Genetic Layout of Melanoma Lesions Is Associated with BRAF/MEK-Targeted Therapy Resistance and Transcriptional Profiles.

The genetic landscape of melanoma resistance to targeted therapy with small molecules inhibiting BRAF and MEK kinases is still largely undefined. In this study, we portrayed in detail the somatic alterations of resistant melanoma and explored the associated biological processes and their integration with transcriptional profiles. By targeted next-generation sequencing and whole-exome sequencing analyses, a list of 101 genes showing imbalance in metastatic tumors from patients with a complete/durable response or disease progression during therapy with vemurafenib or with dabrafenib and trametinib was defined.

microRNA Expression in Sentinel Nodes from Progressing Melanoma Patients Identifies Networks Associated with Dysfunctional Immune Response.

Sentinel node biopsy (SNB) is a main staging biomarker in melanoma and is the first lymph node to drain the tumor, thus representing the immunological site where anti-tumor immune dysfunction is established and where potential prognostic immune markers can be identified. Here we analyzed microRNA (miR) profiles in archival tumor-positive SNBs derived from melanoma patients with different outcomes and performed an integrated analysis of transcriptional data to identify deregulated immune signaling networks. Twenty-six miRs were differentially expressed in melanoma-positive SNB samples between patients with disease progression and non-progressing patients, the majority being previously reported in the regulation of immune responses.

Overcoming melanoma resistance to vemurafenib by targeting CCL2-induced miR-34a, miR-100 and miR-125b.

In melanoma, the adaptative cell response to BRAF inhibitors includes altered patterns of cytokine production contributing to tumor progression and drug resistance. Among the factors produced by PLX4032-resistant melanoma cell lines, CCL2 was higher compared to the sensitive parental cell lines and increased upon drug treatment. CCL2 acted as an autocrine growth factor for melanoma cells, stimulating the proliferation and resistance to apoptosis.

Transcriptional profiling of melanoma sentinel nodes identify patients with poor outcome and reveal an association of CD30(+) T lymphocytes with progression.

Sentinel lymph nodes set the stance of the immune system to a localized tumor and are often the first site to be colonized by neoplastic cells that metastasize. To investigate how the presence of neoplastic cells in sentinel lymph nodes may trigger pathways associated with metastatic progression, we analyzed the transcriptional profiles of archival sentinel node biopsy specimens obtained from melanoma patients. Biopsies from positive nodes were selected for comparable tumor infiltration, presence or absence of further regional node metastases, and relapse at 5-year follow-up.

Identification of MET and SRC activation in melanoma cell lines showing primary resistance to PLX4032.

PLX4032/vemurafenib is a first-in-class small-molecule BRAF(V600E) inhibitor with clinical activity in patients with BRAF mutant melanoma. Nevertheless, drug resistance develops in treated patients, and strategies to overcome primary and acquired resistance are required. To explore the molecular mechanisms involved in primary resistance to PLX4032, we investigated its effects on cell proliferation and signaling in a panel of 27 genetically characterized patient-derived melanoma cell lines.

Heterogeneous phenotype of human melanoma cells with in vitro and in vivo features of tumor-initiating cells.

Melanospheres, the melanoma cells that grow as nonadherent colonies and that show in vitro self-renewing capacity and multipotency, were selected from melanoma specimens or from melanoma cell lines. Melanospheres were highly tumorigenic, and intradermal injections in severe combined immunodeficient (SCID) mice of as few as 100 cells generated tumors that maintained tumorigenic potential into subsequent recipients. Primary and serially transplanted xenografts recapitulated the phenotypic features of the original melanoma of the patient.

PTPRK negatively regulates transcriptional activity of wild type and mutated oncogenic beta-catenin and affects membrane distribution of beta-catenin/E-cadherin complexes in cancer cells.

Previous reports showed that receptor-type protein-tyrosine phosphatase PTPRK co-localizes with beta-catenin at adherens junctions, and in vitro experiments suggested that beta-catenin could be substrate of PTPRK-mediated phosphatase activity. beta-catenin is a molecule endowed with a dual function being involved both in cell adhesion and in Wnt signaling pathway. Here we provide evidence for the role of PTPRK in negatively regulating the beta-catenin transcriptional activity by modulating its intracellular and membrane distribution.

CCN3/nephroblastoma overexpressed matricellular protein regulates integrin expression, adhesion, and dissemination in melanoma.

CCN3/nephroblastoma overexpressed belongs to the CCN family of genes that encode secreted proteins associated with the extracellular matrix (ECM) and exert regulatory effects at the cellular level. Overexpression of CCN3 was shown in metastatic melanoma cells compared with cells of the primary tumor from the same patient. Analysis of short-term cultures from 50 primary and metastatic melanomas revealed a heterogeneous expression pattern of both the 46-kDa full-length cytoplasmic/secreted protein and the 32-kDa nuclear-truncated form.

The apoptosis inhibitor protein survivin induces tumor-specific CD8+ and CD4+ T cells in colorectal cancer patients.

The identification of tumor-associated antigens expressed by colorectal carcinoma remains one of the major goals for designing novel immunological treatments for this tumor. By using a reverse-immunology approach, we show here that the inhibitor of apoptosis protein, survivin, is immunogenic in colorectal cancer patients. In particular, we found that survivin elicited CD8(+) T cell-mediated responses in peripheral blood or in tumor-associated lymphocytes from patients at different disease stage.

Identification of a mutated receptor-like protein tyrosine phosphatase kappa as a novel, class II HLA-restricted melanoma antigen.

Recent studies increasingly point to a pivotal role of CD4(+) T cells in human anti-tumor immune response. Here we show that lymphocytes purified from a tumor-infiltrated lymph node of a melanoma patient that had remained disease free for 10 years after surgical resection of a lymph node metastasis comprised oligoclonal class II HLA-restricted CD4(+) T cells recognizing the autologous tumor cells in vitro. In fact, the CD4(+) T cell clones isolated from these lymphocytes displayed a tumor-specific, cytotoxic activity in addition to a Th1-like cytokine profile.

Induction of lymphocyte apoptosis by tumor cell secretion of FasL-bearing microvesicles.

The hypothesis that FasL expression by tumor cells may impair the in vivo efficacy of antitumor immune responses, through a mechanism known as 'Fas tumor counterattack,' has been recently questioned, becoming the object of an intense debate based on conflicting results. Here we definitely show that FasL is indeed detectable in the cytoplasm of melanoma cells and its expression is confined to multivesicular bodies that contain melanosomes. In these structures FasL colocalizes with both melanosomal (i.