RESUMEN
Recent advances on the development of bumped kinase inhibitors for treatment of cryptosporidiosis have focused on the 5-aminopyrazole-4-carboxamide scaffold, due to analogs that have less hERG inhibition, superior efficacy, and strong in vitro safety profiles. Three compounds, BKI-1770, -1841, and -1708, showed strong efficacy in C. parvum infected mice. Both BKI-1770 and BKI-1841 had efficacy in the C. parvum newborn calf model, reducing diarrhea and oocyst excretion. However, both compounds caused hyperflexion of the limbs seen as dropped pasterns. Toxicity experiments in rats and calves dosed with BKI-1770 showed enlargement of the epiphyseal growth plate at doses only slightly higher than the efficacious dose. Mice were used as a screen to check for bone toxicity, by changes to the tibia epiphyseal growth plate, or neurological causes, by use of a locomotor activity box. These results showed neurological effects from both BKI-1770 and BKI-1841 and bone toxicity in mice from BKI-1770, indicating one or both effects may be contributing to toxicity. However, BKI-1708 remains a viable treatment candidate for further evaluation as it showed no signs of bone toxicity or neurological effects in mice.
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Antineoplásicos , Antiprotozoarios , Criptosporidiosis , Cryptosporidium parvum , Animales , Bovinos , Ratones , Ratas , Criptosporidiosis/tratamiento farmacológico , Antiprotozoarios/farmacología , Antineoplásicos/farmacología , Inhibidores de Proteínas Quinasas/farmacología , OocistosRESUMEN
Cerebellar dysfunction may substantially contribute to the clinical symptoms of Parkinson's disease (PD). The role of cerebellar subregions in tremors and gait disturbances in PD remains unknown. To investigate alterations in cerebellar subregion volumes and functional connectivity (FC), as well as FC between the dentate nucleus (DN) and ventral lateral posterior nucleus (VLp) of the thalamus, which are potentially involved in different PD motor subtypes. We conducted morphometric and resting-state functional connectivity analyses in various cerebellar subregions in 22 tremor-dominant (TD)-PD and 35 postural instability gait difficulty dominant (PIGD)-PD patients and 38 sex- and age-matched healthy controls (HCs). The volume and FC alterations in various cerebellar subregions and the neural correlates of these changes with the clinical severity scores were investigated. The PIGD-PD group showed greater FC between the right motor cerebellum (CBMm) and left postcentral gyrus than the HC group, and a higher FC was associated with less severe PIGD symptoms. In contrast, the TD-PD group had decreased FC between the right DN and left VLp compared with the PIGD-PD and HC groups, and lower FC was associated with worse TD symptoms. Furthermore, the PIGD-PD group had higher FC between the left DN and left inferior temporal gyrus than the TD-PD group. Morphometric analysis revealed that the TD-PD group showed a significantly higher volume of left CBMm than the HC group. Our findings point to differential alteration patterns in cerebellar subregions and offer a new perspective on the pathophysiology of motor subtypes of PD.
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Trastornos Neurológicos de la Marcha , Enfermedad de Parkinson , Humanos , Enfermedad de Parkinson/complicaciones , Enfermedad de Parkinson/diagnóstico por imagen , Imagen por Resonancia Magnética , Temblor/etiología , Temblor/complicaciones , Cerebelo/diagnóstico por imagen , Corteza Somatosensorial , Trastornos Neurológicos de la Marcha/diagnóstico por imagen , Trastornos Neurológicos de la Marcha/etiologíaRESUMEN
MOTIVATION: Non-linear ordinary differential equation (ODE) models that contain numerous parameters are suitable for inferring an emulated gene regulatory network (eGRN). However, the number of experimental measurements is usually far smaller than the number of parameters of the eGRN model that leads to an underdetermined problem. There is no unique solution to the inference problem for an eGRN using insufficient measurements. RESULTS: This work proposes an evolutionary modelling algorithm (EMA) that is based on evolutionary intelligence to cope with the underdetermined problem. EMA uses an intelligent genetic algorithm to solve the large-scale parameter optimization problem. An EMA-based method, GREMA, infers a novel type of gene regulatory network with confidence levels for every inferred regulation. The higher the confidence level is, the more accurate the inferred regulation is. GREMA gradually determines the regulations of an eGRN with confidence levels in descending order using either an S-system or a Hill function-based ODE model. The experimental results showed that the regulations with high-confidence levels are more accurate and robust than regulations with low-confidence levels. Evolutionary intelligence enhanced the mean accuracy of GREMA by 19.2% when using the S-system model with benchmark datasets. An increase in the number of experimental measurements may increase the mean confidence level of the inferred regulations. GREMA performed well compared with existing methods that have been previously applied to the same S-system, DREAM4 challenge and SOS DNA repair benchmark datasets. AVAILABILITY AND IMPLEMENTATION: All of the datasets that were used and the GREMA-based tool are freely available at https://nctuiclab.github.io/GREMA. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
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Algoritmos , Redes Reguladoras de Genes , Evolución Biológica , Biología Computacional , InteligenciaRESUMEN
Macrovascular complications develop in over a half of the diabetic individuals, resulting in high morbidity and mortality. This poses a severe threat to public health and a heavy burden to social economy. It is therefore important to develop effective approaches to prevent or slow down the pathogenesis and progression of macrovascular complications of diabetes (MCD). Oxidative stress is a major contributor to MCD. Nuclear factor (erythroid-derived 2)-like 2 (NRF2) governs cellular antioxidant defence system by activating the transcription of various antioxidant genes, combating diabetes-induced oxidative stress. Accumulating experimental evidence has demonstrated that NRF2 activation protects against MCD. Structural inhibition of Kelch-like ECH-associated protein 1 (KEAP1) is a canonical way to activate NRF2. More recently, novel approaches, such as activation of the Nfe2l2 gene transcription, decreasing KEAP1 protein level by microRNA-induced degradation of Keap1 mRNA, prevention of proteasomal degradation of NRF2 protein and modulation of other upstream regulators of NRF2, have emerged in prevention of MCD. This review provides a brief introduction of the pathophysiology of MCD and the role of oxidative stress in the pathogenesis of MCD. By reviewing previous work on the activation of NRF2 in MCD, we summarize strategies to activate NRF2, providing clues for future intervention of MCD. Controversies over NRF2 activation and future perspectives are also provided in this review.
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Complicaciones de la Diabetes/metabolismo , Diabetes Mellitus/metabolismo , Angiopatías Diabéticas/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , Animales , HumanosRESUMEN
Tumor-infiltrating immune cells play a crucial role in tumor progression and response to treatment. However, the limited studies on infiltrating immune cells have shown inconsistent and even controversial results for osteosarcoma (OS). In addition, the dynamic changes of infiltrating immune cells after neoadjuvant chemotherapy are largely unknown. We downloaded the RNA expression matrix and clinical information of 80 OS patients from the TARGET database. CIBERSORT was used to evaluate the proportion of 22 immune cell types in patients based on gene expression data. M2 macrophages were found to be the most abundant immune cell type and were associated with improved survival in OS. Another cohort of pretreated OS samples was evaluated by immunohistochemistry to validate the results from CIBERSORT analysis. Matched biopsy and surgical samples from 27 patients were collected to investigate the dynamic change of immune cells and factors before and after neoadjuvant chemotherapy. Neoadjuvant chemotherapy was associated with increased densities of CD3+ T cells, CD8+ T cells, Ki67 + CD8+ T cells and PD-L1+ immune cells. Moreover, HLA-DR-CD33+ myeloid-derived suppressive cells (MDSC) were decreased after treatment. We determined that the application of chemotherapy may activate the local immune status and convert OS into an immune "hot" tumor. These findings provide rationale for investigating the schedule of immunotherapy treatment in OS patients in future clinical trials.
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Neoplasias Óseas/inmunología , Linfocitos Infiltrantes de Tumor/inmunología , Osteosarcoma/inmunología , Microambiente Tumoral/efectos de los fármacos , Microambiente Tumoral/inmunología , Adolescente , Adulto , Antineoplásicos Inmunológicos/uso terapéutico , Neoplasias Óseas/tratamiento farmacológico , Neoplasias Óseas/patología , Quimioterapia Adyuvante/métodos , Niño , Femenino , Humanos , Linfocitos Infiltrantes de Tumor/efectos de los fármacos , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Masculino , Células Supresoras de Origen Mieloide/efectos de los fármacos , Células Supresoras de Origen Mieloide/inmunología , Terapia Neoadyuvante/métodos , Osteosarcoma/tratamiento farmacológico , Osteosarcoma/patologíaRESUMEN
Bumped kinase inhibitors (BKIs) have been shown to be potent inhibitors of Toxoplasma gondii calcium-dependent protein kinase 1. Pyrazolopyrimidine and 5-aminopyrazole-4-carboxamide scaffold-based BKIs are effective in acute and chronic experimental models of toxoplasmosis. Through further exploration of these 2 scaffolds and a new pyrrolopyrimidine scaffold, additional compounds have been identified that are extremely effective against acute experimental toxoplasmosis. The in vivo efficacy of these BKIs demonstrates that the cyclopropyloxynaphthyl, cyclopropyloxyquinoline, and 2-ethoxyquinolin-6-yl substituents are associated with efficacy across scaffolds. In addition, a broad range of plasma concentrations after oral dosing resulted from small structural changes to the BKIs. These select BKIs include anti-Toxoplasma compounds that are effective against acute experimental toxoplasmosis and are not toxic in human cell assays, nor to mice when administered for therapy. The BKIs described here are promising late leads for improving anti-Toxoplasma therapy.
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Inhibidores de Proteínas Quinasas/uso terapéutico , Proteínas Protozoarias/antagonistas & inhibidores , Pirazoles/uso terapéutico , Pirimidinas/uso terapéutico , Toxoplasmosis Animal/tratamiento farmacológico , Toxoplasmosis Cerebral/tratamiento farmacológico , Administración Oral , Animales , Área Bajo la Curva , Femenino , Técnicas In Vitro , Ratones , Inhibidores de Proteínas Quinasas/sangre , Inhibidores de Proteínas Quinasas/farmacología , Pirazoles/sangre , Pirazoles/farmacología , Pirimidinas/sangre , Pirimidinas/farmacologíaRESUMEN
Oxidative stress and P53 contribute to the pathogenesis of diabetic kidney disease (DKD). Nuclear factor erythroid 2-related factor 2 (NRF2) is a master regulator of cellular antioxidant defense system, is negatively regulated by P53 and prevents DKD. Recent findings revealed an important role of mouse double minute 2 (MDM2) in protection against DKD. However, the mechanism remained unclear. We hypothesized that MDM2 enhances NRF2 antioxidant signaling in DKD given that MDM2 is a key negative regulator of P53. The MDM2 inhibitor nutlin3a elevated renal P53, inhibited NRF2 signaling and induced oxidative stress, inflammation, fibrosis, DKD-like renal pathology and albuminuria in the wild-type (WT) non-diabetic mice. These effects exhibited more prominently in nutlin3a-treated WT diabetic mice. Interestingly, nutlin3a failed to induce greater renal injuries in the Nrf2 knockout (KO) mice under both the diabetic and non-diabetic conditions, indicating that NRF2 predominantly mediates MDM2's action. On the contrary, P53 inhibition by pifithrin-α activated renal NRF2 signaling and the expression of Mdm2, and attenuated DKD in the WT diabetic mice, but not in the Nrf2 KO diabetic mice. In high glucose-treated mouse mesangial cells, P53 gene silencing completely abolished nutlin3a's inhibitory effect on NRF2 signaling. The present study demonstrates for the first time that MDM2 controls renal NRF2 antioxidant activity in DKD via inhibition of P53, providing MDM2 activation and P53 inhibition as novel strategies in the management of DKD.
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Diabetes Mellitus Experimental/metabolismo , Nefropatías Diabéticas/metabolismo , Factor 2 Relacionado con NF-E2/metabolismo , Estrés Oxidativo , Proteínas Proto-Oncogénicas c-mdm2/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Animales , Células Cultivadas , Diabetes Mellitus Experimental/complicaciones , Diabetes Mellitus Experimental/genética , Diabetes Mellitus Experimental/patología , Nefropatías Diabéticas/genética , Nefropatías Diabéticas/patología , Nefropatías Diabéticas/prevención & control , Silenciador del Gen , Riñón/metabolismo , Riñón/patología , Masculino , Ratones Endogámicos C57BL , Ratones Noqueados , Factor 2 Relacionado con NF-E2/genética , Proteína p53 Supresora de Tumor/antagonistas & inhibidores , Proteína p53 Supresora de Tumor/genéticaRESUMEN
Endothelial dysfunction contributes to diabetic macrovascular complications, resulting in high mortality. Recent findings demonstrate a pathogenic role of P53 in endothelial dysfunction, encouraging the investigation of the effect of P53 inhibition on diabetic endothelial dysfunction. Thus, high glucose (HG)-treated endothelial cells (ECs) were subjected to pifithrin-α (PFT-α)-a specific inhibitor of P53, or P53-small interfering RNA (siRNA), both of which attenuated the HG-induced endothelial inflammation and oxidative stress. Moreover, inhibition of P53 by PFT-α or P53-siRNA prohibited P53 acetylation, decreased microRNA-34a (miR-34a) level, leading to a dramatic increase in sirtuin 1 (SIRT1) protein level. Interestingly, the miR-34a inhibitor (miR-34a-I) and PFT-α increased SIRT1 protein level and alleviated the HG-induced endothelial inflammation and oxidative stress to a similar extent; however, these effects of PFT-α were completely abrogated by the miR-34a mimic. In addition, SIRT1 inhibition by EX-527 or Sirt1-siRNA completely abolished miR-34a-I's protection against HG-induced endothelial inflammation and oxidative stress. Furthermore, in the aortas of streptozotocin-induced diabetic mice, both PFT-α and miR-34a-I rescued the inflammation, oxidative stress and endothelial dysfunction caused by hyperglycaemia. Hence, the present study has uncovered a P53/miR-34a/SIRT1 pathway that leads to endothelial dysfunction, suggesting that P53/miR-34a inhibition could be a viable strategy in the management of diabetic macrovascular diseases.
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Diabetes Mellitus Experimental/genética , Endotelio Vascular/metabolismo , MicroARNs/genética , Sirtuina 1/genética , Proteína p53 Supresora de Tumor/genética , Animales , Benzotiazoles/farmacología , Carbazoles/farmacología , Células Cultivadas , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/fisiopatología , Células Endoteliales/efectos de los fármacos , Células Endoteliales/metabolismo , Endotelio Vascular/fisiopatología , Activación Enzimática/efectos de los fármacos , Activación Enzimática/genética , Regulación de la Expresión Génica/efectos de los fármacos , Glucosa/farmacología , Masculino , Ratones Endogámicos C57BL , Interferencia de ARN , Sirtuina 1/antagonistas & inhibidores , Sirtuina 1/metabolismo , Tolueno/análogos & derivados , Tolueno/farmacología , Proteína p53 Supresora de Tumor/antagonistas & inhibidores , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
Cryptosporidiosis is one of the leading causes of moderate to severe diarrhea in children in low-resource settings. The therapeutic options for cryptosporidiosis are limited to one drug, nitazoxanide, which unfortunately has poor activity in the most needy populations of malnourished children and HIV-infected persons. We describe here the discovery and early optimization of a class of imidazopyridine-containing compounds with potential for treating Cryptosporidium infections. The compounds target the Cryptosporidium methionyl-tRNA synthetase (MetRS), an enzyme that is essential for protein synthesis. The most potent compounds inhibited the enzyme with Ki values in the low picomolar range. Cryptosporidium cells in culture were potently inhibited with 50% effective concentrations as low as 7 nM and >1,000-fold selectivity over mammalian cells. A parasite persistence assay indicates that the compounds act by a parasiticidal mechanism. Several compounds were demonstrated to control infection in two murine models of cryptosporidiosis without evidence of toxicity. Pharmacological and physicochemical characteristics of compounds were investigated to determine properties that were associated with higher efficacy. The results indicate that MetRS inhibitors are excellent candidates for development for anticryptosporidiosis therapy.
Asunto(s)
Antiprotozoarios/farmacología , Criptosporidiosis/tratamiento farmacológico , Cryptosporidium parvum/efectos de los fármacos , Imidazoles/farmacología , Metionina-ARNt Ligasa/antagonistas & inhibidores , Piridinas/farmacología , Animales , Cryptosporidium parvum/genética , Ciclooxigenasa 1/efectos de los fármacos , Modelos Animales de Enfermedad , Descubrimiento de Drogas/métodos , Femenino , Células Hep G2 , Humanos , Imidazoles/química , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Piridinas/químicaRESUMEN
Colorectal cancer (CRC) is the third most common cancer worldwide, with more than 1.3 million new cases and 690 000 deaths each year. In China, the incidence of CRC has increased dramatically due to dietary and lifestyle changes, to become the fifth leading cause of cancer-related death. Here, we performed whole-exome sequencing in 50 rectal cancer cases among the Chinese population as part of the International Cancer Genome Consortium research project. Frequently mutated genes and enriched pathways were identified. Moreover, a previously unreported gene, PCDHB3, was found frequently mutated in 5.19% cases. Additionally, PCDHB3 expression was found decreased in 81.6% of CRC tissues and all eight CRC cell lines tested. Low expression and cytoplasmic localization of PCDHB3 predict poor prognosis in advanced CRC. Copy number decrease and/or CpG island hypermethylation contributes to the pervasive decreased expression of PCDHB3. PCDHB3 inhibits CRC cell proliferation, migration, and epithelial-mesenchymal transition. The tumor-suppressive effects of PCDHB3 are partially due to inhibition of NF-κB transcriptional activity through K63 deubiquitination of p50 at lysine 244/252, which increases the binding affinity of inactive p50 homodimer to κB DNA, resulting in competitive inhibition of the transcription of NF-κB target genes by p65 dimers. Our study identified PCDHB3 as a novel tumor suppressor in CRC via inhibition of the NF-κB pathway, and its expression and localization may serve as prognostic markers for advanced CRC. Copyright © 2018 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.
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Biomarcadores de Tumor/genética , Cadherinas/genética , Neoplasias Colorrectales/genética , Secuenciación del Exoma , Silenciador del Gen , Genes Supresores de Tumor , Mutación , Adulto , Anciano , Animales , Pueblo Asiatico/genética , Biomarcadores de Tumor/metabolismo , Cadherinas/metabolismo , Línea Celular Tumoral , Movimiento Celular , Proliferación Celular , China , Neoplasias Colorrectales/etnología , Neoplasias Colorrectales/metabolismo , Neoplasias Colorrectales/patología , Islas de CpG , Metilación de ADN , Regulación hacia Abajo , Femenino , Dosificación de Gen , Regulación Neoplásica de la Expresión Génica , Predisposición Genética a la Enfermedad , Humanos , Masculino , Ratones Endogámicos BALB C , Ratones Desnudos , Persona de Mediana Edad , FN-kappa B/genética , FN-kappa B/metabolismo , Fenotipo , ProtocadherinasRESUMEN
Rho guanine nucleotide exchange factors (RhoGEFs) are proteins that activate Rho GTPases in response to extracellular stimuli and regulate various biologic processes. ARHGEF19, one of RhoGEFs, was reported to activate RhoA in the Wnt-PCP pathway controlling convergent extension in Xenopus gastrulation. The goal of our study was to identify the role and molecular mechanisms of ARHGEF19 in the tumorigenesis of non-small cell lung cancer (NSCLC). ARHGEF19 expression was significantly elevated in NSCLC tissues, and ARHGEF19 levels were significantly associated with lymph node status, distant metastasis and TNM stage; Patients with high ARHGEF19 levels had poor overall survival (OS) and progression-free survival (PFS). Our investigations revealed that ARHGEF19 overexpression promoted the cell proliferation, invasion and metastasis of lung cancer cells, whereas knockdown of this gene inhibited these processes. Mechanistically, ARHGEF19 activated the mitogen-activated protein kinase (MAPK) pathway in a RhoA-independent manner: ARHGEF19 interacted with BRAF and facilitated the phosphorylation of its downstream kinase MEK1/2; both the Dbl homology (DH) and Pleckstrin homology (PH) domains of ARHGEF19 were indispensable for the phosphorylation of MEK1/2. Furthermore, downregulation of miR-29b was likely responsible for the increased expression of ARHGEF19 in lung cancer tissues and, consequently, the abnormal activation of MAPK signaling. These findings suggest that ARHGEF19 upregulation, due to the low expression of miR-29 in NSCLC tissues, may play a crucial role in NSCLC tumorigenesis by activating MAPK signaling. ARHGEF19 could serve as a negative prognostic marker as well as a therapeutic target for NSCLC patients.
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Carcinogénesis/metabolismo , Carcinoma de Pulmón de Células no Pequeñas/patología , Regulación Neoplásica de la Expresión Génica/fisiología , Factores de Intercambio de Guanina Nucleótido/metabolismo , Neoplasias Pulmonares/patología , Animales , Área Bajo la Curva , Carcinogénesis/genética , Carcinogénesis/patología , Carcinoma de Pulmón de Células no Pequeñas/genética , Carcinoma de Pulmón de Células no Pequeñas/metabolismo , Supervivencia sin Enfermedad , Femenino , Factores de Intercambio de Guanina Nucleótido/genética , Xenoinjertos , Humanos , Estimación de Kaplan-Meier , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/metabolismo , Masculino , Ratones , Ratones Endogámicos BALB C , MicroARNs/biosíntesis , MicroARNs/genética , Persona de Mediana Edad , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Proteínas Proto-Oncogénicas B-raf/metabolismo , Curva ROC , Sensibilidad y Especificidad , Transducción de Señal/fisiologíaRESUMEN
BACKGROUND/AIMS: RBFOX3, an RNA-binding fox protein, plays an important role in the differentiation of neuronal development, but its role in the chemosensitivity of hepatocellular carcinoma (HCC) to 5-FU is unknown. METHODS: In this study, we examined the biological functions of RBFOX3 and its effect on the chemosensitivity of HCC cells to 5-FU in vitro and in a mouse xenograft model. RESULTS: RBFOX3 was found to have elevated expression in HCC cell lines and tissue samples, and its knockdown inhibited HCC cell proliferation. Moreover, knockdown of RBFOX3 improved the inhibitory effect of 5-fluorouracil (5-FU) on cell proliferation, migration and invasion, and enhanced the apoptosis induced by 5-FU. However, overexpression of RBFOX3 reduced the inhibitory effect of 5-fluorouracil (5-FU) on cell proliferation, migration and invasion, and decreased the apoptosis induced by 5-FU. We further elucidated that RBFOX3 knockdown synergized with 5-FU to inhibit the growth and invasion of HCC cells through PI3K/AKT and epithelial-mesenchymal transition (EMT) signaling, and promote apoptosis by activating the cytochrome-c/caspase signaling pathway. Finally, we validated that RBFOX3 regulated 5-FU-mediated cytotoxicity in HCC in mouse xenograft models. CONCLUSIONS: The findings from this study indicate that RBFOX3 regulates the chemosensitivity of HCC to 5-FU in vitro and in vivo. Therefore, targeting RBFOX3 may improve the inhibition of HCC growth and progression by 5-FU, and provide a novel potential therapeutic strategy for HCC.
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Antígenos Nucleares/metabolismo , Resistencia a Antineoplásicos/efectos de los fármacos , Transición Epitelial-Mesenquimal/efectos de los fármacos , Fluorouracilo/toxicidad , Proteínas del Tejido Nervioso/metabolismo , Transducción de Señal/efectos de los fármacos , Animales , Antígenos Nucleares/genética , Apoptosis/efectos de los fármacos , Carcinoma Hepatocelular/tratamiento farmacológico , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patología , Caspasas/metabolismo , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Citocromos c/metabolismo , Fluorouracilo/uso terapéutico , Humanos , Neoplasias Hepáticas/tratamiento farmacológico , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patología , Ratones , Ratones Desnudos , Microscopía Fluorescente , Metástasis de la Neoplasia , Proteínas del Tejido Nervioso/antagonistas & inhibidores , Proteínas del Tejido Nervioso/genética , Fosfatidilinositol 3-Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Trasplante HeterólogoRESUMEN
Motivation: Numerous ubiquitination sites remain undiscovered because of the limitations of mass spectrometry-based methods. Existing prediction methods use randomly selected non-validated sites as non-ubiquitination sites to train ubiquitination site prediction models. Results: We propose an evolutionary screening algorithm (ESA) to select effective negatives among non-validated sites and an ESA-based prediction method, ESA-UbiSite, to identify human ubiquitination sites. The ESA selects non-validated sites least likely to be ubiquitination sites as training negatives. Moreover, the ESA and ESA-UbiSite use a set of well-selected physicochemical properties together with a support vector machine for accurate prediction. Experimental results show that ESA-UbiSite with effective negatives achieved 0.92 test accuracy and a Matthews's correlation coefficient of 0.48, better than existing prediction methods. The ESA increased ESA-UbiSite's test accuracy from 0.75 to 0.92 and can improve other post-translational modification site prediction methods. Availability and Implementation: An ESA-UbiSite-based web server has been established at http://iclab.life.nctu.edu.tw/iclab_webtools/ESAUbiSite/ . Contact: syho@mail.nctu.edu.tw. Supplementary information: Supplementary data are available at Bioinformatics online.
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Biología Computacional/métodos , Programas Informáticos , Máquina de Vectores de Soporte , Ubiquitinación , HumanosRESUMEN
There is a substantial need for novel therapeutics to combat the widespread impact caused by Crytosporidium infection. However, there is a lack of knowledge as to which drug pharmacokinetic (PK) characteristics are key to generate an in vivo response, specifically whether systemic drug exposure is crucial for in vivo efficacy. To identify which PK properties are correlated with in vivo efficacy, we generated physiologically based PK models to simulate systemic and gastrointestinal drug concentrations for a series of bumped kinase inhibitors (BKIs) that have nearly identical in vitro potency against Cryptosporidium but display divergent PK properties. When BKI concentrations were used to predict in vivo efficacy with a neonatal model of Cryptosporidium infection, these concentrations in the large intestine were the sole predictors of the observed in vivo efficacy. The significance of large intestinal BKI exposure for predicting in vivo efficacy was further supported with an adult mouse model of Cryptosporidium infection. This study suggests that drug exposure in the large intestine is essential for generating a superior in vivo response, and that physiologically based PK models can assist in the prioritization of leading preclinical drug candidates for in vivo testing.
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Criptosporidiosis/tratamiento farmacológico , Tracto Gastrointestinal/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacocinética , Animales , Cryptosporidium parvum/efectos de los fármacos , Cryptosporidium parvum/aislamiento & purificación , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Femenino , Tracto Gastrointestinal/metabolismo , Concentración 50 Inhibidora , Ratones , Ratones Noqueados , Modelos Teóricos , Naftalenos/farmacocinética , Piperidinas/farmacocinética , Inhibidores de Proteínas Quinasas/sangre , Pirazoles/farmacocinéticaRESUMEN
BACKGROUND: N-myc (and STAT) interactor (NMI) plays vital roles in tumor growth, progression, and metastasis. In this study, we identified NMI as a potential tumor suppressor in lung cancer and explored its molecular mechanism involved in lung cancer progression. METHODS: Human lung cancer cell lines and a mouse xenograft model was used to study the effect of NMI on tumor growth. The expression of NMI, COX-2 and relevant signaling proteins were examined by Western blot. Tissue microarray immunohistochemical analysis was performed to assess the correlation between NMI and COX-2 expression in lung cancer patients. RESULTS: NMI was highly expressed in normal lung cells and tissues, but lowly expressed in lung cancer cells and tissues. Overexpression of NMI induced apoptosis, suppressed lung cancer cell growth and migration, which were mediated by up-regulation of the cleaved caspase-3/9 and down-regulation of phosphorylated PI3K/AKT, MMP2/MMP9, ß-cadherin, and COX-2/PGE2. In contrast, knockdown of NMI promoted lung cancer cell colony formation and migration, which were correlated with the increased expression of phosphorylated PI3K/AKT, MMP2/MMP9, ß-cadherin and COX-2/PGE2. Further study showed that NMI suppressed COX-2 expression through inhibition of the p50/p65 NF-κB acetylation mediated by p300. The xenograft lung cancer mouse models also confirmed the NMI-mediated suppression of tumor growth by inhibiting COX-2 signaling. Moreover, tissue microarray immunohistochemical analysis of lung adenocarcinomas also demonstrated a negative correlation between NMI and COX-2 expression. Kaplan-Meier analysis indicated that the patients with high level of NMI had a significantly better prognosis. CONCLUSIONS: Our study showed that NMI suppressed tumor growth by inhibiting PI3K/AKT, MMP2/MMP9, COX-2/PGE2 signaling pathways and p300-mediated NF-κB acetylation, and predicted a favorable prognosis in human lung adenocarcinomas, suggesting that NMI was a potential tumor suppressor in lung cancer.
Asunto(s)
Adenocarcinoma/genética , Adenocarcinoma/patología , Regulación Neoplásica de la Expresión Génica , Péptidos y Proteínas de Señalización Intracelular/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Adenocarcinoma/metabolismo , Adenocarcinoma/mortalidad , Adenocarcinoma del Pulmón , Adulto , Anciano , Animales , Apoptosis/genética , Biomarcadores de Tumor , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular , Supervivencia Celular/genética , Ciclooxigenasa 2/genética , Ciclooxigenasa 2/metabolismo , Dinoprostona/biosíntesis , Modelos Animales de Enfermedad , Regulación hacia Abajo , Proteína p300 Asociada a E1A/metabolismo , Femenino , Humanos , Inmunohistoquímica , Péptidos y Proteínas de Señalización Intracelular/metabolismo , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/mortalidad , Masculino , Ratones , Persona de Mediana Edad , FN-kappa B/metabolismo , Estadificación de Neoplasias , Pronóstico , Regiones Promotoras Genéticas , Transducción de Señal , Activación Transcripcional , Carga Tumoral , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Antibiotic-resistant bacteria are widespread and pose a growing threat to human health. New antibiotics acting by novel mechanisms of action are needed to address this challenge. The bacterial methionyl-tRNA synthetase (MetRS) enzyme is essential for protein synthesis, and the type found in Gram-positive bacteria is substantially different from its counterpart found in the mammalian cytoplasm. Both previously published and new selective inhibitors were shown to be highly active against Gram-positive bacteria with MICs of ≤1.3 µg/ml against Staphylococcus, Enterococcus, and Streptococcus strains. Incorporation of radioactive precursors demonstrated that the mechanism of activity was due to the inhibition of protein synthesis. Little activity against Gram-negative bacteria was observed, consistent with the fact that Gram-negative bacterial species contain a different type of MetRS enzyme. The ratio of the MIC to the minimum bactericidal concentration (MBC) was consistent with a bacteriostatic mechanism. The level of protein binding of the compounds was high (>95%), and this translated to a substantial increase in MICs when the compounds were tested in the presence of serum. Despite this, the compounds were very active when they were tested in a Staphylococcus aureus murine thigh infection model. Compounds 1717 and 2144, given by oral gavage, resulted in 3- to 4-log decreases in the bacterial load compared to that in vehicle-treated mice, which was comparable to the results observed with the comparator drugs, vancomycin and linezolid. In summary, the research describes MetRS inhibitors with oral bioavailability that represent a class of compounds acting by a novel mechanism with excellent potential for clinical development.
Asunto(s)
Antibacterianos/química , Antibacterianos/farmacología , Inhibidores Enzimáticos/farmacología , Bacterias Grampositivas/efectos de los fármacos , Metionina-ARNt Ligasa/antagonistas & inhibidores , Animales , Antibacterianos/metabolismo , Antibacterianos/farmacocinética , Proteínas Sanguíneas/metabolismo , Farmacorresistencia Bacteriana/efectos de los fármacos , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/metabolismo , Inhibidores Enzimáticos/farmacocinética , Escherichia coli/efectos de los fármacos , Femenino , Infecciones por Bacterias Grampositivas/tratamiento farmacológico , Infecciones por Bacterias Grampositivas/microbiología , Humanos , Inactivación Metabólica , Ratones , Pruebas de Sensibilidad Microbiana , Microsomas Hepáticos , Staphylococcus aureus/efectos de los fármacosRESUMEN
Cryptosporidium parvum calcium-dependent protein kinase 1 (CpCDPK1) is a promising target for drug development against cryptosporidiosis. We report a series of low-nanomolar CpCDPK1 5-aminopyrazole-4-carboxamide (AC) scaffold inhibitors that also potently inhibit C. parvum growth in vitro Correlation between anti-CpCDPK1 and C. parvum growth inhibition, as previously reported for pyrazolopyrimidines, was not apparent. Nonetheless, lead AC compounds exhibited a substantial reduction of parasite burden in the neonatal mouse cryptosporidiosis model when dosed at 25 mg/kg.
Asunto(s)
Antiprotozoarios/farmacología , Criptosporidiosis/tratamiento farmacológico , Cryptosporidium parvum/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacología , Proteínas Quinasas/metabolismo , Animales , Antiprotozoarios/química , Criptosporidiosis/parasitología , Cryptosporidium parvum/crecimiento & desarrollo , Ratones , Proteínas Protozoarias/metabolismo , Pirazoles/química , Pirazoles/farmacologíaRESUMEN
UNLABELLED: The phosphatidylinositol 3-kinase/phosphatidylinositol 3,4,5-trisphosphate 3-phosphatase/protein kinase B/mammalian target of rapamycin (PI3K-PTEN-AKT-mTOR) pathway is a central controller of cell growth and a key driver for human cancer. MAF1 is an mTOR downstream effector and transcriptional repressor of ribosomal and transfer RNA genes. MAF1 expression is markedly reduced in hepatocellular carcinomas, which is correlated with disease progression and poor prognosis. Consistently, MAF1 displays tumor-suppressor activity toward in vitro and in vivo cancer models. Surprisingly, blocking the synthesis of ribosomal and transfer RNAs is insufficient to account for MAF1's tumor-suppressor function. Instead, MAF1 down-regulation paradoxically leads to activation of AKT-mTOR signaling, which is mediated by decreased PTEN expression. MAF1 binds to the PTEN promoter, enhancing PTEN promoter acetylation and activity. CONCLUSION: In contrast to its canonical function as a transcriptional repressor, MAF1 can also act as a transcriptional activator for PTEN, which is important for MAF1's tumor-suppressor function. These results have implications in disease staging, prognostic prediction, and AKT-mTOR-targeted therapy in liver cancer. (Hepatology 2016;63:1928-1942).
Asunto(s)
Carcinoma Hepatocelular/metabolismo , Regulación Neoplásica de la Expresión Génica , Neoplasias Hepáticas Experimentales/metabolismo , Proteínas Represoras/metabolismo , Adulto , Anciano , Animales , Línea Celular Tumoral , Proliferación Celular , Progresión de la Enfermedad , Regulación hacia Abajo , Femenino , Humanos , Masculino , Ratones Desnudos , Persona de Mediana Edad , Invasividad Neoplásica , Fosfohidrolasa PTEN/metabolismo , Regiones Promotoras Genéticas , Proteínas Proto-Oncogénicas c-akt/metabolismo , Serina-Treonina Quinasas TOR/metabolismoRESUMEN
Many pyrrolizidine alkaloids (PAs) can cause liver injury in animals and humans. Different hepatotoxic PAs can produce similar hepatotoxic effects, but the degree of their toxicities may vary widely. Retrorsine (RTS) and monocrotaline (MCT) share the same core structure (retronecine) and similar metabolic activation pathway. RTS and MCT both produced liver injury, but the former was more hepatotoxic than the latter. Enzyme kinetic study demonstrated that the value of Vmax/Km for RTS was 5.5-fold larger than that of MCT. Additionally, RTS produced higher levels of pyrrole-glutathione (GSH) conjugates and protein covalent binding than MCT at the same dose. Furthermore, RTS induced significant hepatic GSH depletion but MCT did little. This comparative study provides clear evidence that the generation of the reactive pyrrolic intermediates plays a critical role in PA-induced hepatotoxicity.
Asunto(s)
Hígado/efectos de los fármacos , Monocrotalina/toxicidad , Alcaloides de Pirrolicidina/toxicidad , Animales , Glutatión/metabolismo , Cinética , Hígado/enzimología , Hígado/metabolismo , Masculino , Ratones , Microsomas Hepáticos/metabolismoRESUMEN
Methyleugenol (ME), an alkenylbenzene compound, is a natural ingredient of several herbs and is used as flavoring agent in foodstuffs and fragrance in cosmetics. The hepatotoxicity, cytotoxicity, and carcinogenesis of ME have been well documented, and metabolic activation has been suggested to involve in ME-induced toxicities. The objective of this study was to identify chemical identity of interactions of protein with reactive metabolites of ME. Modification of cysteine residues of protein was observed in microsomal incubations and mice after exposure to ME. Three types of protein modification derived from the corresponding epoxide, α,ß-unsaturated aldehyde, and carbonium ion of ME were detected in vitro and in vivo. The protein adduction took place in time- and dose-dependent manners. Dexamethasone, ketoconazole, and l-buthionine sulfoximine increased the protein modification induced by ME, which was proportional to the hepatotoxicity of ME. The findings facilitate the understanding of mechanism action of ME toxicities.