Cardiovascular disease remains a leading cause of death in North America (1). Uncontrolled platelet activation, adhesion and aggregation initiated by vessel wall plaque rupture are thought to be responsible for acute vascular occlusion in many situations (2-5). Although many platelet inhibition drugs are available, all currently available drugs have undesirable toxicity profiles (6-8). Thus, reduction in toxicity and improved management of patients with thrombotic diseases remains an unmet medical need.

Platelet activation plays a pivotal role in the pathogenesis of acute coronary syndromes, strokes and other thrombophilic diseases. Atheromatous plaque rupture changes the shear forces of blood flowing over the injured vessel surface and also exposes collagen as well as other prothrombotic factors (9-11). As the initial hemostatic event, platelets become activated and cover the injured surface. Following platelet activation highly active substances like adenosine diphosphate (ADP) and thromboxane A2 (TxA2) are released from the platelet to promote and recruit further platelet aggregation to the injury site (12). If this process proceeds unabated, as it often does in atherosclerotic diseases, the vessel becomes occluded and infarction may follow.

Lovaza® (Reliant Pharmaceutical Inc., Liberty Corner, NJ), a commercially available formulation that contains 90 % omega-3-acid ethyl esters (46% eicosapentaenoic acid -EPA- and 38% docosohexaenoic acid -DHA-), has the potential ability to modify the recruitment of additional platelets to the growing thrombus by promoting synthesis of thromboxane A3 (TxA3), a poor platelet activator, instead of thromboxane A2, a potent platelet activator. Agents used to inhibit platelet function such as aspirin and clopidogrel are not always effective (13-16). Unfortunately, some patients do not respond to these therapeutics (17-24). Realistic numbers for patient resistance to these drugs are probably 10-15% for ASA and 20-30% for clopidogrel. Almost all resistant patients have less favorable outcomes and are unaware of this potentially life-threatening problem until a severe cardiac adverse event occurs. Lovaza® may add additional therapeutic benefit to these patients.(25,26) Beyond the occasional patient with complaints of eructation or a "fishy" taste in their mouth, Lovaza® has a benign toxicity profile. If Lovaza® can be shown to have a clinically relevant anti-platelet effect, it may have a use to either replace or reduce the dose of more toxic anti-platelet agents.

The proposed biochemical mechanism for the anti-platelet effect of omega n3 fatty acids is based on modifications in platelet prostaglandin metabolism (27-31). Cellular membranes are primarily composed of phospholipids (PL). The backbone of PL's is glycerol. The glycerol hydroxyl groups in position 1 and 2 bind two fatty acid molecules through formation of ester bonds (31). The third hydroxyl binds the so-called head group, which may be choline, inositol, ethanolamine or serine. At least in the case of platelets the fatty acid at the C-2 position is often the unsaturated arachidonic fatty acid (an omega n6 fatty acid). When Lovaza® is ingested (an omega n3 fatty acid), the unsaturated fatty acid at the C2 position can be DHA or EPA. Several important differences result from this substitution including an important effect on platelet function. As part of the platelet activation process, phospholipase A2 clips the fatty acid at the C-2 position, either arachidonic acid or DHA/EPA (31). In the case of the platelet, the fatty acid is then metabolized through an enzyme called COX-1 to a thromboxane (32-35). When the fatty acid is arachidonic acid, thromboxane A2 is synthesized (TxA2). TxA2 is a very potent platelet activator and vasoconstrictor. In the case of DHA or EPA, a series 3 TxA3 is synthesized, a poor platelet activator and vasoconstrictor (32-35). Production of TxA3 underlies the potential anti-platelet effect of Lovaza®.

The second effect of DHA inclusion in PL's is a newly discovered alteration in the cell membrane structure. It is now well established that DHA promotes "lipid raft" formation in cellular membranes (36-38). These rafts, primarily composed of sphingomyelin and cholesterol, form the sites where some transmembrane proteins can be inserted into the membrane. These transmembrane proteins may be sites for ion channels or receptors that define important cellular functions and can be a means to activate cells. Thus, DHA's ability to promote raft formation may have a profound beneficial effect on platelet function.

Since it is the Lovaza®-alteration of the platelet membrane that leads to its clinical benefit, assays to determine how the lipid composition of the platelet membrane changes after ingestion of Lovaza® will be carried out. The concept of these experiments is fairly simple. A standard well-established 1H NMR method will be used to detect changes in the lipid composition of the platelet membrane as a function of the Lovaza® dose (39-41). From these experiments we will be able to prove that DHA or EPA from Lovaza® is actually directly incorporated into a platelet membrane