RESUMO
BACKGROUND: The main drawback of BRAF/MEK inhibitors (BRAF/MEKi)-based targeted therapy in the management of BRAF-mutated cutaneous metastatic melanoma (MM) is the development of therapeutic resistance. We aimed to assess in this context the role of mTORC2, a signaling complex defined by the presence of the essential RICTOR subunit, regarded as an oncogenic driver in several tumor types, including MM. METHODS: After analyzing The Cancer Genome Atlas MM patients' database to explore both overall survival and molecular signatures as a function of intra-tumor RICTOR levels, we investigated the effects of RICTOR downregulation in BRAFV600E MM cell lines on their response to BRAF/MEKi. We performed proteomic screening to identify proteins modulated by changes in RICTOR expression, and Seahorse analysis to evaluate the effects of RICTOR depletion on mitochondrial respiration. The combination of BRAFi with drugs targeting proteins and processes emerged in the proteomic screening was carried out on RICTOR-deficient cells in vitro and in a xenograft setting in vivo. RESULTS: Low RICTOR levels in BRAF-mutated MM correlate with a worse clinical outcome. Gene Set Enrichment Analysis of low-RICTOR tumors display gene signatures suggestive of activation of the mitochondrial Electron Transport Chain (ETC) energy production. RICTOR-deficient BRAFV600E cells are intrinsically tolerant to BRAF/MEKi and anticipate the onset of resistance to BRAFi upon prolonged drug exposure. Moreover, in drug-naïve cells we observed a decline in RICTOR expression shortly after BRAFi exposure. In RICTOR-depleted cells, both mitochondrial respiration and expression of nicotinamide phosphoribosyltransferase (NAMPT) are enhanced, and their pharmacological inhibition restores sensitivity to BRAFi. CONCLUSIONS: Our work unveils an unforeseen tumor-suppressing role for mTORC2 in the early adaptation phase of BRAFV600E melanoma cells to targeted therapy and identifies the NAMPT-ETC axis as a potential therapeutic vulnerability of low RICTOR tumors. Importantly, our findings indicate that the evaluation of intra-tumor RICTOR levels has a prognostic value in metastatic melanoma and may help to guide therapeutic strategies in a personalized manner.
Assuntos
Resistencia a Medicamentos Antineoplásicos , Alvo Mecanístico do Complexo 2 de Rapamicina , Melanoma , Inibidores de Proteínas Quinases , Proteínas Proto-Oncogênicas B-raf , Proteína Companheira de mTOR Insensível à Rapamicina , Animais , Humanos , Camundongos , Linhagem Celular Tumoral , Regulação para Baixo , Resistencia a Medicamentos Antineoplásicos/genética , Regulação Neoplásica da Expressão Gênica , Alvo Mecanístico do Complexo 2 de Rapamicina/metabolismo , Alvo Mecanístico do Complexo 2 de Rapamicina/genética , Melanoma/genética , Melanoma/tratamento farmacológico , Melanoma/metabolismo , Melanoma/patologia , Mutação , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Proteômica/métodos , Proteínas Proto-Oncogênicas B-raf/antagonistas & inibidores , Proteínas Proto-Oncogênicas B-raf/genética , Proteína Companheira de mTOR Insensível à Rapamicina/metabolismo , Proteína Companheira de mTOR Insensível à Rapamicina/genética , Ensaios Antitumorais Modelo de Xenoenxerto , MAP Quinase Quinase Quinases/antagonistas & inibidoresRESUMO
MicroRNAs (miRNAs) are small, non-coding RNAs about 22 nucleotides in length that regulate the expression of target genes post-transcriptionally, and are highly involved in cancer progression. They are able to impact a variety of cell processes such as proliferation, apoptosis and differentiation and can consequently control tumor initiation, tumor progression and metastasis formation. miRNAs can regulate, at the same time, metabolic gene expression which, in turn, influences relevant traits of malignancy such as cell adhesion, migration and invasion. Since the interaction between metabolism and adhesion or cell movement has not, to date, been well understood, in this review, we will specifically focus on miRNA alterations that can interfere with some metabolic processes leading to the modulation of cancer cell movement. In addition, we will analyze the signaling pathways connecting metabolism and adhesion/migration, alterations that often affect cancer cell dissemination and metastasis formation.
Assuntos
MicroRNAs , Neoplasias , Adesão Celular/genética , Movimento Celular/genética , Glucose , Glutamina/genética , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Neoplasias/genética , Neoplasias/patologiaRESUMO
Cancer is a multistep disease based on crucial interactions between tumor cells and the microenvironment (extracellular matrix and stroma/immune cells). In fact, during dissemination, tumor cells have to escape from the primary tumor mass, cross the basal membrane, interact with endothelial cells to enter blood vessels (intravasation), survive in the bloodstream, get in contact with endothelial cells again to exit the bloodstream (extravasation) and seed in distant organs. Interactions between tumor and stroma cells are strongly coordinated by microRNAs (miRNAs), small non-coding RNAs able to silence protein coding genes by binding to specific recognition sites, mostly located at the 3' UTR of mature mRNAs. Relevantly, miRNA expression is often altered (overexpression or downregulation) in tumor cells and influenced by stroma cells. At the same time, miRNAs are abundant and essential in stroma cells during tumor cell dissemination and their expression is influenced by tumor cells. In fact, for instance, conditional ablation of Dicer in the endothelium of tumor bearing-mice leads to reduced tumor growth and microvessel density. In this review, we specifically focus on the role of miRNAs in endothelial cells regarding their positive or negative intervention on tumor angiogenesis or lymphoangiogenesis or when tumor cells detach from the tumor mass and intravasate or extravasate in/out of the blood vessels. Examples of pro-angiogenic miRNAs are miR-9 or miR-494, often overexpressed in tumors, which accumulate in tumor cell microvescicles and, therefore, get transferred to endothelial cells where they induce migration and angiogenesis. Differently, miR-200 and miR-128 are often downregulated in tumors and inhibit angiogenesis and lymphoangiogenesis. Instead, miR-126 controls intravasation while miR-146a, miR-214, miR-148b govern extravasation, in a positive or negative manner. Finally, at the end, we summarize opportunities for therapeutic interventions based on miRNAs acting on endothelial cells.
Assuntos
Comunicação Celular/genética , Células Endoteliais/metabolismo , Regulação Neoplásica da Expressão Gênica , MicroRNAs/genética , Neoplasias/etiologia , Neoplasias/metabolismo , Microambiente Tumoral/genética , Animais , Comunicação Celular/imunologia , Progressão da Doença , Humanos , Estadiamento de Neoplasias , Neoplasias/patologia , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Microambiente Tumoral/imunologiaRESUMO
Telomerase reverse transcriptase (TERT) and telomerase RNA component (TERC) constitute the core telomerase enzyme that maintains the length of telomeres. Telomere maintenance is affected in a broad range of cancer and degenerative disorders. Taking advantage of gain- and loss-of-function approaches, we show that Argonaute 2 (AGO2) promotes telomerase activity and stimulates the association between TERT and TERC AGO2 depletion results in shorter telomeres as well as in lower proliferation rates in vitro and in vivo We also demonstrate that AGO2 interacts with TERC and with a newly identified sRNA (terc-sRNA), arising from the H/ACA box of TERC Notably, terc-sRNA is sufficient to enhance telomerase activity when overexpressed. Analyses of sRNA-Seq datasets show that terc-sRNA is detected in primary human tissues and increases in tumors as compared to control tissues. Collectively, these data uncover a new layer of complexity in the regulation of telomerase activity by AGO2 and might lay the foundation for new therapeutic targets in tumors and telomere diseases.
Assuntos
Proteínas Argonautas/metabolismo , RNA/genética , RNA/metabolismo , Telomerase/metabolismo , Animais , Proteínas Argonautas/genética , Linhagem Celular Tumoral , Proliferação de Células , Modelos Animais de Doenças , Ativação Enzimática , Expressão Gênica , Loci Gênicos , Xenoenxertos , Humanos , Camundongos , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/patologia , Conformação de Ácido Nucleico , Ligação Proteica , RNA/química , Telomerase/química , Telomerase/genéticaRESUMO
We previously demonstrated that miR-214 is upregulated in malignant melanomas and triple-negative breast tumors and promotes metastatic dissemination by affecting a complex pathway including the anti-metastatic miR-148b. Importantly, tumor dissemination could be reduced by blocking miR-214 function or increasing miR-148b expression or by simultaneous interventions. Based on this evidence, with the intent to explore the role of miR-214 as a target for therapy, we evaluated the capability of new chemically modified anti-miR-214, R97/R98, to inhibit miR-214 coordinated metastatic traits. Relevantly, when melanoma or breast cancer cells were transfected with R97/R98, anti-miR-214 reduced miR-214 expression and impaired transendothelial migration were observed. Noteworthy, when the same cells were injected in the tail vein of mice, cell extravasation and metastatic nodule formation in lungs were strongly reduced. Thus, suggesting that R97/R98 anti-miR-214 oligonucleotides were able to inhibit tumor cell escaping through the endothelium. More importantly, when R97/R98 anti-miR-214 compounds were systemically delivered to mice carrying melanomas or breast or neuroendocrine pancreatic cancers, a reduced number of circulating tumor cells and lung or lymph node metastasis formation were detected. Similar results were also obtained when AAV8-miR-214 sponges were used in neuroendocrine pancreatic tumors. Based on this evidence, we propose miR-214 as a promising target for anti-metastatic therapies.
Assuntos
Antagomirs/administração & dosagem , MicroRNAs/genética , Neoplasias/tratamento farmacológico , Regulação para Cima/efeitos dos fármacos , Animais , Antagomirs/farmacologia , Neoplasias da Mama/tratamento farmacológico , Neoplasias da Mama/genética , Carcinoma Neuroendócrino/tratamento farmacológico , Carcinoma Neuroendócrino/genética , Linhagem Celular Tumoral , Progressão da Doença , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Inativação Gênica , Humanos , Melanoma/tratamento farmacológico , Melanoma/genética , Camundongos , MicroRNAs/antagonistas & inibidores , Metástase Neoplásica/tratamento farmacológico , Neoplasias/genética , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/genética , Transfecção , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
BACKGROUND: Biological research increasingly relies on network models to study complex phenomena. Signal Transduction Pathways are molecular circuits that model how cells receive, process, and respond to information from the environment providing snapshots of the overall cell dynamics. Most of the attempts to reconstruct signal transduction pathways are limited to single regulator networks including only genes/proteins. However, networks involving a single type of regulator and neglecting transcriptional and post-transcriptional regulations mediated by transcription factors and microRNAs, respectively, may not fully reveal the complex regulatory mechanisms of a cell. We observed a lack of computational instruments supporting explorative analysis on this type of three-component signal transduction pathways. RESULTS: We have developed CyTRANSFINDER, a new Cytoscape plugin able to infer three-component signal transduction pathways based on user defined regulatory patterns and including miRNAs, TFs and genes. Since CyTRANSFINDER has been designed to support exploratory analysis, it does not rely on expression data. To show the potential of the plugin we have applied it in a study of two miRNAs that are particularly relevant in human melanoma progression, miR-146a and miR-214. CONCLUSIONS: CyTRANSFINDER supports the reconstruction of small signal transduction pathways among groups of genes. Results obtained from its use in a real case study have been analyzed and validated through both literature data and preliminary wet-lab experiments, showing the potential of this tool when performing exploratory analysis.
Assuntos
MicroRNAs/genética , Transdução de Sinais , Progressão da Doença , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Humanos , Melanoma/genética , MicroRNAs/metabolismo , Reprodutibilidade dos Testes , Fatores de Transcrição/genética , Fatores de Transcrição/metabolismoRESUMO
Malignant melanoma is fatal in its metastatic stage. It is therefore essential to unravel the molecular mechanisms that govern disease progression to metastasis. MicroRNAs (miRs) are endogenous non-coding RNAs involved in tumourigenesis. Using a melanoma progression model, we identified a novel pathway controlled by miR-214 that coordinates metastatic capability. Pathway components include TFAP2C, homologue of a well-established melanoma tumour suppressor, the adhesion receptor ITGA3 and multiple surface molecules. Modulation of miR-214 influences in vitro tumour cell movement and survival to anoikis as well as extravasation from blood vessels and lung metastasis formation in vivo. Considering that miR-214 is known to be highly expressed in human melanomas, our data suggest a critical role for this miRNA in disease progression and the establishment of distant metastases.
Assuntos
Regulação da Expressão Gênica , Melanoma/patologia , Melanoma/secundário , MicroRNAs/metabolismo , Metástase Neoplásica/patologia , Fator de Transcrição AP-2/biossíntese , Animais , Movimento Celular , Proliferação de Células , Sobrevivência Celular , Células Cultivadas , Humanos , Integrinas/metabolismo , Pulmão/patologia , Neoplasias Pulmonares/patologia , Camundongos , MicroRNAs/genéticaRESUMO
In an attempt to reveal deregulated miRNAs associated with the progression of carcinomas developed in BALB-neuT transgenic mice, we found increased expression of miR-135b during malignancy. Relevantly, we observed that miR-135b is up-regulated in basal or normal-like human breast cancers, and it correlates with patient survival and early metastatization. Therefore, we investigated its biological functions by modulating its expression (up- or down-regulation) in mammary tumor cells. Although no effect was observed on proliferation in cell culture and in orthotopically injected mice, miR-135b was able to control cancer cell stemness in a mammosphere assay, anchorage-independent growth in vitro, and lung cancer cell dissemination in mice after tail vein injections. Focusing on the miR-135b molecular mechanism, we observed that miR-135b controls malignancy via its direct targets, midline 1 (MID1) and mitochondrial carrier homolog 2 (MTCH2), as proved by biochemical and functional rescuing/phenocopying experiments. Consistently, an anti-correlation between miR-135b and MID1 or MTCH2 was found in human primary tumor samples. In conclusion, our research led us to the identification of miR-135b and its targets, MID1 and MTCH2, as relevant coordinators of mammary gland tumor progression.
Assuntos
Neoplasias Mamárias Experimentais/metabolismo , MicroRNAs/fisiologia , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , Proteínas/metabolismo , Animais , Neoplasias da Mama/metabolismo , Proliferação de Células , Progressão da Doença , Feminino , Perfilação da Expressão Gênica/métodos , Regulação Neoplásica da Expressão Gênica/fisiologia , Genes erbB-2 , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/secundário , Neoplasias Mamárias Experimentais/genética , Neoplasias Mamárias Experimentais/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , MicroRNAs/biossíntese , MicroRNAs/genética , Invasividade Neoplásica , Proteínas de Neoplasias/metabolismo , Células-Tronco Neoplásicas/metabolismo , RNA Neoplásico/genética , Células Tumorais Cultivadas , Ubiquitina-Proteína Ligases , Regulação para Cima/fisiologiaRESUMO
Breast cancer is often fatal during its metastatic dissemination. To unravel the role of microRNAs (miRs) during malignancy, we analyzed miR expression in 77 primary breast carcinomas and identified 16 relapse-associated miRs that correlate with survival and/or distinguish tumor subtypes in different datasets. Among them, miR-148b, down-regulated in aggressive breast tumors, was found to be a major coordinator of malignancy. In fact, it is able to oppose various steps of tumor progression when overexpressed in cell lines by influencing invasion, survival to anoikis, extravasation, lung metastasis formation, and chemotherapy response. miR-148b controls malignancy by coordinating a novel pathway involving over 130 genes and, in particular, it directly targets players of the integrin signaling, such as ITGA5, ROCK1, PIK3CA/p110α, and NRAS, as well as CSF1, a growth factor for stroma cells. Our findings reveal the importance of the identified 16 miRs for disease outcome predictions and suggest a critical role for miR-148b in the control of breast cancer progression.
Assuntos
Neoplasias da Mama/metabolismo , Integrina alfa5/biossíntese , Fator Estimulador de Colônias de Macrófagos/biossíntese , MicroRNAs/metabolismo , Proteína Oncogênica p21(ras)/biossíntese , Fosfatidilinositol 3-Quinases/biossíntese , RNA Neoplásico/metabolismo , Quinases Associadas a rho/biossíntese , Adulto , Idoso , Idoso de 80 Anos ou mais , Neoplasias da Mama/genética , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Classe I de Fosfatidilinositol 3-Quinases , Progressão da Doença , Feminino , Humanos , Integrina alfa5/genética , Fator Estimulador de Colônias de Macrófagos/genética , MicroRNAs/genética , Pessoa de Meia-Idade , Invasividade Neoplásica , Proteína Oncogênica p21(ras)/genética , Fosfatidilinositol 3-Quinases/genética , RNA Neoplásico/genética , Quinases Associadas a rho/genéticaRESUMO
Morgana is a ubiquitous HSP90 co-chaperone protein coded by the CHORDC1 gene. Morgana heterozygous mice develop with age a myeloid malignancy resembling human atypical myeloid leukemia (aCML), now renamed MDS/MPN with neutrophilia. Patients affected by this pathology exhibit low Morgana levels in the bone marrow (BM), suggesting that Morgana downregulation plays a causative role in the human malignancy. A decrease in Morgana expression levels is also evident in the BM of a subgroup of Philadelphia-positive (Ph+) chronic myeloid leukemia (CML) patients showing resistance or an incomplete response to imatinib. Despite the relevance of these data, the mechanism through which Morgana expression is downregulated in patients' bone marrow remains unclear. In this study, we investigated the possibility that Morgana expression is regulated by miRNAs and we demonstrated that Morgana is under the control of four miRNAs (miR-15a/b and miR-26a/b) and that miR-15a may account for Morgana downregulation in CML patients.
Assuntos
Proteínas de Choque Térmico HSP90 , Leucemia Mielogênica Crônica BCR-ABL Positiva , MicroRNAs , Animais , Humanos , Camundongos , Medula Óssea/metabolismo , Medula Óssea/patologia , Regulação para Baixo , Regulação Leucêmica da Expressão Gênica , Proteínas de Choque Térmico HSP90/metabolismo , Proteínas de Choque Térmico HSP90/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Leucemia Mielogênica Crônica BCR-ABL Positiva/patologia , MicroRNAs/genética , MicroRNAs/metabolismo , Chaperonas Moleculares/metabolismo , Chaperonas Moleculares/genéticaRESUMO
miR-21 is overexpressed in tumors and it displays oncogenic activity. Here, we show that expression of miR-21 in primary tumors anticorrelates with KRIT1/CCM1, an interacting partner of the Ras-like GTPase Rap1, involved in Cerebral Cavernous Malformations (CCM). We present evidences that miR-21 silences KRIT1 by targeting its mRNA 3'UTR and that this interaction is involved in tumor growth control. In fact, miR-21 over-expression or KRIT1 knock-down promote anchorage independent tumor cell growth compared to controls, whereas the opposite is observed when anti-miR-21 or KRIT1 overexpression are employed. Our findings suggest that miR-21 promotes tumor cell growth, at least in part, by down-modulating the potential tumor suppressor KRIT1.
Assuntos
Neoplasias da Mama/patologia , Neoplasias da Mama/fisiopatologia , Regulação Neoplásica da Expressão Gênica/genética , MicroRNAs/genética , Proteínas Associadas aos Microtúbulos/metabolismo , Proteínas Proto-Oncogênicas/metabolismo , Linhagem Celular Tumoral , Proliferação de Células , Humanos , Proteína KRIT1 , Proteínas Associadas aos Microtúbulos/genética , Proteínas Proto-Oncogênicas/genéticaRESUMO
Drug resistance that affects patients universally is a major challenge in cancer therapy. The development of drug resistance in cancer cells is a multifactor event, and its process involves numerous mechanisms that allow these cells to evade the effect of treatments. As a result, the need to understand the molecular mechanisms underlying cancer drug sensitivity is imperative. Traditional 2D cell culture systems have been utilized to study drug resistance, but they often fail to mimic the 3D milieu and the architecture of real tissues and cell-cell interactions. As a result of this, 3D cell culture systems are now considered a comprehensive model to study drug resistance in vitro. Cancer cells exhibit an in vivo behavior when grown in a three-dimensional environment and react to therapy more physiologically. In this review, we discuss the relevance of main 3D culture systems in the study of potential approaches to overcome drug resistance and in the identification of personalized drug targets with the aim of developing patient-specific treatment strategies that can be put in place when resistance emerges.
RESUMO
BACKGROUND: Tumor progression is based on a close interaction between cancer cells and Tumor MicroEnvironment (TME). Here, we focus on the role that Cancer Associated Fibroblasts (CAFs), Mesenchymal Stem Cells (MSCs) and microRNAs (miRs) play in breast cancer and melanoma malignancy. METHODS: We used public databases to investigate miR-214 expression in the stroma compartment of primary human samples and evaluated tumor formation and dissemination following tumor cell injections in miR-214 overexpressing (miR-214over) and knock out (miR-214ko) mice. In addition, we dissected the impact of Conditioned Medium (CM) or Extracellular Vesicles (EVs) derived from miR-214-rich or depleted stroma cells on cell metastatic traits. RESULTS: We evidence that the expression of miR-214 in human cancer or metastasis samples mostly correlates with stroma components and, in particular, with CAFs and MSCs. We present data revealing that the injection of tumor cells in miR-214over mice leads to increased extravasation and metastasis formation. In line, treatment of cancer cells with CM or EVs derived from miR-214-enriched stroma cells potentiate cancer cell migration/invasion in vitro. Conversely, dissemination from tumors grown in miR-214ko mice is impaired and metastatic traits significantly decreased when CM or EVs from miR-214-depleted stroma cells are used to treat cells in culture. Instead, extravasation and metastasis formation are fully re-established when miR-214ko mice are pretreated with miR-214-rich EVs of stroma origin. Mechanistically, we also show that tumor cells are able to induce miR-214 production in stroma cells, following the activation of IL-6/STAT3 signaling, which is then released via EVs subsequently up-taken by cancer cells. Here, a miR-214-dependent pro-metastatic program becomes activated. CONCLUSIONS: Our findings highlight the relevance of stroma-derived miR-214 and its release in EVs for tumor dissemination, which paves the way for miR-214-based therapeutic interventions targeting not only tumor cells but also the TME.
Assuntos
Neoplasias da Mama , Células-Tronco Mesenquimais , MicroRNAs , Humanos , Animais , Camundongos , Feminino , MicroRNAs/genética , MicroRNAs/metabolismo , Transdução de Sinais , Neoplasias da Mama/patologia , Células-Tronco Mesenquimais/metabolismo , Células Estromais/metabolismo , Microambiente TumoralRESUMO
INTRODUCTION: Intrinsic plasticity of breast carcinoma cells allows them to undergo a transient and reversible conversion into mesenchymal cells to disseminate into distant organs, where they can re-differentiate to an epithelial-like status to form a cohesive secondary mass. The p130Cas scaffold protein is overexpressed in human ER+ and HER2+ breast cancer where it contributes to cancer progression, invasion and resistance to therapy. However, its role in regulating mesenchymal aggressive breast cancer cells remains to be determined. The aim of this study was to investigate the molecular and functional involvement of this adaptor protein in breast cancer cell plasticity. METHODS: We used silencing strategies and rescue experiments to evaluate phenotypic and biochemical changes from mesenchymal to epithelial traits in breast tumor cell lines. In the mouse A17 cell model previously related to mesenchymal cancer stem cells and basal-like breast cancer, we biochemically dissected the signaling pathways involved and performed functional in vivo tumor growth ability assays. The significance of the signaling platform was assessed in a human setting through the use of specific inhibitors in aggressive MDA-MB-231 subpopulation LM2-4175 cells. To evaluate the clinical relevance of the results, we analyzed publicly available microarray data from the Netherlands Cancer Institute and from the Koo Foundation Sun Yat-Sen Cancer Center. RESULTS: We show that p130Cas silencing induces loss of mesenchymal features, by downregulating Vimentin, Snail, Slug and Twist transcriptional factors, resulting in the acquirement of epithelial-like traits. Mechanistically, p130Cas controls Cyclooxygenase-2 transcriptional expression, which in turn contributes to p130Cas-dependent maintenance of mesenchymal phenotype. This cascade of events also compromises in vivo tumor growth through inhibition of cell signaling controlling cell cycle progression. c-Src and JNK kinases are sequential players in p130Cas/ Cyclooxygenase-2 axis and their pharmacological inhibition is sufficient to downregulate Cyclooxygenase-2 leading to an epithelial phenotype. Finally, in silico microarray data analysis indicates that p130Cas and Cyclooxygenase-2 concomitant overexpression predicts poor survival and high probability of breast tumor recurrence. CONCLUSIONS: Overall, these data identify a new p130Cas/Cyclooxygenase-2 axis as a crucial element in the control of breast tumor plasticity, opening new therapeutic strategies leading to inhibition of these pathways in aggressive breast carcinoma.
Assuntos
Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Proteína Substrato Associada a Crk/metabolismo , Ciclo-Oxigenase 2/metabolismo , Animais , Neoplasias da Mama/genética , Proteína Tirosina Quinase CSK , Linhagem Celular Tumoral , Proteína Substrato Associada a Crk/genética , Ciclo-Oxigenase 2/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Inativação Gênica , Humanos , Proteínas Quinases JNK Ativadas por Mitógeno/metabolismo , Camundongos , Modelos Biológicos , Fenótipo , Característica Quantitativa Herdável , Quinases da Família src/metabolismoRESUMO
Rhabdomyosarcoma (RMS) is the most common childhood sarcoma and is identified as either the embryonal or alveolar (ARMS) subtype. In approximately 75% of cases, ARMSs are characterized by specific chromosomal translocations that involve PAX and FKHR genes. ARMS gene expression signatures vary, depending on the presence or absence of the translocations. Insulin-like growth factor-binding protein 2 (IGFBP2) is strongly overexpressed in translocation-negative RMS. Because IGFBP2 is associated with tumorigenesis, we investigated its functional role in RMS. An analysis of IGFBP2 distribution in RMS cell lines revealed a strong accumulation in the Golgi complex, in which morphological characteristics appeared peculiarly modified. After silencing IGFBP2 expression, our microarray analysis revealed mostly cell cycle and actin cytoskeleton gene modulations. In parallel, IGFBP2-silenced cells showed reduced cell cycle and rates of invasion and decreased seeding in the lungs after tail vein injections in immunodeficient mice. An analysis of IGFBP2 mRNA and protein localization in human tumors showed abnormal protein accumulation in the Golgi complex, mostly in PAX/FKHR-negative RMS. Moreover, an analysis of patients with RMS revealed the presence of conspicuous circulating levels of IGFBP2 proteins in children with highly aggressive RMS tumors. Taken together, our data provide evidence that IGFBP2 contributes to tumor progression and that it could be used as a marker to better classify clinical and biological risks in RMS.
Assuntos
Biomarcadores Tumorais/metabolismo , Fatores de Transcrição Forkhead/metabolismo , Proteína 2 de Ligação a Fator de Crescimento Semelhante à Insulina/metabolismo , Fatores de Transcrição Box Pareados/metabolismo , Rabdomiossarcoma/metabolismo , Animais , Biomarcadores Tumorais/genética , Pontos de Checagem do Ciclo Celular/fisiologia , Linhagem Celular Tumoral , Movimento Celular/fisiologia , Criança , Proteína Forkhead Box O1 , Fatores de Transcrição Forkhead/genética , Expressão Gênica , Inativação Gênica/fisiologia , Complexo de Golgi , Humanos , Proteína 2 de Ligação a Fator de Crescimento Semelhante à Insulina/genética , Camundongos , Invasividade Neoplásica/genética , Inoculação de Neoplasia , Fator de Transcrição PAX3 , Fatores de Transcrição Box Pareados/genética , RNA Interferente Pequeno/farmacologia , Rabdomiossarcoma/genéticaRESUMO
The development of different generations of BCR-ABL1 tyrosine kinase inhibitors (TKIs) has led to the high overall survival of chronic myeloid leukemia (CML) patients. However, there are CML patients who show resistance to TKI therapy and are prone to progress to more advanced phases of the disease. So, implementing an alternative approach for targeting TKIs insensitive cells would be of the essence. Dihydroorotate dehydrogenase (DHODH) is an enzyme in the de novo pyrimidine biosynthesis pathway that is located in the inner membrane of mitochondria. Here, we found that CML cells are vulnerable to DHODH inhibition mediated by Meds433, a new and potent DHODH inhibitor recently developed by our group. Meds433 significantly activates the apoptotic pathway and leads to the reduction of amino acids and induction of huge metabolic stress in CML CD34+ cells. Altogether, our study shows that DHODH inhibition is a promising approach for targeting CML stem/progenitor cells and may help more patients discontinue the therapy.
Assuntos
Di-Hidro-Orotato Desidrogenase , Leucemia Mielogênica Crônica BCR-ABL Positiva , Resistencia a Medicamentos Antineoplásicos , Proteínas de Fusão bcr-abl/metabolismo , Humanos , Leucemia Mielogênica Crônica BCR-ABL Positiva/tratamento farmacológico , Leucemia Mielogênica Crônica BCR-ABL Positiva/genética , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Inibidores de Proteínas Quinases/farmacologiaRESUMO
OBJECTIVE: Inflammatory stimuli released into atherosclerotic plaque microenvironment regulate vessel formation by modulating gene expression and translation. microRNAs are a class of short noncoding RNAs, acting as posttranscriptional regulators of protein-coding genes involved in various biological processes, including vascular cell biology. Among them, microRNA-221/222 (miR-221/222) seem to negatively modulate vascular remodeling by targeting different target genes. Here, we investigated their potential contribution to inflammation-mediated neovessel formation. METHODS AND RESULTS: We used quantitative real-time RT-PCR amplification to analyze expression of 7 microRNAs previously linked to vascular biology, such as miR-17-5p, miR-21, miR-126, miR-210, miR-221, miR-222, and miR-296 and found high levels of expression for all of them in quiescent endothelial cells. However, miR-126, miR-221, miR-222, and miR-296 turned out to be down-modulated in endothelial cells exposed to inflammatory stimuli. Applying a gain-of-function approach, we demonstrated that, among them, only miR-222 was involved in inflammation-mediated vascular remodeling. In addition, we identified signal transducer and activator of transcription 5A (STAT5A) as a bona fide target of miR-222 and observed that miR-222 negatively correlated with STAT5A expression in human endothelial cells from advanced neovascularized atherosclerotic lesions. CONCLUSIONS: We identified STAT5A as a novel miR-222 target, and this finding opens up new perspectives for treatment of vascular diseases.
Assuntos
Aterosclerose/metabolismo , Células Endoteliais/metabolismo , Inflamação/metabolismo , MicroRNAs/metabolismo , Neovascularização Patológica/prevenção & controle , Neovascularização Fisiológica , Fator de Transcrição STAT5/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Regiões 3' não Traduzidas , Animais , Aterosclerose/genética , Aterosclerose/patologia , Aterosclerose/fisiopatologia , Células Cultivadas , Regulação para Baixo , Células Endoteliais/patologia , Células Endoteliais/transplante , Fator 2 de Crescimento de Fibroblastos/metabolismo , Humanos , Inflamação/genética , Inflamação/patologia , Inflamação/fisiopatologia , Mediadores da Inflamação/metabolismo , Interleucina-3/metabolismo , Camundongos , Camundongos SCID , MicroRNAs/genética , Neovascularização Patológica/genética , Neovascularização Patológica/metabolismo , Neovascularização Patológica/fisiopatologia , Neovascularização Fisiológica/genética , Interferência de RNA , Fator de Transcrição STAT5/genética , Fatores de Tempo , Transfecção , Proteínas Supressoras de Tumor/genéticaRESUMO
The metabolism of cancer cells is generally very different from what is found in normal counterparts. However, in a tumor mass, the continuous crosstalk and competition for nutrients and oxygen among different cells lead to metabolic alterations, not only in cancer cells, but also in the different stromal and immune cells of the tumor microenvironment (TME), which are highly relevant for tumor progression. MicroRNAs (miRs) are small non-coding RNAs that silence their mRNA targets post-transcriptionally and are involved in numerous physiological cell functions as well as in the adaptation to stress situations. Importantly, miRs can also be released via extracellular vesicles (EVs) and, consequently, take part in the bidirectional communication between tumor and surrounding cells under stress conditions. Certain miRs are abundantly expressed in stromal and immune cells where they can regulate various metabolic pathways by directly suppressing enzymes or transporters as well as by controlling important regulators (such as transcription factors) of metabolic processes. In this review, we discuss how miRs can induce metabolic reprogramming in stromal (fibroblasts and adipocytes) and immune (macrophages and T cells) cells and, in turn, how the biology of the different cells present in the TME is able to change. Finally, we debate the rebound of miR-dependent metabolic alterations on tumor progression and their implications for cancer management.
RESUMO
An interactive crosstalk between tumor and stroma cells is essential for metastatic melanoma progression. We evidenced that ESDN/DCBLD2/CLCP1 plays a crucial role in endothelial cells during the spread of melanoma. Precisely, increased extravasation and metastasis formation were revealed in ESDN-null mice injected with melanoma cells, even if the primary tumor growth, vessel permeability, and angiogenesis were not enhanced. Interestingly, improved adhesion of melanoma cells to ESDN-depleted endothelial cells was observed, due to the presence of higher levels of E-selectin transcripts/proteins in ESDN-defective cells. In accordance with these results, anticorrelation was observed between ESDN and E-selectin in human endothelial cells. Most importantly, our data revealed that cimetidine, an E-selectin inhibitor, was able to block cell adhesion, extravasation, and metastasis formation in ESDN-null mice, underlying a major role of ESDN in E-selectin transcription upregulation, which according to our data, may presumably be linked to STAT3. Based on our results, we propose a protective role for ESDN during the spread of melanoma and reveal its therapeutic potential.
Assuntos
Selectina E/antagonistas & inibidores , Células Endoteliais/metabolismo , Melanoma/metabolismo , Proteínas de Membrana/metabolismo , Fator de Transcrição STAT3/metabolismo , Animais , Modelos Animais de Doenças , Progressão da Doença , Selectina E/biossíntese , Selectina E/metabolismo , Humanos , Melanoma/genética , Melanoma/patologia , Camundongos , Microambiente TumoralRESUMO
Unbalanced immune responses to pathogens can be life-threatening although the underlying regulatory mechanisms remain unknown. Here, we show a hypoxia-inducible factor 1α-dependent microRNA (miR)-210 up-regulation in monocytes and macrophages upon pathogen interaction. MiR-210 knockout in the hematopoietic lineage or in monocytes/macrophages mitigated the symptoms of endotoxemia, bacteremia, sepsis, and parasitosis, limiting the cytokine storm, organ damage/dysfunction, pathogen spreading, and lethality. Similarly, pharmacologic miR-210 inhibition improved the survival of septic mice. Mechanistically, miR-210 induction in activated macrophages supported a switch toward a proinflammatory state by lessening mitochondria respiration in favor of glycolysis, partly achieved by downmodulating the iron-sulfur cluster assembly enzyme ISCU. In humans, augmented miR-210 levels in circulating monocytes correlated with the incidence of sepsis, while serum levels of monocyte/macrophage-derived miR-210 were associated with sepsis mortality. Together, our data identify miR-210 as a fine-tuning regulator of macrophage metabolism and inflammatory responses, suggesting miR-210-based therapeutic and diagnostic strategies.