RESUMEN
Melanoma of the skin is a common cancer only in Europeans, whereas it arises in internal body surfaces (mucosal sites) and on the hands and feet (acral sites) in people throughout the world. Here we report analysis of whole-genome sequences from cutaneous, acral and mucosal subtypes of melanoma. The heavily mutated landscape of coding and non-coding mutations in cutaneous melanoma resolved novel signatures of mutagenesis attributable to ultraviolet radiation. However, acral and mucosal melanomas were dominated by structural changes and mutation signatures of unknown aetiology, not previously identified in melanoma. The number of genes affected by recurrent mutations disrupting non-coding sequences was similar to that affected by recurrent mutations to coding sequences. Significantly mutated genes included BRAF, CDKN2A, NRAS and TP53 in cutaneous melanoma, BRAF, NRAS and NF1 in acral melanoma and SF3B1 in mucosal melanoma. Mutations affecting the TERT promoter were the most frequent of all; however, neither they nor ATRX mutations, which correlate with alternative telomere lengthening, were associated with greater telomere length. Most melanomas had potentially actionable mutations, most in components of the mitogen-activated protein kinase and phosphoinositol kinase pathways. The whole-genome mutation landscape of melanoma reveals diverse carcinogenic processes across its subtypes, some unrelated to sun exposure, and extends potential involvement of the non-coding genome in its pathogenesis.
Asunto(s)
Genoma Humano/genética , Melanoma/genética , Mutación/genética , ADN Helicasas/genética , GTP Fosfohidrolasas/genética , Genes p16 , Humanos , Melanoma/clasificación , Proteínas de la Membrana/genética , Proteínas Quinasas Activadas por Mitógenos/genética , Neurofibromatosis 1/genética , Proteínas Nucleares/genética , Fosfoproteínas/genética , Proteínas Proto-Oncogénicas B-raf/genética , Factores de Empalme de ARN/genética , Transducción de Señal/efectos de los fármacos , Telomerasa/genética , Telómero/genética , Proteína p53 Supresora de Tumor/genética , Rayos Ultravioleta/efectos adversos , Proteína Nuclear Ligada al Cromosoma XRESUMEN
The individuals carrying melanocortin-1 receptor (MC1R) variants, especially those associated with red hair color, fair skin, and poor tanning ability (RHC trait), are more prone to melanoma; however, the underlying mechanism is poorly defined. Here, we report that UVB exposure triggers phosphatase and tensin homolog (PTEN) interaction with wild-type (WT), but not RHC-associated MC1R variants, which protects PTEN from WWP2-mediated degradation, leading to AKT inactivation. Strikingly, the biological consequences of the failure of MC1R variants to suppress PI3K/AKT signaling are highly context dependent. In primary melanocytes, hyperactivation of PI3K/AKT signaling leads to premature senescence; in the presence of BRAF(V600E), MC1R deficiency-induced elevated PI3K/AKT signaling drives oncogenic transformation. These studies establish the MC1R-PTEN axis as a central regulator for melanocytes' response to UVB exposure and reveal the molecular basis underlying the association between MC1R variants and melanomagenesis.
Asunto(s)
Regulación de la Expresión Génica/efectos de la radiación , Melanocitos/metabolismo , Melanoma Experimental/patología , Fosfohidrolasa PTEN/metabolismo , Receptor de Melanocortina Tipo 1/metabolismo , Pigmentación de la Piel/fisiología , Rayos Ultravioleta , Animales , Western Blotting , Células Cultivadas , Humanos , Técnicas para Inmunoenzimas , Melanocitos/efectos de la radiación , Melanoma Experimental/genética , Melanoma Experimental/metabolismo , Ratones , Mutación/genética , Fosfohidrolasa PTEN/genética , Fosfatidilinositol 3-Quinasas/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Fosforilación , Proteínas Proto-Oncogénicas B-raf/genética , Proteínas Proto-Oncogénicas c-akt/genética , Proteínas Proto-Oncogénicas c-akt/metabolismo , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptor de Melanocortina Tipo 1/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de Señal , Pigmentación de la Piel/efectos de la radiación , alfa-MSH/genética , alfa-MSH/metabolismoRESUMEN
The treatment of melanoma has been markedly improved by the introduction of targeted therapies and checkpoint blockade immunotherapy. Unfortunately, resistance to these therapies remains a limitation. Novel anticancer therapeutics targeting the MCL1 anti-apoptotic protein have shown impressive responses in haematological cancers but are yet to be evaluated in melanoma. To assess the sensitivity of melanoma to new MCL1 inhibitors, we measured the response of 51 melanoma cell lines to the novel MCL1 inhibitor, S63845. Additionally, we assessed combination of this drug with inhibitors of the bromodomain and extra-terminal (BET) protein family of epigenetic readers, which we postulated would assist MCL1 inhibition by downregulating anti-apoptotic targets regulated by NF-kB such as BCLXL, BCL2A1 and XIAP, and by upregulating pro-apoptotic proteins including BIM and NOXA. Only 14% of melanoma cell lines showed sensitivity to S63845, however, combination of S63845 and I-BET151 induced highly synergistic apoptotic cell death in all melanoma lines tested and in an in vivo xenograft model. Cell death was dependent on caspases and BAX/BAK. Although the combination of drugs increased the BH3-only protein, BIM, and downregulated anti-apoptotic proteins such as BCL2A1, the importance of these proteins in inducing cell death varied between cell lines. ABT-199 or ABT-263 inhibitors against BCL2 or BCL2 and BCLXL, respectively, induced further cell death when combined with S63845 and I-BET151. The combination of MCL1 and BET inhibition appears to be a promising therapeutic approach for metastatic melanoma, and presents opportunities to add further BCL2 family inhibitors to overcome treatment resistance.
Asunto(s)
Antineoplásicos/farmacología , Muerte Celular/efectos de los fármacos , Melanoma/tratamiento farmacológico , Melanoma/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Proteínas/metabolismo , Animales , Apoptosis/efectos de los fármacos , Proteínas Reguladoras de la Apoptosis/metabolismo , Línea Celular Tumoral , Regulación hacia Abajo/efectos de los fármacos , Resistencia a Antineoplásicos/efectos de los fármacos , Sinergismo Farmacológico , Humanos , Masculino , Ratones , Ratones Endogámicos NOD , Ratones SCID , Pirimidinas/farmacología , Tiofenos/farmacología , Regulación hacia Arriba/efectos de los fármacosRESUMEN
So far, two genes associated with familial melanoma have been identified, accounting for a minority of genetic risk in families. Mutations in CDKN2A account for approximately 40% of familial cases, and predisposing mutations in CDK4 have been reported in a very small number of melanoma kindreds. Here we report the whole-genome sequencing of probands from several melanoma families, which we performed in order to identify other genes associated with familial melanoma. We identify one individual carrying a novel germline variant (coding DNA sequence c.G1075A; protein sequence p.E318K; rs149617956) in the melanoma-lineage-specific oncogene microphthalmia-associated transcription factor (MITF). Although the variant co-segregated with melanoma in some but not all cases in the family, linkage analysis of 31 families subsequently identified to carry the variant generated a log of odds (lod) score of 2.7 under a dominant model, indicating E318K as a possible intermediate risk variant. Consistent with this, the E318K variant was significantly associated with melanoma in a large Australian case-control sample. Likewise, it was similarly associated in an independent case-control sample from the United Kingdom. In the Australian sample, the variant allele was significantly over-represented in cases with a family history of melanoma, multiple primary melanomas, or both. The variant allele was also associated with increased naevus count and non-blue eye colour. Functional analysis of E318K showed that MITF encoded by the variant allele had impaired sumoylation and differentially regulated several MITF targets. These data indicate that MITF is a melanoma-predisposition gene and highlight the utility of whole-genome sequencing to identify novel rare variants associated with disease susceptibility.
Asunto(s)
Predisposición Genética a la Enfermedad , Melanoma/genética , Factor de Transcripción Asociado a Microftalmía/genética , Mutación , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Persona de Mediana Edad , Sumoilación/genética , Adulto JovenRESUMEN
Synonymous mutations, which do not alter the protein sequence, have been shown to affect protein function [Sauna ZE, Kimchi-Sarfaty C (2011) Nat Rev Genet 12(10):683-691]. However, synonymous mutations are rarely investigated in the cancer genomics field. We used whole-genome and -exome sequencing to identify somatic mutations in 29 melanoma samples. Validation of one synonymous somatic mutation in BCL2L12 in 285 samples identified 12 cases that harbored the recurrent F17F mutation. This mutation led to increased BCL2L12 mRNA and protein levels because of differential targeting of WT and mutant BCL2L12 by hsa-miR-671-5p. Protein made from mutant BCL2L12 transcript bound p53, inhibited UV-induced apoptosis more efficiently than WT BCL2L12, and reduced endogenous p53 target gene transcription. This report shows selection of a recurrent somatic synonymous mutation in cancer. Our data indicate that silent alterations have a role to play in human cancer, emphasizing the importance of their investigation in future cancer genome studies.
Asunto(s)
Apoptosis/genética , Regulación de la Expresión Génica/genética , Genoma Humano/genética , Melanoma/genética , Proteínas Musculares/genética , Proteínas Proto-Oncogénicas c-bcl-2/genética , Secuencia de Bases , Western Blotting , Cartilla de ADN/genética , Exoma/genética , Vectores Genéticos/genética , Células HEK293 , Humanos , Inmunoprecipitación , Lentivirus , MicroARNs/genética , Datos de Secuencia Molecular , Proteínas Musculares/metabolismo , Mutación/genética , Polimorfismo de Nucleótido Simple/genética , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , ARN Interferente Pequeño/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Análisis de Secuencia de ADN , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
High density SNP arrays can be used to identify DNA copy number changes in tumors such as homozygous deletions of tumor suppressor genes and focal amplifications of oncogenes. Illumina Human CNV370 Bead chip arrays were used to assess the genome for unbalanced chromosomal events occurring in 39 cell lines derived from stage III metastatic melanomas. A number of genes previously recognized to have an important role in the development and progression of melanoma were identified including homozygous deletions of CDKN2A (13 of 39 samples), CDKN2B (10 of 39), PTEN (3 of 39), PTPRD (3 of 39), TP53 (1 of 39), and amplifications of CCND1 (2 of 39), MITF (2 of 39), MDM2 (1 of 39), and NRAS (1 of 39). In addition, a number of focal homozygous deletions potentially targeting novel melanoma tumor suppressor genes were identified. Because of their likely functional significance for melanoma progression, FAS, CH25H, BMPR1A, ACTA2, and TFG were investigated in a larger cohort of melanomas through sequencing. Nonsynonymous mutations were identified in BMPR1A (1 of 43), ACTA2 (3 of 43), and TFG (5 of 103). A number of potentially important mutation events occurred in TFG including the identification of a mini mutation "hotspot" at amino acid residue 380 (P380S and P380L) and the presence of multiple mutations in two melanomas. Mutations in TFG may have important clinical relevance for current therapeutic strategies to treat metastatic melanoma.
Asunto(s)
Genes Supresores de Tumor , Melanoma/genética , Melanoma/patología , Proteínas/genética , Línea Celular Tumoral , Amplificación de Genes , Eliminación de Gen , Homocigoto , Humanos , Mutación , Metástasis de la Neoplasia , Estadificación de NeoplasiasRESUMEN
BACKGROUND: Public health sun safety campaigns introduced during the 1980s have successfully reduced skin cancer rates in Australia. Despite this success, high rates of sunburn continue to be reported by youth and young adults. As such, new strategies to reinforce sun protection approaches in this demographic are needed. OBJECTIVE: This study aims to develop a virtual reality (VR) game containing preventive skin cancer messaging and to assess the safety and satisfaction of the design based on end user feedback. METHODS: Using a two-phase design approach, we created a prototype VR game that immersed the player inside the human body while being confronted with growing cancer cells. The first design phase involved defining the problem, identifying stakeholders, choosing the technology platform, brainstorming, and designing esthetic elements. In the second design phase, we tested the prototype VR experience with stakeholders and end users in focus groups and interviews, with feedback incorporated into refining and improving the design. RESULTS: The focus groups and interviews were conducted with 18 participants. Qualitative feedback indicated high levels of satisfaction, with all participants reporting the VR game as engaging. A total of 11% (2/8) of participants reported a side effect of feeling nauseous during the experience. The end user feedback identified game improvements, suggesting an extended multistage experience with visual transitions to other environments and interactions involving cancer causation. The implementation of the VR game identified challenges in sharing VR equipment and hygiene issues. CONCLUSIONS: This study presents key findings highlighting the design and implementation approaches for a VR health intervention primarily aimed at improving sun protection behaviors. This design approach can be applied to other health prevention programs in the future.
RESUMEN
Somatic mutation analysis of melanoma has been performed at the single gene level extensively over the past several decades. This has provided considerable insight into the critical pathways controlling melanoma initiation and progression. During the last 5 yr, next-generation sequencing (NGS) has enabled even more comprehensive mutational screening at the level of multigene panels, exomes and genomes. These studies have uncovered many new and unexpected players in melanoma development. The recent landmark study from The Cancer Genome Atlas (TCGA) consortium describing the genomic architecture of 333 cutaneous melanomas provides the largest and broadest analysis to date on the somatic aberrations underlying melanoma genesis. It thus seems timely to review the mutational landscape of melanoma and highlight the key genes and cellular pathways that appear to drive this cancer.
Asunto(s)
Genoma Humano , Melanoma/genética , Animales , Genes Relacionados con las Neoplasias , Humanos , Tasa de Mutación , Regiones Promotoras Genéticas/genética , Neoplasias Cutáneas , Rayos Ultravioleta , Melanoma Cutáneo MalignoRESUMEN
To explore the distinct genotypic and phenotypic states of melanoma tumors, we applied single-cell RNA sequencing (RNA-seq) to 4645 single cells isolated from 19 patients, profiling malignant, immune, stromal, and endothelial cells. Malignant cells within the same tumor displayed transcriptional heterogeneity associated with the cell cycle, spatial context, and a drug-resistance program. In particular, all tumors harbored malignant cells from two distinct transcriptional cell states, such that tumors characterized by high levels of the MITF transcription factor also contained cells with low MITF and elevated levels of the AXL kinase. Single-cell analyses suggested distinct tumor microenvironmental patterns, including cell-to-cell interactions. Analysis of tumor-infiltrating T cells revealed exhaustion programs, their connection to T cell activation and clonal expansion, and their variability across patients. Overall, we begin to unravel the cellular ecosystem of tumors and how single-cell genomics offers insights with implications for both targeted and immune therapies.
Asunto(s)
Melanoma/genética , Melanoma/secundario , Neoplasias Cutáneas/patología , Microambiente Tumoral , Secuencia de Bases , Comunicación Celular , Ciclo Celular , Resistencia a Antineoplásicos/genética , Células Endoteliales/patología , Genómica , Humanos , Inmunoterapia , Activación de Linfocitos , Melanoma/terapia , Factor de Transcripción Asociado a Microftalmía/metabolismo , Metástasis de la Neoplasia , ARN/genética , Análisis de Secuencia de ARN , Análisis de la Célula Individual , Células del Estroma/patología , Linfocitos T/inmunología , Linfocitos T/patología , TranscriptomaRESUMEN
The epigenetic modifier EZH2 is part of the polycomb repressive complex that suppresses gene expression via histone methylation. Activating mutations in EZH2 are found in a subset of melanoma that contributes to disease progression by inactivating tumor suppressor genes. In this study we have targeted EZH2 with a specific inhibitor (GSK126) or depleted EZH2 protein by stable shRNA knockdown. We show that inhibition of EZH2 has potent effects on the growth of both wild-type and EZH2 mutant human melanoma in vitro particularly in cell lines harboring the EZH2Y646 activating mutation. This was associated with cell cycle arrest, reduced proliferative capacity in both 2D and 3D culture systems, and induction of apoptosis. The latter was caspase independent and mediated by the release of apoptosis inducing factor (AIFM1) from mitochondria. Gene expression arrays showed that several well characterized tumor suppressor genes were reactivated by EZH2 inhibition. This included activating transcription factor 3 (ATF3) that was validated as an EZH2 target gene by ChIP-qPCR. These results emphasize a critical role for EZH2 in the proliferation and viability of melanoma and highlight the potential for targeted therapy against EZH2 in treatment of patients with melanoma.
Asunto(s)
Genes Supresores de Tumor , Melanoma/genética , Melanoma/metabolismo , Complejo Represivo Polycomb 2/genética , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/metabolismo , Factor de Transcripción Activador 3/genética , Apoptosis , Ciclo Celular , Línea Celular Tumoral , Proliferación Celular , Supervivencia Celular , Progresión de la Enfermedad , Proteína Potenciadora del Homólogo Zeste 2 , Epigenómica , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Silenciador del Gen , Genotipo , Histonas/química , Humanos , Indoles/química , Concentración 50 Inhibidora , Mutación , Piridonas/química , Regulación hacia ArribaRESUMEN
Analysis of 501 melanoma exomes identified RASA2, encoding a RasGAP, as a tumor-suppressor gene mutated in 5% of melanomas. Recurrent loss-of-function mutations in RASA2 were found to increase RAS activation, melanoma cell growth and migration. RASA2 expression was lost in ≥30% of human melanomas and was associated with reduced patient survival. These findings identify RASA2 inactivation as a melanoma driver and highlight the importance of RasGAPs in cancer.
Asunto(s)
Biomarcadores de Tumor/genética , Exoma/genética , Melanoma/genética , Mutación/genética , Neoplasias Cutáneas/genética , Proteínas Activadoras de ras GTPasa/genética , Secuenciación de Nucleótidos de Alto Rendimiento/métodos , Humanos , Melanoma/mortalidad , Melanoma/patología , Pronóstico , Neoplasias Cutáneas/mortalidad , Neoplasias Cutáneas/patología , Tasa de SupervivenciaRESUMEN
BACKGROUND: The shelterin complex protects chromosomal ends by regulating how the telomerase complex interacts with telomeres. Following the recent finding in familial melanoma of inactivating germline mutations in POT1, encoding a member of the shelterin complex, we searched for mutations in the other five components of the shelterin complex in melanoma families. METHODS: Next-generation sequencing techniques were used to screen 510 melanoma families (with unknown genetic etiology) and control cohorts for mutations in shelterin complex encoding genes: ACD, TERF2IP, TERF1, TERF2, and TINF 2. Maximum likelihood and LOD [logarithm (base 10) of odds] analyses were used. Mutation clustering was assessed with χ(2) and Fisher's exact tests. P values under .05 were considered statistically significant (one-tailed with Yates' correction). RESULTS: Six families had mutations in ACD and four families carried TERF2IP variants, which included nonsense mutations in both genes (p.Q320X and p.R364X, respectively) and point mutations that cosegregated with melanoma. Of five distinct mutations in ACD, four clustered in the POT1 binding domain, including p.Q320X. This clustering of novel mutations in the POT1 binding domain of ACD was statistically higher (P = .005) in melanoma probands compared with population control individuals (n = 6785), as were all novel and rare variants in both ACD (P = .040) and TERF2IP (P = .022). Families carrying ACD and TERF2IP mutations were also enriched with other cancer types, suggesting that these variants also predispose to a broader spectrum of cancers than just melanoma. Novel mutations were also observed in TERF1, TERF2, and TINF2, but these were not convincingly associated with melanoma. CONCLUSIONS: Our findings add to the growing support for telomere dysregulation as a key process associated with melanoma susceptibility.
Asunto(s)
Codón sin Sentido , Melanoma/genética , Neoplasias Cutáneas/genética , Proteínas de Unión a Telómeros/genética , Adulto , Anciano , ADN de Neoplasias/análisis , Femenino , Predisposición Genética a la Enfermedad , Mutación de Línea Germinal , Humanos , Masculino , Persona de Mediana Edad , Linaje , Mutación Puntual , Análisis de Secuencia de ADN , Complejo Shelterina , Telómero/genética , Proteína 2 de Unión a Repeticiones Teloméricas/genéticaRESUMEN
BACKGROUND: 5-year survival for melanoma metastasis to regional lymph nodes (American Joint Committee on Cancer stage III) is <50%. Knowledge of outcomes following therapeutic lymphadenectomy for stage III melanoma related to BRAF status may guide adjuvant use of BRAF/MEK inhibitors along with established and future therapies. AIMS: To determine patterns of melanoma recurrence and survival following therapeutic lymph node dissection (TLND) associated with oncogenic mutations. METHODS: DNA was obtained from patients who underwent TLND and had ⩾2 positive nodes, largest node >3cm or extracapsular invasion. Mutations were detected using an extended Sequenom MelaCARTA panel. RESULTS: Mutations were most commonly detected in BRAF (57/124 [46%] patients) and NRAS (26/124 [21%] patients). Patients with BRAF mutations had higher 3-year recurrence rate (77%) versus 54% for BRAF wild-type patients (hazard ratio (HR) 1.8, p=0.008). The only prognostically significant mutations occurred in BRAF: median recurrence-free (RFS) and disease-specific survival (DSS) for BRAF mutation patients was 7 months and 16 months, versus 19 months and not reached for BRAF wild-type patients, respectively. Multivariate analysis identified BRAF mutant status and number of positive lymph nodes as the only independent prognostic factors for RFS and DSS. CONCLUSIONS: Patients with BRAF mutations experienced rapid progression of metastatic disease with locoregional recurrence rarely seen in isolation, supporting incorporation of BRAF status into melanoma staging and use of BRAF/MEK inhibitors post-TLND.
Asunto(s)
Melanoma/genética , Mutación , Proteínas Proto-Oncogénicas B-raf/genética , Neoplasias Cutáneas/genética , Análisis Mutacional de ADN , Femenino , Estudios de Seguimiento , Humanos , Inmunoterapia , Estimación de Kaplan-Meier , Escisión del Ganglio Linfático , Metástasis Linfática , Masculino , Melanoma/cirugía , Melanoma/terapia , Persona de Mediana Edad , Análisis Multivariante , Recurrencia Local de Neoplasia , Estadificación de Neoplasias , Evaluación de Resultado en la Atención de Salud/estadística & datos numéricos , Pronóstico , Modelos de Riesgos Proporcionales , Proto-Oncogenes Mas , Radioterapia Adyuvante , Neoplasias Cutáneas/cirugía , Neoplasias Cutáneas/terapiaRESUMEN
The incidence of melanoma continues to rise globally and is increasing at a rate greater than any other cancer. To systematically search for new genes involved in melanomagenesis, we collated exome sequencing data from independent melanoma cohort datasets, including those in the public domain. We identified recurrent mutations that may drive melanoma growth, survival or metastasis, and which may hold promise for the design of novel therapies to treat melanoma. These included a frequent recurrent (i.e. hotspot) mutation in the 5' untranslated region of RPS27 in ~10% of samples. We show that the mutation expands the 5'TOP element, a motif known to regulate the expression of most of the ribosomal protein family, to which RPS27 belongs, and thus might sensitize the mutated transcript to growth-mediated regulation. This finding highlights not only the important role of non-protein coding genetic aberrations in cancer development but also their potential as novel therapeutic targets.
Asunto(s)
Regiones no Traducidas 5' , Melanoma/genética , Metaloproteínas/genética , Proteínas Nucleares/genética , Proteínas de Unión al ARN/genética , Proteínas Ribosómicas/genética , Neoplasias Cutáneas/genética , Secuencia de Bases , Humanos , Datos de Secuencia Molecular , Mutación , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Patients with advanced metastatic melanoma have poor prognosis and the genetics underlying its pathogenesis are poorly understood. High-throughput sequencing has allowed comprehensive discovery of somatic mutations in cancer samples. Here, on analysis of our whole-genome and whole-exome sequencing data of 29 melanoma samples, we identified several genes that harbor recurrent nonsynonymous mutations. These included MAP3K5 (mitogen-activated protein kinase kinase kinase-5), which in a prevalence screen of 288 melanomas was found to harbor a R256C substitution in 5 cases. All MAP3K5-mutated samples were wild type for BRAF, suggesting a mutual exclusivity for these mutations. Functional analysis of the MAP3K5 R256C mutation revealed attenuation of MKK4 (mitogen-activated protein kinase kinase 4) activation through increased binding of the inhibitory protein thioredoxin (TXN/TRX-1/Trx), resulting in increased proliferation and anchorage-independent growth of melanoma cells. This mutation represents a potential target for the design of new therapies to treat melanoma.
Asunto(s)
MAP Quinasa Quinasa Quinasa 5/genética , Melanoma/genética , Melanoma/patología , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/patología , Tiorredoxinas/metabolismo , Apoptosis/fisiología , Proliferación Celular , Células HEK293 , Humanos , MAP Quinasa Quinasa 4/metabolismo , MAP Quinasa Quinasa Quinasa 5/metabolismo , Melanoma/metabolismo , Modelos Genéticos , Mutación Puntual , Unión Proteica , ARN Interferente Pequeño/genética , Transducción de Señal/fisiología , Neoplasias Cutáneas/metabolismo , Células Tumorales CultivadasRESUMEN
We have investigated the potential for the p16-cyclin D-CDK4/6-retinoblastoma protein pathway to be exploited as a therapeutic target in melanoma. In a cohort of 143 patients with primary invasive melanoma, we used fluorescence in situ hybridization to detect gene copy number variations (CNVs) in CDK4, CCND1, and CDKN2A and immunohistochemistry to determine protein expression. CNVs were common in melanoma, with gain of CDK4 or CCND1 in 37 and 18% of cases, respectively, and hemizygous or homozygous loss of CDKN2A in 56%. Three-quarters of all patients demonstrated a CNV in at least one of the three genes. The combination of CCND1 gain with either a gain of CDK4 and/or loss of CDKN2A was associated with poorer melanoma-specific survival. In 47 melanoma cell lines homozygous loss, methylation or mutation of CDKN2A gene or loss of protein (p16(INK) (4A) ) predicted sensitivity to the CDK4/6 inhibitor PD0332991, while RB1 loss predicted resistance.
Asunto(s)
Quinasa 4 Dependiente de la Ciclina/antagonistas & inhibidores , Quinasa 6 Dependiente de la Ciclina/antagonistas & inhibidores , Inhibidor p16 de la Quinasa Dependiente de Ciclina/biosíntesis , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Melanoma/metabolismo , Piperazinas/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Piridinas/farmacología , Adulto , Anciano , Anciano de 80 o más Años , Línea Celular Tumoral , Quinasa 4 Dependiente de la Ciclina/biosíntesis , Quinasa 6 Dependiente de la Ciclina/biosíntesis , Inhibidor p16 de la Quinasa Dependiente de Ciclina/genética , Femenino , Humanos , Masculino , Melanoma/genética , Melanoma/patología , Persona de Mediana Edad , Invasividad NeoplásicaRESUMEN
Melanoma of unknown primary (MUP) is an uncommon phenomenon whereby patients present with metastatic disease without an evident primary site. To determine their likely site of origin, we combined exome sequencing from 33 MUPs to assess the total rate of somatic mutations and degree of UV mutagenesis. An independent cohort of 91 archival MUPs was also screened for 46 hot spot mutations highly prevalent in melanoma including BRAF, NRAS, KIT, GNAQ, and GNA11. Results showed that the majority of MUPs exhibited high somatic mutation rates, high ratios of C>T/G>A transitions, and a high rate of BRAF (45 of 101, 45%) and NRAS (32 of 101, 32%) mutations, collectively indicating a mutation profile consistent with cutaneous sun-exposed melanomas. These data suggest that a significant proportion of MUPs arise from regressed or unrecognized primary cutaneous melanomas or arise de novo in lymph nodes from nevus cells that have migrated from the skin.