We propose to identify an optimal short carbon chain lipid species to be used as a surrogate for membranes in a novel and improved blood coagulation factor assays that will improve rapid diagnosis of coagulation disorders. Blood coagulation protects the integrity of damaged blood vessels and is localized specifically to cells at a site of injury by exposure of a specific type of lipid, phosphatidylserine (PS) on activated platelet membranes. Blood coagulation assays are required to diagnose bleeding disorders such as hemophilia A and B and to provide physicians with information essential to surgery and to emergency medical situations. These assays replace the platelet membranes with a suspension of lipids that contain PS. While most of these assays are routine, a subset of patients have abnormal results. These abnormalities may reflect serious medical problems for the patient or they may reflect interferences in the in vitro assay. Currently these in vitro assays use a lipid membrane containing PS that mimics the material found in the body, meaning that the PS has a fatty acyl chains consisting of 16-22 carbons. But there are problems associated with using long-chain PS due to interference caused by agents that block or consume the lipid membrane. These agents are referred to as Lupus anti-coagulants, and their presence can result in misdiagnosis, which can greatly complicate patient care. There is a pressing need to develop a coagulation assay that is reproducible, reliable and that can be used for patients presenting Lupus anticoagulants. We have introduced and started to study a novel PS compound containing two saturated chains of only six carbons (C6PS), which offers the exciting possibility of overcoming the limitations of current coagulation assays. Because the acyl chain is very short, C6PS can exist in solution as single molecules rather than forming a membrane. Using this soluble PS molecule, we have been able under some circumstances to bypass the limitations caused by Lupus anticoagulants. However, under other assay conditions, C6PS can aggregate to form a micelle, at which point the advantages of C6PS as an assay reagent are lost. We propose to synthesize and test other short acyl chain PS molecules (generically called CNPS, where N is the number of carbons in the acyl chains and ranging from 2 to 5). We believe that shorter chain lengths will keep the PS molecules from forming micelles and so will serve better in assays than C6PS. Our goal is to identify the ideal CXPS compound by the end of Phase I. In Phase II, we will use this CXPS compound to develop a new set of assays for factors IX and VIII that have the potential to provide faster and more reliable tests for bleeding disorders than current clotting-based assays.
Public Health Relevance: Blood coagulation, or clotting, is the means by which the body protects the integrity of blood vessels, and it is very important that physicians know the extent and speed at which a patient's blood will clot. Clinical tests, or coagulation assays, are performed to characterize this information but are limited due to interferences present in the blood of some patients. We have studied a compound which may be used in these clinical assays and is not limited by interferences. We plan to identify and prove effective an optimal compound that can be used in support of better performing clinical coagulation assays. If effective, this will decrease misdiagnoses of bleeding disorders and will remove the need for expensive subsequent testing.
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