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PURPOSE OF REVIEW: Sequential use of radiation therapy before cyclin-dependent kinase (CDK) inhibitors in women with early breast cancer seems reasonable and with a low toxicity rate. This study aimed to evaluate the possible interaction between RT and CDK inhibitors in the adjuvant setting for patients with positive hormone receptors and HER-2 negative, investigating toxicity and the treatment sequencing. RECENT FINDINGS: CDK inhibitors have been studied in patients with localized breast cancer and can improve invasive disease-free survival outcomes. Regarding the time of RT, all trials used CDK inhibitors after the RT. Interruptions in the CDK inhibitors were performed in 27.1% in Pallas, 17.5% in Penelope-B, and 16.6% in Monarch-E trials due to adverse events. Data from the Natalee trial are still not reported. The main adverse event grade III was neutropenia, with good resolution of the symptoms over time. CDK inhibitors applied sequentially and after RT postoperative showed a low profile of acute toxicity and suitable oncological outcomes.
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Neoplasias de la Mama , Femenino , Humanos , Neoplasias de la Mama/tratamiento farmacológico , Neoplasias de la Mama/radioterapia , Quinasas Ciclina-Dependientes , Supervivencia sin Enfermedad , Oncología Médica , Inhibidores de Proteínas Quinasas/uso terapéutico , Quinasa 4 Dependiente de la Ciclina , Quinasa 6 Dependiente de la CiclinaRESUMEN
Cyclin/cyclin-dependent kinase (CDK) heterodimers have multiple phosphorylation targets and may alter the activity of these targets. Proteins from different metabolic processes are among the phosphorylation targets, that is, enzymes of central carbon metabolism. This work explores the interaction of Cyc/CDK complex members with the glycolytic enzymes hexokinase 7 (HXK7) and glyceraldehyde-3-phosphate dehydrogenase (GAP). Both enzymes interacted steadily with CycD2;2, CycB2;1 and CDKA;1 but not with CDKB1;1. However, Cyc/CDKB1;1 complexes phosphorylated both enzymes, decreasing their activities. Treatment with a CDK-specific inhibitor (RO-3306) or with lambda phosphatase after kinase assay restored total HXK7 activity, but not GAP activity. In enzymatic assays, increasing concentrations of CDKB1;1, but not of CycD2;2, CycB2;1 or CycD2;2/CDKB1;1 complex, decreased GAP activity. Cell cycle regulators may modulate carbon channeling in glycolysis by two different mechanisms: Cyc/CDK-mediated phosphorylation of targets (e.g., HXK7; canonical mechanism) or by direct and transient interaction of the metabolic enzyme (e.g., GAP) with CDKB1;1 without a Cyc partner (alternative mechanism).
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Proteínas de Ciclo Celular , Hexoquinasa , Proteínas de Ciclo Celular/metabolismo , Zea mays/metabolismo , Quinasas Ciclina-Dependientes/metabolismo , Gliceraldehído-3-Fosfato Deshidrogenasas/metabolismo , Glucólisis , Ciclo CelularRESUMEN
Alzheimer's disease (AD) is a progressive neuroinflammatory and neurodegenerative disorder that affects different regions of the brain. Its pathophysiology includes the accumulation of ß-amyloid protein, formation of neurofibrillary tangles, and inflammatory processes. Genetic factors are involved in the onset of AD, but they are not fully elucidated. Identification of gene expression in encephalic tissues of patients with AD may help elucidate its development. Our objectives were to characterize and compare the gene expression of CDK10, CDK11, FOXO1, and FOXO3 in encephalic tissue samples from AD patients and elderly controls, from the auditory cortex and cerebellum. RT-qPCR was used on samples from 82 individuals (45 with AD and 37 controls). We observed a statistically significant increase in CDK10 (p = 0.029*) and CDK11 (p = 0.048*) gene expression in the AD group compared to the control, which was most evident in the cerebellum. Furthermore, the Spearman test demonstrated the presence of a positive correlation of gene expression both in the auditory cortex in the AD group (r = 0.046/p = 0.004) and control group (r = 0.454/p = 0.005); and in the cerebellum in the AD group (r = 0.654 /p < 0.001). There was no statistically significant difference and correlation in the gene expression of FOXO1 and FOXO3 in the AD group and the control. In conclusion, CDK10 and CDK11 have high expression in AD patients compared to control, and they present a positive correlation of gene expression in the analyzed groups and tissues, which suggests that they play an important role in the pathogenesis of AD.
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Enfermedad de Alzheimer , Humanos , Anciano , Enfermedad de Alzheimer/metabolismo , Péptidos beta-Amiloides/metabolismo , Ovillos Neurofibrilares/metabolismo , Encéfalo/metabolismo , Expresión Génica , Proteína Forkhead Box O1/genética , Proteína Forkhead Box O1/metabolismo , Proteína Forkhead Box O3/genética , Quinasas Ciclina-Dependientes/genética , Quinasas Ciclina-Dependientes/metabolismoRESUMEN
Cyclin-dependent kinases (CDKs), in association with cyclins, control cell cycle progression by phosphorylating a large number of substrates. In animals, activation of CDKs regularly requires both the association with a cyclin and then phosphorylation of a highly conserved threonine residue in the CDK activation loop (the classical mechanism), mediated by a CDK-activating kinase (CAK). In addition to this typical mechanism of activation, some CDKs can also be activated by the association of a cyclin to a monomeric CDK previously phosphorylated by CAK although not all CDKs can be activated by this mechanism. In animals and yeast, cyclin, in addition to being required for CDK activation, provides substrate specificity to the cyclin/CDK complex; however, in plants both the mechanisms of CDKs activation and the relevance of the CDK-associated cyclin for substrate targeting have been poorly studied. In this work, by co-expressing proteins in E. coli, we studied maize CDKA2;1a and CDKB1;1, two of the main types of CDKs that control the cell cycle in plants. These kinases could be activated by the classical mechanism and by the association of CycD2;2a to a phosphorylated intermediate in its activation loop, a previously unproven mechanism for the activation of plant CDKs. Unlike CDKA2;1a, CDKB1;1 did not require CAK for its activation, since it autophosphorylated in its activation loop. Phosphorylation of CDKB1;1 and association of CycD2;2 was not enough for its full activation as association of maize CKS, a scaffolding protein, differentially stimulated substrate phosphorylation. Our results suggest that both CDKs participate in substrate recognition.
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Proteínas Serina-Treonina Quinasas , Zea mays , Animales , Proteínas Serina-Treonina Quinasas/metabolismo , Zea mays/genética , Escherichia coli/metabolismo , Quinasas Ciclina-Dependientes/metabolismo , Ciclinas/genética , Ciclinas/metabolismo , Saccharomyces cerevisiae/metabolismoRESUMEN
Cell senescence is a state of limited cell proliferation during a stress response or as part of a programmed process. When a senescent cell stops dividing, maintaining metabolic activity contributes to cellular homeostasis maintenance. In this process, the cell cycle is arrested at the G0/G1 phase. p16INK4A protein is a key regulator of this process via its cyclindependent kinase inhibitor (CDKI) function. CDKI 2A (CDKN2A)/p16 gene expression is regulated by DNA methylation and histone acetylation. Sirtuins (SIRTs) are nicotinamide dinucleotide (NAD+)dependent deacetylases that have properties which prevent diseases and reverse certain aspects of aging (such as immune, metabolic and cardiovascular diseases). By performing quantitative PCR, Western blot, ChIP, and siRNAs assays, in this study it was demonstrated that CDKN2A/p16 gene transcriptional activation and repression were accompanied by selective deposition and elimination of histone acetylation during the senescence of MRC5 cells. Specifically, significant H3K9Ac and H3K18Ac enrichment in cells with a senescent phenotype concomitant with CDKN2A/p16 gene overexpression was demonstrated compared with the nonsenescent phenotype. Furthermore, the presence of H3K18Ac in deacetyltransferase SIRT7 knockdown MRC5 cells allowed CDKN2A/p16 promoter activation. These results suggested that SIRT7 served as a critical component of an epigenetic mechanism involved in senescence mediated by the CDKN2A/p16 gene.
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Inhibidor p16 de la Quinasa Dependiente de Ciclina , Sirtuinas , Senescencia Celular/genética , Inhibidor p16 de la Quinasa Dependiente de Ciclina/genética , Inhibidor p16 de la Quinasa Dependiente de Ciclina/metabolismo , Quinasas Ciclina-Dependientes/metabolismo , Histonas/metabolismo , NAD/metabolismo , Niacinamida , Sirtuinas/genética , Sirtuinas/metabolismoRESUMEN
BACKGROUND: Pituitary adenomas (PA) are the second most common intracranial tumors and are classified according to hormone they produce, and the transcription factors they express. The majority of PA occur sporadically, and their molecular pathogenesis is incompletely understood. METHODS: Here we performed transcriptome and proteome analysis of tumors derived from POU1F1 (GH-, TSH-, and PRL-tumors, N = 16), NR5A1 (gonadotropes and null cells adenomas, n = 17) and TBX19 (ACTH-tumors, n = 6) lineages as well as from silent ACTH-tumors (n = 3) to determine expression of kinases, cyclins, CDKs and CDK inhibitors. RESULTS: The expression profiles of genes encoding kinases were distinctive for each of the three PA lineage: NR5A1-derived tumors showed upregulation of ETNK2 and PIK3C2G and alterations in MAPK, ErbB and RAS signaling, POU1F1-derived adenomas showed upregulation of PIP5K1B and NEK10 and alterations in phosphatidylinositol, insulin and phospholipase D signaling pathways and TBX19-derived adenomas showed upregulation of MERTK and STK17B and alterations in VEGFA-VEGFR, EGF-EGFR and Insulin signaling pathways. In contrast, the expression of the different genes encoding cyclins, CDK and CDK inhibitors among NR5A1-, POU1F1- and TBX19-adenomas showed only subtle differences. CDK9 and CDK18 were upregulated in NR5A1-adenomas, whereas CDK4 and CDK7 were upregulated in POUF1-adenomas. CONCLUSIONS: The kinome of PA clusters these lesions into three distinct groups according to the transcription factor that drives their terminal differentiation. And these complexes could be harnessed as molecular therapy targets.
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Adenoma , Neoplasias Hipofisarias , Adenoma/metabolismo , Hormona Adrenocorticotrópica/genética , Proteínas Reguladoras de la Apoptosis/genética , Quinasas Ciclina-Dependientes/genética , Quinasas Ciclina-Dependientes/metabolismo , Ciclinas/genética , Ciclinas/metabolismo , Humanos , Insulina , Neoplasias Hipofisarias/genética , Neoplasias Hipofisarias/metabolismo , Neoplasias Hipofisarias/patología , Proteínas Serina-Treonina Quinasas , Factores de Transcripción/genética , TranscriptomaRESUMEN
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
Cyclin-dependent kinases (CDKs) comprise a family of about 20 serine/threonine kinases whose catalytic activity requires a regulatory subunit known as cyclin; these enzymes play several roles in the cell cycle and transcription. PCTAIRE kinases (PCTKs) are a CDK subfamily, characterized by serine to cysteine mutation in the consensus PSTAIRE motif, involved in binding to the cyclin. One member of this class is PCTK3, which has two isoforms (a and b) and is also known as CDK18. After being activated by cyclin A2 or phosphorylation at Ser12 by PKA, PCTK3 can perform several functions. Among these functions, we may highlight the following: modulation of cargo transport in membrane traffic, p53-responsive gene, regulation of genome integrity. According to different studies, PCTK3 dysfunction is related to a wide range of diseases, such as metabolic diseases, cerebral ischemia, depression, cancer, neurological disorders, and Alzheimer's disease. Although this protein participates in different biological events, we may say that PCTK3 has received far less attention than other CDKs. There are thousands of published articles about other CDKs and less than two hundred articles related to PCTK3. The main objective of this review is to present the selected published studies about this protein. Our focus is on PCTK3 particularities compared to other CDKs. Here we give an overview of the biological functions of PCTK3 and explore its potential as a target for drug design.
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Quinasas Ciclina-Dependientes , Proteínas Serina-Treonina Quinasas , Ciclo Celular , Puntos de Control del Ciclo Celular , Quinasa 2 Dependiente de la Ciclina , Quinasas Ciclina-Dependientes/metabolismo , FosforilaciónRESUMEN
Pigmentation characteristics are well-known risk factors for skin cancer. Polymorphisms in pigmentation genes have been associated with these traits and with the risk of malignancy. However, the functional relationship between genetic variation and disease is still unclear. This study aims to assess whether pigmentation SNPs are associated with pigmentary traits and skin cancer via DNA methylation (DNAm). Using a meta-GWAS of whole-blood DNAm from 36 European cohorts (N = 27,750; the Genetics of DNA Methylation Consortium, GoDMC), we found that 19 out of 27 SNPs in 10 pigmentation genes were associated with 391 DNAm sites across 30 genomic regions. We examined the effect of 25 selected DNAm sites on pigmentation traits, sun exposure phenotypes and skin cancer and on gene expression in whole blood. We uncovered an association of DNAm site cg07402062 with red hair in the Avon Longitudinal Study of Parents and Children (ALSPAC). We also found that the expression of ASIP and CDK10 was associated with hair colour, melanoma and basal cell carcinoma. Our results indicate that DNAm and expression of pigmentation genes may play a role as potential mediators of the relationship between genetic variants, pigmentation phenotypes and skin cancer and thus deserve further scrutiny.
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Proteína de Señalización Agouti/genética , Carcinoma Basocelular/genética , Quinasas Ciclina-Dependientes/genética , Metilación de ADN , ADN de Neoplasias/genética , Melanoma/genética , Proteínas de Neoplasias/genética , Neoplasias Cutáneas/genética , Pigmentación de la Piel/genética , Proteína de Señalización Agouti/metabolismo , Carcinoma Basocelular/metabolismo , Quinasas Ciclina-Dependientes/metabolismo , ADN de Neoplasias/metabolismo , Estudio de Asociación del Genoma Completo , Humanos , Estudios Longitudinales , Melanoma/metabolismo , Proteínas de Neoplasias/metabolismo , Neoplasias Cutáneas/metabolismoRESUMEN
Ambystoma mexicanum (axolotl) has been one of the major experimental models for the study of regeneration during the past 100 years. Axolotl limb regeneration takes place through a multi-stage and complex developmental process called epimorphosis that involves diverse events of cell reprogramming. Such events start with dedifferentiation of somatic cells and the proliferation of quiescent stem cells to generate a population of proliferative cells called blastema. Once the blastema reaches a mature stage, cells undergo progressive differentiation into the diverse cell lineages that will form the new limb. Such pivotal cell reprogramming phenomena depend on the fine-tuned regulation of the cell cycle in each regeneration stage, where cell populations display specific proliferative capacities and differentiation status. The axolotl genome has been fully sequenced and released recently, and diverse RNA-seq approaches have also been generated, enabling the identification and conservatory analysis of core cell cycle regulators in this species. We report here our results from such analyses and present the transcriptional behavior of key regulatory factors during axolotl limb regeneration. We also found conserved protein interactions between axolotl Cyclin Dependent Kinases 2, 4 and 6 and Cyclins type D and E. Canonical CYC-CDK interactions that play major roles in modulating cell cycle progression in eukaryotes.
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Ambystoma mexicanum/crecimiento & desarrollo , Ciclo Celular , Extremidades/crecimiento & desarrollo , Regeneración , Animales , Diferenciación Celular , Linaje de la Célula , Quinasas Ciclina-Dependientes/genética , Ciclinas/genética , RNA-SeqRESUMEN
Cyclin-dependent kinase A (CDKA) is a key component for cell cycle progression. The catalytic kinase activity depends on the protein's ability to form an active complex with cyclins and on phosphoregulatory mechanisms. Cell cycle arrest and plant growth impairment under abiotic stress have been linked to different molecular processes triggered by increased levels of reactive oxygen and nitrogen species (ROS and RNS). Among these, posttranslational modifications (PTMs) of key proteins such as CDKA;1 may be of significance. Herein, isolated maize embryo axes were subjected to sodium nitroprusside (SNP) as an inductor of nitrosative conditions to evaluate if CDKA;1 protein was a target for RNS. A high degree of protein nitration was detected; this included the specific Tyr-nitration of CDKA;1. Tyr15 and Tyr19, located at the ATP-binding site, were the selective targets for nitration according to both in silico analysis using the predictive software GPS-YNO2, and in vitro mass spectrometry studies of recombinant nitrated ZmCDKA;1. Spectrofluorometric measurements demonstrated a reduction of ZmCDKA;1-NO2 affinity for ATP. From these results, we conclude that Tyr nitration in CDKA;1 could act as an active modulator of cell cycle progression during redox stress.
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Quinasas Ciclina-Dependientes/metabolismo , Procesamiento Proteico-Postraduccional , Tirosina/metabolismo , Zea mays/metabolismo , Adenosina Trifosfato/química , Adenosina Trifosfato/metabolismo , Sitios de Unión , Cromatografía Liquida , Quinasas Ciclina-Dependientes/química , Modelos Moleculares , Desarrollo de la Planta , Unión Proteica , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Relación Estructura-Actividad , Espectrometría de Masas en Tándem , Tirosina/química , Zea mays/genéticaRESUMEN
p21Waf/CIP1 is a small unstructured protein that binds and inactivates cyclin-dependent kinases (CDKs). To this end, p21 levels increase following the activation of the p53 tumor suppressor. CDK inhibition by p21 triggers cell-cycle arrest in the G1 and G2 phases of the cell cycle. In the absence of exogenous insults causing replication stress, only residual p21 levels are prevalent that are insufficient to inhibit CDKs. However, research from different laboratories has demonstrated that these residual p21 levels in the S phase control DNA replication speed and origin firing to preserve genomic stability. Such an S-phase function of p21 depends fully on its ability to displace partners from chromatin-bound proliferating cell nuclear antigen (PCNA). Vice versa, PCNA also regulates p21 by preventing its upregulation in the S phase, even in the context of robust p21 induction by irradiation. Such a tight regulation of p21 in the S phase unveils the potential that CDK-independent functions of p21 may have for the improvement of cancer treatments.
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Inhibidor p21 de las Quinasas Dependientes de la Ciclina/genética , Replicación del ADN/genética , Antígeno Nuclear de Célula en Proliferación/genética , Quinasas Ciclina-Dependientes/genética , Humanos , Inhibidores de Proteínas Quinasas/metabolismo , Fase S/genéticaRESUMEN
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
Cyclin dependent kinase A; 1 (CDKA; 1) is essential in G1/S transition of cell cycle and its oxidation has been implicated in cell cycle arrest during plant abiotic stress. In the present study, an evaluation at the molecular level was performed to find possible sites of protein oxidative modifications. In vivo studies demonstrated that carbonylation of maize CDKA,1 is associated with a decrease in complex formation with maize cyclin D (CycD). Control and in vitro oxidized recombinant CDKA; 1 were sequenced by mass spectrometry. Proline at the PSTAIRE cyclin-binding motif was identified as the most susceptible oxidation site by comparative analysis of the resulted peptides. The specific interaction between CDKA; 1 and CycD6; 1, measured by surface plasmon resonance (SPR), demonstrated that the affinity and the kinetic of the interaction depended on the reduced-oxidized state of the CDKA; 1. CDKA; 1 protein oxidative modification would be in part responsible for affecting cell cycle progression, and thus producing plant growth inhibition under oxidative stress.
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Quinasas Ciclina-Dependientes/metabolismo , Ciclinas/metabolismo , Prolina/metabolismo , Zea mays/enzimología , Secuencia de Aminoácidos , Quinasas Ciclina-Dependientes/química , Quinasas Ciclina-Dependientes/genética , Ciclinas/química , Modelos Moleculares , Oxidación-Reducción , Prolina/química , Alineación de SecuenciaRESUMEN
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
BACKGROUND: The essentially unlimited expansion potential and the pluripotency of human embryonic stem cells (hESCs) make them attractive for cell-based therapeutic purposes. Although hESCs can indefinitely proliferate in culture, unlike transformed cancer cells, they are endowed with a cell-intrinsic property termed mitochondrial priming that renders them highly sensitive to apoptotic stimuli. Thus, all attempts to broaden the insights into hESCs apoptosis may be helpful for establishing pro-survival strategies valuable for its in vitro culture and further use in clinical applications. Cyclin-dependent kinases (CDKs), a family of serine/threonine protein kinases originally identified as regulators of the eukaryotic cell cycle, can also regulate transcription and differentiation. Moreover, there are compelling data suggesting that its activities are involved in certain apoptotic programs in different cell types. Currently, it is not completely determined whether CDKs regulate apoptotic processes in rapidly proliferating and apoptosis-prone hESCs. In this study, to elucidate the effect of CDKs inhibition in hESCs we used Roscovitine (ROSC), a purine analogue that selectively inhibits the activities of these kinases. RESULTS: Inhibition of CDKs by ROSC triggers programmed cell death in hESCs but not in proliferating somatic cells (human fibroblasts). The apoptotic process encompasses caspase-9 and -3 activation followed by PARP cleavage. ROSC treatment also leads to p53 stabilization, which coincides with site-specific phosphorylation at serine 46 and decreased levels of Mdm2. Additionally, we observed a transcriptional induction of p53AIP1, a repression of pro-survival factor Mcl-1 and an up-regulation of pro-apoptotic BH3-only proteins NOXA and PUMA. Importantly, we found that the role of CDK2 inhibition appears to be at best accessory as an active CDK2 is not required to ensure hESCs survival. CONCLUSION: Our experimental data reveal that hESCs, contrary to fibroblasts, exhibit a pronounced sensitivity to ROSC.
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Quinasas Ciclina-Dependientes/farmacología , Células Madre Embrionarias Humanas/citología , Inhibidores de Proteínas Quinasas/farmacología , Roscovitina/farmacología , Apoptosis/efectos de los fármacos , Ciclo Celular/efectos de los fármacos , Puntos de Control del Ciclo Celular/efectos de los fármacos , Línea Celular , Regulación hacia Abajo/efectos de los fármacos , Células Madre Embrionarias Humanas/efectos de los fármacos , Humanos , Proteína 1 de la Secuencia de Leucemia de Células Mieloides/metabolismo , Fosforilación/efectos de los fármacos , Dominios Proteicos , ARN Polimerasa II/química , ARN Polimerasa II/metabolismo , Transducción de Señal/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas/farmacología , Proteína p53 Supresora de Tumor/metabolismoRESUMEN
The cyclin-dependent kinase 5 (CDK5) is known as an exceptional component of the CDK family, due to its characteristic regulatory pathways and its atypical roles in comparison to the classical cyclins. Despite its functional uniqueness, CDK5 shares a great part of its structural similarity with other members of the cyclin-dependent kinase family. After its discovery 26 years ago, a progressive set of cellular functions has been associated with this protein kinase, ranging from neuronal migration, axonal guidance, and synaptic plasticity in diverse stages of brain development, including specific and complex cognitive functions. More than 30 substrates for CDK5 have been found in different cellular pathways. Together with its essential physiological roles, a major discovery was the finding twenty years ago that CDK5 participates in neurodegenerative diseases responsible for tau hyperphosphorylations, and, as a consequence, it becomes a neurotoxic factor. This review focuses on the wide roles of CDK5 in the central nervous system, its implications in neurodegeneration, and provides an integrative insight of its involvement in pain modulation, Alzheimer's disease, and other contexts.
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Quinasa 5 Dependiente de la Ciclina/fisiología , Fenómenos Fisiológicos del Sistema Nervioso , Enfermedad de Alzheimer/etiología , Enfermedad de Alzheimer/metabolismo , Animales , Quinasa 5 Dependiente de la Ciclina/metabolismo , Quinasas Ciclina-Dependientes , Humanos , Sistema Nervioso/metabolismo , Dolor/metabolismoRESUMEN
The Proliferating Cell Nuclear Antigen, PCNA, has roles in both G1 and S phases of the cell cycle. Here we show that maize PCNA can be found in cells in structures of a trimer or a dimer of trimer, in complexes of high molecular mass that change in size as germination proceeds, co-eluting with cell cycle proteins as CycD3;1 and CDKs (A/B1;1). Using different methodological strategies, we show that PCNA actually interacts with CycD3;1, CDKA, CDKB1;1, KRP1;1 and KRP4;1, all of which contain PIP or PIP-like motifs. Anti-PCNA immunoprecipitates show kinase activity that is inhibited by KRP1;1 and KRP4;2, indicating the formation of quaternary complexes PCNA-CycD/CDKs-KRPs in which PCNA would act as a platform. This inhibitory effect seems to be differential during the germination process, more pronounced as germination advances, suggesting a complex regulatory mechanism in which PCNA could bind different sets of cyclins/CDKs, some more susceptible to inhibition by KRPs than others.
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Quinasas Ciclina-Dependientes/metabolismo , Antígeno Nuclear de Célula en Proliferación/metabolismo , Zea mays/genética , Proteínas de Ciclo Celular/genética , Proteínas de Ciclo Celular/metabolismo , Quinasas Ciclina-Dependientes/genética , Ciclinas/genética , Ciclinas/metabolismo , Germinación , Fosforilación , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Antígeno Nuclear de Célula en Proliferación/genética , Zea mays/enzimología , Zea mays/fisiologíaRESUMEN
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
We report an unanticipated system of joint regulation by cyclin-dependent kinase (CDK) and mitogen-activated protein kinase (MAPK), involving collaborative multi-site phosphorylation of a single substrate. In budding yeast, the protein Ste5 controls signaling through a G1 arrest pathway. Upon cell-cycle entry, CDK inhibits Ste5 via multiple phosphorylation sites, disrupting its membrane association. Using quantitative time-lapse microscopy, we examined Ste5 membrane recruitment dynamics at different cell-cycle stages. Surprisingly, in S phase, where Ste5 recruitment should be blocked, we observed an initial recruitment followed by a steep drop-off. This delayed inhibition revealed a requirement for both CDK activity and negative feedback from the pathway MAPK Fus3. Mutagenesis, mass spectrometry, and electrophoretic analyses suggest that the CDK and MAPK modify shared sites, which are most extensively phosphorylated when both kinases are active and able to bind their docking sites on Ste5. Such collaborative phosphorylation can broaden regulatory inputs and diversify output dynamics of signaling pathways.