RESUMEN
KIF14 is a mitotic kinesin whose malfunction is associated with cerebral and renal developmental defects and several cancers. Like other kinesins, KIF14 couples ATP hydrolysis and microtubule binding to the generation of mechanical work, but the coupling mechanism between these processes is still not fully clear. Here we report 20 high-resolution (2.7-3.9 Å) cryo-electron microscopy KIF14-microtubule structures with complementary functional assays. Analysis procedures were implemented to separate coexisting conformations of microtubule-bound monomeric and dimeric KIF14 constructs. The data provide a comprehensive view of the microtubule and nucleotide induced KIF14 conformational changes. It shows that: 1) microtubule binding, the nucleotide species, and the neck-linker domain govern the transition between three major conformations of the motor domain; 2) an undocked neck-linker prevents the nucleotide-binding pocket to fully close and dampens ATP hydrolysis; 3) 13 neck-linker residues are required to assume a stable docked conformation; 4) the neck-linker position controls the hydrolysis rather than the nucleotide binding step; 5) the two motor domains of KIF14 dimers adopt distinct conformations when bound to the microtubule; and 6) the formation of the two-heads-bound-state introduces structural changes in both motor domains of KIF14 dimers. These observations provide the structural basis for a coordinated chemo-mechanical kinesin translocation model.
Asunto(s)
Cinesinas/química , Cinesinas/metabolismo , Proteínas Oncogénicas/química , Proteínas Oncogénicas/metabolismo , Adenosina Difosfato/metabolismo , Adenosina Trifosfato/metabolismo , Animales , Sitios de Unión , Microscopía por Crioelectrón , Cinesinas/genética , Ligandos , Ratones , Microtúbulos/química , Microtúbulos/genética , Microtúbulos/metabolismo , Simulación del Acoplamiento Molecular , Proteínas Oncogénicas/genética , Unión Proteica , Conformación Proteica , Dominios ProteicosRESUMEN
Despite the dramatic advances in cancer research over the decades, effective therapeutic strategies are still urgently needed. Increasing evidence indicates that connective tissue growth factor (CTGF), a multifunctional signaling modulator, promotes cancer initiation, progression, and metastasis by regulating cell proliferation, migration, invasion, drug resistance, and epithelial-mesenchymal transition (EMT). CTGF is also involved in the tumor microenvironment in most of the nodes, including angiogenesis, inflammation, and cancer-associated fibroblast (CAF) activation. In this review, we comprehensively discuss the expression of CTGF and its regulation, oncogenic role, clinical relevance, targeting strategies, and therapeutic agents. Herein, we propose that CTGF is a promising cancer therapeutic target that could potentially improve the clinical outcomes of cancer patients.
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Protocolos de Quimioterapia Combinada Antineoplásica/farmacología , Factor de Crecimiento del Tejido Conjuntivo/antagonistas & inhibidores , Neoplasias/tratamiento farmacológico , Proteínas Oncogénicas/antagonistas & inhibidores , Animales , Anticuerpos Monoclonales Humanizados/farmacología , Anticuerpos Monoclonales Humanizados/uso terapéutico , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Movimiento Celular/genética , Proliferación Celular/efectos de los fármacos , Proliferación Celular/genética , Ensayos Clínicos como Asunto , Factor de Crecimiento del Tejido Conjuntivo/genética , Factor de Crecimiento del Tejido Conjuntivo/metabolismo , Curcumina/farmacología , Curcumina/uso terapéutico , Progresión de la Enfermedad , Resistencia a Antineoplásicos/efectos de los fármacos , Resistencia a Antineoplásicos/genética , Transición Epitelial-Mesenquimal/efectos de los fármacos , Transición Epitelial-Mesenquimal/genética , Matriz Extracelular/metabolismo , Regulación Neoplásica de la Expresión Génica , Técnicas de Silenciamiento del Gen , Humanos , Ratones , Neoplasias/genética , Neoplasias/mortalidad , Neoplasias/patología , Proteínas Oncogénicas/genética , Proteínas Oncogénicas/metabolismo , Pronóstico , Transducción de Señal/efectos de los fármacos , Tasa de Supervivencia , Microambiente Tumoral/efectos de los fármacos , Microambiente Tumoral/genética , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Angiogenesis enhances cancer metastasis and progression, however, the roles of transcription regulation in angiogenesis are not fully defined. ZNF322A is an oncogenic zinc-finger transcription factor. Here, we demonstrate a new mechanism of Kras mutation-driven ZNF322A transcriptional activation and elucidate the interplay between ZNF322A and its upstream transcriptional regulators and downstream transcriptional targets in promoting neo-angiogenesis. Methods: Luciferase activity, RT-qPCR and ChIP-qPCR assays were used to examine transcription regulation in cell models. In vitro and in vivo angiogenesis assays were conducted. Immunohistochemistry, Kaplan-Meier method and multivariate Cox regression assays were performed to examine the clinical correlation in tumor specimens from lung cancer patients. Results: We validated that Yin Yang 1 (YY1) upregulated ZNF322A expression through targeting its promoter in the context of Kras mutation. Reconstitution experiments by knocking down YY1 under KrasG13V activation decreased KrasG13V-promoted cancer cell migration, proliferation and ZNF322A promoter activity. Knockdown of YY1 or ZNF322A attenuated angiogenesis in vitro and in vivo. Notably, we validated that ZNF322A upregulated the expression of sonic hedgehog (Shh) gene which encodes a secreted factor that activates pro-angiogenic responses in endothelial cells. Clinically, ZNF322A protein expression positively correlated with Shh and CD31, an endothelial cell marker, in 133 lung cancer patient samples determined using immunohistochemistry analysis. Notably, patients with concordantly high expression of ZNF322A, Shh and CD31 correlated with poor prognosis. Conclusions: These findings highlight the mechanism by which dysregulation of Kras/YY1/ZNF322/Shh transcriptional axis enhances neo-angiogenesis and cancer progression in lung cancer. Therapeutic strategies that target Kras/YY1/ZNF322A/Shh signaling axis may provide new insight on targeted therapy for lung cancer patients.
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Proteínas Hedgehog/genética , Neoplasias Pulmonares/genética , Neovascularización Patológica/genética , Proteínas Oncogénicas/genética , Proteínas Proto-Oncogénicas p21(ras)/genética , Factores de Transcripción/genética , Transcripción Genética/genética , Factor de Transcripción YY1/genética , Animales , Línea Celular Tumoral , Movimiento Celular/genética , Proliferación Celular/genética , Progresión de la Enfermedad , Células Endoteliales/patología , Células Endoteliales de la Vena Umbilical Humana , Humanos , Pulmón/patología , Neoplasias Pulmonares/patología , Ratones Endogámicos C57BL , Ratones Transgénicos , Neovascularización Patológica/patología , Oncogenes/genética , Regiones Promotoras Genéticas/genética , Transducción de Señal/genéticaRESUMEN
The density-regulated protein (DENR) and the malignant T cell-amplified sequence 1 (MCT-1/MCTS1) oncoprotein support noncanonical translation initiation, promote translation reinitiation on a specific set of mRNAs with short upstream reading frames, and regulate ribosome recycling. DENR and MCT-1 form a heterodimer, which binds to the ribosome. We determined the crystal structure of the heterodimer formed by human MCT-1 and the N-terminal domain of DENR at 2.0-Å resolution. The structure of the heterodimer reveals atomic details of the mechanism of DENR and MCT-1 interaction. Four conserved cysteine residues of DENR (C34, C37, C44, C53) form a classical tetrahedral zinc ion-binding site, which preserves the structure of the DENR's MCT-1-binding interface that is essential for the dimerization. Substitution of all four cysteines by alanine abolished a heterodimer formation. Our findings elucidate further the mechanism of regulation of DENR-MCT-1 activities in unconventional translation initiation, reinitiation, and recycling.
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Proteínas de Ciclo Celular/química , Factores Eucarióticos de Iniciación/química , Proteínas Oncogénicas/química , Multimerización de Proteína , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Factores Eucarióticos de Iniciación/genética , Factores Eucarióticos de Iniciación/metabolismo , Humanos , Proteínas Oncogénicas/genética , Proteínas Oncogénicas/metabolismo , Estructura Cuaternaria de ProteínaRESUMEN
Rosmarinic acid (RA), a main phenolic compound contained in rosemary which is used as tea, oil, medicine and so on, has been known to present anti-inflammatory, anti-oxidant and anti-cancer effects. Histone deacetylases (HDACs) are enzymes that play important roles in gene expression by removing the acetyl group from histone. The aberrant expression of HDAC in human tumors is related with the onset of human cancer. Especially, HDAC2, which belongs to HDAC class I composed of HDAC 1, 2, 3 and 8, has been reported to be highly expressed in prostate cancer (PCa) where it downregulates the expression of p53, resulting in an inhibition of apoptosis. The purpose of this study is to investigate the effect of RA in comparison with suberoylanilide hydroxamic acid (SAHA), an HDAC inhibitor used as an anti-cancer agent, on survival and apoptosis of PCa cell lines, PC-3 and DU145, and the expression of HDAC. RA decreased the cell proliferation in cell viability assay, and inhibited the colony formation and tumor spheroid formation. Additionally, RA induced early- and late-stage apoptosis of PC-3 and DU145 cells in Annexin V assay and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, respectively. In western blot analysis, RA inhibited the expression of HDAC2, as SAHA did. Proliferating cell nuclear antigen (PCNA), cyclin D1 and cyclin E1 were downregulated by RA, whereas p21 was upregulated. In addition, RA modulated the protein expression of intrinsic mitochondrial apoptotic pathway-related genes, such as Bax, Bcl-2, caspase-3 and poly (ADP-ribose) polymerase 1 (PARP-1) (cleaved) via the upregulation of p53 derived from HDAC2 downregulation, leading to the increased apoptosis of PC-3 and DU145 cells. Taken together, treatment of RA to PCa cell lines inhibits the cell survival and induces cell apoptosis, and it can be used as a novel therapeutic agent toward PCa.
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Apoptosis/efectos de los fármacos , Puntos de Control del Ciclo Celular/efectos de los fármacos , Cinamatos/análisis , Depsidos/análisis , Histona Desacetilasa 2/metabolismo , Rosmarinus/química , Anexina A5 , Caspasa 3/genética , Caspasa 3/metabolismo , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Ciclina D1/genética , Ciclina D1/metabolismo , Ciclina E/genética , Ciclina E/metabolismo , Fragmentación del ADN/efectos de los fármacos , Regulación de la Expresión Génica , Histona Desacetilasa 2/genética , Inhibidores de Histona Desacetilasas/farmacología , Histona Desacetilasas/metabolismo , Humanos , Etiquetado Corte-Fin in Situ , Masculino , Proteínas Oncogénicas/genética , Proteínas Oncogénicas/metabolismo , Poli(ADP-Ribosa) Polimerasa-1/genética , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Antígeno Nuclear de Célula en Proliferación/genética , Antígeno Nuclear de Célula en Proliferación/metabolismo , Neoplasias de la Próstata/metabolismo , Transducción de Señal , Tés de Hierbas , Tés Medicinales , Vorinostat/análisis , Proteína X Asociada a bcl-2/genética , Proteína X Asociada a bcl-2/metabolismo , Ácido RosmarínicoRESUMEN
The N7-methylguanosine cap is added in the nucleus early in gene transcription and is a defining feature of eukaryotic mRNAs. Mammalian cells also possess cytoplasmic machinery for restoring the cap at uncapped or partially degraded RNA 5' ends. Central to both pathways is capping enzyme (CE) (RNA guanylyltransferase and 5'-phosphatase (RNGTT)), a bifunctional, nuclear and cytoplasmic enzyme. CE is recruited to the cytoplasmic capping complex by binding of a C-terminal proline-rich sequence to the third Src homology 3 (SH3) domain of NCK adapter protein 1 (NCK1). To gain broader insight into the cellular context of cytoplasmic recapping, here we identified the protein interactome of cytoplasmic CE in human U2OS cells through two complementary approaches: chemical cross-linking and recovery with cytoplasmic CE and protein screening with proximity-dependent biotin identification (BioID). This strategy unexpectedly identified 66 proteins, 52 of which are RNA-binding proteins. We found that CE interacts with several of these proteins independently of RNA, mediated by sequences within its N-terminal triphosphatase domain, and we present a model describing how CE-binding proteins may function in defining recapping targets. This analysis also revealed that CE is a client protein of heat shock protein 90 (HSP90). Nuclear and cytoplasmic CEs were exquisitely sensitive to inhibition of HSP90, with both forms declining significantly following treatment with each of several HSP90 inhibitors. Importantly, steady-state levels of capped mRNAs decreased in cells treated with the HSP90 inhibitor geldanamycin, raising the possibility that the cytotoxic effect of these drugs may partially be due to a general reduction in translatable mRNAs.
Asunto(s)
Citoplasma/enzimología , Proteínas HSP90 de Choque Térmico/metabolismo , Nucleotidiltransferasas/metabolismo , Monoéster Fosfórico Hidrolasas/metabolismo , Proteínas de Unión al ARN/metabolismo , Proteínas Adaptadoras Transductoras de Señales/genética , Proteínas Adaptadoras Transductoras de Señales/metabolismo , Citoplasma/genética , Proteínas HSP90 de Choque Térmico/genética , Humanos , Nucleotidiltransferasas/genética , Proteínas Oncogénicas/genética , Proteínas Oncogénicas/metabolismo , Monoéster Fosfórico Hidrolasas/genética , Unión Proteica , Caperuzas de ARN/genética , Caperuzas de ARN/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Proteínas de Unión al ARN/genéticaRESUMEN
PURPOSE: Deregulated MYC drives oncogenesis in many tissues yet direct pharmacologic inhibition has proven difficult. MYC coordinately regulates polyamine homeostasis as these essential cations support MYC functions, and drugs that antagonize polyamine sufficiency have synthetic-lethal interactions with MYC Neuroblastoma is a lethal tumor in which the MYC homologue MYCN, and ODC1, the rate-limiting enzyme in polyamine synthesis, are frequently deregulated so we tested optimized polyamine depletion regimens for activity against neuroblastoma. EXPERIMENTAL DESIGN: We used complementary transgenic and xenograft-bearing neuroblastoma models to assess polyamine antagonists. We investigated difluoromethylornithine (DFMO; an inhibitor of Odc, the rate-limiting enzyme in polyamine synthesis), SAM486 (an inhibitor of Amd1, the second rate-limiting enzyme), and celecoxib (an inducer of Sat1 and polyamine catabolism) in both the preemptive setting and in the treatment of established tumors. In vitro assays were performed to identify mechanisms of activity. RESULTS: An optimized polyamine antagonist regimen using DFMO and SAM486 to inhibit both rate-limiting enzymes in polyamine synthesis potently blocked neuroblastoma initiation in transgenic mice, underscoring the requirement for polyamines in MYC-driven oncogenesis. Furthermore, the combination of DFMO with celecoxib was found to be highly active, alone, and combined with numerous chemotherapy regimens, in regressing established tumors in both models, including tumors harboring highest risk genetic lesions such as MYCN amplification, ALK mutation, and TP53 mutation with multidrug resistance. CONCLUSIONS: Given the broad preclinical activity demonstrated by polyamine antagonist regimens across diverse in vivo models, clinical investigation of such approaches in neuroblastoma and potentially other MYC-driven tumors is warranted. Clin Cancer Res; 22(17); 4391-404. ©2016 AACR.
Asunto(s)
Antineoplásicos/farmacología , Transformación Celular Neoplásica/efectos de los fármacos , Neuroblastoma/etiología , Neuroblastoma/patología , Poliaminas/antagonistas & inhibidores , Animales , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapéutico , Celecoxib/farmacología , Línea Celular Tumoral , Modelos Animales de Enfermedad , Progresión de la Enfermedad , Evaluación Preclínica de Medicamentos , Sinergismo Farmacológico , Eflornitina/farmacología , Genes myc , Homeostasis/efectos de los fármacos , Humanos , Ratones , Ratones Transgénicos , Neuroblastoma/tratamiento farmacológico , Neuroblastoma/mortalidad , Proteínas Oncogénicas/genética , Proteínas Oncogénicas/metabolismo , Poliaminas/metabolismo , Resultado del Tratamiento , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Plantaginaceae, a popular traditional Chinese medicine, has long been used for treating various diseases from common cold to cancer. Linalool is one of the biologically active compounds that can be isolated from Plantaginaceae. Most of the commonly used cytotoxic anticancer drugs have been shown to induce apoptosis in susceptible tumor cells. However, the signaling pathway for apoptosis remains undefined. In this study, the cytotoxic effect of linalool on human cancer cell lines was investigated. Water-soluble tetrazolium salts (WST-1) based colorimetric cellular cytotoxicity assay, was used to test the cytotoxic ability of linalool against U937 and HeLa cells, and flow cytometry (FCM) and genechip analysis were used to investigate the possible mechanism of apoptosis. These results demonstrated that linalool exhibited a good cytotoxic effect on U937 and HeLa cells, with the IC50 value of 2.59 and 11.02 µM, respectively, compared with 5-FU with values of 4.86 and 12.31 µM, respectively. After treating U937 cells with linalool for 6 h, we found an increased sub-G1 peak and a dose-dependent phenomenon, whereby these cells were arrested at the G0/G1 phase. Furthermore, by using genechip analysis, we observed that linalool can promote p53, p21, p27, p16, and p18 gene expression. Therefore, this study verified that linalool can arrest the cell cycle of U937 cells at the G0/G1 phase and can arrest the cell cycle of HeLa cells at the G2/M phase. Its mechanism facilitates the expression of the cyclin-dependent kinases inhibitors (CDKIs) p53, p21, p27, p16, and p18, as well as the non-expression of cyclin-dependent kinases (CDKs) activity.
Asunto(s)
Apoptosis/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Proteínas Inhibidoras de las Quinasas Dependientes de la Ciclina/metabolismo , Leucemia/metabolismo , Monoterpenos/toxicidad , Neoplasias del Cuello Uterino/metabolismo , Monoterpenos Acíclicos , Proteínas Inhibidoras de las Quinasas Dependientes de la Ciclina/genética , Femenino , Células HeLa , Humanos , Proteínas Oncogénicas/genética , Proteínas Oncogénicas/metabolismo , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
NUT midline carcinoma is a rare entity arising primarily in the midline of teenagers and young adults. Genomically, it is associated with a translocation involving a nuclear protein in testis (NUT) gene with other genes, most commonly, the BRD4 gene. The resultant is a partial or near total block in differentiation of tumor cells into mature squamous elements. Such tumors are resistant to conventional therapy with a reported mean survival at less than 1 year. In this study, we investigated two cases with genomic confirmation as NUT midline carcinoma by morphoproteomic analysis using immunohistochemical antibodies. Our results showed overexpression, largely in the undifferentiated cells of the tumors of: 1) Stemness marker, SRY (sex determining region Y)-box 2 (Sox2); 2) Constitutive activation of the mTORC2 pathway with expression of total insulin-like growth factor-1 receptor (IGF-1R[Tyr1165/1166]), and nuclear p-mTOR (Ser 2448) and p-Akt (Ser 473); and 3) c-Myc, silent mating type information regulation 2 homolog 1 (Sirt1) and histone methyltransferase enhancer of Zeste, Drosophila, homolog 2 (EZH2) as molecular impediments to differentiation. These data were analyzed through the use of QIAGEN's Ingenuity(®) Pathway Analysis (IPA(®), QIAGEN Redwood City, www.qiagen.com/ingenuity). The results established the interconnection of these pathways and molecules, and identified several pharmacogenomic agents--melatonin, metformin, vorinostat, curcumin, and sulforaphane--that have the potential to remove the block in differentiation and lead to the establishment of a more benign form of NUT midline carcinoma.
Asunto(s)
Carcinoma/genética , Carcinoma/patología , Neoplasias del Mediastino/genética , Neoplasias del Mediastino/patología , Proteínas Nucleares/genética , Proteínas Oncogénicas/genética , Adolescente , Adulto , Carcinoma/metabolismo , Carcinoma/terapia , Proteína Potenciadora del Homólogo Zeste 2 , Femenino , Humanos , Inmunohistoquímica , Hibridación Fluorescente in Situ , Isotiocianatos/farmacología , Masculino , Neoplasias del Mediastino/metabolismo , Neoplasias del Mediastino/terapia , Metformina/farmacología , Terapia Molecular Dirigida/métodos , Proteínas de Neoplasias , Proteínas Nucleares/metabolismo , Proteínas Oncogénicas/metabolismo , Complejo Represivo Polycomb 2/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Sirtuina 1/metabolismo , Sulfóxidos , Serina-Treonina Quinasas TOR/metabolismoRESUMEN
The herb Momordica cochinchinensis has been used for a variety of purposes, and been shown to have anticancer properties. The present study assessed the potency and the underlying mechanisms of action of the ethyl acetate extract of seeds of Momordica cochinchinensis (ESMC2) on breast cancer cells. Therefore, the effects of ESMC2 on the cell viability, cell cycle and apoptosis of MDAMB231 cells were investigated. The results showed that ESMC2 exerted a marked growth inhibitory effect on the cells. Cell cycle arrest in G2 phase following treatment with ESMC2 was associated with a marked increase in the protein levels of cyclin B1, cyclin E and cyclin-dependent kinase 1 and a decrease in cyclin D1 expression. In addition, ESMC2 dosedependently induced cell apoptosis, which was mediated via upregulation of the apoptosis-associated proteins p53, B-cell lymphoma 2 (Bcl2)associated X protein, Bcl-2 homologous antagonist killer and Bcl-2-associated death promoter expression, as well as downregulation of nuclear factor kappa B, Bcl2 and myeloid cell leukemia1. Furthermore, the activation of extracellular signal-regulated kinase 1/2, p38, c-Jun N-terminal kinase (JNK) and Akt phosphorylation were decreased by ESMC2 in a dosedependent manner, indicating that ESMC2 exerted its effects via the mitogen-activated protein kinase/JNK pathway. Furthermore, nude mouse xenotransplant models were used to evaluate the tumor growth inhibitory effects of ESMC2. The possible chemical components of ESMC2 were analyzed by gas chromatography-mass spectrometry, and 12 compounds were detected from the major peaks based on the similarity index with entries of a compound database. The results of the present study may aid in the development of novel therapies for breast cancer.
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Antineoplásicos Fitogénicos/farmacología , Apoptosis/efectos de los fármacos , Neoplasias de la Mama/patología , Puntos de Control del Ciclo Celular/efectos de los fármacos , Momordica/química , Extractos Vegetales/química , Proteína Quinasa CDC2 , Línea Celular Tumoral/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Ciclina B1/genética , Ciclina B1/metabolismo , Ciclina D1/genética , Ciclina D1/metabolismo , Ciclina E/genética , Ciclina E/metabolismo , Quinasas Ciclina-Dependientes/genética , Quinasas Ciclina-Dependientes/metabolismo , Relación Dosis-Respuesta a Droga , Regulación hacia Abajo , Fase G2/efectos de los fármacos , Cromatografía de Gases y Espectrometría de Masas , Humanos , Proteínas Quinasas JNK Activadas por Mitógenos/genética , Proteínas Quinasas JNK Activadas por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/genética , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/genética , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , FN-kappa B/genética , FN-kappa B/metabolismo , Proteínas Oncogénicas/genética , Proteínas Oncogénicas/metabolismo , Fosforilación , Semillas/química , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo , Proteína X Asociada a bcl-2/genética , Proteína X Asociada a bcl-2/metabolismo , Proteínas Quinasas p38 Activadas por Mitógenos/genética , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
Protein phosphatase 2A (PP2A) is a serine/threonine phosphatase that plays a significant role in mitotic progression and cellular responses to DNA damage. While traditionally viewed as a tumor suppressor, inhibition of PP2A has recently come to attention as a novel therapeutic means of driving senescent cancer cells into mitosis and promoting cell death via mitotic catastrophe. These findings have been corroborated in numerous studies utilizing naturally produced compounds that selectively inhibit PP2A. To overcome the known human toxicities associated with these compounds, a water-soluble small molecule inhibitor, LB100, was recently developed to competitively inhibit the PP2A protein. This review summarizes the pre-clinical studies to date that have demonstrated the anti-cancer activity of LB100 via its chemo- and radio-sensitizing properties. These studies demonstrate the tremendous therapeutic potential of LB100 in a variety of cancer types. The results of an ongoing phase 1 trial are eagerly anticipated.
Asunto(s)
Antineoplásicos/farmacología , Compuestos Bicíclicos Heterocíclicos con Puentes/farmacología , Inhibidores Enzimáticos/farmacología , Piperazinas/farmacología , Proteína Fosfatasa 2/antagonistas & inhibidores , Fármacos Sensibilizantes a Radiaciones/farmacología , Animales , Apoptosis/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Evaluación Preclínica de Medicamentos , Resistencia a Antineoplásicos , Humanos , Mitosis/efectos de los fármacos , Mitosis/genética , Neoplasias/tratamiento farmacológico , Neoplasias/genética , Neoplasias/metabolismo , Proteínas Oncogénicas/antagonistas & inhibidores , Proteínas Oncogénicas/genética , Proteínas Oncogénicas/metabolismo , Proteína Fosfatasa 2/genética , Proteína Fosfatasa 2/metabolismo , Proteína Tumoral Controlada Traslacionalmente 1 , Proteínas Supresoras de Tumor/antagonistas & inhibidores , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo , Vía de Señalización Wnt/efectos de los fármacosRESUMEN
DJ-1 constitutes a ubiquitously expressed, oxidative stress-responsive protein with multiple functions. DJ-1 emerged as a candidate from our previous proteome analysis investigating alterations in the hypothalamus in three mouse strains differing in their susceptibility to diet-induced obesity (DIO). Validation studies demonstrated a high-fat diet (HFD)-induced shift in the DJ-1 isoform pattern in the hypothalamus and several other tissues of mice. Others found HFD-induced alterations in DJ-1 protein abundance in adipose tissue and pancreatic islets in wild-type rodents. Here, we investigated the gene-diet interaction by challenging Dj-1(-/-) mice with a HFD. We demonstrate that the development of diet-induced obesity (DIO) Dj-1(-/-) mice is according to wild-type mice with the exception of transient higher gains in fat mass at the expense of lean mass after 14 weeks of feeding.
Asunto(s)
Dieta Alta en Grasa/efectos adversos , Obesidad/fisiopatología , Proteínas Oncogénicas/deficiencia , Peroxirredoxinas/deficiencia , Tejido Adiposo/metabolismo , Animales , Glucemia/metabolismo , Calorimetría Indirecta , Ingestión de Energía , Femenino , Prueba de Tolerancia a la Glucosa , Hipotálamo/metabolismo , Insulina/sangre , Islotes Pancreáticos/metabolismo , Leptina/administración & dosificación , Leptina/sangre , Masculino , Ratones , Ratones Noqueados , Proteínas Oncogénicas/genética , Estrés Oxidativo , Peroxirredoxinas/genética , Proteína Desglicasa DJ-1 , Reacción en Cadena en Tiempo Real de la Polimerasa , Factor de Transcripción STAT3/genética , Factor de Transcripción STAT3/metabolismo , Análisis de Secuencia de ARNRESUMEN
Neuroblastoma, a childhood cancer that originates from neural crest-derived cells, is the most common deadly solid tumor of infancy. Amplification of the MYCN oncogene, which occurs in approximately 20-25% of human neuroblastomas, is the most prominent genetic marker of high-stage disease. The availability of valid preclinical in vivo models is a prerequisite to develop novel targeted therapies. We here report on the generation of transgenic mice with Cre-conditional induction of MYCN in dopamine ß-hydroxylase-expressing cells, termed LSL-MYCN;Dbh-iCre. These mice develop neuroblastic tumors with an incidence of >75%, regardless of strain background. Molecular profiling of tumors revealed upregulation of the MYCN-dependent miR-17-92 cluster as well as expression of neuroblastoma marker genes, including tyrosine hydroxylase and the neural cell adhesion molecule 1. Gene set enrichment analyses demonstrated significant correlation with MYC-associated expression patterns. Array comparative genome hybridization showed that chromosomal aberrations in LSL-MYCN;Dbh-iCre tumors were syntenic to those observed in human neuroblastomas. Treatment of a cell line established from a tumor derived from a LSL-MYCN;Dbh-iCre mouse with JQ1 or MLN8237 reduced cell viability and demonstrated oncogene addiction to MYCN. Here we report establishment of the first Cre-conditional human MYCN-driven mouse model for neuroblastoma that closely recapitulates the human disease with respect to tumor localization, histology, marker expression and genomic make up. This mouse model is a valuable tool for further functional studies and to assess the effect of targeted therapies.
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Evaluación Preclínica de Medicamentos/métodos , Integrasas/genética , Neuroblastoma/patología , Proteínas Nucleares/genética , Proteínas Oncogénicas/genética , Regiones Promotoras Genéticas , Transgenes , Animales , Modelos Animales de Enfermedad , Femenino , Perfilación de la Expresión Génica , Humanos , Ratones , Ratones Endogámicos C57BL , Ratones Desnudos , Ratones Transgénicos , Análisis por Micromatrices , Proteína Proto-Oncogénica N-Myc , Neuroblastoma/genética , Células Tumorales CultivadasAsunto(s)
Proteínas de Fase Aguda/metabolismo , Encefalomielitis Autoinmune Experimental/tratamiento farmacológico , Encefalomielitis Autoinmune Experimental/metabolismo , Lipocalinas/metabolismo , Esclerosis Múltiple/tratamiento farmacológico , Esclerosis Múltiple/metabolismo , Proteínas Oncogénicas/metabolismo , Preparaciones de Plantas/farmacología , Proteínas de Fase Aguda/genética , Animales , Antiinflamatorios/farmacología , Modelos Animales de Enfermedad , Regulación hacia Abajo/efectos de los fármacos , Encefalomielitis Autoinmune Experimental/genética , Femenino , Factores Inmunológicos/farmacología , Lipocalina 2 , Lipocalinas/genética , Ratones , Ratones Endogámicos C57BL , Esclerosis Múltiple/genética , Proteínas Oncogénicas/genética , Fitoterapia , Médula Espinal/efectos de los fármacos , Médula Espinal/metabolismo , Médula Espinal/patologíaRESUMEN
Neuroblastoma is one of the most common solid tumors and accounts for â¼ 15% of all the cancer related deaths in the children. Despite the standard therapy for advanced disease including chemotherapy, surgery, and radiation, the mortality rate remains high for these patients. Hence, novel therapeutic agents are desperately needed. Here we examined the anticancer activity of a novel plant-derived compound, sparstolonin B (SsnB; 8,5'-dihydroxy-4-phenyl-5,2'-oxidoisocoumarin) using neuroblastoma cell lines of different genetics. SsnB was recently isolated from an aquatic Chinese herb, Sparganium stoloniferum, and tubers of this herb have been used in traditional Chinese medicine for the treatment of several inflammatory diseases and cancers. Our cell viability and morphological analysis indicated that SsnB at 10 µM concentration significantly inhibited the growth of both N-myc amplified (SK-N-BE(2), NGP, and IMR-32 cells) and N-myc nonamplified (SH-SY5Y and SKNF-1 cells) neuroblastoma cells. The flow cytometric analyses suggested that SsnB arrests the cell cycle progression at G2-M phase in all neuroblastoma cell lines tested. Exposure of SsnB inhibited the compact spheroid formation and reduced the tumorigenicity of SH-SY5Y cells and SK-N-BE(2) cells in in vitro 3-D cell culture assays (anchorage-independent colony formation assay and hanging drop assay). SsnB lowers the cellular level of glutathione (GSH), increases generation of reactive oxygen species and activates the cleavage of caspase-3 whereas co-incubation of a GSH precursor, N-acetylcysteine, along with SsnB attenuates the inhibitory effects of SsnB and increases the neuroblastoma cell viability. Our results for the first time demonstrate that SsnB possesses anticancer activity indicating that SsnB-induced reactive oxygen species generation promotes apoptotic cell death in neuroblastoma cells of different genetic background. Thus these data suggest that SsnB can be a promising drug candidate in neuroblastoma therapy.
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Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Compuestos Heterocíclicos de 4 o más Anillos/farmacología , Neuroblastoma/patología , Proteínas Nucleares/genética , Proteínas Oncogénicas/genética , Caspasa 3/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Amplificación de Genes , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Proteína Proto-Oncogénica N-Myc , Neuroblastoma/tratamiento farmacológico , Neuroblastoma/genética , Especies Reactivas de Oxígeno/metabolismoRESUMEN
Competition for iron influences host-pathogen interactions. Pathogens secrete small iron-binding moieties, siderophores, to acquire host iron. In response, the host secretes siderophore-binding proteins, such as lipocalin 24p3, which limit siderophore-mediated iron import into bacteria. Mammals produce 2,5-dihydroxy benzoic acid, a compound that resembles a bacterial siderophore. Our data suggest that bacteria use both mammalian and bacterial siderophores. In support of this idea, supplementation with mammalian siderophore enhances bacterial growth in vitro. In addition, mice lacking the mammalian siderophore resist E. coli infection. Finally, we show that the host responds to infection by suppressing siderophore synthesis while up-regulating lipocalin 24p3 expression via TLR signaling. Thus, reciprocal regulation of 24p3 and mammalian siderophore is a protective mechanism limiting microbial access to iron.
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Infecciones Bacterianas/inmunología , Gentisatos/inmunología , Hidroxibutirato Deshidrogenasa/inmunología , Inmunidad Innata/inmunología , Sideróforos/inmunología , Proteínas de Fase Aguda/genética , Proteínas de Fase Aguda/inmunología , Proteínas de Fase Aguda/metabolismo , Animales , Infecciones Bacterianas/genética , Infecciones Bacterianas/metabolismo , Proteínas de la Membrana Bacteriana Externa/genética , Proteínas de la Membrana Bacteriana Externa/inmunología , Proteínas de la Membrana Bacteriana Externa/metabolismo , Candida albicans/inmunología , Candida albicans/fisiología , Candidiasis/genética , Candidiasis/inmunología , Candidiasis/metabolismo , Proteínas Portadoras/genética , Proteínas Portadoras/inmunología , Proteínas Portadoras/metabolismo , Línea Celular , Enterobactina/inmunología , Enterobactina/metabolismo , Escherichia coli/genética , Escherichia coli/inmunología , Escherichia coli/fisiología , Femenino , Gentisatos/metabolismo , Hidroxibutirato Deshidrogenasa/genética , Hidroxibutirato Deshidrogenasa/metabolismo , Inmunidad Innata/genética , Immunoblotting , Estimación de Kaplan-Meier , Lipocalina 2 , Lipocalinas/genética , Lipocalinas/inmunología , Lipocalinas/metabolismo , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Mutación/inmunología , Proteínas Oncogénicas/genética , Proteínas Oncogénicas/inmunología , Proteínas Oncogénicas/metabolismo , Factor 1 de Unión al Dominio 1 de Regulación Positiva , Interferencia de ARN , Receptores de Superficie Celular/genética , Receptores de Superficie Celular/inmunología , Receptores de Superficie Celular/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Sideróforos/metabolismo , Staphylococcus aureus/inmunologíaRESUMEN
In breast cancer, the cytokine tumor necrosis factor-α (TNF-α) induces cell invasion, although the molecular basis of it has not been clearly elucidated. In this study, we investigated the role of daidzein in regulating TNF-α induced cell invasion and the underlying molecular mechanisms. Daidzein inhibited TNF-α induced cellular migration and invasion in estrogen receptor (ER) negative MCF10DCIS.com human breast cancer cells. TNF-α activated Hedgehog (Hh) signaling by enhancing Gli1 nuclear translocation and transcriptional activity, which resulted in increased invasiveness; these effects were blocked by daidzein and the Hh signaling inhibitors, cyclopamine and vismodegib. Moreover, these compounds suppressed TNF-α induced matrix metalloproteinase (MMP)-9 mRNA expression and activity. Taken together, mammary tumor cell invasiveness was stimulated by TNF-α induced activation of Hh signaling; these effects were abrogated by daidzein, which suppressed Gli1 activation, thereby inhibiting migration and invasion.
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Neoplasias de la Mama/fisiopatología , Glycine max/química , Erizos/metabolismo , Isoflavonas/farmacología , Proteínas Oncogénicas/metabolismo , Extractos Vegetales/farmacología , Transducción de Señal/efectos de los fármacos , Transactivadores/metabolismo , Factor de Necrosis Tumoral alfa/metabolismo , Animales , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/genética , Neoplasias de la Mama/patología , Línea Celular Tumoral , Movimiento Celular/efectos de los fármacos , Nucléolo Celular/genética , Nucléolo Celular/metabolismo , Regulación hacia Abajo/efectos de los fármacos , Femenino , Erizos/genética , Humanos , Invasividad Neoplásica/genética , Proteínas Oncogénicas/genética , Transactivadores/genética , Factor de Necrosis Tumoral alfa/genética , Proteína con Dedos de Zinc GLI1RESUMEN
Genetic and environmental factors mediate via different physiological and molecular processes a shifted energy balance leading to overweight and obesity. To get insights into the underlying processes involved in energy intake and weight gain, we compared hypothalamic tissue of mice kept on a high-fat or control diet for 10 days by a proteomic approach. Using two-dimensional difference gel electrophoresis in combination with LC-MS/MS, we observed significant abundance changes in 15 protein spots. One isoform of the protein DJ-1 was elevated in the high-fat diet group in three different mouse strains SWR/J, C57BL/6N, and AKR/J analyzed. Large-scale validation of DJ-1 isoforms in individual samples and tissues confirmed a shift in the pattern of DJ-1 isoforms toward more acidic isoforms in several brain and peripheral tissues after feeding a high-fat diet for 10 days. The identification of oxidation of cysteine 106 as well as 2-succinyl modification of the same residue by mass spectrometry not only explains the isoelectric shift of DJ-1 but also links our results to similar shifts of DJ-1 observed in neurodegenerative disease states under oxidative stress. We hypothesize that DJ-1 is a common physiological sensor involved in both nutrition-induced effects and neurodegenerative disease states.
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Dieta Alta en Grasa/métodos , Hipotálamo/metabolismo , Proteínas Oncogénicas/metabolismo , Peroxirredoxinas/metabolismo , Proteómica/métodos , Animales , Western Blotting , Cromatografía Liquida , Dieta Alta en Grasa/efectos adversos , Electroforesis en Gel Bidimensional , Punto Isoeléctrico , Masculino , Espectrometría de Masas/métodos , Ratones Endogámicos AKR , Ratones Endogámicos C57BL , Ratones Endogámicos , Obesidad/etiología , Obesidad/metabolismo , Proteínas Oncogénicas/química , Proteínas Oncogénicas/genética , Sobrepeso/etiología , Sobrepeso/metabolismo , Enfermedad de Parkinson/metabolismo , Peroxirredoxinas/química , Peroxirredoxinas/genética , Proteína Desglicasa DJ-1 , Isoformas de Proteínas/química , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Especificidad de la EspecieRESUMEN
KIF14 (kinesin family member 14) is a mitotic kinesin and an important oncogene in several cancers. Tumor KIF14 expression levels are independently predictive of poor outcome, and in cancer cells KIF14 can modulate metastatic behavior by maintaining appropriate levels of cell adhesion and migration proteins at the cell membrane. Thus KIF14 is an exciting potential therapeutic target. Understanding KIF14's regulation in cancer cells is crucial to the development of effective and selective therapies to block its tumorigenic function(s). We previously determined that close to 30% of serous ovarian cancers (OvCa tumors) exhibit low-level genomic gain, indicating one mechanism of KIF14 overexpression in tumors. We now report on transcriptional and epigenetic regulation of KIF14. Through promoter deletion analyses, we identified one cis-regulatory region containing binding sites for Sp1, HSF1 and YY1. siRNA-mediated knockdown of these transcription factors demonstrated endogenous regulation of KIF14 overexpression by Sp1 and YY1, but not HSF1. ChIP experiments confirmed an enrichment of both Sp1 and YY1 binding to the endogenous KIF14 promoter in OvCa cell lines with high KIF14 expression. A strong correlation was seen in primary serous OvCa tumors between Sp1, YY1 and KIF14 expression, further evidence that these transcription factors are important players in KIF14 overexpression. Hypomethylation patterns were observed in primary serous OvCa tumors, suggesting a minor role for promoter methylation in the control of KIF14 gene expression. miRNA expression analysis determined that miR-93, miR-144 and miR-382 had significantly lower levels of expression in primary serous OvCa tumors than normal tissues; treatment of an OvCa cell line with miRNA mimics and inhibitors specifically modulated KIF14 mRNA levels, pointing to potential novel mechanisms of KIF14 overexpression in primary tumors. Our findings reveal multiple mechanisms of KIF14 upregulation in cancer cells, offering new targets for therapeutic interventions to reduce KIF14 in tumors, aiming at improved prognosis.
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Epigénesis Genética , Regulación de la Expresión Génica , Cinesinas/metabolismo , Proteínas Oncogénicas/metabolismo , Neoplasias Ováricas/metabolismo , Transcripción Genética , Línea Celular Tumoral , Inmunoprecipitación de Cromatina , Metilación de ADN , Femenino , Eliminación de Gen , Células HeLa , Humanos , Cinesinas/genética , Mitosis , Metástasis de la Neoplasia , Proteínas Oncogénicas/genética , Neoplasias Ováricas/genética , Pronóstico , Regiones Promotoras Genéticas , ARN Interferente Pequeño/metabolismo , Factor de Transcripción Sp1/metabolismo , Regulación hacia Arriba , Factor de Transcripción YY1/metabolismoRESUMEN
BACKGROUND: Once a promising drug target is identified, the steps to actually discover and optimize a drug are diverse and challenging. OBJECTIVE: The goal of this study was to provide a road map to navigate drug discovery. METHODS: Review general steps for drug discovery and provide illustrating references. RESULTS: A number of approaches are available to enhance and accelerate target identification and validation. Consideration of a variety of potential mechanisms of action of potential drugs can guide discovery efforts. The hit to lead stage may involve techniques such as high-throughput screening, fragment-based screening, and structure-based design, with informatics playing an ever-increasing role. Biologically relevant screening models are discussed, including cell lines, 3-dimensional culture, and in vivo screening. The process of enabling human studies for an investigational drug is also discussed. CONCLUSIONS: Drug discovery is a complex process that has significantly evolved in recent years.