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Tumour Necrosis Factor (TNF) potently induces a transient inflammatory response that must be downregulated once any invasive stimulus has resolved. Yet, how TNF-induced inflammation is shut down in normal cells is incompletely understood. The present study shows that STAT3 was activated in mouse embryo fibroblasts (MEFs) by treatment with TNF or an agonist antibody to TNFR1. STAT3 activation was inhibited by pharmacological inhibition of the Jak2 tyrosine kinase that associates with TNFR1. To identify STAT3 target genes, global transcriptome analysis by RNA sequencing was performed in wild-type MEFs and MEFs from STAT3 knockout (STAT3KO ) mice that were stimulated with TNF, and the results were validated at the protein level by using multiplex cytokine assays and immunoblotting. After TNF stimulation, STAT3KO MEFs showed greater gene and protein induction of the inflammatory chemokines Ccl2, Cxcl1 and Cxcl10 than WT MEFs. These observations show that, by activating STAT3, TNF selectively modulates expression of a cohort of chemokines that promote inflammation. The greater induction by TNF of chemokines in STAT3KO than WT MEFs suggested that TNF induced an inhibitory protein in WT MEFs. Consistent with this possibility, STAT3 activation by TNFR1 increased the expression of Tnfaip3/A20, a ubiquitin modifying enzyme that inhibits inflammation, in WT MEFs but not in STAT3KO MEFs. Moreover, enforced expression of Tnfaip3/A20 in STAT3KO MEFs suppressed proinflammatory chemokine expression induced by TNF. Our observations identify Tnfaip3/A20 as a new downstream target for STAT3 which limits the induction of Ccl2, Cxcl1 and Cxcl10 and inflammation induced by TNF.
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Receptores Tipo I de Factores de Necrosis Tumoral , Factor de Necrosis Tumoral alfa , Animales , Expresión Génica , Inflamación , Janus Quinasa 2/metabolismo , Ratones , Ratones Noqueados , FN-kappa B/metabolismo , Receptores Tipo I de Factores de Necrosis Tumoral/genética , Factor de Transcripción STAT3/metabolismo , Proteína 3 Inducida por el Factor de Necrosis Tumoral alfa/genética , Proteína 3 Inducida por el Factor de Necrosis Tumoral alfa/metabolismo , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Advances in single-cell technologies have highlighted the prevalence and biological significance of cellular heterogeneity. A critical question researchers face is how to design experiments that faithfully capture the true range of heterogeneity from samples of cellular populations. Here we develop a data-driven approach, illustrated in the context of image data, that estimates the sampling depth required for prospective investigations of single-cell heterogeneity from an existing collection of samples.
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Adenocarcinoma/patología , Carcinoma de Pulmón de Células no Pequeñas/patología , Neoplasias Pulmonares/patología , Análisis de la Célula Individual/métodos , Biomarcadores de Tumor , Técnicas de Cultivo de Célula , Línea Celular , Regulación Neoplásica de la Expresión Génica , HumanosRESUMEN
BACKGROUND AND OBJECTIVES: We developed objective measurements of preoperative and residual tumor volume, and debulking rate, to evaluate their prognostic value for neuroendocrine liver metastasis (NELM). METHODS: Seventy-three patients who underwent surgery for NELM were analyzed retrospectively. Indices of preoperative and postoperative residual tumor volume (pre-volume index [VI] and post-VI) were calculated as the sum of the cubes of individual tumor diameters on preoperative and postoperative imaging, respectively. The debulking rate (%) was calculated as 100 - 100 × post-VI/pre-VI. The classification and regression trees method was used to classify pre-VI and post-VI. RESULTS: Overall survival (OS) was discriminated by preoperative tumor volume (5-year OS rates, 87.8% for low pre-VI and 60.1% for high pre-VI; P = .037) and residual tumor volume (5-year OS rates, 88.1% for low post-VI and 24.8% for high post-VI; P < .001). In contrast, debulking rates of 100%, ≥90%, and <90% did not discriminate OS (5-year OS rates, 88.0%, 61.9%, and 58.9%, respectively, not significant). In multivariate analysis, residual tumor volume (high post-VI, hazard ratio, 6.40; 95% confidence interval, 1.45-32.3) was an independent prognostic factor for OS. CONCLUSIONS: Objective measurement of tumor volume demonstrates that residual tumor volume is prognostic after surgery for NELM.
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BACKGROUND AND OBJECTIVES: We classified the extent of mesenteric mass (MM) involvement that predicts challenging mesenteric lymph node dissection (mLND) by minimally invasive surgery (MIS) for ileal neuroendocrine tumors (i-NETs). METHODS: Patients who underwent surgery for i-NETs were retrospectively reviewed. MM involvement was classified as region-0: no MM; region-1: >2 cm from the origins of the ileocolic artery/vein; region-2: ≤2 cm from the origins; and region-3: more proximal superior mesenteric artery/vein. Logistic regression analysis was used to evaluate the predictive value of MM regions for gross positive mesenteric margin (mR2) and/or conversion among the MIS cohort. The open surgery cohort was used as a reference for mR2 rates. RESULTS: Of 108 patients, 83 patients (77%) underwent MIS. MMs in region-2 and region-3 were independent risk factors for mR2 and/or conversion (odds ratio [95% confidence interval]: 4.25 [1.17-16.4] and 8.51 × 107 [11.0-], respectively, against regions-0 and 1]. mR2 rates of MIS and open surgery cohorts per region did not differ significantly (4% and 7% for regions-0 and 1; 17% and 25% for region-2; and 100% and 83% for region-3). CONCLUSIONS: The novel stratification of MM regions was predictive of challenging mLND by MIS. Surgeons should have a low threshold for conversion for MMs in proximal regions.
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Neoplasias del Íleon/patología , Neoplasias del Íleon/cirugía , Ganglios Linfáticos/patología , Ganglios Linfáticos/cirugía , Mesenterio/patología , Tumores Neuroendocrinos/patología , Tumores Neuroendocrinos/cirugía , Estudios de Cohortes , Humanos , Escisión del Ganglio Linfático , Masculino , Márgenes de Escisión , Persona de Mediana Edad , Procedimientos Quirúrgicos Mínimamente Invasivos/métodos , Valor Predictivo de las Pruebas , Estudios RetrospectivosRESUMEN
BACKGROUND AND OBJECTIVES: Ileal neuroendocrine tumors (i-NETs) frequently metastasize to mesenteric lymph nodes and the liver. Regional lymphadenopathy is associated with desmoplasia of the mesentery forming a large mesenteric mass (LMM). Although the latest American Joint Committee on Cancer TNM staging (8th edition) defined LMM >2 cm as N2, the prognostic impact of LMM is ill-defined. We evaluated whether LMM is prognostic for patients with i-NETs. METHODS: This single-institution, retrospective cohort study included 106 patients who underwent resection of i-NETs between 2007 and 2018. Overall survival (OS) and liver progression-free survival (LPFS) were compared between patients with and without LMM. RESULTS: LMM was present in 66 patients (62%) and was not associated with the presence or absence of liver metastasis (P = .969) or the extent of liver involvement (P = .938). OS and LPFS differed significantly between patients with and without LMM (5-year OS rates of 64.8% and 92.9%, respectively, P = .011; 3-year LPFS rates of 45.3% and 67.5%, respectively, P = .025). In multivariate analysis, LMM was an independent prognostic factor for both OS (hazard ratio: 4.69, 95% confidence interval: 1.63-17.6) and LPFS (1.99, 1.08-3.88). CONCLUSION: LMM >2 cm is prognostic for OS and LPFS and represents aggressive tumor biology.
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Neoplasias del Íleon/mortalidad , Neoplasias del Íleon/patología , Ganglios Linfáticos/patología , Mesenterio/patología , Tumores Neuroendocrinos/mortalidad , Tumores Neuroendocrinos/patología , Anciano , Estudios de Cohortes , Femenino , Humanos , Neoplasias del Íleon/cirugía , Neoplasias Hepáticas/secundario , Neoplasias Hepáticas/cirugía , Metástasis Linfática , Masculino , Persona de Mediana Edad , Tumores Neuroendocrinos/cirugía , Pronóstico , Supervivencia sin Progresión , Estudios RetrospectivosRESUMEN
PURPOSE: Tumor levels of thymidylate synthase (TS), a target of 5-fluorouracil (5-FU)-based chemotherapy for colorectal cancer, have been studied as a predictive or prognostic biomarker with mixed results. PATIENTS AND METHODS: Tumor TS levels were prospectively evaluated in two adjuvant therapy trials for patients with resected stage II or III colon cancer. TS expression was determined by standard immunohistochemistry and by automated quantitative analysis. Tumor mismatch repair deficiency (MMR-D) and BRAF c.1799T > A (p.V600E) mutation status were also examined. Relationships between tumor TS, MMR-D, and BRAF mutation status, overall survival (OS), and disease-free survival (DFS) were investigated in the subset of stage III patients. RESULTS: Patients whose tumors demonstrated high TS expression experienced better treatment outcomes, with DFS hazard ratio (HR) = 0.67, 95% confidence interval (CI) = 0.53, 0.84; and OS HR = 0.68, 95% CI = 0.53, 0.88, for high versus low TS expression, respectively. No significant interaction between TS expression and stage was observed (DFS: interaction HR = 0.94; OS: interaction HR = 0.94). Tumors with high TS expression were more likely to demonstrate MMR-D (22.2% vs. 12.8%; p = .0003). Patients whose tumors demonstrated both high TS and MMR-D had a 7-year DFS of 77%, compared with 58% for those whose tumors had low TS and were non-MMR-D (log-rank p = .0006). Tumor TS expression did not predict benefit of a particular therapeutic regimen. CONCLUSION: This large prospective analysis showed that high tumor TS levels were associated with improved DFS and OS following adjuvant therapy for colon cancer, although tumor TS expression did not predict benefit of 5-FU-based chemotherapy. The Oncologist 2017;22:107-114Implications for Practice: This study finds that measurement of tumor levels of thymidylate synthase is not helpful in assigning specific adjuvant treatment for colorectal cancer. It also highlights the importance of using prospective analyses within treatment clinical trials as the optimal method of determining biomarker utility.
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Biomarcadores de Tumor/genética , Neoplasias del Colon/tratamiento farmacológico , Neoplasias del Colon/genética , Timidilato Sintasa/genética , Anciano , Biomarcadores de Tumor/biosíntesis , Neoplasias del Colon/patología , Neoplasias del Colon/cirugía , Terapia Combinada , Supervivencia sin Enfermedad , Femenino , Fluorouracilo/administración & dosificación , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , Inestabilidad de Microsatélites , Persona de Mediana Edad , Estadificación de Neoplasias , Pronóstico , Timidilato Sintasa/antagonistas & inhibidores , Resultado del TratamientoRESUMEN
Proteases responsible for the increased peritumoral proteolysis associated with cancer represent functional biomarkers for monitoring tumorigenesis. One attractive extracellular biomarker is the transmembrane serine protease matriptase. Found on the surface of epithelial cells, the activity of matriptase is regulated by its cognate inhibitor hepatocyte growth factor activator inhibitor-1 (HAI-1). Quantitative mass spectrometry allowed us to show that, in selected cancers, HAI-1 expression decreases, leading to active matriptase. A preclinical probe specific for the measurement of emergent active matriptase was developed. Using an active-site-specific, recombinant human antibody for matriptase, we found that the selective targeting of active matriptase can be used to visualize the tumorigenic epithelium. Live-cell fluorescence imaging validated the selectivity of the antibody in vitro by showing that the probe localized only to cancer cell lines with active matriptase on the surface. Immunofluorescence with the antibody documented significant levels of active matriptase in 68% of primary and metastatic colon cancer sections from tissue microarrays. Labeling of the active form of matriptase in vivo was measured in human colon cancer xenografts and in a patient-derived xenograft model using near-infrared and single-photon emission computed tomography imaging. Tumor uptake of the radiolabeled antibody, (111)In-A11, by active matriptase was high in xenografts (28% injected dose per gram) and was blocked in vivo by the addition of a matriptase-specific variant of ecotin. These findings suggest, through a HAI-1-dependent mechanism, that emergent active matriptase is a functional biomarker of the transformed epithelium and that its proteolytic activity can be exploited to noninvasively evaluate tumorigenesis in vivo.
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Biomarcadores de Tumor/metabolismo , Transformación Celular Neoplásica/metabolismo , Neoplasias del Colon/metabolismo , Células Epiteliales/metabolismo , Imagen Óptica/métodos , Proteínas Inhibidoras de Proteinasas Secretoras/metabolismo , Serina Endopeptidasas/metabolismo , Anticuerpos/inmunología , Humanos , Radioisótopos de Indio/metabolismo , Espectrometría de Masas , Serina Endopeptidasas/inmunologíaRESUMEN
BACKGROUND & AIMS: The CpG island methylator phenotype (CIMP), defined by a high frequency of aberrantly methylated genes, is a characteristic of a subclass of colon tumors with distinct clinical and molecular features. Cohort studies have produced conflicting results on responses of CIMP-positive tumors to chemotherapy. We assessed the association between tumor CIMP status and survival of patients receiving adjuvant fluorouracil and leucovorin alone or with irinotecan (IFL). METHODS: We analyzed data from patients with stage III colon adenocarcinoma randomly assigned to groups given fluorouracil and leucovorin or IFL after surgery, from April 1999 through April 2001. The primary end point of the trial was overall survival and the secondary end point was disease-free survival. DNA isolated from available tumor samples (n = 615) was used to determine CIMP status based on methylation patterns at the CACNA1G, IGF2, NEUROG1, RUNX3, and SOCS1 loci. The effects of CIMP on survival were modeled using Kaplan-Meier and Cox proportional hazards; interactions with treatment and BRAF, KRAS, and mismatch repair (MMR) status were also investigated. RESULTS: Of the tumor samples characterized for CIMP status, 145 were CIMP positive (23%). Patients with CIMP-positive tumors had shorter overall survival times than patients with CIMP-negative tumors (hazard ratio = 1.36; 95% confidence interval: 1.01-1.84). Treatment with IFL showed a trend toward increased overall survival for patients with CIMP-positive tumors, compared with treatment with fluorouracil and leucovorin (hazard ratio = 0.62; 95% CI: 0.37-1.05; P = .07), but not for patients with CIMP-negative tumors (hazard ratio = 1.38; 95% CI: 1.00-1.89; P = .049). In a 3-way interaction analysis, patients with CIMP-positive, MMR-intact tumors benefited most from the addition of irinotecan to fluorouracil and leucovorin therapy (for the interaction, P = .01). CIMP was more strongly associated with response to IFL than MMR status. Results for disease-free survival times were comparable among all analyses. CONCLUSIONS: Patients with stage III, CIMP-positive, MMR-intact colon tumors have longer survival times when irinotecan is added to combination therapy with fluorouracil and leucovorin.
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Adenocarcinoma/tratamiento farmacológico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Neoplasias del Colon/tratamiento farmacológico , Islas de CpG , Metilación de ADN , Adenocarcinoma/genética , Adenocarcinoma/mortalidad , Adenocarcinoma/patología , Adulto , Anciano , Anciano de 80 o más Años , Camptotecina/administración & dosificación , Camptotecina/análogos & derivados , Quimioterapia Adyuvante , Colectomía , Neoplasias del Colon/genética , Neoplasias del Colon/mortalidad , Neoplasias del Colon/patología , Reparación de la Incompatibilidad de ADN , Supervivencia sin Enfermedad , Femenino , Fluorouracilo/administración & dosificación , Humanos , Irinotecán , Estimación de Kaplan-Meier , Leucovorina/administración & dosificación , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Fenotipo , Modelos de Riesgos Proporcionales , Medición de Riesgo , Factores de Riesgo , Factores de Tiempo , Resultado del Tratamiento , Adulto JovenRESUMEN
Tumors rich in stroma are associated with advanced stage and poor prognosis in colorectal adenocarcinoma (CRC). Abundance of stromal cells also has implications for genomic analysis of patient tumors as it may prevent detection of somatic mutations. As part of our efforts to interrogate stroma-cancer cell interactions and to identify actionable therapeutic targets in metastatic CRC, we aimed to determine the proportion of stroma embedded in hepatic CRC metastases by performing computational tumor purity analysis based on whole exome sequencing data (WES). Unlike previous studies focusing on histopathologically prescreened samples, we used an unbiased in-house collection of tumor specimens. WES from CRC liver metastasis samples were utilized to evaluate stromal content and to assess the performance of three in silico tumor purity tools, ABSOLUTE, Sequenza and PureCN. Matching tumor derived organoids were analyzed as a high purity control as they are enriched in cancer cells. Computational purity estimates were compared to those from a histopathological assessment conducted by a board-certified pathologist. According to all computational methods, metastatic specimens had a median tumor purity of 30% whereas the organoids were enriched for cancer cells with a median purity estimate of 94%. In line with this, variant allele frequencies (VAFs) of oncogenes and tumor suppressor genes were undetectable or low in most patient tumors, but higher in matching organoid cultures. Positive correlation was observed between VAFs and in silico tumor purity estimates. Sequenza and PureCN produced concordant results whereas ABSOLUTE yielded lower purity estimates for all samples. Our data shows that unbiased sample selection combined with molecular, computational, and histopathological tumor purity assessment is critical to determine the level of stroma embedded in metastatic colorectal adenocarcinoma.
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Adenocarcinoma , Neoplasias Colorrectales , Neoplasias Hepáticas , Humanos , Secuenciación del Exoma , Mutación , Exoma/genética , Neoplasias Colorrectales/genética , Neoplasias Colorrectales/patología , Adenocarcinoma/genética , Neoplasias Hepáticas/genéticaRESUMEN
ATP-competitive mammalian target of rapamycin (mTOR) inhibitors are in early phase clinical trials. These novel targeted agents, including PP242, are mechanistically distinct from the allosteric, partial mTOR inhibitor, rapamycin. The goal of this study was to evaluate how PP242 best combines with standard chemotherapies for colorectal cancer (CRC), and which subsets of patients are most likely to benefit. The combination index for PP242 plus 5-fluorouracil, oxaliplatin, or irinotecan was determined in CRC cell lines with different mutational backgrounds. In KRAS mutant CRC cell lines, sensitivity to PP242 increases with co-mutation of PIK3CA. Mutation of p53 predicts resistance to chemotherapy, but not PP242. Efficacy of PP242 was comparable to that of standard chemotherapies over the dose range tested. Sensitivity or resistance to PP242 dictates relative synergy or antagonism, respectively, when PP242 is combined with 5-fluorouracil. The same trend exists for PP242 + oxaliplatin, but with a narrower dynamic range. Conversely potency of PP242 and the combination index for PP242 + irinotecan were unrelated, but synergy exists across all dose levels in PP242 and irinotecan sensitive, p53 wild-type cell lines. Overall, our in vitro analysis predicts that mutational status can be used to rank sensitivity to PP242 and standard chemotherapies. Single agent potency can in turn be used to predict the combination index in a drug-specific manner. Our data suggest a clinical trial to determine whether ATP-competitive mTOR inhibitors provide benefit in combination with standard chemotherapies for patients with PIK3CA mutant metastatic CRC, stratified by the presence or absence of KRAS co-mutation.
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Antineoplásicos/administración & dosificación , Camptotecina/análogos & derivados , Fluorouracilo/administración & dosificación , Indoles/administración & dosificación , Compuestos Organoplatinos/administración & dosificación , Purinas/administración & dosificación , Serina-Treonina Quinasas TOR/antagonistas & inhibidores , Adenosina Trifosfato , Camptotecina/administración & dosificación , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Neoplasias Colorrectales/tratamiento farmacológico , Neoplasias Colorrectales/genética , Combinación de Medicamentos , Humanos , Irinotecán , Mutación , Proteínas Nucleares/genética , Oxaliplatino , Proteínas Proto-Oncogénicas/genética , Proteínas Proto-Oncogénicas p21(ras) , Factores de Transcripción/genética , Proteínas ras/genéticaRESUMEN
BACKGROUND: Retroperitoneal tumors are often massive and can involve adjacent organs and/or vital structures, making them difficult to resect. Completeness of resection is within the surgeon's control and critical for long-term survival, particularly for malignant disease. Few studies directly address strategies for complete and safe resection of challenging retroperitoneal tumors. METHODS: Fifty-six patients representing 63 cases of primary or recurrent retroperitoneal tumor resection between 2004-2009 were identified and a retrospective chart review was performed. Rates of complete resection, use of adjunct procedures, and perioperative complications were recorded. RESULTS: In 95% of cases, complete resection was achieved. Fifty-eight percent of these cases required en bloc multi-organ resection, and 8% required major vascular resection. Complete resection rates were higher for primary versus recurrent disease. Adjunct procedures (ureteral stents, femoral nerve monitoring, posterior laminotomy, etc.) were used in 54% of cases. Major postoperative complications occurred in 16% of cases, and one patient died (2% mortality). CONCLUSIONS: Complete resection of challenging retroperitoneal tumors is feasible and can be done safely with important pre- and intraoperative considerations in mind.
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Recurrencia Local de Neoplasia/irrigación sanguínea , Recurrencia Local de Neoplasia/cirugía , Neoplasias Retroperitoneales/irrigación sanguínea , Neoplasias Retroperitoneales/cirugía , Espacio Retroperitoneal/irrigación sanguínea , Espacio Retroperitoneal/cirugía , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Humanos , Masculino , Persona de Mediana Edad , Recurrencia Local de Neoplasia/patología , Complicaciones Posoperatorias , Neoplasias Retroperitoneales/patología , Espacio Retroperitoneal/patología , Estudios Retrospectivos , Resultado del Tratamiento , Adulto JovenRESUMEN
BACKGROUND: Patients with neuroendocrine tumors (NETs) may have metastatic disease and unknown primary site. NETs commonly arise from the bronchopulmonary (BP) and gastrointestinal (GI) tract. The largest subgroups of well-differentiated BP-NETs are typical carcinoids (TCs). The homeodomain transcription factor NKX2.2 regulates development of gut serotonin cells and is a marker of GI-NETs. Previous work on a limited number of samples suggested that BP-TCs do not express NKX2.2. We hypothesized that lack of NKX2.2 expression in BP-TCs might be useful to distinguish BP- from GI-NETs, and evaluated NKX2.2 expression in a larger number of BP-TCs. METHODS: Archived formalin-fixed, paraffin-embedded tissues were obtained from 13 previously undescribed patients with BP-TCs. Expression of NKX2.2, serotonin, and the NE marker chromogranin A (CgA) were assessed by immunohistochemistry. RESULTS: CgA expression was robust in all 13 BP-TCs, confirming the NE phenotype. Serotonin expression was less frequent (9/13; 69%). Two patients with BP-TCs in which serotonin expression was absent exhibited Cushing's syndrome due to ectopic ACTH production. NKX2.2 expression was not observed in any of the 13 tumors. CONCLUSIONS: Bronchopulmonary TCs uniformly express CgA but not NKX2.2. Because most of these tumors express serotonin, our findings suggest that NKX2.2 may not be required for serotonin production by BP-TCs. We conclude that the presence or absence of NKX2.2 expression may assist in the determination of the primary tumor site in patients with NET metastases of unknown origin. NET metastases that are CgA-positive/NKX2.2-negative would suggest a BP primary, whereas those that are CgA-positive/NKX2.2-positive would suggest a GI primary.
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Biomarcadores de Tumor/metabolismo , Tumor Carcinoide/metabolismo , Neoplasias Gastrointestinales/metabolismo , Proteínas de Homeodominio/metabolismo , Neoplasias Pulmonares/metabolismo , Factores de Transcripción/metabolismo , Adolescente , Adulto , Anciano , Tumor Carcinoide/diagnóstico , Femenino , Neoplasias Gastrointestinales/diagnóstico , Proteína Homeobox Nkx-2.2 , Humanos , Inmunohistoquímica , Neoplasias Pulmonares/diagnóstico , Masculino , Persona de Mediana Edad , Proteínas Nucleares , Adulto Joven , Proteínas de Pez CebraRESUMEN
BACKGROUND: In retrospective studies, loss of p27(Kip1) (p27), a cyclin-dependent kinase inhibitor, has been associated with poor prognosis following colorectal cancer treatment. In a prospective study, we validated this relationship in patients enrolled on a trial of adjuvant chemotherapy for stage III colon cancer. METHODS: Cancer and Leukemia Group B protocol 89803 randomized 1,264 stage III colon cancer patients to receive weekly bolus 5-fluorouracil/leucovorin or weekly bolus irinotecan, 5-fluorouracil, and leucovorin (IFL). The primary endpoint was overall survival (OS); disease-free survival was a secondary endpoint. Expression of p27 and DNA mismatch repair proteins were determined by immunohistochemistry in primary tumor and normal tissue from paraffin blocks. Data were analyzed using log-rank test. RESULTS: Of 601 tumors analyzed, 207 (34.4%) showed p27 loss, 377 (62.8%) retained p27, and 17 (2.8%) were indeterminate. Patients with p27-negative tumors showed reduced OS [5-year OS 66%: 95% confidence interval (95% CI), 0.59-0.72 versus 75%: 95% CI, 0.70-0.79; log-rank P = 0.021]. This relationship was not influenced by treatment arm. Combination of p27 status with mismatch repair status, however, identified a small subset of patients that may benefit from IFL (n = 36; 5-year disease-free survival 81%: 95% CI, 0.64-0.98 versus 47%: 95% CI, 0.21-0.72; log-rank P = 0.042; 5-year OS 81%: 95% CI, 0.64-0.98 versus 60%: 95% CI, 0.35-0.85; log-rank P = 0.128). CONCLUSIONS: Loss of p27 is associated with reduced survival in stage III colon cancer but by itself does not indicate a significant difference in outcome between patients treated IFL or 5-fluorouracil/leucovorin.
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Neoplasias del Colon/tratamiento farmacológico , Péptidos y Proteínas de Señalización Intracelular/análisis , Adulto , Anciano , Anciano de 80 o más Años , Quimioterapia Adyuvante , Neoplasias del Colon/química , Neoplasias del Colon/mortalidad , Neoplasias del Colon/patología , Inhibidor p27 de las Quinasas Dependientes de la Ciclina , Reparación de la Incompatibilidad de ADN , Femenino , Humanos , Péptidos y Proteínas de Señalización Intracelular/fisiología , Masculino , Inestabilidad de Microsatélites , Persona de Mediana Edad , Estadificación de Neoplasias , Estudios Prospectivos , Resultado del TratamientoRESUMEN
Identifying master regulators that drive pathologic gene expression is a key challenge in precision oncology. Here, we have developed an analytic framework, named PRADA, that identifies oncogenic RNA-binding proteins through the systematic detection of coordinated changes in their target regulons. Application of this approach to data collected from clinical samples, patient-derived xenografts, and cell line models of colon cancer metastasis revealed the RNA-binding protein RBMS1 as a suppressor of colon cancer progression. We observed that silencing RBMS1 results in increased metastatic capacity in xenograft mouse models, and that restoring its expression blunts metastatic liver colonization. We have found that RBMS1 functions as a posttranscriptional regulator of RNA stability by directly binding its target mRNAs. Together, our findings establish a role for RBMS1 as a previously unknown regulator of RNA stability and as a suppressor of colon cancer metastasis with clinical utility for risk stratification of patients. SIGNIFICANCE: By applying a new analytic approach to transcriptomic data from clinical samples and models of colon cancer progression, we have identified RBMS1 as a suppressor of metastasis and as a post-transcriptional regulator of RNA stability. Notably, RBMS1 silencing and downregulation of its targets are negatively associated with patient survival.See related commentary by Carter, p. 1261.This article is highlighted in the In This Issue feature, p. 1241.
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Neoplasias del Colon/patología , Proteínas de Unión al ADN/metabolismo , Regulación Neoplásica de la Expresión Génica , Neoplasias Hepáticas/genética , ARN Mensajero/genética , Proteínas de Unión al ARN/metabolismo , Animales , Línea Celular Tumoral , Colon/patología , Neoplasias del Colon/diagnóstico , Neoplasias del Colon/genética , Proteínas de Unión al ADN/genética , Técnicas de Silenciamiento del Gen , Humanos , Mucosa Intestinal/patología , Neoplasias Hepáticas/secundario , Masculino , Ratones , Estadificación de Neoplasias , Estabilidad del ARN/genética , Proteínas de Unión al ARN/genética , RNA-Seq , Regulón , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
The homeodomain transcription factor NKX2.2 is necessary for neuroendocrine (NE) differentiation in the central nervous system and pancreas. NE tumors derived from the gut are defined by their NE phenotype, which is used for diagnosis and contributes to tumorigenicity. We hypothesized that NKX2.2 is important for NE differentiation in normal and neoplastic gut. NKX2.2 and NE marker expression was investigated in the small intestine of embryonic and adult mice using immunofluorescence (IF). To determine the role of NKX2.2 in NE differentiation of the intestine, the phenotype of Nkx2.2 (-/-) mice was examined by IF and real-time (RT)-PCR. NKX2.2 and NE marker expression in human NE tumors of the gut and normal tissues were evaluated by immunohistochemistry and qRT-PCR. NKX2.2 expression was detected in the intervillus/crypt regions of embryonic and adult mouse intestine. Co-expression of Nkx2.2 with neurogenin3 (NEUROG3) and hormones was observed in the adult intestinal crypt compartment, suggesting NKX2.2 functions in NEUROG3-positive endocrine progenitors and newly differentiated endocrine cells. In the intestine of Nkx2.2 (-/-) mice, we found a dramatic reduction in the number of cells producing numerous hormones, such as serotonin, gastrin, cholecystokinin, somatostatin, glucagon-like peptide 1 (GLP-1), and secretin, but an increase in cells producing ghrelin. NKX2.2 was expressed in most (24 of 29) human NE tumors derived from diverse primary sites. We conclude NKX2.2 functions in immature endocrine cells to control NE differentiation in normal intestine and is expressed in most NE tumors of the gut, and is therefore a novel target of diagnosis for patients with gastrointestinal NE tumors.
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Neoplasias Gastrointestinales/genética , Regulación del Desarrollo de la Expresión Génica , Regulación Neoplásica de la Expresión Génica , Proteínas de Homeodominio/genética , Proteínas de Homeodominio/metabolismo , Tumores Neuroendocrinos/genética , Factores de Transcripción/genética , Factores de Transcripción/metabolismo , Animales , Apoptosis/fisiología , Diferenciación Celular/fisiología , División Celular/fisiología , Células Endocrinas/citología , Células Endocrinas/fisiología , Neoplasias Gastrointestinales/metabolismo , Neoplasias Gastrointestinales/patología , Ghrelina/metabolismo , Proteína Homeobox Nkx-2.2 , Humanos , Mucosa Intestinal/citología , Mucosa Intestinal/embriología , Intestino Delgado/citología , Intestino Delgado/embriología , Ratones , Ratones Mutantes , Tumores Neuroendocrinos/metabolismo , Tumores Neuroendocrinos/patología , Proteínas Nucleares , Proteínas de Pez CebraRESUMEN
Hdm2 is elevated in numerous types of malignancies and is thought to impede the function of wild-type p53. Reactivation of p53 by disrupting the association with Hdm2 was the impetus for the development of Nutlin3. Although regulation of p53 has been the central focus of Hdm2 activity, it also binds other proteins through its p53-binding domain. Here, we show that hypoxia-inducible factor 1alpha (HIF1alpha) binds to Hdm2 in the domain designated to bind p53. HIF1alpha and p53 share a conserved motif that is required to bind Hdm2. Distinct complexes form between Hdm2-HIF1alpha and Hdm2-p53 as determined by immunoprecipitation of nuclear extracts and in vitro. The Hdm2 antagonist Nutlin3 prevents the association between Hdm2 and HIF1alpha. The vascular endothelial growth factor (VEGF) gene is a transcriptional target of HIF1alpha, and under normoxic or hypoxic conditions, Hdm2 increases HIF1alpha activity to induce VEGF production. Blocking the association of Hdm2 and HIF1alpha by Nutlin3, or ablating Hdm2 expression, diminished the level of VEGF under conditions of normoxia or hypoxia. Our findings establish a unique role for Nutlin3 in attenuating VEGF induction by preventing the association of Hdm2 with HIF1alpha.
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Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Imidazoles/farmacología , Piperazinas/farmacología , Proteínas Proto-Oncogénicas c-mdm2/metabolismo , Factor A de Crecimiento Endotelial Vascular/biosíntesis , Secuencia de Aminoácidos , Animales , Línea Celular Tumoral , Secuencia Conservada , Humanos , Subunidad alfa del Factor 1 Inducible por Hipoxia/antagonistas & inhibidores , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Unión Proteica , Estructura Terciaria de Proteína , Proteínas Proto-Oncogénicas c-mdm2/antagonistas & inhibidores , Proteínas Proto-Oncogénicas c-mdm2/genética , ARN Interferente Pequeño/genética , Conejos , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/genética , Proteínas Recombinantes/farmacología , Alineación de Secuencia , Transfección , Proteína p53 Supresora de Tumor/metabolismo , Factor A de Crecimiento Endotelial Vascular/antagonistas & inhibidores , Factor A de Crecimiento Endotelial Vascular/genéticaRESUMEN
PURPOSE: Prognostic schemes that rely on clinical variables to predict outcome after resection of colorectal metastases remain imperfect. We hypothesized that molecular markers can improve the accuracy of prognostic schemes. METHODS: We screened the transcriptome of matched colorectal liver metastases (CRCLM) and primary tumors from 42 patients with unresected CRCLM to identify differentially expressed genes. Among the differentially expressed genes identified, we looked for associations between expression and time to disease progression or overall survival. To validate such associations, mRNA levels of the candidate genes were assayed by qRT-PCR from CRCLM in 56 additional patients who underwent hepatectomy. RESULTS: Seven candidate genes were selected for validation based on their differential expression between metastases and primary tumors and a correlation between expression and surgical outcome: lumican; tissue inhibitor metalloproteinase 1; basic helix-loop-helix domain containing class B2; fibronectin; transmembrane 4 superfamily member 1; mitogen inducible gene 6 (MIG-6); and serpine 2. In the hepatectomy group, only MIG-6 expression was predictive of poor survival after hepatectomy. Quantitative PCR of MIG-6 mRNA was performed on 25 additional hepatectomy patients to determine if MIG-6 expression could substratify patients beyond the clinical risk score. Patients within defined clinical risk score categories were effectively substratified into distinct groups by relative MIG-6 expression. CONCLUSIONS: MIG-6 expression is inversely associated with survival after hepatectomy and may be used to improve traditional prognostic schemes that rely on clinicopathologic data such as the Clinical Risk Score.
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INTRODUCTION: Prognosis after resection of hepatocellular carcinoma (HCC) is highly variable. Compared to clinicopathologic factors, the use of molecular markers to predict outcome has not been well studied. We investigated the prognostic importance of thymidylate synthase (TS) gene expression and polymorphisms in patients after resection of HCC. METHODS: Patients who underwent complete resection of HCC for whom tissue was available were identified. TS gene expression level and polymorphisms were determined in HCC specimens. Prognostic factors were evaluated using Kaplan-Meier curves and Cox proportional hazard models. RESULTS: The study included 67 patients. In univariate analysis, variables that negatively influenced survival included TNM stage, microvascular invasion, and high TS expression. For the high TS expression group, median survival was 54 months and 5-year actuarial survival was 47%. For the low TS expression group, median survival was not reached and the 5-year actuarial survival was 91%. In multivariate analysis, only high TS expression remained an independent predictor of poor survival (HR = 10.77, 95% CI 1.36-84.91; P = 0.02). TS gene polymorphisms were not associated with TS expression or overall survival. CONCLUSIONS: High TS expression predicts poor outcome after resection of HCC. Molecular markers might be robust predictors of patient outcome after resection of HCC.
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Carcinoma Hepatocelular/cirugía , Neoplasias Hepáticas/cirugía , Timidilato Sintasa/genética , Anciano , Biomarcadores de Tumor/genética , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/patología , Supervivencia sin Enfermedad , Femenino , Regulación Neoplásica de la Expresión Génica/genética , Humanos , Estimación de Kaplan-Meier , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/patología , Masculino , Persona de Mediana Edad , Estadificación de Neoplasias , Pronóstico , Modelos de Riesgos Proporcionales , Resultado del TratamientoRESUMEN
Colorectal cancer (CRC) is the third most frequent neoplastic disorder and is a main cause of tumor-related mortality as many patients progress to stage IV metastatic CRC. Standard care consists of combination chemotherapy (FOLFIRI or FOLFOX). Patients with WT KRAS typing are eligible to receive anti-EGFR therapy combined with chemotherapy. Unfortunately, predicting efficacy of CRC anti-EGFR therapy has remained challenging. Here we uncover that the EGFR-pathway component RasGRP1 acts as CRC tumor suppressor in the context of aberrant Wnt signaling. We find that RasGRP1 suppresses EGF-driven proliferation of colonic epithelial organoids. Having established that RasGRP1 dosage levels impacts biology, we focused on CRC patients next. Mining five different data platforms, we establish that RasGRP1 expression levels decrease with CRC progression and predict poor clinical outcome of patients. Lastly, deletion of one or two Rasgrp1 alleles makes CRC spheroids more susceptible to EGFR inhibition. Retrospective analysis of the CALGB80203 clinical trial shows that addition of anti-EGFR therapy to chemotherapy significantly improves outcome for CRC patients when tumors express low RasGRP1 suppressor levels. In sum, RasGRP1 is a unique biomarker positioned in the EGFR pathway and of potential relevance to anti-EGFR therapy for CRC patients.
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Antineoplásicos Inmunológicos/uso terapéutico , Biomarcadores de Tumor/metabolismo , Neoplasias Colorrectales/tratamiento farmacológico , Proteínas de Unión al ADN/metabolismo , Factores de Intercambio de Guanina Nucleótido/metabolismo , Proteínas Supresoras de Tumor/metabolismo , Animales , Antineoplásicos Inmunológicos/farmacología , Biomarcadores de Tumor/análisis , Biomarcadores de Tumor/genética , Proliferación Celular/efectos de los fármacos , Cetuximab/farmacología , Cetuximab/uso terapéutico , Ensayos Clínicos como Asunto , Neoplasias Colorrectales/mortalidad , Neoplasias Colorrectales/patología , Biología Computacional , Proteínas de Unión al ADN/análisis , Proteínas de Unión al ADN/genética , Conjuntos de Datos como Asunto , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Supervivencia sin Enfermedad , Receptores ErbB/antagonistas & inhibidores , Receptores ErbB/metabolismo , Factores de Intercambio de Guanina Nucleótido/análisis , Factores de Intercambio de Guanina Nucleótido/genética , Humanos , Estimación de Kaplan-Meier , Ratones , Ratones Noqueados , Cultivo Primario de Células , Pronóstico , Transducción de Señal/efectos de los fármacos , Esferoides Celulares , Células Tumorales Cultivadas , Proteínas Supresoras de Tumor/análisis , Proteínas Supresoras de Tumor/genéticaRESUMEN
BACKGROUND: Resection of large retroperitoneal neoplasms may injure the femoral nerve, thereby causing a permanent neurological deficit. We used electrical neurophysiological monitoring to identify, map, and preserve the femoral nerve during surgical resection to reduce the risk of neurological deficit. METHODS: Seven patients with retroperitoneal neoplasms underwent eight resections. Compound muscle action potentials (CMAPs) were recorded from needle electrodes placed in the iliacus, quadriceps, and sartorius muscles. Spontaneous electromyography (EMG) was continuously monitored from the same muscle groups. A handheld monopolar stimulator was used to elicit evoked EMG responses to identify and map the course of the femoral nerve. A stimulating strength of 10 mA was used to map the nerve. The stimulation threshold was tested after neoplasm resection to predict postoperative femoral nerve function. RESULTS: Electrical stimulation with CMAP recording and a stimulating strength of 10 mA successfully localized the femoral nerve in six cases. Monitoring with a stimulating threshold between 0.6 and 1.6 mA predicted postoperative femoral nerve preservation after tumor resection in four of the six cases. CONCLUSION: Neurophysiological monitoring using CMAP and spontaneous EMG can protect the femoral nerve during resection of large retroperitoneal neoplasms.