Structure and Stereochemistry of the Active Metabolite of Clopidogrel

Pereillo, Jean-Marie; Maftouh, Mohamed; Andrieu, Alain; Uzabiaga, Marie-Françoise; Fedeli, Olivier; Savi, Pierre; Pascal, Marc; Herbert, Jean‐Marc; Maffrand, Jean‐Pierre; Picard, Claudine

doi:10.1124/dmd.30.11.1288

Cited by 419 publications

(355 citation statements)

References 11 publications

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“…Clopidogrel [SR25990C, 2-(2-chlorophenyl)-2-(2,4,5,6,7,7a hexahydrothieno [3,2c] pyridine-5yl-acetic acid methyl ester hydrogen sulfate 7S] was from Sanofi-Aventis (Toulouse, France). The active metabolite of clopidogrel (ActMet) was obtained by biotransformation and purified as described (54,55).…”

Section: Methodsmentioning

confidence: 99%

“…Indeed, in vivo studies have demonstrated that clopidogrel has to undergo hepatic metabolization to obtain an active metabolite (53). This active metabolite (Act-Met) has been isolated, and its structure has been elucidated (54). It contains a free thiol function, and its activity is lost when the thiol is derivatized (55), suggesting its possible interaction with cysteine-containing sequences.…”

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confidence: 99%

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The active metabolite of Clopidogrel disrupts P2Y12 receptor oligomers and partitions them out of lipid rafts

Savi

Zachayus

Delesque-Touchard

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

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P2Y12, a G protein-coupled receptor that plays a central role in platelet activation has been recently identified as the receptor targeted by the antithrombotic drug, clopidogrel. In this study, we further deciphered the mechanism of action of clopidogrel and of its active metabolite (Act-Met) on P2Y12 receptors. Using biochemical approaches, we demonstrated the existence of homooligomeric complexes of P2Y12 receptors at the surface of mammalian cells and in freshly isolated platelets. In vitro treatment with Act-Met or in vivo oral administration to rats with clopidogrel induced the breakdown of these oligomers into dimeric and monomeric entities in P2Y12 expressing HEK293 and platelets respectively. In addition, we showed the predominant association of P2Y12 oligomers to cell membrane lipid rafts and the partitioning of P2Y12 out of rafts in response to clopidogrel and Act-Met. The raft-associated P2Y12 oligomers represented the functional form of the receptor, as demonstrated by binding and signal transduction studies. Finally, using a series of receptors individually mutated at each cysteine residue and a chimeric P2Y12͞P2Y13 receptor, we pointed out the involvement of cysteine 97 within the first extracellular loop of P2Y12 in the mechanism of action of Act-Met. mechanism of action ͉ platelet ͉ antiaggregant M any G protein-coupled receptors (GPCRs) have been shown to assemble as homodimers, heterodimers, as well as larger oligomers (1, 2). The existence of such oligomeric entities raises questions as to their functional consequences as well as their physiological relevance. Heterologous expression systems have provided a variety of answers concerning ligand-dependent regulation of GPCR oligomeric states. Ligand binding, depending on the GPCR studied, can promote (3-10) or inhibit (11-13) dimer formation, as well as having no effect on preexisting constitutive homo-or heterodimers (14-25). The fact that heterodimerization may alter the pharmacological properties of a GPCR along with its internalization and signal transduction behavior is of critical importance (26, 27).Clustering, even for nonheptahelical receptors, now appears as a common feature of cell signaling. Specialized structures such as clathrin-coated pits, caveolae, and lipid rafts contain high concentrations of signaling molecules. Rafts represent dynamic assemblies of proteins and lipids, mostly sphingolipids and cholesterol (28,29). Proteins such as glycophosphatidylinositol-anchored proteins, nonreceptor tyrosine kinases, G␣ subunits of heterotrimeric G proteins, and palmitoylated proteins appear to localize to these microdomains (30). In addition, recent studies have shown that partitioning of proteins in and out of rafts can depend on their state of activation or dimerization (31-33). A variety of GPCR have also been identified in caveolae or rafts. These include ␣ and ␤-adrenergic receptors (34, 35), adenosine A1 receptor (36), angiotensin II type 1 receptor (37), muscarinic receptor (38), EDG1 receptor (39), bradykinin B1 and B2 receptor...

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Section: Methodsmentioning

confidence: 99%

mentioning

confidence: 99%

The active metabolite of Clopidogrel disrupts P2Y12 receptor oligomers and partitions them out of lipid rafts

Savi

Zachayus

Delesque-Touchard

et al. 2006

Proc. Natl. Acad. Sci. U.S.A.

Self Cite

277

221

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“…The active metabolite of clopidogrel [molecular weight (MW) 353 Da] has low-BBB permeability (24). Clopidogrel would thus not be expected to interfere with the movement of microglia or their ramified processes under control conditions.…”

Section: Systemic Clopidogrel Suppresses Juxtavascular Microglial Celmentioning

confidence: 99%

Purinergic receptor P2RY12-dependent microglial closure of the injured blood–brain barrier

Lou

Takano

Pei

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

293

227

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Microglia are integral functional elements of the central nervous system, but the contribution of these cells to the structural integrity of the neurovascular unit has not hitherto been assessed. We show here that following blood-brain barrier (BBB) breakdown, P2RY12 (purinergic receptor P2Y, G-protein coupled, 12)-mediated chemotaxis of microglia processes is required for the rapid closure of the BBB. Mice treated with the P2RY12 inhibitor clopidogrel, as well as those in which P2RY12 was genetically ablated, exhibited significantly diminished movement of juxtavascular microglial processes and failed to close laser-induced openings of the BBB. Thus, microglial cells play a previously unrecognized protective role in the maintenance of BBB integrity following cerebrovascular damage. Because clopidogrel antagonizes the platelet P2Y12 receptor, it is widely prescribed for patients with coronary artery and cerebrovascular disease. As such, these observations suggest the need for caution in the postincident continuation of P2RY12-targeted platelet inhibition. purinergic receptors | microglia | blood-brain barrier | stroke | clopidogrel

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“…More importantly, the efficacy enhancement is unlikely to jeopardize the clinical safety records of CPG. Currently, to overcome the disadvantages of CPG, substituting therapies are as follows: (1) dose increase of CPG (e.g., doubling), (2) adjunctive use of cilostazol with CPG, and (3) use of other antiplatelet agents, such as prasugrel or ticagrelor. 21 Being closely related to CPG, the improved deuterated analogues are expected to stay in the narrow therapeutic window and be potentially more beneficial than the above alternative treatments.…”

Section: * S Supporting Informationmentioning

confidence: 99%

Deuterated Clopidogrel Analogues as a New Generation of Antiplatelet Agents

Zhu

Zhou

Jiang

2013

ACS Med. Chem. Lett.

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Clopidogrel (CPG) is an antithrombotic prodrug that needs hepatic cytochrome P450 (CYP) enzymes for its bioactivation. The clinical effects of CPG have been associated with high intersubject variability and a certain level of resistance. Recently, comprehensive biotransformation studies of CPG support that the observed clinical uncertainty stems from the low bioactivation efficiency, which is attributed to extensive attritional metabolism (e.g., hydrolysis of the methyl ester functionality and oxidation of the piperidine moiety). With the goal of potentiating the desired thiophene 2-oxidation through minimal structural modification, we have adopted the strategy of targeted metabolism shift and have designed and synthesized deuterated piperidine analogues of CPG. In vitro studies showed that the prodrug activation percentages have been significantly increased for the deuterated analogues as a result of stability enhancement of the piperidine moiety. In a pharmacological study with a rat model, oral administration of the deuterated analogues also demonstrated higher inhibitory activity than that of CPG against adenosine diphosphate (ADP) induced platelet aggregation. These deuterated analogues represent a new generation of antiplatelet agents with the potential to overcome the major clinical drawbacks of CPG. KEYWORDS: Clopidogrel resistance, clinical variability, prodrug attrition, piperidine deuteration, targeted metabolic shift, bioactivation potentiation C lopidogrel (CPG) is a thienopyridine antiplatelet prodrug that has been widely used in the treatment of cardiovascular diseases, including atherothrombosis, unstable angina, and myocardial infarction. 1 As shown in Scheme 1, CPG (M0) is activated through a two-step cytochrome P450 (CYP)-catalyzed process to form its active metabolite (M13). 2,3 Despite being one of the most prescribed cardiovascular medications of the past decade, CPG is associated with a high clinical uncertainty for its antithrombotic therapy. It has been reported that 20−40% of patients that receive the drug showed poor or no response to it. 4,5 To address the observed clinical variability and resistance, the chemical mechanism of CPG bioactivation has been under extensive investigation. Genetic factors that can impact the therapeutic outcomes are being sought for the design of personalized prodrug treatments. It has been hypothesized that the active metabolite formation from CPG is dependent on genetic polymorphic enzymes, and this leads to the observed intersubject variability. However, research results have shown that both steps of the prodrug activation are catalyzed by various CYPs, and genetic polymorphic enzymes such as paraoxonase-1 (PON-1) or CYP2C19 do not play deciding roles in catalyzing the active metabolite formation. 6−9 Our recent research has revealed that the first step of CPG bioactivation, 2-oxidation of the thiophene motif leading to M2 formation, is significantly attenuated by CYP3A4/5-catalyzed oxidation of the nonactivating piperidine motif and the

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Structure and Stereochemistry of the Active Metabolite of Clopidogrel

Cited by 419 publications

References 11 publications

The active metabolite of Clopidogrel disrupts P2Y12 receptor oligomers and partitions them out of lipid rafts

The active metabolite of Clopidogrel disrupts P2Y12 receptor oligomers and partitions them out of lipid rafts

Purinergic receptor P2RY12-dependent microglial closure of the injured blood–brain barrier

Deuterated Clopidogrel Analogues as a New Generation of Antiplatelet Agents

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