RESUMEN
Melanoma is an aggressive cancer with poor prognosis, requiring personalized management of advanced stages and establishment of molecular markers. Melanomas derive from melanocytes, which specifically express tyrosinase, the rate-limiting enzyme of melanin-synthesis. We demonstrate that melanomas with high levels of DNp73, a cancer-specific variant of the p53 family member p73 and driver of melanoma progression show, in contrast to their less-aggressive low-DNp73 counterparts, hypopigmentation in vivo. Mechanistically, reduced melanin-synthesis is mediated by a DNp73-activated IGF1R/PI3K/AKT axis leading to tyrosinase ER-arrest and proteasomal degradation. Tyrosinase loss triggers reactivation of the EMT signaling cascade, a mesenchymal-like cell phenotype and increased invasiveness. DNp73-induced depigmentation, Slug increase and changes in cell motility are recapitulated in neural crest-derived melanophores of Xenopus embryos, underscoring a previously unnoticed physiological role of tyrosinase as EMT inhibitor. This data provides a mechanism of hypopigmentation accompanying cancer progression, which can be exploited in precision diagnosis of patients with melanoma-associated hypopigmentation (MAH), currently seen as a favorable prognostic factor. The DNp73/IGF1R/Slug signature in colorless lesions might aid to clinically discriminate between patients with MAH-associated metastatic disease and those, where MAH is indeed a sign of regression.
Asunto(s)
Transición Epitelial-Mesenquimal , Hipopigmentación/enzimología , Melaninas/metabolismo , Melanocitos/enzimología , Melanoma/enzimología , Monofenol Monooxigenasa/metabolismo , Neoplasias Cutáneas/enzimología , Proteína Tumoral p73/metabolismo , Animales , Línea Celular Tumoral , Movimiento Celular , Humanos , Hipopigmentación/genética , Hipopigmentación/patología , Melanocitos/patología , Melanoma/genética , Melanoma/patología , Ratones , Monofenol Monooxigenasa/genética , Invasividad Neoplásica , Complejo de la Endopetidasa Proteasomal/metabolismo , Proteolisis , Especies Reactivas de Oxígeno/metabolismo , Receptor IGF Tipo 1 , Receptores de Somatomedina/genética , Receptores de Somatomedina/metabolismo , Transducción de Señal , Neoplasias Cutáneas/genética , Neoplasias Cutáneas/patología , Factores de Transcripción de la Familia Snail/genética , Factores de Transcripción de la Familia Snail/metabolismo , Proteína Tumoral p73/genética , Xenopus laevisRESUMEN
Dissemination of cancer cells from primary tumors is the key event in metastasis, but specific determinants are widely unknown. Here, we show that DNp73, an inhibitor of the p53 tumor suppressor family, drives migration and invasion of nonmetastatic melanoma cells. Knockdown of endogenous DNp73 reduces this behavior in highly metastatic cell lines. Tumor xenografts expressing DNp73 show a higher ability to invade and metastasize, while growth remains unaffected. DNp73 facilitates an EMT-like phenotype with loss of E-cadherin and Slug upregulation. We provide mechanistic insight toward regulation of LIMA1/EPLIN by p73/DNp73 and demonstrate a direct link between the DNp73-EPLIN axis and IGF1R-AKT/STAT3 activation. These findings establish initiation of the invasion-metastasis cascade via EPLIN-dependent IGF1R regulation as major activity of DNp73.
Asunto(s)
Proteínas del Citoesqueleto/metabolismo , Proteínas de Unión al ADN/fisiología , Regulación Neoplásica de la Expresión Génica , Melanoma/metabolismo , Proteínas Nucleares/fisiología , Receptor IGF Tipo 1/metabolismo , Neoplasias Cutáneas/metabolismo , Proteínas Supresoras de Tumor/fisiología , Animales , Línea Celular Tumoral , Proteínas de Unión al ADN/metabolismo , Femenino , Perfilación de la Expresión Génica , Humanos , Melanoma/patología , Ratones , Ratones Desnudos , Metástasis de la Neoplasia , Trasplante de Neoplasias , Proteínas Nucleares/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Factor de Transcripción STAT3/metabolismo , Transducción de Señal , Neoplasias Cutáneas/patología , Proteína Tumoral p73 , Proteína p53 Supresora de Tumor/metabolismo , Proteínas Supresoras de Tumor/metabolismoRESUMEN
Resistance to anti-neoplastic agents is the major cause of therapy failure, leading to disease recurrence and metastasis. E2F1 is a strong inducer of apoptosis in response to DNA damage through its capacity to activate p53/p73 death pathways. Recent evidence, however, showed that E2F1, which is aberrantly expressed in advanced malignant melanomas together with antagonistic p73 family members, drives cancer progression. Investigating mechanisms responsible for dysregulated E2F1 losing its apoptotic function, we searched for genomic signatures in primary and late clinical tumor stages to allow the prediction of downstream effectors associated with apoptosis resistance and survival of aggressive melanoma cells. We identified miR-205 as specific target of p73 and found that upon genotoxic stress, its expression is sufficiently abrogated by endogenous DNp73. Significantly, metastatic cells can be rescued from drug resistance by selective knockdown of DNp73 or overexpression of miR-205 in p73-depleted cells, leading to increased apoptosis and the reduction of tumor growth in vivo. Our data delineate an autoregulatory circuit, involving high levels of E2F1 and DNp73 to downregulate miR-205, which, in turn, controls E2F1 accumulation. Finally, drug resistance associated to this genetic signature is mediated by removing the inhibitory effect of miR-205 on the expression of Bcl-2 and the ATP-binding cassette transporters A2 (ABCA2) and A5 (ABCA5) related to multi-drug resistance and malignant progression. These results define the E2F1-p73/DNp73-miR-205 axis as a crucial mechanism for chemoresistance and, thus, as a target for metastasis prevention.
Asunto(s)
Proteínas de Unión al ADN/metabolismo , Resistencia a Antineoplásicos , Factor de Transcripción E2F1/metabolismo , MicroARNs/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Transportadoras de Casetes de Unión a ATP/genética , Transportadoras de Casetes de Unión a ATP/metabolismo , Adenoviridae/genética , Adenoviridae/metabolismo , Antineoplásicos/farmacología , Apoptosis , Secuencia de Bases , Transformación Celular Neoplásica/metabolismo , Transformación Celular Neoplásica/patología , Cicloheximida/farmacología , Proteínas de Unión al ADN/antagonistas & inhibidores , Proteínas de Unión al ADN/genética , Factor de Transcripción E2F1/genética , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Vectores Genéticos , Células HEK293 , Humanos , Melanoma/metabolismo , Melanoma/patología , MicroARNs/genética , Metástasis de la Neoplasia/patología , Estadificación de Neoplasias , Proteínas Nucleares/antagonistas & inhibidores , Proteínas Nucleares/genética , Proteínas Proto-Oncogénicas c-bcl-2/genética , Proteína Tumoral p73 , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo , Proteínas Supresoras de Tumor/antagonistas & inhibidores , Proteínas Supresoras de Tumor/genéticaRESUMEN
Malignant melanoma of the skin is one of the most aggressive human cancers with increasing incidence, despite efforts to improve primary prevention. In particular, the prognosis of patients at late stages of the disease has not significantly improved in the last three decades, because systemic therapies have proven disappointing. Thus, metastatic melanoma continues to be a daunting clinical problem. The increasingly high rates of lethal outcome associated with advanced melanoma rely on the acquisition of invasiveness, early metastatic dissemination of tumor cells from their primary sites, and generation of chemoresistance as a consequence of alteration of key molecules involved in the regulation of cell survival. Thus far, extensive studies have been conducted to understand the molecular mechanisms that drive tumor progression, but the specific requirements underlying the aggressive behavior are still widely unknown. Understanding the determinants of this process is key to unveiling its dynamics, especially those that promote invasiveness, and may open new routes for the development of therapeutic strategies that control metastatic spread, and eventually the prevention of life-threatening metastases. Here, we review recent advances on molecular aspects, particularly of E2F1 transcription factor function, in the context of patient data, and discuss the implications for targeting melanoma cells when they begin to invade and metastasize.
Asunto(s)
Factor de Transcripción E2F1/fisiología , Melanoma/patología , Invasividad Neoplásica , Biomarcadores de Tumor , Progresión de la Enfermedad , Humanos , Melanoma/fisiopatología , Pronóstico , Medición de RiesgoRESUMEN
Metastases are responsible for cancer deaths, but the molecular alterations leading to tumor progression are unclear. Overexpression of the E2F1 transcription factor is common in high-grade tumors that are associated with poor patient survival. To investigate the association of enhanced E2F1 activity with aggressive phenotype, we performed a gene-specific silencing approach in a metastatic melanoma model. Knockdown of endogenous E2F1 via E2F1 small hairpin RNA (shRNA) expression increased E-cadherin expression of metastatic SK-Mel-147 melanoma cells and reduced their invasive potential but not their proliferative activity. Although growth rates of SK-Mel-147 and SK-Mel-103 xenograft tumors expressing E2F1 shRNA or control shRNA were similar, mice implanted with cells expressing E2F1 shRNA had a smaller area of metastases per lung than control mice (n = 3 mice per group; 5% vs 46%, difference = 41%, 95% confidence interval = 15% to 67%; P = .01; one-way analysis of variance). We identified epidermal growth factor receptor as a direct target of E2F1 and demonstrated that inhibition of receptor signaling abrogates E2F1-induced invasiveness, emphasizing the importance of the E2F1-epidermal growth factor receptor interaction as a driving force in melanoma progression that may serve as a paradigm for E2F1-induced metastasis in other human cancers.
Asunto(s)
Factor de Transcripción E2F1/metabolismo , Receptores ErbB/metabolismo , Melanoma/secundario , Neoplasias Cutáneas/patología , Progresión de la Enfermedad , Regulación Neoplásica de la Expresión Génica , Humanos , Melanoma/metabolismo , Invasividad Neoplásica , Neoplasias Cutáneas/metabolismoRESUMEN
The E2F1 transcription factor enhances apoptosis by DNA damage in tumors lacking p53. To elucidate the mechanism of a potential cooperation between E2F1 and chemotherapy, whole-genome microarrays of chemoresistant tumor cell lines were performed focusing on the identification of cooperation response genes (CRG). This gene class is defined by a synergistic expression response upon endogenous E2F1 activation and drug treatment. Cluster analysis revealed an expression pattern of CRGs similar to E2F1 mono-therapy, suggesting that chemotherapeutics enhance E2F1-dependent gene expression at the transcriptional level. Using this approach as a tool to explore E2F1-driven gene expression in response to anticancer drugs, we identified novel apoptosis genes such as the tumor suppressor TIEG1/KLF10 as direct E2F1 targets. We show that TIEG1/KLF10 is transcriptionally activated by E2F1 and crucial for E2F1-mediated chemosensitization of cancer cells. Our results provide a broader picture of E2F1-regulated genes in conjunction with cytotoxic treatment that allows the design of more rational therapeutics.