RESUMO
Whereas immunotherapies have revolutionized the treatment of different solid and hematological cancers, their efficacy in nodal peripheral T-cell lymphomas (PTCLs) is limited, due to a lack of understanding of the immune response they trigger. To fully characterize the immune tumor microenvironment (TME) of PTCLs, we performed spectral flow cytometry analyses on 11 angioimmunoblastic T-cell lymphomas (AITL), 7 PTCL, not otherwise specified (PTCL, NOS) lymph node samples, and 10 non-tumoral control samples. The PTCL TME contained a larger proportion of regulatory T cells and exhausted CD8+ T cells, with enriched expression of druggable immune checkpoints. Interestingly, CD39 expression was up-regulated at the surface of most immune cells, and a multi-immunofluorescence analyses on a retrospective cohort of 43 AITL patients demonstrated a significant association between high CD39 expression by T cells and poor patient prognosis. Together, our study unravels the complex TME of nodal PTCLs, identifies targetable immune checkpoints, and highlights CD39 as a novel prognostic factor.
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Human skin is particularly vulnerable to age-related deterioration and undergoes profound structural and functional changes, reflected in the external skin appearance. Skin ageing is characterized by features such as wrinkling or loss of elasticity. Even if research advances have been done concerning the molecular mechanisms that underlie these changes, very few studies have been conducted concerning the structure stiffness of the skin organ as a whole. In this study, we showed, thanks to human skin reconstructs and the Japanese Medaka fish model, that biomechanics is a new biomarker of skin ageing. We revealed that global stiffness measurement by Atomic Force Microscopy, since modulated through ageing in these models, can be a new biomarker of skin ageing, and reflects the profound reorganization of the dermis extracellular matrix, as shown by Transmission Electron Microscopy. Moreover, our data unveiled that the Japanese Medaka fish could represent a highly relevant integrated model to study skin ageing in vivo.
Assuntos
Elasticidade , Modelos Animais , Envelhecimento da Pele/fisiologia , Pele/diagnóstico por imagem , Animais , Biomarcadores , Fenômenos Biomecânicos , Catalase/genética , Técnicas de Imagem por Elasticidade , Proteína Forkhead Box O1/genética , Glucuronidase/genética , Humanos , Proteínas Klotho , Microscopia de Força Atômica , Microscopia Eletrônica de Transmissão , Oryzias , RNA/metabolismo , Pele/metabolismo , Superóxido Dismutase/genética , beta-Galactosidase/metabolismoRESUMO
We describe here the evaluation of the cytotoxic efficacy of two platinum (II) complexes bearing an N-heterocyclic carbene (NHC) ligand, a pyridine ligand and bromide or iodide ligands on a panel of human metastatic cutaneous melanoma cell lines representing different genetic subsets including BRAF-inhibitor-resistant cell lines, namely A375, SK-MEL-28, MeWo, HMCB, A375-R, SK-MEL-5-R and 501MEL-R. Cisplatin and dacarbazine were also studied for comparison purposes. Remarkably, the iodine-labelled Pt-NHC complex strongly inhibited proliferation of all tested melanoma cells after 1-h exposure, likely due to its rapid uptake by melanoma cells. The mechanism of this inhibitory activity involves the formation of DNA double-strand breaks and apoptosis. Considering the intrinsic chemoresistance of metastatic melanoma cells of current systemic treatments, these findings are promising and could give research opportunities in the future to improve the prognosis of patients suffering from unresectable metastatic melanoma that are not eligible or that do not respond to the most effective drugs available to date, namely BRAF inhibitors and the anti-PD-1 monoclonal antibody (mAb).
Assuntos
Antineoplásicos/farmacologia , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Melanoma/tratamento farmacológico , Compostos Organoplatínicos/farmacologia , Neoplasias Cutâneas/tratamento farmacológico , Antineoplásicos/química , Antineoplásicos/farmacocinética , Morte Celular/efeitos dos fármacos , Morte Celular/fisiologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Quebras de DNA de Cadeia Dupla/efeitos dos fármacos , Dano ao DNA , Resistencia a Medicamentos Antineoplásicos/genética , Ensaios de Seleção de Medicamentos Antitumorais , Compostos Heterocíclicos/química , Compostos Heterocíclicos/farmacologia , Humanos , Melanoma/patologia , Metano/análogos & derivados , Metano/química , Compostos Organoplatínicos/química , Compostos Organoplatínicos/farmacocinética , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Proteínas Proto-Oncogênicas B-raf/genética , Neoplasias Cutâneas/patologia , Proteína bcl-X/metabolismo , Melanoma Maligno CutâneoAssuntos
Biomarcadores Tumorais , Expressão Gênica , Melanoma/etiologia , Melanoma/mortalidade , Neoplasias Cutâneas/etiologia , Neoplasias Cutâneas/mortalidade , Tetraspaninas/genética , Humanos , Imuno-Histoquímica , Melanoma/patologia , Melanoma/terapia , Mutação , Metástase Neoplásica , Estadiamento de Neoplasias , Prognóstico , Proteínas Proto-Oncogênicas B-raf/genética , RNA Mensageiro , Neoplasias Cutâneas/patologia , Neoplasias Cutâneas/terapia , Tetraspaninas/metabolismo , Melanoma Maligno CutâneoRESUMO
The intrinsic biomechanical properties of cancer cells remain poorly understood. To decipher whether cell stiffness modulation could increase melanoma cells' invasive capacity, we performed both in vitro and in vivo experiments exploring cell stiffness by atomic force microscopy (AFM). We correlated stiffness properties with cell morphology adaptation and the molecular mechanisms underlying epithelial-to-mesenchymal (EMT)-like phenotype switching. We found that melanoma cell stiffness reduction was systematically associated with the acquisition of invasive properties in cutaneous melanoma cell lines, human skin reconstructs, and Medaka fish developing spontaneous MAP-kinase-induced melanomas. We observed a systematic correlation of stiffness modulation with cell morphological changes towards mesenchymal characteristic gains. We accordingly found that inducing melanoma EMT switching by overexpressing the ZEB1 transcription factor, a major regulator of melanoma cell plasticity, was sufficient to decrease cell stiffness and transcriptionally induce tetraspanin-8-mediated dermal invasion. Moreover, ZEB1 expression correlated with Tspan8 expression in patient melanoma lesions. Our data suggest that intrinsic cell stiffness could be a highly relevant marker for human cutaneous melanoma development.
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The need for reliable biomarkers to predict clinical benefit from anti-PD1 treatment in metastatic melanoma (MM) patients remains unmet. Several parameters have been considered in the tumor environment or the blood, but none has yet achieved sufficient accuracy for routine clinical practice. Whole blood samples from MM patients receiving second-line anti-PD1 treatment (NCT02626065), collected longitudinally, were analyzed by flow cytometry to assess the immune cell subsets absolute numbers, the expression of immune checkpoints or ligands on T cells and the functionality of innate immune cells and T cells. Clinical response was assessed according to Progression-Free Survival (PFS) status at one-year following initiation of anti-PD1 (responders: PFS > 1 year; non-responders: PFS ≤ 1 year). At baseline, several phenotypic and functional alterations in blood immune cells were observed in MM patients compared to healthy donors, but only the proportion of polyfunctional memory CD4+ T cells was associated with response to anti-PD1. Under treatment, a decreased frequency of HVEM on CD4+ and CD8+ T cells after 3 months of treatment identified responding patients, whereas its receptor BTLA was not modulated. Both reduced proportion of CD69-expressing CD4+ and CD8+ T cells and increased number of polyfunctional blood memory T cells after 3 months of treatment were associated with response to anti-PD1. Of upmost importance, the combination of changes of all these markers accurately discriminated between responding and non-responding patients. These results suggest that drugs targeting HVEM/BTLA pathway may be of interest to improve anti-PD1 efficacy.
Assuntos
Melanoma , Receptor de Morte Celular Programada 1 , Receptores Imunológicos , Membro 14 de Receptores do Fator de Necrose Tumoral , Adulto , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/metabolismo , Inibidores de Checkpoint Imunológico/uso terapêutico , Inibidores de Checkpoint Imunológico/farmacologia , Melanoma/tratamento farmacológico , Melanoma/imunologia , Melanoma/patologia , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Receptor de Morte Celular Programada 1/metabolismo , Receptores Imunológicos/metabolismo , Membro 14 de Receptores do Fator de Necrose Tumoral/metabolismo , Resultado do TratamentoRESUMO
Cell plasticity sustains intra-tumor heterogeneity and treatment resistance in melanoma. Deciphering the transcriptional mechanisms governing reversible phenotypic transitions between proliferative/differentiated and invasive/stem-like states is required. Expression of the ZEB1 transcription factor is frequently activated in melanoma, where it fosters adaptive resistance to targeted therapies. Here, we performed a genome-wide characterization of ZEB1 transcriptional targets, by combining ChIP-sequencing and RNA-sequencing, upon phenotype switching in melanoma models. We identified and validated ZEB1 binding peaks in the promoter of key lineage-specific genes crucial for melanoma cell identity. Mechanistically, ZEB1 negatively regulates SOX10-MITF dependent proliferative/melanocytic programs and positively regulates AP-1 driven invasive and stem-like programs. Comparative analyses with breast carcinoma cells revealed lineage-specific ZEB1 binding, leading to the design of a more reliable melanoma-specific ZEB1 regulon. We then developed single-cell spatial multiplexed analyses to characterize melanoma cell states intra-tumoral heterogeneity in human melanoma samples. Combined with scRNA-Seq analyses, our findings confirmed increased ZEB1 expression in Neural-Crest-like cells and mesenchymal cells, underscoring its significance in vivo in both populations. Overall, our results define ZEB1 as a major transcriptional regulator of cell states transitions and provide a better understanding of lineage-specific transcriptional programs sustaining intra-tumor heterogeneity in melanoma.
Assuntos
Regulação Neoplásica da Expressão Gênica , Melanoma , Homeobox 1 de Ligação a E-box em Dedo de Zinco , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genética , Homeobox 1 de Ligação a E-box em Dedo de Zinco/metabolismo , Melanoma/genética , Melanoma/patologia , Melanoma/metabolismo , Humanos , Linhagem Celular Tumoral , Linhagem da Célula/genética , Fatores de Transcrição SOXE/genética , Fatores de Transcrição SOXE/metabolismo , Fator de Transcrição Associado à Microftalmia/genética , Fator de Transcrição Associado à Microftalmia/metabolismo , Camundongos , Animais , Proliferação de Células/genética , Transcrição Gênica/genéticaRESUMO
The outcome of cancer and autoimmunity is often dictated by the effector functions of CD4+ conventional T cells (Tconv). Although activation of the NF-κB signaling pathway has long been implicated in Tconv biology, the cell-autonomous roles of the separate NF-κB transcription-factor subunits are unknown. Here, we dissected the contributions of the canonical NF-κB subunits RelA and c-Rel to Tconv function. RelA, rather than c-Rel, regulated Tconv activation and cytokine production at steady-state and was required for polarization toward the TH17 lineage in vitro. Accordingly, RelA-deficient mice were fully protected against neuroinflammation in a model of multiple sclerosis due to defective transition to a pathogenic TH17 gene-expression program. Conversely, Tconv-restricted ablation of c-Rel impaired their function in the microenvironment of transplanted tumors, resulting in enhanced cancer burden. Moreover, Tconv required c-Rel for the response to PD-1-blockade therapy. Our data reveal distinct roles for canonical NF-κB subunits in different disease contexts, paving the way for subunit-targeted immunotherapies.
Assuntos
Esclerose Múltipla , Neoplasias , Animais , Camundongos , Linfócitos T CD4-Positivos , NF-kappa B , Transdução de Sinais , Microambiente Tumoral , Proteínas Proto-Oncogênicas c-rel/metabolismoRESUMO
A growing body of evidence suggests that the multifunctional protein E4F1 is involved in signaling pathways that play essential roles during normal development and tumorigenesis. We generated E4F1 conditional knockout mice to address E4F1 functions in vivo in newborn and adult skin. E4F1 inactivation in the entire skin or in the basal compartment of the epidermis induces skin homeostasis defects, as evidenced by transient hyperplasia in the interfollicular epithelium and alteration of keratinocyte differentiation, followed by loss of cellularity in the epidermis and severe skin ulcerations. E4F1 depletion alters clonogenic activity of epidermal stem cells (ESCs) ex vivo and ends in exhaustion of the ESC pool in vivo, indicating that the lesions observed in the E4F1 mutant skin result, at least in part, from cell-autonomous alterations in ESC maintenance. The clonogenic potential of E4F1 KO ESCs is rescued by Bmi1 overexpression or by Ink4a/Arf or p53 depletion. Skin phenotype of E4F1 KO mice is also delayed in animals with Ink4a/Arf and E4F1 compound gene deficiencies. Our data identify a regulatory axis essential for ESC-dependent skin homeostasis implicating E4F1 and the Bmi1-Arf-p53 pathway.
Assuntos
Proteínas de Ligação a DNA/fisiologia , Células Epidérmicas , Homeostase , Células-Tronco/fisiologia , Fatores de Transcrição/fisiologia , Fatores Etários , Animais , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Camundongos , Camundongos Knockout , Proteínas Nucleares/metabolismo , Fenótipo , Complexo Repressor Polycomb 1 , Proteínas Proto-Oncogênicas/metabolismo , Proteínas Repressoras/metabolismo , Células-Tronco/citologia , Proteína Supressora de Tumor p53/metabolismo , Ubiquitina-Proteína LigasesRESUMO
Metastatic melanoma patients carrying a BRAFV600 mutation can be treated with a combination of BRAF and MEK inhibitors (BRAFi/MEKi), but innate and acquired resistance invariably occurs. Predicting patient response to targeted therapies is crucial to guide clinical decision. We describe here the development of a highly efficient patient-derived xenograft model adapted to patient melanoma biopsies, using the avian embryo as a host (AVI-PDXTM ). In this in vivo paradigm, we depict a fast and reproducible tumor engraftment of patient samples within the embryonic skin, preserving key molecular and phenotypic features. We show that sensitivity and resistance to BRAFi/MEKi can be reliably modeled in these AVI-PDXTM , as well as synergies with other drugs. We further provide proof-of-concept that the AVI-PDXTM models the diversity of responses of melanoma patients to BRAFi/MEKi, within days, hence positioning it as a valuable tool for the design of personalized medicine assays and for the evaluation of novel combination strategies.
Assuntos
Melanoma , Proteínas Proto-Oncogênicas B-raf , Animais , Humanos , Proteínas Proto-Oncogênicas B-raf/genética , Melanoma/patologia , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Quinases de Proteína Quinase Ativadas por Mitógeno , Mutação , Modelos Animais de DoençasRESUMO
Melanoma is the deadliest form of skin cancer due to its propensity to metastasize. It arises from melanocytes, which are attached to keratinocytes within the basal epidermis. Here, we hypothesize that, in addition to melanocyte-intrinsic modifications, dysregulation of keratinocyte functions could initiate early-stage melanoma cell invasion. We identified the lysolipid sphingosine 1-phosphate (S1P) as a tumor paracrine signal from melanoma cells that modifies the keratinocyte transcriptome and reduces their adhesive properties, leading to tumor invasion. Mechanistically, tumor cell-derived S1P reduced E-cadherin expression in keratinocytes via S1P receptor dependent Snail and Slug activation. All of these effects were blocked by S1P2/3 antagonists. Importantly, we showed that epidermal E-cadherin expression was inversely correlated with the expression of the S1P-producing enzyme in neighboring tumors and the Breslow thickness in patients with early-stage melanoma. These findings support the notion that E-cadherin loss in the epidermis initiates the metastatic cascade in melanoma.
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Melanoma , Humanos , Melanoma/patologia , Esfingolipídeos/metabolismo , Comunicação Parácrina , Queratinócitos/metabolismo , Caderinas/metabolismo , Esfingosina/metabolismo , Lisofosfolipídeos/metabolismoRESUMO
Immunotherapies blocking negative immune checkpoints are now approved for the treatment of a growing number of cancers. However, even in metastatic melanoma, where sustained responses are observed, a significant number of patients still do not respond or display resistance. Increasing evidence indicates that non-genetic cancer cell-intrinsic alterations play a key role in resistance to therapies and immune evasion. Cancer cell plasticity, mainly associated with the epithelial-to-mesenchymal transition in carcinoma, relies on transcriptional, epigenetic or translational reprogramming. In melanoma, an EMT-like dedifferentiation process is characterized by the acquisition of invasive or neural crest stem cell-like features. Herein, we discuss recent findings on the specific roles of phenotypic reprogramming of melanoma cells in driving immune evasion and resistance to immunotherapies. The mechanisms by which dedifferentiated melanoma cells escape T cell lysis, mediate T cell exclusion or remodel the immune microenvironment will be detailed. The expanded knowledge on tumor cell plasticity in melanoma should contribute to the development of novel therapeutic combination strategies to further improve outcomes in this deadly metastatic cancer.
Assuntos
Melanoma , Transição Epitelial-Mesenquimal , Humanos , Imunoterapia , Melanoma/tratamento farmacológico , Crista Neural/patologia , Fenótipo , Microambiente TumoralRESUMO
BACKGROUND: The efficacy of immunotherapies in metastatic melanoma depends on a robust T cell infiltration. Oncogenic alterations of tumor cells have been associated to T cell exclusion. Identifying novel cancer cell-intrinsic non-genetic mechanisms of immune escape, the targeting of which would reinstate T cell recruitment, would allow to restore the response to anti-programmed cell death protein 1 (PD-1) antibody therapy. The epithelial-to-mesenchymal transition (EMT)-inducing transcription factor ZEB1 is a major regulator of melanoma cell plasticity, driving resistance to mitogen-activated protein kinase (MAPK) targeted therapies. We thus wondered whether ZEB1 signaling in melanoma cells may promote immune evasion and resistance to immunotherapy. METHODS: We evaluated the putative correlation between ZEB1 expression in melanoma cells and the composition of the immune infiltrate in a cohort of 60 human melanoma samples by combining transcriptomic (RNA-sequencing) and seven-color spatial multi-immunofluorescence analyses. Algorithm-based spatial reconstitution of tumors allowed the quantification of CD8+, CD4+ T cells number and their activation state (PD-1, Ki67). ZEB1 gain-of-function or loss-of-function approaches were then implemented in syngeneic melanoma mouse models, followed by monitoring of tumor growth, quantification of immune cell populations frequency and function by flow cytometry, cytokines secretion by multiplex analyses. Chromatin-immunoprecipitation was used to demonstrate the direct binding of this transcription factor on the promoters of cytokine-encoding genes. Finally, the sensitivity to anti-PD-1 antibody therapy upon ZEB1 gain-of-function or loss-of-function was evaluated. RESULTS: Combined spatial and transcriptomic analyses of the immune infiltrates in human melanoma samples demonstrated that ZEB1 expression in melanoma cells is associated with decreased CD8+ T cell infiltration, independently of ß-catenin pathway activation. ZEB1 ectopic expression in melanoma cells impairs CD8+ T cell recruitment in syngeneic mouse models, resulting in tumor immune evasion and resistance to immune checkpoint blockade. Mechanistically, we demonstrate that ZEB1 directly represses the secretion of T cell-attracting chemokines, including CXCL10. Finally, Zeb1 knock-out, by promoting CD8+ T cell infiltration, synergizes with anti-PD-1 antibody therapy in promoting tumor regression. CONCLUSIONS: We identify the ZEB1 transcription factor as a key determinant of melanoma immune escape, highlighting a previously unknown therapeutic target to increase efficacy of immunotherapy in melanoma. TRIAL REGISTRATION NUMBER: NCT02828202.
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Melanoma , Homeobox 1 de Ligação a E-box em Dedo de Zinco , Animais , Transição Epitelial-Mesenquimal/fisiologia , Humanos , Imunoterapia , Melanoma/tratamento farmacológico , Melanoma/genética , Melanoma/patologia , Camundongos , Oncogenes , Homeobox 1 de Ligação a E-box em Dedo de Zinco/genéticaRESUMO
The WHO classification identifies nine classes of melanocytic proliferations according to location, UV exposure, histological, and genetic features. Only a minority of lesions remain unclassified. We describe five cases that harbored either an ERBIN-RASGRF2 or an ATP2B4-RASGRF2 in-frame fusion transcript. These lesions were collected from different studies, unified only by the lack of identifiable known mutations, with a highly variable phenotype. One case was a large abdominal congenital nevus, three were slowly growing pigmented nodules, and the last was an ulcerated nodule arising on the site of a preexisting small nevus, known since childhood. The latter was diagnosed as a 4 mm thick melanoma with loss of BAP1 expression. The four other cases were compound, melanocytic proliferations with an unusual deep pattern of small dense nests of bland melanocytes encased in a fibrous background. The RASGRF2 fusion was confirmed by a break-apart FISH technique. Array CGH performed in three cases found non-recurrent secondary copy number alterations. Follow-up was uneventful. In silico analysis identified a single RASGRF2 fusion in the TCGA pan-cancer database, whereas RASGRF2 variants were stochastically distributed in all cancer subtypes.
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Melanócitos , Melanoma , Proteínas de Fusão Oncogênica , Neoplasias Cutâneas , Fatores ras de Troca de Nucleotídeo Guanina , Adulto , Criança , Feminino , Humanos , Masculino , Melanócitos/metabolismo , Melanócitos/patologia , Melanoma/genética , Melanoma/metabolismo , Melanoma/patologia , Pessoa de Meia-Idade , Proteínas de Fusão Oncogênica/genética , Proteínas de Fusão Oncogênica/metabolismo , Neoplasias Cutâneas/genética , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/patologia , Fatores ras de Troca de Nucleotídeo Guanina/genética , Fatores ras de Troca de Nucleotídeo Guanina/metabolismoRESUMO
T cell development proceeds under the influence of a network of transcription factors (TFs). The precise role of Zeb1, a member of this network, remains unclear. Here, we report that Zeb1 expression is induced early during T cell development in CD4-CD8- double-negative (DN) stage 2 (DN2). Zeb1 expression was further increased in the CD4+CD8+ double-positive (DP) stage before decreasing in more mature T cell subsets. We performed an exhaustive characterization of T cells in Cellophane mice that bear Zeb1 hypomorphic mutations. The Zeb1 mutation profoundly affected all thymic subsets, especially DN2 and DP cells. Zeb1 promoted the survival and proliferation of both cell populations in a cell-intrinsic manner. In the periphery of Cellophane mice, the number of conventional T cells was near normal, but invariant NKT cells, NK1.1+ γδ T cells and Ly49+ CD8 T cells were virtually absent. This suggested that Zeb1 regulates the development of unconventional T cell types from DP progenitors. A transcriptomic analysis of WT and Cellophane DP cells revealed that Zeb1 regulated the expression of multiple genes involved in the cell cycle and TCR signaling, which possibly occurred in cooperation with Tcf1 and Heb. Indeed, Cellophane DP cells displayed stronger signaling than WT DP cells upon TCR engagement in terms of the calcium response, phosphorylation events, and expression of early genes. Thus, Zeb1 is a key regulator of the cell cycle and TCR signaling during thymic T cell development. We propose that thymocyte selection is perturbed in Zeb1-mutated mice in a way that does not allow the survival of unconventional T cell subsets.
Assuntos
Receptores de Antígenos de Linfócitos T alfa-beta , Subpopulações de Linfócitos T , Animais , Diferenciação Celular , Proliferação de Células , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Receptores de Antígenos de Linfócitos T alfa-beta/genética , Transdução de Sinais/genética , TimoRESUMO
Transcription factors, extensively described for their role in epithelial-mesenchymal transition (EMT-TFs) in epithelial cells, also display essential functions in the melanocyte lineage. Recent evidence has shown specific expression patterns and functions of these EMT-TFs in neural crest-derived melanoma compared to carcinoma. Herein, we present an update of the specific roles of EMT-TFs in melanocyte differentiation and melanoma progression. As major regulators of phenotype switching between differentiated/proliferative and neural crest stem cell-like/invasive states, these factors appear as major drivers of intra-tumor heterogeneity and resistance to treatment in melanoma, which opens new avenues in terms of therapeutic targeting.
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Due to its high proclivity to metastasize, and despite the recent development of targeted and immune therapy strategies, melanoma is still the deadliest form of skin cancer. Therefore, understanding the molecular mechanisms underlying melanoma invasion remains crucial. We previously characterized Tspan8 for its ability to prompt melanoma cell detachment from their microenvironment and trigger melanoma cell invasiveness, but the signaling events by which Tspan8 regulates the invasion process still remain unknown. Here, we demonstrated that ß-catenin stabilization is a molecular signal subsequent to the onset of Tspan8 expression, and that, in turn, ß-catenin triggers the direct transcriptional activation of Tspan8 expression, leading to melanoma invasion. Moreover, we showed that ß-catenin activation systematically correlates with a high expression of Tspan8 protein in melanoma lesions from transgenic Nras; bcat* mice, as well as in deep penetrating naevi, a type of human pre-melanoma neoplasm characterized by a combined activation of ß-catenin and MAP kinase signaling. Overall, our data suggest that ß-catenin and Tspan8 are part of a positive feedback loop, which sustains a high Tspan8 expression level, conferring to melanoma cells the invasive properties required for tumor progression and dissemination.
Assuntos
Melanoma/metabolismo , Melanoma/patologia , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/patologia , Tetraspaninas/metabolismo , beta Catenina/metabolismo , Animais , Retroalimentação Fisiológica , Regulação Neoplásica da Expressão Gênica , Humanos , Melanoma/genética , Camundongos Transgênicos , Regiões Promotoras Genéticas , Estabilidade Proteica , Neoplasias Cutâneas/genética , Tetraspaninas/genética , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto , beta Catenina/genética , Melanoma Maligno CutâneoRESUMO
Phenotypic plasticity and subsequent generation of intratumoral heterogeneity underly key traits in malignant melanoma such as drug resistance and metastasis. Melanoma plasticity promotes a switch between proliferative and invasive phenotypes characterized by different transcriptional programs of which MITF is a critical regulator. Here, we show that the acid ceramidase ASAH1, which controls sphingolipid metabolism, acted as a rheostat of the phenotypic switch in melanoma cells. Low ASAH1 expression was associated with an invasive behavior mediated by activation of the integrin alphavbeta5-FAK signaling cascade. In line with that, human melanoma biopsies revealed heterogeneous staining of ASAH1 and low ASAH1 expression at the melanoma invasive front. We also identified ASAH1 as a new target of MITF, thereby involving MITF in the regulation of sphingolipid metabolism. Together, our findings provide new cues to the mechanisms underlying the phenotypic plasticity of melanoma cells and identify new anti-metastatic targets.
Assuntos
Ceramidase Ácida/genética , Proliferação de Células/genética , Melanoma/genética , Fator de Transcrição Associado à Microftalmia/genética , Linhagem Celular Tumoral , Feminino , Quinase 1 de Adesão Focal/genética , Regulação Neoplásica da Expressão Gênica , Humanos , Masculino , Melanoma/patologia , Invasividade Neoplásica/genética , Proteínas Proto-Oncogênicas B-raf , Receptores de Vitronectina/genética , Transdução de SinaisRESUMO
Several mechanisms have been described to elucidate the emergence of resistance to MAPK inhibitors in melanoma and there is a crucial need for biomarkers to identify patients who are likely to achieve a better and long-lasting response to BRAF inhibitors therapy. In this study, we developed a targeted approach combining both mRNA and DNA alterations analysis focusing on relevant gene alterations involved in acquired BRAF inhibitor resistance. We collected baseline tumor samples from 64 melanoma patients at BRAF inhibitor treatment initiation and showed that the presence, prior to treatment, of mRNA over-expression of genes' subset was significantly associated with improved progression free survival and overall survival. The presence of DNA alterations was in favor of better overall survival. The genomic analysis of relapsed-matched tumor samples from 20 patients allowed us to uncover the largest landscape of resistance mechanisms reported to date as at least one resistance mechanism was identified for each patient studied. Alterations in RB1 have been most frequent and hence represent an important additional acquired resistance mechanism. Our targeted genomic analysis emerges as a relevant tool in clinical practice to identify those patients who are more likely to achieve durable response to targeted therapies and to exhaustively describe the spectrum of resistance mechanisms. Our approach can be adapted to new targeted therapies by including newly identified genetic alterations.