RESUMEN
Growth hormone (GH) and insulin-like growth factor-1 (IGF1) play an important role in mammalian development, cell proliferation and lifespan. Especially in cases of tumor growth there is an urgent need to control the GH/IGF1 axis. In this study we screened a 38,480-compound library, and in two consecutive rounds of analogues selection, we identified active lead compounds based on the following criteria: inhibition the GH receptor (GHR) activity and its downstream effectors Jak2 and STAT5, and inhibition of growth of breast and colon cancer cells. The most active small molecule (BM001) inhibited both the GH/IGF1 axis and cell proliferation with an IC50 of 10-30 nM of human cancer cells. BM001 depleted GHR in human lymphoblasts. In preclinical xenografted experiments, BM001 showed a strong decrease in tumor volume in mice transplanted with MDA-MB-231 breast cancer cells. Mechanistically, the drug acts on the synthesis of the GHR. Our findings open the possibility to inhibit the GH/IGF1 axis with a small molecule.
Asunto(s)
Hormona de Crecimiento Humana , Receptores de Somatotropina , Animales , Proliferación Celular , Hormona del Crecimiento/fisiología , Humanos , Factor I del Crecimiento Similar a la Insulina , Mamíferos , RatonesRESUMEN
Transformed epithelial cells can activate programs of epithelial plasticity and switch from a sessile, epithelial phenotype to a motile, mesenchymal phenotype. This process is linked to the acquisition of an invasive phenotype and the formation of distant metastases. The development of compounds that block the acquisition of an invasive phenotype or revert the invasive mesenchymal phenotype into a more differentiated epithelial phenotype represent a promising anticancer strategy. In a high-throughput assay based on E-cadherin (re)induction and the inhibition of tumor cell invasion, 44,475 low molecular weight (LMW) compounds were screened. The screening resulted in the identification of candidate compounds from the PROAM02 class. Selected LMW compounds activated E-cadherin promoter activity and inhibited cancer cell invasion in multiple metastatic human cancer cell lines. The intraperitoneal administration of selected LMW compounds reduced the tumor burden in human prostate and breast cancer in vivo mouse models. Moreover, selected LMW compounds decreased the intra-bone growth of xenografted human prostate cancer cells. This study describes the identification of the PROAM02 class of small molecules that can be exploited to reduce cancer cell invasion and metastases. Further clinical evaluation of selected candidate inhibitors is warranted to address their safety, bioavailability and antitumor efficacy in the management of patients with aggressive cancers.
Asunto(s)
Neoplasias de la Mama/patología , Movimiento Celular , Descubrimiento de Drogas , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Neoplasias de la Próstata/patología , Bibliotecas de Moléculas Pequeñas/farmacología , Animales , Apoptosis , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Proliferación Celular , Femenino , Ensayos Analíticos de Alto Rendimiento , Humanos , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Desnudos , Invasividad Neoplásica , Neoplasias de la Próstata/tratamiento farmacológico , Neoplasias de la Próstata/genética , Neoplasias de la Próstata/metabolismo , Células Tumorales Cultivadas , Ensayos Antitumor por Modelo de XenoinjertoAsunto(s)
Alérgenos/inmunología , Asma/inmunología , Asma/terapia , Desensibilización Inmunológica , Péptidos/inmunología , Proteínas de Plantas/inmunología , Ribonucleasas/inmunología , Animales , Desensibilización Inmunológica/métodos , Modelos Animales de Enfermedad , Humanos , Inyecciones Subcutáneas , Ratones , Resultado del TratamientoRESUMEN
Mutations in the gene encoding nucleophosmin (NPM1) carry a prognostic value for patients with acute myeloid leukemia (AML). Various techniques are currently being used to detect these mutations in routine molecular diagnostics. Incorporation of accurate NPM1 mutation detection on a gene expression platform would enable simultaneous detection with various other expression biomarkers. Here we present an array-based mutation detection using custom probes for NPM1 WT mRNA and NPM1 type A, B, and D mutant mRNA. This method was 100% accurate on a training cohort of 505 newly diagnosed unselected AML cases. Validation on an independent cohort of 143 normal-karyotype AML cases revealed no false-negative results, and one false positive (sensitivity 100.0% and specificity 98.7%). Based on this, we conclude that this method provides a reliable method for NPM1 mutation detection. The method can be applied to other genes/mutations as long as the mutant alleles are sufficiently highly expressed.
Asunto(s)
Algoritmos , Leucemia Mieloide Aguda/genética , Mutación , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Alelos , Sondas de ADN , Femenino , Expresión Génica , Humanos , Leucemia Mieloide Aguda/metabolismo , Masculino , Nucleofosmina , Pronóstico , ARN Mensajero/genética , ARN Mensajero/metabolismo , Sensibilidad y EspecificidadRESUMEN
Double (bi-allelic) mutations in the gene encoding the CCAAT/enhancer-binding protein-alpha (CEBPA) transcription factor have a favorable prognostic impact in acute myeloid leukemia (AML). Double mutations in CEBPA can be detected using various techniques, but it is a notoriously difficult gene to sequence due to its high GC-content. Here we developed a two-step gene expression classifier for accurate and standardized detection of CEBPA double mutations. The key feature of the two-step classifier is that it explicitly removes cases with low CEBPA expression, thereby excluding CEBPA hypermethylated cases that have similar gene expression profiles as a CEBPA double mutant, which would result in false-positive predictions. In the second step, we have developed a 55 gene signature to identity the true CEBPA double-mutation cases. This two-step classifier was tested on a cohort of 505 unselected AML cases, including 26 CEBPA double mutants, 12 CEBPA single mutants, and seven CEBPA promoter hypermethylated cases, on which its performance was estimated by a double-loop cross-validation protocol. The two-step classifier achieves a sensitivity of 96.2% (95% confidence interval [CI] 81.1 to 99.3) and specificity of 100.0% (95% CI 99.2 to 100.0). There are no false-positive detections. This two-step CEBPA double-mutation classifier has been incorporated on a microarray platform that can simultaneously detect other relevant molecular biomarkers, which allows for a standardized comprehensive diagnostic assay. In conclusion, gene expression profiling provides a reliable method for CEBPA double-mutation detection in patients with AML for clinical use.
Asunto(s)
Proteínas Potenciadoras de Unión a CCAAT/genética , Perfilación de la Expresión Génica , Leucemia Mieloide Aguda/diagnóstico , Leucemia Mieloide Aguda/genética , Mutación , Análisis de Secuencia por Matrices de Oligonucleótidos/métodos , Proteína alfa Potenciadora de Unión a CCAAT/metabolismo , Metilación de ADN , Regulación Leucémica de la Expresión Génica/genética , Humanos , Datos de Secuencia Molecular , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , PronósticoRESUMEN
High levels of BAALC, ERG, EVI1 and MN1 expression have been associated with shorter overall survival in AML but standardized and clinically validated assays are lacking. We have therefore developed and optimized an assay for standardized detection of these prognostic genes for patients with intermediate cytogenetic risk AML. In a training set of 147 intermediate cytogenetic risk cases we performed cross validations at 5 percentile steps of expression level and observed a bimodal significance profile for BAALC expression level and unimodal significance profiles for ERG and MN1 levels with no statistically significant cutoff points near the median expression level of BAALC, ERG or MN1. Of the possible cutoff points for expression levels of BAALC, ERG and MN1, just the 30th and 75th percentile of BAALC expression level and the 30th percentile of MN1 expression level cutoff points showed clinical significance. Of these only the 30th percentile of BAALC expression level reproduced in an independent verification (extended training) data set of 242 cytogenetically normal AML cases and successfully validated in an external cohort of 215 intermediate cytogenetic risk AML cases. Finally, we show independent prognostic value for high EVI1 and low BAALC in multivariate analysis with other clinically relevant molecular AML markers. We have developed a highly standardized molecular assay for the independent gene expression markers EVI1 and BAALC.