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1.
Nat Commun ; 11(1): 6339, 2020 12 11.
Artigo em Inglês | MEDLINE | ID: mdl-33311482

RESUMO

Ferroptosis is a more recently recognized form of cell death that relies on iron-mediated oxidative damage. Here, we evaluate the impact of high-iron diets or depletion of Gpx4, an antioxidant enzyme reported as an important ferroptosis suppressor, in the pancreas of mice with cerulean- or L-arginine-induced pancreatitis, and in an oncogenic Kras murine model of spontaneous pancreatic ductal adenocarcinoma (PDAC). We find that either high-iron diets or Gpx4 depletion promotes 8-OHG release and thus activates the TMEM173/STING-dependent DNA sensor pathway, which results in macrophage infiltration and activation during Kras-driven PDAC in mice. Consequently, the administration of liproxstatin-1 (a ferroptosis inhibitor), clophosome-mediated macrophage depletion, or pharmacological and genetic inhibition of the 8-OHG-TMEM173 pathway suppresses Kras-driven pancreatic tumorigenesis in mice. GPX4 is also a prognostic marker in patients with PDAC. These findings provide pathological and mechanistic insights into ferroptotic damage in PDAC tumorigenesis in mice.


Assuntos
Carcinogênese/metabolismo , Transformação Celular Neoplásica/metabolismo , Ferroptose/fisiologia , Proteínas de Membrana/metabolismo , Pâncreas/metabolismo , Animais , Biomarcadores Tumorais , Carcinogênese/genética , Carcinoma Ductal Pancreático/metabolismo , Carcinoma Ductal Pancreático/patologia , Morte Celular/fisiologia , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/patologia , DNA , Dieta , Modelos Animais de Doenças , Feminino , Ferroptose/efeitos dos fármacos , Humanos , Ferro/metabolismo , Macrófagos , Masculino , Proteínas de Membrana/genética , Camundongos , Camundongos Knockout , Pâncreas/patologia , Pancreatite/genética , Fosfolipídeo Hidroperóxido Glutationa Peroxidase/genética , Fosfolipídeo Hidroperóxido Glutationa Peroxidase/metabolismo , Proteínas Proto-Oncogênicas p21(ras)/metabolismo , Quinoxalinas/farmacologia , Compostos de Espiro/farmacologia , Microambiente Tumoral
2.
Yakugaku Zasshi ; 140(10): 1207-1212, 2020.
Artigo em Japonês | MEDLINE | ID: mdl-32999199

RESUMO

T-type calcium channels are low-threshold voltage-gated calcium channel and characterized by unique electrophysiological properties such as fast inactivation and slow deactivation kinetics. All subtypes of T-type calcium channel (Cav3.1, 3.2 and 3.3) are widely expressed in the central nerve system, and they have an important role in homeostasis of sleep, pain response, and development of epilepsy. Recently, several reports suggest that T-type calcium channels may mediate neuronal plasticity in the mouse brain. We succeeded to develop T-type calcium channel enhancer ethyl 8'-methyl-2',4-dioxo-2-(piperidin-1-yl)-2'H-spiro[cyclopentane-1,3'-imidazo[1,2-a]pyridine]-2-ene-3-carboxylate (SAK3) which enhances Cav3.1 and 3.3 currents in each-channel expressed neuro2A cells. SAK3 can promote acetylcholine (ACh) release in the mouse hippocampus via enhancing T-type calcium channel. In this review, we have introduced the role of T-type calcium channel, especially Cav3.1 channel in the mouse hippocampus based on our previous data using SAK3 and Cav3.1 knockout mice.


Assuntos
Canais de Cálcio Tipo T/efeitos dos fármacos , Canais de Cálcio Tipo T/fisiologia , Imidazóis/farmacologia , Neurônios/fisiologia , Compostos de Espiro/farmacologia , Acetilcolina/metabolismo , Animais , Encéfalo/fisiologia , Canais de Cálcio Tipo T/genética , Canais de Cálcio Tipo T/metabolismo , Células Cultivadas , Sistema Nervoso Central/metabolismo , Fenômenos Eletrofisiológicos , Epilepsia/etiologia , Expressão Gênica/efeitos dos fármacos , Hipocampo/metabolismo , Homeostase , Camundongos , Plasticidade Neuronal , Dor/etiologia , Ratos , Sono/fisiologia
3.
Anticancer Res ; 40(9): 5049-5057, 2020 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-32878793

RESUMO

BACKGROUND/AIM: Studies with acridine compounds have reported anticancer effects. Herein, we evaluated the toxicity and antitumor effect of the (E)-1'-((4-chlorobenzylidene)amino)-5'-oxo-1',5'-dihydro-10H-spiro[acridine-9,2'-pyrrole]-4'-carbonitrile (AMTAC-06), a promising anticancer spiro-acridine compound. MATERIALS AND METHODS: The toxicity of AMTAC-06 was evaluated on zebrafish and mice. Antitumor activity was assessed in Ehrlich ascites carcinoma model. Effects on angiogenesis, cytokine levels and cell cycle were also investigated. RESULTS: AMTAC-06 did not induce toxicity on zebrafish and mice (LD50 approximately 5000 mg/kg, intraperitoneally). No genotoxicity was observed on micronucleus assay. AMTAC-06 significantly reduced the total viable Ehrlich tumor cells and increased sub-G1 peak, suggesting apoptosis was triggered. Moreover, the compound significantly decreased the density of peritumoral microvessels, indicating an anti-angiogenic action, possibly dependent on the cytokine modulation (TNF-α, IL-1ß and IFN-γ). No significant toxicological effects were recorded for AMTAC-06 on tumor transplanted animals. CONCLUSION: AMTAC-06 has low toxicity and a significant antitumor activity.


Assuntos
Acridinas/farmacologia , Inibidores da Angiogênese/farmacologia , Antineoplásicos/farmacologia , Fatores Imunológicos/farmacologia , Compostos de Espiro/farmacologia , Acridinas/química , Inibidores da Angiogênese/química , Animais , Antineoplásicos/química , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Citocinas/metabolismo , Modelos Animais de Doenças , Humanos , Fatores Imunológicos/química , Imunomodulação/efeitos dos fármacos , Camundongos , Estrutura Molecular , Compostos de Espiro/química , Ensaios Antitumorais Modelo de Xenoenxerto , Peixe-Zebra
4.
Mol Pharmacol ; 98(4): 462-474, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32958572

RESUMO

Opioid receptors (ORs) convert extracellular messages to signaling events by coupling to the heterotrimeric G proteins, Gα•ßγ Classic pharmacological methods, such as [35S]GTPγS binding and inhibition of cyclic AMP production, allow for general opioid characterization, but they are subject to the varying endogenous Gα proteins in a given cell type. Bioluminescence resonance energy transfer (BRET) technology offers new insight by allowing the direct observation of Gα subunit-specific effects on opioid pharmacology. Using a Venus-tagged Gßγ and nanoluciferase-tagged truncated G protein receptor kinase 3, an increase in BRET signal correlated with OR activation mediated by a specific Gα protein. The magnitude of the BRET signal was normalized to the maximum response obtained with 10 µM 2-(3,4-dichlorophenyl)-N-methyl-N-[(1R,2R)-2-pyrrolidin-1-ylcyclohexyl]acetamide (U50,488) for the kappa OR (KOR). Opioids reached equilibrium with the KOR, and concentration-response curves were generated. Although the full agonists U50,488, salvinorin A, nalfurafine, and dynorphin peptides were equally efficacious regardless of the Gα subunit present, the concentration-response curves were leftward shifted when the KOR was signaling through Gαz compared with other Gαi/o subunits. In contrast, the Gα subunit distinctly affected both the efficacy and potency of partial kappa agonists, such as the benzomorphans, and the classic mu opioid antagonists, naloxone, naltrexone, and nalmefene. For example, (-)pentazocine had EC50 values of 7.3 and 110 nM and maximal stimulation values of 79% and 35% when the KOR signaled through Gαz and Gαi1, respectively. Together, these observations suggest KOR pharmacology varies based on the specific Gα subunit coupled to the KOR. SIGNIFICANCE STATEMENT: Opioid receptors couple to various heterotrimeric Gαßγ proteins to convert extracellular cues to precise intracellular events. This paper focuses on how the various inhibitory Gα subunits influence the pharmacology of full and partial agonists at the kappa opioid receptor. Using a bioluminescent assay, the efficacy and potency of kappa opioids was determined. Opioid signaling was more potent through Gαz compared with other Gα proteins. These observations suggest that Gαz may impact opioid pharmacology and cellular physiology more than previously thought.


Assuntos
Analgésicos Opioides/farmacologia , Técnicas de Transferência de Energia por Ressonância de Bioluminescência/métodos , Subunidades alfa de Proteínas de Ligação ao GTP/metabolismo , Receptores Opioides kappa/metabolismo , (trans)-Isômero de 3,4-dicloro-N-metil-N-(2-(1-pirrolidinil)-ciclo-hexil)-benzenoacetamida/farmacologia , Diterpenos Clerodânicos/farmacologia , Dinorfinas/farmacologia , Células HEK293 , Humanos , Morfinanos/farmacologia , Transdução de Sinais/efeitos dos fármacos , Compostos de Espiro/farmacologia
5.
Anesthesiology ; 133(3): 559-568, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32788558

RESUMO

BACKGROUND: To improve understanding of the respiratory behavior of oliceridine, a µ-opioid receptor agonist that selectively engages the G-protein-coupled signaling pathway with reduced activation of the ß-arrestin pathway, the authors compared its utility function with that of morphine. It was hypothesized that at equianalgesia, oliceridine will produce less respiratory depression than morphine and that this is reflected in a superior utility. METHODS: Data from a previous trial that compared the respiratory and analgesic effects of oliceridine and morphine in healthy male volunteers (n = 30) were reanalyzed. A population pharmacokinetic-pharmacodynamic analysis was performed and served as basis for construction of utility functions, which are objective functions of probability of analgesia, P(analgesia), and probability of respiratory depression, P(respiratory depression). The utility function = P(analgesia ≥ 0.5) - P(respiratory depression ≥ 0.25), where analgesia ≥ 0.5 is the increase in hand withdrawal latency in the cold pressor test by at least 50%, and respiratory depression ≥ 0.25 is the decrease of the hypercapnic ventilatory response by at least 25%. Values are median ± standard error of the estimate. RESULTS: The two drugs were equianalgesic with similar potency values (oliceridine: 27.9 ± 4.9 ng/ml; morphine 34.3 ± 9.7 ng/ml; potency ratio, 0.81; 95% CI, 0.39 to 1.56). A 50% reduction of the hypercapnic ventilatory response by morphine occurred at an effect-site concentration of 33.7 ± 4.8 ng/ml, while a 25% reduction by oliceridine occurred at 27.4 ± 3.5 ng/ml (potency ratio, 2.48; 95% CI, 1.65 to 3.72; P < 0.01). Over the clinically relevant concentration range of 0 to 35 ng/ml, the oliceridine utility function was positive, indicating that the probability of analgesia exceeds the probability of respiratory depression. In contrast, the morphine function was negative, indicative of a greater probability of respiratory depression than analgesia. CONCLUSIONS: These data indicate a favorable oliceridine safety profile over morphine when considering analgesia and respiratory depression over the clinical concentration range.


Assuntos
Analgésicos Opioides/farmacologia , Morfina/farmacologia , Insuficiência Respiratória/induzido quimicamente , Compostos de Espiro/farmacologia , Tiofenos/farmacologia , Adulto , Analgésicos Opioides/efeitos adversos , Voluntários Saudáveis , Humanos , Masculino , Pessoa de Meia-Idade , Morfina/efeitos adversos , Valores de Referência , Medição de Risco , Compostos de Espiro/efeitos adversos , Tiofenos/efeitos adversos , Adulto Jovem
6.
Mol Pharmacol ; 98(4): 475-486, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32680919

RESUMO

Methadone is a synthetic opioid agonist with notoriously unique properties, such as lower abuse liability and induced relief of withdrawal symptoms and drug cravings, despite acting on the same opioid receptors triggered by classic opioids-in particular the µ-opioid receptor (MOR). Its distinct pharmacologic properties, which have recently been attributed to the preferential activation of ß-arrestin over G proteins, make methadone a standard-of-care maintenance medication for opioid addiction. Although a recent biophysical study suggests that methadone stabilizes different MOR active conformations from those stabilized by classic opioid drugs or G protein-biased agonists, how this drug modulates the conformational equilibrium of MOR and what specific active conformation of the receptor it stabilizes are unknown. Here, we report the results of submillisecond adaptive sampling molecular dynamics simulations of a predicted methadone-bound MOR complex and compare them with analogous data obtained for the classic opioid morphine and the G protein-biased ligand TRV130. The model, which is supported by existing experimental data, is analyzed using Markov state models and transfer entropy analysis to provide testable hypotheses of methadone-specific conformational dynamics and activation kinetics of MOR. SIGNIFICANCE STATEMENT: Opioid addiction has reached epidemic proportions in both industrialized and developing countries. Although methadone maintenance treatment represents an effective therapeutic approach for opioid addiction, it is not as widely used as needed. In this study, we contribute an atomic-level understanding of how methadone exerts its unique function in pursuit of more accessible treatments for opioid addiction. In particular, we present details of a methadone-specific active conformation of the µ-opioid receptor that has thus far eluded experimental structural characterization.


Assuntos
Analgésicos Opioides/farmacologia , Metadona/farmacologia , Receptores Opioides mu/química , Receptores Opioides mu/metabolismo , Compostos de Espiro/farmacologia , Tiofenos/farmacologia , Analgésicos Opioides/química , Animais , Sítios de Ligação , Entropia , Humanos , Cadeias de Markov , Metadona/química , Camundongos , Modelos Moleculares , Simulação de Dinâmica Molecular , Ligação Proteica , Conformação Proteica/efeitos dos fármacos , Compostos de Espiro/química , Tiofenos/química
7.
Sci Adv ; 6(28): eabb8097, 2020 07.
Artigo em Inglês | MEDLINE | ID: mdl-32691011

RESUMO

The prevalence of respiratory illness caused by the novel SARS-CoV-2 virus associated with multiple organ failures is spreading rapidly because of its contagious human-to-human transmission and inadequate globalhealth care systems. Pharmaceutical repurposing, an effective drug development technique using existing drugs, could shorten development time and reduce costs compared to those of de novo drug discovery. We carried out virtual screening of antiviral compounds targeting the spike glycoprotein (S), main protease (Mpro), and the SARS-CoV-2 receptor binding domain (RBD)-angiotensin-converting enzyme 2 (ACE2) complex of SARS-CoV-2. PC786, an antiviral polymerase inhibitor, showed enhanced binding affinity to all the targets. Furthermore, the postfusion conformation of the trimeric S protein RBD with ACE2 revealed conformational changes associated with PC786 drug binding. Exploiting immunoinformatics to identify T cell and B cell epitopes could guide future experimental studies with a higher probability of discovering appropriate vaccine candidates with fewer experiments and higher reliability.


Assuntos
Antivirais/farmacologia , Betacoronavirus/imunologia , Infecções por Coronavirus/prevenção & controle , Cisteína Endopeptidases/química , Desenho de Fármacos , Pandemias/prevenção & controle , Peptidil Dipeptidase A/química , Pneumonia Viral/prevenção & controle , Glicoproteína da Espícula de Coronavírus/química , Proteínas não Estruturais Virais/química , Benzamidas , Benzazepinas , Betacoronavirus/efeitos dos fármacos , Betacoronavirus/metabolismo , Sítios de Ligação , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/virologia , Cisteína Endopeptidases/imunologia , Cisteína Endopeptidases/metabolismo , Avaliação Pré-Clínica de Medicamentos , Epitopos de Linfócito B/efeitos dos fármacos , Epitopos de Linfócito B/imunologia , Epitopos de Linfócito T/efeitos dos fármacos , Epitopos de Linfócito T/imunologia , Humanos , Simulação de Acoplamento Molecular , Peptidil Dipeptidase A/imunologia , Peptidil Dipeptidase A/metabolismo , Pneumonia Viral/imunologia , Pneumonia Viral/virologia , Ligação Proteica , Conformação Proteica , Domínios Proteicos , Domínios e Motivos de Interação entre Proteínas , Glicoproteína da Espícula de Coronavírus/imunologia , Glicoproteína da Espícula de Coronavírus/metabolismo , Compostos de Espiro/farmacologia , Proteínas não Estruturais Virais/imunologia , Proteínas não Estruturais Virais/metabolismo
8.
PLoS Pathog ; 16(6): e1008485, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32589689

RESUMO

Ozonide antimalarials, OZ277 (arterolane) and OZ439 (artefenomel), are synthetic peroxide-based antimalarials with potent activity against the deadliest malaria parasite, Plasmodium falciparum. Here we used a "multi-omics" workflow, in combination with activity-based protein profiling (ABPP), to demonstrate that peroxide antimalarials initially target the haemoglobin (Hb) digestion pathway to kill malaria parasites. Time-dependent metabolomic profiling of ozonide-treated P. falciparum infected red blood cells revealed a rapid depletion of short Hb-derived peptides followed by subsequent alterations in lipid and nucleotide metabolism, while untargeted peptidomics showed accumulation of longer Hb-derived peptides. Quantitative proteomics and ABPP assays demonstrated that Hb-digesting proteases were increased in abundance and activity following treatment, respectively. Ozonide-induced depletion of short Hb-derived peptides was less extensive in a drug-treated K13-mutant artemisinin resistant parasite line (Cam3.IIR539T) than in the drug-treated isogenic sensitive strain (Cam3.IIrev), further confirming the association between ozonide activity and Hb catabolism. To demonstrate that compromised Hb catabolism may be a primary mechanism involved in ozonide antimalarial activity, we showed that parasites forced to rely solely on Hb digestion for amino acids became hypersensitive to short ozonide exposures. Quantitative proteomics analysis also revealed parasite proteins involved in translation and the ubiquitin-proteasome system were enriched following drug treatment, suggestive of the parasite engaging a stress response to mitigate ozonide-induced damage. Taken together, these data point to a mechanism of action involving initial impairment of Hb catabolism, and indicate that the parasite regulates protein turnover to manage ozonide-induced damage.


Assuntos
Adamantano/análogos & derivados , Antimaláricos/farmacologia , Eritrócitos , Hemoglobinas/metabolismo , Compostos Heterocíclicos com 1 Anel/farmacologia , Peróxidos/farmacologia , Plasmodium falciparum/metabolismo , Compostos de Espiro/farmacologia , Adamantano/farmacologia , Eritrócitos/metabolismo , Eritrócitos/parasitologia , Hemoglobinas/genética , Compostos Heterocíclicos/química , Compostos Heterocíclicos/farmacologia , Humanos , Plasmodium falciparum/genética , Proteômica
9.
Psychopharmacology (Berl) ; 237(7): 2075-2087, 2020 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-32372348

RESUMO

RATIONALE: Kappa-opioid receptor (KOR) agonists are antinociceptive but have side effects that limit their therapeutic utility. New KOR agonists have been developed that are fully efficacious at the KOR but may produce fewer or reduced side effects that are typical of KOR agonists. OBJECTIVES: We determined behavioral profiles for typical and atypical KOR agonists purported to differ in intracellular-signaling profiles as well as a mu-opioid receptor (MOR) agonist, oxycodone, using a behavioral scoring system based on Novak et al. (Am J Primatol 28:124-138, 1992, Am J Primatol 46:213-227, 1998) and modified to quantify drug-induced effects (e.g., Duke et al. J Pharmacol Exp Ther 366:145-157, 2018). METHODS: Six adult male rhesus monkeys were administered a range of doses of the typical KOR agonists, U50-488H (0.0032-0.1 mg/kg) and salvinorin A (0.00032-0.01 mg/kg); the atypical KOR agonists, nalfurafine (0.0001-0.001 mg/kg) and triazole 1.1 (0.01-0.32 mg/kg); the MOR agonist, oxycodone (0.0032-0.32 mg/kg); and as controls, cocaine (0.032-0.32 mg/kg) and ketamine (0.32-10 mg/kg). For time-course determinations, the largest dose of each KOR agonist or MOR agonist was administered across timepoints (10-320 min). In mixture conditions, oxycodone (0.1 mg/kg) was followed by KOR-agonist administration. RESULTS: Typical KOR agonists produced sedative-like and motor-impairing effects. Nalfurafine was similar to typical KOR agonists on most outcomes, and triazole 1.1 produced no effects on its own except for reducing scratch during time-course determinations. In the mixture, all KOR agonists reduced oxycodone-induced scratching, U50-488H and nalfurafine reduced species-typical activity, and U50-488H increased rest/sleep posture. CONCLUSIONS: Atypical "biased" KOR agonists produce side-effect profiles that are relatively benign (triazole 1.1) or reduced (nalfurafine) compared to typical KOR agonists.


Assuntos
Analgésicos Opioides/farmacologia , Atividade Motora/efeitos dos fármacos , Atividade Motora/fisiologia , Receptores Opioides kappa/agonistas , Receptores Opioides kappa/fisiologia , (trans)-Isômero de 3,4-dicloro-N-metil-N-(2-(1-pirrolidinil)-ciclo-hexil)-benzenoacetamida/farmacologia , Animais , Comportamento Animal/efeitos dos fármacos , Comportamento Animal/fisiologia , Diterpenos Clerodânicos/farmacologia , Relação Dose-Resposta a Droga , Macaca mulatta , Masculino , Morfinanos/farmacologia , Oxicodona/farmacologia , Compostos de Espiro/farmacologia
10.
Eur J Med Chem ; 194: 112240, 2020 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-32248003

RESUMO

Discovery and optimization of selective liver X receptor ß (LXRß) agonists are challenging due to the high homology of LXRα and LXRß in the ligand-binding domain. There is only one different residue (Val versus Ile) at the ligand-binding pocket of LXRs. With machine learning methods, we identified pan LXR agonists with a novel scaffold (spiro[pyrrolidine-3,3'-oxindole]). Then, we figured out the mechanism of LXR isoform selectivity from co-crystal structures. Based on the mechanism and the new scaffold, LXRß selective agonists were designed and synthesized. This led to the discovery of LXRß agonists 4-7rr, 4-13 and 4-13rr with IC50 values ranging from 1.78 to 6.36 µM against glioblastoma in vitro. Treatment with 50 mg/kg/day of 4-13 for 15 days significantly reduced tumor growth using an in vivo xenograft glioblastoma model.


Assuntos
Antineoplásicos/farmacologia , Descoberta de Drogas , Glioblastoma/tratamento farmacológico , Receptores X do Fígado/agonistas , Oxindois/farmacologia , Pirrolidinas/farmacologia , Compostos de Espiro/farmacologia , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Glioblastoma/metabolismo , Células HEK293 , Humanos , Receptores X do Fígado/metabolismo , Aprendizado de Máquina , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Estrutura Molecular , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Oxindois/síntese química , Oxindois/química , Pirrolidinas/síntese química , Pirrolidinas/química , Compostos de Espiro/síntese química , Compostos de Espiro/química , Relação Estrutura-Atividade , Células Tumorais Cultivadas
11.
Pharmacol Rep ; 72(2): 427-434, 2020 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-32002826

RESUMO

BACKGROUND: In our previous work, spiroxatrine was taken as reference compound to develop selective NOP ligands. Therefore, several triazaspirodecanone derivatives were synthesized. Here, we verify their selectivity towards other 5-HT1 receptor subtypes and with respect to α2-AR (Adrenergic Receptors). METHODS: Binding affinities were determined on cells expressing human cloned receptors for 5-HT1A/B/D and α2A/B/C subtypes. The Ki values were determined for those with at least 50% radioligand inhibition. RESULTS: All our derivatives show a moderate affinity for α2 subtypes, spanning from 5 to 7.5 pKi values. Moreover, they show affinity values in a µM-nM range at the 5-HT1A receptor, while they are practically inactive at 5-HT1B and 5-HT1D subtypes. Compound 11, the best of the series, has a 5-HT1A pKi value of 8.43 similar to spiroxatrine but, notably, it has a 5-HT1A favorable selectivity ratio of 52, 8 and 29, respectively over α2A, α2B and α2C adrenoceptor subtypes. CONCLUSIONS: In this SAR study, a 5-HT1A selective ligand has been identified in which a tetralone moiety replaced the 1,4-benzodioxane of spiroxatrine and the methylene linker to the triazaspirodecanone portion was maintained in position 2.


Assuntos
Dioxanos/farmacologia , Descoberta de Drogas , Receptor 5-HT1A de Serotonina/metabolismo , Compostos de Espiro/farmacologia , Animais , Ligação Competitiva , Células CHO , Cricetulus , Dioxanos/química , Dioxanos/metabolismo , Humanos , Ligantes , Estrutura Molecular , Ligação Proteica , Ensaio Radioligante , Receptor 5-HT1A de Serotonina/genética , Receptores Adrenérgicos alfa 2/genética , Receptores Adrenérgicos alfa 2/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Compostos de Espiro/química , Compostos de Espiro/metabolismo , Relação Estrutura-Atividade
12.
J Med Chem ; 63(7): 3552-3562, 2020 04 09.
Artigo em Inglês | MEDLINE | ID: mdl-32073266

RESUMO

We report the discovery of a novel indoleamine 2,3-dioxygenase-1 (IDO1) inhibitor class through the affinity selection of a previously unreported indole-based DNA-encoded library (DEL). The DEL exemplar, spiro-chromane 1, had moderate IDO1 potency but high in vivo clearance. Series optimization quickly afforded a potent, low in vivo clearance lead 11. Although amorphous 11 was highly bio-available, crystalline 11 was poorly soluble and suffered disappointingly low bio-availability because of solubility-limited absorption. A prodrug approach was deployed and proved effective in discovering the highly bio-available phosphonooxymethyl 31, which rapidly converted to 11 in vivo. Obtaining crystalline 31 proved problematic, however; thus salt screening was performed in an attempt to circumvent this obstacle and successfully delivered greatly soluble and bio-available crystalline tris-salt 32. IDO1 inhibitor 32 is characterized by a low calculated human dose, best-in-class potential, and an unusual inhibition mode by binding the IDO1 heme-free (apo) form.


Assuntos
DNA/química , Inibidores Enzimáticos/farmacologia , Indolamina-Pirrol 2,3,-Dioxigenase/antagonistas & inibidores , Pró-Fármacos/farmacologia , Compostos de Espiro/farmacologia , Animais , Descoberta de Drogas , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/farmacocinética , Eutérios , Masculino , Estrutura Molecular , Pró-Fármacos/síntese química , Pró-Fármacos/farmacocinética , Compostos de Espiro/síntese química , Compostos de Espiro/farmacocinética , Relação Estrutura-Atividade
13.
Eur J Med Chem ; 191: 112143, 2020 Apr 01.
Artigo em Inglês | MEDLINE | ID: mdl-32078865

RESUMO

Herein, we communicate our recent medicinal chemistry efforts which have culminated in a series of PI3Kδ/γ dual inhibitors structurally featuring a seven-membered spirocyclic spacer. Compound 26, the most potent one among them, exhibited superior PI3Kδ inhibitory activity (IC50 = 1.0 nM) to that of the approved PI3Kδ inhibitor Idelalisib. Besides, it exerted remarkable anti-proliferative efficacy against human malignant B-cell line SU-DHL-6 with GI50 value of 33 nM. The biochemical assay against the other three class I PI3K isoforms identified compound 26 as a potent PI3Kδ/γ dual inhibitor with considerable selectivity over PI3Kα and PI3Kß. In SU-DHL-6 cells, a dramatic down-regulation of PI3K signaling was observed following compound 26-treatment at the concentration as low as 10 nM. Inspiringly, the pharmacokinetic (PK) study in Sprague-Dawley (SD) rats revealed it was orally available with a favorable bioavailability (F = 87.5%). Overall, compound 26, a promising PI3Kδ/γ dual inhibitor, has the potential to emerge as a clinical candidate for the treatment of leukocyte-mediated malignancies after extensive functional investigation.


Assuntos
Antineoplásicos/farmacologia , Fosfatidilinositol 3-Quinases/metabolismo , Inibidores de Fosfoinositídeo-3 Quinase/farmacologia , Compostos de Espiro/farmacologia , Antineoplásicos/síntese química , Antineoplásicos/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Simulação de Acoplamento Molecular , Estrutura Molecular , Inibidores de Fosfoinositídeo-3 Quinase/síntese química , Inibidores de Fosfoinositídeo-3 Quinase/química , Compostos de Espiro/síntese química , Compostos de Espiro/química , Relação Estrutura-Atividade
14.
Org Biomol Chem ; 18(5): 931-940, 2020 02 07.
Artigo em Inglês | MEDLINE | ID: mdl-31922157

RESUMO

The design of glycogen phosphorylase (GP) inhibitors targeting the catalytic site of the enzyme is a promising strategy for a better control of hyperglycaemia in the context of type 2 diabetes. Glucopyranosylidene-spiro-heterocycles have been demonstrated as potent GP inhibitors, and more specifically spiro-oxathiazoles. A new synthetic route has now been elaborated through 1,3-dipolar cycloaddition of an aryl nitrile oxide to a glucono-thionolactone affording in one step the spiro-oxathiazole moiety. The thionolactone was obtained from the thermal rearrangement of a thiosulfinate precursor according to Fairbanks' protocols, although with a revisited outcome and also rationalised with DFT calculations. The 2-naphthyl substituted glucose-based spiro-oxathiazole 5h, identified as one of the most potent GP inhibitors (Ki = 160 nM against RMGPb) could be produced on the gram-scale from this strategy. Further evaluation in vitro using rat and human hepatocytes demonstrated that compound 5h is a anti-hyperglycaemic drug candidates performing slightly better than DAB used as a positive control. Investigation in Zucker fa/fa rat model in acute and subchronic assays further confirmed the potency of compound 5h since it lowered blood glucose levels by ∼36% at 30 mg kg-1 and ∼43% at 60 mg kg-1. The present study is one of the few in vivo investigations for glucose-based GP inhibitors and provides data in animal models for such drug candidates.


Assuntos
Inibidores Enzimáticos/farmacologia , Glucose/metabolismo , Glicogênio Fosforilase/antagonistas & inibidores , Hipoglicemiantes/farmacologia , Compostos de Espiro/farmacologia , Tiazóis/farmacologia , Animais , Glicemia/metabolismo , Ciclização , Teoria da Densidade Funcional , Glicogênio/metabolismo , Glicogênio Fosforilase/metabolismo , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Humanos , Hipoglicemiantes/síntese química , Hipoglicemiantes/química , Concentração Inibidora 50 , Cinética , Lactonas/síntese química , Lactonas/química , Oxirredução , Ratos Zucker , Compostos de Espiro/síntese química , Compostos de Espiro/química , Estereoisomerismo , Temperatura , Tiazóis/síntese química , Tiazóis/química
15.
Am J Physiol Heart Circ Physiol ; 318(3): H508-H518, 2020 03 01.
Artigo em Inglês | MEDLINE | ID: mdl-31975626

RESUMO

Cigarette smoking is a major risk factor for aortic aneurysm and dissection; however, no causative link between smoking and these aortic disorders has been proven. In the present study, we investigated the mechanism by which cigarette smoke affects vascular wall cells and found that cigarette smoke extract (CSE) induced a novel form of regulated cell death termed ferroptosis in vascular smooth muscle cells (VSMCs). CSE markedly induced cell death in A7r5 cells and primary rat VSMCs, but not in endothelial cells, which was completely inhibited by specific ferroptosis inhibitors [ferrostatin-1 (Fer-1) and Liproxstatin-1] and an iron chelator (deferoxamine). CSE-induced VSMC death was partially inhibited by a GSH precursor (N-acetyl cysteine) and an NADPH oxidase inhibitor [diphenyleneiodonium chloride (DPI)], but not by inhibitors of pan-caspases (Z-VAD), caspase-1 (Z-YVAD), or necroptosis (necrostatin-1). CSE also upregulated IL-1ß, IL-6, TNF-α, matrix metalloproteinase (MMP)-2, MMP-9, and TIMP-1 (tissue inhibitor of metalloproteinase)in A7r5 cells, which was inhibited by Fer-1. Furthermore, CSE induced the upregulation of Ptgs2 mRNA, lipid peroxidation, and intracellular GSH depletion, which are key features of ferroptosis. VSMC ferroptosis was induced by acrolein and methyl vinyl ketone, major constituents of CSE. Furthermore, CSE caused medial VSMC loss in ex vivo aortas. Electron microscopy analysis showed mitochondrial damage and fragmentation in medial VSMCs of CSE-treated aortas. All of these manifestations were partially restored by Fer-1. These findings demonstrate that ferroptosis is responsible for CSE-induced VSMC death and suggest that ferroptosis is a potential therapeutic target for preventing aortic aneurysm and dissection.NEW & NOTEWORTHY Cigarette smoke extract (CSE)-induced cell death in rat vascular smooth muscle cells (VSMCs) was completely inhibited by specific ferroptosis inhibitors and an iron chelator. CSE also induced the upregulation of Ptgs2 mRNA, lipid peroxidation, and intracellular GSH depletion, which are key features of ferroptosis. CSE caused medial VSMC loss in ex vivo aortas. These findings demonstrate that ferroptosis is responsible for CSE-induced VSMC death.


Assuntos
Ferroptose/efeitos dos fármacos , Músculo Liso Vascular/metabolismo , Miócitos de Músculo Liso/metabolismo , Fumaça , Animais , Morte Celular/efeitos dos fármacos , Linhagem Celular , Cicloexilaminas/farmacologia , Desferroxamina/farmacologia , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/metabolismo , Masculino , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/metabolismo , Miócitos de Músculo Liso/efeitos dos fármacos , NADPH Oxidases/metabolismo , Fenilenodiaminas/farmacologia , Quinoxalinas/farmacologia , Ratos , Ratos Sprague-Dawley , Sideróforos/farmacologia , Compostos de Espiro/farmacologia , Inibidor Tecidual de Metaloproteinase-1/metabolismo
16.
J Antibiot (Tokyo) ; 73(5): 290-298, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-31992865

RESUMO

The emergence of antibiotic resistance necessitates not only the identification of new compounds with antimicrobial properties, but also new strategies and combination therapies to circumvent this growing problem. Here, we report synergistic activity against methicillin-resistant Staphylococcus aureus (MRSA) of the ß-lactam antibiotic oxacillin combined with 7,8-dideoxygriseorhodin C in vitro. Ongoing efforts to identify antibiotics from marine mollusk-associated bacteria resulted in the isolation of 7,8-dideoxygriseorhodin C from a Streptomyces sp. strain cultivated from a marine gastropod tissue homogenate. Despite the long history of 7,8-dideoxygriseorhodin C in the literature, the absolute configuration has never been previously reported. A comparison of measured and calculated ECD spectra resolved the configuration of the spiroketal carbon C6, and 2D ROESY NMR spectroscopy established the absolute configuration as 6s,6aS. The compound is selective against Gram-positive bacteria including MRSA and Enterococcus faecium with an MIC range of 0.125-0.5 µg ml-1. Moreover, the compound synergizes with oxacillin against MRSA as observed in the antimicrobial microdilution and time-kill assays. Simultaneous treatment of the compound with oxacillin resulted in an approximately tenfold decrease in MIC with a combination index of <0.5, indicating synergistic anti-MRSA activity.


Assuntos
Antibacterianos/farmacologia , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Oxacilina/farmacologia , Antibacterianos/administração & dosagem , Antibacterianos/isolamento & purificação , Sinergismo Farmacológico , Enterococcus faecium/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Naftoquinonas/administração & dosagem , Naftoquinonas/química , Naftoquinonas/isolamento & purificação , Naftoquinonas/farmacologia , Oxacilina/administração & dosagem , Compostos de Espiro/administração & dosagem , Compostos de Espiro/química , Compostos de Espiro/isolamento & purificação , Compostos de Espiro/farmacologia , Streptomyces/metabolismo
17.
Eur J Med Chem ; 188: 111977, 2020 Feb 15.
Artigo em Inglês | MEDLINE | ID: mdl-31927313

RESUMO

a series of 2-oxospiro[indoline-3,4'-pyran]derivatives 4 and 7 were obtained in good yield under mild conditions from the one-pot reaction of indole-2,3-dione derivatives 1, appropriate methylene active nitriles 2 and ß-dicarbonyl compound 3 or 6. The newly synthesized compounds were characterized and evaluated for their in vitro antibacterial, antifungal as well as immunomodulatory activity. According to MIC values, the most potent compounds 4f, 4h, 7a, 7c, 7e, 7f, 7g, 8a, and 8c were evaluated for MBC and displayed high activity to killing pathogens with a good MBC value against norfloxacin as well as investigated against an extended panel of multidrug resistance bacteria (MDRB) and exhibited promising to moderate multidrug resistance activities, compounds 7f showed the much better than norfloxacin with higher potency results. Furthermore, the most potent compounds showed an increase in the intracellular killing activity of neutrophils which confirmed the immunostimulatory power. Eight of the nine active compounds exhibited inhibitory activities with IC50 ranged between (18.07 ± 0.18) to (27.03 ± 0.24) µM stronger than ciprofloxacin (26.43 ± 0.64 µM) for S. aureus DNA gyrase. Molecular docking was performed inside the active site of S. aureus DNA gyrase to predict the binding mode.


Assuntos
Antibacterianos/farmacologia , Antifúngicos/farmacologia , Sulfonatos de Arila/farmacologia , Fatores Imunológicos/farmacologia , Compostos de Espiro/farmacologia , Inibidores da Topoisomerase/farmacologia , Antibacterianos/síntese química , Antibacterianos/química , Antifúngicos/síntese química , Antifúngicos/química , Sulfonatos de Arila/síntese química , Sulfonatos de Arila/química , DNA Girase/metabolismo , Relação Dose-Resposta a Droga , Fungos/efeitos dos fármacos , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Fatores Imunológicos/síntese química , Fatores Imunológicos/química , Testes de Sensibilidade Microbiana , Simulação de Acoplamento Molecular , Estrutura Molecular , Compostos de Espiro/síntese química , Compostos de Espiro/química , Relação Estrutura-Atividade , Inibidores da Topoisomerase/síntese química , Inibidores da Topoisomerase/química
18.
Sci Rep ; 10(1): 1323, 2020 Jan 28.
Artigo em Inglês | MEDLINE | ID: mdl-31992750

RESUMO

Herbs may contain pesticide residues which are an important discriminator of food security and food quality. The challenge of the research was to assess the fate of the herbicide clethodim (CLE) and the insecticide spirotetramat (SPI) applied in herbs (BBCH 11-21) during herb growth and processing under controlled greenhouse trial conditions. The metabolic profile of CLE and SPI and their degradation products in basil (Ocimum basilicum L.), peppermint (Mentha × piperita L.) and sage (Salvia officinalis L.) was also presented. The half-lives of CLE and SPI in herbs were 1.10-1.56 days and 0.51-0.83 days, respectively. The terminal residues of SPI (SPI-enol, SPI-ketohydroxy, SPI-monohydroxy and SPI-enol-glucoside) and CLE (CLE-sulfone and CLE-sulfoxide) in herbal matrices were measured below EU maximum residue limits. In this paper, we aimed to assess the impact of washing and dehydratation pretreatment and calculated processing factors (PFs) which can be applied to more accurate food safety assessments. The PF values of CLE and SPI after drying prior washing was below 1 indicating reduction of initial residues. Drying process without washing demonstrated increases of SPI concentrations (PF up to 1.50). The lowest PFs were obtained when raw herbal plants were washed before drying showing almost complete degradation of parent compound (93-99%).


Assuntos
Compostos Aza/farmacologia , Cicloexanonas/farmacologia , Inseticidas/farmacologia , Metaboloma/efeitos dos fármacos , Desenvolvimento Vegetal/efeitos dos fármacos , Plantas Medicinais/efeitos dos fármacos , Plantas Medicinais/metabolismo , Compostos de Espiro/farmacologia , Cromatografia Líquida , Meio Ambiente , Metabolômica/métodos , Plantas Medicinais/crescimento & desenvolvimento , Reprodutibilidade dos Testes , Espectrometria de Massas em Tandem
19.
J Pharmacol Exp Ther ; 373(1): 34-43, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-31937563

RESUMO

The peptide nociceptin/orphanin FQ (N/OFQ) is the natural ligand of the N/OFQ receptor (NOP), which is widely expressed in the central and peripheral nervous system. Selective NOP antagonists are worthy of testing as innovative drugs to treat depression, Parkinson disease, and drug abuse. The aim of this study was to perform a detailed in vitro characterization of BTRX-246040 (also known as LY2940094, [2-[4-[(2-chloro-4,4-difluoro-spiro[5H-thieno[2,3-c]pyran-7,4'-piperidine]-1'-yl)methyl]-3-methyl-pyrazol-1-yl]-3-pyridyl]methanol), a novel NOP antagonist that has been already studied in humans. BTRX-246040 has been tested in vitro in the following assays: calcium mobilization in cells expressing NOP and classic opioid receptors and chimeric G proteins, bioluminescence resonance energy transfer assay measuring NOP interaction with G proteins and ß-arrestins, the label-free dynamic mass redistribution assay, and the electrically stimulated mouse vas deferens. BTRX-246040 was systematically compared with the standard NOP antagonist SB-612111. In all assays, BTRX-246040 behaves as a pure and selective antagonist at human recombinant and murine native NOP receptors displaying 3-10-fold higher potency than the standard antagonist SB-612111. BTRX-246040 is an essential pharmacological tool to further investigate the therapeutic potential of NOP antagonists in preclinical and clinical studies. SIGNIFICANCE STATEMENT: NOP antagonists may be innovative antidepressant drugs. In this research, the novel clinically viable NOP antagonist BTRX-246040 has been deeply characterized in vitro in a panel of assays. BTRX-246040 resulted a pure, potent, and selective NOP antagonist.


Assuntos
Piranos/farmacologia , Receptores Opioides/fisiologia , Compostos de Espiro/farmacologia , Animais , Células CHO , Cricetinae , Cricetulus , Relação Dose-Resposta a Droga , Células HEK293 , Humanos , Masculino , Camundongos
20.
J Neurochem ; 152(5): 523-541, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-31376158

RESUMO

N-methyl-d-aspartate receptors (NMDARs) mediate both physiological and pathophysiological processes, although selective ligands lack broad clinical utility. NMDARs are composed of multiple subunits, but N-methyl-d-aspartate receptor subunit 2 (GluN2) is predominately responsible for functional heterogeneity. Specifically, the GluN2A- and GluN2B-containing subtypes are enriched in adult hippocampus and cortex and impact neuronal communication via dynamic trafficking into and out of the synapse. We sought to understand if ((2S, 3R)-3-hydroxy-2-((R)-5-isobutyryl-1-oxo-2,5-diazaspiro[3,4]octan-2-yl) butanamide (NYX-2925), a novel NMDAR modulator, alters synaptic levels of GluN2A- or GluN2B-containing NMDARs. Low-picomolar NYX-2925 increased GluN2B colocalization with the excitatory post-synaptic marker post-synaptic density protein 95 (PSD-95) in rat primary hippocampal neurons within 30 min. Twenty-four hours following oral administration, 1 mg/kg NYX-2925 increased GluN2B in PSD-95-associated complexes ex vivo, and low-picomolar NYX-2925 regulated numerous trafficking pathways in vitro. Because the NYX-2925 concentration that increases synaptic GluN2B was markedly below that which enhances long-term potentiation (mid-nanomolar), we sought to elucidate the basis of this effect. Although NMDAR-dependent, NYX-2925-mediated colocalization of GluN2B with PSD-95 occurred independent of ion flux, as colocalization increased in the presence of either the NMDAR channel blocker (5R,10S)-(-)-5-Methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate or glycine site antagonist 7-chlorokynurenic acid. Moreover, while mid-nanomolar NYX-2925 concentrations, which do not increase synaptic GluN2B, enhanced calcium transients, functional plasticity was only enhanced by picomolar NYX-2925. Thus, NYX-2925 concentrations that increase synaptic GluN2B facilitated the chemical long-term potentiation induced insertion of synaptic α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor GluA1 subunit levels. Basal (unstimulated by chemical long-term potentiation) levels of synaptic GluA1 were only increased by mid-nanomolar NYX-2925. These data suggest that NYX-2925 facilitates homeostatic plasticity by initially increasing synaptic GluN2B via metabotropic-like NMDAR signaling. Cover Image for this issue: doi: 10.1111/jnc.14735.


Assuntos
Receptores de AMPA/metabolismo , Receptores de N-Metil-D-Aspartato/metabolismo , Compostos de Espiro/farmacologia , Sinapses/metabolismo , Animais , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Potenciação de Longa Duração/efeitos dos fármacos , Potenciação de Longa Duração/fisiologia , Masculino , Ratos , Ratos Sprague-Dawley , Receptores de AMPA/efeitos dos fármacos , Receptores de N-Metil-D-Aspartato/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos , Sinapses/efeitos dos fármacos
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