RESUMEN
The presence of cancer stem-like cells (CSCs) is one of the mechanisms responsible for chemoresistance that has been a major hindrance towards lung adenocarcinoma (LAD) treatment. Recently, we have identified microRNA (miR)-200b as a key regulator of chemoresistance in human docetaxel-resistant LAD cells. However, whether miR-200b has effects on regulating CSCs remains largely unclear and needs to be further elucidated. Here, we showed that miR-200b was significantly downregulated in CD133+/CD326+ cells that exhibited properties of CSCs derived from docetaxel-resistant LAD cells. Also, restoration of miR-200b could inhibit maintenance and reverse chemoresistance of CSCs. Furthermore, suppressor of zeste-12 (Suz-12) was identified as a direct and functional target of miR-200b, and silencing of Suz-12 phenocopied the effects of miR-200b on CSCs. Additionally, overexpression of histone deacetylase (HDAC) 1 was identified as a pivotal mechanism responsible for miR-200b repression in CSCs through a specificity protein (Sp) 1-dependent mechanism, and restoration of miR-200b by HDAC1 repression significantly suppressed CSCs formation and reversed chemoresistance of CSCs by regulating Suz-12-E-cadherin signaling. Also, downregulation of HDAC1 or upregulation of miR-200b reduced the in vivo tumorigenicity of CSCs. Finally, Suz-12 was inversely correlated with miR-200b, positively correlated with HDAC1 and up-regulated in docetaxel-resistant LAD tissues compared with docetaxel-sensitive tissues. Taken together, the HDAC1/miR-200b/Suz-12-E-cadherin signaling might account for maintenance of CSCs and formation of chemoresistant phenotype in docetaxel-resistant LAD cells.
Asunto(s)
Adenocarcinoma/patología , Resistencia a Antineoplásicos/genética , Histona Desacetilasa 1/metabolismo , Neoplasias Pulmonares/patología , MicroARNs/genética , Células Madre Neoplásicas/patología , Factor de Transcripción Sp1/metabolismo , Adenocarcinoma/genética , Adenocarcinoma/metabolismo , Animales , Apoptosis , Western Blotting , Cadherinas/genética , Cadherinas/metabolismo , Proliferación Celular , Inmunoprecipitación de Cromatina , Citometría de Flujo , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Histona Desacetilasa 1/genética , Humanos , Técnicas para Inmunoenzimas , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Proteínas de Neoplasias , Células Madre Neoplásicas/efectos de los fármacos , Células Madre Neoplásicas/metabolismo , Complejo Represivo Polycomb 2/genética , Complejo Represivo Polycomb 2/metabolismo , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transducción de Señal/efectos de los fármacos , Factor de Transcripción Sp1/genética , Factores de Transcripción , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Chemoresistance is one of the most significant obstacles in lung adenocarcinoma (LAD) treatment, and this process involves genetic and epigenetic dysregulation of chemoresistance-related genes. Previously, we have shown that restoration of microRNA (miR)-200b significantly reverses chemoresistance of human LAD cells by targeting E2F3. However, the molecular mechanisms involved in the silencing of miR-200b are still unclear. Here we showed that histone deacetylase (HDAC) inhibitors could restore the expression of miR-200b and reverse chemoresistant phenotypes of docetaxel-resistant LAD cells. HDAC1/4 repression significantly increased miR-200b expression by upregulating histone-H3 acetylation level at the two miR-200b promoters partially via a Sp1-dependent pathway. Furthermore, silencing of HDAC1/4 suppressed cell proliferation, promoted cell apoptosis, induced G2/M cell cycle arrest and ultimately reversed in vitro and in vivo chemoresistance of docetaxel-resistant LAD cells, at least partially in a miR-200b-dependent manner. HDAC1/4 suppression-induced rescue of miR-200b contributed to downregulation of E2F3, survivin and Aurora-A, and upregulation of cleaved-caspase-3. HDAC1/4 levels in docetaxel-insensitive human LAD tissues, inversely correlated with miR-200b, were upregulated compared with docetaxel-sensitive tissues. Taken together, our findings suggest that the HDAC1/4/Sp1/miR-200b/E2F3 pathway is responsible for chemoresistance of docetaxel-resistant LAD cells.
Asunto(s)
Adenocarcinoma/genética , Resistencia a Antineoplásicos/genética , Factor de Transcripción E2F3/genética , Histona Desacetilasa 1/genética , Histona Desacetilasas/genética , Neoplasias Pulmonares/genética , MicroARNs/genética , Proteínas Represoras/genética , Adenocarcinoma/mortalidad , Adenocarcinoma del Pulmón , Western Blotting , Línea Celular Tumoral , Inmunoprecipitación de Cromatina , Supervivencia sin Enfermedad , Citometría de Flujo , Silenciador del Gen , Humanos , Inmunoprecipitación , Estimación de Kaplan-Meier , Neoplasias Pulmonares/mortalidad , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , TransfecciónRESUMEN
Chemoresistant tumors usually fail to respond to radiotherapy. However, the mechanisms involved in chemo- and radiotherapy cross resistance are not fully understood. Previously, we have identified microRNA (miR)-451 as a tumor suppressor in lung adenocarcinoma (LAD). However, whether miR-451 plays critical roles in chemo- and radiotherapy cross resistance in LAD is unclear. Here, we established two docetaxel-resistant LAD cell models (SPC-A1/DTX and H1299/DTX), and showed that miR-451 was significantly downregulated in docetaxel-resistant LAD cells. Gain - and loss - of - function assays indicated that re-expression of miR-451 could reverse radioresistance of docetaxel-resistant LAD cells both in vitro and in vivo through promoting apoptosis and DNA double-strand breaks (DSBs). The proto-oncogene c-Myc was identified as a direct target of miR-451, and re-expression of miR-451 inhibited survivin and rad-51 expression by reducing the amount of c-Myc protein binding to their promoters. Silencing of c-Myc could phenocopy the effects of miR-451 upregulation, and restoration of c-Myc could partially rescue the effect of miR-451 upregulation on radiosensitivity of docetaxel-resistant LAD cells. Therefore, dysregulation of miR-451/c-Myc-survivin/rad-51 signaling is responsible for radioresistance of docetaxel-resistant LAD cells, and targeting it will be a potential strategy for reversing chemo- and radiotherapy cross resistance of LAD patients.
Asunto(s)
Adenocarcinoma/genética , Neoplasias Pulmonares/genética , MicroARNs/genética , Adenocarcinoma del Pulmón , Línea Celular Tumoral , Docetaxel , Resistencia a Antineoplásicos/genética , Humanos , Proto-Oncogenes Mas , Transducción de Señal , TaxoidesRESUMEN
Angiopoietin-like protein 4 (ANGPTL4) has been reported to promote tumor growth, metastasis, and angiogenesis under certain conditions. The aim of this study was to examine ANGPTL4 expression in tumor and serum tissues from esophageal squamous cell carcinoma (ESCC) patients. A total of 78 ESCC patients treated with radical resection were enrolled in this study. Immunohistochemistry was used to detect ANGPTL4 expression in ESCC tissues. Serum ANGPTL4 levels were determined via enzyme-linked immunosorbent assay (ELISA). The receiver operating characteristics curve was constructed to describe diagnostic specificity and sensitivity. There were 52 cases (69.2 %) showing a higher level of ANGPTL4 expression in tumor tissues than that in normal tissues, and the rate of ANGPTL4 protein high/moderate expression in ESCC and normal tissues was 55.1 % (43/78) and 6.4 % (5/78), respectively, with a significant difference (P < 0.001). Moreover, the high/moderate of ANGPTL4 protein was significantly associated with lymph metastasis, clinical stage, and adverse 2-year progression-free survival. In addition, serum ANGPTL4 level in ESCC patients was much higher than that in patients with benign esophageal disease (P < 0.001), and area under the curve was 0.94 (95 % CI 0.886390-0.978173, P < 0.001). But serum ANGPTL4 level was significantly decreased at post-operative 7-10 days (P = 0.004). ANGPTL4 upregulation may play an important role in ESCC development, and serum ANGPTL4 level may be a potential tumor marker for ESCC diagnosis and prognosis.
Asunto(s)
Angiopoyetinas/genética , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/patología , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/patología , Adulto , Anciano , Anciano de 80 o más Años , Proteína 4 Similar a la Angiopoyetina , Supervivencia sin Enfermedad , Carcinoma de Células Escamosas de Esófago , Femenino , Humanos , Metástasis Linfática/genética , Metástasis Linfática/patología , Masculino , Persona de Mediana Edad , Pronóstico , Curva ROC , Regulación hacia Arriba/genéticaRESUMEN
The RUNX3 gene has been shown to function as a tumor suppressor gene implicated in various cancers, but its association with tumor chemoresistance has not been fully understood. Here, we investigated the effect of epigenetic downregulation of RUNX3 in docetaxel resistance of human lung adenocarcinoma and its possible molecular mechanisms. RUNX3 was found to be downregulated by hypermethylation in docetaxel-resistant lung adenocarcinoma cells. Its overexpression could resensitize cells to docetaxel both in vitro and in vivo by growth inhibition, enhancement of apoptosis and G1 phase arrest. Conversely, knockdown of RUNX3 could lead to the decreased sensitivity of parental human lung adenocarcinoma cells to docetaxel by enhancing proliferative capacity. Furthermore, we showed that overexpression of RUNX3 could inactivate the AKT/GSK3ß/ß-catenin signaling pathway in the docetaxel-resistant cells. Importantly, co-transfection of RUNX3 and constitutively active Akt1 could reverse the effects of RUNX3 overexpression, while treatment with the MK-2206 (AKT inhibitor) mimicked the effects of RUNX3 overexpression in docetaxel-resistant human lung adenocarcinoma cells. Immunohistochemical analysis revealed that decreased RUNX3 expression was correlated with high expression of Akt1 and decreased sensitivity of patients to docetaxel-based chemotherapy. Taken together, our results suggest that epigenetic downregulation of RUNX3 can induce docetaxel resistance in human lung adenocarcinoma cells by activating AKT signaling and increasing expression of RUNX3 may represent a promising strategy for reversing docetaxel resistance in the future.