RESUMO
INTRODUCTION: Clinical studies suggest that obesity, in addition to promoting breast cancer aggressiveness, is associated with a decrease in chemotherapy efficacy, although the mechanisms involved remain elusive. As chemotherapy is one of the main treatments for aggressive or metastatic breast cancer, we investigated whether adipocytes can mediate resistance to doxorubicin (DOX), one of the main drugs used to treat breast cancer, and the mechanisms associated. METHODS: We used a coculture system to grow breast cancer cells with in vitro differentiated adipocytes as well as primary mammary adipocytes isolated from lean and obese patients. Drug cellular accumulation, distribution, and efflux were studied by immunofluorescence, flow cytometry, and analysis of extracellular vesicles. Results were validated by immunohistochemistry in a series of lean and obese patients with cancer. RESULTS: Adipocytes differentiated in vitro promote DOX resistance (with cross-resistance to paclitaxel and 5-fluorouracil) in a large panel of human and murine breast cancer cell lines independently of their subtype. Subcellular distribution of DOX was altered in cocultivated cells with decreased nuclear accumulation of the drug associated with a localized accumulation in cytoplasmic vesicles, which then are expelled into the extracellular medium. The transport-associated major vault protein (MVP), whose expression was upregulated by adipocytes, mediated both processes. Coculture with human mammary adipocytes also induced chemoresistance in breast cancer cells (as well as the related MVP-induced DOX efflux) and their effect was amplified by obesity. Finally, in a series of human breast tumors, we observed a gradient of MVP expression, which was higher at the invasive front, where tumor cells are at close proximity to adipocytes, than in the tumor center, highlighting the clinical relevance of our results. High expression of MVP in these tumor cells is of particular interest since they are more likely to disseminate to give rise to chemoresistant metastases. CONCLUSIONS: Collectively, our study shows that adipocytes induce an MVP-related multidrug-resistant phenotype in breast cancer cells, which could contribute to obesity-related chemoresistance.
Assuntos
Adipócitos/metabolismo , Antineoplásicos/farmacologia , Neoplasias da Mama/tratamento farmacológico , Obesidade/complicações , Partículas de Ribonucleoproteínas em Forma de Abóbada/metabolismo , Células 3T3 , Tecido Adiposo/citologia , Adulto , Idoso , Animais , Antineoplásicos/uso terapêutico , Mama/citologia , Mama/patologia , Mama/cirurgia , Neoplasias da Mama/patologia , Neoplasias da Mama/cirurgia , Linhagem Celular Tumoral , Técnicas de Cocultura , Doxorrubicina/farmacologia , Doxorrubicina/uso terapêutico , Resistência a Múltiplos Medicamentos , Resistencia a Medicamentos Antineoplásicos , Feminino , Fluoruracila/farmacologia , Fluoruracila/uso terapêutico , Humanos , Mastectomia , Camundongos , Pessoa de Meia-Idade , Paclitaxel/farmacologia , Paclitaxel/uso terapêutico , Cultura Primária de Células , RNA Interferente Pequeno/metabolismo , Partículas de Ribonucleoproteínas em Forma de Abóbada/genéticaRESUMO
In addition to the major subcutaneous and visceral adipose tissues (AT), other adipose depots are dispersed throughout the body and are found in close interaction with proximal organs such as mammary and periprostatic AT (MAT and PPAT respectively). These ATs have an effect on proximal organ function during physiological processes and diseases such as cancer. We highlighted here some of their most distinctive features in terms of tissular organization and responses to external stimuli and discussed how obesity affects them based on our current knowledge.
Assuntos
Tecido Adiposo , Obesidade , Humanos , Tecido Adiposo/fisiologia , Feminino , Obesidade/fisiopatologia , Neoplasias/patologia , Animais , Mama/fisiologia , Mama/patologia , Neoplasias da Mama/patologia , Glândulas Mamárias Humanas/fisiologia , Glândulas Mamárias Humanas/patologia , Gordura Intra-Abdominal , Gordura Subcutânea/fisiologia , Gordura Subcutânea/patologiaRESUMO
DNA-dependent protein kinase (DNA-PK) is involved in DNA double-strand break (DSB) signalling and repair. We report that DNA-PK is activated by mild hypoxia conditions (0.1-1% O2) as shown by (1) its autophosphorylation on Ser2056, and (2) its mobilisation from a soluble nucleoplasmic compartment to a less extractable nuclear fraction. The recruitment of DNA-PK was not followed by activation and recruitment of the XRCC4-DNA-ligase-IV complex, suggesting that DSBs are not responsible for activation of DNA-PK. To unravel the mechanism of DNA-PK activation, we show that exposure of cells to trichostatin A, a histone deacetylase inhibitor, leads to DNA-PK autophosphorylation and relocalisation to DNA. Histone acetylation (mainly H3K14) is increased in hypoxic cells and treatment with anacardic acid, an inhibitor of histone acetyl transferase, prevented both histone modifications and DNA-PK activation in hypoxic conditions. Importantly, in using either silenced DNA-PK cells or cells exposed to a specific DNA-PK inhibitor (NU7026), we demonstrated that hypoxic DNA-PK activation positively regulates the key transcription factor HIF-1 and one subsequent target gene, GLUT1. Our results show that hypoxia initiates chromatin modification and consequently DNA-PK activation, which positively regulate cellular oxygen-sensing and oxygen-signalling pathways.
Assuntos
Cromatina/metabolismo , Proteína Quinase Ativada por DNA/metabolismo , Estresse Fisiológico , Acetilação , Adaptação Fisiológica , Aminoglicosídeos/farmacologia , Antígenos Nucleares/metabolismo , Hipóxia Celular , Linhagem Celular , Cromonas/farmacologia , Dano ao DNA , Proteína Quinase Ativada por DNA/antagonistas & inibidores , Proteínas de Ligação a DNA/metabolismo , Enedi-Inos/farmacologia , Ativação Enzimática , Transportador de Glucose Tipo 1/metabolismo , Histonas/metabolismo , Humanos , Ácidos Hidroxâmicos/farmacologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Autoantígeno Ku , Morfolinas/farmacologia , Fosforilação , Processamento de Proteína Pós-Traducional , Transdução de SinaisRESUMO
Obesity is a negative prognosis factor for breast cancer. Yet, the biological mechanisms underlying this effect are still largely unknown. An emerging hypothesis is that the transfer of free fatty acids (FFA) between adipocytes and tumor cells might be altered under obese conditions, contributing to tumor progression. Currently there is a paucity of models to study human mammary adipocytes (M-Ads)-cancer crosstalk. As for other types of isolated white adipocytes, herein, we showed that human M-Ads die within 2-3 days by necrosis when grown in 2D. As an alternative, M-Ads were grown in a fibrin matrix, a 3D model that preserve their distribution, integrity and metabolic function for up to 5 days at physiological glucose concentrations (5 mM). Higher glucose concentrations frequently used in in vitro models promote lipogenesis during M-Ads culture, impairing their lipolytic function. Using transwell inserts, the matrix embedded adipocytes were cocultured with breast cancer cells. FFA transfer between M-Ads and cancer cells was observed, and this event was amplified by obesity. Together these data show that our 3D model is a new tool for studying the effect of M-Ads on tumor cells and beyond with all the components of the tumor microenvironment including the immune cells.
Assuntos
Adipócitos , Neoplasias da Mama , Ácidos Graxos não Esterificados , Glândulas Mamárias Humanas , Obesidade , Magreza , Técnicas de Cultura de Células em Três Dimensões , Adipócitos/metabolismo , Adipócitos/patologia , Cultura Primária de Células , Glândulas Mamárias Humanas/patologia , Neoplasias da Mama/patologia , Obesidade/metabolismo , Obesidade/patologia , Magreza/metabolismo , Magreza/patologia , Humanos , Células MDA-MB-231 , Ácidos Graxos não Esterificados/metabolismo , PrognósticoRESUMO
Obesity increases the occurrence of post-menopausal breast cancer and negatively affects prognosis independently of menopausal status. After summarizing the available epidemiological data concerning these associations, we will show that a deleterious crosstalk is established during tumor progression between cancer cells and the surrounding mammary adipose tissue (MAT). In obesity, the chronic sub-inflammatory state of MAT could amplify the negative effect of this crosstalk although other mechanisms also warrant further study. Finally, we will discuss the efficiency of weight loss in both primary prevention and recurrence, a strategy that could be more complex that initially thought.
Assuntos
Tecido Adiposo/fisiologia , Neoplasias da Mama/etiologia , Obesidade/complicações , Adiposidade/fisiologia , Animais , Neoplasias da Mama/diagnóstico , Neoplasias da Mama/epidemiologia , Neoplasias da Mama/metabolismo , Feminino , Humanos , Menopausa/fisiologia , Obesidade/diagnóstico , Obesidade/epidemiologia , Obesidade/metabolismo , Prognóstico , Fatores de RiscoRESUMO
Growing evidence has raised the important roles of adipocytes as an active player in the tumor microenvironment. In many tumors adipocytes are in close contact with cancer cells. They secrete various factors that can mediate local and systemic effects. The adipocyte-cancer cell crosstalk leads to phenotypical and functional changes of both cell types, which can further enhance tumor progression. Moreover, obesity, which is associated with an increase in adipose mass and an alteration of adipose tissue, has been established as a risk factor for cancer incidence and cancer-related mortality. In this review, we summarize the mechanisms of the adipocyte-cancer cell crosstalk in both obese and lean conditions as well as its impact on cancer cell growth, local invasion, metastatic spread and resistance to treatments. Better characterization of cancer-associated adipocytes and the key molecular events in the adipocyte-cancer cell crosstalk will provide insights into tumor biology and suggest efficient therapeutic opportunities.
RESUMO
CYP1A1 is an extrahepatic enzyme largely involved in the bioactivation of various procarcinogens such as polycyclic aromatic hydrocarbons (PAHs) and arylamines. CYP1A1 expression is mainly regulated by AhR. Our laboratory has recently shown a new CYP1A1 regulation pathway involving PPARalpha. The aim of this study was to evaluate, in a Caco-2 cell line, the effect of a coexposure to 3-methylcholanthrene (3MC, AhR ligand) and WY-14643 (WY, PPARalpha ligand) on CYP1A1 expression (enzymatic activity, mRNA level and promoter activity). An additive effect on CYP1A1 expression was shown in cells coexposed with 3MC (0.1 or 1 microM) and a low WY concentration (30 microM) whereas a potentiating effect was observed after coexposure with 3MC (0.1 or 1 microM) and a high WY concentration (200 microM). Furthermore, 200 microM WY, alone or with 3MC, was able to increase the AhR protein level (two-fold). In conclusion, coexposure with 3MC and the PPARalpha agonist WY leads to an additive or potentiating effect on CYP1A1 inducibility, depending on the WY concentration. Furthermore, at high concentration (200 microM), WY induced AhR expression, which could explain the potentiating effect on CYP1A1 inducibility observed after addition of an AhR ligand (3MC). This phenomenon should be taken into account for risk assessment involving CYP1A1 induction.
Assuntos
Citocromo P-450 CYP1A1/biossíntese , PPAR alfa/metabolismo , Receptores de Hidrocarboneto Arílico/metabolismo , Fatores de Transcrição Hélice-Alça-Hélice Básicos , Western Blotting , Células CACO-2 , Citocromo P-450 CYP1A1/efeitos dos fármacos , Citocromo P-450 CYP1A1/genética , Citocromo P-450 CYP1A1/metabolismo , Relação Dose-Resposta a Droga , Sinergismo Farmacológico , Indução Enzimática/efeitos dos fármacos , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Humanos , Metilcolantreno/farmacologia , Regiões Promotoras Genéticas/efeitos dos fármacos , Pirimidinas/farmacologia , RNA Mensageiro/efeitos dos fármacos , Receptores de Hidrocarboneto Arílico/químicaRESUMO
Ataxia Telangiectasia (AT) is an autosomal recessive disorder characterized by a wide variety of progressive clinical symptoms. This includes neuronal degeneration, oculocutaneous telangiectasias, diabetes mellitus, immunodeficiency, increased risk of cancer and sensitivity to ionizing radiation. The gene mutated in this disease, ATM (Ataxia Telangiectasia Mutated), encodes a protein kinase involved in DNA double strand breaks signalling and repair. ATM deficient cells also display an increase in oxidative stress, by poorly characterized mechanism(s), which clearly contributes to the neurodegenerative aspect of the disease. Despite these advances, the occurrence of the vascular abnormalities, glucose intolerance and insulin resistance remains poorly understood. In different cellular models where ATM expression was disrupted, we demonstrated that the absence of ATM leads to an increased expression of both subunits of the transcription factor Hypoxia Inducible Factor 1 (HIF-1). We also observed enhanced trans-activating functions of HIF-1. HIF-1 is the central regulator of responses to hypoxia which induces the transcription of genes involved in angiogenesis (e.g., VEGF-Vascular Endothelial Growth Factor) and cellular metabolism (e.g., GLUT-1). Interestingly, we demonstrated that ATM disruption positively regulates both expression and function of the basal glucose transporter GLUT-1 as well as the proangiogenic factor, VEGF. In addition, our results suggest that the absence of ATM increases HIF-1 proteins biosynthesis, and this effect is dependant on the oxidative stress existing in ATM deficient cells. Our compelling results highlight a new link between ATM deficiency and the clinical features of the disease and provide a molecular link between ATM downregulation and the increase in tumor angiogenesis observed in human breast cancers.
Assuntos
Ataxia Telangiectasia/etiologia , Proteínas de Ciclo Celular/metabolismo , Proteínas de Ligação a DNA/metabolismo , Fator 1 Induzível por Hipóxia/metabolismo , Estresse Oxidativo , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Proteínas Mutadas de Ataxia Telangiectasia , Proteínas de Ciclo Celular/genética , Linhagem Celular , Proteínas de Ligação a DNA/deficiência , Proteínas de Ligação a DNA/genética , Transportador de Glucose Tipo 1/genética , Transportador de Glucose Tipo 1/metabolismo , Células HeLa , Humanos , Proteínas Serina-Treonina Quinases/deficiência , Proteínas Serina-Treonina Quinases/genética , Proteínas Supressoras de Tumor/deficiência , Proteínas Supressoras de Tumor/genética , Fator A de Crescimento do Endotélio Vascular/genética , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
CYP1A1 is largely involved in carcinogenesis through the bioactivation of numerous procarcinogens. Exposure to environmental pollutants such as polycyclic aromatic hydrocarbons (PAHs) leads to induction of CYP1A1 via AhR pathway. We have previously demonstrated that fetal bovine serum (FBS) induces CYP1A1 gene transcription. In this work, we show evidence that the serum does not contain an AhR ligand and we evaluated the effect of a 3-methylcholanthrene (3-MC) and FBS cotreatment on CYP1A1 expression. CYP1A1 activity was potentiated by this treatment. This potentiation was at least in part associated with an increase of the CYP1A1 mRNA and gene transcription levels. FBS potentiation of CYP1A1 PAH-mediated induction was related to a significant increase of single strand breaks of DNA as compared to a single 3-MC treatment. Moreover, we demonstrated that human serum induces CYP1A1 with a high interindividual variability. The potentiation by serum of polycyclic aromatic hydrocarbon CYP1A1 induction could be involved in the etiology of some human cancers.
Assuntos
Proteínas Sanguíneas/farmacologia , Citocromo P-450 CYP1A1/biossíntese , Metilcolantreno/metabolismo , Adenocarcinoma , Animais , Benzo(a)Antracenos/farmacologia , Benzo(a)pireno/farmacologia , Carcinoma , Bovinos , Neoplasias do Colo , Citocromo P-450 CYP1A1/genética , Dano ao DNA , Indução Enzimática/efeitos dos fármacos , Humanos , RNA Mensageiro/metabolismo , Células Tumorais CultivadasRESUMO
CYP1A1 isoform is mainly regulated by the transcription factor AhR and to a lesser extent by the nuclear receptor RAR. The effect of a coexposure with 3MC, a AhR ligand, and RA, a RAR ligand, which are, respectively, strong and weak CYP1A1 inducers, is poorly known. We showed in Caco-2 cells that addition of RA significantly decreased 3MC-induced CYP1A1 expression by -55% for mRNA level and -30% for promoter and enzymatic activities. We further showed that RA decreased AhR protein level. Moreover, a physical interaction between AhR and the RAR-corepressor SMRT has been described in vitro. Using the corepressor inhibitor TSA, transfected-cells with SMRT cDNA, and coimmunoprecipitation experiments, we demonstrated that RA addition repressed AhR function through a marked AhR/SMRT physical interaction. This interaction explains the decrease of 3MC-induced CYP1A1 expression. This new mechanism involving the repression of AhR-induced CYP1A1 expression by retinoids allows better knowledge of the CYP1A1 regulation.