RESUMEN
BACKGROUND: Glucocorticoids (GCs) are widely used to treat inflammatory or autoimmune diseases. However, several studies have reported that the use of GCs can lead to numerous complications, the most serious of which are osteoporosis and osteonecrosis of the femoral head (ONFH). Osteoblast apoptosis has been identified as an important event in the development of GC-induced osteoporosis and ONFH. However, the mechanisms underlying the regulation of these processes have not yet been explored. OBJECTIVES: To observe the effect of dexamethasone (Dex) on the apoptosis of osteoblasts and explore its mechanism, as well as provide a new therapeutic idea for GCinduced osteoporosis and ONFH. MATERIAL AND METHODS: Cell proliferation and apoptosis of MC3T3-E1 cells after Dex treatment were determined using the CellTiter-Glo® Luminescent Cell Viability Assay kit and Annexin V-FITC/PI Double Staining Apoptosis Detection Kit, respectively. The expression of caspase-3/cleaved caspase-3 and poly(ADP-ribose) polymerase (PARP)/cleaved PARP in MC3T3-E1 cells after Dex treatment was determined with western blotting. The expression of p53 and checkpoint kinase 2 (Chk2) in MC3T3-E1 cells after Dex treatment was analyzed using western blotting and polymerase chain reaction (PCR). The effects of p53 knockdown and Chk2 knockdown on Dex-induced apoptosis of MC3T3-E1 cells were also characterized. RESULTS: Dexamethasone remarkably inhibited cell growth and induced the apoptosis of MC3T3-E1 cells. We also observed that Dex induced osteoblast apoptosis by promoting p53 expression. The regulatory effect of Dex on p53 expression is mediated by the upregulation of Chk2, which interacted with p53 and inhibited p53 degradation. The knockdown of p53 alleviated Dex-induced MC3T3-E1 cell apoptosis by decreasing the expression of cleaved caspase-3 and cleaved PARP. CONCLUSIONS: We demonstrated that Dex increased Chk2 protein expression, which stabilized the protein expression of p53, and in turn promoted osteoblast apoptosis.
Asunto(s)
Dexametasona , Osteoblastos , Osteoporosis , Humanos , Apoptosis , Caspasa 3/metabolismo , Caspasa 3/farmacología , Quinasa de Punto de Control 2/efectos de los fármacos , Quinasa de Punto de Control 2/metabolismo , Dexametasona/efectos adversos , Dexametasona/farmacología , Glucocorticoides/efectos adversos , Glucocorticoides/farmacología , Osteoblastos/efectos de los fármacos , Osteoblastos/metabolismo , Transducción de Señal , Proteína p53 Supresora de Tumor/efectos de los fármacos , Proteína p53 Supresora de Tumor/metabolismo , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Poli(ADP-Ribosa) Polimerasas/metabolismoRESUMEN
Novel PARP inhibitors with selective mode-of-action have been approved for clinical use. Herein, oxadiazole based ligands that are predicted to target PARP-1 have been synthesized and screened for the loss of cell viability in mammary carcinoma cells, wherein seven compounds were observed to possess significant IC50 values in the range of 1.4 to 25 µM. Furthermore, compound 5u, inhibited the viability of MCF-7 cells with an IC50 value of 1.4µM, when compared to Olaparib (IC50 = 3.2 µM). Compound 5s also decreased cell viability in MCF-7 and MDA-MB-231 cells with IC50 values of 15.3 and 19.2 µM, respectively. Treatment of MCF-7 cells with compounds 5u and 5s produced PARP cleavage, H2AX phosphorylation and CASPASE-3 activation comparable to that observed with Olaparib. Compounds 5u and 5s also decreased foci-formation and 3D Matrigel growth of MCF-7 cells equivalent to or greater than that observed with Olaparib. Finally, in silico analysis demonstrated binding of compound 5s towardsthe catalytic site of PARP-1, indicating that these novel oxadiazoles synthesized herein may serve as exemplars for the development of new therapeutics in cancer.
Asunto(s)
Diseño de Fármacos/métodos , Oxadiazoles/farmacología , Poli(ADP-Ribosa) Polimerasas/metabolismo , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Ensayos de Selección de Medicamentos Antitumorales , Femenino , Humanos , Células MCF-7 , Oxadiazoles/química , Poli(ADP-Ribosa) Polimerasa-1/efectos de los fármacos , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacosRESUMEN
Increased levels of reactive oxygen and nitrogen species play an important role in the development and progression of neurodegenerative diseases, such as Alzheimer's and Parkinson's disease. The overproduction of these highly reactive chemical species leads to DNA damage and subsequent activation of the poly(ADP-ribose)polymerase (PARP) enzyme. Several studies have demonstrated the potential use of PARP inhibitors for neuroprotection. We previously reported that the dual Src/Abl kinase inhibitor bosutinib (BOS) decreases PARP activity and acts as a chemosensitizer in cancer cells. In this study, we evaluated the neuroprotective potential of BOS with respect to its inhibitory effect on cellular poly(ADP-ribos)ylation (PARylation) using a 3-morpholinosydnonimine (SIN1)-mediated cellular toxicity model. Our data suggest that pretreatment with BOS, especially at lower doses, significantly decreased the level of SIN1-induced cellular PARylation. This regulation pattern of PARylation was found to be associated with the protective effect of BOS against SIN1 on the viability of retinoic acid-differentiated SH-SY5Y cells. Furthermore, while PARP-1 expression was decreased, phosphorylation of SAPK/JNK was not reverted at the observed neuroprotective doses of BOS. In conclusion, we suggest a novel mechanism for the neuroprotective effect of BOS involving the inhibition of cellular PARylation.
Asunto(s)
Proteínas Adaptadoras Transductoras de Señales/fisiología , Compuestos de Anilina/farmacología , Fármacos Neuroprotectores/farmacología , Nitrilos/farmacología , Poli ADP Ribosilación/efectos de los fármacos , Quinolinas/farmacología , Diferenciación Celular/efectos de los fármacos , Línea Celular Tumoral , Humanos , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Tretinoina/farmacologíaRESUMEN
Poly-ADP-ribose polymerase (PARP)-family ADP-ribosyltransferases function in various signaling pathways, predominantly in the nucleus and cytosol. Although PARP inhibitors are in clinical practice for cancer therapy, the enzymatic activities of individual PARP family members are yet insufficiently understood. We studied PARP10, a mono-ADP-ribosyltransferase and potential drug target. Using acid-urea gel electrophoresis, we found that the isolated catalytic domain of PARP10 auto-ADP-ribosylates (MARylates) at eight or more acceptor residues. We isolated individual species with either singular or several modifications and then analyzed them by mass spectrometry. The results confirmed multi-site MARylation in a random order and identified four acceptor residues. The mutagenesis of singular acceptor residues had a minor impact on the overall auto-MARylation level and no effect on the MARylation of histone H3.1. Together, our results suggest that PARP10 automodification may have functions in the regulation of intramolecular or partner binding events, rather than of its enzymatic catalysis. This contributes to a better understanding of PARP10 functions, and, in the long run, to gauging the consequences of PARP inhibitor actions.
Asunto(s)
ADP Ribosa Transferasas/metabolismo , Electroforesis , Histonas/metabolismo , Poli(ADP-Ribosa) Polimerasas/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , ADP Ribosa Transferasas/genética , Antineoplásicos/farmacología , Electroforesis/métodos , Humanos , Poli(ADP-Ribosa) Polimerasa-1/metabolismo , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Proteínas Proto-Oncogénicas/efectos de los fármacosRESUMEN
Breast cancer is the most common and significant cancers in females regarding the loss of life quality. Similar to other cancers, one of the etiologic factors in breast cancer is DNA damage. A plethora of molecules are responsible for sensing DNA damage and mediating actions which lead to DNA repair, senescence, cell cycle arrest and if damage is unbearable to apoptosis. In each of these, aberrations leading to unrepaired damage was resulted in uncontrolled proliferation and cancer. Another cellular function is autophagy defined as a process eliminating of unnecessary proteins in stress cases involved in pathogenesis of cancer. Knowing their role in cancer, scholars have tried to develop strategies in order to target DDR and autophagy. Further, the interactions of DDR and autophagy plus their regulatory role on each other have been focused simultaneously. The present review study has aimed to illustrate the importance of DDR and autophagy in breast cancer according to the related studies and uncover the relation between DDR and autophagy and its significance in breast cancer therapy.
Asunto(s)
Antineoplásicos/uso terapéutico , Neoplasias de la Mama/tratamiento farmacológico , Daño del ADN , Reparación del ADN , Transducción de Señal , Antineoplásicos/farmacología , Proteínas de la Ataxia Telangiectasia Mutada/antagonistas & inhibidores , Autofagia , Proteína BRCA1/antagonistas & inhibidores , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/fisiopatología , ADN/metabolismo , Femenino , Humanos , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Inhibidores de Poli(ADP-Ribosa) Polimerasas/uso terapéutico , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Inhibidores de Proteínas Quinasas/farmacología , Inhibidores de Proteínas Quinasas/uso terapéuticoRESUMEN
Inhibitors of poly-ADP-ribose polymerase 1 (PARPi) are highly effective in killing cells deficient in homologous recombination (HR); thus, PARPi have been clinically utilized to successfully treat BRCA2-mutant tumors. However, positive response to PARPi is not universal, even among patients with HR-deficiency. Here, we present the results of genome-wide CRISPR knockout and activation screens which reveal genetic determinants of PARPi response in wildtype or BRCA2-knockout cells. Strikingly, we report that depletion of the ubiquitin ligase HUWE1, or the histone acetyltransferase KAT5, top hits from our screens, robustly reverses the PARPi sensitivity caused by BRCA2-deficiency. We identify distinct mechanisms of resistance, in which HUWE1 loss increases RAD51 levels to partially restore HR, whereas KAT5 depletion rewires double strand break repair by promoting 53BP1 binding to double-strand breaks. Our work provides a comprehensive set of putative biomarkers that advance understanding of PARPi response, and identifies novel pathways of PARPi resistance in BRCA2-deficient cells.
Asunto(s)
Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas , Inhibidores de Poli(ADP-Ribosa) Polimerasas/aislamiento & purificación , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Proteína BRCA2/genética , Proteína BRCA2/metabolismo , Biomarcadores , Daño del ADN , Reparación del ADN , Técnicas de Inactivación de Genes , Células HeLa , Recombinación Homóloga/efectos de los fármacos , Humanos , Lisina Acetiltransferasa 5/metabolismo , Proteínas Mad2/metabolismo , Recombinasa Rad51/genética , Recombinasa Rad51/metabolismo , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/metabolismo , Proteína 1 de Unión al Supresor Tumoral P53 , Ubiquitina-Proteína Ligasas/genética , Ubiquitina-Proteína Ligasas/metabolismoRESUMEN
PURPOSE: PARP inhibitors (PARPi) are standard-of-care therapy for high-grade serous ovarian cancer (HGSOC). We investigated combining cediranib (antiangiogenic) with olaparib (PARPi) at emergence of PARPi resistance. PATIENTS AND METHODS: The proof-of-concept EVOLVE study (NCT02681237) assessed cediranib-olaparib combination therapy after progression on a PARPi. Women with HGSOC and radiographic evidence of disease progression were enrolled into one of three cohorts: platinum sensitive after PARPi; platinum resistant after PARPi; or progression on standard chemotherapy after progression on PARPi (exploratory cohort). Patients received olaparib tablets 300 mg twice daily with cediranib 20 mg once daily until progression or unacceptable toxicity. The coprimary endpoints were objective response rate (RECIST v1.1) and progression-free survival (PFS) at 16 weeks. Archival tissue (PARPi-naïve) and baseline biopsy (post-PARPi) samples were mandatory. Genomic mechanisms of resistance were assessed by whole-exome and RNA sequencing. RESULTS: Among 34 heavily pretreated patients, objective responses were observed in 0 of 11 (0%) platinum-sensitive patients, 2 of 10 (20%) platinum-resistant patients, and 1 of 13 (8%) in the exploratory cohort. Sixteen-week PFS rates were 55%, 50%, and 39%, respectively. The most common grade 3 toxicities were diarrhea (12%) and anemia (9%). Acquired genomic alterations at PARPi progression were reversion mutations in BRCA1, BRCA2, or RAD51B (19%); CCNE1 amplification (16%); ABCB1 upregulation (15%); and SLFN11 downregulation (7%). Patients with reversion mutations in homologous recombination genes and/or ABCB1 upregulation had poor outcomes. CONCLUSIONS: This is currently the largest post-PARPi study identifying genomic mechanisms of resistance to PARPis. In this setting, the activity of cediranib-olaparib varied according to the PARPi resistance mechanism.
Asunto(s)
Neoplasias Ováricas/tratamiento farmacológico , Ftalazinas/administración & dosificación , Piperazinas/administración & dosificación , Poli(ADP-Ribosa) Polimerasas/genética , Quinazolinas/administración & dosificación , Anciano , Protocolos de Quimioterapia Combinada Antineoplásica/administración & dosificación , Protocolos de Quimioterapia Combinada Antineoplásica/efectos adversos , Proteína BRCA1/genética , Proteína BRCA2/genética , Resistencia a Antineoplásicos/genética , Femenino , Humanos , Persona de Mediana Edad , Recurrencia Local de Neoplasia/tratamiento farmacológico , Recurrencia Local de Neoplasia/genética , Recurrencia Local de Neoplasia/patología , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Ftalazinas/efectos adversos , Piperazinas/efectos adversos , Inhibidores de Poli(ADP-Ribosa) Polimerasas/administración & dosificación , Inhibidores de Poli(ADP-Ribosa) Polimerasas/efectos adversos , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Supervivencia sin Progresión , Quinazolinas/efectos adversosRESUMEN
Poly (ADP-ribose) polymerase (PARP) inhibitors have transformed the therapeutic management of solid tumors, particularly ovarian cancer. Initially studied in BRCA deficient tumors, the Food and Drug Administration (FDA) indications have expanded to include other homologous recombination deficient tumors as well as biomarker-wildtype tumors. They have also gained momentum not only as a treatment strategy, but as a maintenance strategy as well. While PARP inhibitors were initially ev aluated in the recurrent setting, they have now moved to frontline therapy. This review will discuss the current FDA indications of the clinically available PARP inhibitors for treatment and maintenance therapies. We will then review the recently completed and ongoing clinical trials which may inform future clinical approvals.
Asunto(s)
Antineoplásicos/administración & dosificación , Neoplasias Ováricas/tratamiento farmacológico , Inhibidores de Poli(ADP-Ribosa) Polimerasas/administración & dosificación , Antineoplásicos/farmacología , Biomarcadores de Tumor/metabolismo , Femenino , Humanos , Recurrencia Local de Neoplasia , Neoplasias Ováricas/patología , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Poli(ADP-Ribosa) Polimerasas/metabolismoRESUMEN
Mono(ADP-ribosylation) (MARylation) and poly(ADP-ribosylation) (PARylation) are posttranslational modifications found on multiple amino acids. There are 12 enzymatically active mono(ADP-ribose) polymerase (monoPARP) enzymes and 4 enzymatically active poly(ADP-ribose) polymerase (polyPARP) enzymes that use nicotinamide adenine dinucleotide (NAD+) as the ADP-ribose donating substrate to generate these modifications. While there are approved drugs and clinical trials ongoing for the enzymes that perform PARylation, MARylation is gaining recognition for its role in immune function, inflammation, and cancer. However, there is a lack of chemical probes to study the function of monoPARPs in cells and in vivo. An important first step to generating chemical probes for monoPARPs is to develop biochemical assays to enable hit finding, and determination of the potency and selectivity of inhibitors. Complicating the development of enzymatic assays is that it is poorly understood how monoPARPs engage their substrates. To overcome this, we have developed a family-wide approach to developing robust high-throughput monoPARP assays where the enzymes are immobilized and forced to self-modify using biotinylated-NAD+, which is detected using a dissociation-enhanced lanthanide fluorescence immunoassay (DELFIA) readout. Herein we describe the development of assays for 12 monoPARPs and 3 polyPARPs and apply them to understand the potency and selectivity of a focused library of inhibitors across this family.
Asunto(s)
ADP Ribosa Transferasas/antagonistas & inhibidores , Inhibidores Enzimáticos/aislamiento & purificación , Ensayos Analíticos de Alto Rendimiento , Inhibidores de Poli(ADP-Ribosa) Polimerasas/aislamiento & purificación , Procesamiento Proteico-Postraduccional/genética , ADP Ribosa Transferasas/química , ADP Ribosa Transferasas/genética , ADP-Ribosilación/genética , Adenosina Difosfato Ribosa/genética , Inhibidores Enzimáticos/farmacología , Humanos , NAD/química , Poli ADP Ribosilación/genética , Inhibidores de Poli(ADP-Ribosa) Polimerasas/química , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Poli(ADP-Ribosa) Polimerasas/genética , Especificidad por SustratoRESUMEN
Introduction: The implementation of poly-ADP-ribose polymerase (PARP) inhibitors for therapy has created potential treatments for a wide spectrum of malignancies involving DNA damage repair gene abnormalities. PARPs are a group of enzymes that are responsible for detecting and repairing DNA damage and therefore play a key role in maintaining cell function and integrity. PARP inhibitors are drugs that target DNA repair deficiencies. Inhibiting PARP activity in cancer cells causes cell death. Areas covered: This review summarizes the role of PARP inhibitors in the treatment of cancer. We performed a systematic literature search in February 2019 in the electronic databases PubMed and EMBASE. Our search terms were the following: PARP, PARP inhibitors, PARPi, Poly ADP ribose polymerase, cancer treatment. We discuss PARP inhibitors currently being investigated in cancer clinical trials, their safety profiles, clinical resistance, combined therapeutic approaches and future challenges. Expert Opinion: The future could bring novel PARP inhibitors with greater DNA trapping potential, better safety profiles and improved combined therapies involving hormonal, chemo-, radio- or immunotherapies. Progress may afford wider indications for PARP inhibitors in the treatment of cancer and the utilization for cancer prevention in high-risk mutation carriers. Research efforts should focus on identifying novel drugs that target DNA repair deficiencies.
Asunto(s)
Terapia Molecular Dirigida , Neoplasias/tratamiento farmacológico , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Animales , Antineoplásicos/efectos adversos , Antineoplásicos/farmacología , Daño del ADN/efectos de los fármacos , Reparación del ADN/efectos de los fármacos , Humanos , Neoplasias/patología , Inhibidores de Poli(ADP-Ribosa) Polimerasas/efectos adversos , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Poli(ADP-Ribosa) Polimerasas/metabolismoRESUMEN
ADP-ribosylation is a unique posttranslational modification catalyzed by poly(ADP-ribose) polymerases (PARPs) using NAD+ as ADP-ribose donor. PARPs play an indispensable role in DNA damage repair and small molecule PARP inhibitors have emerged as potent anticancer drugs. However, to date, PARP inhibitor treatment has been restricted to patients with BRCA1/2 mutation-associated breast and ovarian cancer. One of the major challenges to extend the therapeutic potential of PARP inhibitors to other cancer types is the absence of predictive biomarkers. Here, we show that ovarian cancer cells with higher level of NADP+, an NAD+ derivative, are more sensitive to PARP inhibitors. We demonstrate that NADP+ acts as a negative regulator and suppresses ADP-ribosylation both in vitro and in vivo. NADP+ impairs ADP-ribosylation-dependent DNA damage repair and sensitizes tumor cell to chemically synthesized PARP inhibitors. Taken together, our study identifies NADP+ as an endogenous PARP inhibitor that may have implications in cancer treatment.
Asunto(s)
Antineoplásicos/farmacología , Daño del ADN/efectos de los fármacos , NADP/antagonistas & inhibidores , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , ADP-Ribosilación , Animales , Biomarcadores , Línea Celular Tumoral/efectos de los fármacos , Reparación del ADN , Proteínas del Grupo de Complementación de la Anemia de Fanconi/genética , Femenino , Humanos , Ratones , NAD/farmacología , Neoplasias Ováricas , Fosfotransferasas (Aceptor de Grupo Alcohol)/efectos de los fármacos , Poli ADP Ribosilación/efectos de los fármacos , ARN Helicasas/genéticaRESUMEN
Poly(ADP-ribose)polymerase, member 14 (PARP14, alternatively named ARTD8, BAL2, and COAST6) is an intracellular mono(ADP-ribosyl) transferase. PARP14 transfers a negatively charged ADP-ribose unit from a donor NAD+ molecule onto a target protein, post-translationally. PARP14's domain architecture consists of three macrodomains (Macro1, Macro2 and Macro3), a WWE domain and an ARTD (or catalytic domain). The Macro2 and Macro3 domains bind ADPribose (ADPr) with high affinity, whereas the WWE domain stabilizes the protein structure by binding to ADPr derivatives. The catalytic domain is involved in binding the NAD+ and catalyzing the mono- ADP-ribosylation reaction. PARP14 has been identified as a possible anti-cancer and antiinflammatory target. Acting as a transcriptional co-activator for STAT6, PARP14 acts to promote the over activation of the Th2 immune response, thus promoting the metabolic change to an anaerobic state (Warburg effect) and activation of cell survival pathways through JNK2 and the PGI/AMF complex. These changes are consistent with the metabolic sophistication observed in cancer, and the immune imbalance in inflammatory diseases. Current literature on selective and unselective PARP14 inhibitors are reviewed and discussed. Although there is no evidence that selective PARP inhibitors would be advantageous we have proposed some strategies for future design of selective PARP14 inhibitors.
Asunto(s)
Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Animales , Dominio Catalítico , Humanos , NAD/metabolismo , Poli(ADP-Ribosa) Polimerasas/química , Poli(ADP-Ribosa) Polimerasas/metabolismo , Relación Estructura-ActividadRESUMEN
Poly(ADP-ribose) polymerase inhibitors (PARPi) are selectively active in cells with homologous recombination (HR) deficiency (HRD) caused by mutations in BRCA1, BRCA2, and other pathway members. We sought small molecules that induce HRD in HR-competent cells to induce synthetic lethality with PARPi and extend the utility of PARPi. We demonstrated that inhibition of bromodomain containing 4 (BRD4) induced HRD and sensitized cells across multiple tumor lineages to PARPi regardless of BRCA1/2, TP53, RAS, or BRAF mutation status through depletion of the DNA double-stand break resection protein CtIP (C-terminal binding protein interacting protein). Importantly, BRD4 inhibitor (BRD4i) treatment reversed multiple mechanisms of resistance to PARPi. Furthermore, PARPi and BRD4i are synergistic in multiple in vivo models.
Asunto(s)
Recombinación Homóloga/genética , Proteínas Nucleares/antagonistas & inhibidores , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Factores de Transcripción/antagonistas & inhibidores , Animales , Proteínas de Ciclo Celular , Línea Celular Tumoral , Daño del ADN/efectos de los fármacos , Reparación del ADN/efectos de los fármacos , Reparación del ADN/genética , Femenino , Recombinación Homóloga/efectos de los fármacos , Humanos , Ratones Transgénicos , Mutación/efectos de los fármacos , Mutación/genética , Proteínas Nucleares/genética , Neoplasias Ováricas/tratamiento farmacológico , Neoplasias Ováricas/genética , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Poli(ADP-Ribosa) Polimerasas/genética , Factores de Transcripción/genéticaRESUMEN
The polyadenosine-diphosphate-ribose polymerase 14 (PARP14) has been implicated in DNA damage response pathways for homologous recombination. PARP14 contains three (ADP ribose binding) macrodomains (MD) whose exact contribution to overall PARP14 function in pathology remains unclear. A medium throughput screen led to the identification of N-(2(-9H-carbazol-1-yl)phenyl)acetamide (GeA-69, 1) as a novel allosteric PARP14 MD2 (second MD of PARP14) inhibitor. We herein report medicinal chemistry around this novel chemotype to afford a sub-micromolar PARP14 MD2 inhibitor. This chemical series provides a novel starting point for further development of PARP14 chemical probes.
Asunto(s)
Cisteína Endopeptidasas/química , Descubrimiento de Drogas , Inhibidores de Poli(ADP-Ribosa) Polimerasas/síntesis química , Poli(ADP-Ribosa) Polimerasas/química , Regulación Alostérica , Carbazoles/química , Humanos , Concentración 50 Inhibidora , Modelos Biológicos , Simulación del Acoplamiento Molecular , Estructura Molecular , Inhibidores de Poli(ADP-Ribosa) Polimerasas/química , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Relación Estructura-ActividadRESUMEN
The PARP-2 selective inhibitor is important for clarifying specific roles of PARP-2 in the pathophysiological process and developing desired drugs with reduced off-target side effects. In this work, a series of novel quinazoline-2,4(1H,3H)-dione derivatives was designed and synthesized to explore isoform selective PARP inhibitors. As a result, compound 11a (PARP-1 IC50=467nM, PARP-2 IC50=11.5nM, selectivity PARP-1/PARP-2=40.6) was disclosed as the most selective PARP-2 inhibitor with high potency to date. The binding features of compound 11a within PARP-1 and PARP-2 were investigated respectively to provide useful insights for the further construction of new isoform selective inhibitors of PARP-1 and PARP-2 by using CDOCKER program.
Asunto(s)
Descubrimiento de Drogas , Inhibidores Enzimáticos/química , Inhibidores Enzimáticos/farmacología , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Quinazolinas/química , Quinazolinas/farmacología , Espectroscopía de Resonancia Magnética con Carbono-13 , Humanos , Concentración 50 Inhibidora , Modelos Moleculares , Espectroscopía de Protones por Resonancia Magnética , Espectrometría de Masa por Ionización de ElectrosprayRESUMEN
The present study investigated the protective effect of diethylcarbamazine in inhibiting nuclear factor (NF)-κB activation in isoproterenolinduced acute myocardial infarction (AMI) rats through the poly ADP ribose polymerase (PARP) pathway. Male albino Wistar rats were injected subcutaneously with isoproterenol (100 mg/kg/day) for 2 days to induce an AMI model. Diethylcarbamazine (50 mg/kg) was administered by gavage for 12 days prior to the isoproterenol-induced AMI. It was noted that diethylcarbamazine significantly inhibited AMIinduced casein kinase and lactate dehydrogenase levels, and reduced the AMIinduced wet heart weight to body weight ratio in AMI rats. Diethylcarbamazine treatment significantly weakened reactive oxygen species production and reduced the levels of tumor necrosis factor (TNF)α, interleukin6 and NFκB/p65 in AMI rats. Western blotting demonstrated that diethylcarbamazine significantly suppressed the AMIinduced inducible nitric oxide synthase (iNOS), transforming growth factor (TGF)ß1, cyclooxygenase2 (COX2) and PARP protein expression in AMI rats. The results demonstrated that the protective effect of diethylcarbamazine inhibited isoproterenolinduced AMI through the suppression of inflammation, iNOS, TGFß1, COX2 and the PARP pathway, and revealed the clinical potential of diethylcarbamazine for therapeutic and clinical applications.
Asunto(s)
Dietilcarbamazina/antagonistas & inhibidores , Dietilcarbamazina/uso terapéutico , Isoproterenol/farmacología , Infarto del Miocardio/tratamiento farmacológico , FN-kappa B/antagonistas & inhibidores , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Poli(ADP-Ribosa) Polimerasas/metabolismo , Animales , Caseína Quinasas/metabolismo , Ciclooxigenasa 2/metabolismo , Dietilcarbamazina/administración & dosificación , Modelos Animales de Enfermedad , Proteínas I-kappa B/metabolismo , Interleucina-6/metabolismo , L-Lactato Deshidrogenasa/metabolismo , Masculino , Infarto del Miocardio/inducido químicamente , Infarto del Miocardio/metabolismo , Óxido Nítrico Sintasa de Tipo II/metabolismo , Ratas , Ratas Wistar , Especies Reactivas de Oxígeno/metabolismo , Factor de Crecimiento Transformador beta1/metabolismo , Factor de Necrosis Tumoral alfa/metabolismoRESUMEN
Poly(ADP-ribose) polymerase (PARP) enzyme contributes to nephropathy, a serious diabetic complication which may lead to end-stage renal disease. The study aims to investigate the effect of PARP over-activation on kidney functions in a type 2 diabetic rat model. The study also tests the therapeutic use of PARP inhibitors in diabetic nephropathy. Type 2 diabetes was induced in adult male rats by high-fructose/high-fat diet and low streptozotocin dose. Then, the PARP inhibitor 4-aminobenzamide (4-AB) was administered daily for 10 weeks. At the end, urine samples were collected to measure urine creatinine, albumin, and total proteins. PARP activity, superoxide dismutase (SOD) activity, and nitrite content were measured in kidney tissue homogenate. Glucose, fructosamine, insulin, and tumor necrosis factor-alpha (TNF-α) were measured in serum. Furthermore, histological studies, collagen deposition, and immunofluorescence of nuclear factor kappa B (NFκB) and transforming growth factor beta1 (TGF-ß1) were carried out. PARP enzyme activity was significantly higher in the diabetic group and was significantly reduced by 4-AB administration. Diabetic animals had clear nephropathy indicated by proteinuria and increased albumin excretion rate (AER) which were significantly decreased by PARP inhibition. In addition, PARP inhibition increased creatinine clearance in diabetic animals and reduced renal TGF-ß1 and glomerular fibrosis. Moreover, PARP inhibition alleviated the elevated serum TNF-α level, renal NFκB, nitrite, and the decrease in SOD activity in diabetic animals. However, PARP inhibition did not significantly affect neither hyperglycemia nor insulin sensitivity. PARP enzyme inhibition alleviates diabetic nephropathy through decreasing inflammation, oxidative stress, and renal fibrosis.
Asunto(s)
Diabetes Mellitus Experimental/tratamiento farmacológico , Diabetes Mellitus Tipo 2/tratamiento farmacológico , Nefropatías Diabéticas/tratamiento farmacológico , Inhibidores de Poli(ADP-Ribosa) Polimerasas/farmacología , Animales , Benzamidas/farmacología , Diabetes Mellitus Experimental/complicaciones , Diabetes Mellitus Tipo 2/complicaciones , Nefropatías Diabéticas/patología , Dieta Alta en Grasa , Fibrosis/tratamiento farmacológico , Fibrosis/patología , Inflamación/tratamiento farmacológico , Inflamación/patología , Masculino , FN-kappa B/metabolismo , Estrés Oxidativo/efectos de los fármacos , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Poli(ADP-Ribosa) Polimerasas/metabolismo , Ratas , Ratas Sprague-Dawley , Estreptozocina/toxicidad , Superóxido Dismutasa/metabolismo , Factor de Necrosis Tumoral alfa/sangre , para-Aminobenzoatos/farmacologíaRESUMEN
BACKGROUND: Nucleophosmin 1 (NPM1) is a nucleocytoplasmic shuttling protein mainly localized in the nucleolus. NPM1 is frequently mutated in acute myeloid leukemia (AML). NPM1c oligomerizes with wild-type nucleophosmin 1 (wt-NPM1), and this leads to its continuous cytoplasmic delocalization and contributes to leukemogenesis. Recent studies have shown that Cytoplasmic NPM1 (NPM1c) degradation leads to growth arrest and apoptosis of NPM1c AML cells and corrects wt-NPM1 normal nucleolar localization. METHODS: AML cells expressing wt-NPM1 or NPM1c or transfected with wt-NPM1 or NPM1c as well as wt-NPM1 and NPM1c AML xenograft mice were used. Cell growth was assessed with trypan blue or a CellTiter 96 proliferation kit. The cell cycle was studied with a propidium iodide (PI) assay. Caspase-mediated intrinsic apoptosis was assessed with annexin V/PI, the mitochondrial membrane potential, and poly(adenosine diphosphate ribose) polymerase cleavage. The expression of NPM1, p53, phosphorylated p53, and p21 was analyzed via immunoblotting. Localization was performed with confocal microscopy. The leukemia burden was evaluated by flow cytometry with an anti-human CD45 antibody. RESULTS: The imidazoquinoxaline 1-(3-methoxyphenyl)-N-methylimidazo[1,2-a]quinoxalin-4-amine (EAPB0503) induced selective proteasome-mediated degradation of NPM1c, restored wt-NPM1 nucleolar localization in NPM1c AML cells, and thus yielded selective growth arrest and apoptosis. Introducing NPM1c to cells normally harboring wt-NPM1 sensitized them to EAPB0503 and led to their growth arrest. Moreover, EAPB0503 selectively reduced the leukemia burden in NPM1c AML xenograft mice. CONCLUSIONS: These findings further reinforce the idea of targeting the NPM1c oncoprotein to eradicate leukemic cells and warrant a broader preclinical evaluation and then a clinical evaluation of this promising drug. Cancer 2017;123:1662-1673. © 2017 American Cancer Society.
Asunto(s)
Antineoplásicos/farmacología , Apoptosis/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Leucemia Mieloide Aguda/tratamiento farmacológico , Proteínas Mutantes/efectos de los fármacos , Proteínas Nucleares/efectos de los fármacos , Quinoxalinas/farmacología , Animales , Anexina A5/efectos de los fármacos , Anexina A5/metabolismo , Línea Celular Tumoral , Nucléolo Celular/metabolismo , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/efectos de los fármacos , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/metabolismo , Citoplasma/metabolismo , Citometría de Flujo , Humanos , Immunoblotting , Leucemia Mieloide Aguda/genética , Ratones , Microscopía Confocal , Proteínas Mutantes/metabolismo , Mutación , Proteínas Nucleares/genética , Proteínas Nucleares/metabolismo , Nucleofosmina , Fosforilación/efectos de los fármacos , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Proteína p53 Supresora de Tumor/efectos de los fármacos , Proteína p53 Supresora de Tumor/metabolismo , Ensayos Antitumor por Modelo de XenoinjertoRESUMEN
Baicalein (BA), one of the major compounds isolated from the root of Scutellaria baicalensis Gerogi, exhibits various pharmacological effects, such as anti-oxidant, anti-inflammatory, and anticancer effects. In this study, we found that BA reduced cell viability and increased apoptosis in ovarian cancer cells. Treatment of cells with BA enhanced microtubule-associated protein light chain 3-II (LC3-II) expression, acidic vesicular organelle and GFP-LC3 fluorescence dot accumulation. Combined treatment with chloroquine and BA apparently reduced cell viability and increased the cleavage of poly (ADPribose) polymerase (PARP) in both HEY and A2780 ovarian cancer cell lines, indicating that BA induces a protective autophagy in these cells. Knockdown of Beclin 1 by siRNA remarkably decreased BA-induced LC3-II lipidation. In addition, we found an increase in the phosphorylation of extracellular signal-regulated kinase (ERK, Thr202/Thr204) and AKT (Ser473) after BA treatment, and inhibition of ERK activation by the pharmacological inhibitor U0126 or ERK siRNA blocked BA-induced autophagy. Taken together, these results suggest that BA induces Beclin 1- and ERK-dependent autophagy in ovarian cancer cells.
Asunto(s)
Autofagia/efectos de los fármacos , Beclina-1/efectos de los fármacos , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Inhibidores Enzimáticos/farmacología , Flavanonas/farmacología , Sistema de Señalización de MAP Quinasas/efectos de los fármacos , Proteínas Asociadas a Microtúbulos/efectos de los fármacos , Antimaláricos/farmacología , Beclina-1/metabolismo , Western Blotting , Línea Celular Tumoral , Cloroquina/farmacología , Femenino , Técnicas de Silenciamiento del Gen , Humanos , Técnicas In Vitro , Microscopía Confocal , Microscopía Fluorescente , Proteínas Asociadas a Microtúbulos/metabolismo , Neoplasias Ováricas , Poli(ADP-Ribosa) Polimerasas/efectos de los fármacos , Poli(ADP-Ribosa) Polimerasas/metabolismo , ARN Interferente PequeñoRESUMEN
OBJECTIVE: Cisplatin is commonly used to treat solid tumors. However, permanent hearing loss is a major side effect of cisplatin chemotherapy and often results in dose reduction of the cisplatin chemotherapy. Peanut sprouts show cytoprotective properties owing to their antioxidant activities. This study was designed to investigate the effect of peanut sprout extract (PSE) on cisplatin-induced ototoxicity in an auditory cell line, HEI-OC1 cells. METHODS: Cells were exposed to cisplatin for 24 h, with or without pre-treatment with PSE, cell viability was examined using the MTT assay. Apoptotic cells were identified by double staining with Hoechst 33258 and propidium iodide. Western blot analysis was performed to examine apoptotic proteins including C-PARP and C-caspase, anti-apoptotic protein Bcl-2, and Nrf2 redox system activation. Mitochondrial reactive oxygen species (ROS) were investigated to examine whether PSE could scavenge cisplatin-induced ROS. Real-time PCR analyses were performed to investigate the mRNA levels of antioxidant enzymes including NQO1, HO-1, GPx2, Gclc, and catalase. RESULTS: The cisplatin-treated group showed reduced cell viability, increased apoptotic properties and markers, and increased ROS levels. PSE pre-treatment before cisplatin exposure significantly increased cell viability and reduced apoptotic properties and ROS production. These effects resulted from the up-regulation of antioxidant genes, including NQO1, HO-1, GPx2, Gclc, and catalase through Akt phosphorylation and Nrf2 activation. CONCLUSION: Our results demonstrate that PSE protects from cisplatin-induced cytotoxicity by activating the antioxidant effects via the Akt/Nrf-2 pathway in this auditory cell line, and indicate that PSE may provide novel treatment to prevent cisplatin-induced ototoxicity.