Systematic analyses of cancer genomes promise to unveil patterns of genetic alterations linked to the genesis and spread of human cancers. High-density single-nucleotide polymorphism (SNP) arrays enable detailed and genome-wide identification of both loss-of-heterozygosity events and copy-number alterations in cancer. Here, by integrating SNP array-based genetic maps with gene expression signatures derived from NCI60 cell lines, we identified the melanocyte master regulator MITF (microphthalmia-associated transcription factor) as the target of a novel melanoma amplification. We found that MITF amplification was more prevalent in metastatic disease and correlated with decreased overall patient survival. BRAF mutation and p16 inactivation accompanied MITF amplification in melanoma cell lines. Ectopic MITF expression in conjunction with the BRAF(V600E) mutant transformed primary human melanocytes, and thus MITF can function as a melanoma oncogene. Reduction of MITF activity sensitizes melanoma cells to chemotherapeutic agents. Targeting MITF in combination with BRAF or cyclin-dependent kinase inhibitors may offer a rational therapeutic avenue into melanoma, a highly chemotherapy-resistant neoplasm. Together, these data suggest that MITF represents a distinct class of 'lineage survival' or 'lineage addiction' oncogenes required for both tissue-specific cancer development and tumour progression.

Microphthalmia-associated transcription factor (MITF) acts as a master regulator of melanocyte development, function and survival by modulating various differentiation and cell-cycle progression genes. It has been demonstrated that MITF is an amplified oncogene in a fraction of human melanomas and that it also has an oncogenic role in human clear cell sarcoma. However, MITF also modulates the state of melanocyte differentiation. Several closely related transcription factors also function as translocated oncogenes in various human malignancies. These data place MITF between instructing melanocytes towards terminal differentiation and/or pigmentation and, alternatively, promoting malignant behavior. In this review, we survey the roles of MITF as a master lineage regulator in melanocyte development and its emerging activities in malignancy. Understanding the molecular function of MITF and its associated pathways will hopefully shed light on strategies for improving therapeutic approaches for these diseases.

Extant evidence implicates growth factor signaling in the pathogenesis of many tumor types, including cutaneous melanoma. Recently, reciprocal activating mutations of NRAS and BRAF were found in benign melanocytic nevi and cutaneous melanomas. We had previously reported a similar epistatic relationship between activating NRAS mutations and inactivating PTEN/MMAC1 alterations. We thus hypothesized that BRAF and PTEN/MMAC1 mutations may cooperate to promote melanoma tumorigenesis. Overall, 40 of 47 (85%) melanoma cell lines and 11 of 16 (69%) uncultured melanoma metastases had mutations in NRAS, BRAF, or PTEN/MMAC1. NRAS was exclusively mutated in nine of 47 (19%) cell lines and two of 16 (13%) metastases, whereas BRAF was solely mutated in 28 of 47 (60%) cell lines and nine of 16 (56%) metastases. In the 12 of 15 melanoma cell lines (80%) and two of two melanoma metastases with PTEN alterations, BRAF was also mutated. These findings suggest the existence of possible cooperation between BRAF activation and PTEN loss in melanoma development.

Episodic exposure of fair-skinned individuals to intense sunlight is thought to be responsible for the steadily increasing melanoma incidence worldwide over recent decades. Rarely, melanoma susceptibility is increased more than tenfold by heritable mutations in the cell cycle regulatory genes CDKN2A and CDK4. Effective treatment requires early diagnosis followed by surgical excision with adequately wide margins. Sentinel lymph node biopsy provides accurate staging, but no published results are yet available from clinical trials designed to assess the therapeutic efficacy of early complete regional node dissection in those with metastatic disease in a sentinel node. Magnetic resonance spectroscopy is one technique under investigation for non-invasive, in-situ assessment of sentinel nodes. Localised metastatic disease is best treated surgically. No postoperative adjuvant therapy is of proven value for improving overall survival, although numerous clinical trials of vaccines and cytokines are in progress. Medical therapies have contributed little to the control of established metastatic disease, but molecular pathways recently identified as being central to melanoma growth and apoptosis are under intense investigation for their potential as therapeutic targets.

OBJECTIVE

Recently, it was reported that BRAF mutations are frequent in melanoma. Previously, we analyzed a large series of paired primary and metastatic melanomas for NRAS codon 61 mutations and showed that they arise early and are preserved during tumor progression. Here, we have screened the same tumor samples for BRAF mutations.

METHODS

Primary melanomas (n = 71) and corresponding metastases (n = 88) from 71 patients were screened for BRAF exon 11 and exon 15 mutations using single-strand conformational polymorphism and nucleotide sequence analysis

RESULTS

BRAF mutations were found in 42 of 71 patients (59%). Thirty-seven patients had mutations that lead to a Val599Glu change, whereas mutations resulting in Gly468Ser, Val599Arg, Val599Lys, and Lys600Glu changes were detected in one patient each. Furthermore, one patient had a 6-bp insertion between codons 598 and 599, encoding two threonine residues. In most cases, paired primary and metastatic lesions had the same BRAF genotype (i.e., mutations present in the primary tumors were preserved in the corresponding metastases, and mutations did not arise at the metastatic stage if they were not present in the primary lesion). Using laser-capture microdissection, BRAF mutations were found in the radial growth phase of the primary lesions. BRAF mutations occurred exclusively in tumors that were wild type for NRAS, and in total, 89% of the patients analyzed (63 of 71) had mutations in either of these two genes.

CONCLUSIONS

The Ras-Raf-mitogen-activated protein kinase/extracellular signal-regulated kinase-extracellular signal-regulated kinase signaling pathway is activated in the vast majority of melanomas. Activation occurs through either NRAS or BRAF mutations, both of which arise early during melanoma pathogenesis and are preserved throughout tumor progression.

Tumour development and progression involves the expression of oncogenes and inactivation of tumour suppressor genes, leading to the appearance of multiple malignant characteristics. Malignant melanoma cells express different growth factors and cytokines and their receptors in respective stages of tumour progression, which by autocrine and paracrine effects enable them to grow autonomously and confer competence to metastasis. Autocrine growth factors (bFGF, MGSA/GRO, IL-8 and sometimes IL-6, PDGF-A, IL-10) produced by melanoma cells stimulate proliferation of the producing cell itself, while paracrine growth factors (for example PDGF, EGF, TGF-beta, IL-1, GM-CSF, IGF-I, NGF, VEGF) modulate the microenvironment to the benefit of tumour growth and invasion. Paracrine effects include angiogenesis, stroma formation, modulation of host immune response, activation of proteolytic enzymes, adhesion or motility and metastasis formation. Some growth factors have inhibitory effects on melanocytes and early lesions (IL-1, IL-6, TGF-beta, OSM, TNF and IFN) but not on advanced stage melanomas, and in some cases they switch to autocrine stimulator (IL-6, TGF-beta). Understanding the involvement of different growth factors and cytokines in the molecular mechanism of melanoma progression will help to provide an insight into new future therapeutic approaches for melanoma.

Therapeutic resistance and proclivity for metastasis are hallmarks of malignant melanoma. Genetic, epidemiological and genomic investigations are uncovering the spectrum of stereotypical mutations that are associated with melanoma and how these mutations relate to risk factors such as ultraviolet exposure. The ability to validate the pathogenetic relevance of these mutations in the mouse, coupled with advances in rational drug design, has generated optimism for the development of effective prevention programmes, diagnostic measures and targeted therapeutics in the near future.

Melanoma is a cancer of the neural crest-derived cells that provide pigmentation to skin and other tissues. Over the past 4 decades, the incidence of melanoma has increased more rapidly than that of any other malignancy in the United States. No current treatments substantially enhance patient survival once metastasis has occurred. This review focuses on recent insights into melanoma genetics and new therapeutic approaches being developed based on these advances.

In the last decade, significant progress has been made in our understanding of the genetic alterations in melanocytic tumors. The most exciting finding is the discovery of oncogenic BRAF mutations in both malignant melanoma and melanocytic nevi. This finding indicates that activation of the mitogen-activated protein kinase pathway may be a critical initiating step of melanocytic neoplasia, and that the fundamental difference between melanoma and nevi may lie in the inhibitory machinery for this oncogenic signaling. In addition, different genetic alterations identified in melanomas at different sites and with different levels of sun exposure have been shown, indicating that there are several distinct genetic pathways in the development of melanoma. Different patterns of genetic alterations have also been identified among different kinds of melanocytic nevi. While acquired nevi and small congenital nevi show a high frequency of BRAF mutations regardless of their anatomic localization, the mutations were rare in medium-sized congenital nevi and giant congenital nevi. Spitz nevi show no BRAF mutations, while a subset of cases show HRAS mutations, often associated with a copy number increase of chromosome 11p. The clear differences in genetic aberration patterns have significant clinical implications in the diagnosis and treatment of melanocytic tumors.

Cancer is generally viewed as the result of disrupted intra-and intercellular homeostatic regulation. Once the homeostatic balance is lost and malignant transformation has occurred, microenvironmental factors such as degradation of matrix components and host-tumor interactions are essential for survival and growth of malignant cells. Within the previous year, cadherins and matrixins (matrix metalloproteinases) have emerged as key factors in these processes. The pathways involved are interconnected and detailed knowledge about the biologic significance of each member in a given pathway is essential for our understanding of oncogenesis. Restoration of E-cadherin-mediated control over melanoma cells and modulation of the involved regulation pathways are promising novel therapeutic strategies. Another approach is the rational design of inhibitors that perturb matrix metalloproteinases in a particular cell type and interrupt tumor-specific proteinase activation cascades. Advances in these fields will lead to the development of better tools for prevention, diagnosis, and therapy.

Mutations in the TP53 gene are found in about 11% of melanomas. Although nearly 600 papers have been published with varying degrees of consensus, there does not appear to be any comparable analysis that facilitates more than a glimpse into the role of p53 in melanomagenesis. This article reviews p53 alterations (at the gene and protein levels) in melanocytic skin lesions and discusses the following points: (i) p53 alterations commence as early as at the stage of benign and dysplastic nevi; (ii) these alterations are frequent in melanomas, and gradually increase with their progression; (iii) there is no concordance between the frequent p53 protein expression and the rarity of both TP53 gene mutations in melanomas, and (iv) the entire p53 pathway is a more critical determinant of the fate of the melanocytic skin lesions than the status of the p53 protein or the gene itself.