Chemistry: molecular biology and microbiology – Measuring or testing process involving enzymes or... – Involving antigen-antibody binding – specific binding protein...
Utility Patent
1999-02-08
2001-01-02
Saunders, David (Department: 1644)
Chemistry: molecular biology and microbiology
Measuring or testing process involving enzymes or...
Involving antigen-antibody binding, specific binding protein...
C435S007210, C435S007240, C530S388220, C530S388700, C530S388250
Utility Patent
active
06168925
ABSTRACT:
The invention relates to a method of analysing the platelet GPIIb/IIIa receptors occupied by an anti-platelet aggregation compound or by fibrinogen and the receptors which have remained in the free state.
The platelets play a key role in the equilibrium of the haemostatic balance. Activating signals can cause, at their level, both morphological and biochemical modifications together with variations in the levels of expression of surface glycoproteins. These glycoproteins are, for most ligand receptors, involved in adhesion (eg. the receptor for the von Willebrand factor: the molecule GPIb), in aggregation (eg. the receptor for fibrinogen: the molecule GPIIb/IIIa) whereas others indicate the state of platelet activation (eg. the molecule GMP 140 or P-selectin).
The activation of platelets and the aggregation resulting therefrom are physiological phenomena which, after exceeding a threshold, are associated with various pathological conditions such as arterial thrombotic accidents. This explains the reason for the interest which the pharmaceutical industry has for developing new anti-aggregation therapeutic molecules.
The preferred target in the new therapies developed is the GPIIb/IIIa receptor which is specifically expressed on the platelets. Under the effect of an activation of the platelets, this receptor binds, with a very high affinity, fibrinogen and other adhesion proteins, which causes aggregation of the platelets with each other.
The GPIIb/IIIa receptor belongs to the group comprising the integrins, which is composed of various molecules involved in the phenomena of cell adhesion. The integrins are a/p heterodimers and are classified into several families according to their &bgr;-subunit. About twenty integrins are currently known, of which 9 bind to their ligand via the amino acid sequence R—G—D.
GPIIb/IIIa (&agr;II&bgr;3) is a member of the family of &bgr;3 integrins. It has the same &bgr; subunit (but a different &agr; subunit) as the vitronectin receptor (&agr;v&bgr;3) present especially on the platelets and, in a large quantity, on the endothelial cells. The IIb and IIIa subunits are classified respectively into the CD 41 and CD 61 groups according to the international nomenclature.
The anti-platelet GPIIb-IIIa anti-aggregation agents belong to two different classes: a) the monoclonal antibodies which are antagonists of fibrinogen, including C7E3Fab of the humanized 7E3 monoclonal antibody directed against an epitope of CD 61 and marketed under the name a 7E3 monoclonal antibody solid under the tradename REOPRO®, b) the cyclic peptides, of the integrelin type, which possess the sequence RGD or KGD, close to the RGD sequence (Arg—Gly—Asp) common to the platelet receptor ligands, and the peptidomimetics, of the lamifiban or tirofiban type. These currently developed substances are only active by the parenteral route. Other molecules which can be administered by the oral route are being developed. These new molecules may, however, induce side effects which result in the patients treated in haemorrhagic effects at the points of access or of internal location. For these new molecules which, by their nature, remain in circulation for a long time, there is currently no antidote. It is therefore important, in the context of an anti-platelet therapy, to be able not only to adapt the therapeutic dose of the agent administered, but also to rapidly have available, during the monitoring of the treatment, information on the number of GPIIb/IIIa receptors occupied by the antagonist, so as to be able to control the haemorrhagic complications while preserving a sufficient quantity of bound antagonist in order to obtain an anti-aggregation effect.
Accordingly, the present invention relates to a method of determining, on a test sample, the occupation of the GPIIb/IIIa receptor, by an antagonist of fibrinogen, characterized in that the number of receptors occupied is determined with the aid of an antibody [MAb1] which is a competitor for the antagonist, and the total number of occupied or unoccupied receptors is determined with the aid of a so-called noncompetitor antibody [MAb2] specific for the occupied or unoccupied receptor and the occupation of the GPIIb/IIIa receptors in the sample is deduced therefrom.
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B.S. Coller et al., “Rapid and Simple Platelet Function Assay to Assess Glycoprotein IIb/IIIa Receptor Blockade”, Circulation Vo. 95, No. 4, Feb. 18, 1997, pp. 860-867 (XP-002057028).
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Besson-Faure Isabelle
Canton Michel
Biocytex
DeCloux Amy
Foley & Lardner
Saunders David
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