RESUMEN

Objective: To determine the rate of adverse events (AE) in women who self-manage their vaginal ring pessary on a monthly basis. We hypothesised that the AE rate would be lower compared to previously published traditional management protocols. Study design: Audit study of 75 women with pelvic organ prolapse and/or stress incontinence, who were fitted with a vaginal ring pessary during a five-year period, and who have self-managed their vaginal rings for at least two years, in a tertiary referral urogynaecology clinic. Main outcome measures: AEs included vaginal bleeding, malodorous vaginal discharge, extrusion of the device, pain/discomfort, and disorders of defaecation or de novo urinary incontinence. AEs that led to discontinuation of usage were termed "major". Results: Of the 75 women who were taught to self-manage their ring pessary, 68 were initially successful. At a median follow-up of 50.5 months [IQR 43-76 months; median 4.2 years], 36 women (52.9%) were still using their ring pessary. Five women (7.4%) had vaginal erosions and bleeding leading them to cease pessary use (four proceeded to surgery). Three minor AEs were identified (4.4%), resolving after discontinuation of ring use two weeks. Thus, the overall AE rate was 11.8% (8/68). Conclusions: In contrast to previous published AE rates of 43-56% in women having ring changes at a clinic every 4-6 months, the AE rate was 12% in the women who performed monthly self-management of vaginal ring pessaries. Such information should be made available to patients considering a vaginal ring pessary.

RESUMEN

INTRODUCTION AND HYPOTHESIS: The continence dish has been a treatment option since 2002 for women with stress urinary incontinence (SUI) who decline surgery, but few quantitative objective efficacy data are published. We aimed to determine the efficacy and acceptability of this device for pure SUI or mixed incontinence (MUI). METHODS: Prospective interventional cohort study of 100 women with SUI or stress-predominant MUI who were interested to use the device; International Consultation on Incontinence Questionnaire (ICIQ) was primary outcome measure; 24-h pad test and Incontinence Impact Questionnaire (IIQ) were secondary outcomes. Acceptability was determined by device retention for 4 weeks, adverse events and ability to self-insert the device. RESULTS: Of 100 suitable women, 9 were not actually fitted, and 27 did not complete (acceptability: 64/100). The rate of adverse events was 7.7%, with 62.5% of users able to self-insert the device: 22 (34%) had pure SUI; 66% had MUI. In SUI, 68% were 'dry' on ICIQ median value 4.0 (IQR 2.5-8.5); 88% were dry on 24-h pad test (median 0.0, IQR 0.0-8.5). The "dry rate" was lower in MUI: 36% for ICIQ (median 9.0, IQR 5.0-15.0) and 62% for 24-h pad test (median 6.2, IQR 0.95-19.7). A "good" response on IIQ occurred in 88% of SUI and 69% of MUI. CONCLUSION: These new data showing strong objective benefits of the continence dish should be further validated by randomized trials, but this information should be made available to women seeking treatment options for SUI/MUI (particularly in view of concerns regarding mesh mid-urethral slings).

Asunto(s)

Cabestrillo Suburetral , Incontinencia Urinaria de Esfuerzo , Incontinencia Urinaria , Estudios de Cohortes , Femenino , Humanos , Estudios Prospectivos , Resultado del Tratamiento , Incontinencia Urinaria/cirugía , Incontinencia Urinaria de Esfuerzo/cirugía

RESUMEN

AIM: Because bacterial cystitis is common in women with refractory detrusor overactivity, the aim was to compare the efficacy of 6 weeks of rotating antibiotics versus placebo, in conjunction with an anticholinergic, in controlling the symptoms of urge incontinence. METHODS: In a multicenter phase IIb double-blinded randomized placebo-controlled trial, women with urodynamically proven refractory detrusor overactivity were randomized in a 2:1 ratio of antibiotics versus placebo for 6 weeks, in addition to darifenacin for 6 months. Any woman with disabling cystitis symptoms was given appropriate antibiotics ("clinical override"). The primary outcome was the degree of urge incontinence change at 6 weeks and 6 months on 24-h pad test. Secondary outcomes were changes in leaks and voids per day measured on 3-day bladder diary and quality of life measures. Microbiological data were collected at all visits. RESULTS: Although 278 women were screened, only 36 were randomized and 33 (91.7%) completed the trial. Leakage on 24-h pad test decreased at 6 months by 75 g in patients receiving antibiotics versus 35 g in placebo. Cure of urge incontinence occurred at 6 months in 10/21 (48%) of antibiotics versus 2/12 (17%) of placebo. Clinical override, necessitating treatment of cystitis, occurred in 41.6% of placebo versus 16.7% of the antibiotic group by 6 months. CONCLUSION: Despite the small sample size, the study showed a significant reduction in pad leakage and leaks per day over 24 h in the active treatment group over a 6-month period. Nearly half of patients on placebo had disabling urinary tract infection symptoms that required clinical override treatment.

Asunto(s)

Antibacterianos/uso terapéutico , Vejiga Urinaria Hiperactiva/tratamiento farmacológico , Método Doble Ciego , Femenino , Humanos , Persona de Mediana Edad , Resultado del Tratamiento

RESUMEN

RAS mutant (MT) metastatic colorectal cancer (mCRC) is resistant to MEK1/2 inhibition and remains a difficult-to-treat group. Therefore, there is an unmet need for novel treatment options for RASMT mCRC. RALA and RALB GTPases function downstream of RAS and have been found to be key regulators of several cell functions implicated in KRAS-driven tumorigenesis. However, their role as regulators of the apoptotic machinery remains to be elucidated. Here, we found that inhibition of RALB expression, but not RALA, resulted in Caspase-8-dependent cell death in KRASMT CRC cells, which was not further increased following MEK1/2 inhibition. Proteomic analysis and mechanistic studies revealed that RALB depletion induced a marked upregulation of the pro-apoptotic cell surface TRAIL Death Receptor 5 (DR5) (also known as TRAIL-R2), primarily through modulating DR5 protein lysosomal degradation. Moreover, DR5 knockdown or knockout attenuated siRALB-induced apoptosis, confirming the role of the extrinsic apoptotic pathway as a regulator of siRALB-induced cell death. Importantly, TRAIL treatment resulted in the association of RALB with the death-inducing signalling complex (DISC) and targeting RALB using pharmacologic inhibition or RNAi approaches triggered a potent increase in TRAIL-induced cell death in KRASMT CRC cells. Significantly, high RALB mRNA levels were found in the poor prognostic Colorectal Cancer Intrinsic Subtypes (CRIS)-B CRC subgroup. Collectively, this study provides to our knowledge the first evidence for a role for RALB in apoptotic priming and suggests that RALB inhibition may be a promising strategy to improve response to TRAIL treatment in poor prognostic RASMT CRIS-B CRC.

Asunto(s)

Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/metabolismo , GTP Fosfohidrolasas/metabolismo , Proteínas Proto-Oncogénicas p21(ras)/genética , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Ligando Inductor de Apoptosis Relacionado con TNF/farmacología , Proteínas de Unión al GTP ral/metabolismo , Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Bencimidazoles/administración & dosificación , Neoplasias Colorrectales/genética , Humanos , Mutación , Proteínas Proto-Oncogénicas p21(ras)/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/agonistas , Proteínas Recombinantes/farmacología , Ligando Inductor de Apoptosis Relacionado con TNF/administración & dosificación , Transfección , Proteínas de Unión al GTP ral/antagonistas & inhibidores , Proteínas de Unión al GTP ral/biosíntesis , Proteínas de Unión al GTP ral/genética

RESUMEN

p53 is the most frequently mutated, well-studied tumor-suppressor gene, yet the molecular basis of the switch from p53-induced cell-cycle arrest to apoptosis remains poorly understood. Using a combination of transcriptomics and functional genomics, we unexpectedly identified a nodal role for the caspase-8 paralog and only human pseudo-caspase, FLIP(L), in regulating this switch. Moreover, we identify FLIP(L) as a direct p53 transcriptional target gene that is rapidly up-regulated in response to Nutlin-3A, an MDM2 inhibitor that potently activates p53. Genetically or pharmacologically inhibiting expression of FLIP(L) using siRNA or entinostat (a clinically relevant class-I HDAC inhibitor) efficiently promoted apoptosis in colorectal cancer cells in response to Nutlin-3A, which otherwise predominantly induced cell-cycle arrest. Enhanced apoptosis was also observed when entinostat was combined with clinically relevant, p53-activating chemotherapy in vitro, and this translated into enhanced in vivo efficacy. Mechanistically, FLIP(L) inhibited p53-induced apoptosis by blocking activation of caspase-8 by the TRAIL-R2/DR5 death receptor; notably, this activation was not dependent on receptor engagement by its ligand, TRAIL. In the absence of caspase-8, another of its paralogs, caspase-10 (also transcriptionally up-regulated by p53), induced apoptosis in Nutlin-3A-treated, FLIP(L)-depleted cells, albeit to a lesser extent than in caspase-8-proficient cells. FLIP(L) depletion also modulated transcription of canonical p53 target genes, suppressing p53-induced expression of the cell-cycle regulator p21 and enhancing p53-induced up-regulation of proapoptotic PUMA. Thus, even in the absence of caspase-8/10, FLIP(L) silencing promoted p53-induced apoptosis by enhancing PUMA expression. Thus, we report unexpected, therapeutically relevant roles for FLIP(L) in determining cell fate following p53 activation.

Asunto(s)

Proteína Reguladora de Apoptosis Similar a CASP8 y FADD/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Acetilación , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/genética , Benzamidas/farmacología , Caspasa 8/metabolismo , Ciclo Celular/efectos de los fármacos , Ciclo Celular/genética , Línea Celular Tumoral , Sinergismo Farmacológico , Regulación de la Expresión Génica , Humanos , Imidazoles/metabolismo , Modelos Biológicos , Piperazinas/metabolismo , Unión Proteica , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas c-mdm2/metabolismo , Piridinas/farmacología , Receptores del Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Ligando Inductor de Apoptosis Relacionado con TNF/metabolismo , Proteína p53 Supresora de Tumor/genética

RESUMEN

BRAFV600E mutations occur in ∼10% of colorectal cancer cases, are associated with poor survival, and have limited responses to BRAF/MEK inhibition with or without EGFR inhibition. There is an unmet need to understand the biology of poor prognostic BRAFMT colorectal cancer. We have used differential gene expression and pathway analyses of untreated stage II and stage III BRAFMT (discovery set: n = 31; validation set: n = 26) colorectal cancer, and an siRNA screen to characterize the biology underpinning the BRAFMT subgroup with poorest outcome. These analyses identified the unfolded protein response (UPR) as a novel and druggable pathway associated with the BRAFMT colorectal cancer subgroup with poorest outcome. We also found that oncogenic BRAF drives endoplasmic reticulum (ER) stress and UPR pathway activation through MEK/ERK. Furthermore, inhibition of GRP78, the master regulator of the UPR, using siRNA or small molecule inhibition, resulted in acute ER stress and apoptosis, in particular in BRAFMT colorectal cancer cells. In addition, dual targeting of protein degradation using combined Carfilzomib (proteasome inhibitor) and ACY-1215 (HDAC6-selective inhibitor) treatment resulted in marked accumulation of protein aggregates, acute ER stress, apoptosis, and therapeutic efficacy in BRAFMT in vitro and xenograft models. Mechanistically, we found that the apoptosis following combined Carfilzomib/ACY-1215 treatment is mediated through increased CHOP expression. Taken together, our findings indicate that oncogenic BRAF induces chronic ER stress and that inducers of acute ER stress could be a novel treatment strategy for poor prognostic BRAFMT colorectal cancer. Mol Cancer Ther; 17(6); 1280-90. ©2018 AACR.

Asunto(s)

Neoplasias Colorrectales/genética , Neoplasias Colorrectales/metabolismo , Mutación , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas B-raf/antagonistas & inhibidores , Proteínas Proto-Oncogénicas B-raf/genética , Respuesta de Proteína Desplegada/efectos de los fármacos , Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/genética , Biomarcadores de Tumor , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/mortalidad , Chaperón BiP del Retículo Endoplásmico , Estrés del Retículo Endoplásmico/efectos de los fármacos , Estrés del Retículo Endoplásmico/genética , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Humanos , Ácidos Hidroxámicos/farmacología , Sistema de Señalización de MAP Quinasas , Modelos Biológicos , Oligopéptidos/farmacología , Pronóstico , Biosíntesis de Proteínas , Proteínas Proto-Oncogénicas B-raf/metabolismo , Pirimidinas/farmacología , Transducción de Señal/efectos de los fármacos , Factor de Transcripción CHOP/genética , Factor de Transcripción CHOP/metabolismo

Asunto(s)

Medios de Contraste/efectos adversos , Extravasación de Materiales Terapéuticos y Diagnósticos/prevención & control , Yohexol/efectos adversos , Yopamidol/efectos adversos , Mejoramiento de la Calidad , Tomografía Computarizada por Rayos X , Sistemas de Registro de Reacción Adversa a Medicamentos , Anciano , Femenino , Humanos , Masculino , Persona de Mediana Edad , Dimensión del Dolor , Sistema de Registros , Estudios Retrospectivos , Factores de Riesgo

Asunto(s)

Aneurisma Coronario/etiología , Fiebre de Origen Desconocido/etiología , Síndrome Mucocutáneo Linfonodular/diagnóstico , Sinovitis/diagnóstico , Recuento de Células Sanguíneas , Preescolar , Diagnóstico Diferencial , Articulación de la Cadera , Humanos , Masculino , Síndrome Mucocutáneo Linfonodular/complicaciones , Sinovitis/etiología

RESUMEN

A recent phase II study of patients with metastatic colorectal carcinoma showed that mismatch repair gene status was predictive of clinical response to PD-1-targeting immune checkpoint blockade. Further examination revealed strong correlation between PD-L1 protein expression and microsatellite instability (MSI) in stage IV colorectal carcinoma, suggesting that the amount of PD-L1 protein expression could identify late-stage patients who might benefit from immunotherapy. To assess whether the clinical associations between PD-L1 gene expression and MSI identified in metastatic colorectal carcinoma are also present in stage II/III colorectal carcinoma, we used in silico analysis to elucidate the cell types expressing the PD-L1 gene. We found a statistically significant association of PD-L1 gene expression with MSI in early-stage colorectal carcinoma (P < 0.001) and show that, unlike in non-colorectal carcinoma tumors, PD-L1 is derived predominantly from the immune infiltrate. We demonstrate that PD-L1 gene expression has positive prognostic value in the adjuvant disease setting (PD-L1(low) vs. PD-L1(high) HR = 9.09; CI, 2.11-39.10). PD-L1 gene expression had predictive value, as patients with high PD-L1 expression appear to be harmed by standard-of-care treatment (HR = 4.95; CI, 1.10-22.35). Building on the promising results from the metastatic colorectal carcinoma PD-1-targeting trial, we provide compelling evidence that patients with PD-L1(high)/MSI/immune(high) stage II/III colorectal carcinoma should not receive standard chemotherapy. This conclusion supports the rationale to clinically evaluate this patient subgroup for PD-1 blockade treatment. Cancer Immunol Res; 4(7); 582-91. ©2016 AACR.

Asunto(s)

Antígeno B7-H1/genética , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/inmunología , Expresión Génica , Anciano , Anciano de 80 o más Años , Quimioterapia Adyuvante , Análisis por Conglomerados , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/patología , Bases de Datos de Ácidos Nucleicos , Femenino , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Humanos , Linfocitos Infiltrantes de Tumor/inmunología , Linfocitos Infiltrantes de Tumor/metabolismo , Masculino , Inestabilidad de Microsatélites , Persona de Mediana Edad , Estadificación de Neoplasias , Pronóstico , Análisis de Supervivencia , Resultado del Tratamiento

RESUMEN

PURPOSE: A number of independent gene expression profiling studies have identified transcriptional subtypes in colorectal cancer with potential diagnostic utility, culminating in publication of a colorectal cancer Consensus Molecular Subtype classification. The worst prognostic subtype has been defined by genes associated with stem-like biology. Recently, it has been shown that the majority of genes associated with this poor prognostic group are stromal derived. We investigated the potential for tumor misclassification into multiple diagnostic subgroups based on tumoral region sampled. EXPERIMENTAL DESIGN: We performed multiregion tissue RNA extraction/transcriptomic analysis using colorectal-specific arrays on invasive front, central tumor, and lymph node regions selected from tissue samples from 25 colorectal cancer patients. RESULTS: We identified a consensus 30-gene list, which represents the intratumoral heterogeneity within a cohort of primary colorectal cancer tumors. Using a series of online datasets, we showed that this gene list displays prognostic potential HR = 2.914 (confidence interval 0.9286-9.162) in stage II/III colorectal cancer patients, but in addition, we demonstrated that these genes are stromal derived, challenging the assumption that poor prognosis tumors with stem-like biology have undergone a widespread epithelial-mesenchymal transition. Most importantly, we showed that patients can be simultaneously classified into multiple diagnostically relevant subgroups based purely on the tumoral region analyzed. CONCLUSIONS: Gene expression profiles derived from the nonmalignant stromal region can influence assignment of colorectal cancer transcriptional subtypes, questioning the current molecular classification dogma and highlighting the need to consider pathology sampling region and degree of stromal infiltration when employing transcription-based classifiers to underpin clinical decision making in colorectal cancer. Clin Cancer Res; 22(16); 4095-104. ©2016 AACRSee related commentary by Morris and Kopetz, p. 3989.

Asunto(s)

Biomarcadores de Tumor , Neoplasias Colorrectales/diagnóstico , Neoplasias Colorrectales/genética , Perfilación de la Expresión Génica , Perfilación de la Expresión Génica/métodos , Regulación Neoplásica de la Expresión Génica , Humanos , Metástasis Linfática , Estadificación de Neoplasias , Especificidad de Órganos/genética , Células del Estroma/metabolismo , Transcriptoma

RESUMEN

There are currently no approved targeted therapies for advanced KRAS mutant (KRASMT) colorectal cancer (CRC). Using a unique systems biology approach, we identified JAK1/2-dependent activation of STAT3 as the key mediator of resistance to MEK inhibitors in KRASMT CRC in vitro and in vivo. Further analyses identified acute increases in c-MET activity following treatment with MEK inhibitors in KRASMT CRC models, which was demonstrated to promote JAK1/2-STAT3-mediated resistance. Furthermore, activation of c-MET following MEK inhibition was found to be due to inhibition of the ERK-dependent metalloprotease ADAM17, which normally inhibits c-MET signaling by promoting shedding of its endogenous antagonist, soluble "decoy" MET. Most importantly, pharmacological blockade of this resistance pathway with either c-MET or JAK1/2 inhibitors synergistically increased MEK-inhibitor-induced apoptosis and growth inhibition in vitro and in vivo in KRASMT models, providing clear rationales for the clinical assessment of these combinations in KRASMT CRC patients.

Asunto(s)

Proteínas ADAM/metabolismo , Neoplasias Colorrectales/tratamiento farmacológico , Quinasas Quinasa Quinasa PAM/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Proto-Oncogénicas c-met/metabolismo , Proteínas Proto-Oncogénicas/genética , Factor de Transcripción STAT3/metabolismo , Proteínas ras/genética , Proteínas ADAM/genética , Proteína ADAM17 , Animales , Apoptosis/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Neoplasias Colorrectales/enzimología , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/metabolismo , Resistencia a Antineoplásicos , Femenino , Células HCT116 , Humanos , Quinasas Quinasa Quinasa PAM/metabolismo , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Proteínas Proto-Oncogénicas/metabolismo , Proteínas Proto-Oncogénicas p21(ras) , Transducción de Señal , Ensayos Antitumor por Modelo de Xenoinjerto , Proteínas ras/metabolismo

RESUMEN

PURPOSE: Pre-operative chemoradiation (CRT) is currently the standard of care for patients with clinical stage II and III rectal cancer but only about 45% of patients achieve tumor downstaging and <20% of patients achieve a pathologic complete response. Better methods to stratify patients according to potential neoadjuvant treatment response are needed. We used microarray analysis to identify a genetic signature that correlates with a pathological complete response (pCR) to neoadjuvant CRT. We performed a gene network analysis to identify potential signaling pathways involved in determining response to neoadjuvant treatment. PATIENTS AND METHODS: We identified 31 T3-4 N0-1 rectal cancer patients who were treated with neoadjuvant fluorouracil-based CRT. Eight patients were identified to have achieved a pCR to treatment while 23 patients did not. mRNA expression was analyzed using cDNA microarrays. The correlation between mRNA expression and pCR from pre-treatment tumor biopsies was determined. Gene network analysis was performed for the genes represented by the predictive signature. RESULTS: A genetic signature represented by expression levels of the three genes EHBP1, STAT1, and GAPDH was found to correlate with a pCR to neoadjuvant treatment. The difference in expression levels between patients who achieved a pCR and those who did not was greatest for EHBP1. Gene network analysis showed that the three genes can be connected by the gene ubiquitin C (UBC). CONCLUSION: This study identifies a 3-gene signature expressed in pre-treatment tumor biopsies that correlates with a pCR to neoadjuvant CRT in patients with clinical stage II and III rectal cancer. These three genes can be connected by the gene UBC, suggesting that ubiquitination is a molecular mechanism involved in determining response to treatment. Validating this genetic signature in a larger number of patients is proposed.

RESUMEN

AIMS: In the bladder, ATP is an important signaling molecule, which is released by bladder stretch and acid. We hypothesized that ATP might play a unique role in patients with OAB, characterized by low bladder volumes at first desire to void (FDV) and maximal cystometric capacity (MCC) and symptoms of frequency/urgency [mild bladder pain syndrome (BPS)]. Our aim was to investigate the correlation between ATP release and urodynamic parameters, as well as urine pH, in OAB patients. METHODS: Routine cystometry was performed in a consecutive series of 249 women. The voided urodynamic fluid (VUF) was stored at -20°C and ATP measured using bioluminescence. Catheter urine was collected for pH measurement. Correlations between two factors were tested by linear regression analysis. RESULTS: Subjects with urinary tract infection, voiding dysfunction, and detrusor overactivity (DO) were excluded. For OAB patients (n = 25), there was an inverse correlation between ATP concentration in VUF and FDV (r(2) = 0.25; P = 0.01) but not MCC. This was not seen in controls (n = 69). In OAB, but not controls, there was a significant reverse correlation (r(2) = 0.16; P = 0.047) between ATP in VUF and urine pH. Urine pH was not significantly correlated with MCC in either group. CONCLUSIONS: In OAB patients, ATP is an important factor for initial perception of need to urinate (as indicated by FDV). This is similar to our previous findings in patients with DO, suggesting that ATP may mediate initial afferent sensation in patients with bladder dysfunctions characterized by urgency. ATP release was also strongly affected by urine pH, in patients with OAB (at FDV).

Asunto(s)

Adenosina Trifosfato/orina , Vejiga Urinaria Hiperactiva/diagnóstico , Vejiga Urinaria/metabolismo , Vejiga Urinaria/fisiopatología , Urodinámica , Adulto , Anciano , Anciano de 80 o más Años , Biomarcadores/orina , Estudios de Casos y Controles , Femenino , Humanos , Concentración de Iones de Hidrógeno , Modelos Lineales , Mediciones Luminiscentes , Persona de Mediana Edad , Valor Predictivo de las Pruebas , Vejiga Urinaria Hiperactiva/fisiopatología , Vejiga Urinaria Hiperactiva/orina , Cateterismo Urinario , Micción

RESUMEN

PURPOSE: A major factor limiting the effective clinical management of colorectal cancer (CRC) is resistance to chemotherapy. Therefore, the identification of novel, therapeutically targetable mediators of resistance is vital. EXPERIMENTAL DESIGN: We used a CRC disease-focused microarray platform to transcriptionally profile chemotherapy-responsive and nonresponsive pretreatment metastatic CRC liver biopsies and in vitro samples, both sensitive and resistant to clinically relevant chemotherapeutic drugs (5-FU and oxaliplatin). Pathway and gene set enrichment analyses identified candidate genes within key pathways mediating drug resistance. Functional RNAi screening identified regulators of drug resistance. RESULTS: Mitogen-activated protein kinase signaling, focal adhesion, cell cycle, insulin signaling, and apoptosis were identified as key pathways involved in mediating drug resistance. The G-protein-coupled receptor galanin receptor 1 (GalR1) was identified as a novel regulator of drug resistance. Notably, silencing either GalR1 or its ligand galanin induced apoptosis in drug-sensitive and resistant cell lines and synergistically enhanced the effects of chemotherapy. Mechanistically, GalR1/galanin silencing resulted in downregulation of the endogenous caspase-8 inhibitor FLIP(L), resulting in induction of caspase-8-dependent apoptosis. Galanin mRNA was found to be overexpressed in colorectal tumors, and importantly, high galanin expression correlated with poor disease-free survival of patients with early-stage CRC. CONCLUSION: This study shows the power of systems biology approaches to identify key pathways and genes that are functionally involved in mediating chemotherapy resistance. Moreover, we have identified a novel role for the GalR1/galanin receptor-ligand axis in chemoresistance, providing evidence to support its further evaluation as a potential therapeutic target and biomarker in CRC.

Asunto(s)

Neoplasias Colorrectales , Resistencia a Antineoplásicos/genética , Galanina/genética , Receptor de Galanina Tipo 1/genética , Biomarcadores Farmacológicos/metabolismo , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/genética , Fluorouracilo/administración & dosificación , Galanina/metabolismo , Regulación Neoplásica de la Expresión Génica , Células HCT116 , Células HT29 , Humanos , Análisis de Secuencia por Matrices de Oligonucleótidos , Compuestos Organoplatinos/administración & dosificación , Oxaliplatino , ARN Interferente Pequeño , Receptor de Galanina Tipo 1/metabolismo , Transducción de Señal

RESUMEN

The topoisomerase I inhibitor irinotecan is used to treat advanced colorectal cancer and has been shown to have p53-independent anticancer activity. The aim of this study was to identify the p53-independent signaling mechanisms activated by irinotecan. Transcriptional profiling of isogenic HCT116 p53 wild-type and p53 null cells was carried out following treatment with the active metabolite of irinotecan, SN38. Unsupervised analysis methods showed that p53 status had a highly significant impact on gene expression changes in response to SN38. Pathway analysis indicated that pathways involved in cell motility [adherens junction, focal adhesion, mitogen-activated protein kinase (MAPK), and regulation of the actin cytoskeleton] were significantly activated in p53 null cells, but not p53 wild-type cells, following SN38 treatment. In functional assays, SN38 treatment increased the migratory potential of p53 null and p53-mutant colorectal cancer cell lines, but not p53 wild-type lines. Moreover, p53 null SN38-resistant cells were found to migrate at a faster rate than parental drug-sensitive p53 null cells, whereas p53 wild-type SN38-resistant cells failed to migrate. Notably, cotreatment with inhibitors of the MAPK pathway inhibited the increased migration observed following SN38 treatment in p53 null and p53-mutant cells. Thus, in the absence of wild-type p53, SN38 promotes migration of colorectal cancer cells, and inhibiting MAPK blocks this potentially prometastatic adaptive response to this anticancer drug.

Asunto(s)

Antineoplásicos Fitogénicos/farmacología , Camptotecina/análogos & derivados , Movimiento Celular/genética , Neoplasias Colorrectales/genética , Proteínas Quinasas Activadas por Mitógenos/antagonistas & inhibidores , Inhibidores de Topoisomerasa I/farmacología , Proteína p53 Supresora de Tumor/genética , Camptotecina/farmacología , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Neoplasias Colorrectales/metabolismo , Perfilación de la Expresión Génica , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Células HCT116 , Humanos , Irinotecán , Proteínas Quinasas Activadas por Mitógenos/metabolismo , Reproducibilidad de los Resultados , Transducción de Señal/efectos de los fármacos

RESUMEN

Chemotherapy response rates for advanced colorectal cancer remain disappointingly low, primarily because of drug resistance, so there is an urgent need to improve current treatment strategies. To identify novel determinants of resistance to the clinically relevant drugs 5-fluorouracil (5-FU) and SN38 (the active metabolite of irinotecan), transcriptional profiling experiments were carried out on pretreatment metastatic colorectal cancer biopsies and HCT116 parental and chemotherapy-resistant cell line models using a disease-specific DNA microarray. To enrich for potential chemoresistance-determining genes, an unsupervised bioinformatics approach was used, and 50 genes were selected and then functionally assessed using custom-designed short interfering RNA (siRNA) screens. In the primary siRNA screen, silencing of 21 genes sensitized HCT116 cells to either 5-FU or SN38 treatment. Three genes (RAPGEF2, PTRF, and SART1) were selected for further analysis in a panel of 5 colorectal cancer cell lines. Silencing SART1 sensitized all 5 cell lines to 5-FU treatment and 4/5 cell lines to SN38 treatment. However, silencing of RAPGEF2 or PTRF had no significant effect on 5-FU or SN38 sensitivity in the wider cell line panel. Further functional analysis of SART1 showed that its silencing induced apoptosis that was caspase-8 dependent. Furthermore, silencing of SART1 led to a downregulation of the caspase-8 inhibitor, c-FLIP, which we have previously shown is a key determinant of drug resistance in colorectal cancer. This study shows the power of systems biology approaches for identifying novel genes that regulate drug resistance and identifies SART1 as a previously unidentified regulator of c-FLIP and drug-induced activation of caspase-8.

Asunto(s)

Antígenos de Neoplasias/genética , Camptotecina/análogos & derivados , Neoplasias Colorrectales/metabolismo , Resistencia a Antineoplásicos/genética , Fluorouracilo/farmacología , Ribonucleoproteínas Nucleares Pequeñas/genética , Antígenos de Neoplasias/metabolismo , Antimetabolitos Antineoplásicos/farmacología , Antineoplásicos Fitogénicos/farmacología , Apoptosis/genética , Proteína Reguladora de Apoptosis Similar a CASP8 y FADD/metabolismo , Camptotecina/farmacología , Caspasa 8/metabolismo , Línea Celular Tumoral , Neoplasias Colorrectales/patología , Humanos , Irinotecán , Interferencia de ARN , ARN Interferente Pequeño , Ribonucleoproteínas Nucleares Pequeñas/metabolismo , Biología de Sistemas

RESUMEN

INTRODUCTION AND HYPOTHESIS: This is a prospective randomized controlled trial of cough versus no cough test in the tension-free vaginal tape (TVT) procedure to determine its effect upon voiding dysfunction and 12-month efficacy. METHODS: The trial was conducted in a single tertiary urogynecology unit. Women ≥21 years old with primary urodynamic stress incontinence without voiding dysfunction were considered eligible. Participants were randomized to undergo the TVT procedure using either an intraoperative cough test or using no intraoperative cough test. Our hypothesis was that postoperative voiding dysfunction would be more common in the "no cough test" arm. The primary outcome was proportion of patients successfully completing a trial of void (TOV) within 24 h of catheter removal. Efficacy at 12 months comprised the secondary outcome. Participants were randomized using a computer-generated randomization sequence by an independent party who was not the operating surgeon. Due to the nature of the intervention to be tested, neither the patients nor the operating surgeons were blinded to the randomization process during the procedure. RESULTS: This trial is reported according to the recommendations of the 2010 CONSORT statement. In total, 94 women were recruited over a 4-year study period. Of these, 92 women were randomized (47 in the "cough" group and 45 in the "no cough" group). In one case, the TVT procedure was abandoned intraoperatively, leaving 91 women who underwent analysis. There was no significant difference in the proportion of women with a successful TOV within 24 h between the two arms (79% in the "cough" group versus 71% in the "no cough" group; p = 0.47). Efficacy data at 12 months were not significantly different between groups. CONCLUSION: Our data suggest that the performance of the intraoperative cough test during the TVT procedure does not reduce the incidence of postoperative voiding dysfunction (as determined by successful TOV within 24 h) nor affect efficacy. The removal of the cough test from the standard TVT technique may be appropriate.

Asunto(s)

Tos , Prueba de Esfuerzo/métodos , Procedimientos Quirúrgicos Ginecológicos/métodos , Cabestrillo Suburetral , Incontinencia Urinaria de Esfuerzo/cirugía , Trastornos Urinarios/epidemiología , Adulto , Femenino , Humanos , Incidencia , Periodo Intraoperatorio , Estudios Longitudinales , Persona de Mediana Edad , Estudios Prospectivos , Factores de Tiempo , Resultado del Tratamiento

RESUMEN

The role of the calcium binding protein, Calbindin 2 (CALB2), in regulating the response of colorectal cancer (CRC) cells to 5-Fluorouracil (5-FU) was investigated. Real-time RT-PCR and Western blot analysis revealed that CALB2 mRNA and protein expression were down-regulated in p53 wild-type and p53 null isogenic HCT116 CRC cell lines following 48 h and 72 h 5-FU treatment. Moreover, 5-FU-induced apoptosis was significantly reduced in HCT116 and LS174T CRC cell lines in which CALB2 expression had been silenced. Further investigation revealed that CALB2 translocated to the mitochondria following 5-FU treatment and that 5-FU-induced loss of mitochondrial membrane potential (Δψ(m)) was abrogated in CALB2-silenced cells. Furthermore, CALB2 silencing decreased 5-FU-induced cytochrome c and smac release from the mitochondria and also decreased 5-FU-induced activation of caspases 9 and 3/7. Of note, co-silencing of XIAP overcame 5-FU resistance in CALB2-silenced cells. Collectively, these results suggest that following 5-FU treatment in CRC cell lines, CALB2 is involved in apoptosis induction through the intrinsic mitochondrial pathway. This indicates that CALB2 may be an important mediator of 5-FU-induced cell death. Moreover, down-regulation of CALB2 in response to 5-FU may represent an intrinsic mechanism of resistance to this anti-cancer drug.

Asunto(s)

Apoptosis/efectos de los fármacos , Neoplasias Colorrectales/metabolismo , Fluorouracilo/farmacología , Antineoplásicos/farmacología , Apoptosis/genética , Western Blotting , Caspasa 3/genética , Caspasa 3/metabolismo , Caspasa 7/genética , Caspasa 7/metabolismo , Caspasa 9/genética , Caspasa 9/metabolismo , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Citocromos c/genética , Citocromos c/metabolismo , Células HCT116 , Humanos , Potencial de la Membrana Mitocondrial/efectos de los fármacos , Análisis de Secuencia por Matrices de Oligonucleótidos , ARN Interferente Pequeño , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Proteína Inhibidora de la Apoptosis Ligada a X/genética , Proteína Inhibidora de la Apoptosis Ligada a X/metabolismo

RESUMEN

Over the past two decades, several protein and genomic markers have refined the prognostic information of colorectal cancer (CRC) and helped to predict which patient group may benefit most from systemic treatment or targeted therapies. Of all these markers, KRAS represents the first biomarker integrated into clinical practice for CRC. Microarray-based gene-expression profiling has been used to identify prognostic signatures and to a lesser degree predictive signatures in CRC; however, common challenges with these types of studies are clinical study design, reproducibility, interpretation and reporting of the results. We focus on the clinical application of a range of published prognostic and predictive protein and genomic markers in CRC and discuss the different challenges associated with microarray-based gene-expression profiling. While none of these genomic signatures is currently in routine clinical use in CRC, novel adaptive clinical trial designs that incorporate putative genomic prognostic/predictive markers in prospective randomized trials, will enable a clinical validation of these markers and may facilitate the implementation of these biomarkers into routine medical practice.

Asunto(s)

Antineoplásicos/uso terapéutico , Biomarcadores de Tumor/genética , Neoplasias Colorrectales/tratamiento farmacológico , Resistencia a Antineoplásicos/genética , Ensayos Clínicos como Asunto , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/mortalidad , Humanos , Pronóstico , Proyectos de Investigación

RESUMEN

BACKGROUND: To date, there are no clinically reliable predictive markers of response to the current treatment regimens for advanced colorectal cancer. The aim of the current study was to compare and assess the power of transcriptional profiling using a generic microarray and a disease-specific transcriptome-based microarray. We also examined the biological and clinical relevance of the disease-specific transcriptome. METHODS: DNA microarray profiling was carried out on isogenic sensitive and 5-FU-resistant HCT116 colorectal cancer cell lines using the Affymetrix HG-U133 Plus2.0 array and the Almac Diagnostics Colorectal cancer disease specific Research tool. In addition, DNA microarray profiling was also carried out on pre-treatment metastatic colorectal cancer biopsies using the colorectal cancer disease specific Research tool. The two microarray platforms were compared based on detection of probesets and biological information. RESULTS: The results demonstrated that the disease-specific transcriptome-based microarray was able to out-perform the generic genomic-based microarray on a number of levels including detection of transcripts and pathway analysis. In addition, the disease-specific microarray contains a high percentage of antisense transcripts and further analysis demonstrated that a number of these exist in sense:antisense pairs. Comparison between cell line models and metastatic CRC patient biopsies further demonstrated that a number of the identified sense:antisense pairs were also detected in CRC patient biopsies, suggesting potential clinical relevance. CONCLUSIONS: Analysis from our in vitro and clinical experiments has demonstrated that many transcripts exist in sense:antisense pairs including IGF2BP2, which may have a direct regulatory function in the context of colorectal cancer. While the functional relevance of the antisense transcripts has been established by many studies, their functional role is currently unclear; however, the numbers that have been detected by the disease-specific microarray would suggest that they may be important regulatory transcripts. This study has demonstrated the power of a disease-specific transcriptome-based approach and highlighted the potential novel biologically and clinically relevant information that is gained when using such a methodology.

Asunto(s)

Neoplasias Colorrectales/genética , Resistencia a Antineoplásicos/genética , Perfilación de la Expresión Génica/métodos , Regulación Neoplásica de la Expresión Génica , Análisis de Secuencia por Matrices de Oligonucleótidos , Antimetabolitos Antineoplásicos/farmacología , Antimetabolitos Antineoplásicos/uso terapéutico , Biopsia , Supervivencia Celular/efectos de los fármacos , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/patología , Fluorouracilo/farmacología , Genotipo , Células HCT116 , Humanos , Proteína 2 de Unión a Factor de Crecimiento Similar a la Insulina/genética , Fenotipo , Pronóstico , ARN sin Sentido/metabolismo , Reproducibilidad de los Resultados , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa