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1.
Nat Commun ; 11(1): 4296, 2020 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-32855387

RESUMO

Assays to study cancer cell responses to pharmacologic or genetic perturbations are typically restricted to using simple phenotypic readouts such as proliferation rate. Information-rich assays, such as gene-expression profiling, have generally not permitted efficient profiling of a given perturbation across multiple cellular contexts. Here, we develop MIX-Seq, a method for multiplexed transcriptional profiling of post-perturbation responses across a mixture of samples with single-cell resolution, using SNP-based computational demultiplexing of single-cell RNA-sequencing data. We show that MIX-Seq can be used to profile responses to chemical or genetic perturbations across pools of 100 or more cancer cell lines. We combine it with Cell Hashing to further multiplex additional experimental conditions, such as post-treatment time points or drug doses. Analyzing the high-content readout of scRNA-seq reveals both shared and context-specific transcriptional response components that can identify drug mechanism of action and enable prediction of long-term cell viability from short-term transcriptional responses to treatment.


Assuntos
Perfilação da Expressão Gênica/métodos , Neoplasias/genética , Análise de Célula Única/métodos , Antineoplásicos/farmacologia , Sequência de Bases , Linhagem Celular Tumoral , Sobrevivência Celular/genética , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Modelos Estatísticos , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Polimorfismo de Nucleotídeo Único , Piridonas/farmacologia , Pirimidinonas/farmacologia
2.
Adv Exp Med Biol ; 1207: 569-579, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32671775

RESUMO

Pulmonary fibrosis is a progressive chronic inflammatory disease with a poor clinical outcome. Although pirfenidone and nintedanib have been approved by FDA to treat idiopathic pulmonary fibrosis (IPF), these drugs can only slow the progression of IPF. Autophagy plays an important role in the pathogenesis of pulmonary fibrosis. Whether the autophagic flux is blocked or not is directly related to the development direction of pulmonary fibrosis. Defining how autophagy activity regulates the pathogenesis of pulmonary fibrosis will greatly advance the progression of pulmonary fibrosis therapy.


Assuntos
Autofagia , Fibrose Pulmonar , Progressão da Doença , Humanos , Indóis/farmacologia , Indóis/uso terapêutico , Fibrose Pulmonar/tratamento farmacológico , Piridonas/farmacologia , Piridonas/uso terapêutico
3.
Prostate ; 80(12): 917-925, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32569423

RESUMO

BACKGROUND: Chronic prostatitis/chronic pelvic pain syndrome (CP/CPPS) is an intractable problem of the urogenital system. The aetiopathogenesis and effective treatments for CP/CPPS are needed to be untangled. Pirfenidone is a molecule that exhibits anti-inflammatory, antifibrotic, and antioxidative stress capacities in a variety of animal experiments and clinical trials. This study was aimed to investigate the therapeutic effect of pirfenidone on CP/CPPS and to identify the mechanism responsible for it. METHODS: A CP/CPPS model was induced in rats by intraprostatic injection of complete Freund's adjuvant (CFA). Blood and prostatic tissues were harvested for assessment after the administration of pirfenidone or vehicle for 4 weeks. RESULTS: The findings revealed that pirfenidone significantly ameliorated chronic pelvic pain and inhibited prostatic inflammation and fibrosis. Further study found that pirfenidone suppressed the expression of proinflammatory mediators, including tumor necrosis factor-α, interleukin-1ß (IL-1ß), IL-6, IL-8. Pirfenidone exhibited a potent antioxidant capacity through improving the activities of glutathione, catalase, total superoxide dismutase, and reducing the production of malondialdehyde. Furthermore, pirfenidone also facilitated the polarization of M2 macrophages and suppressed the activation of the nuclear factor-κB (NF-κB) signaling pathway. CONCLUSIONS: Pirfenidone can exert a beneficial effect against CFA-induced CP/CPPS by anti-inflammatory, antioxidative, antifibrotic properties, and the function is mediated at least partly through the M2 polarization of macrophages and the inhibition of NF-κB signaling pathway. These findings suggest that pirfenidone holds promise as a potential therapeutic for the treatment of CP/CPPS.


Assuntos
Dor Crônica/tratamento farmacológico , Dor Pélvica/tratamento farmacológico , Prostatite/tratamento farmacológico , Piridonas/farmacologia , Animais , Anti-Inflamatórios não Esteroides/farmacologia , Antioxidantes/farmacologia , Polaridade Celular/efeitos dos fármacos , Doença Crônica/tratamento farmacológico , Dor Crônica/metabolismo , Dor Crônica/patologia , Citocinas/metabolismo , Modelos Animais de Doenças , Mediadores da Inflamação/antagonistas & inibidores , Mediadores da Inflamação/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Macrófagos/efeitos dos fármacos , Macrófagos/patologia , Masculino , NF-kappa B/metabolismo , Dor Pélvica/metabolismo , Dor Pélvica/patologia , Fosforilação/efeitos dos fármacos , Prostatite/metabolismo , Prostatite/patologia , Ratos , Síndrome
4.
Eur J Pharmacol ; 882: 173237, 2020 Sep 05.
Artigo em Inglês | MEDLINE | ID: mdl-32525005

RESUMO

Pirfenidone (PFD), a pyridone compound, is well recognized as an antifibrotic agent tailored for the treatment of idiopathic pulmonary fibrosis. Recently, through its anti-inflammatory and anti-oxidant effects, PFD based clinical trial has also been launched for the treatment of coronavirus disease (COVID-19). To what extent this drug can perturb membrane ion currents remains largely unknown. Herein, the exposure to PFD was observed to depress the amplitude of hyperpolarization-activated cation current (Ih) in combination with a considerable slowing in the activation time of the current in pituitary GH3 cells. In the continued presence of ivabradine or zatebradine, subsequent application of PFD decreased Ih amplitude further. The presence of PFD resulted in a leftward shift in Ih activation curve without changes in the gating charge. The addition of this compound also led to a reduction in area of voltage-dependent hysteresis evoked by long-lasting inverted triangular (downsloping and upsloping) ramp pulse. Neither the amplitude of M-type nor erg-mediated K+ current was altered by its presence. In whole-cell potential recordings, addition of PFD reduced the firing frequency, and this effect was accompanied by the depression in the amplitude of sag voltage elicited by hyperpolarizing current stimulus. Overall, this study highlights evidence that PFD is capable of perturbing specific ionic currents, revealing a potential additional impact on functional activities of different excitable cells.


Assuntos
Membrana Celular/efeitos dos fármacos , Infecções por Coronavirus/tratamento farmacológico , Pneumonia Viral/tratamento farmacológico , Piridonas/farmacologia , Animais , Betacoronavirus/metabolismo , Cátions/metabolismo , Linhagem Celular Tumoral , Membrana Celular/metabolismo , Infecções por Coronavirus/virologia , Humanos , Canais Iônicos/efeitos dos fármacos , Canais Iônicos/metabolismo , Transporte de Íons/efeitos dos fármacos , Potenciais da Membrana/efeitos dos fármacos , Pandemias , Pneumonia Viral/virologia , Potássio/metabolismo , Piridonas/uso terapêutico , Ratos , Sódio/metabolismo
5.
Life Sci ; 256: 117963, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32535080

RESUMO

The new Coronavirus (SARS-CoV-2) is the cause of a serious infection in the respiratory tract called COVID-19. Structures of the main protease of SARS-CoV-2 (Mpro), responsible for the replication of the virus, have been solved and quickly made available, thus allowing the design of compounds that could interact with this protease and thus to prevent the progression of the disease by avoiding the viral peptide to be cleaved, so that smaller viral proteins can be released into the host's plasma. These structural data are extremely important for in silico design and development of compounds as well, being possible to quick and effectively identify potential inhibitors addressed to such enzyme's structure. Therefore, in order to identify potential inhibitors for Mpro, we used virtual screening approaches based with the structure of the enzyme and two compounds libraries, targeted to SARS-CoV-2, containing compounds with predicted activity against Mpro. In this way, we selected, through docking studies, the 100 top-ranked compounds, which followed to subsequent studies of pharmacokinetic and toxicity predictions. After all the simulations and predictions here performed, we obtained 10 top-ranked compounds that were again in silico analyzed inside the Mpro catalytic site, together some drugs that are being currently investigated for treatment of COVID-19. After proposing and analyzing the interaction modes of these compounds, we submitted one molecule then selected as template to a 2D similarity study in a database containing drugs approved by FDA and we have found and indicated Apixaban as a potential drug for future treatment of COVID-19.


Assuntos
Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Infecções por Coronavirus/tratamento farmacológico , Desenho de Fármacos , Pneumonia Viral/tratamento farmacológico , Antivirais/efeitos adversos , Antivirais/farmacocinética , Betacoronavirus/isolamento & purificação , Simulação por Computador , Infecções por Coronavirus/virologia , Desenvolvimento de Medicamentos , Reposicionamento de Medicamentos , Humanos , Simulação de Acoplamento Molecular , Pandemias , Pneumonia Viral/virologia , Pirazóis/farmacologia , Piridonas/farmacologia
6.
Life Sci ; 256: 117917, 2020 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-32525001

RESUMO

AIMS: Methamphetamine (METH) is an abused psychostimulant causing public health concern worldwide. While most studies have focused on the neurotoxic effects of METH, METH-induced cerebrovascular dysfunction has recently drawn attention as an important facet of METH-related pathophysiology. In this study, we investigated the protective role of GKT136901, a NOX1/4 inhibitor, against METH-induced blood-brain barrier (BBB) dysfunction. MAIN METHODS: Primary human brain microvascular endothelial cells (HBMECs) were used as an in vitro BBB model. HBMECs were treated with GKT136901, followed by METH exposure for 24 h. The generation of reactive oxidative species (ROS) was measured using 2',7'-dichlorofluorescin diacetate (DCF-DA) staining. To examine the BBB function, paracellular permeability of HBMEC monolayer was measured using FITC-labeled dextran. To evaluate structural properties of BBB in HBMECs, tight junction (TJ), adherent junction (AJ), and cytoskeletal proteins were stained and analyzed by confocal microscopy. KEY FINDINGS: METH treatment rapidly increased ROS generation in HBMECs but GKT136901 treatment inhibited METH-induced ROS generation. Although METH increased the permeability of HBMEC monolayer, this effect was abolished upon GKT136901 treatment. Following METH exposure, the proteins Zonula occludens-1 (ZO-1) and vascular endothelial cadherin (VE-cadherin) were translocalized from the cell membrane to the cytoplasm, thereby destroying intercellular tight junction (TJ) and adherent junction (AJ) structures, which were ameliorated upon GKT136901 treatment. METH exposure altered the cellular morphology of HBMECs and induced stress fiber formation. However, GKT136901 prevented METH-induced morphological and cytoskeletal changes in HBMECs. SIGNIFICANCE: These results suggest that GKT136901 prevents METH-induced BBB dysfunction in HBMECs through the inhibition of ROS generation.


Assuntos
Barreira Hematoencefálica/efeitos dos fármacos , Células Endoteliais/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Metanfetamina/efeitos adversos , NADPH Oxidases/antagonistas & inibidores , Substâncias Protetoras/farmacologia , Pirazóis/farmacologia , Piridonas/farmacologia , Antígenos CD/metabolismo , Barreira Hematoencefálica/citologia , Caderinas/metabolismo , Capilares/citologia , Permeabilidade Capilar , Descoberta de Drogas , Humanos , Espécies Reativas de Oxigênio/metabolismo , Junções Íntimas/metabolismo , Proteína da Zônula de Oclusão-1/metabolismo
7.
Proc Natl Acad Sci U S A ; 117(27): 15846-15851, 2020 07 07.
Artigo em Inglês | MEDLINE | ID: mdl-32561648

RESUMO

Combination use of BRAF V600E inhibitor dabrafenib and MEK inhibitor trametinib has become a standard treatment for human cancers harboring BRAF V600E. Its anticancer efficacies vary, however, with dramatic efficacy in some patients and drug resistance/tumor recurrence in others, which is poorly understood. Using thyroid cancer, melanoma, and colon cancer cell models, we showed that dabrafenib and trametinib induced robust apoptosis of cancer cells harboring both BRAF V600E and TERT promoter mutations but had little proapoptotic effect in cells harboring only BRAF V600E. Correspondingly, the inhibitors nearly completely abolished the growth of in vivo tumors harboring both mutations but had little effect on tumors harboring only BRAF V600E. Upon drug withdrawal, tumors harboring both mutations remained hardly measurable but tumors harboring only BRAF V600E regrew rapidly. BRAF V600E/MAP kinase pathway is known to robustly activate mutant promoter of TERT, a strong apoptosis suppressor. Thus, for survival, cancer cells harboring both mutations may have evolved to rely on BRAF V600E-promoted and high-TERT expression-mediated suppression of apoptosis. As such, inhibition of BRAF/MEK can trigger strong apoptosis-induced cell death and hence tumor abolishment. This does not happen in cells harboring only BRAF V600E as they have not developed reliance on TERT-mediated suppression of apoptosis due to the lack of mutant promoter-driven high-TERT expression. TERT promoter mutation governs BRAF-mutant cancer cells' apoptotic and hence therapeutic responses to BRAF/MEK inhibitors. Thus, the genetic duet of BRAF V600E and TERT promoter mutation represents an Achilles Heel for effective therapeutic targeting and response prediction in cancer.


Assuntos
Apoptose/efeitos dos fármacos , Mutação , Regiões Promotoras Genéticas/efeitos dos fármacos , Regiões Promotoras Genéticas/genética , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas B-raf/efeitos dos fármacos , Proteínas Proto-Oncogênicas B-raf/genética , Telomerase/genética , Animais , Morte Celular , Linhagem Celular Tumoral , Neoplasias do Colo , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Feminino , Humanos , Imidazóis/farmacologia , Melanoma/genética , Camundongos Nus , Proteínas Quinases Ativadas por Mitógeno/metabolismo , Oximas/farmacologia , Piridonas/farmacologia , Pirimidinonas/farmacologia , Neoplasias da Glândula Tireoide/genética , Ensaios Antitumorais Modelo de Xenoenxerto
8.
J Physiol Pharmacol ; 71(1)2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-32350148

RESUMO

Data concerning the impact of direct oral anticoagulants (DOACs) on the thromboelastography (TEG) indices in venousthromboembolism (VTE) patients are limited. The goal of this study was to compare the impact of DOACs on clot properties measured in whole blood using TEG kaolin test and in plasma obtained from real-life VTE patients. We assessed 53 patients, including 20 on rivaroxaban, 20 on apixaban, and 13 on dabigatran. Using the TEG® 5000, coagulation status was evaluated in whole blood samples, while plasma fibrin clot permeability (Ks) and its susceptibility to lysis (CLT) were assessed in citrated plasma. Plasma concentrations of rivaroxaban (99 [48 - 311] ng/ml) and apixaban (85 [40 - 105] ng/ml) were positively associated with Ks and inversely with CLT, while dabigatran concentrations (71 [39 - 98] ng/ml) correlated positively with Ks. Moreover, Ks was associated with clot formation times (R and K), time to maximum clot strength (TMA), coagulation index (CI) reflecting the overall coagulation status, and whole blood clot lysis time (TEG-CLT). Blood clot lysis index (CL30) was associated with plasma CLT in all patients on DOACs, while TMA correlated with CLT only in patients on apixaban. Higher DOAC concentrations were associated with longer retardation of whole blood clot formation and longer time needed to reach maximum level of its strength as well as with more permeable clots. We concluded that plasma fibrin clot properties correlate with corresponding TEG indices suggesting that TEG might be helpful in providing prognostic information about DOAC influence in VTE patients.


Assuntos
Anticoagulantes/farmacologia , Coagulação Sanguínea/efeitos dos fármacos , Tromboembolia Venosa/fisiopatologia , Administração Oral , Adulto , Anticoagulantes/uso terapêutico , Dabigatrana/farmacologia , Dabigatrana/uso terapêutico , Feminino , Fibrina/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Pirazóis/farmacologia , Pirazóis/uso terapêutico , Piridonas/farmacologia , Piridonas/uso terapêutico , Rivaroxabana/farmacologia , Rivaroxabana/uso terapêutico , Tromboelastografia/efeitos dos fármacos , Tromboembolia Venosa/tratamento farmacológico
9.
Nat Commun ; 11(1): 1923, 2020 04 21.
Artigo em Inglês | MEDLINE | ID: mdl-32317663

RESUMO

Drug resistance mediated by clonal evolution is arguably the biggest problem in cancer therapy today. However, evolving resistance to one drug may come at a cost of decreased fecundity or increased sensitivity to another drug. These evolutionary trade-offs can be exploited using 'evolutionary steering' to control the tumour population and delay resistance. However, recapitulating cancer evolutionary dynamics experimentally remains challenging. Here, we present an approach for evolutionary steering based on a combination of single-cell barcoding, large populations of 108-109 cells grown without re-plating, longitudinal non-destructive monitoring of cancer clones, and mathematical modelling of tumour evolution. We demonstrate evolutionary steering in a lung cancer model, showing that it shifts the clonal composition of the tumour in our favour, leading to collateral sensitivity and proliferative costs. Genomic profiling revealed some of the mechanisms that drive evolved sensitivity. This approach allows modelling evolutionary steering strategies that can potentially control treatment resistance.


Assuntos
Antineoplásicos/farmacologia , Resistencia a Medicamentos Antineoplásicos , Evolução Molecular , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Evolução Clonal , Biologia Computacional , Simulação por Computador , Gefitinibe/farmacologia , Genótipo , Humanos , Neoplasias Pulmonares/patologia , Modelos Teóricos , Medicina Molecular , Piridonas/farmacologia , Pirimidinonas/farmacologia , Processos Estocásticos
10.
Pediatr Blood Cancer ; 67(6): e28267, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32307821

RESUMO

BACKGROUND: The treatment of high-risk neuroblastoma continues to present a formidable challenge to pediatric oncology. Previous studies have shown that Bromodomain and extraterminal (BET) inhibitors can inhibit MYCN expression and suppress MYCN-amplified neuroblastoma in vivo. Furthermore, alterations within RAS-MAPK (mitogen-activated protein kinase) signaling play significant roles in neuroblastoma initiation, maintenance, and relapse, and mitogen-activated extracellular signal-regulated kinase (MEK) inhibitors demonstrate efficacy in subsets of neuroblastoma preclinical models. Finally, hyperactivation of RAS-MAPK signaling has been shown to promote resistance to BET inhibitors. Therefore, we examined the antitumor efficacy of combined BET/MEK inhibition utilizing I-BET726 or I-BET762 and trametinib in high-risk neuroblastoma. PROCEDURE: Utilizing a panel of genomically annotated neuroblastoma cell line models, we investigated the in vitro effects of combined BET/MEK inhibition on cell proliferation and apoptosis. Furthermore, we evaluated the effects of combined inhibition in neuroblastoma xenograft models. RESULTS: Combined BET and MEK inhibition demonstrated synergistic effects on the growth and survival of a large panel of neuroblastoma cell lines through augmentation of apoptosis. A combination therapy slowed tumor growth in a non-MYCN-amplified, NRAS-mutated neuroblastoma xenograft model, but had no efficacy in an MYCN-amplified model harboring a loss-of-function mutation in NF1. CONCLUSIONS: Combinatorial BET and MEK inhibition was synergistic in the vast majority of neuroblastoma cell lines in the in vitro setting but showed limited antitumor activity in vivo. Collectively, these data do not support clinical development of this combination in high-risk neuroblastoma.


Assuntos
Antineoplásicos/farmacologia , Benzodiazepinas/farmacologia , MAP Quinase Quinase 1/antagonistas & inibidores , Neuroblastoma/tratamento farmacológico , Proteínas/antagonistas & inibidores , Piridonas/farmacologia , Pirimidinonas/farmacologia , Animais , Apoptose , Proliferação de Células , Feminino , Humanos , Camundongos , Camundongos SCID , Neuroblastoma/metabolismo , Neuroblastoma/patologia , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
11.
J Med Chem ; 63(8): 3956-3975, 2020 04 23.
Artigo em Inglês | MEDLINE | ID: mdl-32208600

RESUMO

The bromodomain and extra-terminal (BET) family proteins have recently emerged as promising drug targets for cancer therapy. In this study, identification of an 8-methyl-pyrrolo[1,2-a]pyrazin-1(2H)-one fragment (47) as a new binder to the BET bromodomains and the subsequent incorporation of fragment 47 to the scaffold of ABBV-075, which recently entered Phase I clinical trials, enabled the generation of a series of highly potent BET bromodomain inhibitors. Further druggability optimization led to the discovery of compound 38 as a potential preclinical candidate. Significantly, compared with ABBV-075, which exhibits a 63-fold selectivity for BRD4(1) over EP300, compound 38 demonstrates an excellent selectivity for the BET bromodomain family over other bromodomains, with an ∼1500-fold selectivity for BRD4(1) over EP300. Orally administered 38 achieves a complete inhibition of tumor growth with a tumor growth inhibition (TGI) of 99.7% accompanied by good tolerability.


Assuntos
Acetanilidas/química , Acetanilidas/farmacologia , Proteínas de Ciclo Celular/antagonistas & inibidores , Descoberta de Drogas/métodos , Compostos Heterocíclicos com 3 Anéis/química , Compostos Heterocíclicos com 3 Anéis/farmacologia , Piridonas/química , Piridonas/farmacologia , Sulfonamidas/química , Sulfonamidas/farmacologia , Fatores de Transcrição/antagonistas & inibidores , Animais , Proteínas de Ciclo Celular/metabolismo , Cães , Relação Dose-Resposta a Droga , Proteína p300 Associada a E1A/antagonistas & inibidores , Proteína p300 Associada a E1A/metabolismo , Haplorrinos , Humanos , Camundongos , Microssomos Hepáticos/efeitos dos fármacos , Microssomos Hepáticos/metabolismo , Domínios Proteicos/efeitos dos fármacos , Domínios Proteicos/fisiologia , Estrutura Secundária de Proteína , Ratos , Relação Estrutura-Atividade , Fatores de Transcrição/metabolismo
12.
Int J Oncol ; 56(3): 761-771, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32124956

RESUMO

Pancreatic cancer is a lethal solid malignancy with limited therapeutic options. The development of novel therapeutic drugs requires adequate new cell line models. A new pancreatic cancer cell line, designated PDXPC1, was established from one pancreatic ductal adenocarcinoma (PDAC) patient­derived xenograft. The PDXPC1 cells were stably cultured for >2 years and had a stable short tandem repeat profile. The PDXPC1 cell line retained the key mutations of the primary tumor, along with the epithelial origin and other important protein expression. The PDXPC1 cells induced rapid in vivo tumor growth, both subcutaneously and orthotopically, in a mouse model with an elevated CA199 level. The PDXPC1 cells showed weak growth, invasion and migration potency compared to another pancreatic cancer cell line, but were relatively resistant to multiple anti­cancer drugs. Interestingly, the MEK inhibitor trametinib significantly inhibited the proliferation of PDXPC1 cells, and not that of Panc­1 cells, by inactivating MEK/ERK/MYC signaling and activating the apoptotic pathway via Bcl­2 degradation. In conclusion, the PDXPC1 cell line, capturing the major characteristics of the primary tumor, may be a suitable tool for studying the underlying mechanisms of chemo­resistance in PDAC and developing new targeted therapeutic options.


Assuntos
Carcinoma Ductal Pancreático/tratamento farmacológico , Técnicas de Cultura de Células/métodos , Neoplasias Pancreáticas/tratamento farmacológico , Inibidores de Proteínas Quinases/administração & dosagem , Piridonas/administração & dosagem , Pirimidinonas/administração & dosagem , Animais , Carcinoma Ductal Pancreático/genética , Carcinoma Ductal Pancreático/patologia , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Feminino , Humanos , Camundongos , Mutação , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/patologia , Inibidores de Proteínas Quinases/farmacologia , Piridonas/farmacologia , Pirimidinonas/farmacologia , Transdução de Sinais/efeitos dos fármacos , Células Tumorais Cultivadas , Ensaios Antitumorais Modelo de Xenoenxerto
13.
Artigo em Chinês | MEDLINE | ID: mdl-32062902

RESUMO

Pulmonary fibrosis is the terminal manifestation of a variety of interstitial lung diseases. Idiopathic pulmonary fibrosis (IPF) is one of the chronic, progressive, fibrotic lung disease with high incidence and poor prognosis. Nintedanib and pirfenidone are currently marketed anti-pulmonary fibrosis drugs, and their efficacy and safety are recognized in patients with IPF. This article reviews the targets and clinical trials of the two drugs, and provides a basis for the expansion of indications for anti-pulmonary fibrosis drugs.


Assuntos
Indóis/farmacologia , Piridonas/farmacologia , Ensaios Clínicos Controlados Aleatórios como Assunto , Humanos
14.
Ann Rheum Dis ; 79(4): 507-517, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32041748

RESUMO

BACKGROUND: Systemic sclerosis (SSc) is characterised by autoimmune activation, tissue and vascular fibrosis in the skin and internal organs. Tissue fibrosis is driven by myofibroblasts, that are known to maintain their phenotype in vitro, which is associated with epigenetically driven trimethylation of lysine 27 of histone 3 (H3K27me3). METHODS: Full-thickness skin biopsies were surgically obtained from the forearms of 12 adult patients with SSc of recent onset. Fibroblasts were isolated and cultured in monolayers and protein and RNA extracted. HOX transcript antisense RNA (HOTAIR) was expressed in healthy dermal fibroblasts by lentiviral induction employing a vector containing the specific sequence. Gamma secretase inhibitors were employed to block Notch signalling. Enhancer of zeste 2 (EZH2) was blocked with GSK126 inhibitor. RESULTS: SSc myofibroblasts in vitro and SSc skin biopsies in vivo display high levels of HOTAIR, a scaffold long non-coding RNA known to direct the histone methyltransferase EZH2 to induce H3K27me3 in specific target genes. Overexpression of HOTAIR in dermal fibroblasts induced EZH2-dependent increase in collagen and α-SMA expression in vitro, as well as repression of miRNA-34A expression and consequent NOTCH pathway activation. Consistent with these findings, we show that SSc dermal fibroblast display decreased levels of miRNA-34a in vitro. Further, EZH2 inhibition rescued miRNA-34a levels and mitigated the profibrotic phenotype of both SSc and HOTAIR overexpressing fibroblasts in vitro. CONCLUSIONS: Our data indicate that the EZH2-dependent epigenetic phenotype of myofibroblasts is driven by HOTAIR and is linked to miRNA-34a repression-dependent activation of NOTCH signalling.


Assuntos
Proteína Potenciadora do Homólogo 2 de Zeste/metabolismo , Fibroblastos/metabolismo , MicroRNAs/metabolismo , Miofibroblastos/metabolismo , RNA Longo não Codificante/metabolismo , Receptores Notch/metabolismo , Escleroderma Sistêmico/metabolismo , Secretases da Proteína Precursora do Amiloide/antagonistas & inibidores , Epigênese Genética , Fibrose , Código das Histonas , Humanos , Indóis/farmacologia , Fenótipo , Piridonas/farmacologia , Receptores Notch/antagonistas & inibidores , Escleroderma Sistêmico/genética , Escleroderma Sistêmico/patologia , Transdução de Sinais , Pele/citologia , Pele/metabolismo , Pele/patologia
15.
Chem Pharm Bull (Tokyo) ; 68(1): 77-90, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31902903

RESUMO

Novel 3,5-dimethylpyridin-4(1H)-one scaffold compounds were synthesized and evaluated as AMP-activated protein kinase (AMPK) activators. Unlike direct AMPK activators, this series of compounds showed selective cell growth inhibitory activity against human breast cancer cell lines. By optimizing the lead compound (4a) from our library, 2-[({1'-[(4-fluorophenyl)methyl]-2-methyl-1',2',3',6'-tetrahydro[3,4'-bipyridin]-6-yl}oxy)methyl]-3,5-dimethylpyridin-4(1H)-one (25) was found to have potent AMPK activating activity. Compound 25 also showed good aqueous solubility while maintaining the unique selectivity in cell growth inhibitory activity.


Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Piridonas/química , Proteínas Quinases Ativadas por AMP/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos , Humanos , Piridonas/síntese química , Piridonas/farmacologia , Solubilidade , Relação Estrutura-Atividade
16.
Mol Immunol ; 119: 35-45, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31962268

RESUMO

Loss of CD58 is a common mechanism for tumor immune evasion in lymphoid malignancies. CD58 loss is known to occur due to both genetic and non-genetic causes; therefore, we hypothesized that restoring CD58 expression in lymphoma cells may be an effective treatment approach. To explore the potential for restoring CD58 expression, we first screened 11 B-cell lymphoma lines and found that 3 had decreased CD58 expression. Among these, CD58 was genetically damaged in two lines but not in the third line. Using the cell line with downregulated CD58 without a genetic abnormality, we performed epigenetic library screening and found that two EZH2 inhibitors, EPZ6438 and GSK126, specifically enhanced CD58 expression. By examining the effect of three EZH2 inhibitors with different selectivity profiles in different B-cell lines, EZH2 inhibition was shown to have a common activity in upregulating CD58 expression. Restoring the expression of CD58 in lymphoma cells using an EZH2 inhibitor was shown to enhance interferon-γ production of T and NK cells against lymphoma cells. H3K27 was shown to be highly trimethylated in the CD58 promoter region, and EZH2 inhibition induced its demethylation and activated transcription of the CD58 gene. These results indicated that EZH2 is involved in the epigenetic silencing of CD58 in lymphoma cells as a mechanism for tumor immune escape, and EZH2 inhibitors are able to restore epigenetically suppressed CD58 expression. Our findings provide a molecular basis for the combination of an EZH2 inhibitor and immunotherapy for lymphoma treatment.


Assuntos
Antineoplásicos/farmacologia , Benzamidas/farmacologia , Antígenos CD58/genética , Proteína Potenciadora do Homólogo 2 de Zeste/antagonistas & inibidores , Epigênese Genética , Indóis/farmacologia , Linfoma de Células B/genética , Piridonas/farmacologia , Antígenos CD58/biossíntese , Antígenos CD58/imunologia , Linhagem Celular Tumoral , Epigênese Genética/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Inativação Gênica , Humanos , Ativação Linfocitária , Linfoma de Células B/tratamento farmacológico , Linfoma de Células B/imunologia , Linfócitos T/imunologia
17.
Am J Respir Cell Mol Biol ; 62(4): 413-422, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-31967851

RESUMO

Pirfenidone (PFD) is a pharmacological compound with therapeutic efficacy in idiopathic pulmonary fibrosis. It has been chiefly characterized as an antifibrotic agent, although it was initially developed as an antiinflammatory compound because of its ability to diminish the accumulation of inflammatory cells and cytokines. Despite recent studies that have elucidated key mechanisms, the precise molecular activities of PFD remain incompletely understood. PFD modulates fibrogenic growth factors, thereby attenuating fibroblast proliferation, myofibroblast differentiation, collagen and fibronectin synthesis, and deposition of extracellular matrix. This effect is mediated by suppression of TGF-ß1 (transforming growth factor-ß1) and other growth factors. Here, we appraise the impact of PFD on TGF-ß1 production and its downstream pathways. Accumulating evidence indicates that PFD also downregulates inflammatory pathways and therefore has considerable potential as a viable and innovative antiinflammatory compound. We examine the effects of PFD on inflammatory cells and the production of pro- and antiinflammatory cytokines in the lung. In this context, recent evidence that PFD can target inflammasome pathways and ensuing lung inflammation is highlighted. Finally, the antioxidant properties of PFD, such as its ability to inhibit redox reactions and regulate oxidative stress-related genes and enzymes, are detailed. In summary, this narrative review examines molecular mechanisms underpinning PFD and its recognized benefits in lung fibrosis. We highlight preclinical data that demonstrate the potential of PFD as a nonsteroidal antiinflammatory agent and outline areas for future research.


Assuntos
Pneumopatias/tratamento farmacológico , Piridonas/farmacologia , Piridonas/uso terapêutico , Animais , Anti-Inflamatórios não Esteroides/farmacologia , Anti-Inflamatórios não Esteroides/uso terapêutico , Proliferação de Células/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Humanos , Pulmão/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos
18.
Expert Opin Ther Pat ; 30(3): 195-208, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31944142

RESUMO

Introduction: Studies presented in the patent applications demonstrate that a new integrase strand transfer inhibitor cabotegravir might be used as long-acting antiretroviral formulation or delivery system that reduces dosing frequency and may therefore increase adherence and thus pre-exposure prophylaxis (PrEP) and treatment efficacy against HIV. As announced in 2019, the developer ViiV Healthcare seeks US and EU approval of long-acting, injectable HIV treatment.Area covered: This review covers all the patent applications published until October 2019 with cabotegravir in the examples or claim section of the patent application document. The patent applications cover drug substance synthesis, solid-state forms, therapeutic applications, in vitro and in vivo efficacy as well as the potential formulations of cabotegravir alone or in combination with other anti-HIV agents.Expert opinion: The results from multiple clinical studies suggest that cabotegravir can be used as PrEP agent and treatment agent against HIV. Multiple studies use cabotegravir in combination with other anti-HIV agents such as rilpivirine. Cabotegravir in combination with rilpivirine is an interesting therapeutic, due to the possibility of formulating long-acting formulation with dosing interval of every 4 weeks or less, thus reducing daily pill burden and improving patient's compliance.


Assuntos
Fármacos Anti-HIV/administração & dosagem , Infecções por HIV/tratamento farmacológico , Piridonas/administração & dosagem , Animais , Fármacos Anti-HIV/farmacologia , Quimioterapia Combinada , Humanos , Adesão à Medicação , Patentes como Assunto , Profilaxia Pré-Exposição , Piridonas/farmacologia , Rilpivirina/administração & dosagem
19.
Am J Physiol Renal Physiol ; 318(1): F183-F192, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31760771

RESUMO

Angiotensin II exacerbates oxidative stress in part by increasing superoxide (O2-) production by many renal tissues. However, whether it does so in proximal tubules and the source of O2- in this segment are unknown. Dietary fructose enhances the stimulatory effect of angiotensin II on proximal tubule Na+ reabsorption, but whether this is true for oxidative stress is unknown. We hypothesized that angiotensin II causes proximal nephron oxidative stress in part by stimulating NADPH oxidase (NOX)4-dependent O2- production and decreasing the amount of the antioxidant glutathione, and this is exacerbated by dietary fructose. We measured basal and angiotensin II-stimulated O2- production with and without inhibitors, NOX1 and NOX4 expression, and total and reduced glutathione (GSH) in proximal tubules from rats drinking either tap water (control) or 20% fructose. Angiotensin II (10 nM) increased O2- production by 113 ± 42 relative light units·mg protein-1·s-1 in controls and 401 ± 74 relative light units·mg protein-1·s-1 with 20% fructose (n = 11 for each group, P < 0.05 vs. control). Apocynin and the Nox1/4 inhibitor GKT136901 prevented angiotensin II-induced increases in both groups. NOX4 expression was not different between groups. NOX1 expression was undetectable. Angiotensin II decreased GSH by 1.8 ± 0.8 nmol/mg protein in controls and by 4.2 ± 0.9 nmol/mg protein with 20% fructose (n = 18 for each group, P < 0.047 vs. control). We conclude that 1) angiotensin II causes oxidative stress in proximal tubules by increasing O2- production by NOX4 and decreasing GSH and 2) dietary fructose enhances the ability of angiotensin II to stimulate O2- and diminish GSH, thereby exacerbating oxidative stress in this segment.


Assuntos
Angiotensina II/farmacologia , Glutationa/metabolismo , Túbulos Renais Proximais/efeitos dos fármacos , Superóxidos/metabolismo , Acetofenonas/farmacologia , Animais , Antioxidantes/farmacologia , Açúcares da Dieta , Frutose , Túbulos Renais Proximais/metabolismo , Masculino , NADPH Oxidases/metabolismo , Néfrons/efeitos dos fármacos , Néfrons/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Pirazóis/farmacologia , Piridonas/farmacologia , Ratos , Ratos Sprague-Dawley
20.
Eur J Med Chem ; 186: 111855, 2020 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-31740051

RESUMO

There is an increasing interest in the field of cancer therapy for small compounds targeting pyrimidine biosynthesis, and in particular dihydroorotate dehydrogenase (DHODH), the fourth enzyme of this metabolic pathway. Three available DHODH structures, featuring three different known inhibitors, were used as templates to screen in silico an original chemical library from Erevan University. This process led to the identification of P1788, a compound chemically related to the alkaloid cerpegin, as a new class of pyrimidine biosynthesis inhibitors. In line with previous reports, we investigated the effect of P1788 on the cellular innate immune response. Here we show that pyrimidine depletion by P1788 amplifies cellular response to both type-I and type II interferons, but also induces DNA damage as assessed by γH2AX staining. Moreover, the addition of inhibitors of the DNA damage response led to the suppression of the P1788 stimulatory effects on the interferon pathway. This demonstrates that components of the DNA damage response are bridging the inhibition of pyrimidine biosynthesis by P1788 to the interferon signaling pathway. Altogether, these results provide new insights on the mode of action of novel pyrimidine biosynthesis inhibitors and their development for cancer therapies.


Assuntos
Furanos/farmacologia , Piridinas/farmacologia , Piridonas/farmacologia , Pirimidinas/antagonistas & inibidores , Células Cultivadas , Dano ao DNA , Relação Dose-Resposta a Droga , Furanos/síntese química , Furanos/química , Células HEK293 , Humanos , Modelos Moleculares , Estrutura Molecular , Piridinas/síntese química , Piridinas/química , Piridonas/química , Pirimidinas/biossíntese , Relação Estrutura-Atividade
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