RESUMEN
MUC4 mucin is a high molecular weight transmembrane glycoprotein that plays important roles in tumour biology. It is aberrantly expressed in pancreatic adenocarcinoma, while not being detectable in the normal pancreas. Previous studies have demonstrated that the cystic fibrosis transmembrane conductance regulator (CFTR), a chloride channel that is defective in CF, is implicated in multiple cellular functions, including gene regulation. In the present study, using a CFTR-defective pancreatic cancer cell line and its derived subline expressing functional CFTR, we report that MUC4 expression is negatively regulated by CFTR. Short-interfering RNA (siRNA)-mediated silencing of CFTR also leads to an increased expression of MUC4. Additionally, our results suggest that CFTR-mediated regulation of MUC4 is cell density-dependent and is achieved by transcriptional and posttranslational mechanisms. Moreover, in a panel of pancreatic cancer cell lines and normal pancreas, we observed that CFTR was downregulated in pancreatic cancer cells and negatively correlated with MUC4 in most cases. An aberrant expression of MUC4 was also detected in the CF pancreas. Downregulation of CFTR in pancreatic adenocarcinoma and its inverse association with the tumour-linked mucin, MUC4, indicate novel function(s) of CFTR in pancreatic tumour biology and suggest the implication of new signalling pathway(s) in MUC4 regulation.
Asunto(s)
Adenocarcinoma/metabolismo , Regulador de Conductancia de Transmembrana de Fibrosis Quística/fisiología , Mucinas/genética , Neoplasias Pancreáticas/metabolismo , Procesamiento Proteico-Postraduccional , Adenocarcinoma/patología , Secuencia de Bases , Northern Blotting , Línea Celular Tumoral , Cartilla de ADN , Humanos , Mucina 4 , Neoplasias Pancreáticas/patología , Regiones Promotoras Genéticas , ARN Mensajero/genética , ARN Interferente Pequeño , Reacción en Cadena de la Polimerasa de Transcriptasa InversaRESUMEN
Treatment of SENCAR mouse skin with dibenzo[a,l]pyrene results in abundant formation of abasic sites that undergo error-prone excision repair, forming oncogenic H-ras mutations in the early preneoplastic period. To examine whether the abundance of abasic sites causes repair infidelity, we treated SENCAR mouse skin with estradiol-3,4-quinone (E(2)-3,4-Q) and determined adduct levels 1 h after treatment, as well as mutation spectra in the H-ras gene between 6 h and 3 days after treatment. E(2)-3,4-Q formed predominantly (> or =99%) the rapidly-depurinating 4-hydroxy estradiol (4-OHE(2))-1-N3Ade adduct and the slower-depurinating 4-OHE(2)-1-N7Gua adduct. Between 6 h and 3 days, E(2)-3,4-Q induced abundant A to G mutations in H-ras DNA, frequently in the context of a 3'-G residue. Using a T.G-DNA glycosylase (TDG)-PCR assay, we determined that the early A to G mutations (6 and 12 h) were in the form of G.T heteroduplexes, suggesting misrepair at A-specific depurination sites. Since G-specific mutations were infrequent in the spectra, it appears that the slow rate of depurination of the N7Gua adducts during active repair may not generate a threshold level of G-specific abasic sites to affect repair fidelity. These results also suggest that E(2)-3,4-Q, a suspected endogenous carcinogen, is a genotoxic compound and could cause mutations.
Asunto(s)
Aductos de ADN/genética , Daño del ADN/genética , Reparación del ADN/genética , Estradiol/análogos & derivados , Genes ras/genética , Mutagénesis/genética , Piel/metabolismo , Animales , Artefactos , Secuencia de Bases , Aductos de ADN/química , Aductos de ADN/efectos de los fármacos , Daño del ADN/efectos de los fármacos , Análisis Mutacional de ADN , Reparación del ADN/efectos de los fármacos , Estradiol/química , Estradiol/farmacología , Femenino , Ratones , Ratones Endogámicos SENCAR , Mutágenos/química , Mutágenos/farmacología , Ácidos Nucleicos Heterodúplex/efectos de los fármacos , Ácidos Nucleicos Heterodúplex/genética , Mutación Puntual/genética , Reacción en Cadena de la Polimerasa , Piel/efectos de los fármacosRESUMEN
PURPOSE: The study goal was to improve outcome in children with rhabdomyosarcoma by comparing risk-based regimens of surgery, radiotherapy (RT) and chemotherapy. PATIENTS AND METHODS: Eight hundred eighty-three previously untreated eligible patients with nonmetastatic rhabdomyosarcoma entered the Intergroup Rhabdomyosarcoma Study-IV (IRS-IV) (1991 to 1997) after surgery and were randomized treatment by primary tumor site, group (1 to 3), and stage (I to III). Failure-free survival (FFS) rates and survival were the end points used in comparisons between randomized groups and between patient subgroups treated on IRS-III and IRS-IV. Most patients were randomized to receive vincristine and dactinomycin (VA) and cyclophosphamide (VAC, n = 235), or VA and ifosfamide (VAI, n = 222), or vincristine, ifosfamide, and etoposide (VIE, n = 236). Patients with group 3 tumors were randomized to receive conventional RT (C-RT) versus hyperfractionated RT (HF-RT). RESULTS: Overall 3-year FFS and survival were 77% and 86%, respectively. Three-year FFS rates with VAC, VAI, and VIE were 75%, 77%, and 77%, respectively (P =.42). No significant difference in outcome was noted with HF-RT versus C-RT (P =.85 and P =.90, respectively). Overall, patients with embryonal tumors benefited from intensive three-drug chemotherapy in IRS-IV (3-year FFS, 83%). The improvement was seen for patients with stage I or stage II/III, group 1/2 disease, many of whom received VA chemotherapy on IRS-III. Patients with stage 2/3, group 3 disease had similar outcomes on IRS-III and IRS-IV. Three-year FFS for the nonrandomized patient subsets was 75% with renal abnormalities; 81% for paratesticular, group 1 cases; and 91% for group 1/2 orbit or eyelid tumors. Patients with paratesticular primaries had poorer outcomes if they were more than 10 years old (3-year FFS, 63% v 90%). Myelosuppression occurred in most patients, but toxic deaths occurred in less than 1%. CONCLUSION: VAC and VAI or VIE with surgery (with or without RT), are equally effective for patients with local or regional rhabdomyosarcoma and are more effective for embryonal tumors than therapies used previously. Younger patients with group 1 paratesticular embryonal tumors and all patients with group 1/2 orbit or eyelid tumors can usually be cured with VA chemotherapy along with postoperative RT for group 2 disease.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Radioterapia/métodos , Rabdomiosarcoma/terapia , Adolescente , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Niño , Preescolar , Terapia Combinada/métodos , Supervivencia sin Enfermedad , Fraccionamiento de la Dosis de Radiación , Neoplasias de los Párpados/mortalidad , Neoplasias de los Párpados/patología , Neoplasias de los Párpados/terapia , Femenino , Humanos , Lactante , Masculino , Neoplasias Orbitales/mortalidad , Neoplasias Orbitales/patología , Neoplasias Orbitales/terapia , Pronóstico , Estudios Retrospectivos , Rabdomiosarcoma/mortalidad , Rabdomiosarcoma/patología , Tasa de Supervivencia , Neoplasias Testiculares/mortalidad , Neoplasias Testiculares/patología , Neoplasias Testiculares/terapiaRESUMEN
Lenalidomide (LEN) and metronomic cyclophosphamide (CTX) regulate angiogenesis and immunosuppressive cells linked to the progression of metastatic castration-resistant prostate cancer (mCRPC). A phase-I/II, dose-escalation trial of LEN plus oral CTX was conducted in patients with previously treated mCRPC. In the phase-I study, CTX was given at 50 mg (day 1-28) and LEN at 10-25 mg (day 1-21) on a 28-day cycle using a "3+3" study design. In phase II, patients received LEN at 25 mg (day 1-21) with CTX at 50 mg PO QD (day 1-28) on a 28-day cycle. Nineteen patients in phase I were evaluable for toxicity. The maximum tolerated dose (MTD) was not observed at any of the dose levels (DLs) tested. Six patients received treatment in phase II before the trial was closed. A ≥ 50% reduction in PSA was observed in 31.7% evaluable patients. Radiographically, one patient had a partial response. Stable disease was documented in 68% of evaluable patients after two therapy cycles. Circulating tumor cells (CTCs) decreased in 22.7% and remained stable in 31.8% of patients. Baseline numbers of peripheral MDSCs (MDSC; Lin-DR(-)CD11b(+)) were significantly increased in patients versus normal donors, and were decreased by chemotherapy. At baseline, MDSCs correlated directly with CTCs, and inversely with T- and B cell frequency supporting their immunosuppressive activity. The combination of LEN and metronomic CTX can be safely administered, reversing cellular immunosuppression in this group of elderly patients with mCRPC. Further research is required to identify responsive subgroup(s) and validate the biomarkers.
Asunto(s)
Ciclofosfamida/uso terapéutico , Regulación Neoplásica de la Expresión Génica , Inmunosupresores , Neoplasias de la Próstata Resistentes a la Castración/tratamiento farmacológico , Talidomida/análogos & derivados , Anciano , Anciano de 80 o más Años , Antineoplásicos/uso terapéutico , Citocinas/metabolismo , Relación Dosis-Respuesta a Droga , Ensayo de Inmunoadsorción Enzimática , Citometría de Flujo , Humanos , Inmunosupresores/uso terapéutico , Estimación de Kaplan-Meier , Lenalidomida , Leucocitos/citología , Masculino , Dosis Máxima Tolerada , Persona de Mediana Edad , Células Mieloides/citología , Células Neoplásicas Circulantes , Antígeno Prostático Específico/metabolismo , Neoplasias de la Próstata Resistentes a la Castración/inmunología , Talidomida/uso terapéutico , Factores de Tiempo , Resultado del TratamientoRESUMEN
1. The electrophysiological responsivity to desipramine (DMI) applied by systemic or local route was tested in Wistar female rats submitted to a wide lesion in the lateral septal area on the 8th day after birth. 2. One year after a septal lesion, the dorsal raphe nucleus (DRN) stimulation produced an initial brief response in lateral septal and CA1/CA3 neurons. In the control and sham-lesion groups, most recordings showed only this initial brief response (non-late responding neurons); however, in the lesion group most of the recorded neurons showed an afterdischarge. 3. DMI (2.14 mg/kg, 21 days, i.p.) increased the firing rate in septal and CA1/CA3 non-late responding neurons. In the equivalent septal neurons from lesion group, DMI produced the inverse effect, i.e., a decreased firing rate. 4. In septal non-late responding neurons, DMI (2 mM; 10-15 nAmps, 0.5 sec) applied by microiontophoresis increased the firing rate only after long-term systemic DMI impregnation. The response to locally applied DMI did not occur in the lesion group even after long-term DMI treatment. 5. In conclusion, an early lateral septal lesion canceled the response of intermediate-dorsal septal neurons to DMI applied by systemic and local routes.
Asunto(s)
Antidepresivos Tricíclicos/farmacología , Lesiones Encefálicas/fisiopatología , Desipramina/farmacología , Neuronas/efectos de los fármacos , Núcleos del Rafe/fisiología , Animales , Estimulación Eléctrica , Electrofisiología , Femenino , Iontoforesis , Potenciales de la Membrana/fisiología , Núcleos del Rafe/citología , Ratas , Ratas WistarRESUMEN
MUC4 is a large, heavily glycosylated transmembrane mucin, that is implicated in the pathogenesis of various types of cancers. To date, no extensive study has been done to check the expression and functional significance of MUC4 in different types of gastric adenocarcinomas. Here, we report the expression profile of MUC4 in gastric adenocarcinomas and its function in poorly differentiated gastric non-signet ring cell carcinoma (non-SRCC) type cells. Immunohistochemical analysis using tissue microarray (TMA) showed a significant difference in MUC4 expression between normal adjacent (n = 45) and gastric adenocarcinoma (n = 83; P < 0.001). MUC4 expression was not associated with tumour type, stage or with the degree of differentiation. To gain further insight into the significance of MUC4 expression in gastric non-SRCC cells, MUC4 was ectopically expressed in AGS, a poorly differentiated gastric non-signet ring cell line. The MUC4 overexpressing cells (AGS-MUC4) showed a significant increase (P < 0.005) in cell motility and a decrease in cellular aggregation as compared with the vector-transfected cells. Furthermore, in vivo tumorigenicity analysis revealed that animals transplanted with the MUC4 overexpressing cells (AGS-MUC4) had a greater incidence of tumours (83%) in comparison to empty vector control (17%). In addition, the expression of MUC4 resulted in enhanced expression of total cellular ErbB2 and phosphorylated ErbB2. In conclusion, our results showed that MUC4 is overexpressed in gastric adenocarcinoma tissues, and that it has a role in promoting aggressive properties in poorly differentiated gastric non-SRCC cells through the activation of the ErbB2 oncoprotein.
Asunto(s)
Adenocarcinoma/metabolismo , Carcinoma de Células en Anillo de Sello/metabolismo , Diferenciación Celular , Mucina 4/metabolismo , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patología , Adenocarcinoma/secundario , Animales , Biomarcadores de Tumor/genética , Biomarcadores de Tumor/metabolismo , Carcinoma de Células en Anillo de Sello/secundario , Movimiento Celular/fisiología , Técnica del Anticuerpo Fluorescente , Mucosa Gástrica/metabolismo , Mucosa Gástrica/patología , Humanos , Immunoblotting , Técnicas para Inmunoenzimas , Inmunoprecipitación , Ratones , Ratones Desnudos , Estadificación de Neoplasias , Receptor ErbB-2/metabolismo , Análisis de Matrices Tisulares , Transfección , Células Tumorales CultivadasRESUMEN
MUC4 is a type-1 transmembrane glycoprotein and is overexpressed in many carcinomas. It is a heterodimeric protein of 930 kDa, composed of a mucin-type subunit, MUC4alpha, and a membrane-bound growth factor-like subunit, MUC4beta. MUC4 mRNA contains unique 5' and 3' coding sequences along with a large variable number of tandem repeat (VNTR) domain of 7-19 kb. A direct association of MUC4 overexpression has been established with the degree of invasiveness and poor prognosis of pancreatic cancer. To understand the precise role of MUC4 in pancreatic cancer, we engineered a MUC4 complementary DNA construct, mini-MUC4, whose deduced protein (320 kDa) is comparable with that of wild-type MUC4 (930 kDa) but represents only 10% of VNTR. Stable ectopic expression of mini-MUC4 in two human pancreatic cancer cell lines, Panc1 and MiaPaCa, showed that MUC4 minigene expression follows a biosynthesis and localisation pattern similar to the wild-type MUC4. Expression of MUC4 resulted in increased growth, motility, and invasiveness of the pancreatic cancer cells in vitro. Ultra-structural examination of MUC4-transfected cells showed the presence of increased number and size of mitochondria. The MUC4-expressing cells also demonstrated an enhanced tumorigenicity in an orthotopic xenograft nude mice model, further supporting a direct role of MUC4 in inducing the cancer properties. In conclusion, our results suggest that MUC4 promotes tumorigenicity and is directly involved in growth and survival of the cancer cells.