Drug – bio-affecting and body treating compositions – Preparations characterized by special physical form – Liposomes
Reexamination Certificate
1995-04-24
2001-03-20
Kishore, Gollamudi S. (Department: 1615)
Drug, bio-affecting and body treating compositions
Preparations characterized by special physical form
Liposomes
C264S004100, C264S004300, C436S069000, C514S021800
Reexamination Certificate
active
06203816
ABSTRACT:
BACKGROUND OF THE INVENTION
1. Field of the Invention
The present invention relates to a prothrombin time (PT) reagent using purified, reconstituted natural or recombinant human tissue factor (rTF). More particularly, the invention relates to the reconstitution of tissue factor into phospholipid vesicles to produce a tissue factor based PT reagent. Such a reagent allows specific monitoring of oral anticoagulant therapy and deficiencies in the extrinsic pathway of coagulation.
2. Related Art
In 1935, thromboplastin (procoagulant tissue factor) use was first described in a one stage PT test (Quick,
J. Biol. Chem.,
109:73-74, 1935). This test employed thromboplastin derived from mammalian tissue and a standard curve prepared with dilutions of pooled normal human plasma. The modern version of this test is easy to perform and can be automated. The PT test is the most commonly performed assay in the coagulation laboratory. Variants of this test have a number of uses (White, et al.,
Hemostasis and Thrombosis, Basic Principles and Clinical Practice
, Coleman, et al., eds., J.B. Lippencott Co., Philadephia, pp. 1048-1060, 1987). One use is to assess deficiencies in the extrinsic pathway of coagulation (Factors VII, X, V, and prothrombin). A second use is to monitor patients undergoing long term oral anticoagulant therapy for disorders such as recurrent venous thrombosis and cancer. A third use is to evaluate liver dysfunction.
The therapeutic range of anticoagulant therapy is based on the avoidance of bleeding and thrombolic complications. When monitoring oral anticoagulant therapy, as well as for a variety of other conditions by the PT test, an elongation of prothrombin time by a factor of 2 is most desirable for long term therapy (O'Reilly,
Hemostasis and Thrombosis, Basic Principles and Clinical Practice
, Coleman, et al., eds., J.B. Lippencott Co., Philadephia, pp. 1367-1372, 1987). This elongation factor is defined as the prothrombin ratio (PR) and is calculated by dividing the PT of a patient plasma by the PT of a pool of plasmas from normal individuals. A higher PR indicates a more sensitive PT reagent. The benefits of a more sensitive reagent for monitoring anticoagulation therapy is the use of lower doses of anticoagulant drug. These lower doses still provide adequate protection against thromboembolic disease while minimizing bleeding complications.
Several reagents for determining PT times are continually available. These include Thromborel S (Curtis Matheson Scientific, Inc., Yorba Linda, Calif.) and Simplastin (Organon Teknika Corp., Charlotte, N.C.). These reagents yield very different PT times for the same patient plasma with Thromborel S exhibiting longer times than Simplastin. Lower doses of anticoagulant drug are required to maintain extended PT times (high PR) when the PT times are monitored using Thromborel S instead of Simplastin.
A need exists for an even more sensitive tissue factor based PT reagent to monitor anticoagulant therapy and other conditions. The present invention provides just such a sensitive reagent with its highly desirable PR.
SUMMARY OF THE INVENTION
The present invention relates to liposome compositions and methods for their preparation with tissue factor incorporated in the lipid bilayer. The composition of the phospholipids in the liposomes has been adjusted to allow maximum coagulant activity and sensitivity to extrinsic coagulation factors of the incorporated tissue factor. In the preferred aspect, the method of the invention utilizes the zwitterionic detergent {3-[(3-cholamidopropyl)-dimethylammonio]-1-propanesulfonate} (CHAPS) to solubilize highly purified phospholipids. Tissue factor, also solubilized in CHAPS, is added with a carrier protein and the detergent removed by dialysis. Liposomes with tissue factor reconstituted into the lipid bilayer are formed spontaneously as the detergent concentration is lowered during dialysis.
REFERENCES:
patent: 3179567 (1965-04-01), Owren
patent: 4737276 (1988-04-01), Adamich
patent: 4761288 (1988-08-01), Mezei
patent: 4857319 (1989-08-01), Crowe et al.
patent: 4865984 (1989-09-01), Nemerson
patent: 4981685 (1991-01-01), Healey
patent: 5223427 (1993-06-01), Edgington
Anchordoguy, Cryobiology 24, pp. 324 1987.*
Deamer et al. Liposome prep. methods & mechanism, Chap. I, 1983, pp. 27-51.*
Journal of Parenteral Science and Technology, Wang et al., 42 (52) pp. 53-526.*
Neugebauer, J.,A Guide to the Properties and Uses of Detergents in Biology and Biochemistry, pp. 5-6, 27 and 35, (Calbiochem-Novabiochem, 1988).
Corvas International Inc.
Kishore Gollamudi S.
Lyon & Lyon LLP
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