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1.
Yonsei Med J ; 61(9): 762-773, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32882760

RESUMO

PURPOSE: Pharmacological inhibition of mutant isocitrate dehydrogenase (IDH) reduces R-2-hydroxyglutarate (2-HG) levels and restores cellular differentiation in vivo and in vitro. The IDH2 inhibitor enasidenib (AG-221) has been approved by the FDA as a first-in-class inhibitor for the treatment of relapsed or refractory (R/R) IDH2-mutant acute myeloid leukemia (AML). In this study, the effects of a combination of all-trans retinoic acid (ATRA) and AG-221 on AML cell differentiation was explored, along with the mechanisms employed by IDH2-mutant cells in AML. MATERIALS AND METHODS: We treated the human AML cell line, IDH2-mutant-TF-1, and primary human AML cells carrying IDH2 mutation with 30 µM AG-221 and 100 nM ATRA, alone or in combination. RESULTS: Combined treatment with AG-221 and ATRA inhibited 2-HG production and resulted in synergistic effects on differentiation among IDH2-mutant AML cells and primary AML cells expressing IDH2 mutation. Combined treatment with AG-221 and ATRA altered autophagic activity. AG-221 and ATRA treatment-induced differentiation of IDH2-mutant AML cells was associated with autophagy induction, without suppressing autophagy flux at maturation and degradation stages. A RAF-1/MEK/ERK pathway was founded to be associated with AG-221 and ATRA-induced differentiation in IDH2-mutant AML cells. IDH-associated changes in histone methylation markers decreased after AG-221 and ATRA combination treatment. CONCLUSION: Our preliminary evidence indicates that the addition of ATRA to treatments with IDH2 inhibitor may lead to further improvements or increases in response rates in IDH2-mutant AML patients who do not appear to benefit from treatments with IDH2 inhibitor alone.


Assuntos
Aminopiridinas/farmacologia , Antineoplásicos/farmacologia , Diferenciação Celular/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Isocitrato Desidrogenase/genética , Leucemia Mieloide Aguda/tratamento farmacológico , Tretinoína/farmacologia , Triazinas/farmacologia , Resistencia a Medicamentos Antineoplásicos/genética , Humanos , Isocitrato Desidrogenase/antagonistas & inibidores , Isocitrato Desidrogenase/metabolismo , Leucemia Mieloide Aguda/genética , Mutação
2.
Zhonghua Wei Chang Wai Ke Za Zhi ; 23(9): 911-916, 2020 Sep 25.
Artigo em Chinês | MEDLINE | ID: mdl-32927518

RESUMO

Imatinib has created the era of precise treatment of gastrointestinal stromal tumor (GIST). However, in recent years, there has been a lack of satisfactory drugs for imatinib resistance. Recently, the emergence of a series of new agents not only brings new hope to break this situation, but also verifies that the drug development of GIST treatment still needs to lock the driving gene as the main target. New drugs, such as avapritinib and ripretinib, have shown surprising therapeutic efficacy, which may lead to a new situation in the treatment of GIST. The precise treatment of GIST guided by different primary gene mutations and secondary drug resistance mutations will replace the traditional guidelines based on the number of treatment lines.


Assuntos
Antineoplásicos/uso terapêutico , Resistencia a Medicamentos Antineoplásicos/genética , Tumores do Estroma Gastrointestinal/tratamento farmacológico , Tumores do Estroma Gastrointestinal/genética , Antineoplásicos/farmacologia , Humanos , Mesilato de Imatinib/farmacologia , Mesilato de Imatinib/uso terapêutico , Naftiridinas/farmacologia , Naftiridinas/uso terapêutico , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Pirazóis/farmacologia , Pirazóis/uso terapêutico , Pirróis/farmacologia , Pirróis/uso terapêutico , Triazinas/farmacologia , Triazinas/uso terapêutico , Ureia/análogos & derivados , Ureia/farmacologia , Ureia/uso terapêutico
3.
Proc Natl Acad Sci U S A ; 117(34): 20803-20813, 2020 08 25.
Artigo em Inglês | MEDLINE | ID: mdl-32764148

RESUMO

Virus entry is a multistep process. It initiates when the virus attaches to the host cell and ends when the viral contents reach the cytosol. Genetically unrelated viruses can subvert analogous subcellular mechanisms and use similar trafficking pathways for successful entry. Antiviral strategies targeting early steps of infection are therefore appealing, particularly when the probability for successful interference through a common step is highest. We describe here potent inhibitory effects on content release and infection by chimeric vesicular stomatitis virus (VSV) containing the envelope proteins of Zaire ebolavirus (VSV-ZEBOV) or severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) (VSV-SARS-CoV-2) elicited by Apilimod and Vacuolin-1, small-molecule inhibitors of the main endosomal phosphatidylinositol-3-phosphate/phosphatidylinositol 5-kinase, PIKfyve. We also describe potent inhibition of SARS-CoV-2 strain 2019-nCoV/USA-WA1/2020 by Apilimod. These results define tools for studying the intracellular trafficking of pathogens elicited by inhibition of PIKfyve kinase and suggest the potential for targeting this kinase in developing small-molecule antivirals against SARS-CoV-2.


Assuntos
Betacoronavirus/efeitos dos fármacos , Ebolavirus/efeitos dos fármacos , Compostos Heterocíclicos de 4 ou mais Anéis/farmacologia , Morfolinas/farmacologia , Fosfatidilinositol 3-Quinases , Triazinas/farmacologia , Internalização do Vírus/efeitos dos fármacos , Animais , Betacoronavirus/fisiologia , Células Cultivadas , Infecções por Coronavirus , Ebolavirus/fisiologia , Edição de Genes , Humanos , Pandemias , Inibidores de Fosfoinositídeo-3 Quinase/farmacologia , Pneumonia Viral , Proteínas do Envelope Viral/genética
4.
Nature ; 586(7827): 113-119, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32707573

RESUMO

The emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in 2019 has triggered an ongoing global pandemic of the severe pneumonia-like disease coronavirus disease 2019 (COVID-19)1. The development of a vaccine is likely to take at least 12-18 months, and the typical timeline for approval of a new antiviral therapeutic agent can exceed 10 years. Thus, repurposing of known drugs could substantially accelerate the deployment of new therapies for COVID-19. Here we profiled a library of drugs encompassing approximately 12,000 clinical-stage or Food and Drug Administration (FDA)-approved small molecules to identify candidate therapeutic drugs for COVID-19. We report the identification of 100 molecules that inhibit viral replication of SARS-CoV-2, including 21 drugs that exhibit dose-response relationships. Of these, thirteen were found to harbour effective concentrations commensurate with probable achievable therapeutic doses in patients, including the PIKfyve kinase inhibitor apilimod2-4 and the cysteine protease inhibitors MDL-28170, Z LVG CHN2, VBY-825 and ONO 5334. Notably, MDL-28170, ONO 5334 and apilimod were found to antagonize viral replication in human pneumocyte-like cells derived from induced pluripotent stem cells, and apilimod also demonstrated antiviral efficacy in a primary human lung explant model. Since most of the molecules identified in this study have already advanced into the clinic, their known pharmacological and human safety profiles will enable accelerated preclinical and clinical evaluation of these drugs for the treatment of COVID-19.


Assuntos
Antivirais/análise , Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Infecções por Coronavirus/tratamento farmacológico , Infecções por Coronavirus/virologia , Avaliação Pré-Clínica de Medicamentos , Reposicionamento de Medicamentos , Pneumonia Viral/tratamento farmacológico , Pneumonia Viral/virologia , Monofosfato de Adenosina/análogos & derivados , Monofosfato de Adenosina/farmacologia , Alanina/análogos & derivados , Alanina/farmacologia , Células Epiteliais Alveolares/citologia , Células Epiteliais Alveolares/efeitos dos fármacos , Betacoronavirus/crescimento & desenvolvimento , Linhagem Celular , Inibidores de Cisteína Proteinase/análise , Inibidores de Cisteína Proteinase/farmacologia , Relação Dose-Resposta a Droga , Sinergismo Farmacológico , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Modelos Biológicos , Morfolinas/análise , Morfolinas/farmacologia , Pandemias , Reprodutibilidade dos Testes , Bibliotecas de Moléculas Pequenas/análise , Bibliotecas de Moléculas Pequenas/farmacologia , Triazinas/análise , Triazinas/farmacologia , Internalização do Vírus/efeitos dos fármacos , Replicação Viral/efeitos dos fármacos
5.
Nat Commun ; 11(1): 2750, 2020 06 02.
Artigo em Inglês | MEDLINE | ID: mdl-32487990

RESUMO

Influenza viruses annually kill 290,000-650,000 people worldwide. Antivirals can reduce death tolls. Baloxavir, the recently approved influenza antiviral, inhibits initiation of viral mRNA synthesis, whereas oseltamivir, an older drug, inhibits release of virus progeny. Baloxavir blocks virus replication more rapidly and completely than oseltamivir, reducing the duration of infectiousness. Hence, early baloxavir treatment may indirectly prevent transmission. Here, we estimate impacts of ramping up and accelerating baloxavir treatment on population-level incidence using a new model that links viral load dynamics from clinical trial data to between-host transmission. We estimate that ~22 million infections and >6,000 deaths would have been averted in the 2017-2018 epidemic season by administering baloxavir to 30% of infected cases within 48 h after symptom onset. Treatment within 24 h would almost double the impact. Consequently, scaling up early baloxavir treatment would substantially reduce influenza morbidity and mortality every year. The development of antivirals against the SARS-CoV2 virus that function like baloxavir might similarly curtail transmission and save lives.


Assuntos
Antivirais/uso terapêutico , Epidemias , Influenza Humana/tratamento farmacológico , Orthomyxoviridae/efeitos dos fármacos , Oxazinas/uso terapêutico , Piridinas/uso terapêutico , Tiepinas/uso terapêutico , Triazinas/uso terapêutico , Antivirais/farmacologia , Betacoronavirus/efeitos dos fármacos , Proliferação de Células , Infecções por Coronavirus/tratamento farmacológico , Humanos , Influenza Humana/virologia , Oseltamivir/farmacologia , Oseltamivir/uso terapêutico , Oxazinas/farmacologia , Pandemias , Pneumonia Viral/tratamento farmacológico , Saúde Pública , Piridinas/farmacologia , RNA Mensageiro/metabolismo , Estações do Ano , Tiepinas/farmacologia , Triazinas/farmacologia , Carga Viral , Replicação Viral/efeitos dos fármacos
6.
Toxicol Appl Pharmacol ; 398: 115018, 2020 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-32333917

RESUMO

The incorporation of mesenchymal-epithelial transition factor (c-Met) inhibitors with conventional chemotherapeutics may increase the anticancer efficacy of chemotherapeutic agents, but bears the risk of enhancing the adverse effects. To test the hypothesis, co-administration of the novel c-Met inhibitor capmatinib with cisplatin (CIS) or doxorubicin (DOX) was investigated on nephrotoxicity and cardiotoxicity induced by these agents in mice, as well as their in vitro cytotoxicities. The results demonstrated that capmatinib in vivo offered protection against nephrotoxicity and cardiotoxicity by both CIS and DOX, respectively. The underlying mechanisms behind capmatinib protective effect were found to be i) limiting excessive generation of reactive oxygen species by decreasing the level of lipid peroxidation and nitrosative stress products; and ii) suppressing overproduction of pro-inflammatory mediators like TNF-α and IL-6 that coincided with less inflammatory cell infiltration as denoted by lower levels of serum MCP-1 and Ly6G immunostaining. Besides, capmatinib effectively improved the in vivo anticancer efficacy of both CIS and DOX against solid tumors. In vitro, capmatinib increased the apoptotic activity of DOX against cancerous cells, but did not affect that of CIS. This effect might be linked to capmatinib and DOX abilities to lower IL-12(p40) that has an inhibitory effect on IL-12(p70)/IFN-γ-mediated apoptotic activity. In conclusion, the favorable effects of capmatinib can be applied clinically to decrease the toxicity of DOX and CIS chemotherapeutic agents.


Assuntos
Antineoplásicos/efeitos adversos , Antineoplásicos/farmacologia , Cardiotoxicidade/tratamento farmacológico , Imidazóis/farmacologia , Rim/efeitos dos fármacos , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-met/antagonistas & inibidores , Triazinas/farmacologia , Animais , Cisplatino/efeitos adversos , Cisplatino/farmacologia , Doxorrubicina/efeitos adversos , Doxorrubicina/farmacologia , Feminino , Rim/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Camundongos , Estresse Oxidativo/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo
7.
Zoolog Sci ; 37(2): 103-108, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32282140

RESUMO

Ogerin is a positive allosteric modulator of human and mouse ovarian cancer G protein-coupled receptors (OGR1s). In the present study, we found that ogerin differentially enhances the activation of OGR1 in various animal species. Amino acid residues of OGR1 that are associated with ogerin are conserved among the species. This suggests that other amino acid residues may be involved in the action of ogerin. Chimeric receptors between human and zebrafish OGR1s showed that the amino acid residues that determine the species specificity of ogerin-induced enhancement reside in the transmembrane and/or intracellular regions of OGR1. This result highlights the importance of first verifying the effectiveness of ogerin to the OGR1 of the species of interest at the cellular level prior to analyzing the physiological and pathophysiological roles of OGR1 in the species.


Assuntos
Álcoois Benzílicos/farmacologia , Prótons , Receptores Acoplados a Proteínas-G/genética , Triazinas/farmacologia , Animais , Galinhas , Feminino , Células HEK293 , Humanos , Concentração de Íons de Hidrogênio , Manganês/administração & dosagem , Camundongos , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/metabolismo , Síndrome Respiratória e Reprodutiva Suína , Ratos , Receptores Acoplados a Proteínas-G/metabolismo , Análise de Sequência de Proteína , Suínos , Xenopus , Peixe-Zebra
8.
Arch Biochem Biophys ; 685: 108352, 2020 05 30.
Artigo em Inglês | MEDLINE | ID: mdl-32240637

RESUMO

Rab1A, a member of the Ras-like protein in rat brain (Rab) family, acts as an oncogene in a variety of malignant tumors. Previous studies reported that Rab1A was highly expressed in GC tissues. However, the function and molecular mechanism of Rab1A in gastric cancer (GC) development remain far from being addressed. Rab1A mRNA and protein levels were detected by qRT-PCR and western blot, respectively. Cell proliferation was evaluated by CCK-8 and BrdU incorporation assays. Apoptosis was estimated by flow cytometry analysis and western blot analysis of B cell lymphoma 2 (Bcl-2), myeloid cell leukemia 1 (Mcl-1), Bcl-2 associated X (Bax), and Bcl-2 homologous antagonist/killer (Bak) expression. Alteration of the mammalian target of rapamycin (mTOR)/p70 ribosomal protein S6 kinase (p70S6K) signaling pathway was detected by western blot. We found that Rab1A expression at both mRNA and protein was upregulated in GC cells. Rab1A knockdown significantly inhibited cell proliferation and induced apoptosis in GC cells. Rab1A overexpression promoted proliferation, inhibited cisplatin-induced apoptosis, and increased xenograft growth. In addition, we found that Rab1A knockdown suppressed the mTOR/p70S6K pathway in GC cells. Moreover, activation of mTOR/p70S6K pathway by MHY1485 abolished the effects of Rab1A knockdown on cell proliferation and apoptosis. In conclusion, Rab1A knockdown repressed proliferation and promoted apoptosis in GC cells by inhibition of the mTOR/p70S6K pathway.


Assuntos
Apoptose/fisiologia , Proliferação de Células/fisiologia , Transdução de Sinais/fisiologia , Neoplasias Gástricas/metabolismo , Proteínas rab1 de Ligação ao GTP/metabolismo , Animais , Apoptose/genética , Linhagem Celular Tumoral , Proliferação de Células/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Camundongos Endogâmicos BALB C , Camundongos Nus , Morfolinas/farmacologia , Proteínas Quinases S6 Ribossômicas 70-kDa/metabolismo , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Serina-Treonina Quinases TOR/metabolismo , Triazinas/farmacologia , Regulação para Cima , Proteínas rab1 de Ligação ao GTP/genética
9.
PLoS Pathog ; 16(4): e1008395, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32294137

RESUMO

Influenza viruses cause seasonal outbreaks and pose a continuous pandemic threat. Although vaccines are available for influenza control, their efficacy varies each season and a vaccine for a novel pandemic virus manufactured using current technology will not be available fast enough to mitigate the effect of the first pandemic wave. Antivirals can be effective against many different influenza viruses but have not thus far been used extensively for outbreak control. Baloxavir, a recently licensed antiviral drug that targets the influenza virus endonuclease, has been shown to reduce virus shedding more effectively than oseltamivir, a widely used neuraminidase inhibitor drug. Thus it is possible that treatment with baloxavir might also interrupt onward virus transmission. To test this, we utilized the ferret model, which is the most commonly used animal model to study influenza virus transmission. We established a subcutaneous baloxavir administration method in ferrets which achieved similar pharmacokinetics to the approved human oral dose. Transmission studies were then conducted in two different locations with different experimental setups to compare the onward transmission of A(H1N1)pdm09 virus from infected ferrets treated with baloxavir, oseltamivir or placebo to naïve sentinel ferrets exposed either indirectly in adjacent cages or directly by co-housing. We found that baloxavir treatment reduced infectious viral shedding in the upper respiratory tract of ferrets compared to placebo, and reduced the frequency of transmission amongst sentinels in both experimental setups, even when treatment was delayed until 2 days post-infection. In contrast, oseltamivir treatment did not substantially affect viral shedding or transmission compared to placebo. We did not detect the emergence of baloxavir-resistant variants in treated animals or in untreated sentinels. Our results support the concept that antivirals which decrease viral shedding could also reduce influenza transmission in the community.


Assuntos
Antivirais/farmacologia , Vírus da Influenza A Subtipo H1N1/efeitos dos fármacos , Neuraminidase/antagonistas & inibidores , Infecções por Orthomyxoviridae/tratamento farmacológico , Oxazinas/farmacologia , Piridinas/farmacologia , Tiepinas/farmacologia , Triazinas/farmacologia , Replicação Viral/efeitos dos fármacos , Eliminação de Partículas Virais/efeitos dos fármacos , Animais , Feminino , Furões , Infecções por Orthomyxoviridae/transmissão , Infecções por Orthomyxoviridae/virologia
10.
Proc Natl Acad Sci U S A ; 117(15): 8593-8601, 2020 04 14.
Artigo em Inglês | MEDLINE | ID: mdl-32217734

RESUMO

Baloxavir marboxil (BXM) was approved in 2018 for treating influenza A and B virus infections. It is a first-in-class inhibitor targeting the endonuclease activity of the virus polymerase acidic (PA) protein. Clinical trial data revealed that PA amino acid substitutions at residue 38 (I38T/F/M) reduced BXM potency and caused virus rebound in treated patients, although the fitness characteristics of the mutant viruses were not fully defined. To determine the fitness impact of the I38T/F/M substitutions, we generated recombinant A/California/04/2009 (H1N1)pdm09, A/Texas/71/2017 (H3N2), and B/Brisbane/60/2008 viruses with I38T/F/M and examined drug susceptibility in vitro, enzymatic properties, replication efficiency, and transmissibility in ferrets. Influenza viruses with I38T/F/M substitutions exhibited reduced baloxavir susceptibility, with 38T causing the greatest reduction. The I38T/F/M substitutions impaired PA endonuclease activity as compared to that of wild-type (I38-WT) PA. However, only 38T/F A(H3N2) substitutions had a negative effect on polymerase complex activity. The 38T/F substitutions decreased replication in cells among all viruses, whereas 38M had minimal impact. Despite variable fitness consequences in vitro, all 38T/M viruses disseminated to naive ferrets by contact and airborne transmission, while 38F-containing A(H3N2) and B viruses failed to transmit via the airborne route. Reversion of 38T/F/M to I38-WT was rare among influenza A viruses in this study, suggesting stable retention of 38T/F/M genotypes during these transmission events. BXM reduced susceptibility-associated mutations had variable effects on in vitro fitness of influenza A and B viruses, but the ability of these viruses to transmit in vivo indicates a risk of their spreading from BXM-treated individuals.


Assuntos
Farmacorresistência Viral , Vírus da Influenza A Subtipo H1N1/efeitos dos fármacos , Vírus da Influenza A Subtipo H3N2/efeitos dos fármacos , Vírus da Influenza B/efeitos dos fármacos , Infecções por Orthomyxoviridae/transmissão , Oxazinas/farmacologia , Piridinas/farmacologia , Tiepinas/farmacologia , Triazinas/farmacologia , Replicação Viral , Substituição de Aminoácidos , Animais , Antivirais/farmacologia , Furões , Masculino , Testes de Sensibilidade Microbiana , Infecções por Orthomyxoviridae/tratamento farmacológico , Infecções por Orthomyxoviridae/veterinária , Infecções por Orthomyxoviridae/virologia , Proteínas Virais/genética , Proteínas Virais/metabolismo
11.
Biochim Biophys Acta Biomembr ; 1862(6): 183235, 2020 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-32126232

RESUMO

The adenosine 2A receptor (A2AR), a G-protein-coupled receptor (GPCR), was solubilised and purified encapsulated in styrene maleic acid lipid particles (SMALPs). The purified A2AR-SMALP was associated with phospholipids characteristic of the plasma membrane of Pichia pastoris, the host used for its expression, confirming that the A2AR-SMALP encapsulated native lipids. The fluorescence spectrum of the A2AR-SMALP showed a characteristic broad emission peak at 330 nm, produced by endogenous Trp residues. The inverse agonist ZM241385 caused 30% increase in fluorescence emission, unusually accompanied by a red-shift in the emission wavelength. The emission spectrum also showed sub-peaks at 321 nm, 335 nm and 350 nm, indicating that individual Trp inhabited different environments following ZM241385 addition. There was no effect of the agonist NECA on the A2AR-SMALP fluorescence spectrum. Substitution of two Trp residues by Tyr suggested that ZM241385 affected the environment and mobility of Trp2466.48 in TM6 and Trp2687.33 at the extracellular face of TM7, causing transition to a more hydrophobic environment. The fluorescent moiety IAEDANS was site-specifically introduced at the intracellular end of TM6 (residue 2316.33) to report on the dynamic cytoplasmic face of the A2AR. The inverse agonist ZM241385 caused a concentration-dependent increase in fluorescence emission as the IAEDANS moved to a more hydrophobic environment, consistent with closing the G-protein binding crevice. NECA generated only 30% of the effect of ZM241385. This study provides insight into the SMALP environment; encapsulation supported constitutive activity of the A2AR and ZM241385-induced conformational transitions but the agonist NECA generated only small effects.


Assuntos
Lipídeos/química , Receptor A2A de Adenosina/química , Estireno/química , Interações Hidrofóbicas e Hidrofílicas , Ligantes , Maleatos/química , Pichia/química , Conformação Proteica , Espectrometria de Fluorescência/métodos , Triazinas/farmacologia , Triazóis/farmacologia , Triptofano/química
12.
Life Sci ; 248: 117474, 2020 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-32112869

RESUMO

BACKGROUND/OBJECTIVES: Nicotinamide N-methyltransferase (NNMT) is a novel regulator of energy homeostasis in adipocytes. NNMT expression in adipose tissue is increased in obesity and diabetes. Knockdown of NNMT prevents mice from developing diet-induced obesity, which is closely linked to insulin resistance. An early sign of systemic insulin resistance is reduced expression of glucose transporter 4 (GLUT4) selectively in adipose tissue. Adipose tissue-specific knockout and overexpression of GLUT4 cause reciprocal changes in NNMT expression. The aim of the current study was to elucidate the mechanism that regulates NNMT expression in adipocytes. METHODS: 3T3-L1 adipocytes were cultured in media with varying glucose concentrations or activators and inhibitors of intracellular pathways. NNMT mRNA and protein levels were measured with quantitative polymerase chain reaction and Western blotting. RESULTS: Glucose deprivation of 3T3-L1 adipocytes induced a 2-fold increase in NNMT mRNA and protein expression. This effect was mimicked by inhibition of glucose transport with phloretin, and by inhibition of glycolysis with the phosphoglucose isomerase inhibitor 2-deoxyglucose. Conversely, inhibition of the pentose phosphate pathway did not affect NNMT expression. Pharmacological activation of the cellular energy sensor AMP-activated protein kinase (AMPK) and inhibition of the mammalian target of rapamycin (mTOR) pathway caused an increase in NNMT levels that was similar to the effect of glucose deprivation. Activation of mTOR with MHY1485 prevented the effect of glucose deprivation on NNMT expression. Furthermore, upregulation of NNMT levels depended on functional autophagy and protein translation. CONCLUSION: Glucose availability regulates NNMT expression via an mTOR-dependent mechanism.


Assuntos
Adipócitos/efeitos dos fármacos , Transportador de Glucose Tipo 4/genética , Glucose/farmacologia , Nicotinamida N-Metiltransferase/genética , Serina-Treonina Quinases TOR/genética , Células 3T3-L1 , Proteínas Quinases Ativadas por AMP/genética , Proteínas Quinases Ativadas por AMP/metabolismo , Adipócitos/citologia , Adipócitos/metabolismo , Animais , Autofagia/efeitos dos fármacos , Autofagia/genética , Transporte Biológico/efeitos dos fármacos , Diferenciação Celular , Desoxiglucose/farmacologia , Metabolismo Energético/genética , Regulação da Expressão Gênica , Glucose/metabolismo , Transportador de Glucose Tipo 4/antagonistas & inibidores , Transportador de Glucose Tipo 4/metabolismo , Glucose-6-Fosfato Isomerase/antagonistas & inibidores , Glucose-6-Fosfato Isomerase/genética , Glucose-6-Fosfato Isomerase/metabolismo , Homeostase/genética , Camundongos , Morfolinas/farmacologia , Nicotinamida N-Metiltransferase/antagonistas & inibidores , Nicotinamida N-Metiltransferase/metabolismo , Via de Pentose Fosfato/efeitos dos fármacos , Via de Pentose Fosfato/genética , Floretina/farmacologia , Biossíntese de Proteínas , RNA Mensageiro/antagonistas & inibidores , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Triazinas/farmacologia
13.
Parasitol Res ; 119(5): 1653-1661, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32219548

RESUMO

Ethanamizuril (EZL) is a novel triazine compound with excellent anticoccidial activity. We carried out a preliminary investigation of the effects of EZL on the different life cycle stages of Eimeria tenella. EZL mainly acted on the schizogony stage, with peak activity during the second-generation merozoite stage. We also studied the possible target of EZL by identifying the majorly differentially expressed gene affected by EZL in second-generation merozoites using real-time polymerase chain reaction, and screening for surface antigen proteins (SAGs). The relative expression levels of SAGs were compared by Western blot analysis showing that expression levels of surface antigen family member (SAGfm) and SAG19 were significantly downregulated by EZL. Immunofluorescence analysis indicated that SAGfm and SAG19 were localized on the surface of second-generation merozoites. In addition, fluorescence signals were significantly stronger in second-generation merozoites of infected non-medicated control (INC) group compared with that of the EZL group. Therefore, it was speculated that SAGs might be a potential target of EZL action. The inhibitory effects of anticoccidial drugs on SAG levels in coccidia thus warrant further research.


Assuntos
Coccidiose/tratamento farmacológico , Eimeria tenella/efeitos dos fármacos , Doenças das Aves Domésticas/prevenção & controle , Triazinas/farmacologia , Animais , Antígenos de Superfície/metabolismo , Western Blotting , Galinhas/parasitologia , Coccidiose/parasitologia , Estágios do Ciclo de Vida/efeitos dos fármacos , Merozoítos/efeitos dos fármacos , Proteínas de Protozoários/genética , RNA Mensageiro/genética , Reação em Cadeia da Polimerase em Tempo Real
14.
Chemosphere ; 244: 125505, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32050329

RESUMO

Melamine (MEL) and its derivatives are widely used in many consumer products, including furniture, kitchenware, and plastics. However, very limited knowledge exists on human exposure to MEL and its derivatives, especially in the indoor environment. Here, we determined the occurrence and distribution of 11 MEL derivatives in childcare facilities and estimated children's exposure through dust ingestion and dermal absorption. We analyzed dust and samples of nap mats, a commonly used item in many childcares, from eight facilities located in the United States. Eight MEL-based compounds were detected in dust, and total MEL concentrations ranged from 429 to 117,000 ng/g. The most abundant compounds found in the dust samples were MEL, cyanuric acid (CYA), ammeline (AMN), and ammelide (AMD), with median concentrations of 1620, 585, 1060, and 299 ng/g, respectively. MEL, CYA, AMN and 2,4,6-tris[bis(methoxymethyl)amino]-1,3,5-triazine (TBMMAT) were also detected in nap mats with median concentrations of 45.6, 19.8, 1510 and 2.5 ng/g, respectively. ΣMEL concentrations in mat covers (median 709 ng/g) were significantly higher than those in mat foam (median 15.1 ng/g). Estimated daily intakes (EDIs) of MEL and its derivatives via dust ingestion were two orders of magnitude higher than the EDIs through dermal absorption, but both were below the established tolerable daily intake levels. This is the first report on exposure to MEL and its derivatives in the childcare environment.


Assuntos
Creches/normas , Exposição Ambiental/análise , Triazinas/farmacologia , Poluição do Ar em Ambientes Fechados/análise , Criança , Poeira/análise , Humanos , Absorção Cutânea , Triazinas/análise , Estados Unidos
15.
BMC Infect Dis ; 20(1): 88, 2020 Jan 30.
Artigo em Inglês | MEDLINE | ID: mdl-32000698

RESUMO

BACKGROUND: Carbapenem-resistant Enterobacteriaceae (CRE) infections have become a global health threat. Controlling CRE transmission in hospitals is increasingly dependent on the use of disinfectants to restrict the risk of infection. Here, the susceptibility of patient-derived carbapenem resistant Klebsiella pneumoniae (CRKP) and Escherichia coli (CREC) strains against three common disinfectants and the determinants of resistance to disinfectants were investigated. METHODS: The minimum inhibitory concentrations (MICs) and the minimum bactericidal concentrations (MBCs) of three common chemical disinfectants: chlorhexidine, trichloroisocyanuric (TCCA) acid and Povidone iodine (PVP-I) against 50 CRE strains were measured. The drug-resistance genes -qacEΔ1, qacA/B and cepA-were determined using polymerase chain reaction. RESULTS: A total of 36 CRKP and 14 CREC strains were collected in our hospital from 2016 to 2018. The MIC ranges of 36 CRKP strains against chlorhexidine, TCCA and PVP-I were 8~512 mg/L, 64~128 mg/L and 8~128 mg/L, respectively. For 14 CREC strains, the MIC ranges against chlorhexidine, TCCA and PVP-I were 4~128 mg/L, 64~128 mg/L and 4~128 mg/L, respectively. Moreover, against chlorhexidine and PVP-I, the MIC90 of 36 CRKP strains was higher than that of 50 CSKP strains. The qacE△1 gene was detected in 15 isolates among 36 CRKP strains (41.7%), and 8 isolates among 14 CREC strains (57.1%); while the qacA/B gene was not detected. Specifically, the cepA gene was much more prevalent than the qacEΔ1; it reached over 80% among CRKP strains. Compared to the CSKP strains, the presence of the qacEΔ1 and cepA genes was significantly higher among the CRKP strains (p < 0.05). CONCLUSION: CRE strains collected from patients in our hospital exhibit various degree of resistance to the commonly used chemical disinfectants. It is of great help to keep monitoring the tendency of the reduced susceptibility of the pan-resistant strains against disinfectants, in order to effectively control and prevent the spread of the super resistant bacteria.


Assuntos
Enterobacteriáceas Resistentes a Carbapenêmicos/efeitos dos fármacos , Desinfetantes/farmacologia , Farmacorresistência Bacteriana/genética , Escherichia coli/efeitos dos fármacos , Klebsiella pneumoniae/efeitos dos fármacos , Antibacterianos/farmacologia , Enterobacteriáceas Resistentes a Carbapenêmicos/genética , Enterobacteriáceas Resistentes a Carbapenêmicos/isolamento & purificação , China , Clorexidina/farmacologia , Farmacorresistência Bacteriana/efeitos dos fármacos , Escherichia coli/genética , Escherichia coli/isolamento & purificação , Humanos , Infecções por Klebsiella/microbiologia , Klebsiella pneumoniae/genética , Klebsiella pneumoniae/isolamento & purificação , Testes de Sensibilidade Microbiana , Povidona-Iodo/farmacologia , Centros de Atenção Terciária , Triazinas/farmacologia
16.
Sci Rep ; 10(1): 1614, 2020 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-32005941

RESUMO

The biological function of non-thermal atmospheric pressure plasma has been widely accepted in several types of cancer. We previously developed plasma-activated medium (PAM) for clinical use, and demonstrated that PAM exhibits a metastasis-inhibitory effect on ovarian cancer through reduced MMP-9 secretion. However, the anti-tumor effects of PAM on endometrial cancer remain unknown. In this study, we investigated the inhibitory effect of PAM on endometrial cancer cell viability in vitro. Our results demonstrated that AMEC and HEC50 cell viabilities were reduced by PAM at a certain PAM ratio, and PAM treatment effectively increased autophagic cell death in a concentration dependent manner. In addition, we evaluated the molecular mechanism of PAM activity and found that the mTOR pathway was inactivated by PAM. Moreover, our results demonstrated that the autophagy inhibitor MHY1485 partially inhibited the autophagic cell death induced by PAM treatment. These findings indicate that PAM decreases the viability of endometrial cancer cells along with alteration of the mTOR pathway, which is critical for cancer cell viability. Collectively, our data suggest that PAM inhibits cell viability while inducing autophagic cell death in endometrial cancer cells, representing a potential novel treatment for endometrial cancer.


Assuntos
Morte Celular Autofágica/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Fatores Biológicos/farmacologia , Plasma/metabolismo , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismo , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Neoplasias do Endométrio/tratamento farmacológico , Neoplasias do Endométrio/metabolismo , Feminino , Humanos , Morfolinas/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Triazinas/farmacologia
17.
Biomolecules ; 10(2)2020 01 25.
Artigo em Inglês | MEDLINE | ID: mdl-31991782

RESUMO

: GAL-021 has recently been developed as a novel breathing control modulator. However, modifications of ionic currents produced by this agent remain uncertain, although its efficacy in suppressing the activity of big-conductance Ca2+-activated K+ (BKCa) channels has been reported. In pituitary tumor (GH3) cells, we found that the presence of GAL-021 decreased the amplitude of macroscopic Ca2+-activated K+ current (IK(Ca)) in a concentration-dependent manner with an effective IC50 of 2.33 µM. GAL-021-mediated reduction of IK(Ca) was reversed by subsequent application of verteporfin or ionomycin; however, it was not by that of diazoxide. In inside-out current recordings, the addition of GAL-021 to the bath markedly decreased the open-state probability of BKCa channels. This agent also resulted in a rightward shift in voltage dependence of the activation curve of BKCa channels; however, neither the gating charge of the curve nor single-channel conductance of the channel was changed. There was an evident lengthening of the mean closed time of BKCa channels in the presence of GAL-021, with no change in mean open time. The GAL-021 addition also suppressed M-type K+ current with an effective IC50 of 3.75 µM; however, its presence did not alter the amplitude of erg-mediated K+ current, or mildly suppressed delayed-rectifier K+ current. GAL-021 at a concentration of 30 µM could also suppress hyperpolarization-activated cationic current. In HEK293T cells expressing α-hSlo, the addition of GAL-021 was also able to suppress the BKCa-channel open probabilities, and GAL-021-mediated suppression of BKCa-channel activity was attenuated by further addition of BMS-191011. Collectively, the GAL-021 effects presented herein do not exclusively act on BKCa channels and these modifications on ionic currents exert significant influence on the functional activities of electrically excitable cells occurring in vivo.


Assuntos
Neoplasias Hipofisárias/tratamento farmacológico , Respiração/efeitos dos fármacos , Triazinas/farmacologia , Cálcio/metabolismo , Diazóxido/farmacologia , Células HEK293 , Humanos , Ionomicina/farmacologia , Neoplasias Hipofisárias/patologia , Verteporfina/farmacologia
18.
Nat Commun ; 11(1): 164, 2020 01 09.
Artigo em Inglês | MEDLINE | ID: mdl-31919360

RESUMO

Host dependency factors that are required for influenza A virus infection may serve as therapeutic targets as the virus is less likely to bypass them under drug-mediated selection pressure. Previous attempts to identify host factors have produced largely divergent results, with few overlapping hits across different studies. Here, we perform a genome-wide CRISPR/Cas9 screen and devise a new approach, meta-analysis by information content (MAIC) to systematically combine our results with prior evidence for influenza host factors. MAIC out-performs other meta-analysis methods when using our CRISPR screen as validation data. We validate the host factors, WDR7, CCDC115 and TMEM199, demonstrating that these genes are essential for viral entry and regulation of V-type ATPase assembly. We also find that CMTR1, a human mRNA cap methyltransferase, is required for efficient viral cap snatching and regulation of a cell autonomous immune response, and provides synergistic protection with the influenza endonuclease inhibitor Xofluza.


Assuntos
Predisposição Genética para Doença/genética , Interações Hospedeiro-Patógeno/genética , Vírus da Influenza A/patogenicidade , Influenza Humana/genética , Influenza Humana/patologia , Células A549 , Proteínas Adaptadoras de Transdução de Sinal/genética , Antivirais/farmacologia , Sistemas CRISPR-Cas , Linhagem Celular , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética , Estudo de Associação Genômica Ampla , Humanos , Proteínas de Membrana/genética , Metiltransferases/metabolismo , Proteínas do Tecido Nervoso/genética , Oxazinas/farmacologia , Piridinas/farmacologia , Tiepinas/farmacologia , Triazinas/farmacologia , ATPases Vacuolares Próton-Translocadoras/metabolismo , Internalização do Vírus
19.
Arch Pharm (Weinheim) ; 353(2): e1900108, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31894866

RESUMO

Two series of thieno[2,3-d][1,2,3]triazine derivatives were designed, synthesized, and biologically evaluated as potential epidermal growth factor receptor (EGFR) inhibitors targeting the non-small-cell lung cancer cell line H1299. Most of the synthesized compounds displayed IC50 values ranging from 25 to 58 nM against H1299, which are superior to that of gefitinib (40 µM). 3-(5,6,7,8-Tetrahydro-7H-cyclohexa[4:5]thieno[2,3-d]-1,2,3-triazin-4-ylamino)benzene-1,3-diamine (6b) achieved the highest cytotoxic activity against H1299 with an IC50 value of 25 nM; it had the ability to decrease the EGFR concentration in H1299 cells from 7.22 to 2.67 pg/ml. In vitro, the IC50 value of compound 6b was 0.33 nM against EGFR, which is superior to that of gefitinib at 1.9 nM and erlotinib at 4 nM. The three-dimensional quantitative structure-activity relationships and molecular modeling studies revealed comparable binding modes of compound 6b, gefitinib, and erlotinib in the EGFR active site. The in silico ADME (absorption, distribution, metabolism, and excretion) prediction parameters of this compound revealed promising pharmacokinetic and physicochemical properties. Moreover, DFT (density functional theory) calculations showed the high reactivity of compound 6b toward the EGFR compared with other compounds. The designed compound 6b might serve as an encouraging lead compound for the discovery of promising anti-lung cancer agents targeting EGFR.


Assuntos
Antineoplásicos/farmacologia , Neoplasias Pulmonares/tratamento farmacológico , Relação Quantitativa Estrutura-Atividade , Triazinas/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Proliferação de Células/efeitos dos fármacos , Teoria da Densidade Funcional , Relação Dose-Resposta a Droga , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/metabolismo , Humanos , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patologia , Modelos Moleculares , Estrutura Molecular , Triazinas/síntese química , Triazinas/química , Células Tumorais Cultivadas
20.
J Infect Dis ; 221(1): 63-70, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31419295

RESUMO

BACKGROUND: Baloxavir is a cap-dependent inhibitor of the polymerase acid (PA) protein of influenza viruses. While appearing virologically superior to oseltamivir, baloxavir exhibits a low barrier of resistance. We sought to assess the impact of the common baloxavir-resistant I38T PA substitution on in vitro properties and virulence. METHODS: Influenza A/Quebec/144147/2009 (H1N1)pdm09 and A/Switzerland/9715293/2013 (H3N2) recombinant viruses and their I38T PA mutants were compared in single and competitive infection experiments in ST6GalI-MDCK cells and C57/BL6 mice. Virus titers in cell culture supernatants and lung homogenates were determined by virus yield assays. Ratios of wild-type (WT) and I38T mutant were assessed by digital RT-PCR. RESULTS: I38T substitution did not alter the replication kinetics of A(H1N1)pdm09 and A(H3N2) viruses. In competition experiments, a 50%:50% mixture evolved to 70%:30% (WT/mutant) for A(H1N1) and 88%:12% for A(H3N2) viruses after a single cell passage. The I38T substitution remained stable after 4 passages in vitro. In mice, the WT and its I38T mutant induced similar weight loss with comparable lung titers in both viral subtypes. The mutant virus tended to predominate over the WT in mouse competition experiments. CONCLUSION: The fitness of baloxavir-resistant I38T PA mutants appears relatively unaltered in seasonal subtypes warranting surveillance for its dissemination.


Assuntos
Antivirais/farmacologia , Farmacorresistência Viral/genética , Vírus da Influenza A Subtipo H1N1/genética , Vírus da Influenza A Subtipo H3N2/genética , Infecções por Orthomyxoviridae/tratamento farmacológico , Oxazinas/farmacologia , Piridinas/farmacologia , RNA Replicase/genética , Tiepinas/farmacologia , Triazinas/farmacologia , Proteínas Virais/genética , Substituição de Aminoácidos , Animais , Antivirais/uso terapêutico , Cães , Feminino , Vírus da Influenza A Subtipo H1N1/enzimologia , Vírus da Influenza A Subtipo H1N1/patogenicidade , Vírus da Influenza A Subtipo H3N2/enzimologia , Vírus da Influenza A Subtipo H3N2/patogenicidade , Concentração Inibidora 50 , Pulmão/virologia , Células Madin Darby de Rim Canino , Camundongos , Camundongos Endogâmicos C57BL , Mutação , Infecções por Orthomyxoviridae/virologia , Oxazinas/uso terapêutico , Fenótipo , Piridinas/uso terapêutico , RNA Replicase/metabolismo , Tiepinas/uso terapêutico , Triazinas/uso terapêutico , Carga Viral/efeitos dos fármacos , Proteínas Virais/metabolismo , Replicação Viral/efeitos dos fármacos
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