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BACKGROUND: Trastuzumab is part of the standard treatment for patients with human epidermal growth factor receptor 2 (HER-2)-positive breast cancer, but not all patients respond to trastuzumab. Altered microRNA (miR) expression levels in cancer cells have been correlated with prognosis and response to chemotherapy. The authors of this report hypothesized that altered miR expression levels in plasma are associated with sensitivity to trastuzumab in patients with HER-2 positive breast cancer. METHODS: Quantitative reverse transcriptase-polymerase chain reaction was used to analyze plasma samples, including samples from patients with breast cancer who were enrolled in a clinical trial of neoadjuvant trastuzumab-based chemotherapy. Expression levels of miR-210, miR-21, miR-29a, and miR-126 were analyzed according to the type of response (pathologic complete response [n = 18] vs residual disease [n = 11]). MicroRNA expression levels also were compared in trastuzumab-sensitive and trastuzumab-resistant breast cancer cells derived from BT474 cells and in an independent set of preoperative plasma samples (n = 39) and postoperative plasma samples (n = 30) from 43 breast cancer patients who did not receive any treatment. RESULTS: At baseline before patients received neoadjuvant chemotherapy combined with trastuzumab, circulating miR-210 levels were significantly higher in those who had residual disease than in those who achieved a pathologic complete response (P = .0359). The mean expression ratio for miR-210 was significantly higher in trastuzumab-resistant BT474 cells, and miR-210 expression was significantly higher before surgery than after surgery (P = .0297) and in patients whose cancer metastasized to the lymph nodes (P = .0030). CONCLUSIONS: Circulating miR-210 levels were associated with trastuzumab sensitivity, tumor presence, and lymph node metastases. These results suggest that plasma miR-210 may be used to predict and perhaps monitor response to therapies that contain trastuzumab.
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Anticuerpos Monoclonales Humanizados/uso terapéutico , Antineoplásicos/uso terapéutico , Neoplasias de la Mama/tratamiento farmacológico , MicroARNs/sangre , Adulto , Anciano , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Resistencia a Antineoplásicos , Femenino , Humanos , Metástasis Linfática , Persona de Mediana Edad , TrastuzumabRESUMEN
INTRODUCTION: The c-Jun coactivator, Jun activation-domain binding protein 1 (Jab1) also known as the fifth component of the COP9 signalosome complex (CSN5), is a novel candidate oncogene whose aberrant expression contributes to the progression of breast carcinoma and other human cancers. The mechanism of Jab1 gene expression and its deregulation in cancer cells remains to be identified. We therefore investigated the transcriptional regulatory mechanisms of Jab1 expression in human breast carcinoma cells. METHODS: To identify potential regulators of Jab1 transcription, we cloned the 5' upstream region of the human Jab1 gene and mapped its transcriptional start site. We identified binding sequences for the CCAAT/enhancer binding protein (C/EBP) and GATA, as well as a signal transducer and activator of transcription-3 (Stat3) consensus sequence overlapping the C/EBP site, using 5'- deletion analysis and a gene reporter assay. Mutational analysis of these binding sites was performed to confirm their roles in promoting Jab1 transcription in breast cancer cells. We further confirmed these binding sites using electrophoretic mobility shift assays (EMSAs) and chromatin immunoprecipitation (ChIP) assays. We also analyzed whether the siRNA-mediated inactivation of Stat3 and Src could reduce Jab1-promoter activity and whether interleukine-6 (IL-6) could mediate increased Jab1 expression through Stat3 signaling. RESULTS: We identified binding sequences for C/EBP, GATA, as well as a Stat3 consensus sequence overlapping the C/EBP site in the promoter region of Jab1. C/EBP-beta2 is a potential transcriptional activator of Jab1 and mutation of the C/EBP/Stat3 binding site significantly reduced Jab1-promoter activity. In addition, inhibiting Stat3 significantly reduced Jab1-promoter activation. EMSA and ChIP assays confirmed that C/EBP, GATA1 and Stat3 bind to Jab1 promoter in breast carcinoma cells. We also found that Src, an activator of Stat3, is involved in Jab1-promoter activation. siRNA knockdown of Src reduced the Jab1-promoter activity, similar to the results seen when Stat3 was inhibited in breast carcinoma cells. Interestingly, reactivation of Stat3 in normal mammary epithelial cells (MCF-10A, MCF-10F) is sufficient to reactivate Jab1 expression. Treatment with the cytokine IL-6 resulted in increased Jab1 expression that was blocked by inhibition of Stat3. CONCLUSIONS: These findings reveal a novel mechanism of Jab1 gene regulation and provide functional and mechanistic links between the Src/Stat3 and IL-6/Stat3 signaling axes that are involved in the activation of Jab1 transcription and regulation of this novel oncogenic protein.
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Neoplasias de la Mama/metabolismo , Proteínas Potenciadoras de Unión a CCAAT/metabolismo , Regulación Neoplásica de la Expresión Génica , Péptidos y Proteínas de Señalización Intracelular/genética , Péptido Hidrolasas/genética , Regiones Promotoras Genéticas , Factor de Transcripción STAT3/metabolismo , Secuencia de Bases , Sitios de Unión , Proteína beta Potenciadora de Unión a CCAAT , Complejo del Señalosoma COP9 , Línea Celular Tumoral , Inmunoprecipitación de Cromatina , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Ensayo de Cambio de Movilidad Electroforética , Femenino , Factor de Transcripción GATA1/metabolismo , Humanos , Interleucina-6/farmacología , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Péptido Hidrolasas/metabolismo , Interferencia de ARN , ARN Interferente Pequeño , Secuencias Reguladoras de Ácidos Nucleicos , Factor de Transcripción STAT3/antagonistas & inhibidores , Análisis de Secuencia de ADN , Transducción de Señal/genética , Transcripción Genética , Activación Transcripcional , Familia-src Quinasas/genética , Familia-src Quinasas/metabolismoRESUMEN
AIMS: Dedicator of cytokinesis I (Dock180) is a novel guanine nucleotide exchange factor for Rho guanosine triphosphates (GTPases) important for cell migration. The aim of this study was to evaluate the role of Dock180 in ovarian carcinogenesis. METHODS AND RESULTS: Using immunohistochemistry, real-time polymerase chain reaction and Western blotting, overexpression of Dock180 RNA and protein was demonstrated in the nucleus and cytoplasm of ovarian cancer cell lines (n = 5) and clinical samples of ovarian borderline tumours (n = 21) and invasive cancers (n = 108) when compared with ovarian epithelial cell lines (n = 3) and benign cystadenomas (n = 10) (P < 0.05). High Dock180 cytoplasmic expression in ovarian cancer (n = 108) was associated significantly with serous histological type, high-grade cancer and advanced stage (P < 0.05), as well as poor overall and disease-free survival (P = 0.004). Using multivariate progression analysis, high Dock180 cytoplasmic expression and advanced cancer stage were found to be independent prognostic factors for short overall survival and disease-free survival (P < 0.05). Exogenous expression of Dock180 by transient transfection enhanced cancer cell migration and invasion, whereas knockdown of Dock180 by an siRNA approach retarded cancer cell migration and invasion in association with down-regulation of matrix metalloproteinase 2. CONCLUSIONS: Our findings suggest that Dock180 contributes to ovarian carcinogenesis and dissemination and is a potential prognostic marker and therapeutic target.
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Carcinoma/enzimología , Cistoadenoma/enzimología , Neoplasias Ováricas/enzimología , Proteínas de Unión al GTP rac/biosíntesis , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores de Tumor/análisis , Western Blotting , Carcinoma/mortalidad , Carcinoma/patología , Movimiento Celular/genética , Cistoadenoma/mortalidad , Cistoadenoma/patología , Femenino , Humanos , Inmunohistoquímica , Microscopía Confocal , Persona de Mediana Edad , Clasificación del Tumor , Invasividad Neoplásica/genética , Invasividad Neoplásica/patología , Neoplasias Ováricas/mortalidad , Neoplasias Ováricas/patología , Fenotipo , Pronóstico , ARN Interferente Pequeño , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Transfección , Regulación hacia Arriba , Adulto Joven , Proteínas de Unión al GTP rac/análisisRESUMEN
BACKGROUND: FBI-1 (factor that binds to the inducer of short transcripts of human immunodeficiency virus-1) is a member of the POK (POZ and Kruppel) family of transcription factors and play important roles in cellular differentiation and oncogenesis. Recent evidence suggests that FBI-1 is expressed at high levels in a subset of human lymphomas and some epithelial solid tumors. However, the function of FBI-1 in human ovarian cancers remains elusive. RESULTS: In this study, we investigated the role of FBI-1 in human ovarian cancers, in particularly, its function in cancer cell invasion via modulating membrane type 1-matrix metalloproteinase (MT1-MMP). Significantly higher FBI-1 protein and mRNA expression levels were demonstrated in ovarian cancers samples and cell lines compared with borderline tumors and benign cystadenomas. Increased FBI-1 mRNA expression was correlated significantly with gene amplification (P = 0.037). Moreover, higher FBI-1 expression was found in metastatic foci (P = 0.036) and malignant ascites (P = 0.021), and was significantly associated with advanced stage (P = 0.012), shorter overall survival (P = 0.032) and disease-free survival (P = 0.016). In vitro, overexpressed FBI-1 significantly enhanced cell migration and invasion both in OVCA 420 and SKOV-3 ovarian carcinoma cells, irrespective of p53 status, accompanied with elevated expression of MT1-MMP, but not MMP-2 or TIMP-2. Moreover, knockdown of MT1-MMP abolished FBI-1-mediated cell migration and invasion. Conversely, stable knockdown of FBI-1 remarkably reduced the motility of these cells with decreased expression of MT1-MMP. Promoter assay and chromatin immunoprecipitation study indicated that FBI-1 could directly interact with the promoter spanning ~600 bp of the 5'-flanking sequence of MT1-MMP and enhanced its expression in a dose-dependent manner. Furthermore, stable knockdown and ectopic expression of FBI-1 decreased and increased cell proliferation respectively in OVCA 420, but not in the p53 null SKOV-3 cells. CONCLUSIONS: Our results suggested an important role of FBI-1 in ovarian cancer cell proliferation, cell mobility, and invasiveness, and that FBI-1 can be a potential target of chemotherapy.
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Proteínas de Unión al ADN/metabolismo , Metaloproteinasa 14 de la Matriz/metabolismo , Neoplasias Ováricas/enzimología , Neoplasias Ováricas/patología , Factores de Transcripción/metabolismo , Adulto , Anciano , Western Blotting , Línea Celular , Línea Celular Tumoral , Movimiento Celular/genética , Movimiento Celular/fisiología , Proliferación Celular , Inmunoprecipitación de Cromatina , Proteínas de Unión al ADN/genética , Femenino , Humanos , Inmunohistoquímica , Técnicas In Vitro , Metaloproteinasa 14 de la Matriz/genética , Persona de Mediana Edad , Reacción en Cadena de la Polimerasa , Unión Proteica , Proto-Oncogenes Mas , Factores de Transcripción/genética , Adulto JovenRESUMEN
OBJECTIVE: To compare the incidence and risk factors of superior facet joint violation (FJV) during cortical bone trajectory screw placement in robot-assisted approach versus conventional technique. METHODS: A retrospective study, including 69 patients having cortical bone trajectory (CBT) screw instrumentation for symptomatic degenerated diseases or trauma, was conducted between June 2015 to January 2019. All patients underwent CBT surgery performed by the same team of experienced surgeons. Patients were randomly divided into two groups: a conventional group (CG, 46 cases) and a robot group (RG, 23 cases). The surgical robotic system was used for screw instrumentation in the robot group and the traditional screw instrumentation with fluoroscopic guidance was used in the conventional group. Cortical screws followed a medio-to-lateral path in the transverse plane and a caudal-to-cephalad path in the sagittal plane. Preoperative and postoperative computed tomography (CT) scans were obtained to determine the degree and incidence of FJV. The violation status of facet joint was evaluated according to the modified classification: grade 0, no violation; grade 1, screw shaft, screw head or rod within 1 mm of or abutting the facet joint, but did not enter the articular facet joint; grade 2, screw shaft, screw head or rod clearly in the facet joint. The following factors that may contribute to the occurrence of FJV were analyzed: age, sex, body mass index (BMI), proximal fusion level, fusion length, the side of screw, preoperative vertebral slip, superior facet angle, and degenerative scoliosis. The chi-squared test and Student's t-test were used for analysis of the variables for significance (P < 0.05). RESULTS: FJV occurred in 41.3% of patients in CG and 17.3% of patients in RG. A chi-squared analysis revealed a significantly lower rate of FJV for RG compared with CG (P = 0.04). In the CG, 17 of the 109 cephalad screws were grade 1 (15.6%), and five were grade 2 (4.6%). In the RG, three of the 46 cephalad screws were grade 1 (6.5%), and three were grade 2 (6.5%). There was a statistically significant difference in the incidence of FJV between the left and right screw with fluoroscopy-assisted CBT screw instrumentation (P < 0.05). A significant correlation between scoliosis with the FJV was found in CG (P < 0.05) and in RG (P < 0.05). With regard to superior facet angle, a measurement ≥45° was a significant risk factor of FJV in CG (P < 0.05) and in RG (P < 0.05). CONCLUSIONS: A robot-assisted approach could reduce the incidence of FJV compared with the conventional approach in CBT technique.
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Hueso Cortical/cirugía , Vértebras Lumbares/cirugía , Tornillos Pediculares , Procedimientos Quirúrgicos Robotizados/métodos , Fusión Vertebral/métodos , Articulación Cigapofisaria/lesiones , Anciano , Femenino , Fluoroscopía , Humanos , Masculino , Persona de Mediana Edad , Complicaciones Posoperatorias/prevención & control , Estudios Retrospectivos , Factores de RiesgoRESUMEN
OBJECTIVE: To compare one-time accuracy rate between simulated freehand (SFH) and navigation simulated (NS) pedicle screw insertion, assuming no second chance to correct screws. METHODS: A simulated, comparative, cross-sectional study was conducted on 69 patients undergoing lumbar spine surgery. An intraoperative registration system captured the planned point of entry and trajectory of pedicle screws for both SFH under direct visualization and NS under navigation-aided visualization. Pedicle screw insertion was simulated for each captured image (370 screws) using Surgimap. Rajasekaran's method helped evaluate the point of entry accuracy and trajectory. RESULTS: Accuracy rate was better for the NS method (97.8%) than for the SFH method (63.8%). Of 370 screws in the SFH group, 134 penetrated the cortex, with 31 resulting in >4 mm penetration. Of 370 screws in the NS group, 8 penetrated the cortex, <4 mm penetration. Of 134 misplaced screws in the SFH group, 64 were due to error in the point of entry, 63 were due to error in the trajectory angle, and 7 were due to both errors. Of 8 errors in the NS group, 7 were due to the point of entry. CONCLUSIONS: Intraoperative navigation had significantly better one-time accuracy of pedicle screw insertion than freehand insertion and should be used to avoid injury to the pedicle and surrounding tissue from screw reinsertion.
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Vértebras Lumbares/cirugía , Neuronavegación/métodos , Procedimientos Neuroquirúrgicos/métodos , Tornillos Pediculares , Anciano , Simulación por Computador , Estudios Transversales , Femenino , Humanos , Imagenología Tridimensional , Desplazamiento del Disco Intervertebral/cirugía , Masculino , Persona de Mediana Edad , Reoperación , Estenosis Espinal/cirugía , Espondilolistesis/cirugíaRESUMEN
OBJECTIVE: To compare the superior-level facet joint violations (FJV) between robot-assisted (RA) percutaneous pedicle screw placement and conventional open fluoroscopic-guided (FG) pedicle screw placement in a prospective cohort study. METHODS: This was a prospective cohort study without randomization. One-hundred patients scheduled to undergo RA (n = 50) or FG (n = 50) transforaminal lumbar interbody fusion were included from February 2016 to May 2018. The grade of FJV, the distance between pedicle screws and the corresponding proximal facet joint, and intra-pedicle accuracy of the top screw were evaluated based on postoperative CT scan. Patient demographics, perioperative outcomes, and radiation exposure were recorded and compared. Perioperative outcomes include surgical time, intraoperative blood loss, postoperative length of stay, conversion, and revision surgeries. RESULTS: Of the 100 screws in the RA group, 4 violated the proximal facet joint, while 26 of 100 in the FG group had FJV (P = 0.000). In the RA group, 3 and 1 screws were classified as grade 1 and 2, respectively. Of the 26 FJV screws in the FG group, 17 screws were scored as grade 1, 6 screws were grade 2, and 3 screws were grade 3. Significantly more severe FJV were noted in the FG group than in the RA group (P = 0.000). There was a statistically significant difference between RA and FG for overall violation grade (0.05 vs 0.38, P = 0.000). The average distance of pedicle screws from facet joints in the RA group (4.16 ± 2.60 mm) was larger than that in the FG group (1.92 ± 1.55 mm; P = 0.000). For intra-pedicle accuracy, the rate of perfect screw position was greater in the RA group than in the FG group (85% vs 71%; P = 0.017). No statistically significant difference was found between the clinically acceptable screws between groups (P = 0.279). The radiation dose was higher in the FG group (30.3 ± 11.3 vs 65.3 ± 28.3 µSv; P = 0.000). The operative time in the RA group was significantly longer (184.7 ± 54.3 vs 117.8 ± 36.9 min; P = 0.000). CONCLUSIONS: Compared to the open FG technique, minimally invasive RA spine surgery was associated with fewer proximal facet joint violations, larger facet to screw distance, and higher intra-pedicle accuracy.
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Vértebras Lumbares/cirugía , Tornillos Pediculares , Procedimientos Quirúrgicos Robotizados/métodos , Fusión Vertebral/métodos , Articulación Cigapofisaria/cirugía , Adulto , Anciano , Femenino , Fluoroscopía , Humanos , Masculino , Persona de Mediana Edad , Estudios ProspectivosRESUMEN
PURPOSE: Paclitaxel is an integral component of primary therapy for breast and epithelial ovarian cancers, but less than half of these cancers respond to the drug. Enhancing the response to primary therapy with paclitaxel could improve outcomes for women with both diseases.Experimental Design: Twelve kinases that regulate metabolism were depleted in multiple ovarian and breast cancer cell lines to determine whether they regulate sensitivity to paclitaxel in Sulforhodamine B assays. The effects of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 2 (PFKFB2) depletion on cell metabolomics, extracellular acidification rate, nicotinamide adenine dinucleotide phosphate, reactive oxygen species (ROS), and apoptosis were studied in multiple ovarian and breast cancer cell lines. Four breast and ovarian human xenografts and a breast cancer patient-derived xenograft (PDX) were used to examine the knockdown effect of PFKFB2 on tumor cell growth in vivo. RESULTS: Knockdown of PFKFB2 inhibited clonogenic growth and enhanced paclitaxel sensitivity in ovarian and breast cancer cell lines with wild-type TP53 (wtTP53). Silencing PFKFB2 significantly inhibited tumor growth and enhanced paclitaxel sensitivity in four xenografts derived from two ovarian and two breast cancer cell lines, and prolonged survival in a triple-negative breast cancer PDX. Transfection of siPFKFB2 increased the glycolysis rate, but decreased the flow of intermediates through the pentose-phosphate pathway in cancer cells with wtTP53, decreasing NADPH. ROS accumulated after PFKFB2 knockdown, which stimulated Jun N-terminal kinase and p53 phosphorylation, and induced apoptosis that depended upon upregulation of p21 and Puma. CONCLUSIONS: PFKFB2 is a novel target whose inhibition can enhance the effect of paclitaxel-based primary chemotherapy upon ovarian and breast cancers retaining wtTP53.
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Neoplasias de la Mama/metabolismo , Resistencia a Antineoplásicos , Neoplasias Ováricas/metabolismo , Paclitaxel/farmacología , Fosfofructoquinasa-2/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Animales , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Modelos Animales de Enfermedad , Resistencia a Antineoplásicos/genética , Femenino , Expresión Génica , Silenciador del Gen , Humanos , Inmunohistoquímica , Redes y Vías Metabólicas , Ratones , Mutación , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/genética , Estrés Oxidativo , Fosfofructoquinasa-2/genética , Interferencia de ARN , Especies Reactivas de Oxígeno/metabolismo , Proteína p53 Supresora de Tumor/genética , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The CCNG2 gene that encodes the unconventional cyclin G2 was one of the few genes up-regulated on anti-human epidermal growth factor receptor 2 (HER2) antibody-mediated inhibition of HER2 signaling. The purpose of this study was to explore how HER2 signaling modulates cyclin G2 expression and the effect of elevated cyclin G2 on breast cancer cell growth. Treatment of breast cancer cells that overexpress HER2 (BT474, SKBr3, and MDAMB453) with the anti-HER2 antibody trastuzumab or its precursor 4D5 markedly up-regulated cyclin G2 mRNA in vitro and in vivo, as shown by real-time PCR. Immunoblot and immunofluorescence analysis with specific antibodies against cyclin G2 showed that anti-HER2 antibody significantly increased cyclin G2 protein expression and translocated the protein to the nucleus. Trastuzumab was not able to induce cyclin G2 expression in cells weakly expressing HER2 (MCF7) or in cells that had developed resistance to trastuzumab. Enforced expression of HER2 in T47D and MDAMB435 breast cancer cells reduced cyclin G2 levels. Collectively, these data suggest that HER2-mediated signaling negatively regulates cyclin G2 expression. Inhibition of phosphoinositide 3-kinase (LY294002), c-jun NH(2)-terminal kinase (SP600125), and mammalian target of rapamycin (mTOR)/p70 S6 kinase (p70S6K; rapamycin) increased cyclin G2 expression. In contrast, treatment with inhibitors of p38 mitogen-activated protein kinase (SB203580), mitogen-activated protein kinase/extracellular signal-regulated kinase kinase 1/2 (U0126), or phospholipase Cgamma (U73122) did not affect cyclin G2 expression. Anti-HER2 antibody in combination with LY294002, rapamycin, or SP600125 induced greater cyclin G2 expression than either agent alone. Ectopic expression of cyclin G2 inhibited cyclin-dependent kinase 2 activity, Rb phosphorylation, cell cycle progression, and cellular proliferation without affecting p27(Kip1) expression. Thus, cyclin G2 expression is modulated by HER2 signaling through multiple pathways including phosphoinositide 3-kinase, c-jun NH(2)-terminal kinase, and mTOR signaling. The negative effects of cyclin G2 on cell cycle and cell proliferation, which occur without altering p27(Kip1) levels, may contribute to the ability of trastuzumab to inhibit breast cancer cell growth.
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Neoplasias de la Mama/enzimología , Neoplasias de la Mama/patología , Ciclinas/genética , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Fosfatidilinositol 3-Quinasas/metabolismo , Receptor ErbB-2/metabolismo , Proteínas Quinasas S6 Ribosómicas 70-kDa/metabolismo , Animales , Anticuerpos Monoclonales/farmacología , Neoplasias de la Mama/genética , Ciclo Celular/efectos de los fármacos , Línea Celular Tumoral , Núcleo Celular/efectos de los fármacos , Núcleo Celular/patología , Proliferación Celular/efectos de los fármacos , Ciclina G2 , Quinasa 2 Dependiente de la Ciclina/metabolismo , Inhibidores Enzimáticos/farmacología , Quinasas MAP Reguladas por Señal Extracelular/antagonistas & inhibidores , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Proteínas Quinasas JNK Activadas por Mitógenos/antagonistas & inhibidores , Ratones , Inhibidores de las Quinasa Fosfoinosítidos-3 , Fosfolipasa C gamma/antagonistas & inhibidores , Fosforilación/efectos de los fármacos , Proteínas Quinasas/metabolismo , Transporte de Proteínas/efectos de los fármacos , Proteína de Retinoblastoma/metabolismo , Serina-Treonina Quinasas TOR , Proteínas Quinasas p38 Activadas por Mitógenos/antagonistas & inhibidoresRESUMEN
Minimally invasive (MI) transforaminal lumbar interbody fusion (TLIF) is a challenging technique with a long learning curve. We combined computer-assisted navigation and MI TLIF (CAMISS TLIF) to treat lumbar degenerative disease. This study aimed to evaluate the learning curve associated with computer-assisted navigation MI spine surgery (CAMISS) and TLIF for the surgical treatment of lumbar degenerative disease. Seventy four consecutive patients with lumbar degenerative disease underwent CAMISS TLIF between March 2011 and May 2015; all surgeries were performed by a single surgeon. According to the plateau of the asymptote, the initial 25 patients constituted the early group and the remaining patients comprised the latter group. The clinical evaluation data included operative times, anesthesia times, intraoperative blood losses, days until ambulation, postoperative hospital stays, visual analog scale (VAS) leg and back pain scores, Oswestry disability index (ODI) values, Macnab outcome scale scores, complications, radiological outcomes, and rates of conversion to open surgery. The complexity of the cases increased over the series, but the complication rate decreased (12.00%-6.12%). There were significant differences between the early and late groups with respect to the average surgical times and durations of anesthesia, but no differences in intraoperative blood losses, days until ambulation, postoperative hospital stays, complication rate, VAS, ODI, Macnab outcome scale scores, or solid fusion rates. There was no need for conversion to open procedures in either group. Our study showed that a plateau asymptote for CAMISS TLIF was reached after 25 operations. The later patients experienced shorter operative times and anesthesia durations.
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Vértebras Lumbares/cirugía , Procedimientos Quirúrgicos Mínimamente Invasivos/métodos , Fusión Vertebral/métodos , Cirujanos/educación , Cirugía Asistida por Computador/métodos , Adulto , Competencia Clínica/estadística & datos numéricos , Estudios de Cohortes , Evaluación de la Discapacidad , Femenino , Humanos , Curva de Aprendizaje , Tiempo de Internación/estadística & datos numéricos , Masculino , Persona de Mediana Edad , Procedimientos Quirúrgicos Mínimamente Invasivos/efectos adversos , Procedimientos Quirúrgicos Mínimamente Invasivos/educación , Tempo Operativo , Complicaciones Posoperatorias/epidemiología , Estudios Retrospectivos , Enfermedades de la Columna Vertebral/cirugía , Fusión Vertebral/efectos adversos , Fusión Vertebral/educación , Cirugía Asistida por Computador/efectos adversos , Cirugía Asistida por Computador/educación , Resultado del Tratamiento , Escala Visual AnalógicaRESUMEN
Purpose: High levels of ROS and ineffective antioxidant systems contribute to oxidative stress, which affects the function of hematopoietic cells in acute myeloid leukemia (AML); however, the mechanisms by which ROS lead to malignant transformation in relapsed AML-M5 are not completely understood. We hypothesized that alterations in intracellular ROS would trigger AML-M5 relapse by activating the intrinsic pathway.Experimental Design: We studied ROS levels and conducted c-Jun activation domain-binding protein-1 (JAB1/COPS5) and thioredoxin (TRX) gene expression analyses with blood samples obtained from 60 matched AML-M5 patients at diagnosis and relapse and conducted mechanism studies of Jab1's regulation of Trx in leukemia cell lines.Results: Our data showed that increased production of ROS and a low capacity of antioxidant enzymes were characteristics of AML-M5, both at diagnosis and at relapse. Consistently, increased gene expression levels of TRX and JAB1/COPS5 were associated with low overall survival rates in patients with AML-M5. In addition, stimulating AML-M5 cells with low concentrations of hydrogen peroxide led to increased Jab1 and Trx expression. Consistently, transfection of ectopic Jab1 into leukemia cells increased Trx expression, whereas silencing of Jab1 in leukemia cells reduced Trx expression. Mechanistically, Jab1 interacted with Trx and stabilized Trx protein. Moreover, Jab1 transcriptionally regulated Trx. Furthermore, depletion of Jab1 inhibited leukemia cell growth both in vitro and in vivoConclusions: We identified a novel Jab1-Trx axis that is a key cellular process in the pathobiologic characteristics of AML-M5. Targeting the ROS/Jab1/Trx pathway could be beneficial in the treatment of AML-M5. Clin Cancer Res; 23(15); 4450-61. ©2017 AACR.
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Complejo del Señalosoma COP9/sangre , Péptidos y Proteínas de Señalización Intracelular/sangre , Leucemia Monocítica Aguda/sangre , Estrés Oxidativo/genética , Péptido Hidrolasas/sangre , Tiorredoxinas/sangre , Adolescente , Adulto , Anciano , Complejo del Señalosoma COP9/genética , Línea Celular Tumoral , Proliferación Celular , Transformación Celular Neoplásica/genética , Niño , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Masculino , Persona de Mediana Edad , Péptido Hidrolasas/genética , Especies Reactivas de Oxígeno/metabolismo , Recurrencia , Transducción de Señal/genética , Tiorredoxinas/genéticaRESUMEN
ARHI, an imprinted putative tumor suppressor gene, encodes a M(r) 26,000 GTP-binding protein that is 60% homologous to ras and rap but has a dramatically different function. ARHI expression is down-regulated in a majority of breast and ovarian cancers. Using a dual adenovirus system, we have reexpressed ARHI in ovarian cancer and breast cancer cells that have lost ARHI expression. Reexpression of ARHI inhibited growth, decreased invasiveness, and induced apoptosis. At 5 days after infection with ARHI adenovirus, 30-45% of MDA-MB-231 breast cancer cells and 5-11% of SKOv3 ovarian cancer cells were apoptotic as judged by a terminal deoxynucleotidyl transferase-mediated nick end labeling assay and by Annexin V staining with flow cytometric analysis. Although poly(ADP-ribose) polymerase could be detected immunohistochemically in the nuclei of apoptotic cells, no activation of the effector caspases (caspase 3, 6, 7, or 12) or the initiator caspases (caspase 8 or 9) could be detected in cell lysates using Western blotting. When gene expression was analyzed on a custom cDNA array that contained 2304 known genes, infection with ARHI adenovirus up-regulated 15 genes relative to control cells infected with LacZ adenovirus. The greatest degree of mRNA up-regulation was observed in a Homo sapiens calpain-like protease. On Western blot analysis, calpain protein was increased 2-3-fold at 3-5 days after infection with ARHI adenovirus. No increase in calpain protein was observed after LacZ adenovirus infection. Calpain cleavage could be detected after ARHI reexpression, and inhibitors of calpain, but not inhibitors of caspase, partially prevented ARHI-induced apoptosis. Consequently, reexpression of ARHI in breast and ovarian cancer cells appears to induce apoptosis through a caspase-independent, calpain-dependent mechanism.
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Apoptosis/fisiología , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Calpaína/metabolismo , Terapia Genética/métodos , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Proteínas de Unión al GTP rho/fisiología , Adenoviridae/genética , Animales , Apoptosis/genética , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/terapia , Calpaína/antagonistas & inhibidores , Inhibidores de Caspasas , Caspasas/metabolismo , Ciclo Celular/genética , Ciclo Celular/fisiología , División Celular/genética , División Celular/fisiología , Femenino , Genes Supresores de Tumor , Humanos , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Neoplasias Ováricas/metabolismo , Neoplasias Ováricas/terapia , Transducción de Señal , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de Xenoinjerto , Proteínas de Unión al GTP rho/biosíntesis , Proteínas de Unión al GTP rho/genéticaRESUMEN
The 70 kDa ribosomal S6 kinase (p70S6K) is important for cell growth and survival. Activation of p70S6K requires sequential phosphorylation of multiple serine and threonine sites often triggered by growth factors and hormones. Here, we report that paclitaxel, a microtubule-damaging agent, induces phosphorylation of p70S6K at threonine 421 and serine 424 (T421/S424) in a concentration- and time-dependent manner in multiple breast and ovarian cancer cell lines demonstrated by a T421/S424 phospho-p70S6K antibody. Phosphoamino-acid analysis and Western blot analysis by serine-/threonine-specific antibodies further confirms that both serine and threonine residues are phosphorylated in p70S6K following treatment with paclitaxel. Paclitaxel-induced p70S6K(T421/S424) phosphorylation requires both de novo RNA and protein synthesis via multiple signaling pathways including ERK1/2 MAP kinase, JNK, PKC, Ca(++), PI3K, and mammalian target of rapamycin (mTOR). Despite phosphorylation of p70S6K(T421/S424), paclitaxel inactivates this kinase in a concentration- and time-dependent manner as illustrated by in vitro kinase assay. Inhibitors of mTOR, PI3K, and Ca(++) impair p70S6K activity, whereas inhibitors of JNK and PKC stimulate p70S6K activity. Inhibition of PKC and JNK prevents paclitaxel-induced p70S6K inactivation. Moreover, the paclitaxel-induced phosphorylation and low activity of p70S6K mainly occurs during mitosis. In summary, paclitaxel is able to induce p70S6K(T421/S424) phosphorylation and decrease its activity in mitotic cells via multiple signaling pathways. Our data suggest that paclitaxel-induced p70S6K(T421/S424) phosphorylation and kinase inactivation are differentially regulated. Our data also indicate that paclitaxel may exert its antitumor effect, at least in part, via inhibition of p70S6K.
Asunto(s)
Antineoplásicos Fitogénicos/farmacología , Mitosis/efectos de los fármacos , Paclitaxel/farmacología , Proteínas Quinasas S6 Ribosómicas/antagonistas & inhibidores , Serina/metabolismo , Transducción de Señal/efectos de los fármacos , Treonina/metabolismo , Western Blotting , Calcio/metabolismo , Humanos , Isoenzimas/antagonistas & inhibidores , Isoenzimas/metabolismo , Proteínas Quinasas Activadas por Mitógenos/antagonistas & inhibidores , Fosfolipasa C gamma , Fosforilación , Pruebas de Precipitina , Células Tumorales Cultivadas , Fosfolipasas de Tipo C/antagonistas & inhibidores , Fosfolipasas de Tipo C/metabolismoRESUMEN
microRNAs (miRNAs/miRs) belong to a class of small noncoding RNAs that can negatively regulate messenger RNA (mRNA) expression of target genes. miRNAs are involved in multiple aspects of ovarian cancer cell dysfunction and the phenotype of ovarian cancer cells can be modified by targeting miRNA expression. miRNA profiling has detected a number of candidate miRNAs with the potential to regulate many important biologic functions in ovarian cancer, but their role still needs to be clarified, given the remarkable heterogeneity among ovarian cancers and the context-dependent role of miRNAs. This review summarizes the data collected from The Cancer Genome Atlas (TCGA) and several other genome-wide projects to identify dysregulated miRNAs in ovarian cancers. Copy number variations (CNVs), epigenetic alterations, and oncogenic mutations are also discussed that affect miRNA levels in ovarian disease. Emphasis is given to the role of particular miRNAs in altering expression of genes in human ovarian cancers with the potential to provide diagnostic, prognostic, and therapeutic targets. Particular attention has been given to TP53, BRCA1/2, CA125 (MUC16), HE4 (WFDC2), and imprinted genes such as ARHI (DIRAS3). A better understanding of the abnormalities in miRNA expression and downstream transcriptional and biologic consequences will provide leads for more effective biomarkers and translational approaches in the management of ovarian cancer.
Asunto(s)
Variación Genética , MicroARNs/genética , Neoplasias Ováricas/genética , ARN Mensajero/genética , Regiones no Traducidas 3' , Variaciones en el Número de Copia de ADN , Epigénesis Genética , Femenino , Regulación Neoplásica de la Expresión Génica , Impresión Genómica , Humanos , Mutación , Neoplasias Ováricas/patologíaRESUMEN
Cyclin-dependent kinase 5 (CDK5) is a cytoplasmic serine/ threonine kinase. Knockdown of CDK5 enhances paclitaxel sensitivity in human ovarian cancer cells. This study explores the mechanisms by which CDK5 regulates paclitaxel sensitivity in human ovarian cancers. Multiple ovarian cancer cell lines and xenografts were treated with CDK5 small interfering RNA (siRNA) with or without paclitaxel to examine the effect on cancer cell viability, cell cycle arrest and tumor growth. CDK5 protein was measured by immunohistochemical staining of an ovarian cancer tissue microarray to correlate CDK5 expression with overall patient survival. Knockdown of CDK5 with siRNAs inhibits activation of AKT which significantly correlates with decreased cell growth and enhanced paclitaxel sensitivity in ovarian cancer cell lines. In addition, CDK5 knockdown alone and in combination with paclitaxel induced G1 cell cycle arrest and caspase 3 dependent apoptotic cell death associated with post-translational upregulation and nuclear translocation of TP53 and p27(Kip1) as well as TP53-dependent transcriptional induction of p21(Cip1) in wild type TP53 cancer cells. Treatment of HEYA8 and A2780 wild type TP53 xenografts in nu/nu mice with CDK5 siRNA and paclitaxel produced significantly greater growth inhibition than either treatment alone. Increased expression of CDK5 in human ovarian cancers correlates inversely with overall survival. CDK5 modulates paclitaxel sensitivity by regulating AKT activation, the cell cycle and caspase-dependent apoptosis. CDK5 inhibition can potentiate paclitaxel activity in human ovarian cancer cells.
Asunto(s)
Antineoplásicos Fitogénicos/farmacología , Apoptosis , Quinasa 5 Dependiente de la Ciclina/fisiología , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Inhibidor p27 de las Quinasas Dependientes de la Ciclina/metabolismo , Fase G1 , Neoplasias Ováricas/patología , Paclitaxel/farmacología , Proteínas Proto-Oncogénicas c-akt/metabolismo , Animales , Línea Celular Tumoral , Quinasa 5 Dependiente de la Ciclina/genética , Activación Enzimática , Femenino , Humanos , Ratones , Ratones Desnudos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
BACKGROUND: 8-chloro-adenosine (8-Cl-Ado) is a unique ribonucleoside analog which is currently in a phase I clinical trial for hematological malignancies. Previously, we demonstrated in breast cancer cells that a 3-day treatment with 10 µM 8-Cl-Ado causes a 90% loss of clonogenic survival. In contrast, there was only a modest induction of apoptosis under these conditions, suggesting an alternative mechanism for the tumoricidal activity of 8-Cl-Ado. METHODS: Cellular metabolism, AMP-activated protein kinase (AMPK) and mammalian target of rapamycin (mTOR) pathway signaling, as well as autophagy induction was evaluated in breast cancer cell lines treated with 8-Cl-Ado. The effects of knocking down essential autophagy factors with small interfering RNA on 8-Cl-Ado-inhibited cell survival was assessed in breast cancer cells by examining apoptosis induction and clonogenic survival. In vivo efficacy of 8-Cl-Ado was measured in two breast cancer orthotopic model systems. RESULTS: We demonstrate that in breast cancer cell lines, the metabolism of 8-Cl-Ado results in depletion of endogenous ATP that subsequently induces the phosphorylation and activation of the energy sensor, AMPK. This was associated with an attenuation of mTOR signaling and an induction of the phosphorylation of the autophagy factor, Unc51-like kinase 1 on Ser555. 8-Cl-Ado-mediated induction of autophagy was evident by increased aggregates of microtubule-associated protein 1 light chain 3B (LC3B) which was associated with its conversion to its lipidated form, LC3B-II, p62 degradative flux, and increased formation of acidic vesicular organelles. Additionally, transfection of MCF-7 cells with siRNA to ATG7 or beclin 1 provided partial protection of the cells to 8-Cl-Ado cytotoxicity as measured by clonogenicity. In vivo, 8-Cl-Ado inhibited growth of both MCF-7 and BT-474 xenograft tumors. Moreover, in 9 of 22 BT-474 tumors treated with 100 mg/kg/day 3 times a week, there was an absence of macroscopically detectable tumor after 3 weeks of treatment. CONCLUSIONS: Our data demonstrates that 8-Cl-Ado treatment activates the AMPK pathway leading to autophagy induction of in breast cancer cells, eliciting, in part, its tumoricidal effects. Additionally, 8-Cl-Ado effectively inhibited in vivo tumor growth in mice. Based on this biological activity, we are planning to test 8-Cl-Ado in the clinic for patients with breast cancer.
Asunto(s)
2-Cloroadenosina/análogos & derivados , Proteínas Quinasas Activadas por AMP/metabolismo , Apoptosis/efectos de los fármacos , Neoplasias de la Mama/tratamiento farmacológico , 2-Cloroadenosina/farmacología , Animales , Autofagia/efectos de los fármacos , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Línea Celular Tumoral , Modelos Animales de Enfermedad , Femenino , Humanos , Células MCF-7 , Ratones , Fosforilación/efectos de los fármacos , Distribución Aleatoria , Transducción de Señal/efectos de los fármacos , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
AIM: Deregulation of FOXM1 has been documented in various cancers. The aim of this study was to evaluate the role of FOXM1 in ovarian cancer tumorigenesis and paclitaxel resistance. EXPERIMENTAL DESIGN: Expression of FOXM1 was examined in 119 clinical samples by immunohistochemistry and correlated with clinicopathological parameters. Effects of FOXM1 knockdown on ovarian cancer cell migration, invasion and mitotic catastrophe were also studied. qPCR and ChIP-qPCR were used to establish KIF2C as a novel FOXM1 target gene implicated in chemoresistance. RESULTS: High nuclear FOXM1 expression in ovarian cancer patient samples was significantly associated with advanced stages (P = 0.035), shorter overall (P = 0.019) and disease-free (P = 0.014) survival. Multivariate analysis confirmed FOXM1 expression as an independent prognostic factor for ovarian cancer. FOXM1 knockdown significantly inhibited migration and invasion of ovarian cancer cells and enhanced paclitaxel-mediated cell death and mitotic catastrophe in a p53-independent manner. Bioinformatics analysis suggested a number of potential transcription targets of FOXM1. One of the potential targets, KIF2C, exhibited similar expression pattern to FOXM1 in chemosensitive and chemoresistant cells in response to paclitaxel treatment. FOXM1 could be detected at the promoter of KIF2C and FOXM1 silencing significantly down-regulated KIF2C. CONCLUSION: Our findings suggest that FOXM1 is associated with poor patient outcome and contributes to paclitaxel resistance by blocking mitotic catastrophe. KIF2C is identified as a novel FOXM1 transcriptional target that may be implicated in the acquisition of chemoresistance. FOXM1 should be further investigated as a potential prognostic marker and therapeutic target for ovarian cancer.
Asunto(s)
Antineoplásicos Fitogénicos/farmacología , Apoptosis/efectos de los fármacos , Factores de Transcripción Forkhead/metabolismo , Neoplasias Ováricas/patología , Paclitaxel/farmacología , Antineoplásicos Fitogénicos/uso terapéutico , Línea Celular Tumoral , Movimiento Celular , Supervivencia sin Enfermedad , Regulación hacia Abajo/efectos de los fármacos , Resistencia a Antineoplásicos , Femenino , Estudios de Seguimiento , Proteína Forkhead Box M1 , Factores de Transcripción Forkhead/antagonistas & inhibidores , Factores de Transcripción Forkhead/genética , Puntos de Control de la Fase G2 del Ciclo Celular , Humanos , Inmunohistoquímica , Estimación de Kaplan-Meier , Cinesinas/genética , Estadificación de Neoplasias , Neoplasias Ováricas/diagnóstico , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/mortalidad , Paclitaxel/uso terapéutico , Pronóstico , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
Baicalein is a purified flavonoid extracted from the roots of Scutellaria baicalensis or Scutellaria radix. Although previous studies have suggested that Baicalein possesses an in vitro anti-hepatocellular carcinoma activity, its in vivo effects and mechanisms of action are still not completely understood. In this study, Baicalein at concentrations of 40-120 µM exhibited significant cytotoxicity to three hepatocellular carcinoma (HCC) cell lines but marginal cytotoxicity to a normal liver cell line in vitro. Compared to a standard chemotherapy drug, 5-fluorouracil (5-FU), Baicalein had greater effect on HCC cells but less toxicity on normal liver cells. Treatment with Baicalein dramatically reduced mitochondrial transmembrane potential, and activated caspase-9 and caspase-3. Blockade of Baicalein-induced apoptosis with a pan-caspase inhibitor partially attenuated Baicalein-induced growth inhibition in HCC. Baicalein treatment significantly inhibited tumor growth of HCC xenografts in mice. Induction of apoptosis was demonstrated in Baicalein-treated xenograft tumors by the terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay. Furthermore, Baicalein treatment dramatically decreased the levels of phosphorylation of MEK1, ERK1/2 and Bad in vitro and in vivo. Overexpression of human MEK1 partially blocked Baicalein-induced growth inhibition. Consequently, these findings suggest that Baicalein preferentially inhibits HCC tumor growth through inhibition of MEK-ERK signaling and by inducing intrinsic apoptosis.
Asunto(s)
Carcinoma Hepatocelular/tratamiento farmacológico , Inhibidores Enzimáticos/farmacología , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Flavanonas/farmacología , Neoplasias Hepáticas/tratamiento farmacológico , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Animales , Apoptosis/efectos de los fármacos , Caspasa 3/biosíntesis , Caspasa 3/efectos de los fármacos , Caspasa 3/metabolismo , Caspasa 9/biosíntesis , Caspasa 9/efectos de los fármacos , Caspasa 9/metabolismo , Línea Celular Tumoral , Fluorouracilo/farmacología , Humanos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Ratones , Fosforilación , Extractos Vegetales , Scutellaria baicalensisRESUMEN
Trastuzumab, a humanized monoclonal antibody directed against the extracellular domain of the HER2 oncoprotein, can effectively target HER2-positive breast cancer through several mechanisms. Although the effects of trastuzumab on cancer cell proliferation, angiogenesis and apoptosis have been investigated in depth, the effect of trastuzumab on microRNA (miRNA) has not been extensively studied. We have performed miRNA microarray profiling before and after trastuzumab treatment in SKBr3 and BT474 human breast cancer cells that overexpress HER2. We found that trastuzumab treatment of SKBr3 cells significantly decreased five miRNAs and increased three others, whereas treatment of BT474 cells significantly decreased two miRNAs and increased nine. The only change in miRNA expression observed in both cell lines following trastuzumab treatment was upregulation of miRNA-194 (miR-194) that was further validated in vitro and in vivo. Forced expression of miR-194 in breast cancer cells that overexpress HER2 produced no effect on apoptosis, modest inhibition of proliferation, significant inhibition of cell migration/invasion in vitro and significant inhibition of xenograft growth in vivo. Conversely, knockdown of miR-194 promoted cell migration. Increased miR-194 expression markedly reduced levels of the cytoskeletal protein talin2 and specifically inhibited luciferase reporter activity of a talin2 wild-type 3'-untranslated region, but not that of a mutant reporter, indicating that talin2 is a direct downstream target of miR-194. Trastuzumab treatment inhibited breast cancer cell migration and reduced talin2 expression in vitro and in vivo. Knockdown of talin2 inhibited cell migration/invasion. Knockdown of trastuzumab-induced miR-194 expression with a miR-194 inhibitor compromised trastuzumab-inhibited cell migration in HER2-overexpressing breast cancer cells. Consequently, trastuzumab treatment upregulates miR-194 expression and may exert its cell migration-inhibitory effect through miR-194-mediated downregulation of cytoskeleton protein talin2 in HER2-overexpressing human breast cancer cells.