RESUMO
PURPOSE: Ticagrelor and dapagliflozin can suppress the activation of the NOD-like receptor 3 (NLRP3)-inflammasome and activate AMP-activated protein kinase (AMPK). The anti-inflammatory effects of dapagliflozin has been shown to depend on AMPK activation. Dapagliflozin and ticagrelor have been shown to have additive effects on the progression of diabetic cardiomyopathy in BTBR ob/ob mice with type-2 diabetes. We assessed whether dapagliflozin and ticagrelor have additive effects on the activation of the NLRP3-inflammasome and the progression of diabetic nephropathy in mice with type-2 diabetes. METHODS: Eight-week-old BTBR received either no-drug, dapagliflozin (1.5 mg/kg/d), ticagrelor (100 mg/kg/d), or their combination for 12 weeks. Blood was assessed weekly for glucose and urine for glucose and albumin. After 12 weeks, blood creatinine, cystatin C, inflammasome activation, and insulin were assessed by ELISA. Renal cortex samples were assessed by hematoxylin and eosin and periodic acid-Schiff staining. RT-PCR and immunoblotting were used to evaluate fibrosis and the activation of Akt, AMPK and the inflammasome. RESULTS: Both ticagrelor and dapagliflozin reduced serum creatinine and cystatin C levels and urinary albumin. Both drugs attenuated the increase in glomerular area and mesangial matrix index. Both drugs decreased collagen-1 and collagen-3 expression and the activation of the NLRP3-inflammasome. Both drugs increased P-AMPK levels, but only dapagliflozin increased P-Akt levels. Overall, the protective effects of dapagliflozin and ticagrelor were additive. CONCLUSIONS: Dapagliflozin and ticagrelor attenuated the progression of diabetic nephropathy in BTBR ob/ob mice with additive effects of the combination. This was associated with AMPK activation and reduced activation of the NLRP3 inflammasome, whereas only dapagliflozin increased Akt activation.
Assuntos
Diabetes Mellitus Tipo 2 , Nefropatias Diabéticas , Insulinas , Proteínas Quinases Ativadas por AMP/metabolismo , Albuminas/metabolismo , Albuminas/farmacologia , Albuminas/uso terapêutico , Animais , Anti-Inflamatórios/farmacologia , Compostos Benzidrílicos , Creatinina/metabolismo , Creatinina/farmacologia , Creatinina/uso terapêutico , Cistatina C/metabolismo , Cistatina C/farmacologia , Cistatina C/uso terapêutico , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/tratamento farmacológico , Diabetes Mellitus Tipo 2/metabolismo , Nefropatias Diabéticas/tratamento farmacológico , Nefropatias Diabéticas/prevenção & controle , Amarelo de Eosina-(YS)/metabolismo , Amarelo de Eosina-(YS)/farmacologia , Amarelo de Eosina-(YS)/uso terapêutico , Glucose/metabolismo , Glucosídeos , Hematoxilina/metabolismo , Hematoxilina/farmacologia , Hematoxilina/uso terapêutico , Inflamassomos/metabolismo , Insulinas/metabolismo , Insulinas/farmacologia , Insulinas/uso terapêutico , Rim , Camundongos , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Ácido Periódico/metabolismo , Ácido Periódico/farmacologia , Ácido Periódico/uso terapêutico , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ticagrelor/farmacologia , Ticagrelor/uso terapêuticoRESUMO
PURPOSE: Ticagrelor, a P2Y12 receptor antagonist, and dapagliflozin, a sodium-glucose-cotransporter-2 inhibitor, suppress the activation of the NLRP3 inflammasome. The anti-inflammatory effects of dapagliflozin depend on AMPK activation. Also, ticagrelor can activate AMPK. We assessed whether dapagliflozin and ticagrelor have additive effects in attenuating the progression of diabetic cardiomyopathy in T2DM mice. METHODS: Eight-week-old BTBR and wild-type mice received no drug, dapagliflozin (1.5 mg/kg/day), ticagrelor (100 mg/kg/day), or their combination for 12 weeks. Heart function was evaluated by echocardiography and heart tissue samples were assessed for fibrosis, apoptosis, qRT-PCR, and immunoblotting. RESULTS: Both drugs attenuated the progression of diabetic cardiomyopathy as evident by improvements in left ventricular end-systolic and end-diastolic volumes and left ventricular ejection fraction, which were further improved by the combination. Both drugs attenuated the activation of the NOD-like receptor 3 (NLRP3) inflammasome and fibrosis. The effect of the combination was significantly greater than each drug alone on myocardial tissue necrotic factorα (TNFα) and interleukin-6 (IL-6) levels, suggesting additive effects. The combination had also a greater effect on ASC, collagen-1, and collagen-3 mRNA levels than each drug alone. While both drugs activated adenosine mono-phosphate kinase (AMPK), only dapagliflozin activated mTOR and increased RICTOR levels. Moreover, only dapagliflozin decreased myocardial BNP and Caspase-1 mRNA levels, and the effects of dapagliflozin on NLRP3 and collagen-3 mRNA levels were significantly greater than those of ticagrelor. CONCLUSIONS: Both dapagliflozin and ticagrelor attenuated the progression of diabetic cardiomyopathy, the activation of the NLRP3 inflammasome, and fibrosis in BTBR mice with additive effects of the combination. While both dapagliflozin and ticagrelor activated AMPK, only dapagliflozin activated mTOR complex 2 (mTORC2) in hearts of BTBR mice.
Assuntos
Proteínas Quinases Ativadas por AMP/metabolismo , Compostos Benzidrílicos/farmacologia , Diabetes Mellitus Tipo 2/tratamento farmacológico , Cardiomiopatias Diabéticas/prevenção & controle , Glucosídeos/farmacologia , Inflamassomos/metabolismo , Miócitos Cardíacos/efeitos dos fármacos , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Antagonistas do Receptor Purinérgico P2Y/farmacologia , Inibidores do Transportador 2 de Sódio-Glicose/farmacologia , Serina-Treonina Quinases TOR/metabolismo , Ticagrelor/farmacologia , Animais , Diabetes Mellitus Tipo 2/complicações , Diabetes Mellitus Tipo 2/enzimologia , Cardiomiopatias Diabéticas/enzimologia , Cardiomiopatias Diabéticas/etiologia , Cardiomiopatias Diabéticas/fisiopatologia , Modelos Animais de Doenças , Progressão da Doença , Ativação Enzimática , Fibrose , Masculino , Alvo Mecanístico do Complexo 2 de Rapamicina/metabolismo , Camundongos Endogâmicos C57BL , Miócitos Cardíacos/enzimologia , Transdução de Sinais , Volume Sistólico/efeitos dos fármacos , Função Ventricular Esquerda/efeitos dos fármacos , Remodelação Ventricular/efeitos dos fármacosRESUMO
BACKGROUND/AIMS: We assessed the effects of ticagrelor, aspirin and prasugrel, started 7days after myocardial ischemia-reperfusion injury on remodeling, inflammation and fibrosis in the rat. We examined whether ticagrelor can affect the number of progenitor cells in the border zone. Ticagrelor, started 24h after myocardial ischemia-reperfusion injury, attenuates the decrease in heart function and adverse remodeling, an effect which is blocked by aspirin. METHODS: Rats underwent 40min ischemia followed by reperfusion. Oral dosing with vehicle, ticagrelor (300mg/kg/d), aspirin (20mg/kg/d), their combination or prasugrel (15mg/kg/d) started 7days after infarction. Echocardiography was used to assess systolic function. Heart tissue were analyzed by rt-PCR, immunoblotting, ELISA and immunohistochemistry 2weeks after infarction. RESULTS: Both ticagrelor and aspirin attenuated the decrease in systolic function and remodeling, an effect that was blocked by their combination. Ticagrelor and aspirin attenuated the increase in ANP, BNP, collagen-I and collagen-III. Again, the effect was blocked by their combination. Ticagrelor increased c-Kit, Sca-1, Ki-67, CD34, attenuated the decrease in CD105 mRNA levels, and attenuated the increase in CD31, whereas aspirin increased Ki-67, suppressed the increase in CD31 and attenuated the decrease in CD105 mRNA levels. Prasugrel did not display any effects. CONCLUSION: Ticagrelor attenuated adverse remodeling and deterioration of left ventricular systolic function despite starting treatment after the myocardial ischemia-reperfusion injury is completed. Aspirin had similar effects; however, when combined with ticagrelor, the protective effects were significantly attenuated. Ticagrelor increased the levels of several markers of stem cells and regeneration, suggesting cardiac healing by recruiting regenerative cells into the infarct.
Assuntos
Apoptose/efeitos dos fármacos , Infarto do Miocárdio/patologia , Inibidores da Agregação Plaquetária/farmacologia , Ticagrelor/farmacologia , Remodelação Ventricular/efeitos dos fármacos , Animais , Aspirina/farmacologia , Aspirina/uso terapêutico , Fator Natriurético Atrial/genética , Fator Natriurético Atrial/metabolismo , Modelos Animais de Doenças , Quimioterapia Combinada , Endoglina/genética , Endoglina/metabolismo , Fibrose , Regulação da Expressão Gênica/efeitos dos fármacos , Infarto do Miocárdio/tratamento farmacológico , Traumatismo por Reperfusão Miocárdica/metabolismo , Traumatismo por Reperfusão Miocárdica/patologia , Inibidores da Agregação Plaquetária/uso terapêutico , Cloridrato de Prasugrel/farmacologia , Cloridrato de Prasugrel/uso terapêutico , Proteínas Proto-Oncogênicas c-kit/genética , Proteínas Proto-Oncogênicas c-kit/metabolismo , Ratos , Ratos Sprague-Dawley , Células-Tronco/citologia , Células-Tronco/efeitos dos fármacos , Células-Tronco/metabolismo , Ticagrelor/uso terapêutico , Função Ventricular Esquerda/efeitos dos fármacosRESUMO
Accurate determination of in vivo circulating concentrations of extracellular adenosine in blood samples is challenging due to the rapid formation and rapid clearance of adenosine in blood. A blood collection protocol was developed based on direct sampling of venous blood into, and instant mixing with, a STOP solution developed to conserve in vivo adenosine concentrations by completely preventing both its formation and clearance in collected blood. Stable isotope labeled AMP and adenosine spiked into blood ex vivo were used in combination with mass spectrometry to evaluate conservation of adenosine and prevention of its formation. A number of approved drugs, including the P2Y12 antagonist ticagrelor, have been described to increase extracellular adenosine. This may contribute to its clinical profile, highlighting the importance of accurate measurement of in vivo adenosine concentrations.A high sensitive ultra performance liquid chromatography-tandem- mass spectrometry (UPLC-tandem-MS) analytical method for plasma adenosine was developed and validated with a lower limit of quantification of 2 nmol/L. The method demonstrated plasma adenosine stability during sample processing and analytical method performance relevant to human blood samples. The final STOP solution proved able to conserve exogenous adenosine and to prevent adenosine formation from exogenous AMP added in vitro to human blood over 15 minutes. The mean endogenous adenosine concentration in plasma prepared from venous blood collected from 10 healthy volunteers was 13 ± 7 nmol/L. Finally, the method was used to demonstrate the previously described concentration-dependent ability of ticagrelor to conserve extracellular adenosine at clinically relevant exposures. In conclusion, we report an optimized sampling protocol and a validated analytical method for accurate measurement of in vivo circulating adenosine concentrations in human blood, suitable for use in clinical trials.
Assuntos
Adenosina/sangue , Coleta de Amostras Sanguíneas/métodos , Antagonistas do Receptor Purinérgico P2Y/farmacologia , Ticagrelor/farmacologia , Adenosina/metabolismo , Cromatografia Líquida de Alta Pressão/métodos , Feminino , Voluntários Saudáveis , Humanos , Limite de Detecção , Masculino , Reprodutibilidade dos Testes , Espectrometria de Massas em Tandem/métodosRESUMO
Ticagrelor, a P2Y12 antagonist, is approved for prevention of thromboembolic events. MEDI2452 is a potential reversal agent for ticagrelor and ticagrelor active metabolite (TAM). The total plasma exposure of ticagrelor and TAM in patients are roughly 0.5-1 and 0.2-0.5 µmol/L, respectively. Both have similar high potency vs. P2Y12 (Ki 2 nmol/L) but are plasma protein-bound to 99.8% and only the 0.2% free fraction is able to inhibit the P2Y12 receptor. Thus, for unbound concentration measurements to be a proof of mechanism biomarker for MEDI2452 a very high sensitivity is required. Using established techniques as equilibrium dialysis and LC-MS/MS, made it possible to evaluate the efficacy of the reversal agent by measuring reduction of unbound concentration of ticagrelor in the presence of MEDI2452. With challenges such as ultra-low concentrations, small sample volumes, recovery issues and adsorption to plastic we managed to develop a highly sensitive assay for determining unbound concentration levels of ticagrelor and TAM in plasma with a quantification limit of 30 pmol/L and 45 pmol/L, respectively. With this method we were able to detect close to a 100-fold MEDI2452 mediated reduction in the unbound concentration of both ticagrelor and TAM. The assay provided proof of mechanism as MEDI2452 concentration- and dose-dependently eliminated unbound concentration of ticagrelor and reversed its antiplatelet activity in preclinical models and will support future development of MEDI2452.
Assuntos
Anticorpos Neutralizantes/sangue , Antídotos , Análise Química do Sangue , Inibidores da Agregação Plaquetária/sangue , Antagonistas do Receptor Purinérgico P2Y/sangue , Ticagrelor/sangue , Animais , Anticorpos Neutralizantes/farmacologia , Antídotos/farmacologia , Coleta de Amostras Sanguíneas , Anticorpos Amplamente Neutralizantes , Cromatografia Líquida , Diálise , Relação Dose-Resposta a Droga , Humanos , Espectrometria de Massas , Camundongos , Agregação Plaquetária/efeitos dos fármacos , Inibidores da Agregação Plaquetária/farmacologia , Antagonistas do Receptor Purinérgico P2Y/farmacologia , Sus scrofa , Ticagrelor/farmacologiaRESUMO
PURPOSE: We explored the effects of rosuvastatin, aspirin, ticagrelor, and clopidogrel, alone or in combinations on the progression of atherosclerosis and inflammasome activation in diabetic mice. Statins and ticagrelor increase the production of 15-epi-lipoxin A4 via cyclooxygenase-2. Aspirin alone increases 15-epi-lipoxin A4, but when combined with statins, cyclooxygenase-2 is completely blocked. METHODS: ApoE-/-/db+/db+ double-knockout mice received rosuvastatin (5 mg/kg/day), aspirin (25 mg/kg/day), ticagrelor (300 mg/kg/day), clopidogrel (75 mg/kg/day), or their combination for 14 weeks. Serum 15-epi-lipoxin A4 levels and aortic wall cholesterol content, IL-1ß, IL-6, and TNF-α levels, and plaque area were assessed. RESULTS: Aspirin, ticagrelor, and rosuvastatin increased 15-epi-lipoxin A4 levels. The combination of rosuvastatin + ticagrelor provided an additive effect. Aspirin attenuated the effect of both ticagrelor and rosuvastatin. Aspirin, ticagrelor, and rosuvastatin reduced the area of the atherosclerotic plaque. The combination of ticagrelor + rosuvastatin provided additive effects. There was a negative interaction when aspirin was combined with ticagrelor or rosuvastatin. Aspirin, ticagrelor, and rosuvastatin decreased serum IL-1ß and IL-6 levels. There was no interaction between aspirin and ticagrelor or aspirin and rosuvastatin, whereas combining rosuvastatin and ticagrelor provided an additive effect. Aspirin, ticagrelor, and rosuvastatin all decreased TNF-α levels. Aspirin attenuated the effect of both ticagrelor and rosuvastatin, and there was no additive effect of combining ticagrelor + rosuvastatin. CONCLUSIONS: We found an intricate interaction between aspirin, ticagrelor, and rosuvastatin, as aspirin reduced both ticagrelor and rosuvastatin ability to ameliorate inflammation and atherosclerosis. In contrast, we found additive effects when ticagrelor and rosuvastatin were combined.
Assuntos
Adenosina/análogos & derivados , Aspirina/uso terapêutico , Aterosclerose/prevenção & controle , Diabetes Mellitus Experimental/imunologia , Rosuvastatina Cálcica/uso terapêutico , Adenosina/administração & dosagem , Adenosina/uso terapêutico , Animais , Apolipoproteínas E/genética , Aspirina/administração & dosagem , Aterosclerose/imunologia , Diabetes Mellitus Experimental/tratamento farmacológico , Progressão da Doença , Interações Medicamentosas , Camundongos Knockout , Rosuvastatina Cálcica/administração & dosagem , TicagrelorRESUMO
BACKGROUND: Ticagrelor inhibits the equilibrative-nucleoside-transporter-1 and thereby, adenosine cell re-uptake. Ticagrelor limits infarct size (IS) in non-diabetic rats and the effect is adenosine-dependent. Statins, via ecto-5'-nucleotidase activation, also increase adenosine levels and limit IS. HYPOTHESIS: Ticagrelor and rosuvastatin have additive effects on myocardial adenosine levels, and therefore, on IS and post-reperfusion activation of the NLRP3-inflammasome. METHODS: Diabetic ZDF rats received via oral gavage; water (control), ticagrelor (150 mg/kg/d), prasugrel (7.5 mg/kg/d), rosuvastatin (5 mg/kg/d), ticagrelor + rosuvastatin and prasugrel + rosuvastatin for 3d. On day 4, rats underwent 30 min coronary artery occlusion and 24 h of reperfusion. Two additional groups received, ticagrelor + rosuvastatin or water in combination with CGS15943 (CGS, an adenosine receptor antagonist, 10 mg/kg i.p. 1 h before ischemia). RESULTS: Both ticagrelor and rosuvastatin increased myocardial adenosine levels with an additive effect of the combination whereas prasugrel had no effect. Similarly, both ticagrelor and rosuvastatin significantly reduced IS with an additive effect of the combination whereas prasugrel had no effect. The effect on IS was adenosine dependent as CGS15943 reversed the effect of ticagrelor + rosuvastatin. The ischemia-reperfusion injury increased myocardial mRNA levels of NLRP3, ASC, IL-1ß and IL-6. Ticagrelor and rosuvastatin, but not prasugrel, significantly decreased these pro-inflammatory mediators with a trend to an additive effect of the combination. The combination also increased the levels of anti-inflammatory 15-epilipoxin A4. CONCLUSIONS: Ticagrelor and rosuvastatin when given in combination have an additive effect on local myocardial adenosine levels in the setting of ischemia reperfusion. This translates into an additive cardioprotective effect mediated by adenosine-induced effects including downregulation of pro- but upregulation of anti-inflammatory mediators.
Assuntos
Adenosina/análogos & derivados , Cardiotônicos/uso terapêutico , Traumatismo por Reperfusão Miocárdica/tratamento farmacológico , Rosuvastatina Cálcica/uso terapêutico , Adenosina/sangue , Adenosina/farmacocinética , Adenosina/farmacologia , Adenosina/uso terapêutico , Animais , Proteínas Reguladoras de Apoptose/genética , Proteínas Adaptadoras de Sinalização CARD , Cardiotônicos/farmacologia , Ciclo-Oxigenase 2/metabolismo , Diabetes Mellitus Experimental/tratamento farmacológico , Diabetes Mellitus Experimental/metabolismo , Sinergismo Farmacológico , Inibidores de Hidroximetilglutaril-CoA Redutases/farmacologia , Inibidores de Hidroximetilglutaril-CoA Redutases/uso terapêutico , Interleucina-1beta/genética , Interleucina-6/genética , Lipoxinas/metabolismo , Masculino , Traumatismo por Reperfusão Miocárdica/metabolismo , Miocárdio/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Obesidade/tratamento farmacológico , Obesidade/metabolismo , Inibidores da Agregação Plaquetária/farmacologia , Inibidores da Agregação Plaquetária/uso terapêutico , Cloridrato de Prasugrel/farmacologia , Cloridrato de Prasugrel/uso terapêutico , Antagonistas do Receptor Purinérgico P2Y/farmacologia , Antagonistas do Receptor Purinérgico P2Y/uso terapêutico , RNA Mensageiro/metabolismo , Ratos Zucker , Rosuvastatina Cálcica/farmacologia , TicagrelorRESUMO
Ticagrelor is a potent antagonist of the P2Y12 receptor (P2Y12R) and consequently an inhibitor of platelet activity effective in the treatment of atherothrombosis. Here, we sought to further characterize its molecular mechanism of action. Initial studies showed that ticagrelor promoted a greater inhibition of adenosine 5'-diphosphate (ADP)-induced Ca2+ release in washed platelets vs other P2Y12R antagonists. This additional effect of ticagrelor beyond P2Y12R antagonism was in part as a consequence of ticagrelor inhibiting the equilibrative nucleoside transporter 1 (ENT1) on platelets, leading to accumulation of extracellular adenosine and activation of Gs-coupled adenosine A2A receptors. This contributed to an increase in basal cyclic adenosine monophosphate (cAMP) and vasodilator-stimulated phosphoprotein phosphorylation (VASP-P). In addition, ticagrelor increased platelet cAMP and VASP-P in the absence of ADP in an adenosine receptor-independent manner. We hypothesized that this increase originated from a direct effect on basal agonist-independent P2Y12R signaling, and this was validated in 1321N1 cells stably transfected with human P2Y12R. In these cells, ticagrelor blocked the constitutive agonist-independent activity of the P2Y12R, limiting basal Gi-coupled signaling and thereby increasing cAMP levels. These data suggest that ticagrelor has the pharmacological profile of an inverse agonist. Based on our results showing insurmountable inhibition of ADP-induced Ca2+ release and forskolin-induced cAMP, the mode of antagonism of ticagrelor also appears noncompetitive, at least functionally. In summary, our studies describe 2 novel modes of action of ticagrelor, inhibition of platelet ENT1 and inverse agonism at the P2Y12R that contribute to its effective inhibition of platelet activation.
Assuntos
Adenosina/análogos & derivados , Plaquetas/metabolismo , Transportador Equilibrativo 1 de Nucleosídeo/antagonistas & inibidores , Ativação Plaquetária/efeitos dos fármacos , Receptores Purinérgicos P2Y12/metabolismo , Adenosina/farmacologia , Difosfato de Adenosina/farmacologia , Plaquetas/citologia , Cálcio/metabolismo , Sinalização do Cálcio/efeitos dos fármacos , Moléculas de Adesão Celular/metabolismo , Colforsina/farmacologia , AMP Cíclico/metabolismo , Transportador Equilibrativo 1 de Nucleosídeo/metabolismo , Feminino , Humanos , Masculino , Proteínas dos Microfilamentos/metabolismo , Fosfoproteínas/metabolismo , Fosforilação/efeitos dos fármacos , TicagrelorRESUMO
Ticagrelor is an antagonist of the platelet P2Y12 receptor for ADP, approved for the prevention of thromboembolic events in patients with acute coronary syndrome. Previous studies showed that ticagrelor has no significant activity versus P1 receptors for adenosine and other known P2Y receptors, with the exception of P2Y13, which was not tested. The P2Y12 antagonist cangrelor has been shown to also inhibit P2Y13 and to decrease the P2Y13-regulated capacity of megakaryocytes to produce pro-platelets. We tested whether or not ticagrelor inhibits P2Y13 signalling and function. The in vitro effects of ticagrelor, its active (TAM) and inactive (TIM) metabolites, cangrelor and the P2Y13 antagonist MRS2211 were tested in two experimental models: 1) a label-free cellular response assay in P2Y13-transfected HEK293 T-REx cells; and 2) pro-platelet formation by human megakaryocytes in culture. Ticagrelor, TAM, cangrelor and MRS2211, but not TIM, inhibited the cellular responses in P2Y13-transfected cells. In contrast, only MRS2211 and cangrelor, confirming previous results, inhibited pro-platelet formation by megakaryocytes in vitro. The platelet count of patients randomised to treatment with ticagrelor in the PLATO trial did not change during treatment and was comparable to those of patients randomised to clopidogrel. In conclusion, ticagrelor and TAM act as P2Y13 antagonists in a transfected cell system in vitro but this does not translate into any impact on pro-platelet formation in vitro or altered platelet count in patients.
Assuntos
Adenosina/análogos & derivados , Megacariócitos/citologia , Inibidores da Agregação Plaquetária/farmacologia , Receptores Purinérgicos P2/metabolismo , Síndrome Coronariana Aguda/tratamento farmacológico , Adenosina/farmacologia , Plaquetas , Células HEK293 , Humanos , Megacariócitos/efeitos dos fármacos , Antagonistas do Receptor Purinérgico P2Y/farmacologia , Ticagrelor , Ticlopidina/farmacologiaRESUMO
Antiplatelet therapy is given to patients with acute coronary syndrome to reduce the risk for thrombotic events, but may increase the risk for bleeding. Ticagrelor was administered intravenously to mice. Cumulative blood loss and bleeding time were measured after cutting 5âmm of the tail, 20âmin after the start of ticagrelor infusion. The tail was placed in a hemoglobin-sensitive device measuring light absorbance (abs) over time for 35âmin. Activated recombinant human factor VII (rhFVIIa; NovoSeven; NovoNordisk A/S, Bagsvaerd, Denmark) 1âmg/kg (study 1); recombinant human prothrombin (rhFII, MEDI8111) 10âmg/kg (study 2); or vehicle was given intravenously once bleeding had commenced, within 90s after tail cut. Ticagrelor resulted in more than 98% inhibition of ex-vivo ADP-induced platelet aggregation. In study 1, the median blood loss in the ticagrelor, vehicle, and rhFVIIa groups were 909, 122, and 397âabs*s, respectively (Pâ<â0.05 for both comparisons, including the ticagrelor group). Similar pattern was seen for bleeding time. The median bleeding time in the ticagrelor, vehicle, and rhFVIIa groups were 2003, 449, and 884s, respectively (Pâ<â0.05 for both comparisons, including the ticagrelor group). In study 2, the median blood loss and bleeding time in the ticagrelor group were 362âabs*s and 1847s. The corresponding numbers for the vehicle and rhFII groups were 71âabs*s and 613s, and 178âabs*s and 701s, respectively (Pâ<â0.05 for comparisons between ticagrelor and vehicle for both blood loss and bleeding time). In mice dosed to complete P2Y12 inhibition, boosting coagulation by administration of rhFVIIa or rhFII within 90s after bleeding initiation can partly reverse ticagrelor-enhanced bleeding.
Assuntos
Adenosina/análogos & derivados , Coagulação Sanguínea/efeitos dos fármacos , Hemostasia/genética , Antagonistas do Receptor Purinérgico P2Y/uso terapêutico , Adenosina/administração & dosagem , Adenosina/uso terapêutico , Animais , Camundongos , Camundongos Endogâmicos C57BL , Antagonistas do Receptor Purinérgico P2Y/administração & dosagem , TicagrelorRESUMO
The effect and clinical benefit of P2Y12 receptor antagonists may not be limited to platelet inhibition and the prevention of arterial thrombus formation. Potential additional effects include reduction of the pro-inflammatory role of activated platelets and effects related to P2Y12 receptor inhibition on other cells apart from platelets. P2Y12 receptor antagonists, thienopyridines and ticagrelor, differ in their mode of action being prodrugs instead of direct acting and irreversibly instead of reversibly binding to P2Y12 . These key differences may provide different potential when it comes to additional effects. In addition to P2Y12 receptor blockade, ticagrelor is unique in having the only well-documented additional target of inhibition, the equilibrative nucleoside transporter 1. The current review will address the effects of P2Y12 receptor antagonists beyond platelets and the protection against arterial thrombosis. The discussion will include the potential for thienopyridines and ticagrelor to mediate anti-inflammatory effects, to conserve vascular function, to affect atherosclerosis, to provide cardioprotection and to induce dyspnea.
Assuntos
Adenosina/análogos & derivados , Fenômenos Fisiológicos Cardiovasculares/efeitos dos fármacos , Inibidores da Agregação Plaquetária/farmacologia , Antagonistas do Receptor Purinérgico P2Y/farmacologia , Receptores Purinérgicos P2Y12/metabolismo , Tienopiridinas/farmacologia , Adenosina/farmacologia , Adenosina/uso terapêutico , Animais , Aterosclerose/fisiopatologia , Aterosclerose/prevenção & controle , Plaquetas/efeitos dos fármacos , Plaquetas/fisiologia , Cardiotônicos/farmacologia , Cardiotônicos/uso terapêutico , Transportador Equilibrativo 1 de Nucleosídeo/antagonistas & inibidores , Humanos , Inflamação/sangue , Inibidores da Agregação Plaquetária/uso terapêutico , Antagonistas do Receptor Purinérgico P2Y/uso terapêutico , Tienopiridinas/uso terapêutico , TicagrelorRESUMO
OBJECTIVE: The objective of this study was to assess the pharmacokinetic and pharmacodynamic behavior of ticagrelor administered either as crushed (in the semi-upright sitting position) or as integral (in the supine position) tablets in ST-segment elevation myocardial infarction (STEMI) patients undergoing primary percutaneous coronary intervention (PCI). METHODS: We randomized 20 patients to ticagrelor 180 mg either as 2 integral tablets administered in the supine position (standard administration) or as 2 tablets crushed and dispersed, administered in the semi-upright sitting position. Blood samples were drawn for pharmacokinetic and pharmacodynamic assessment at randomization (0 h) and at 0.5, 1, 2, and 4 h. RESULTS: At 1 h, ticagrelor plasma exposure and area under the plasma concentration-time curve from time zero to 1 h (AUC1) (co-primary endpoints) were higher in the crushed versus integral tablets group (median 586 vs. 70.1 ng/mL and 234 vs. 24.4 ng·h/mL, respectively), with a ratio of adjusted geometric means (95% confidence interval [CI]) of 12.67 (2.34-68.51) [p = 0.005] and 19.28 (3.51-106.06) [p = 0.002], respectively. Time to maximum plasma concentration was shorter in the crushed versus integral tablets group (median 2 vs. 4 h), with a ratio of adjusted geometric means (95% CI) of 0.69 (0.49-0.97) [p = 0.035]. Parallel findings were observed with AR-C124910XX (active metabolite). Platelet reactivity (VerifyNow(®)) at 1 h was lower with crushed versus standard administration with least squares estimates mean difference (95% CI) of 92 (-158.4 to 26.6) P2Y12 reaction units (p = 0.009). CONCLUSIONS: In patients with STEMI undergoing primary PCI, ticagrelor crushed tablets administered in the semi-upright sitting position seems to lead to a faster-compared with standard administration-absorption, with stronger antiplatelet activity within the first hour. TRIAL REGISTRATION: ClinicalTrials.gov identifier: NCT02046486.
Assuntos
Adenosina/análogos & derivados , Infarto do Miocárdio/metabolismo , Inibidores da Agregação Plaquetária/administração & dosagem , Postura , Antagonistas do Receptor Purinérgico P2Y/administração & dosagem , Adenosina/administração & dosagem , Adenosina/sangue , Adenosina/farmacocinética , Adenosina/farmacologia , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Infarto do Miocárdio/tratamento farmacológico , Infarto do Miocárdio/cirurgia , Intervenção Coronária Percutânea , Inibidores da Agregação Plaquetária/farmacocinética , Inibidores da Agregação Plaquetária/farmacologia , Antagonistas do Receptor Purinérgico P2Y/sangue , Antagonistas do Receptor Purinérgico P2Y/farmacocinética , Antagonistas do Receptor Purinérgico P2Y/farmacologia , Método Simples-Cego , Comprimidos , TicagrelorRESUMO
INTRODUCTION: Ticagrelor, a novel platelet inhibitor acting on the ADP-dependent P2Y12 receptor, is currently approved for treating adults with acute coronary syndrome. The effect of ticagrelor in children has not been explored. As a first step, we here evaluate if the in vitro anti-platelet potency of ticagrelor in blood samples from children of different age is different as compared with in blood samples from adults. MATERIALS AND METHODS: Blood samples from 36 healthy children grouped by age (0-2 months, n=6; 2-6 months, n=6; 6months-2years, n=6; 2-6 years, n=10; 6-12 years, n=8) and 13 adults were collected for in vitro analysis using vasodilator stimulated phosphoprotein phosphorylation (VASP) assay in whole blood and ADP-induced light transmission aggregometry (LTA) in platelet rich plasma. Ticagrelor (0.01 - 10µmol/L) was added in vitro and its potency was assessed by calculating the concentration that provided 50% inhibition of the maximum response (IC50). RESULTS: The in vitro potency of ticagrelor in blood from adults and in blood from children of any age group were comparable, both when analyzed with LTA and with VASP. CONCLUSIONS: These in vitro results are consistent with the hypothesis that ticagrelor would achieve a comparable anti-platelet effect in children of different ages as in adults at equal plasma exposure.
Assuntos
Adenosina/análogos & derivados , Envelhecimento/fisiologia , Plaquetas/efeitos dos fármacos , Plaquetas/fisiologia , Agregação Plaquetária/efeitos dos fármacos , Agregação Plaquetária/fisiologia , Adenosina/administração & dosagem , Adulto , Células Cultivadas , Criança , Pré-Escolar , Relação Dose-Resposta a Droga , Feminino , Humanos , Lactente , Recém-Nascido , Masculino , Inibidores da Agregação Plaquetária/administração & dosagem , Ticagrelor , Resultado do TratamentoRESUMO
OBJECTIVE: In addition to P2Y12 receptor antagonism, ticagrelor inhibits adenosine cell uptake. Prior data show that 7-day pretreatment with ticagrelor limits infarct size. We explored the acute effects of ticagrelor and clopidogrel on infarct size and potential long-term effects on heart function. APPROACH AND RESULTS: Rats underwent 30-minute ischemia per 24-hour reperfusion. (1) Ticagrelor (10 or 30 mg/kg) or clopidogrel (12.5 mg/kg) was given via intraperitoneal injection 5 minutes before reperfusion. (2) Rats received ticagrelor acute (intraperitoneal; 30 mg/kg), chronic (oral; 300 mg/kg per day) for 4 weeks starting 1 day after reperfusion or the combination (acute+chronic). Another group received clopidogrel (intraperitoneal [12.5 mg/kg]+oral [62.5 mg/kg per day]) for 4 weeks. (1) Ticagrelor dose-dependently reduced infarct size, 10 mg/kg (31.5%±1.8%; P<0.001) and 30 mg/kg (21.4%±2.6%; P<0.001) versus control (45.3±1.7%), whereas clopidogrel had no effect (42.4%±2.6%). Ticagrelor, but not clopidogrel, increased myocardial adenosine levels, increased phosphorylation of Akt, endothelial NO synthase, and extracellular-signal-regulated kinase 1/2 4 hours after reperfusion and decreased apoptosis. (2) After 4 weeks, left ventricular ejection fraction was reduced in the vehicle-treated group (44.8%±3.5%) versus sham (77.6%±0.9%). All ticagrelor treatments improved left ventricular ejection fraction, acute (69.5%±1.6%), chronic (69.2%±1.0%), and acute+chronic (76.3%±1.2%), whereas clopidogrel had no effect (37.4%±3.7%). Ticagrelor, but not clopidogrel, attenuated fibrosis and decreased collagen-III mRNA levels 4 weeks after ischemia/reperfusion. Ticagrelor, but not clopidogrel, attenuated the increase in proinflammatory tumor necrosis factor-α, interleukin-1ß, and interleukin-18, and increased anti-inflammatory 15-epi-lipoxin-A4 levels. CONCLUSIONS: Ticagrelor, but not clopidogrel, administered just before reperfusion protects against reperfusion injury. This acute treatment or chronic ticagrelor for 4 weeks or their combination improved heart function, whereas clopidogrel, despite achieving a similar degree of platelet inhibition, had no effect.
Assuntos
Adenosina/análogos & derivados , Cardiotônicos/farmacologia , Infarto do Miocárdio/tratamento farmacológico , Traumatismo por Reperfusão Miocárdica/prevenção & controle , Miocárdio/patologia , Antagonistas do Receptor Purinérgico P2Y/farmacologia , Função Ventricular Esquerda/efeitos dos fármacos , Remodelação Ventricular/efeitos dos fármacos , Adenosina/administração & dosagem , Adenosina/farmacologia , Administração Oral , Animais , Apoptose/efeitos dos fármacos , Cardiotônicos/administração & dosagem , Clopidogrel , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Fibrose , Injeções Intraperitoneais , Masculino , Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Infarto do Miocárdio/fisiopatologia , Traumatismo por Reperfusão Miocárdica/metabolismo , Traumatismo por Reperfusão Miocárdica/patologia , Traumatismo por Reperfusão Miocárdica/fisiopatologia , Miocárdio/metabolismo , Antagonistas do Receptor Purinérgico P2Y/administração & dosagem , Ratos Sprague-Dawley , Transdução de Sinais/efeitos dos fármacos , Ticagrelor , Ticlopidina/análogos & derivados , Ticlopidina/farmacologia , Fatores de TempoRESUMO
In patients with acute coronary syndrome, dual antiplatelet therapy with aspirin and a P2Y12 inhibitor like prasugrel is prescribed for one year. Here, we investigated how the hemostatic function of platelets recovers after discontinuation of prasugrel treatment. Therefore, 16 patients who suffered from ST-elevation myocardial infarction were investigated. Patients were treated with aspirin (100 mg/day, long-term) and stopped taking prasugrel (10 mg/day) after one year. Blood was collected at the last day of prasugrel intake and at 1, 2, 5, 12 and 30 days later. Platelet function in response to ADP was normalized between five and 30 days after treatment cessation and in vitro addition of the reversible P2Y12 receptor antagonist ticagrelor fully suppressed the regained activation response. Discontinuation of prasugrel resulted in the formation of an emerging subpopulation of ADP-responsive platelets, exhibiting high expression of active integrin αIIbß3. Two different mRNA probes, thiazole orange and the novel 5'Cy5-oligo-dT probe revealed that this subpopulation consisted of juvenile platelets, which progressively contributed to platelet aggregation and thrombus formation under flow. During offset, juvenile platelets were overall more reactive than older platelets. Interestingly, the responsiveness of both juvenile and older platelets increased in time, pointing towards a residual inhibitory effect of prasugrel on the megakaryocyte level. In conclusion, the gradual increase in thrombogenicity after cessation of prasugrel treatment is due to the increased activity of juvenile platelets.
Assuntos
Síndrome Coronariana Aguda/tratamento farmacológico , Síndrome Coronariana Aguda/metabolismo , Coagulação Sanguínea/efeitos dos fármacos , Plaquetas/metabolismo , Infarto do Miocárdio/tratamento farmacológico , Infarto do Miocárdio/metabolismo , Cloridrato de Prasugrel/administração & dosagem , Difosfato de Adenosina/metabolismo , Adulto , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Testes de Função Plaquetária , Complexo Glicoproteico GPIIb-IIIa de Plaquetas/metabolismo , Receptores Purinérgicos P2Y12/metabolismoRESUMO
Ticagrelor is a direct-acting reversibly binding P2Y12 antagonist and is widely used as an antiplatelet therapy for the prevention of cardiovascular events in acute coronary syndrome patients. However, antiplatelet therapy can be associated with an increased risk of bleeding. Here, we present data on the identification and the in vitro and in vivo pharmacology of an antigen-binding fragment (Fab) antidote for ticagrelor. The Fab has a 20 pM affinity for ticagrelor, which is 100 times stronger than ticagrelor's affinity for its target, P2Y12. Despite ticagrelor's structural similarities to adenosine, the Fab is highly specific and does not bind to adenosine, adenosine triphosphate, adenosine 5'-diphosphate, or structurally related drugs. The antidote concentration-dependently neutralized the free fraction of ticagrelor and reversed its antiplatelet activity both in vitro in human platelet-rich plasma and in vivo in mice. Lastly, the antidote proved effective in normalizing ticagrelor-dependent bleeding in a mouse model of acute surgery. This specific antidote for ticagrelor may prove valuable as an agent for patients who require emergency procedures.
Assuntos
Adenosina/análogos & derivados , Anticorpos Neutralizantes/química , Anticorpos Neutralizantes/farmacologia , Antídotos/química , Antídotos/farmacologia , Adenosina/antagonistas & inibidores , Adenosina/imunologia , Animais , Anticorpos/isolamento & purificação , Anticorpos/metabolismo , Especificidade de Anticorpos , Anticorpos Amplamente Neutralizantes , Células CHO , Cricetinae , Cricetulus , Cristalografia por Raios X , Hemorragia/prevenção & controle , Humanos , Fragmentos Fab das Imunoglobulinas/farmacologia , Camundongos , Modelos Moleculares , Agregação Plaquetária/efeitos dos fármacos , Engenharia de Proteínas , TicagrelorRESUMO
OBJECTIVE: In a phase III clinical trial (PLATelet inhibition and patient Outcomes, PLATO), ticagrelor provided better clinical outcomes than clopidogrel in patients with acute coronary syndromes. In addition to P2Y12-receptor antagonism, ticagrelor prevents cell uptake of adenosine and has proven able to augment adenosine effects. Adenosine protects the heart against ischemia-reperfusion injury. We compared the effects of clopidogrel and ticagrelor on myocardial infarct size (IS). APPROACH AND RESULTS: Rats received oral ticagrelor (0, 75, 150, or 300 mg/kg/d) or clopidogrel (30 or 90 mg/kg/d) for 7 days and underwent 30-minute coronary artery ligation and 24-hour reperfusion. Area at risk was assessed by blue dye and IS by 2,3,5-triphenyl-tetrazolium-chloride. Cyclooxygenase-2 (COX2) enzyme activity was assessed by ELISA and expression by real-time polymerase chain reaction. Mechanism responsible was explored using adenosine-receptor antagonist (CGS15943, an A2A/A1 antagonist) or cyclooxygenase inhibition by either aspirin (5, 10, or 25 mg/kg) or specific cyclooxygenase-1 (SC560) or COX2 (SC5815) inhibitors. Ticagrelor, dose-dependently, reduced IS, whereas clopidogrel had no effect. Adenosine-receptor antagonism blocked the ticagrelor effect and COX2 inhibition by SC5815, or high-dose aspirin attenuated the IS-limiting effect of ticagrelor, whereas cyclooxygenase-1 inhibition or low-dose aspirin had no effect. Ticagrelor, but not clopidogrel, upregulated COX2 expression and activity. Also this effect was blocked by adenosine-receptor antagonism. Ticagrelor, but not clopidogrel, increased Akt and endothelial nitric oxide synthase phosphorylation. CONCLUSIONS: Ticagrelor, but not clopidogrel, reduces myocardial IS. The protective effect of ticagrelor was dependent on adenosine-receptor activation with downstream upregulation of endothelial nitric oxide synthase and COX2 activity.
Assuntos
Adenosina/análogos & derivados , Adenosina/fisiologia , Cardiotônicos/uso terapêutico , Ciclo-Oxigenase 2/fisiologia , Infarto do Miocárdio/tratamento farmacológico , Traumatismo por Reperfusão Miocárdica/prevenção & controle , 6-Cetoprostaglandina F1 alfa/metabolismo , Adenosina/farmacologia , Adenosina/uso terapêutico , Antagonistas do Receptor A1 de Adenosina/farmacologia , Antagonistas do Receptor A2 de Adenosina/farmacologia , Animais , Aspirina/farmacologia , Cardiotônicos/farmacologia , Clopidogrel , Ciclo-Oxigenase 2/biossíntese , Ciclo-Oxigenase 2/genética , Inibidores de Ciclo-Oxigenase 2/farmacologia , Avaliação Pré-Clínica de Medicamentos , Indução Enzimática/efeitos dos fármacos , Lipoxinas/metabolismo , Masculino , Infarto do Miocárdio/patologia , Traumatismo por Reperfusão Miocárdica/tratamento farmacológico , Traumatismo por Reperfusão Miocárdica/patologia , Óxido Nítrico Sintase Tipo III/biossíntese , Óxido Nítrico Sintase Tipo III/genética , Óxido Nítrico Sintase Tipo III/fisiologia , Pirazóis/farmacologia , Quinazolinas/farmacologia , Ratos , Ratos Sprague-Dawley , Receptor A1 de Adenosina/fisiologia , Receptor A2A de Adenosina/fisiologia , Ticagrelor , Ticlopidina/análogos & derivados , Ticlopidina/uso terapêutico , Triazóis/farmacologia , Regulação para Cima/efeitos dos fármacosRESUMO
This review constitutes a critical evaluation of recent publications that have described an additional mode of action of the P2Y12 receptor antagonist ticagrelor. The effect is mediated by inhibition of the adenosine transporter ENT1 (type 1 equilibrative nucleoside transporter), which provides protection for adenosine from intracellular metabolism, thus increasing its concentration and biological activity, particularly at sites of ischemia and tissue injury where it is formed. Understanding the mode of action of ticagrelor is of particular interest given that its clinical profile, both in terms of efficacy and adverse events, differs from that of thienopyridine P2Y12 antagonists.
Assuntos
Síndrome Coronariana Aguda/tratamento farmacológico , Adenosina/análogos & derivados , Adenosina/metabolismo , Antagonistas do Receptor Purinérgico P2Y/uso terapêutico , Síndrome Coronariana Aguda/metabolismo , Adenosina/uso terapêutico , Humanos , Ticagrelor , Resultado do TratamentoRESUMO
OBJECTIVE: The effect of proton pump inhibitors (PPIs) on the pharmacokinetics and pharmacodynamics of clopidogrel was assessed in two healthy volunteer crossover studies. SUBJECTS AND METHODS: Study 1: subjects received clopidogrel alone (300-mg loading dose, then 75 mg/day for 28 days) and two of three PPIs (omeprazole 80 mg, esomeprazole 40 mg or lansoprazole 60 mg) plus clopidogrel for 29 days in three treatment periods (randomized treatment sequence assignment). Study 2: subjects received clopidogrel alone (75 mg/day for 9 days) and clopidogrel alone for 4 days followed by clopidogrel plus fixed-combination esomeprazole 20 mg/low-dose acetylsalicylic acid (ASA) 81 mg for 5 days in two treatment periods (randomized treatment sequence assignment). Pharmacokinetic effects were estimated by measuring active metabolite of clopidogrel, and pharmacodynamic effects by inhibition of adenosine diphosphate (ADP)-induced platelet aggregation. RESULTS: There was a relative decrease of up to 50 % in exposure to the active metabolite of clopidogrel with the different PPIs (study 1), and close to 40 % with esomeprazole/low-dose ASA (study 2), compared with clopidogrel alone. There was an absolute decrease of up to 17 % in inhibition of ADP-induced platelet aggregation with co-administration of different PPIs, compared with clopidogrel alone; however, no differences in platelet inhibition were observed during co-administration with the esomeprazole/low-dose ASA fixed-dose combination. CONCLUSION: Omeprazole, esomeprazole and lansoprazole decreased systemic exposure to the active metabolite of clopidogrel in healthy volunteers, leading to modest decreases in its antiplatelet effect. However, no apparent differences in platelet inhibition were observed when esomeprazole was co-administered with low-dose ASA as a fixed-dose combination.
Assuntos
Inibidores da Bomba de Prótons/farmacologia , Ticlopidina/análogos & derivados , Adolescente , Adulto , Área Sob a Curva , Aspirina/administração & dosagem , Aspirina/farmacologia , Clopidogrel , Estudos Cross-Over , Combinação de Medicamentos , Interações Medicamentosas , Quimioterapia Combinada , Esomeprazol/administração & dosagem , Esomeprazol/farmacologia , Feminino , Voluntários Saudáveis , Humanos , Lansoprazol/farmacologia , Masculino , Pessoa de Meia-Idade , Omeprazol/farmacologia , Agregação Plaquetária/efeitos dos fármacos , Inibidores da Bomba de Prótons/administração & dosagem , Ticlopidina/farmacocinética , Adulto JovemRESUMO
INTRODUCTION: Studies have shown that ticagrelor has a further adenosine-mediated mechanism of action in addition to its potent inhibition of the P2Y12 receptor, which may explain some of ticagrelor's clinical characteristics. This study aimed to further characterize the adenosine pharmacology of ticagrelor, its major metabolites, and other P2Y12 receptor antagonists. METHODS: Inhibition of nucleoside transporter-mediated [(3)H]adenosine uptake by ticagrelor, its major metabolites, and alternative P2Y12 antagonists was examined in recombinant Madin-Darby canine kidney (MDCK) cells. The pharmacology of ticagrelor and its major metabolites at adenosine A1, A2A, A2B, and A3 receptor subtypes was examined using in vitro radioligand binding and functional assays and ex vivo C-fiber experiments in rat and guinea pig vagus nerves. RESULTS: Ticagrelor (and less effectively its metabolites) and the main cangrelor metabolite inhibited [(3)H]adenosine uptake in equilibrative nucleoside transporter (ENT) 1-expressing MDCK cells, whereas cangrelor and the active metabolites of prasugrel or clopidogrel had no effect. No significant inhibitory activity was observed in MDCK cells expressing ENT2 or concentrative nucleoside transporters 2/3. Ticagrelor demonstrated high affinity (inhibition constant [Ki] = 41 nmol/L) for ENT1. In adenosine receptor-binding experiments, ticagrelor and its major circulating metabolite, AR-C124910XX, had low affinity (Ki > 6 µmol/L) for each of the adenosine A1, A2A, and A2B receptors, whereas ticagrelor had a submicromolar (Ki = 190 nmol/L) affinity for the adenosine A3 receptor. However, in functional assays, at high concentrations (10 µmol/L) ticagrelor only partially inhibited 3 mmol/L adenosine-induced depolarizations in the guinea pig and rat vagus nerve preparations (by 35% and 49%, respectively). CONCLUSIONS: Ticagrelor inhibits cellular adenosine uptake selectively via ENT1 inhibition at concentrations of clinical relevance. However, the low-binding affinity and functional inhibition of adenosine receptors observed with ticagrelor or its metabolites indicate that they possess a negligible adenosine-like activity at clinically relevant concentrations.