country and sun behavior. In countries with a low-medium incidence of melanoma, genetic testing should be offered to families with two cases of melanoma or an individual with two primary melanomas. In countries with a high incidence, families with three cases of melanoma, with two melanomas and one pancreatic adenocarcinoma, or patients with three primary melanomas, may benefit from genetic testing.

Cutaneous phenotype and MC1R variants as modifying factors for the development of melanoma in CDKN2A G101W mutation carriers from 4 countries.

The G101W founder mutation is the most common CDKN2A mutation in Italy, Spain, and France. As the background of modifying genes, environmental exposures, and sun behavior vary across countries, studying G101W carriers from distinct countries offers a unique opportunity to evaluate possible modifying factors in melanoma development. We evaluated 76 G101W cases and 59 carrier controls from France, Italy, Spain, and the United States. Hair color and dysplastic nevi distributions differed significantly in cases and controls across the 4 study groups. Cases also varied significantly for eye color, freckling, and nevi. The distribution of MC1R variants in cases differed significantly across study groups because 12% of Italian melanoma patients had > or =2 MC1R variants vs. >50% for the other case groups. Several MC1R covariates showed significant associations with melanoma risk in all groups combined and in the American, French, and Spanish samples; no significant findings were observed in the Italian sample. In multiple-case families, the number and type of MC1R variants varied significantly between multiple-primary-melanoma and single-primary-melanoma patients from the 4 groups; there was also a significant decrease in median age at melanoma diagnosis as the number or type of MC1R variants increased. The variation in the effects of the cutaneous phenotypic and MC1R factors across the study sample suggests that these factors differentially contribute to development of melanoma even on a common genetic background of a germline CDKN2A mutation. Differences in melanoma risk across geographic regions justify the need for individual studies in each country before counseling should be considered.

Features associated with germline CDKN2A mutations: a GenoMEL study of melanoma-prone families from three continents.

BACKGROUND: The major factors individually reported to be associated with an increased frequency of CDKN2A mutations are increased number of patients with melanoma in a family, early age at melanoma diagnosis, and family members with multiple primary melanomas (MPM) or pancreatic cancer. METHODS: These four features were examined in 385 families with > or =3 patients with melanoma pooled by 17 GenoMEL groups, and these attributes were compared across continents. RESULTS: Overall, 39% of families had CDKN2A mutations ranging from 20% (32/162) in Australia to 45% (29/65) in North America to 57% (89/157) in Europe. All four features in each group, except pancreatic cancer in Australia (p = 0.38), individually showed significant associations with CDKN2A mutations, but the effects varied widely across continents. Multivariate examination also showed different predictors of mutation risk across continents. In Australian families, > or =2 patients with MPM, median age at melanoma diagnosis < or =40 years and > or =6 patients with melanoma in a family jointly predicted the mutation risk. In European families, all four factors concurrently predicted the risk, but with less stringent criteria than in Australia. In North American families, only > or =1 patient with MPM and age at diagnosis < or =40 years simultaneously predicted the mutation risk. CONCLUSIONS: The variation in CDKN2A mutations for the four features across continents is consistent with the lower melanoma incidence rates in Europe and higher rates of sporadic melanoma in Australia. The lack of a pancreatic cancer-CDKN2A mutation relationship in Australia probably reflects the divergent spectrum of mutations in families from Australia versus those from North America and Europe. GenoMEL is exploring candidate host, genetic and/or environmental risk factors to better understand the variation observed.

High-risk melanoma susceptibility genes and pancreatic cancer, neural system tumors, and uveal melanoma across GenoMEL.

GenoMEL, comprising major familial melanoma research groups from North America, Europe, Asia, and Australia has created the largest familial melanoma sample yet available to characterize mutations in the high-risk melanoma susceptibility genes CDKN2A/alternate reading frames (ARF), which encodes p16 and p14ARF, and CDK4 and to evaluate their relationship with pancreatic cancer (PC), neural system tumors (NST), and uveal melanoma (UM). This study included 466 families (2,137 patients) with at least three melanoma patients from 17 GenoMEL centers. Overall, 41% (n = 190) of families had mutations; most involved p16 (n = 178). Mutations in CDK4 (n = 5) and ARF (n = 7) occurred at similar frequencies (2-3%). There were striking differences in mutations across geographic locales. The proportion of families with the most frequent founder mutation(s) of each locale differed significantly across the seven regions (P = 0.0009). Single founder CDKN2A mutations were predominant in Sweden (p.R112_L113insR, 92% of family's mutations) and the Netherlands (c.225_243del19, 90% of family's mutations). France, Spain, and Italy had the same most frequent mutation (p.G101W). Similarly, Australia and United Kingdom had the same most common mutations (p.M53I, c.IVS2-105A>G, p.R24P, and p.L32P). As reported previously, there was a strong association between PC and CDKN2A mutations (P < 0.0001). This relationship differed by mutation. In contrast, there was little evidence for an association between CDKN2A mutations and NST (P = 0.52) or UM (P = 0.25). There was a marginally significant association between NST and ARF (P = 0.05). However, this particular evaluation had low power and requires confirmation. This GenoMEL study provides the most extensive characterization of mutations in high-risk melanoma susceptibility genes in families with three or more melanoma patients yet available.