Molecular Mechanisms involved in Collagen mediated Platelet Activation and their Modulation by Anti-platelet Compounds

Abstract

Atherothrombotic diseases are a major healthcare problem [1] and are responsible for >25% of all deaths worldwide[2]. Long considered a disease of industrialized countries, recent World Health Organization statistics have highlighted the true global impact of this disease [2] with ~80% of the world’s deaths from atherothrombosis occurring in low-and middle-income countries. The development of a clot in the coronary or cerebral circulation (causing acute myocardial infarction or ischemic stroke, respectively) is now the single most common cause of morbidity and mortality globally, and the prevalence of these diseases continues to rise, particularly in developing nations. Platelets have a central role in cardiovascular thrombosis. They adhere to the sub-endothelial matrix after endothelial damage due to a ruptured atherosclerotic plaque, and then aggregate with each other to form a prothrombotic surface that promotes clot formation and subsequently vascular occlusion. As a result, therapies targeting key pathways of platelet activation — including thromboxane A2 synthesis, ADP-mediated signalling and integrin αIIbβ3 (also know as GPIIb–IIIa) signalling — have established a role in the treatment of cardiovascular arterial disease. The most common of these anti-platelet agents include aspirin, clopidogrel and integrin αIIbβ3 antagonists. The most clear-cut evidence that acute myocardial infarction is a platelet-related disease is the ability of anti-platelet therapy to reduce mortality and morbidity in this clinical setting. Despite intense investigation over the last 40 years into the discovery and development of more effective antithrombotic drugs, the effect of these therapies on mortality rates has remained disappointingly small, with less than one in four individuals taking antithrombotic therapies avoiding a fatal thrombotic event. Limitations of current therapies include weak inhibition of platelet function (for example, by aspirin), blockade of only one pathway of ADP-mediated signalling (for example, by clopidogrel), slow onset of action (for example, of clopidogrel), inter-patient response variability with poor inhibition of platelet response in some patients (for example, to clopidogrel), the inability to transform the success of intravenous integrin αIIbβ3 antagonist therapy into oral therapy and the inability to completely separate a reduction in thrombotic events from an increase in bleeding events. This situation has become more challenging as the incidence of obesity, diabetes and the metabolic syndrome is rapidly increasing. These conditions are typically more resistant to the benefits of anti-thrombotic therapy [3], and, thus, there is a need for the identification and development of more effective approaches to combat this global cardiovascular epidemic. Angiographic studies of individuals who have undergone successful pharmacological thrombolysis have showed that >50% of the culprit lesions cause <50% of the coronary stenosis [4, 5]. This indicates that in a high proportion of individuals with acute myocardial infarction, an exaggerated thrombotic response at sites of plaque disruption is probably a major factor in the development of disease. Multiple factors are known to contribute to an exaggerated platelet response at sites of atherosclerotic plaque rupture including the presence of potent thrombogenic elements within the atherosclerotic plaque, heightened platelet reactivity and a loss of the normal control mechanisms dampening platelet activation, as well as localized blood flow disturbances at the atherothrombotic site. Among the multiple factors that contribute to the heightened thrombogenic potential of ruptured atherosclerotic lesions, includes the presence of tissue factor, and fibrillar collagens in the lesion, as well as the presence of platelet-activating lipids in the necrotic lipid core. Type I and III collagens are among the most potent activators of platelets and support efficient platelet adhesion and aggregation under the rapid blood flow conditions that operate in stenosed arteries. Blockade or deficiency of the platelet collagen receptors (GPVI and integrin α2β1) in humans and animals does not produce a marked bleeding tendency, raising the possibility that targeting platelet collagen receptors may be a well-tolerated approach to reduce excess platelet deposition on disrupted atherosclerotic plaques. There is compelling evidence for a crucial role of GPVI in arterial thrombosis from studies in mice. Massberg et al [6] demonstrated that thrombus formation in the injured carotid artery in mice is virtually abolished in the absence of functional GPVI. This agrees with a recent study by Konishi et al [7] who found markedly reduced platelet attachment and subsequent neointimal hyperplasia at sites of vascular injury in FcRγ chain–deficient mice that lack GPVI. These developments implicate GPVI as a potential pharmacologic target for the treatment of ischemic cardiovascular disease. Such a strategy might have a number of advantages. First, GPVI is exclusively expressed on platelets and megakaryocytes, which prevent the risk of side effects of anti-GPVI agents on other cell types. Second, GPVI deficiency is not associated with a major bleeding risk in humans and mice, suggesting that anti-GPVI therapy might be well tolerated. The last decade has seen major advances in understanding how platelets interact with collagen and the events that initiate primary hemostasis and arterial thrombosis. The cloning of GPVI, the generation of mice deficient in collagen receptors and their associated signaling molecules, and the availability of new antibodies and collagen-derived peptides allowed detailed in vitro and in vivo studies on the role of the individual candidate receptors in the complex process of platelet tethering, activation, adhesion, aggregation, and procoagulant activity [8] [9]. These studies have changed the long-standing concept of platelet-collagen interactions, the so called 2-site, 2-step model, which proposed α2β1 as the major collagen receptor in hemostasis and thrombosis. This hypothesis had been based on the assumption that the integrin α2β1 is constitutively in a high-affinity conformation and is essential for the initial firm arrest of platelets on collagen. However, it is now recognized that α2β1, like the other integrins, is in a low-affinity state on resting platelets and requires inside-out signals to efficiently bind to collagen. It is also now established that the initial platelet contact with collagen and the subsequent initiation of integrin activation (ie, adhesion and thrombus growth) are strictly dependent on functional GPVI. These developments identify a new sequence of events in the initial phase of hemostasis and thrombosis and therefore, place GPVI in a central position in this complex process [8]. Thus, there is considerable interest in developing novel tools, such as receptor antagonists, depleting antibodies, or recombinant peptides to evaluate GPVI function in humans, because in contrast to the existing drugs, which inhibits all platelets and therefore incur markedly increased risk of bleeding, compounds that specifically interfere in the collagen induced activation of platelets hold promise for the reduced risk of complications.