RESUMEN
Cyclin-dependent kinases (CDKs) play a key regulating role in the cell cycle, which is almost universally altered in cancer, leading to sustained proliferation. Early pan-CDK inhibitors showed poor results in clinical trials for solid malignancies, as the lack of selectivity produced significant toxicity. The production of more selective inhibitors led to significant developments in cancer therapy, as CDK4/6 inhibitors in combination with endocrine therapy changed the landscape of the treatment of hormone-receptor positive (HR +) metastatic breast cancer. Recently, Trilaciclib demonstrated benefits regarding hematological toxicity compared to placebo when administered in combination with chemotherapy in small cell lung cancer. Newer agents, such as SY-5609, a selective CDK7 inhibitor, have also shown promising results in early clinical trials. In this paper, we review the data from clinical trials of CDK inhibitors in solid tumors, either as a monotherapy or in combination with other agents, with an emphasis on novel agents and potential new indications for this drug class.
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Ensayos Clínicos como Asunto , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Neoplasias/tratamiento farmacológico , HumanosRESUMEN
INTRODUCTION AND OBJECTIVES: HBV covalently closed circular (ccc) DNA is the key player in viral persistence and an important predictive biomarker for hepatitis relapse. Precise quantification of intracellular cccDNA is challenging because cccDNA is present in very low levels in hepatocytes, where it also co-exists with a large excess amount of relaxed circular (rc) DNA. We aimed to develop a highly sensitive cccDNA detection method for cccDNA quantification by digital PCR (dPCR). PATIENTS OR MATERIALS AND METHODS: A standard plasmid containing the whole HBV genome in the closed circular conformation was employed to characterize the performance of dPCR. rcDNA in the growth medium of HBV-producing HepAD38 cells was used as a matrix for cccDNA detection. Intrahepatic cccDNA measurement by dPCR and qPCR was performed to determine the correlation of the analysis results for the two methods. RESULTS: The limit of detection (LOD) of the cccDNA dPCR was 1.05copy/µl, and the linear range of detection was 1.02×104copies/µl, achieving a dynamic detection range of 104-fold. cccDNA measurement using excess rcDNA as the matrix did not reveal false-positive detection, indicating that dPCR was highly specific. In the HepAD38 cells, the cccDNA levels measured by dPCR were highly correlated with those measured by qPCR but had a higher sensitivity. The CDK inhibitor AZD-5438 was found to block intracellular cccDNA synthesis. CONCLUSIONS: Dpcr greatly improved the sensitivity and specificity of cccDNA detection. Host CDK activities are likely required for cccDNA synthesis. dPCR can potentially be applied for drug screening for effective cccDNA inhibitors.
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Quinasas Ciclina-Dependientes/metabolismo , ADN Circular/análisis , ADN Viral/análisis , Virus de la Hepatitis B/genética , Hepatocitos/metabolismo , Reacción en Cadena de la Polimerasa/métodos , Línea Celular , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , ADN Circular/biosíntesis , ADN Viral/biosíntesis , Virus de la Hepatitis B/efectos de los fármacos , Hepatocitos/efectos de los fármacos , Humanos , Imidazoles/farmacología , Pirimidinas/farmacologíaRESUMEN
Two mononuclear copper(II) compounds, [Cu(isad)(H2O)Cl]Cl 1 and [Cu(isah)(H2O)Cl]Cl 2, and its corresponding heterobinuclear species containing also platinum(II), [CuCl(isad)Pt(NH3)Cl2] 3 and [CuCl(isah)Pt(NH3)Cl2] 4 (where isad and isah are oxindolimine ligands, (E)-3-(2-(3-aminopropylamino)ethylimino)indolin-2-one, and (E)-3-(3-amino-2-hydroxypropylimino)indolin-2-one, respectively), have been previously synthesized and characterized by different spectroscopic techniques in our laboratory. Cytotoxicity assays performed with B16F10 murine cancer cells, and MES-SA human uterine sarcoma cells, showed IC50 values lower or in the same order of cisplatin. Herein, in order to better elucidate their probable modes of action, possible interaction and damage to DNA, as well as their effect on the activity of crucial proteins were verified. Both mononuclear complexes and the binuclear compound 4 displayed a significant cleavage activity toward plasmid DNA, while compound 3 tends to protect DNA from oxidative damage, avoiding degradation. Complementary experiments indicated a significant inhibition activity toward cyclin-dependent kinase (CDK1/cyclinB) activity in the phosphorylation of histone H1, and only moderate inhibition concerning alkaline phosphatase. Results also revealed that the reactivity is reliant on the ligand structure and on the nature of the metal present, in a synergistic effect. Simulation studies complemented and supported our results, indicating different bindings of the binuclear compounds to DNA. Therefore, the verified cytotoxicity of these complexes comprises multiple modes of action, including modification of DNA conformation, scission of DNA strands by reactive oxygen species, and inhibition of selected proteins that are crucial to the cellular cycle.
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Fosfatasa Alcalina/antagonistas & inhibidores , Complejos de Coordinación/farmacología , ADN/metabolismo , Iminas/farmacología , Oxindoles/farmacología , Inhibidores de Proteínas Quinasas/farmacología , Animales , Línea Celular Tumoral , Complejos de Coordinación/metabolismo , Cobre/química , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , División del ADN/efectos de los fármacos , Humanos , Iminas/metabolismo , Ligandos , Ratones , Simulación del Acoplamiento Molecular , Oxindoles/metabolismo , Platino (Metal)/química , Inhibidores de Proteínas Quinasas/metabolismoRESUMEN
Cyclin-dependent kinases (CDKs) play a key role in cell cycle regulation, which makes them a clear therapeutic target to interfere with cell division and proliferation in cancer patients. Palbociclib, a specific inhibitor of CDK4/6 with outstanding clinical efficacy data and limited toxicity, has been recently approved for the treatment of hormone receptor (HR)-positive human epidermal growth factor receptor 2 (HER2)-negative locally advanced or metastatic breast cancer, either in combination with an aromatase inhibitor or in combination with fulvestrant in women who have received prior endocrine therapy. This review describes the mechanism of action, preclinical experiences and clinical data of palbociclib, with a special focus on integrating this data with the positioning of palbociclib in the current clinical guidelines for advanced HR-positive/HER2-negative breast cancer. Aspects of the ongoing major studies are also presented, as well as future prospects in the development of palbociclib.
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Antineoplásicos/uso terapéutico , Neoplasias de la Mama/tratamiento farmacológico , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Piperazinas/uso terapéutico , Inhibidores de Proteínas Quinasas/uso terapéutico , Piridinas/uso terapéutico , Animales , Ciclo Celular/efectos de los fármacos , Ensayos Clínicos como Asunto , Humanos , Piperazinas/farmacología , Piridinas/farmacologíaRESUMEN
BACKGROUND: Cyclin-dependent kinases (CDKs) comprise an important protein family for development of drugs, mostly aimed for use in treatment of cancer but there is also potential for development of drugs for neurodegenerative diseases and diabetes. Since the early 1990s, structural studies have been carried out on CDKs, in order to determine the structural basis for inhibition of this protein target. OBJECTIVE: Our goal here is to review recent structural studies focused on CDKs. We concentrate on latest developments in the understanding of the structural basis for inhibition of CDKs, relating structures and ligand-binding information. METHOD: Protein crystallography has been successfully applied to elucidate over 400 CDK structures. Most of these structures are complexed with inhibitors. We use this richness of structural information to describe the major structural features determining the inhibition of this enzyme. RESULTS: Structures of CDK1, 2, 4-9, 12 13, and 16 have been elucidated. Analysis of these structures in complex with a wide range of different competitive inhibitors, strongly indicate some common features that can be used to guide the development of CDK inhibitors, such as a pattern of hydrogen bonding and the presence of halogen atoms in the ligand structure. CONCLUSION: Nowadays we have structural information for hundreds of CDKs. Combining the structural and functional information we may say that a pattern of intermolecular hydrogen bonds is of pivotal importance for inhibitor specificity. In addition, machine learning techniques have shown improvements in predicting binding affinity for CDKs.
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Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/farmacología , Humanos , Modelos Moleculares , Estructura Molecular , Inhibidores de Proteínas Quinasas/químicaRESUMEN
Temporary meiosis arrest with cyclin-dependent kinases inhibitors has been proposed in order to improve the quality of in vitro matured oocytes. In sheep, however, this phenomenon has been rarely investigated. Therefore, the present study aimed to evaluate the effect of different incubation times with roscovitine on nuclear maturation and cumulus cell expansion of sheep cumulus-oocyte complexes (COCs). For this, COCs were cultured for 0, 6, 12 or 20 h in basic maturation medium (Control) containing 75 µM roscovitine (Rosco). After, they were in vitro matured (IVM) for 18 h in the presence of luteinizing hormone (LH) and follicle-stimulating hormone (FSH). At the end of each treatment, cumulus cell expansion and nuclear maturation were assessed under a stereomicroscope and by Hoechst 33342 staining, respectively. In the Control and Rosco groups, the absence of cumulus cell expansion prevailed at 0, 6, 12 and 20 h. After IVM for 18 h, total cumulus cell expansion in the Rosco treatments was dependent on the exposure time to roscovitine. A significantly high percentage of oocytes treated with roscovitine for 6 h (87%), 12 h or 20 h (65%) were arrested at the germinal vesicle (GV) stage. In contrast, 23% GVBD, 54% metaphase I (MI) and 61% MII oocytes were observed in the Control groups at 6, 12 and 20 h, respectively. In all treatments, a significant percentage of oocytes reached MII after IVM for 18 h. Therefore, roscovitine reversibly arrested the meiosis of sheep oocytes during different culture times with the maximal efficiency of meiotic inhibition reached at 6 h. In addition, reversibility of its inhibitory action on cumulus cells was exposure-time dependent.
Asunto(s)
Células del Cúmulo/efectos de los fármacos , Meiosis/efectos de los fármacos , Oocitos/efectos de los fármacos , Purinas/farmacología , Animales , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Células del Cúmulo/citología , Células del Cúmulo/metabolismo , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Femenino , Hormona Folículo Estimulante/farmacología , Técnicas de Maduración In Vitro de los Oocitos/métodos , Hormona Luteinizante/farmacología , Oocitos/citología , Oocitos/metabolismo , Inhibidores de Proteínas Quinasas/farmacología , Roscovitina , Ovinos , Factores de TiempoRESUMEN
Oxindolimine-copper(II) and zinc(II) complexes that previously have shown to induce apoptosis, with DNA and mitochondria as main targets, exhibit here significant inhibition of kinase CDK1/cyclin B protein. Copper species are more active than the corresponding zinc, and the free ligand shows to be less active, indicating a major influence of coordination in the process, and a further modulation by the coordinated ligand. Molecular docking and classical molecular dynamics provide a better understanding of the effectiveness and kinase inhibition mechanism by these compounds, showing that the metal complex provides a stronger interaction than the free ligand with the ATP-binding site. The metal ion introduces charge in the oxindole species, giving it a more rigid conformation that then becomes more effective in its interactions with the protein active site. Analogous experiments resulted in no significant effect regarding phosphatase inhibition. These results can explain the cytotoxicity of these metal complexes towards different tumor cells, in addition to its capability of binding to DNA, and decreasing membrane potential of mitochondria.
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Complejos de Coordinación/química , Complejos de Coordinación/farmacología , Cobre/química , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Iminas/farmacología , Simulación del Acoplamiento Molecular , Zinc/química , Estabilidad de Medicamentos , Activación Enzimática/efectos de los fármacos , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Iminas/química , Indoles/química , Indoles/farmacología , Ligandos , Modelos Moleculares , OxindolesRESUMEN
The effects of tamoxifen (TAM) on anxiety and depression-like behavior in ovariectomized (OVX) and naïve female rats were investigated. The animals were divided into Sham-TAM, OVX-TAM, Sham and OVX groups. Tamoxifen (1 mg/kg) was administered for 4 weeks. In the forced swimming test, the immobility times in the OVX and Sham-TAM groups were higher than in the Sham group. In the open field, the numbers of central crossings in the OVX and Sham-TAM groups were lower than the number in the Sham group, and the number of peripheral crossings in the OVX group was lower than the number in the Sham group. In the elevated plus maze, the numbers of entries to the open arm among the animals in the Sham-TAM and OVX groups were lower than the number in the Sham group, while the number of entries to the open arm in the OVX-TAM group was higher than the number in the OVX group. It was shown that deletion of ovarian hormones induced anxiety and depression-like behavior. Administration of tamoxifen in naïve rats led to anxiety and depression-like behavior that was comparable with the effects of ovarian hormone deletion. It can be suggested that tamoxifen antagonizes the effects of ovarian hormones. It also seems that tamoxifen has anxiolytic effects on ovariectomized rats.
Foram investigados os efeitos do tamoxifeno (TAM) no comportamento semelhante a ansiedade de depressão de ratas ooforectomizadas (OVX) e controles. Os animais foram divididos em Sham-TAM, OVX-TAM, Sham e OVX groups. Tamoxifeno (1 mg/kg) foi administrado por quatro semanas. No teste de natação forçada, os tempos de imobilidade nos grupos OVX e Sham-TAM foram maiores que aqueles do grupo Sham. No campo aberto, os números de cruzamento no centro nos grupos OVX e Sham-TAM foram menores que aquele do grupo Sham, e o número dos cruzamentos na periferia no grupo OVX foi menor que o número no grupo Sham. No labirinto elevado, os números de entradas com braços abertos entre os animais nos grupos Sham-TAM e OVX foram menores do que aqueles do grupo Sham, enquanto o número de entradas com os braços abertos no grupo OVX-TAM foi maior que aquele no grupo OVX. Foi observado que a deleção dos hormônios ovarianos induziu comportamento similar a ansiedade e depressão. A administração de tamoxifeno em ratos controle induziu a um comportamento que era comparável aos efeitos da deleção do hormônio ovariano. Pode ser sugerido que o tamoxifeno antagoniza os efeitos dos hormônios ovarianos. Parece também que o tamoxifeno tem efeito ansiolítico nas ratas ooforectomizadas.
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Animales , Masculino , Ratas , Cocaína/farmacología , Quinasas Ciclina-Dependientes/metabolismo , Dendritas/efectos de los fármacos , Dendritas/metabolismo , Inhibidores de Captación de Dopamina/farmacología , Núcleo Accumbens/efectos de los fármacos , Núcleo Accumbens/enzimología , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Inhibidores Enzimáticos/farmacología , Microscopía Confocal , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Purinas/farmacología , Ratas Sprague-DawleyRESUMEN
Cyclin-dependent kinases (CDKs) are a family of serine/threonine kinases essential for cell cycle progression. Herein, we describe the participation of CDKs in the physiology of Rhipicephalus microplus, the southern cattle tick and an important disease vector. Firstly, amino acid sequences homologous with CDKs of other organisms were identified from a R. microplus transcriptome database in silico. The analysis of the deduced amino acid sequences of CDK1 and CDK10 from R. microplus showed that both have caspase-3/7 cleavage motifs despite their differences in motif position and length of encoded proteins. CDK1 has two motifs (DKRGD and SAKDA) located opposite to the ATP binding site while CDK10 has only one motif (SLLDN) for caspase 3-7 near the ATP binding site. Roscovitine (Rosco), a purine derivative that inhibits CDK/cyclin complexes by binding to the catalytic domain of the CDK molecule at the ATP binding site, which prevents the transfer of ATP's γphosphoryl group to the substrate. To determine the effect of Rosco on tick CDKs, BME26 cells derived from R. microplus embryo cells were utilized in vitro inhibition assays. Cell viability decreased in the Rosco-treated groups after 24 hours of incubation in a concentration-dependent manner and this was observed up to 48 hours following incubation. To our knowledge, this is the first report on characterization of a cell cycle protein in arachnids, and the sensitivity of BME26 tick cell line to Rosco treatment suggests that CDKs are potential targets for novel drug design to control tick infestation.
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Proteínas de Artrópodos/química , Proteína Quinasa CDC2/química , Quinasas Ciclina-Dependientes/química , Inhibidores de Proteínas Quinasas/farmacología , Purinas/farmacología , Rhipicephalus/efectos de los fármacos , Adenosina Trifosfato/química , Adenosina Trifosfato/metabolismo , Secuencias de Aminoácidos , Animales , Proteínas de Artrópodos/antagonistas & inhibidores , Proteínas de Artrópodos/clasificación , Proteínas de Artrópodos/metabolismo , Proteína Quinasa CDC2/antagonistas & inhibidores , Proteína Quinasa CDC2/clasificación , Proteína Quinasa CDC2/metabolismo , Caspasas/química , Caspasas/metabolismo , Dominio Catalítico , Bovinos , Línea Celular , Supervivencia Celular/efectos de los fármacos , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Quinasas Ciclina-Dependientes/clasificación , Quinasas Ciclina-Dependientes/metabolismo , Escherichia coli/química , Escherichia coli/genética , Simulación del Acoplamiento Molecular , Simulación de Dinámica Molecular , Datos de Secuencia Molecular , Filogenia , Unión Proteica , Inhibidores de Proteínas Quinasas/química , Purinas/química , Proteínas Recombinantes/química , Proteínas Recombinantes/clasificación , Proteínas Recombinantes/metabolismo , Rhipicephalus/citología , Rhipicephalus/enzimología , Roscovitina , Glándulas Salivales/citología , Glándulas Salivales/efectos de los fármacos , Alineación de Secuencia , Homología Estructural de ProteínaRESUMEN
The recent finding of a tissue-specific cell cycle regulator (SCI1) that inhibits cell proliferation/differentiation in the upper pistil points to an unanticipated way of controlling plant morphogenesis. The similarity between the SCI1 RNAi-silenced plants and some auxin-related phenotypes suggested that SCI1 could be involved in the auxin signaling pathway. To address this hypothesis, we analyzed the expression of three auxin-related genes in transgenic plants in which SCI1 was silenced and overexpressed. The results showed that the expression levels of the auxin-related genes largely correlated with the SCI1 expression level. Additionally, we analyzed the Arabidopsis SCI1 upstream regulatory region and found putative cis-acting elements also present in the AtCYCB1;1 AtYUC1, AtYUC2 and AtYUC4 URRs, suggesting a cell cycle- and auxin-related transcriptional regulation. Based on our previous and the current studies, we propose SCI1 as a signal transducer engaging auxin signaling and cell division/differentiation.
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Proteínas de Arabidopsis/fisiología , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Ácidos Indolacéticos/metabolismo , Péptidos y Proteínas de Señalización Intracelular/fisiología , Inhibidores de Proteínas Quinasas/farmacología , Transducción de SeñalRESUMEN
This study aimed to assess the effects of cyclin-dependent kinase (CDK) inhibition on factors involved in the control of meiosis in bovine oocytes: maturation promoting factor (MPF) (p34(cdc2) and cyclin B1) and mitogen activated protein kinase (MAPK). Oocytes were maintained at germinal vesicle (GV) stage in vitro with 10 µM of the CDK inhibitor butyrolactone I (BLI) for 24 h (inhibited). After this period, some of the oocytes were transferred to in vitro maturation (IVM) culture for 24 h (inhibited and matured). Control oocytes were assessed immediately after follicle aspiration (immature) or after in vitro maturation for 24 h (matured). Real-time PCR analyses showed that transcripts for p34(cdc2) and MAPK were detected in immature and inhibited oocytes and decreased after maturation, irrespective of CDK inhibition with BLI. Cyclin B1 was detected at similar levels in all oocyte groups. The p34(cdc2) and MAPK proteins were detected by Western blotting at similar levels in all oocyte groups, and cyclin B1 protein was detected only after maturation. Immunofluorescence detection showed that p34(cdc2) was localized in the cytoplasm and GV of immature oocytes, and then throughout the cytoplasm after maturation. Cyclin B1 and MAPK were detected in the cytoplasm in all oocyte groups. Maturation promoting factor and MAPK activities were similar throughout most of maturation for oocytes treated with or without BLI. In conclusion, CDK inhibition did not affect the expression (mRNA and protein levels) and localization of MPF and MAPK, and had nearly no effect on kinase activities during maturation.
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4-Butirolactona/análogos & derivados , Bovinos/fisiología , Regulación de la Expresión Génica/efectos de los fármacos , Factor Promotor de Maduración/metabolismo , Quinasas de Proteína Quinasa Activadas por Mitógenos/metabolismo , Oocitos/efectos de los fármacos , 4-Butirolactona/farmacología , Animales , Ciclina B1/genética , Ciclina B1/metabolismo , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Factor Promotor de Maduración/genética , Oocitos/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismoRESUMEN
The complex CDK3-cyclin is involved in the control of the progression of G0. While the mechanisms governing early and late G1 progression are well understood, very little is known about the G0-G1 transition. Human CDK3 is closely related to cyclin-dependent kinase 2 (CDK2). Since there is no crystallographic structure of human CDK3, this work describes for the first time a molecular model of human CDK3 complexed with several inhibitors. Comparison of the binary complexes with different inhibitors strongly indicates that those inhibitors should inhibit CDK3 as well as CDK2.
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Quinasas Ciclina-Dependientes/química , Inhibidores de Proteínas Quinasas/química , Secuencia de Aminoácidos , Quinasa 3 Dependiente de Ciclina , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Humanos , Modelos Moleculares , Datos de Secuencia Molecular , Inhibidores de Proteínas Quinasas/farmacología , Homología de Secuencia de AminoácidoRESUMEN
Among the different strategies to treat cancer, chemotherapy approaches are the subject of intense research efforts. There is still a high demand for new anticancer drugs exhibiting improved efficiency and selectivity for their use in combined therapy strategies. The high development of molecular and cellular biology tools has made possible the set up of simple in vitro assays, susceptible to automation, thus bringing about the possibility of rapid screening of hundreds of compounds. Chemistry has reacted to this challenge by developing a new technology: combinatorial chemistry. By this procedure large collections of compounds, known as chemical libraries, can be prepared in a rapid and efficient manner. In recent years, combinatorial chemistry has had a great impact on drug discovery programmes addressed to tackling cancer pharmaceutical targets. In this review, the contribution of this technology to the discovery of anticancer drugs that are currently in clinical trials or already in the market is discussed.
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Investigación Biomédica/métodos , Técnicas Químicas Combinatorias , Neoplasias/tratamiento farmacológico , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Resistencia a Antineoplásicos , Humanos , Proteínas Tirosina Quinasas/antagonistas & inhibidoresRESUMEN
The inhibitory activity towards p34(cdc2)/cyclin b kinase (CBK) enzyme of 30 cytokinin-derived compounds has been successfully modelled using 2D spatial autocorrelation vectors. Predictive linear and non-linear models were obtained by forward stepwise multi-linear regression analysis (MRA) and artificial neural network (ANN) approaches respectively. A variable selection routine that selected relevant non-linear information from the data set was employed prior to networks training. The best ANN with three input variables was able to explain about 87% data variance in comparison with 80% by the linear equation using the same number of descriptors. Similarly, the neural network had higher predictive power. The MRA model showed a linear dependence between the inhibitory activities and the spatial distributions of masses, electronegativities and van der Waals volumes on the inhibitors molecules. Meanwhile, ANN model evidenced the occurrence of non-linear relationships between the inhibitory activity and the mass distribution at different topological distance on the cytokinin-derived compounds. Furthermore, inhibitors were well distributed regarding its activity levels in a Kohonen self-organizing map (SOM) built using the input variables of the best neural network.
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Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Citocininas/farmacología , Modelos Biológicos , Animales , Proteína Quinasa CDC2/antagonistas & inhibidores , Citocininas/química , Femenino , Técnicas In Vitro , Matemática , Redes Neurales de la Computación , Inhibidores de Proteínas Quinasas/química , Inhibidores de Proteínas Quinasas/farmacología , Análisis de Regresión , Estrellas de Mar/enzimología , Relación Estructura-ActividadRESUMEN
Tumor cell proliferation is frequently associated to genetic or epigenetic alterations in key cell cycle regulators. Most human tumors deregulate this pathway to sustain proliferation with independence of external mitogenic factors. In addition, the alteration of cell cycle proteins may confer genomic instability that results in additional mutations in these tumor cells. The frequent alteration of the cell cycle in tumor cells has launched the identification for critical cell cycle regulators as anticancer targets. The inhibition of some cell cycle kinases such as cyclin-dependent kinases (CDKs) or the Aurora and Polo mitotic kinases is currently under study in several preclinical and clinical trials. Similarly, the clinical success of microtubule poisons such as taxol has promoted new applied research in mitosis regulation. Recent investigations have suggested new targets of interest including additional kinases, phosphatases and other mitotic regulators such as microtubule motor proteins (kinesins). Current research in this area will undoubtedly result in new and improved targeted therapies for cancer treatment.
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Proteínas de Ciclo Celular/antagonistas & inhibidores , Ciclo Celular/efectos de los fármacos , Proteínas de Neoplasias/antagonistas & inhibidores , Neoplasias/tratamiento farmacológico , Huso Acromático/efectos de los fármacos , Animales , Aurora Quinasas , Ciclo Celular/genética , Ciclo Celular/fisiología , Proteínas de Ciclo Celular/fisiología , Segregación Cromosómica/efectos de los fármacos , Segregación Cromosómica/fisiología , Proteínas Inhibidoras de las Quinasas Dependientes de la Ciclina/fisiología , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Quinasas Ciclina-Dependientes/fisiología , Daño del ADN , Replicación del ADN/efectos de los fármacos , Diseño de Fármacos , Predicción , Fase G1/efectos de los fármacos , Genes cdc , Humanos , Ratones , Modelos Biológicos , Proteínas Motoras Moleculares , Proteínas de Neoplasias/fisiología , Neoplasias/genética , Neoplasias/fisiopatología , Fosfoproteínas Fosfatasas/fisiología , Fosforilación/efectos de los fármacos , Procesamiento Proteico-Postraduccional/efectos de los fármacos , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/fisiología , Proteínas Proto-Oncogénicas/antagonistas & inhibidores , Proteínas Proto-Oncogénicas/fisiología , Transducción de Señal/efectos de los fármacos , Quinasa Tipo Polo 1RESUMEN
Artificial neural network ensembles were used for modeling the cyclin-dependent kinase inhibition of 1H-pyrazolo[3,4-d]pyrimidine derivatives. The structural characteristics of these inhibitors were encoded in relevant 3D-spatial descriptors extracted by genetic algorithm feature selection. Bayesian-regularized multilayer neural networks, trained by the back-propagation algorithm, were developed using these variables as inputs. The predictive power of the model was tested by leave-one-out cross validation. In addition, for a more rigorous measure of the predictive capacity, multiple validation sets were randomly generated as members of neural network ensembles, which makes doing averaged predictions feasible. In this way, the predictive power was analyzed accounting for the averaged test set R values and test set mean-square errors. Otherwise, Kohonen self-organizing maps were used as an additional tool for the same modeling. The location of the inhibitors in a map facilitates the analysis of the connection between compounds and serves as a useful tool for qualitative predictions.
Asunto(s)
Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Redes Neurales de la Computación , Pirazoles/farmacología , Pirimidinas/farmacología , Algoritmos , Inteligencia Artificial , Teorema de Bayes , Bases de Datos Factuales , Modelos Moleculares , Relación Estructura-Actividad CuantitativaRESUMEN
Cyclin-dependent kinase inhibitors (CDKIs) such as butyrolactone I (BL-I) and roscovitine (ROS) maintain bovine oocytes blocked at the germinal vesicle (GV) stage. Bohemine (BOH), another CDKI, has been used for oocyte activation. The objective of this study was to determine whether BOH blocks meiosis and to compare its efficiency with other CDKIs (ROS and BL-I). Oocytes were cultured for 24 h in 0, 50, 100 and 150 microM BOH to determine the best concentration for blocking meiosis (experiment 1). GV rates were 3.3%, 64.5%, 83.3% and 88.9% (0,50, 100 and 150 microM, respectively). Experiment 2 compared meiotic inhibition efficiency of BOH (100 microM), ROS (25 microM) and BL-I (100 microM). BL-I presented the highest GV rates (97.5%). BOH and ROS were similar to each other (85.4% and 79.9%, respectively). To assess the reversibility of meiotic inhibition (experiment 3), oocytes underwent in vitro maturation (IVM) for 18 h after the 24 h inhibition. Control oocytes were submitted to IVM for 18 h (C18) or 24 h (C24). Maturation rates were either similar to (ROS and BL-I: 96.0% and 93.6%, respectively) or superior to (BOH, 96.9%) C24 (91.0%). All groups were superior to C18 (82.5%). In experiment 4, oocytes were treated as in experiment 3 and then in vitro fertilized and cultured for 8 days. Blastocyst rates for BL-I (32.3%) were similar to C24 (35.0%), while those for BOH (20.2%) and ROS (24.2%) were inferior. All groups were inferior to C18 (43.4%). The results show that: (a) BOH inhibits meiosis resumption; (b) BL-I is the most effective of the CDKIs tested for blocking meiosis; (c) culture of oocytes with meiosis inhibitors is fully reversible in terms of nuclear maturation but they may either decrease (BOH and ROS) or maintain (BL-I) embryo development rates.
Asunto(s)
4-Butirolactona/análogos & derivados , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Embrión de Mamíferos/efectos de los fármacos , Meiosis/efectos de los fármacos , Oocitos/efectos de los fármacos , 4-Butirolactona/farmacología , Animales , Bovinos , Purinas/farmacología , RoscovitinaRESUMEN
The CRK3 cyclin-dependent kinase of Leishmania has been shown by genetic manipulation of the parasite to be essential for proliferation. We present data which demonstrate that chemical inhibition of CRK3 impairs the parasite's viability within macrophages, thus further validating CRK3 as a potential drug target. A microtiter plate-based histone H1 kinase assay was developed to screen CRK3 against a chemical library enriched for protein kinase inhibitors. Twenty-seven potent CRK3 inhibitors were discovered and screened against Leishmania donovani amastigotes in vitro. Sixteen of the CRK3 inhibitors displayed antileishmanial activity, with a 50% effective dose (ED50) of less than 10 microM. These compounds fell into four chemical classes: the 2,6,9-trisubstituted purines, including the C-2-alkynylated purines; the indirubins; the paullones; and derivatives of the nonspecific kinase inhibitor staurosporine. The paullones and staurosporine derivatives were toxic to macrophages. The 2,6,9-trisubstituted purines inhibited CRK3 in vitro, with 50% inhibitory concentrations ranging from high nanomolar to low micromolar concentrations. The most potent inhibitors of CRK3 (compounds 98/516 and 97/344) belonged to the indirubin class; the 50% inhibitory concentrations for these inhibitors were 16 and 47 nM, respectively, and the ED50s for these inhibitors were 5.8 and 7.6 microM, respectively. In culture, the indirubins caused growth arrest, a change in DNA content, and aberrant cell types, all consistent with the intracellular inhibition of a cyclin-dependent kinase and disruption of cell cycle control. Thus, use of chemical inhibitors supports genetic studies to confirm CRK3 as a validated drug target in Leishmania and provides pharmacophores for further drug development.
Asunto(s)
Antiprotozoarios/farmacología , Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Quinasas Ciclina-Dependientes/genética , Leishmania mexicana/efectos de los fármacos , Leishmania mexicana/genética , Animales , Antiprotozoarios/uso terapéutico , Proteína Quinasa CDC2 , Ciclo Celular/efectos de los fármacos , Células Cultivadas , ADN Protozoario/genética , Evaluación Preclínica de Medicamentos , Citometría de Flujo , Colorantes Fluorescentes , Biblioteca de Genes , Humanos , Immunoblotting , Indoles/farmacología , Cariotipificación , Leishmaniasis Cutánea/tratamiento farmacológico , Macrófagos Peritoneales/parasitología , Ratones , Proteínas Quinasas/metabolismo , Proteínas ProtozoariasRESUMEN
Pharmacological blockers of cyclin-dependent kinases (CDKs) can inhibit cell cycle progression. Deferoxamine (DFO) and mimosine (MIMO) arrest cells reversibly at the G1/S transition and olomoucine (OLO) inhibits the cell cycle at both G1/S and G2/M. We investigated the effect of these drugs upon cell death in histotypical explants taken from the retina of neonatal rats. Degeneration of retinal ganglions cells (RGC) induced by axotomy was inhibited by OLO (100 microM) but not by DFO (up to 2 mM) or MIMO (up to 1 mM). On the other hand, after 1 day in vitro, all cell cycle inhibitors induced cell death in the neuroblastic layer (NBL) of the explants. DFO and MIMO induced cell death only of proliferating cells, identified either by their incorporation of bromodeoxyuridine or by immunolabeling the proliferating cell nuclear antigen. In turn, OLO induced cell death of both proliferating and post-mitotic cells. However, the post-mitotic cells were unlabeled with markers of retinal differentiation. Our results indicate that cyclin-dependent kinases are involved in the control of sensitivity to cell death in the retina, and that retinal cells present differentiation-dependent responses to modulation of CDK activity.
Asunto(s)
Quinasas Ciclina-Dependientes/antagonistas & inhibidores , Inhibidores Enzimáticos/farmacología , Retina/citología , Retina/crecimiento & desarrollo , Adenilil Ciclasas/metabolismo , Animales , Anisomicina/farmacología , Antivirales , Axotomía , Bromodesoxiuridina , Recuento de Células , Ciclo Celular/efectos de los fármacos , Muerte Celular/efectos de los fármacos , Colforsina/farmacología , Técnicas de Cultivo , ADN/biosíntesis , ADN/genética , Activación Enzimática/efectos de los fármacos , Inmunohistoquímica , Inhibidores de la Síntesis de la Proteína/farmacología , Ratas , Retina/efectos de los fármacos , Degeneración Retiniana/patologíaRESUMEN
Flavopiridol has been shown to potently inhibit CDK1 and 2 (cyclin-dependent kinases 1 and 2) and most recently it has been found that it also inhibits CDK9. The complex CDK9-cyclin T1 controls the elongation phase of transcription by RNA polymerase II. The present work describes a molecular model for the binary complex CDK9-flavopiridol. This structural model indicates that the inhibitor strongly binds to the ATP-binding pocket of CDK9 and the structural comparison of the complex CDK2-flavopiridol correlates the structural differences with differences in inhibition of these CDKs by flavopiridol. This structure opens the possibility of testing new inhibitor families, in addition to new substituents for the already known leading structures such as flavones and adenine derivatives.