Binding of [3H]-2-methylthio ADP to rat platelets--effect of clopidogrel and ticlopidine.
Thienopyridine compounds, including ticlopidine and clopidogrel, have been found to selectively inhibit adenosine 5' diphosphate (ADP)-induced platelet aggregation and adenylyl cyclase ex vivo, but the mechanism of their antiplatelet action remains to be determined. This study was aimed at investigating the effect of clopidogrel and ticlopidine on the binding of [3H]-2-methylthio- adenosine-5'-diphosphate (2-MeS-ADP) to rat platelets. Binding of [3H]-2-MeS-ADP to rat platelets was time-dependent and saturable. Scatchard analysis of the saturation binding data indicated that [3H]-2-MeS-ADP bound to one population of specific binding sites with high affinity (KD = 0.78 +/- 0.05 nM; Bmax = 156.3 +/- 4.8 fmole/10(8) cells) (n = 3). Unlabeled 2-MeS-ADP and ADP competitively and selectively inhibited the specific binding of [3H]-2-MeS-ADP with IC50 values of 11.3 +/- 1.2 nM and 11.3 +/- 0.7 microM, respectively (n = 3). Other nucleotide analogs such as ADP-beta S, ATP and ATP-alpha S also antagonized [3H]-2-MeS-ADP binding. When administered orally at doses ranging from 1 to 25 mg/kg, clopidogrel inhibited ADP- or 2-MeS-ADP-induced platelet aggregation as well as ADP or 2-MeS-ADP-induced inhibition of intraplatelet adenylyl cyclase. When measured in parallel, clopidogrel reduced in a dose-dependent manner the binding of [3H]-2-MeS-ADP to rat platelets ex vivo. Clopidogrel administration resulted in the decrease of [3H]-2-MeS-ADP binding sites on platelets without any significant change in the affinity; this indicates noncompetitive binding. Ticlopidine (200 mg/kg a day for 3 days) behaved in the same way.(ABSTRACT TRUNCATED AT 250 WORDS)
Savi
P
P
Sanofi Recherche, Toulouse, France.
Laplace
M C
MC
Maffrand
J P
JP
Herbert
J M
JM
D000244:Adenosine Diphosphate / Q000031:analogs & derivatives* / Q000097:blood; D000262:Adenylyl Cyclases / Q000378:metabolism; D000818:Animals; D001665:Binding Sites / Q000187:drug effects; D001792:Blood Platelets / Q000187:drug effects* / Q000201:enzymology / Q000378:metabolism; D000077144:Clopidogrel; D005260:Female; D066298:In Vitro Techniques; D010975:Platelet Aggregation Inhibitors / Q000494:pharmacology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D018048:Receptors, Purinergic P2 / Q000187:drug effects; D013873:Thionucleotides / Q000097:blood*; D013988:Ticlopidine / Q000031:analogs & derivatives* / Q000494:pharmacology*
D010975:Platelet Aggregation Inhibitors; D018048:Receptors, Purinergic P2; D013873:Thionucleotides; C002480:methylthio-ADP; D000244:Adenosine Diphosphate; D000077144:Clopidogrel; D000262:Adenylyl Cyclases; D013988:Ticlopidine