RESUMO
Recent evidence indicates that the presence of a primary cilium (PC), and of selective cAMP signaling within this smallest of organelles, promotes adipogenic differentiation of 3T3-L1 preadipocytes incubated in media supplemented with either a natural (docosahexaenoic acid, DHA), or a synthetic (TUG-891), free fatty acid receptor 4 (FFAR4) agonist. Indeed, in this earlier work, activation of ciliary FFAR4 in 3T3-L1 cells was correlated with selective increases in PC cAMP and adipogenesis in these cells. However, this study was silent on the role of local PC cAMP phosphodiesterases (PDEs)-mediated events in regulating these adipogenic responses and on the identity of cAMP PDEs that could regulate the "pool" of ciliary cAMP accessed by FFAR4 agonists. In this context, we have identified the PDEs expressed by 3T3-L1 preadipocytes and showed that of these, only PDE4 inhibition promotes FFAR4-mediated adipogenesis. We propose that this work will identify more selective therapeutic targets through which to control adipogenesis, and perhaps the differentiation of other stem cells in which ciliary cAMP is critical.
Assuntos
Adipogenia , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4 , Camundongos , Animais , Células 3T3-L1 , Diferenciação Celular , Ácidos Docosa-Hexaenoicos , PPAR gamaRESUMO
Medicinal herbs have been used as traditional medicines for centuries. The molecular mechanism of action of their bioactive molecules against various diseases or therapeutic targets is still being explored. Here, the active compounds (withanolides) of a well-known Indian medicinal herb, Ashwagandha (Withania somnifera), have been studied for their most potential therapeutic targets and their mechanism of action using ligand-based screening and receptor-based approaches. Ligand-based screening predicted the six top therapeutic targets, namely, Protein kinase C alpha (PRKCA), Protein kinase C delta (PRKCD), Protein kinase C epsilon (PRKCE), Androgenic Receptor (AR), Cycloxygenase-2 (PTGS-2) and Phosphodiesterase-4D (PDE4D). Further, when these predictions were validated using receptor-based studies, i.e. molecular docking, molecular dynamics simulation and free energy calculations, it was found that PDE4D was the most potent target for four withanolides, namely, Withaferin-A, 17-Hydroxywithaferin-A, 27-Hydroxywithanone and Withanolide-R. These compounds had a better binding affinity and similar interactions as that of an already known inhibitor (Zardaverine) of PDE4D. These results warrant further in-vitro and in-vivo investigations to examine their therapeutic potential as an inhibitor of PDE4D.Communicated by Ramaswamy H. Sarma.
Assuntos
Inibidores da Fosfodiesterase 4 , Plantas Medicinais , Withania , Vitanolídeos , Vitanolídeos/farmacologia , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Simulação de Acoplamento Molecular , Inibidores da Fosfodiesterase 4/farmacologia , Ligantes , Withania/químicaRESUMO
Excessive alcohol use often results in alcoholic liver disease (ALD). An early change in the liver due to excessive drinking is hepatic steatosis, which may ultimately progress to hepatitis, liver fibrosis, cirrhosis, and liver cancer. Among these debilitating processes, hepatic steatosis is reversible with the appropriate treatment. Therefore, it is important to find treatments and foods that reverse hepatic steatosis. Black carrot has antioxidant and anti-inflammatory effects. In this study, we examined the effectiveness of black carrot extract (BCE) on hepatic steatosis in in vivo and in vitro ethanol-induced liver injury models. For the in vivo experiments, serum aminotransferase activities enhanced by ethanol- and carbon tetrachloride were significantly suppressed by the BCE diet. Furthermore, morphological changes in the liver hepatic steatosis and fibrosis were observed in the in vivo ethanol-induced liver injury model, however, BCE feeding resulted in the recovery to an almost normal liver morphology. In the in vitro experiments, ethanol treatment induced reactive oxygen species (ROS) levels in hepatocytes at 9 h. Conversely, ROS production was suppressed to control levels and hepatic steatosis was suppressed when hepatocyte culture with ethanol were treated with BCE. Furthermore, we investigated enzyme activities, enzyme protein levels, and messenger RNA levels of alcohol dehydrogenase (ADH), cytochrome p450 2E1 (CYP2E1), and aldehyde dehydrogenase (ALDH) using enzyme assays, western blot, and quantitative reverse transcription-polymerase chain reaction analyses. We found that the activities of ADH, CYP2E1, and ALDH were regulated through the cAMP-PKA pathway at different levels, namely, translational, posttranslational, and transcriptional levels, respectively. The most interesting finding of this study is that BCE increases cAMP levels by suppressing the Pde4b mRNA and PDE4b protein levels in ethanol-treated hepatocytes, suggesting that BCE may prevent ALD.
Assuntos
Doença Hepática Crônica Induzida por Substâncias e Drogas , Daucus carota , Fígado Gorduroso , Hepatopatias Alcoólicas , Etanol/metabolismo , Citocromo P-450 CYP2E1/genética , Citocromo P-450 CYP2E1/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Daucus carota/metabolismo , Álcool Desidrogenase/genética , Álcool Desidrogenase/metabolismo , Álcool Desidrogenase/farmacologia , Antioxidantes/farmacologia , RNA Mensageiro/metabolismo , Tetracloreto de Carbono , Fígado/metabolismo , Fígado Gorduroso/metabolismo , Aldeído Desidrogenase/metabolismo , Aldeído Desidrogenase/farmacologia , Cirrose Hepática , Transaminases/metabolismo , Anti-Inflamatórios/farmacologiaRESUMO
We studied the epigenetic regulation of how black carrot extract (BCE) protects against ethanol-induced hepatic damage. We have shown that the butanol-extracted fraction of BCE (BCE-BuOH) increased intracellular cyclic adenosine monophosphate (cAMP) levels by suppressing the expression of phosphodiesterase 4b (PDE4b); however, the detailed mechanism remains to be elucidated. We focused on changes in histone modifications involved in the suppression of pde4 expression. The methylation level of histone H3 lysine 9 (H3K9), which regulates gene expression of PDE4b, decreased after treatment with 100 mM ethanol but was significantly increased by treatment with 400 µg/ml BCE-BuOH. In contrast, ethanol induced an increase in H3K9 acetylation. However, treatment with BCE-BuOH inhibited the increase in acetylation through an increase in Sirtuin 1 (Sirt1), a histone deacetylase. Furthermore, BCE-BuOH treatment increased the level of methionine adenosyltransferase (MAT) 2a mRNA and increased intracellular S-adenosylmethionine. The present results indicate that BCE-BuOH is useful for protection against alcohol-induced hepatic injury. PRACTICAL APPLICATIONS: We have reported that black carrot extract (BCE) suppressed liver steatosis and liver fibrosis on a rat alcoholic liver disease model. The results from this study have shown that BCE regulated the alcoholic-induced hepatic injury at the level of epigenetic modifications. These results suggested that BCE is useful for protection against alcoholic-induced hepatic injury.
Assuntos
Daucus carota , Epigênese Genética , Monofosfato de Adenosina , Animais , Butanóis , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Daucus carota/genética , Etanol , Histonas/metabolismo , Lisina/metabolismo , Metionina Adenosiltransferase/metabolismo , Extratos Vegetais , RNA Mensageiro , Ratos , S-Adenosilmetionina/metabolismo , Sirtuína 1/genética , Sirtuína 1/metabolismoRESUMO
Asthma affects millions of people worldwide and its prevalence is increasing. It is characterized by chronic airway inflammation, airway remodeling, and pathologic bronchoconstriction, and it poses a continuous treatment challenge with very few new therapeutics available. Thus, many asthmatics turn to plant-based complementary products, including ginger, for better symptom control, indicating an unmet need for novel therapies. Previously, we demonstrated that 6-shogaol (6S), the primary bioactive component of ginger, relaxes human airway smooth muscle (hASM) likely by inhibition of phosphodiesterases (PDEs) in the ß-adrenergic (cyclic nucleotide PDEs), and muscarinic (phospholipase C, PLC) receptor pathways. However, oral 6S is extensively metabolized and it is unknown if the resulting metabolites remain bioactive. Here, we screened all the known human metabolites of 6S and several metabolite-based synthetic derivatives to better understand their mechanism of action and structure-function relationships. We demonstrate that several metabolites and metabolite-based synthetic derivatives are able to prevent Gq-coupled stimulation of intracellular calcium [Ca2+]i and inositol trisphosphate (IP3) synthesis by inhibiting PLC, similar to the parent compound 6S. We also show that these compounds prevent recontraction of ASM after ß-agonist relaxation likely by inhibiting PDEs. Furthermore, they potentiate isoproterenol-induced relaxation. Importantly, moving beyond cell-based assays, metabolites also retain the functional ability to relax Gq-coupled-contractions in upper (human) and lower (murine) airways. The current study indicates that, although oral ginger may be metabolized rapidly, it retains physiological activity through its metabolites. Moreover, we are able to use naturally occurring metabolites as inspiration to develop novel therapeutics for brochoconstrictive diseases.
Assuntos
Cálcio/metabolismo , Relaxamento Muscular/efeitos dos fármacos , Miócitos de Músculo Liso/efeitos dos fármacos , Zingiber officinale , Animais , Asma/induzido quimicamente , Asma/metabolismo , Broncoconstrição/efeitos dos fármacos , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Zingiber officinale/metabolismo , Humanos , Isoproterenol/farmacologia , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Camundongos Endogâmicos C57BL , Relaxamento Muscular/fisiologia , Músculo Liso/metabolismo , Miócitos de Músculo Liso/metabolismoRESUMO
Membrane glycoprotein is the most abundant protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), but its role in coronavirus disease 2019 (COVID-19) has not been fully characterized. Mice intranasally inoculated with membrane glycoprotein substantially increased the interleukin (IL)-6, a hallmark of the cytokine storm, in bronchoalveolar lavage fluid (BALF), compared to mice inoculated with green fluorescent protein (GFP). The high level of IL-6 induced by membrane glycoprotein was significantly diminished in phosphodiesterase 4 (PDE4B) knockout mice, demonstrating the essential role of PDE4B in IL-6 signaling. Mycelium fermentation of Lactobacillus rhamnosus (L. rhamnosus) EH8 strain yielded butyric acid, which can down-regulate the PDE4B expression and IL-6 secretion in macrophages. Feeding mice with mycelia increased the relative abundance of commensal L. rhamnosus. Two-week supplementation of mice with L. rhamnosus plus mycelia considerably decreased membrane glycoprotein-induced PDE4B expression and IL-6 secretion. The probiotic activity of L. rhamnosus plus mycelia against membrane glycoprotein was abolished in mice treated with GLPG-0974, an antagonist of free fatty acid receptor 2 (Ffar2). Activation of Ffar2 in the gut-lung axis for down-regulation of the PDE4B-IL-6 signalling may provide targets for development of modalities including probiotics for treatment of the cytokine storm in COVID-19.
Assuntos
Proteínas M de Coronavírus/farmacologia , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Interleucina-6/metabolismo , Lacticaseibacillus rhamnosus/fisiologia , Probióticos/farmacologia , SARS-CoV-2/metabolismo , Animais , Ácido Butírico , Linhagem Celular , Clonagem Molecular , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/genética , Feminino , Fermentação , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Interleucina-6/genética , Camundongos , Camundongos Endogâmicos ICR , Receptores Acoplados a Proteínas G/metabolismoRESUMO
LY2775240 is a highly selective, potent and orally-administered inhibitor of phosphodiesterase 4 (PDE4), and is being investigated as a treatment option for inflammatory disorders, such as psoriasis. LY2775240 was investigated in rodent and rhesus monkey nonclinical models. Treatment with LY2775240 led to significant reductions in TNFα production, a marker of PDE4 engagement upon immune activation, in both nonclinical models. In the first part of a 2-part first-in-human randomized study, a wide dose range of LY2775240 was safely evaluated and found to be well-tolerated with common adverse events (AEs) of nausea, diarrhea, and headache. No serious AEs were reported. The pharmacokinetic profile of LY2775240 was well-characterized, with a half-life that can support once-a-day dosing. An ex vivo pharmacodynamic (PD) assay demonstrated dose-dependent PDE4 target engagement as assessed by reduction in TNFα production. A 20 mg dose of LY2775240 led to near-maximal TNFα inhibition in this PD assay in the first part of the study and was selected for comparison with the clinical dose of apremilast (30 mg) in the crossover, second part of this study. The 20 mg dose of LY2775240 demonstrated sustained maximal (50%-80%) inhibition of TNFα over all timepoints over the 24-h duration. The comparator apremilast achieved peak inhibition of ~ 50% at only 4 h postdose with a return to about 10% inhibition within 12 h of dosing. In summary, the nonclinical data and safety, tolerability, and PK/PD data in healthy subjects supports further investigation of LY2775240 in inflammatory indications. Study Highlights WHAT IS THE CURRENT KNOWLEDGE ON THE TOPIC? Phosphodiesterase 4 (PDE4) inhibitors, such as apremilast, are currently approved to treat autoimmune disorders, such as psoriasis. LY2775240 is an oral PDE4 inhibitor being developed for treatment of a variety of inflammatory disorders. The degree of enzymatic inhibition achieved by PDE4 inhibitors clinically is poorly understood. WHAT QUESTION DID THIS STUDY ADDRESS? This study investigated single ascending doses of LY2775240, a highly selective oral PDE4 inhibitor, in healthy subjects. LY2775240 was well-tolerated over the dose range evaluated, and pharmacokinetic/pharmacodynamic (PD) profiles were well-characterized. WHAT DOES THIS STUDY ADD TO OUR KNOWLEDGE? This study evaluated different doses of LY2775240 and subsequently compared a selected LY2775240 dose with the clinical dose of apremilast with an ex vivo assay. This information builds a connection between target engagement and clinical efficacy. HOW MIGHT THIS CHANGE CLINICAL PHARMACOLOGY OR TRANSLATIONAL SCIENCE? This is the first report of an ex vivo PD assay that has been systematically implemented in a PDE4 inhibitor Phase 1 study. Early investigation of exposure-response relationships versus a comparator can support evaluation of clinically meaningful doses of investigational agents.
Assuntos
Drogas em Investigação/farmacologia , Inibidores da Fosfodiesterase 4/farmacologia , Administração Oral , Adulto , Animais , Estudos Cross-Over , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Avaliação Pré-Clínica de Medicamentos , Drogas em Investigação/uso terapêutico , Ensaios Enzimáticos , Feminino , Voluntários Saudáveis , Humanos , Macaca mulatta , Masculino , Camundongos , Pessoa de Meia-Idade , Inibidores da Fosfodiesterase 4/uso terapêutico , Psoríase/tratamento farmacológico , Talidomida/análogos & derivados , Talidomida/farmacologia , Talidomida/uso terapêuticoRESUMO
Diabetic nephropathy (DN) is a leading cause of end-stage renal disease (ESRD). Hypertension increases kidney stress, which deteriorates function, and leads to peripheral renal vascular resistance. Long-term hypoperfusion promotes interstitial fibrosis and glomerular sclerosis, resulting in nephrosclerosis. Although hypertension and DN are frequent ESRD complications, relevant animal models remain unavailable. We generated a deoxycorticosterone acetate (DOCA)-salt hypertensive uni-nephrectomized (UNx) KKAy mouse model demonstrating hypertension, hyperglycemia, cardiac hypertrophy, kidney failure, increased urinary albumin creatinine ratio (UACR), and increased renal PDE4D and cardiac PDE5A mRNA levels. We hypothesized that the novel PDE4 selective inhibitor, compound A, and PDE5 inhibitor, sildenafil, exhibit nephroprotective, and cardioprotective effects in this new model. Compound A, sildenafil, and the angiotensin II receptor blocker, irbesartan, significantly reduced ventricular hypertrophy and pleural effusion volume. Meanwhile, compound A and sildenafil significantly suppressed the UACR, urinary kidney injury molecule-1, and monocyte chemoattractant protein-1 levels, as well as that of renal pro-fibrotic marker mRNAs, including collagen 1A1, fibronectin, and transforming growth factor-beta (TGF-ß). Moreover, compound A significantly suppressed TGF-ß-induced pro-fibrotic mRNA expression in vitro in all major kidney lesions, including within the glomerular mesangial region, podocytes, and epithelial region. Hence, PDE4 and PDE5 inhibitors may be promising treatments, in combination with irbesartan, for DN with hypertension as they demonstrate complementary mechanisms.
Assuntos
Cardiomegalia/tratamento farmacológico , Desoxicorticosterona/toxicidade , Hiperglicemia/tratamento farmacológico , Hipertensão/tratamento farmacológico , Inibidores da Fosfodiesterase 5/farmacologia , Insuficiência Renal/tratamento farmacológico , Citrato de Sildenafila/farmacologia , Acetatos/farmacologia , Animais , Anti-Inflamatórios não Esteroides/farmacologia , Cardiomegalia/induzido quimicamente , Cardiomegalia/enzimologia , Cardiomegalia/patologia , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/química , Nucleotídeo Cíclico Fosfodiesterase do Tipo 5/química , Feminino , Hiperglicemia/induzido quimicamente , Hiperglicemia/enzimologia , Hiperglicemia/patologia , Hipertensão/induzido quimicamente , Hipertensão/enzimologia , Hipertensão/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mineralocorticoides/toxicidade , Insuficiência Renal/induzido quimicamente , Insuficiência Renal/enzimologia , Insuficiência Renal/patologia , Cloreto de Sódio/toxicidade , Tiramina/análogos & derivados , Tiramina/farmacologiaRESUMO
Despite the great search for an effective approach to minimize secondary injury in spinal cord injury (SCI) setting, there have been limited advances. Roflumilast is a selective inhibitor of phosphodiesterase 4 with potent anti-inflammatory properties. Here, we sought to explore Roflumilast efficacy in the improvement of locomotor and sensory deficits of SCI. In an animal setting, 50 male rats were randomly assigned to five groups: an SCI group receiving Placebo, three SCI groups receiving Roflumilast at the doses of 0.25, 0.5, and 1 mg/kg prior to T9 vertebra laminectomy, and a sham-operated group. Locomotor, mechanical, and thermal activities were evaluated for 28 days. At the end of the study, spinal cord samples were taken to assess the relative ratio of microglial subtypes, including M1 and M2, histopathological changes, levels of pro-inflammatory (TNF-α and IL-1ß) and anti-inflammatory (IL-10) biomarkers, and cAMP level. Repeated measure analysis revealed significant effect for time-treatment interaction on locomotion [F (24, 270) = 280.7, p < 0.001], thermal sensitivity [F (16, 180) = 4.35, p < 0.001], and mechanical sensitivity [F (16, 180) = 7.96, p < 0.001]. As expected, Roflumilast significantly increased the expression of spinal cAMP. H&E staining exhibited lesser histopathological disruptions in Roflumilast-treated rodents. We also observed a significant reduction in the M1/M2 ratio (p values < 0.001) as well as in pro-inflammatory biomarkers following the administration of Roflumilast to the injured rats. Furthermore, IL-10 level was increased in rodents receiving 1 mg/kg of the reagent. In conclusion, the increased spinal cAMP following Roflumilast therapy might attenuate neuroinflammation via altering microglial activity; therefore, it could be considered as an alternative therapeutic agent for SCI complications.
Assuntos
Agnosia/metabolismo , Aminopiridinas/uso terapêutico , Benzamidas/uso terapêutico , Microglia/metabolismo , Inibidores da Fosfodiesterase 4/uso terapêutico , Traumatismos da Medula Espinal/metabolismo , Medula Espinal/metabolismo , Tabes Dorsal/metabolismo , Agnosia/etiologia , Agnosia/prevenção & controle , Animais , AMP Cíclico/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Ciclopropanos/uso terapêutico , Modelos Animais de Doenças , Humanos , Masculino , Microglia/patologia , Inflamação Neurogênica , Ratos , Medula Espinal/patologia , Traumatismos da Medula Espinal/complicações , Traumatismos da Medula Espinal/tratamento farmacológico , Tabes Dorsal/etiologia , Tabes Dorsal/prevenção & controleRESUMO
OBJECTIVE: Salt-induced kinase 1 (SIK1) acts as a key modulator in many physiological processes. However, the effects of SIK1 on gluconeogenesis and the underlying mechanisms have not been fully elucidated. In this study, we found that a natural compound phanginin A could activate SIK1 and further inhibit gluconeogenesis. The mechanisms by which phanginin A activates SIK1 and inhibits gluconeogenesis were explored in primary mouse hepatocytes, and the effects of phanginin A on glucose homeostasis were investigated in ob/ob mice. METHODS: The effects of phanginin A on gluconeogenesis and SIK1 phosphorylation were examined in primary mouse hepatocytes. Pan-SIK inhibitor and siRNA-mediated knockdown were used to elucidate the involvement of SIK1 activation in phanginin A-reduced gluconeogenesis. LKB1 knockdown was used to explore how phanginin A activated SIK1. SIK1 overexpression was used to evaluate its effect on gluconeogenesis, PDE4 activity, and the cAMP pathway. The acute and chronic effects of phanginin A on metabolic abnormalities were observed in ob/ob mice. RESULTS: Phanginin A significantly increased SIK1 phosphorylation through LKB1 and further suppressed gluconeogenesis by increasing PDE4 activity and inhibiting the cAMP/PKA/CREB pathway in primary mouse hepatocytes, and this effect was blocked by pan-SIK inhibitor HG-9-91-01 or siRNA-mediated knockdown of SIK1. Overexpression of SIK1 in hepatocytes increased PDE4 activity, reduced cAMP accumulation, and thereby inhibited gluconeogenesis. Acute treatment with phanginin A reduced gluconeogenesis in vivo, accompanied by increased SIK1 phosphorylation and PDE4 activity in the liver. Long-term treatment of phanginin A profoundly reduced blood glucose levels and improved glucose tolerance and dyslipidemia in ob/ob mice. CONCLUSION: We discovered an unrecognized effect of phanginin A in suppressing hepatic gluconeogenesis and revealed a novel mechanism that activation of SIK1 by phanginin A could inhibit gluconeogenesis by increasing PDE4 activity and suppressing the cAMP/PKA/CREB pathway in the liver. We also highlighted the potential value of phanginin A as a lead compound for treating type 2 diabetes.
Assuntos
Diterpenos/farmacologia , Proteínas Serina-Treonina Quinases/metabolismo , Animais , Caesalpinia/metabolismo , AMP Cíclico/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/genética , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Diabetes Mellitus Tipo 2/metabolismo , Gluconeogênese/efeitos dos fármacos , Glucose/metabolismo , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Fígado/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fosforilação , Extratos Vegetais/farmacologia , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/efeitos dos fármacos , Transdução de SinaisRESUMO
Phosphodiesterase (PDE) inhibitors are currently a widespread and extensively studied group of anti-inflammatory and anti-fibrotic compounds which may find use in the treatment of numerous lung diseases, including asthma and chronic obstructive pulmonary disease. Several PDE inhibitors are currently in clinical development, and some of them, e.g., roflumilast, are already recommended for clinical use. Due to numerous reports indicating that elevated intracellular cAMP levels may contribute to the alleviation of inflammation and airway fibrosis, new and effective PDE inhibitors are constantly being sought. Recently, a group of 7,8-disubstituted purine-2,6-dione derivatives, representing a novel and prominent pan-PDE inhibitors has been synthesized. Some of them were reported to modulate transient receptor potential ankyrin 1 (TRPA1) ion channels as well. In this study, we investigated the effect of selected derivatives (832-a pan-PDE inhibitor, 869-a TRPA1 modulator, and 145-a pan-PDE inhibitor and a weak TRPA1 modulator) on cellular responses related to airway remodeling using MRC-5 human lung fibroblasts. Compound 145 exerted the most considerable effect in limiting fibroblast to myofibroblasts transition (FMT) as well as proliferation, migration, and contraction. The effect of this compound appeared to depend mainly on its strong PDE inhibitory properties, and not on its effects on TRPA1 modulation. The strong anti-remodeling effects of 145 required activation of the cAMP/protein kinase A (PKA)/cAMP response element-binding protein (CREB) pathway leading to inhibition of transforming growth factor type ß1 (TGF-ß1) and Smad-dependent signaling in MRC-5 cells. These data suggest that the TGF-ß pathway is a major target for PDE inhibitors leading to inhibitory effects on cell responses involved in airway remodeling. These potent, pan-PDE inhibitors from the group of 7,8-disubstituted purine-2,6-dione derivatives, thus represent promising anti-remodeling drug candidates for further research.
Assuntos
Proteínas Quinases Dependentes de AMP Cíclico/metabolismo , AMP Cíclico/metabolismo , Fibroblastos/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Inibidores de Fosfodiesterase/farmacologia , Fator de Crescimento Transformador beta1/metabolismo , 3',5'-AMP Cíclico Fosfodiesterases/metabolismo , Cálcio/metabolismo , Movimento Celular , Proliferação de Células , Sobrevivência Celular , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 7/metabolismo , Desenho de Fármacos , Avaliação Pré-Clínica de Medicamentos , Fibroblastos/metabolismo , Fibrose , Humanos , Pulmão/metabolismo , Miofibroblastos/metabolismo , Transdução de Sinais , Canal de Cátion TRPA1/metabolismoRESUMO
The incidence and prevalence of ulcerative colitis are increasing globally. Although the exact cause and pathogenesis of this disease is unclear, research has led to a better understanding of the condition and to identification of new targets for therapy, which in turn has encouraged the development of new therapies. As well as biologic therapies, which have changed the way inflammatory bowel disease is managed, small molecules have been developed for the treatment of ulcerative colitis. These small molecule treatments are orally administered and are likely to bring a substantial shift in the way this chronic disease is treated. Oral therapies offer many advantages over infusion therapies, such as ease of use, increased acceptability by patients, and reduction of cost. This Review focuses not only on oral therapies that have been approved for use in ulcerative colitis, but also on those that are in development, providing a comprehensive overview for clinicians of available oral therapies and drugs that are likely to become available. We have also reviewed drugs that have shown promise in preclinical studies and could be effective future therapies.
Assuntos
Colite Ulcerativa/tratamento farmacológico , Doenças Inflamatórias Intestinais/tratamento farmacológico , Terapia de Alvo Molecular/métodos , Administração Oral , Idoso , Doença Crônica/tratamento farmacológico , Ensaios Clínicos como Assunto , Colite Ulcerativa/epidemiologia , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/efeitos dos fármacos , Avaliação Pré-Clínica de Medicamentos/métodos , Humanos , Imunomodulação , Incidência , Inibidores de Janus Quinases/farmacologia , Janus Quinases/antagonistas & inibidores , Janus Quinases/efeitos dos fármacos , Pessoa de Meia-Idade , Inibidores da Fosfodiesterase 4/farmacologia , Prevalência , Receptores de Esfingosina-1-Fosfato/agonistas , Receptores de Esfingosina-1-Fosfato/efeitos dos fármacosRESUMO
The aim of the present study was to evaluate the possible gut inhibitory role of the phosphodiesterase (PDE) inhibitor roflumilast. Increasing doses of roflumilast were tested against castor oil-induced diarrhea in mice, whereas the pharmacodynamics of the same effect was determined in isolated rabbit jejunum tissues. For in silico analysis, the identified PDE protein was docked with roflumilast and papaverine using the Autodock vina program from the PyRx virtual screening tool. Roflumilast protected against diarrhea significantly at 0.5 and 1.5 mg/kg doses, with 40% and 80% protection. Ex vivo findings from jejunum tissues show that roflumilast possesses an antispasmodic effect by inhibiting spontaneous contractions in a concentration-dependent manner. Roflumilast reversed carbachol (CCh, 1 µM)-mediated and potassium (K+, 80 mM)-mediated contractile responses with comparable efficacies but different potencies. The observed potency against K+ was significantly higher in comparison to CCh, similar to verapamil. Experiments were extended to further confirm the inhibitory effect on Ca++ channels. Interestingly, roflumilast deflected Ca++ concentration-response curves (CRCs) to the right with suppression of the maximum peak at both tested doses (0.001-0.003 mg/mL), similar to verapamil. The PDE-inhibitory effect was authenticated when pre-incubation of jejunum tissues with roflumilast (0.03-0.1 mg/mL) produced a leftward deflection of isoprenaline-mediated inhibitory CRCs and increased the tissue level of cAMP, similar to papaverine. This idea was further strengthened by molecular docking studies, where roflumilast exhibited a better binding affinity (-9.4 kcal/mol) with the PDE protein than the standard papaverine (-8.3 kcal/mol). In conclusion, inhibition of Ca++ channels and the PDE-4 enzyme explains the pharmacodynamics of the gut inhibitory effect of roflumilast.
Assuntos
Aminopiridinas/farmacologia , Antidiarreicos/farmacologia , Benzamidas/farmacologia , Bloqueadores dos Canais de Cálcio/farmacologia , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Diarreia/prevenção & controle , Parassimpatolíticos/farmacologia , Inibidores da Fosfodiesterase 4/farmacologia , Aminopiridinas/química , Aminopiridinas/farmacocinética , Animais , Antidiarreicos/química , Antidiarreicos/farmacocinética , Benzamidas/química , Benzamidas/farmacocinética , Sítios de Ligação , Bloqueadores dos Canais de Cálcio/química , Bloqueadores dos Canais de Cálcio/farmacocinética , Carbacol/farmacologia , Óleo de Rícino/administração & dosagem , AMP Cíclico/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/química , Ciclopropanos/química , Ciclopropanos/farmacocinética , Ciclopropanos/farmacologia , Diarreia/induzido quimicamente , Diarreia/metabolismo , Diarreia/fisiopatologia , Isoproterenol/farmacologia , Jejuno/efeitos dos fármacos , Jejuno/metabolismo , Camundongos , Simulação de Acoplamento Molecular , Papaverina/farmacologia , Parassimpatolíticos/química , Parassimpatolíticos/farmacocinética , Inibidores da Fosfodiesterase 4/química , Inibidores da Fosfodiesterase 4/farmacocinética , Ligação Proteica , Domínios e Motivos de Interação entre Proteínas , Estrutura Secundária de Proteína , Coelhos , Verapamil/farmacologiaRESUMO
BACKGROUND & AIMS: New oral therapeutic agents are needed for patients with ulcerative colitis (UC) who are unresponsive or intolerant to conventional therapy. METHODS: We performed a double-blind, phase 2 trial of adults with active UC for 3 months or more who were naïve to biologic therapy or had been failed by, could not tolerate, or had contraindications to conventional therapies. The study was performed at 61 sites in 14 countries (screening from January 2015 through May 2017). Patients were randomly assigned to groups given apremilast 30 mg (n = 57), apremilast 40 mg (n = 55), or placebo (n = 58) twice daily for 12 weeks; patients were then randomly assigned to groups that received apremilast, 30 or 40 mg twice daily, for an additional 40 weeks. Endoscopies were performed and biopsies were collected during the screening phase, at week 12, and at week 52. Blood and fecal samples were also collected and analyzed throughout the study. The primary endpoint was clinical remission at week 12, defined as a total Mayo score of 2 or less, with no individual subscore above 1. RESULTS: Clinical remission was achieved at week 12 by 31.6% of patients in the 30 mg apremilast group and 12.1% of patients in the placebo group (P = .01). However, only 21.8% of patients in the 40 mg apremilast group achieved clinical remission at week 12 (P = .27 compared with placebo). Differences in clinical remission between the 30 mg and 40 mg apremilast groups were associated with differences in endoscopic improvement. Both apremilast groups had similar improvements from baseline in Mayo score components (stool frequency score, rectal bleeding score, physician's global assessment). The 30 mg and 40 mg apremilast groups had greater median percent reductions in C-reactive protein (measured by a high-sensitivity blood test) and fecal calprotectin through week 12 than the placebo group. At week 52, clinical remission was achieved by 40.4% of patients initially assigned to the apremilast 30 mg group and 32.7% of patients initially assigned to the apremilast 40 mg group. The most frequent apremilast-associated adverse events were headache and nausea. CONCLUSIONS: Although the primary endpoint of clinical remission was not met in this phase 2 trial, a greater proportion of patients with active UC who received apremilast (30 mg or 40 mg) had improvements in clinical and endoscopic features, and markers of inflammation, at 12 weeks. Clinical remission was maintained to week 52 in up to 40% of patients who continued apremilast until that time point. ClinicalTrials.gov no: NCT02289417.
Assuntos
Colite Ulcerativa , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4 , Adulto , Terapia Biológica , Colite Ulcerativa/tratamento farmacológico , Método Duplo-Cego , Humanos , Indução de Remissão , Talidomida/efeitos adversos , Talidomida/análogos & derivados , Resultado do TratamentoRESUMO
Despite the recent advances in the life sciences and the remarkable investment in drug discovery research, the success rate of small-molecule drug development remains low. Safety is the second most influential factor of drug attrition in clinical studies; thus, the selection of compounds with fewer toxicity concerns is crucial to increase the success rate of drug discovery. Compounds that promiscuously bind to multiple targets are likely to cause unexpected pharmacological activity that may lead to adverse effects. Therefore, avoiding such compounds during early research stages would contribute to identifying compounds with a higher chance of success in the clinic. To evaluate the interaction profile against a wide variety of targets, we constructed a small-scale promiscuity panel (PP) consisting of eight targets (ROCK1, PDE4D2, GR, PPARγ, 5-HT2B, adenosine A3, M1, and GABAA) that were selected from diverse gene families. The validity of this panel was confirmed by comparison with the promiscuity index evaluated from larger-scale panels. Analysis of data from the PP revealed that both lipophilicity and basicity are likely to increase promiscuity, while the molecular weight does not significantly contribute. Additionally, the promiscuity assessed using our PP correlated with the occurrence of both in vitro cytotoxicity and in vivo toxicity, suggesting that the PP is useful to identify compounds with fewer toxicity concerns. In summary, this small-scale and cost-effective PP can contribute to the identification of safer compounds that would lead to a reduction in drug attrition due to safety issues.
Assuntos
Avaliação Pré-Clínica de Medicamentos/métodos , Animais , Sobrevivência Celular , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/genética , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/prevenção & controle , Células Hep G2 , Humanos , Camundongos , PPAR gama/genética , Ratos , Receptor A3 de Adenosina/genética , Receptor Muscarínico M1/genética , Receptor 5-HT2B de Serotonina/genética , Receptores de GABA-A/genética , Receptores de Glucocorticoides/genética , Quinases Associadas a rho/genéticaRESUMO
Obesity is associated with an increased risk of depression. The aim of the present study was to investigate whether obesity is a causative factor for the development of depression and what is the molecular pathway(s) that link these two disorders. Using lipidomic and transcriptomic methods, we identified a mechanism that links exposure to a high-fat diet (HFD) in mice with alterations in hypothalamic function that lead to depression. Consumption of an HFD selectively induced accumulation of palmitic acid in the hypothalamus, suppressed the 3', 5'-cyclic AMP (cAMP)/protein kinase A (PKA) signaling pathway, and increased the concentration of free fatty acid receptor 1 (FFAR1). Deficiency of phosphodiesterase 4A (PDE4A), an enzyme that degrades cAMP and modulates stimulatory regulative G protein (Gs)-coupled G protein-coupled receptor signaling, protected animals either from genetic- or dietary-induced depression phenotype. These findings suggest that dietary intake of saturated fats disrupts hypothalamic functions by suppressing cAMP/PKA signaling through activation of PDE4A. FFAR1 inhibition and/or an increase of cAMP signaling in the hypothalamus could offer potential therapeutic targets to counteract the effects of dietary or genetically induced obesity on depression.
Assuntos
AMP Cíclico/metabolismo , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Depressão/fisiopatologia , Dieta Hiperlipídica/efeitos adversos , Hipotálamo/fisiopatologia , Obesidade/fisiopatologia , Animais , Comportamento Animal , Depressão/etiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Obesidade/etiologia , Transdução de SinaisRESUMO
Psoriasis is a common, chronic inflammatory disease characterized by abnormal skin plaques, and the effectiveness of phosphodiesterase 4 (PDE4) inhibitor to lessen the symptoms of psoriasis has been proved. Aiming to find a novel PDE4 inhibitor acting as an effective, safe, and convenient therapeutic agent, we constructed a library consisting of berberine analogues, and compound 2 with a tetrahydroisoquinoline scaffold was identified as a novel and potent hit. The structure-aided and cell-based structure-activity relationship studies on a series of tetrahydro-isoquinolines lead to efficient discovery of a qualified lead compound (16) with the high potency and selectivity, well-characterized binding mechanism, high cell permeability, good safety and pharmacokinetic profile, and impressive in vivo efficacy on antipsoriasis, in particular with a topical application. Thus, our study presents a prime example for efficient discovery of novel, potent lead compounds derived from natural products using a combination of medicinal chemistry, biochemical, biophysical, and pharmacological approaches.
Assuntos
Desenho de Fármacos , Inibidores da Fosfodiesterase 4/química , Inibidores da Fosfodiesterase 4/farmacologia , Psoríase/tratamento farmacológico , Tetra-Hidroisoquinolinas/química , Tetra-Hidroisoquinolinas/farmacologia , Animais , Células CACO-2 , Domínio Catalítico , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/química , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/metabolismo , Avaliação Pré-Clínica de Medicamentos , Feminino , Células HEK293 , Humanos , Masculino , Camundongos , Modelos Moleculares , Inibidores da Fosfodiesterase 4/farmacocinética , Inibidores da Fosfodiesterase 4/uso terapêutico , Ratos , Estereoisomerismo , Relação Estrutura-Atividade , Tetra-Hidroisoquinolinas/farmacocinética , Tetra-Hidroisoquinolinas/uso terapêutico , Distribuição TecidualRESUMO
BACKGROUND: Newer classes of targeted drugs for moderate to severe plaque psoriasis are more effective and more expensive than older classes, posing a difficult and potentially costly decision about whether to use them as initial targeted treatments. OBJECTIVE: To estimate the clinical and economic outcomes of initial targeted treatment for the following drugs: adalimumab, etanercept, and infliximab (TNFα inhibitors); apremilast (PDE4 inhibitor); ustekinumab (IL-12/23 inhibitor); and ixekizumab, secukinumab, and brodalumab (IL-17 inhibitors). METHODS: We developed a Markov model to simulate patient outcomes as measured by quality-adjusted life-years (QALYs) and health care costs over a 10-year period. We assumed that patients who fail initial targeted treatment either proceed to subsequent therapy or discontinue targeted treatment. Effectiveness estimates for initial treatment were defined as improvement in Psoriasis Area and Severity Index (PASI) from baseline and derived from a 2018 network meta-analysis. Wholesale acquisition drug costs were discounted by a class-specific, empirically derived rebate percentage off of 2016 costs. We conducted one-way and probabilistic sensitivity analyses to assess uncertainty in results. RESULTS: The incremental benefits compared with no targeted treatment were, in descending order: ixekizumab 1.68 QALYs (95% credible range [CR] = 1.11-2.02), brodalumab 1.64 QALYs (95% CR = 1.08-1.98), secukinumab 1.51 QALYs (95% CR = 1.00-1.83), ustekinumab 1.43 QALYs (95% CR=0.94-1.74), infliximab 1.27 QALYs (95% CR = 0.89-1.55), adalimumab 1.15 QALYs (95% CR = 0.76-1.44), etanercept 0.97 QALYs (95% CR = 0.61-1.25), and apremilast 0.87 QALYs (95% CR = 0.52-1.17). Costs of care without targeted treatment totaled $66,451, and costs of targeted treatment ranged from $137,080 (apremilast) to $255,422 (ustekinumab). Probabilistic sensitivity analysis results indicated that infliximab and apremilast are likely to be the most cost-effective initial treatments at willingness-to-pay thresholds around $100,000 per QALY, while IL-17 drugs are more likely to be cost-effective at thresholds approaching $150,000 per QALY. Acquisition cost of the initial targeted drug and utility of clinical response were the most influential parameters. CONCLUSIONS: Our findings suggest that initial targeted treatment with IL-17 inhibitors is the most effective treatment strategy for plaque psoriasis patients who have failed methotrexate and phototherapy. Apremilast, brodalumab, infliximab, ixekizumab, and secukinumab are cost-effective at different willingness-to-pay thresholds. Additional research is needed on whether the effectiveness of targeted agents changes when used after previously targeted agents. DISCLOSURES: Funding for this study was contributed by the Institute for Clinical and Economic Review (ICER). Ollendorf, Chapman, Pearson, and Kumar are current employees, and Loos and Liu are former employees, of ICER, an independent organization that evaluates the evidence on the value of health care interventions, which is funded by grants from the Laura and John Arnold Foundation, Blue Shield of California Foundation, and the California HealthCare Foundation. ICER's annual policy summit is supported by dues from Aetna, AHIP, Anthem, Alnylam, AstraZeneca, Blue Shield of California, Cambia Health Solutions and MedSavvy, CVS Caremark, Editas, Express Scripts, Genentech, GlaxoSmithKline, Harvard Pilgrim Health Care, Health Care Service Corporation, OmedaRx, United Healthcare, Johnson & Johnson, Kaiser Permanente, Premera Blue Cross, Merck, National Pharmaceutical Council, Takeda, Pfizer, Novartis, Lilly, Humana, Prime Therapeutics, Sanofi, and Spark Therapeutics. Linder owns stock in Amgen, Biogen, and Eli Lilly; has contingent value rights in Sanofi Genzyme (related to alemtuzumab for multiple sclerosis); has received grant support from Astellas Pharma not related to this study and Clintrex, which was supported by AstraZeneca on an unrelated topic; and has received an honorarium from the Society of Healthcare Epidemiology of America (SHEA) as part of the SHEA Antimicrobial Stewardship Research Workshop Planning Committee, an educational activity supported by Merck. No other authors have potential conflicts of interest.
Assuntos
Análise Custo-Benefício , Fármacos Dermatológicos/uso terapêutico , Custos de Medicamentos , Psoríase/tratamento farmacológico , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/imunologia , Fármacos Dermatológicos/economia , Fármacos Dermatológicos/farmacologia , Humanos , Interleucina-12/antagonistas & inibidores , Interleucina-12/imunologia , Interleucina-17/antagonistas & inibidores , Interleucina-17/imunologia , Interleucina-23/antagonistas & inibidores , Interleucina-23/imunologia , Cadeias de Markov , Pessoa de Meia-Idade , Modelos Econômicos , Terapia de Alvo Molecular/economia , Terapia de Alvo Molecular/métodos , Inibidores da Fosfodiesterase 4/economia , Inibidores da Fosfodiesterase 4/uso terapêutico , Psoríase/economia , Psoríase/imunologia , Anos de Vida Ajustados por Qualidade de Vida , Resultado do Tratamento , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/imunologiaRESUMO
Recently, antiobesity studies using the method of inhibiting enzymatic activity of obesity-related enzymes as targets have received considerable attention. The aims of the current study were to investigate whether p-hydroxybenzyl alcohol (HBA), identified from Cudrania tricuspidata fruits with antiobesity effects, inhibits the activity of digestive and obesity-related enzymes and acts as an inhibitor against four target enzymes in kinetic studies. In vitro enzyme assays showed HBA at the highest concentration significantly reduced the enzymatic activity of four targets: pancreatic lipase (IC50 = 2.34-3.70 µM), α-glycosidase (IC50 = 9.08 µM), phosphodiesterase IV (IC50 = 4.99 µM), and citrate synthase (IC50 = 2.07 µM) enzymes. Based on the results of kinetic assays, the types of inhibition were investigated. Our findings indicate that HBA could have antiobesity efficacy, and it deserves further study.
Assuntos
Fármacos Antiobesidade/farmacologia , Álcoois Benzílicos/farmacologia , Citrato (si)-Sintase/antagonistas & inibidores , Nucleotídeo Cíclico Fosfodiesterase do Tipo 4/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Lipase/antagonistas & inibidores , Obesidade/enzimologia , alfa-Glucosidases/efeitos dos fármacos , Animais , Humanos , Concentração Inibidora 50 , Cinética , Moraceae/química , Extratos Vegetais/farmacologia , SuínosRESUMO
Our previous study revealed that the ethanolic extract of Justicia procumbens ameliorates ovalbumin-induced airway inflammation and airway hyper-responsiveness in a mouse model of asthma. However, the mechanism of action of the extract remains unknown. In this study, we prepared DW2008S, an optimized and standardized powder extracted from J. procumbens using anhydrous ethanol, and investigated its anti-asthmatic effect and mechanism of action. Our results showed that DW2008S contains two major ingredients, justicidin A (JA) and justicidin B (JB), which selectively inhibit T helper 2 (Th2) cell responses in concanavalin A-activated spleen cells and polarized Th2 cells. Blockade of T cell immunoreceptor with immunoglobulin and immunoreceptor tyrosine-based inhibition motif domains (TIGIT) using a neutralizing antibody also selectively inhibited Th2 cell responses. Furthermore, DW2008S regulated TIGIT expression in the mice and cultured cells. Additionally, DW2008S and JA antagonized human adenosine receptor A3 (A3 AR), which mediates mast cell-dependent inflammation and bronchoconstriction. DW2008S and JB inhibited human phosphodiesterase 4 (PDE4), which is known to cause bronchoconstriction; however, the required concentrations were higher than those needed to affect TIGIT . These findings suggest that DW2008S can potentially ameliorate Th2-driven airway inflammation and bronchoconstriction through negative regulation of TIGIT and blockade of A3 AR and PDE4 activities.