CA2117495A1 - Fibrin sealant delivery method - Google Patents
Fibrin sealant delivery methodInfo
- Publication number
- CA2117495A1 CA2117495A1 CA002117495A CA2117495A CA2117495A1 CA 2117495 A1 CA2117495 A1 CA 2117495A1 CA 002117495 A CA002117495 A CA 002117495A CA 2117495 A CA2117495 A CA 2117495A CA 2117495 A1 CA2117495 A1 CA 2117495A1
- Authority
- CA
- Canada
- Prior art keywords
- thrombin
- fibrin sealant
- clotting activity
- inhibitor
- thrombin clotting
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Abandoned
Links
- 108010080379 Fibrin Tissue Adhesive Proteins 0.000 title claims abstract description 46
- 238000002716 delivery method Methods 0.000 title description 2
- 108090000190 Thrombin Proteins 0.000 claims abstract description 81
- 229960004072 thrombin Drugs 0.000 claims abstract description 81
- 230000035602 clotting Effects 0.000 claims abstract description 58
- 108010049003 Fibrinogen Proteins 0.000 claims abstract description 43
- 102000008946 Fibrinogen Human genes 0.000 claims abstract description 43
- 229940012952 fibrinogen Drugs 0.000 claims abstract description 43
- 238000000034 method Methods 0.000 claims abstract description 42
- 229940012444 factor xiii Drugs 0.000 claims abstract description 32
- 108010071289 Factor XIII Proteins 0.000 claims abstract description 31
- 239000000243 solution Substances 0.000 claims description 27
- 239000003112 inhibitor Substances 0.000 claims description 24
- 230000000694 effects Effects 0.000 claims description 19
- 238000002360 preparation method Methods 0.000 claims description 13
- 230000015572 biosynthetic process Effects 0.000 claims description 11
- 239000000725 suspension Substances 0.000 claims description 10
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 claims description 9
- 239000011575 calcium Substances 0.000 claims description 9
- 229910052791 calcium Inorganic materials 0.000 claims description 9
- 210000001124 body fluid Anatomy 0.000 claims description 8
- 239000010839 body fluid Substances 0.000 claims description 8
- 239000000872 buffer Substances 0.000 claims description 8
- 239000002738 chelating agent Substances 0.000 claims description 6
- BHPQYMZQTOCNFJ-UHFFFAOYSA-N Calcium cation Chemical compound [Ca+2] BHPQYMZQTOCNFJ-UHFFFAOYSA-N 0.000 claims description 5
- 239000000337 buffer salt Substances 0.000 claims description 5
- 229910001424 calcium ion Inorganic materials 0.000 claims description 5
- 239000003795 chemical substances by application Substances 0.000 claims description 4
- 239000000463 material Substances 0.000 claims description 3
- 239000007864 aqueous solution Substances 0.000 claims description 2
- 159000000007 calcium salts Chemical class 0.000 claims description 2
- 230000003472 neutralizing effect Effects 0.000 claims description 2
- 239000000203 mixture Substances 0.000 abstract description 27
- 239000002244 precipitate Substances 0.000 abstract description 19
- 238000009472 formulation Methods 0.000 abstract 2
- 206010053567 Coagulopathies Diseases 0.000 description 20
- 206010052428 Wound Diseases 0.000 description 18
- 208000027418 Wounds and injury Diseases 0.000 description 18
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 10
- 239000007983 Tris buffer Substances 0.000 description 10
- 239000007921 spray Substances 0.000 description 9
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 9
- 239000000565 sealant Substances 0.000 description 8
- 239000001509 sodium citrate Substances 0.000 description 6
- NLJMYIDDQXHKNR-UHFFFAOYSA-K sodium citrate Chemical compound O.O.[Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NLJMYIDDQXHKNR-UHFFFAOYSA-K 0.000 description 6
- 238000001356 surgical procedure Methods 0.000 description 6
- KWPACVJPAFGBEQ-IKGGRYGDSA-N (2s)-1-[(2r)-2-amino-3-phenylpropanoyl]-n-[(3s)-1-chloro-6-(diaminomethylideneamino)-2-oxohexan-3-yl]pyrrolidine-2-carboxamide Chemical compound C([C@@H](N)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CCCNC(N)=N)C(=O)CCl)C1=CC=CC=C1 KWPACVJPAFGBEQ-IKGGRYGDSA-N 0.000 description 5
- SLXKOJJOQWFEFD-UHFFFAOYSA-N 6-aminohexanoic acid Chemical compound NCCCCCC(O)=O SLXKOJJOQWFEFD-UHFFFAOYSA-N 0.000 description 5
- 241000283690 Bos taurus Species 0.000 description 5
- BWGVNKXGVNDBDI-UHFFFAOYSA-N Fibrin monomer Chemical compound CNC(=O)CNC(=O)CN BWGVNKXGVNDBDI-UHFFFAOYSA-N 0.000 description 5
- 230000002378 acidificating effect Effects 0.000 description 5
- 230000003213 activating effect Effects 0.000 description 5
- 239000011780 sodium chloride Substances 0.000 description 5
- 108010073385 Fibrin Proteins 0.000 description 4
- 102000009123 Fibrin Human genes 0.000 description 4
- 229940088598 enzyme Drugs 0.000 description 4
- 229950003499 fibrin Drugs 0.000 description 4
- 230000005764 inhibitory process Effects 0.000 description 4
- 210000001519 tissue Anatomy 0.000 description 4
- 108090000790 Enzymes Proteins 0.000 description 3
- 102000004190 Enzymes Human genes 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 3
- 229940122388 Thrombin inhibitor Drugs 0.000 description 3
- 230000004913 activation Effects 0.000 description 3
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 3
- 239000011159 matrix material Substances 0.000 description 3
- 238000006116 polymerization reaction Methods 0.000 description 3
- 230000002028 premature Effects 0.000 description 3
- 239000002904 solvent Substances 0.000 description 3
- 239000003868 thrombin inhibitor Substances 0.000 description 3
- NTUPOKHATNSWCY-PMPSAXMXSA-N (2s)-2-[[(2s)-1-[(2r)-2-amino-3-phenylpropanoyl]pyrrolidine-2-carbonyl]amino]-5-(diaminomethylideneamino)pentanoic acid Chemical compound C([C@@H](N)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CCCN=C(N)N)C(O)=O)C1=CC=CC=C1 NTUPOKHATNSWCY-PMPSAXMXSA-N 0.000 description 2
- 239000004475 Arginine Substances 0.000 description 2
- 102000008186 Collagen Human genes 0.000 description 2
- 108010035532 Collagen Proteins 0.000 description 2
- MTCFGRXMJLQNBG-UHFFFAOYSA-N Serine Natural products OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 description 2
- 101000712605 Theromyzon tessulatum Theromin Proteins 0.000 description 2
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- 239000000443 aerosol Substances 0.000 description 2
- ODKSFYDXXFIFQN-UHFFFAOYSA-N arginine Natural products OC(=O)C(N)CCCNC(N)=N ODKSFYDXXFIFQN-UHFFFAOYSA-N 0.000 description 2
- 239000008280 blood Substances 0.000 description 2
- 210000004369 blood Anatomy 0.000 description 2
- 230000023555 blood coagulation Effects 0.000 description 2
- 210000000988 bone and bone Anatomy 0.000 description 2
- 210000000038 chest Anatomy 0.000 description 2
- -1 cinnamoyl Chemical class 0.000 description 2
- 239000007979 citrate buffer Substances 0.000 description 2
- ZYGHJZDHTFUPRJ-UHFFFAOYSA-N coumarin Chemical compound C1=CC=C2OC(=O)C=CC2=C1 ZYGHJZDHTFUPRJ-UHFFFAOYSA-N 0.000 description 2
- 239000013078 crystal Substances 0.000 description 2
- 238000004090 dissolution Methods 0.000 description 2
- 229940079593 drug Drugs 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 239000013038 irreversible inhibitor Substances 0.000 description 2
- 239000007788 liquid Substances 0.000 description 2
- 238000006386 neutralization reaction Methods 0.000 description 2
- 239000006174 pH buffer Substances 0.000 description 2
- 150000003839 salts Chemical class 0.000 description 2
- RSGFPIWWSCWCFJ-VAXZQHAWSA-N 2-hydroxypropane-1,2,3-tricarboxylic acid;(2r,3s,4r,5r)-2,3,4,5,6-pentahydroxyhexanal;phosphoric acid Chemical compound OP(O)(O)=O.OC[C@@H](O)[C@@H](O)[C@H](O)[C@@H](O)C=O.OC(=O)CC(O)(C(O)=O)CC(O)=O RSGFPIWWSCWCFJ-VAXZQHAWSA-N 0.000 description 1
- 229930024421 Adenine Natural products 0.000 description 1
- 102000015081 Blood Coagulation Factors Human genes 0.000 description 1
- 108010039209 Blood Coagulation Factors Proteins 0.000 description 1
- 206010006956 Calcium deficiency Diseases 0.000 description 1
- MNAFYRLSNBPFDC-UHFFFAOYSA-N Cl.C(N)(=N)C1=CC=C(C=C1)C(=C(C(=O)O)C)C1=C(C=C(C=C1)N(CC)CC)O Chemical compound Cl.C(N)(=N)C1=CC=C(C=C1)C(=C(C(=O)O)C)C1=C(C=C(C=C1)N(CC)CC)O MNAFYRLSNBPFDC-UHFFFAOYSA-N 0.000 description 1
- 108010000196 Factor XIIIa Proteins 0.000 description 1
- 208000032843 Hemorrhage Diseases 0.000 description 1
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 1
- 206010071229 Procedural haemorrhage Diseases 0.000 description 1
- 108010094028 Prothrombin Proteins 0.000 description 1
- 102100027378 Prothrombin Human genes 0.000 description 1
- 239000003929 acidic solution Substances 0.000 description 1
- 239000000654 additive Substances 0.000 description 1
- 229960000643 adenine Drugs 0.000 description 1
- 239000000853 adhesive Substances 0.000 description 1
- 230000001070 adhesive effect Effects 0.000 description 1
- 229960002684 aminocaproic acid Drugs 0.000 description 1
- 239000003242 anti bacterial agent Substances 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- 230000000740 bleeding effect Effects 0.000 description 1
- 239000003114 blood coagulation factor Substances 0.000 description 1
- 239000004202 carbamide Substances 0.000 description 1
- 238000013130 cardiovascular surgery Methods 0.000 description 1
- 238000004113 cell culture Methods 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 229920001436 collagen Polymers 0.000 description 1
- 238000002316 cosmetic surgery Methods 0.000 description 1
- 229960000956 coumarin Drugs 0.000 description 1
- 235000001671 coumarin Nutrition 0.000 description 1
- 150000004775 coumarins Chemical class 0.000 description 1
- 239000003995 emulsifying agent Substances 0.000 description 1
- 239000002532 enzyme inhibitor Substances 0.000 description 1
- 125000001495 ethyl group Chemical group [H]C([H])([H])C([H])([H])* 0.000 description 1
- 108010073651 fibrinmonomer Proteins 0.000 description 1
- 230000008014 freezing Effects 0.000 description 1
- 238000007710 freezing Methods 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 239000008187 granular material Substances 0.000 description 1
- 239000003102 growth factor Substances 0.000 description 1
- 230000023597 hemostasis Effects 0.000 description 1
- 238000005286 illumination Methods 0.000 description 1
- 230000000415 inactivating effect Effects 0.000 description 1
- 230000002779 inactivation Effects 0.000 description 1
- 230000007774 longterm Effects 0.000 description 1
- 230000000399 orthopedic effect Effects 0.000 description 1
- 238000006349 photocyclization reaction Methods 0.000 description 1
- 238000007699 photoisomerization reaction Methods 0.000 description 1
- 239000003504 photosensitizing agent Substances 0.000 description 1
- 235000020004 porter Nutrition 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 229940039716 prothrombin Drugs 0.000 description 1
- 230000007420 reactivation Effects 0.000 description 1
- 238000007789 sealing Methods 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 239000011550 stock solution Substances 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 238000010257 thawing Methods 0.000 description 1
- 229940033618 tisseel Drugs 0.000 description 1
- 238000004448 titration Methods 0.000 description 1
- 230000002792 vascular Effects 0.000 description 1
- 238000003260 vortexing Methods 0.000 description 1
- 230000029663 wound healing Effects 0.000 description 1
- NWONKYPBYAMBJT-UHFFFAOYSA-L zinc sulfate Chemical compound [Zn+2].[O-]S([O-])(=O)=O NWONKYPBYAMBJT-UHFFFAOYSA-L 0.000 description 1
- 229960001763 zinc sulfate Drugs 0.000 description 1
- 229910000368 zinc sulfate Inorganic materials 0.000 description 1
Classifications
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61B—DIAGNOSIS; SURGERY; IDENTIFICATION
- A61B17/00—Surgical instruments, devices or methods, e.g. tourniquets
- A61B17/00491—Surgical glue applicators
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L24/00—Surgical adhesives or cements; Adhesives for colostomy devices
- A61L24/04—Surgical adhesives or cements; Adhesives for colostomy devices containing macromolecular materials
- A61L24/10—Polypeptides; Proteins
- A61L24/106—Fibrin; Fibrinogen
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P7/00—Drugs for disorders of the blood or the extracellular fluid
- A61P7/04—Antihaemorrhagics; Procoagulants; Haemostatic agents; Antifibrinolytic agents
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P9/00—Drugs for disorders of the cardiovascular system
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61B—DIAGNOSIS; SURGERY; IDENTIFICATION
- A61B17/00—Surgical instruments, devices or methods, e.g. tourniquets
- A61B17/00491—Surgical glue applicators
- A61B2017/00495—Surgical glue applicators for two-component glue
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61B—DIAGNOSIS; SURGERY; IDENTIFICATION
- A61B17/00—Surgical instruments, devices or methods, e.g. tourniquets
- A61B17/22—Implements for squeezing-off ulcers or the like on the inside of inner organs of the body; Implements for scraping-out cavities of body organs, e.g. bones; Calculus removers; Calculus smashing apparatus; Apparatus for removing obstructions in blood vessels, not otherwise provided for
- A61B2017/22082—Implements for squeezing-off ulcers or the like on the inside of inner organs of the body; Implements for scraping-out cavities of body organs, e.g. bones; Calculus removers; Calculus smashing apparatus; Apparatus for removing obstructions in blood vessels, not otherwise provided for after introduction of a substance
Abstract
The present invention relates to a method for the formulation of fibrin sealant in a single delivery system. The method involves mixing a fibrinogen/Factor XIII precipitate solution with thrombin under conditions such that thrombin clotting activity is inhibited and said mixture is applied to a body site under conditions which activate the thrombin to convert fibrinogen into fibrin sealant. A single device, syringe or container, can be used to apply the fibrin sealant formulation.
Description
W O 93/19805 ~A ~ 7~ 9~ PCT/~S93/02228 FIBRIN SEALANT DELIVERY METHOD
8A~K~UN~ OF THE INVENTION
This is a continuation-in-part application of Serial No. 07/460,379 filed on January 3, 1990, which is hereby in-~o.~ ted herein in its entirity.
Technical Field The present invention relates to a method of fibrin sealant preparation and delivery, which permits use of a single delivery device. The method may be used for autologous, single-donor, pooled-donor or cell culture-derived fibrin sealant for various human and veterinary surgical procedures.
The invention further relates to a kit suitable for use in such a method.
R~r~TJNn INFORMATION
The blood coagulation system is a complex series of proteins and factors which are activated sequentially to produce a fibrin gel or clot. In the final stages of the process, fibrinogen is cleaved by thrombin to generate fibrin monomer, which rapidly polymerizes and is cross-linked by activated Factor XIII to form a fibrin matrix.
Preparations of human coagulation factors, including fibrinogen and thrombin, have been used extensively in surgery over the last ten years (Schlag et al (eds), Fibrin Sealant in Operative Medicine, vol 1-7, Springer-Verlag, H~ lh~rg).
These biological fibrin sealants promote hemostasis and wound healing by sealing leakage from tissues, sutures, staples, and prostheses, and are W O 93/19805 C A 2 1 1 7 4 9 5 P(~r/US93/02228 particularly useful during open heart surgery in heparinized patients. The sealants also have use as an adhesive for the bonding of tissues and they reduce the amount of blood reguired for transfusions by controlling intraoperative bleeding. Their effectiveness is reflected in the extensive range of surgical applications for which they have been used, inr~U~ing cardiovascular surgery, plastic surgery, orthopedics, urology, obstetrics and gynecology, dentistry, maxillofacial and ophthalmic surgery.
Fibrin sealant p~u~Ls prepared from pooled human plasma fibrinogen/Factor XIII are available commercially in Europe (Tissucol/Tisseel, Immuno AG, Vienna, Austria and Beriplast P, Hoechst, West Germany) but such products have not received U.S. Food and Drug Administration approval. As an alternative, some hospitals are preparing fibrin sealant in-house using the patient's own blood (autologous) or single-donor (homologous) plasma as a source of fibrinogen and Factor XIII.
The plasma fibrinogen/Factor XIII
~ ;L of fibrin sealant is typically prepared by freezing plasma at a t~ LULe below -20-C
overnight, slowly thawing the material at 0-4 C, centrifuging, and transferring the cryoprecipitate to a syringe or spray container (Dresdale et al, Ann. Thorac. Surg. 40:385 1985: and U.S. Patent 4,627,879). The thrombin , -nt, usually purified from bovine plasma, can be obtained commercially and is typically prepared in a separate syringe or spray container. In use, the two solutions are delivered simult~n~oncly or alternately to generate fibrin sealant at the site W093/1980~ C A 2 1 1 7 4 q 5 PCT/US93/02228 of the wound; alternatively, the sealant is applied to a collagen matrix (e.g. Gelfoam or Avitene) and then pressed against the site (Lupinetti et al, J.
Thorac. Cardiovasc. Surg. 90:502 1985; and U.S.
Patent 4,453,939).
The use of fibrin sealant in surgery has been limited by problems associated with mixing and delivery of the sealant to the wound. Generation of fibrin sealant at the wound site is currently achieved using a two syringe or spray container system to prevent pl~ LuLe mixing and clotting of the ~ ~s. Such two syringe systems are, however, unsatisfactory due to the awkwardness of filling and manipulating the delivery devices at the wound site. In addition, the syringe system is A~_ -nied by problems of inA~ Ate mixing of the two solutions, resulting in the formation of a weak clot. Alternatively, the two syringes can be placed into a holder designed such that the solutions are permitted to mix before entering the needle (V.S.
Patents 4,735,616, 4,359,049, and 4,631,05~). The mixing chamber directs the flow of the separate ~ Ls through two narrow ~hAnn~lc and forces mixing at the top of the outflow needle. Although the ~LlellyLh of the clot obtained using this method is reproducible, the needle frequently clogs and must repeatedly be replaced.
In view of the problems inherent in the methodologies currently available for delivering fibrin sealant, the need for a simple, reproducible technique is clear. Such a technique must be convenient to use and must result in the formation, at a specific site, of a clot of appropriate W O 93/19805 C A 2 1 1 7 4 9 5 P~r/US93/02228 strength. Such a delivery technique is provided by the invention disclosed herein.
S~MARY OF THE INVENTION
It is a general object of the present S invention to provide a method of forming a fibrin sealant from blood coagulation Ls that overcomes the problems associated with methods known in the art.
It is a specific object of the invention to provide a method of delivering fibrin sealant to a wound site, in which method a fibrinogen/Factor XIII-enriched precipitate (or a fibrinogen/Factor XIII mixture) and thrombin are mixed together under conditions such that clotting is prever.ted until such time as sealant formation is desired.
It is another object of the present invention to provide means of reversibly inactivating thrombin and s~hce~ .Lly restoring activity which can be used to prevent pl Lul_ clot formation.
It is a further object of the invention to provide a kit suitable for use in the above-described method.
A more complete appreciation of the present invention and the advantages thereof will be readily understood by one skilled in the art from a reading of the description that follows.
In one : J~;- L, the present invention relates to a method of effecting the ~ormation of fibrin sealant at a body site. The method comprises: i) mixing, in a container means, an w093/l9805 C A 2 1 1 7 4 ~ 5 PCT/~S93/02228 aqueous solution comprising fibrinogen, Factor XIII
and mature thrombin under conditions such that ~ thrombin clotting activity is inhibited; and ii) applying a preparation resulting from step (i) to the body site under conditions such that thrombin clotting activity is restored and the fibrin sealant is formed.
In another ~ , the present invention relates to a method of effecting the formation of fibrin sealant at a body site comprising: i) forming a suspension comprising a first phase which comprises fibrinogen and Factor XIII and a second phase which comprises thrombin, and ii) applying the suspension to the body site under conditions such that mixing of the fibrinogen, Factor XIII and thrombin is effected so that the fibrin sealant is formed.
In a further : '-'i- ~, the present invention relates to a kit for use in the preparation of a fibrin sealant. The kit includes an applicator comprising: i) a container means having ~i~posed therein a solution comprising fibrinogen, Factor XIII and mature thrombin; and ii) an outlet means operably connected to said container means.
nTTATTT~n DESCRIPTION OF INVENTION
The present invention relates to a method of delivering the ents of a fibrin sealant (mature thrombin (as opposed to prothrombin) and the plasma-derived fibrinogen/Factor XIII precipitate) to a body site in a manner such that clot formation W O 93/19805 CA21 ~ 5 P~r/~S93/02228 is effected, and to a kit suitable for use in such a method. (The term "body site" as used herein includes the tissue in the area of a wound or incision as well as implantable tissues or ~s to be inserted into the area, e.g., vascular prostheses, bone or collAg~n pads.) In the description that follows, it will be appreciated that a combination of isolated forms of fibrinogen and Factor XIII can be used in place of the plasma-derived precipitate.
In the method of the present invention, a fibrinogen/Factor XIII-enriched precipitate and mature thrombin are mixed together under conditions such that thrombin and/or Factor XIII are/is inactivated (or under conditions such that thrombin is present in an active form but is rendered unavailable, as in the calcium depletion '-'i described below) and clotting thereby prevented.
The mixture is then delivered to the body site under conditions such that the enzyme activity is restored (or thrombin availability restored).
In one '~ , the mixture of thrombin and fibrinogen/FaCtor XIII precipitate is prepared in a low pH buffer (the clotting of fibrinogen by ~~ in being inhibited by low pH (less than 5.5)).
In this : ' 'i- ~, thrombin activity is restored and clotting rapidly initiated upon neutralization of the mixture with a phArr~-~utically acceptable buffer, or alternatively, upon contact of the mixture with the patient's own body fluids. In this -'i- ~, the fibrinogen/Factor XIII precipitate can be prepared at a low pH or, alternatively, a low pH buffer can be used to dissolve the plasma W O 93/19805 C A 2 1 1 7 4 9 5 P~r/~S93/02228 precipitate and the lyophilized thrombin. In either case, the mixture can be transferred to a delivery container (such as a spray bottle or syringe) and applied to the body site directly, if conditions are ~ 5 such that the patient's body fluids are sufficient to increase the pH to a point where clotting occurs.
Where conditions are such that the patient's body fluids are not sufficient to raise the pH of the precipitate/thrombin mixture to a point where thrombin activity is restored, a delivery device can be used that is designed such that, as the acidic mixture passes out of the device, it is contacted with buffer salts coated on an interior portion of the device. The buffer salts are selected such that when contact is made with the acidic mixture, dissolution occurs with the result that the pH is raised to a point where clotting takes place. For example, a syringe can be used as the delivery device (applicator), where the syringe is fitted with a ~;cpoc~hle tip, the interior surface of which is coated with appropriate buffer salts. As the acidic mixture passes through the coated tip, the buffer (in the form, for example, of crystals or a gel) neutralizes the acidic mixture, thus restoring thrombin activity and effecting the formation of a clot at the desired site. Should clot formation occur in the tip, the tip can simply be removed and a new coated tip attached.
In another o~lir ---t, the fibrinogen and Factor XIII precipitate/thrombin mixture can be ~ prepared in a buffer that is depleted of calcium.
Rapid clot formation requires the presence of calcium ions: thus, if the calcium is removed, W O 93/19805 C A ~ 14 ~ 5 PC~r/~S93/02228 fibrin polymerization is inhibited (see Carr et al Biochem J. (1986) 239:513; KAminck; et al J. Biol.
Chem (1983) 258:10530: Kanaide et al (1982) 13:229).
Calcium chelators (. -c such as sodium citrate or ethyl~nP~iAminetetraacetic acid, which tightly bind calcium and make it inAcc~ccible) can be added to the solution used to precipitate the fibrinogen and Factor XIII and/or the dissolving buffer. To restore activity, the container (for example, a syringe) can be attached to a ~icpocAhle sterile tip, the interior surface of which is coated internally with sufficient calcium salt to saturate the chelator. As the free calcium ~u..~--LL~tion increases upon passage of the mixture through the tip, clotting is effected at the body site.
In a further ~ L, the clotting activity of thrombin, in the precipitate/thrombin mixture, can be inhibited using a photosensitive inhibitor. For example, light sensitive ri yl derivatives can be used to inactivate thrombin, at room temperature in the absence of light, for more than 26 hours (Turner et al J. Am. Chem. Soc. 109:
1274-1275 (1987); Turner et al J. Am. Chem. Soc.
110: 244-250 (1988)). These same thrombin inhibitor complexes can generate active thrombin within 1-2 seconds of irradiation (low intensity). These inhibitors are known to form acyl-enzyme complexes involving the active site serine hydroxyl (SER 195).
Upon irradiation, the cinnamoyl derivative undergoes photoisomerization to release coumarin and regenerate the active serine hydroxyl. Since coumarin derivatives are not good thrombin inhibitors, this photocyclization reaction WO93/19805 C~2 i 1 7495 i PCT/US93/0222X
effectively removes inhibitor from the enzyme solution. Thus, a solution of the fibrinogen/Factor XIII-enriched precipitate can be mixed with lyoph;li7~ inhibitor:thrombin complex in a dark environment (such as an opaque or colored syringe or container) and delivered to the wound site.
Activation of the enzyme and thus clot formation occurs upon delivery to the wound due to the ~X~O~UL~ of the solution to normal room light.
Alternatively, activation can be controlled by a light source, for example, one built directly into the applicator, so that variations in lighting conditions will not result in variable clotting times.
Preferably, the clotting activity of thrombin, in the precipitate/thrombin mixture is inhibited by the use of a photosensitive inhibitor in combination with an irreversible inhibitor. Such "doubly-inhibited" thrombin offers several advantages including increased stability of the double inhibitor:thrombin complex and fibrinogen/Factor XIII precipitate which reduces premature clot formation during long term storage as well as during use. The doubly inhibited thrombin also provides for increased control of clot activation, increased clot strength and increased time for mixing with additives (such as, for example, bone granules, antibiotics or growth factors).
For example, thrombin activity can be inhibited by the use of the photosensitive inhibitor 4-amidino-phenyl-2-hydroxy-4-diethylamino-alpha-methylcinnamate hydrochloride, "I-l" and the W o 93/19805 C A 2 i 1 14 9 5 P(~r/~S93/02228 irreversible inhibitor D-phenylalanyl-L-prolyl-L-arginine chloromethyl ketone, "PPACK". I-l inhibits approximately 99% of the thrombin activity, however, the residual 1% activity can not be reduced by additional I-1. This residual activity, which is sufficient to cause clotting after 15-20 minutes in the absence of light, can be reduced or titrated by the addition of the second inhibitor ("PPACK" in the present example). In the absence of activating irradiation (light of approximately 360 nm as provided by a Caulk W Polymerization Unit), the doubly inhibited thrombin solution is subse~ue.-Lly added to the fibrinogen c L and stored frozen, lyophilized or used within hours. Fibrin sealant prepared from this double inhibitor:Lh , I in complex and fibrinogen/Factor XIII precipitate can be maintained in the dark without clotting for over 2 hours. When the fibrin sealant is needed, the mixture can be dispensed onto the wound site or appropriate delivery container (such as a spray bottle or syringe) and activated by light to clot in less than 1-2 minutes.
~ h~ . ' ;n inhibited by the combination of such inhibitors may be useful for other applications. For example, the double inhibitor:thrombin complex can be used in situations requiring careful control of clotting activity or when thrombin is used alone to stop bleeding. In addition, the inhibited thrombin can be used to localize clotting activity (for example, eye surgery). The inhibited thrombin may also be used with other components or in other forms (for W O 93/19805 C A 2 1 1 7 4 9 5 P(~r/US93/02228 example, with gel matrix or on solid support) to improve hAn~l ing of delivery to the wound site.
In yet another ' ~;- L, premature clot formation can be prevented prior to delivery of the fibrinogen and Factor XIII/thrombin mixture by physically separating the thrombin from the fibrinogen/Factor XIII precipitate. In this : '-'; ~, physical separation is effected using a two-phase system. Liquids suitable for use in this : ' 'i- t are non-miscible and readily separable into two phases. The two phases are mixed into a suspension before each application and delivered to the wound. Where conditions are such that the patient's body fluids extract the soluble ~
of the nonaqueous phase, mixing occurs at the body ~site and clotting is thus initiated. If conditions will not elicit proper mixing of - ts, a delivery device can be used that is designed such that, as the suspension passes out of the device, it is contacted with a solubilizing agent coated on an interior portion of the device~ The solubilizing agent is selected such that when contact is made with the suspension, dissolution occurs with the result that mixing occurs to a degree where clotting takes place. For example, a syringe is fitted with a ~;epnc~hle tip, the interior surface of which is coated with an appropriate phase transfer agent(s).
As the suspension passes through the coated tip, the phase transfer agent (in the form, for example, of crystals) assists in the mixing process, thus allowing clot formation. Should clot formation occur in the tip, the tip can simply be removed and a new coated tip attached.
W O 93/19805 ~ A 2 i 1 7 4 ~ 5 P~r/~S93/02228 The present invention also relates to a kit suitable for use in the above de~e~ibed method of delivering fibrin sealant _ ~s to a wound site. In a preferred A ' ~ the kit includes an applicator designed so as to permit mixing of the fibrinogen/Factor XIII precipitate and thrombin in a single system. The applicator can be one that permits the application at the body site of, for example, a film, or a thin line of the , ~ s of fibrin sealant. Alternatively, a pump or aerosol spray applicator can be used.
As suggested above, the applicator can, for example, take the form of a glass or plastic syringe with ~;~pos~hle tips. The shape of the tip will determine the form in which the ,~ ~s are delivered. A tip with a flat, broad end can be used to deliver a thin wide streak of fibrin sealant whereas a narrow tubular end can be used to deliver a round thread of sealant. Applying pleS~ule to force the mixture through a tip constricted with, for example, a mesh screen can be used to produce a spray, resulting in a fine glaze of fibrin sealant.
In another : ' 'i- L, particularly suitable for use with the above-described photosensitive thrombin inhibitor, the applicator can take the form of a pump or aerosol spray device having a built-in light source situated such that, as the sealant components exit the device, they are irradiated with the light.
The wavelength of light used would depend on the photosensitizer.
The kit can be structured so as to include individual storage containers for the separate fibrin sealant ~ , ents. The kit can also include W O 93/19805 C A 2 1 1 7 4 9 5 ~ PC~r/US93/02228 one or more other storage containers ~icpose~ within which are any nPcPCsAry reagents, including solvents, buffers, etc.
The present invention will be understood in greater detail by reference to the following non-limiting Examples.
EXAMPLES
ExamPle 1 A precipitate containing fibrinogen and Factor XIII was prepared as follows:
Four hundred fifty microliters of a stock 1 M zinc sulfate solution were added to 5 ml of anticoagulated (citrate phosphate dextrose adenine (CPDA-1)) human plasma. The solution was mixed well without vortexing and centrifuged at 2,000 to 9,000 g for 5 minutes. The supernatant was decanted and discarded.
Inhibition of clotting and reactivation was achieved by any of the following methods:
A. Acid Tnh;bition - Lyophilized bovine thrombin was dissolved in citrate buffer (500 mM
citric acid, 150 mM NaCl, and 20 mM EACA, pH 4.5) to a final conce,l~.Gtion of 100 U/ml. Precipitated fibrinogen was dissolved in Tris buffer (50 mM Tris, 250 mM sodium citrate, 150 mM sodium chloride, 50 mM
Arginine (Arg), and 20 mM ~-amino-caproic acid (EACA), pH 7.4) to a concentration of approximately 15.0 mg/ml. This fibrinogen stock was then diluted 25-fold in citrate buffer. The clotting time for 200 microliters of this fibrinogen solution plus 100 microliters thrombin ~YceP~Pd 90 seconds in a Becton W O 93/19805 C A 2 1 1 7 4 9 5 PC~r/~S93/02228 Dic-lc;n con BBL Fibrosystem fibrometer under standard conditions indicating no clot formation. Addition of 70 microliters of lN sodium hydroxide resulted in clot formation in 3.8 seconds (average of 10 samples).
The following p-~cedu.es can be used for application to a wound site:
Where body fluids are sufficient to neutralize the acidic mixture of precipitate and thrombin, the mixture can be applied directly to the wound site. Alternatively, the delivery device can be connected to dicroc~hle tips coated internally with a neutralizing salt or gel (e.g. Tris).
Neutralization of the acidic solution by the buffer salts activatss thrombin and restores clotting activity.
B. Chelator Inhibition - Precipitated fibrinogen and lyophili7sd bovine thrombin were dissolved in Tris buffer (50 mM Tris, 250 mM sodium citrate, 150 mM sodium chloride, 50 mM Arg, and 20 mM EACA, p~ 7.4) to a c~"~e..LL~tion of approximately 15.0 mg/ml and 100 U/ml, respectively.
The fibrinogen stock solution was then diluted 25-fold in Tris buffer containing 500 mM sodium citrate. The clotting time for 200 microliters of this fibrinogen solution plus 100 microliters thrombin ~Ycee~d 90 seconds in a Becton Dickinson BBL fibrosystem fibrometer under standard conditions. Addition of 50 microliters of a lM
CaCl, solution resulted in clot formation in 1.8 seconds (average of 10 samples).
W O 93/19805 C A 2 i 1 / 4 9 5 PC~r/~S93/02228 The following procedure can be used for application to a wound site:
- The delivery device is connected to ~i~pn~hle tips coated internally with a calcium - 5 salt or gel. As the mixture passes through the tip, the molar excess of calcium saturates the chelator and clotting is thereby promoted.
C. Photosensitive Inhibition - In the absence of light, a 5 to 20-fold excess of 4-10 amidino phe~l-2-hydroxy-4-diethylamino-alpha-methylcinnamate hydrochloride (Porter et al, J.
Amer. Chem. Soc. 111:7616 (1989)) was added to thrombin in buffer (approximately 100 U/ml in 50 mM
Tris, 250 mM sodium chloride, 250 mM sodium citrate, 20 mM EACA, 50 mM arginine (or urea), pH 7.4, final methanol cu.,~e--Llation <10%). The inhibition was allowed to proceed for at least 1 hour at room t~ a LUL ~.
A minimal quantity of this solution was used to dissolve the precipitated fibrinogen/Factor XIII. This sealant required approximately 2 to 3 minutes illumination under standard operating lights to clot completely, whereas a sample mixture kept in the dark did not clot after 90 min.
The following pLuceduL~s can be used for application to a wound site:
The photosensitive inhibitor-thrombin complex can be mixed with the precipitated fibrinogen/Factor XIII in a colored delivery device that does not transmit light of the activating wavelengths. Delivery of the mixture to an illuminated wound site results in clot formation.
W O 93/19805 C A 2 1 1 7 4 q 5 PC~r/US93/02228 D. Two Phase Suspension - Lyophilized bovine thrombin is dissolved in an emulsifying agent to a final c~"cel,L-~tion of about 100 U/ml.
Precipitated fibrinogen/Factor XIII is dissolved in a minimal volume of buffer (50 mM Tris, 150 mN
sodium chloride, 250 mM sodium citrate, 20 mM EACA, 50mM Arg, pH 7.4). A sllqppn~i~n of the ; i~c;hle liquids is formed. On a wound surface, body fluids may be sufficient to dissolve both components and promote proper mixing and clot formation.
ExamDle 2 A fibrin sealant was prepared using the two enzyme inhibitors as follows. All inactivation and transfer steps were performed in the absence of activating light (366 nm), using a dark room illuminated by a red light.
Thrombin was inactivated sequentially using 1) the light sensitive inhibitor 4-Am;~in~ph~nyl-2 }Iy~lv~ 4-diethylamino-alpha-methylcinnamate hydrochloride and 2) D-Phenylalanyl-L-prolyl-L-arginine Chloromethyl Ketone, PPACK.
Bovine thrombin (0.1 ml of 1,000 units/ml Armour Thrombinar) was mixed with 0.8 ml of 50 mM tris buffer, pH 7.3, and 0.1 ml of 31.2~ ~g/ml inhibitor in 2.5% ethanol, 50 mM tris, pH 7.3. The solution was incubated at room temperature for 2 hours.
Residual thrombin activity of the thrombin:inhibitor-l complex (approximately 1%
original activity) was inactivated by titration with PPACK until residual activity was such that the resulting fibrin sealant could be maintained in the W O 93/19805 C A 2 i 1 7 4 q 5 PC~r/~S93/02228 dark for 2-4 hours without premature clotting. The above solution (0.1 ml) was mixed with 0.01 ml of ~ 2.5 ~g/ml PPACK at room temperature and thrombin activity of the thrombin:inhibitor-2 was negligible ~ 5 within minutes.
Fibrin sealant was prepared by mixing 1 ml fibrinogen stock (e.g. single unit human cryoprecipitate) with 0.1 ml of the thrombin:inhibitor-2 in the absence of activating light (e.g. opaque container) and transferred to an amber syringe or spray container for delivery.
Clotting activity was easily controlled using light. In the absence of activating light, the mixture was stable for at least 2 hours at room temperature before clotting occurred. A 20 second pulse of light (that is, light of 360 nm, by Caulk MAX polymerization unit) at a distance of less than 5 cm from the dispensed solution resulted in clotting within seconds.
* * * *
The entire contents of each of the references cited above are hereby incorporated by reference.
While the present invention has been described in some detail for purposes of clarity and understanding, it will be clear to one skilled in the art from a reading of this disclosure that various changes can be made in form and detail without departing from the true scope of the invention.
8A~K~UN~ OF THE INVENTION
This is a continuation-in-part application of Serial No. 07/460,379 filed on January 3, 1990, which is hereby in-~o.~ ted herein in its entirity.
Technical Field The present invention relates to a method of fibrin sealant preparation and delivery, which permits use of a single delivery device. The method may be used for autologous, single-donor, pooled-donor or cell culture-derived fibrin sealant for various human and veterinary surgical procedures.
The invention further relates to a kit suitable for use in such a method.
R~r~TJNn INFORMATION
The blood coagulation system is a complex series of proteins and factors which are activated sequentially to produce a fibrin gel or clot. In the final stages of the process, fibrinogen is cleaved by thrombin to generate fibrin monomer, which rapidly polymerizes and is cross-linked by activated Factor XIII to form a fibrin matrix.
Preparations of human coagulation factors, including fibrinogen and thrombin, have been used extensively in surgery over the last ten years (Schlag et al (eds), Fibrin Sealant in Operative Medicine, vol 1-7, Springer-Verlag, H~ lh~rg).
These biological fibrin sealants promote hemostasis and wound healing by sealing leakage from tissues, sutures, staples, and prostheses, and are W O 93/19805 C A 2 1 1 7 4 9 5 P(~r/US93/02228 particularly useful during open heart surgery in heparinized patients. The sealants also have use as an adhesive for the bonding of tissues and they reduce the amount of blood reguired for transfusions by controlling intraoperative bleeding. Their effectiveness is reflected in the extensive range of surgical applications for which they have been used, inr~U~ing cardiovascular surgery, plastic surgery, orthopedics, urology, obstetrics and gynecology, dentistry, maxillofacial and ophthalmic surgery.
Fibrin sealant p~u~Ls prepared from pooled human plasma fibrinogen/Factor XIII are available commercially in Europe (Tissucol/Tisseel, Immuno AG, Vienna, Austria and Beriplast P, Hoechst, West Germany) but such products have not received U.S. Food and Drug Administration approval. As an alternative, some hospitals are preparing fibrin sealant in-house using the patient's own blood (autologous) or single-donor (homologous) plasma as a source of fibrinogen and Factor XIII.
The plasma fibrinogen/Factor XIII
~ ;L of fibrin sealant is typically prepared by freezing plasma at a t~ LULe below -20-C
overnight, slowly thawing the material at 0-4 C, centrifuging, and transferring the cryoprecipitate to a syringe or spray container (Dresdale et al, Ann. Thorac. Surg. 40:385 1985: and U.S. Patent 4,627,879). The thrombin , -nt, usually purified from bovine plasma, can be obtained commercially and is typically prepared in a separate syringe or spray container. In use, the two solutions are delivered simult~n~oncly or alternately to generate fibrin sealant at the site W093/1980~ C A 2 1 1 7 4 q 5 PCT/US93/02228 of the wound; alternatively, the sealant is applied to a collagen matrix (e.g. Gelfoam or Avitene) and then pressed against the site (Lupinetti et al, J.
Thorac. Cardiovasc. Surg. 90:502 1985; and U.S.
Patent 4,453,939).
The use of fibrin sealant in surgery has been limited by problems associated with mixing and delivery of the sealant to the wound. Generation of fibrin sealant at the wound site is currently achieved using a two syringe or spray container system to prevent pl~ LuLe mixing and clotting of the ~ ~s. Such two syringe systems are, however, unsatisfactory due to the awkwardness of filling and manipulating the delivery devices at the wound site. In addition, the syringe system is A~_ -nied by problems of inA~ Ate mixing of the two solutions, resulting in the formation of a weak clot. Alternatively, the two syringes can be placed into a holder designed such that the solutions are permitted to mix before entering the needle (V.S.
Patents 4,735,616, 4,359,049, and 4,631,05~). The mixing chamber directs the flow of the separate ~ Ls through two narrow ~hAnn~lc and forces mixing at the top of the outflow needle. Although the ~LlellyLh of the clot obtained using this method is reproducible, the needle frequently clogs and must repeatedly be replaced.
In view of the problems inherent in the methodologies currently available for delivering fibrin sealant, the need for a simple, reproducible technique is clear. Such a technique must be convenient to use and must result in the formation, at a specific site, of a clot of appropriate W O 93/19805 C A 2 1 1 7 4 9 5 P~r/US93/02228 strength. Such a delivery technique is provided by the invention disclosed herein.
S~MARY OF THE INVENTION
It is a general object of the present S invention to provide a method of forming a fibrin sealant from blood coagulation Ls that overcomes the problems associated with methods known in the art.
It is a specific object of the invention to provide a method of delivering fibrin sealant to a wound site, in which method a fibrinogen/Factor XIII-enriched precipitate (or a fibrinogen/Factor XIII mixture) and thrombin are mixed together under conditions such that clotting is prever.ted until such time as sealant formation is desired.
It is another object of the present invention to provide means of reversibly inactivating thrombin and s~hce~ .Lly restoring activity which can be used to prevent pl Lul_ clot formation.
It is a further object of the invention to provide a kit suitable for use in the above-described method.
A more complete appreciation of the present invention and the advantages thereof will be readily understood by one skilled in the art from a reading of the description that follows.
In one : J~;- L, the present invention relates to a method of effecting the ~ormation of fibrin sealant at a body site. The method comprises: i) mixing, in a container means, an w093/l9805 C A 2 1 1 7 4 ~ 5 PCT/~S93/02228 aqueous solution comprising fibrinogen, Factor XIII
and mature thrombin under conditions such that ~ thrombin clotting activity is inhibited; and ii) applying a preparation resulting from step (i) to the body site under conditions such that thrombin clotting activity is restored and the fibrin sealant is formed.
In another ~ , the present invention relates to a method of effecting the formation of fibrin sealant at a body site comprising: i) forming a suspension comprising a first phase which comprises fibrinogen and Factor XIII and a second phase which comprises thrombin, and ii) applying the suspension to the body site under conditions such that mixing of the fibrinogen, Factor XIII and thrombin is effected so that the fibrin sealant is formed.
In a further : '-'i- ~, the present invention relates to a kit for use in the preparation of a fibrin sealant. The kit includes an applicator comprising: i) a container means having ~i~posed therein a solution comprising fibrinogen, Factor XIII and mature thrombin; and ii) an outlet means operably connected to said container means.
nTTATTT~n DESCRIPTION OF INVENTION
The present invention relates to a method of delivering the ents of a fibrin sealant (mature thrombin (as opposed to prothrombin) and the plasma-derived fibrinogen/Factor XIII precipitate) to a body site in a manner such that clot formation W O 93/19805 CA21 ~ 5 P~r/~S93/02228 is effected, and to a kit suitable for use in such a method. (The term "body site" as used herein includes the tissue in the area of a wound or incision as well as implantable tissues or ~s to be inserted into the area, e.g., vascular prostheses, bone or collAg~n pads.) In the description that follows, it will be appreciated that a combination of isolated forms of fibrinogen and Factor XIII can be used in place of the plasma-derived precipitate.
In the method of the present invention, a fibrinogen/Factor XIII-enriched precipitate and mature thrombin are mixed together under conditions such that thrombin and/or Factor XIII are/is inactivated (or under conditions such that thrombin is present in an active form but is rendered unavailable, as in the calcium depletion '-'i described below) and clotting thereby prevented.
The mixture is then delivered to the body site under conditions such that the enzyme activity is restored (or thrombin availability restored).
In one '~ , the mixture of thrombin and fibrinogen/FaCtor XIII precipitate is prepared in a low pH buffer (the clotting of fibrinogen by ~~ in being inhibited by low pH (less than 5.5)).
In this : ' 'i- ~, thrombin activity is restored and clotting rapidly initiated upon neutralization of the mixture with a phArr~-~utically acceptable buffer, or alternatively, upon contact of the mixture with the patient's own body fluids. In this -'i- ~, the fibrinogen/Factor XIII precipitate can be prepared at a low pH or, alternatively, a low pH buffer can be used to dissolve the plasma W O 93/19805 C A 2 1 1 7 4 9 5 P~r/~S93/02228 precipitate and the lyophilized thrombin. In either case, the mixture can be transferred to a delivery container (such as a spray bottle or syringe) and applied to the body site directly, if conditions are ~ 5 such that the patient's body fluids are sufficient to increase the pH to a point where clotting occurs.
Where conditions are such that the patient's body fluids are not sufficient to raise the pH of the precipitate/thrombin mixture to a point where thrombin activity is restored, a delivery device can be used that is designed such that, as the acidic mixture passes out of the device, it is contacted with buffer salts coated on an interior portion of the device. The buffer salts are selected such that when contact is made with the acidic mixture, dissolution occurs with the result that the pH is raised to a point where clotting takes place. For example, a syringe can be used as the delivery device (applicator), where the syringe is fitted with a ~;cpoc~hle tip, the interior surface of which is coated with appropriate buffer salts. As the acidic mixture passes through the coated tip, the buffer (in the form, for example, of crystals or a gel) neutralizes the acidic mixture, thus restoring thrombin activity and effecting the formation of a clot at the desired site. Should clot formation occur in the tip, the tip can simply be removed and a new coated tip attached.
In another o~lir ---t, the fibrinogen and Factor XIII precipitate/thrombin mixture can be ~ prepared in a buffer that is depleted of calcium.
Rapid clot formation requires the presence of calcium ions: thus, if the calcium is removed, W O 93/19805 C A ~ 14 ~ 5 PC~r/~S93/02228 fibrin polymerization is inhibited (see Carr et al Biochem J. (1986) 239:513; KAminck; et al J. Biol.
Chem (1983) 258:10530: Kanaide et al (1982) 13:229).
Calcium chelators (. -c such as sodium citrate or ethyl~nP~iAminetetraacetic acid, which tightly bind calcium and make it inAcc~ccible) can be added to the solution used to precipitate the fibrinogen and Factor XIII and/or the dissolving buffer. To restore activity, the container (for example, a syringe) can be attached to a ~icpocAhle sterile tip, the interior surface of which is coated internally with sufficient calcium salt to saturate the chelator. As the free calcium ~u..~--LL~tion increases upon passage of the mixture through the tip, clotting is effected at the body site.
In a further ~ L, the clotting activity of thrombin, in the precipitate/thrombin mixture, can be inhibited using a photosensitive inhibitor. For example, light sensitive ri yl derivatives can be used to inactivate thrombin, at room temperature in the absence of light, for more than 26 hours (Turner et al J. Am. Chem. Soc. 109:
1274-1275 (1987); Turner et al J. Am. Chem. Soc.
110: 244-250 (1988)). These same thrombin inhibitor complexes can generate active thrombin within 1-2 seconds of irradiation (low intensity). These inhibitors are known to form acyl-enzyme complexes involving the active site serine hydroxyl (SER 195).
Upon irradiation, the cinnamoyl derivative undergoes photoisomerization to release coumarin and regenerate the active serine hydroxyl. Since coumarin derivatives are not good thrombin inhibitors, this photocyclization reaction WO93/19805 C~2 i 1 7495 i PCT/US93/0222X
effectively removes inhibitor from the enzyme solution. Thus, a solution of the fibrinogen/Factor XIII-enriched precipitate can be mixed with lyoph;li7~ inhibitor:thrombin complex in a dark environment (such as an opaque or colored syringe or container) and delivered to the wound site.
Activation of the enzyme and thus clot formation occurs upon delivery to the wound due to the ~X~O~UL~ of the solution to normal room light.
Alternatively, activation can be controlled by a light source, for example, one built directly into the applicator, so that variations in lighting conditions will not result in variable clotting times.
Preferably, the clotting activity of thrombin, in the precipitate/thrombin mixture is inhibited by the use of a photosensitive inhibitor in combination with an irreversible inhibitor. Such "doubly-inhibited" thrombin offers several advantages including increased stability of the double inhibitor:thrombin complex and fibrinogen/Factor XIII precipitate which reduces premature clot formation during long term storage as well as during use. The doubly inhibited thrombin also provides for increased control of clot activation, increased clot strength and increased time for mixing with additives (such as, for example, bone granules, antibiotics or growth factors).
For example, thrombin activity can be inhibited by the use of the photosensitive inhibitor 4-amidino-phenyl-2-hydroxy-4-diethylamino-alpha-methylcinnamate hydrochloride, "I-l" and the W o 93/19805 C A 2 i 1 14 9 5 P(~r/~S93/02228 irreversible inhibitor D-phenylalanyl-L-prolyl-L-arginine chloromethyl ketone, "PPACK". I-l inhibits approximately 99% of the thrombin activity, however, the residual 1% activity can not be reduced by additional I-1. This residual activity, which is sufficient to cause clotting after 15-20 minutes in the absence of light, can be reduced or titrated by the addition of the second inhibitor ("PPACK" in the present example). In the absence of activating irradiation (light of approximately 360 nm as provided by a Caulk W Polymerization Unit), the doubly inhibited thrombin solution is subse~ue.-Lly added to the fibrinogen c L and stored frozen, lyophilized or used within hours. Fibrin sealant prepared from this double inhibitor:Lh , I in complex and fibrinogen/Factor XIII precipitate can be maintained in the dark without clotting for over 2 hours. When the fibrin sealant is needed, the mixture can be dispensed onto the wound site or appropriate delivery container (such as a spray bottle or syringe) and activated by light to clot in less than 1-2 minutes.
~ h~ . ' ;n inhibited by the combination of such inhibitors may be useful for other applications. For example, the double inhibitor:thrombin complex can be used in situations requiring careful control of clotting activity or when thrombin is used alone to stop bleeding. In addition, the inhibited thrombin can be used to localize clotting activity (for example, eye surgery). The inhibited thrombin may also be used with other components or in other forms (for W O 93/19805 C A 2 1 1 7 4 9 5 P(~r/US93/02228 example, with gel matrix or on solid support) to improve hAn~l ing of delivery to the wound site.
In yet another ' ~;- L, premature clot formation can be prevented prior to delivery of the fibrinogen and Factor XIII/thrombin mixture by physically separating the thrombin from the fibrinogen/Factor XIII precipitate. In this : '-'; ~, physical separation is effected using a two-phase system. Liquids suitable for use in this : ' 'i- t are non-miscible and readily separable into two phases. The two phases are mixed into a suspension before each application and delivered to the wound. Where conditions are such that the patient's body fluids extract the soluble ~
of the nonaqueous phase, mixing occurs at the body ~site and clotting is thus initiated. If conditions will not elicit proper mixing of - ts, a delivery device can be used that is designed such that, as the suspension passes out of the device, it is contacted with a solubilizing agent coated on an interior portion of the device~ The solubilizing agent is selected such that when contact is made with the suspension, dissolution occurs with the result that mixing occurs to a degree where clotting takes place. For example, a syringe is fitted with a ~;epnc~hle tip, the interior surface of which is coated with an appropriate phase transfer agent(s).
As the suspension passes through the coated tip, the phase transfer agent (in the form, for example, of crystals) assists in the mixing process, thus allowing clot formation. Should clot formation occur in the tip, the tip can simply be removed and a new coated tip attached.
W O 93/19805 ~ A 2 i 1 7 4 ~ 5 P~r/~S93/02228 The present invention also relates to a kit suitable for use in the above de~e~ibed method of delivering fibrin sealant _ ~s to a wound site. In a preferred A ' ~ the kit includes an applicator designed so as to permit mixing of the fibrinogen/Factor XIII precipitate and thrombin in a single system. The applicator can be one that permits the application at the body site of, for example, a film, or a thin line of the , ~ s of fibrin sealant. Alternatively, a pump or aerosol spray applicator can be used.
As suggested above, the applicator can, for example, take the form of a glass or plastic syringe with ~;~pos~hle tips. The shape of the tip will determine the form in which the ,~ ~s are delivered. A tip with a flat, broad end can be used to deliver a thin wide streak of fibrin sealant whereas a narrow tubular end can be used to deliver a round thread of sealant. Applying pleS~ule to force the mixture through a tip constricted with, for example, a mesh screen can be used to produce a spray, resulting in a fine glaze of fibrin sealant.
In another : ' 'i- L, particularly suitable for use with the above-described photosensitive thrombin inhibitor, the applicator can take the form of a pump or aerosol spray device having a built-in light source situated such that, as the sealant components exit the device, they are irradiated with the light.
The wavelength of light used would depend on the photosensitizer.
The kit can be structured so as to include individual storage containers for the separate fibrin sealant ~ , ents. The kit can also include W O 93/19805 C A 2 1 1 7 4 9 5 ~ PC~r/US93/02228 one or more other storage containers ~icpose~ within which are any nPcPCsAry reagents, including solvents, buffers, etc.
The present invention will be understood in greater detail by reference to the following non-limiting Examples.
EXAMPLES
ExamPle 1 A precipitate containing fibrinogen and Factor XIII was prepared as follows:
Four hundred fifty microliters of a stock 1 M zinc sulfate solution were added to 5 ml of anticoagulated (citrate phosphate dextrose adenine (CPDA-1)) human plasma. The solution was mixed well without vortexing and centrifuged at 2,000 to 9,000 g for 5 minutes. The supernatant was decanted and discarded.
Inhibition of clotting and reactivation was achieved by any of the following methods:
A. Acid Tnh;bition - Lyophilized bovine thrombin was dissolved in citrate buffer (500 mM
citric acid, 150 mM NaCl, and 20 mM EACA, pH 4.5) to a final conce,l~.Gtion of 100 U/ml. Precipitated fibrinogen was dissolved in Tris buffer (50 mM Tris, 250 mM sodium citrate, 150 mM sodium chloride, 50 mM
Arginine (Arg), and 20 mM ~-amino-caproic acid (EACA), pH 7.4) to a concentration of approximately 15.0 mg/ml. This fibrinogen stock was then diluted 25-fold in citrate buffer. The clotting time for 200 microliters of this fibrinogen solution plus 100 microliters thrombin ~YceP~Pd 90 seconds in a Becton W O 93/19805 C A 2 1 1 7 4 9 5 PC~r/~S93/02228 Dic-lc;n con BBL Fibrosystem fibrometer under standard conditions indicating no clot formation. Addition of 70 microliters of lN sodium hydroxide resulted in clot formation in 3.8 seconds (average of 10 samples).
The following p-~cedu.es can be used for application to a wound site:
Where body fluids are sufficient to neutralize the acidic mixture of precipitate and thrombin, the mixture can be applied directly to the wound site. Alternatively, the delivery device can be connected to dicroc~hle tips coated internally with a neutralizing salt or gel (e.g. Tris).
Neutralization of the acidic solution by the buffer salts activatss thrombin and restores clotting activity.
B. Chelator Inhibition - Precipitated fibrinogen and lyophili7sd bovine thrombin were dissolved in Tris buffer (50 mM Tris, 250 mM sodium citrate, 150 mM sodium chloride, 50 mM Arg, and 20 mM EACA, p~ 7.4) to a c~"~e..LL~tion of approximately 15.0 mg/ml and 100 U/ml, respectively.
The fibrinogen stock solution was then diluted 25-fold in Tris buffer containing 500 mM sodium citrate. The clotting time for 200 microliters of this fibrinogen solution plus 100 microliters thrombin ~Ycee~d 90 seconds in a Becton Dickinson BBL fibrosystem fibrometer under standard conditions. Addition of 50 microliters of a lM
CaCl, solution resulted in clot formation in 1.8 seconds (average of 10 samples).
W O 93/19805 C A 2 i 1 / 4 9 5 PC~r/~S93/02228 The following procedure can be used for application to a wound site:
- The delivery device is connected to ~i~pn~hle tips coated internally with a calcium - 5 salt or gel. As the mixture passes through the tip, the molar excess of calcium saturates the chelator and clotting is thereby promoted.
C. Photosensitive Inhibition - In the absence of light, a 5 to 20-fold excess of 4-10 amidino phe~l-2-hydroxy-4-diethylamino-alpha-methylcinnamate hydrochloride (Porter et al, J.
Amer. Chem. Soc. 111:7616 (1989)) was added to thrombin in buffer (approximately 100 U/ml in 50 mM
Tris, 250 mM sodium chloride, 250 mM sodium citrate, 20 mM EACA, 50 mM arginine (or urea), pH 7.4, final methanol cu.,~e--Llation <10%). The inhibition was allowed to proceed for at least 1 hour at room t~ a LUL ~.
A minimal quantity of this solution was used to dissolve the precipitated fibrinogen/Factor XIII. This sealant required approximately 2 to 3 minutes illumination under standard operating lights to clot completely, whereas a sample mixture kept in the dark did not clot after 90 min.
The following pLuceduL~s can be used for application to a wound site:
The photosensitive inhibitor-thrombin complex can be mixed with the precipitated fibrinogen/Factor XIII in a colored delivery device that does not transmit light of the activating wavelengths. Delivery of the mixture to an illuminated wound site results in clot formation.
W O 93/19805 C A 2 1 1 7 4 q 5 PC~r/US93/02228 D. Two Phase Suspension - Lyophilized bovine thrombin is dissolved in an emulsifying agent to a final c~"cel,L-~tion of about 100 U/ml.
Precipitated fibrinogen/Factor XIII is dissolved in a minimal volume of buffer (50 mM Tris, 150 mN
sodium chloride, 250 mM sodium citrate, 20 mM EACA, 50mM Arg, pH 7.4). A sllqppn~i~n of the ; i~c;hle liquids is formed. On a wound surface, body fluids may be sufficient to dissolve both components and promote proper mixing and clot formation.
ExamDle 2 A fibrin sealant was prepared using the two enzyme inhibitors as follows. All inactivation and transfer steps were performed in the absence of activating light (366 nm), using a dark room illuminated by a red light.
Thrombin was inactivated sequentially using 1) the light sensitive inhibitor 4-Am;~in~ph~nyl-2 }Iy~lv~ 4-diethylamino-alpha-methylcinnamate hydrochloride and 2) D-Phenylalanyl-L-prolyl-L-arginine Chloromethyl Ketone, PPACK.
Bovine thrombin (0.1 ml of 1,000 units/ml Armour Thrombinar) was mixed with 0.8 ml of 50 mM tris buffer, pH 7.3, and 0.1 ml of 31.2~ ~g/ml inhibitor in 2.5% ethanol, 50 mM tris, pH 7.3. The solution was incubated at room temperature for 2 hours.
Residual thrombin activity of the thrombin:inhibitor-l complex (approximately 1%
original activity) was inactivated by titration with PPACK until residual activity was such that the resulting fibrin sealant could be maintained in the W O 93/19805 C A 2 i 1 7 4 q 5 PC~r/~S93/02228 dark for 2-4 hours without premature clotting. The above solution (0.1 ml) was mixed with 0.01 ml of ~ 2.5 ~g/ml PPACK at room temperature and thrombin activity of the thrombin:inhibitor-2 was negligible ~ 5 within minutes.
Fibrin sealant was prepared by mixing 1 ml fibrinogen stock (e.g. single unit human cryoprecipitate) with 0.1 ml of the thrombin:inhibitor-2 in the absence of activating light (e.g. opaque container) and transferred to an amber syringe or spray container for delivery.
Clotting activity was easily controlled using light. In the absence of activating light, the mixture was stable for at least 2 hours at room temperature before clotting occurred. A 20 second pulse of light (that is, light of 360 nm, by Caulk MAX polymerization unit) at a distance of less than 5 cm from the dispensed solution resulted in clotting within seconds.
* * * *
The entire contents of each of the references cited above are hereby incorporated by reference.
While the present invention has been described in some detail for purposes of clarity and understanding, it will be clear to one skilled in the art from a reading of this disclosure that various changes can be made in form and detail without departing from the true scope of the invention.
Claims (24)
1. A method of effecting the formation of fibrin sealant at a body site comprising:
i) mixing, in a container means, an aqueous solution comprising fibrinogen, Factor XIII
and mature thrombin under conditions such that thrombin clotting activity is inhibited; and ii) applying a preparation resulting from step (i) to said body site under conditions such that thrombin clotting activity is restored and said fibrin sealant is formed.
i) mixing, in a container means, an aqueous solution comprising fibrinogen, Factor XIII
and mature thrombin under conditions such that thrombin clotting activity is inhibited; and ii) applying a preparation resulting from step (i) to said body site under conditions such that thrombin clotting activity is restored and said fibrin sealant is formed.
2. The method according to claim 1 wherein step (i) is carried out at a pH of less than 5.5, whereby thrombin clotting activity is inhibited.
3. The method according to claim 2 wherein, in step (ii), the pH of said preparation resulting from step (i) is increased such that thrombin clotting activity is restored.
4. The method according to claim 3 wherein the pH of said preparation resulting from step (i) is increased upon contact of said preparation with body fluids of said patient present at said body site.
5. The method according to claim 3 wherein, in step (ii), the pH of said preparation resulting from step (i) is increased upon contact with a buffer capable of increasing the pH.
6. The method according to claim 1 wherein said solution of step (i) includes an amount of a photosensitive inhibitor of thrombin clotting activity sufficient to inhibit thrombin clotting activity.
7. The method according to claim 6 wherein, in step (ii), said preparation resulting from step (i) is irradiated with light of a wavelength that inactivates said photosensitive inhibitor, whereby thrombin clotting activity is restored.
8. The method according to claim 1 wherein said solution of step (i) includes an amount of a photosensitive inhibitor of thrombin clotting activity and an amount of a second inhibitor of thrombin clotting activity sufficient to inhibit thrombin clotting activity.
9. The method according to claim 8 wherein, in step (ii), said preparation resulting from step (i) is irradiated with light of a wavelength that inactivates said photosensitive inhibitor, whereby thrombin clotting activity is restored.
10. The method according to claim 1 wherein the conditions of step (i) are such that thrombin clotting activity is inhibited by the absence of calcium ions sufficient to effect fibrin sealant formation.
11. The method according to claim 10 wherein, in step (ii), the conditions are such that calcium ions sufficient to effect fibrin sealant formation are present.
12. The method according to claim 10 wherein said solution of step (i) includes an amount of a chelator of calcium sufficient to reduce free calcium ions in said solution to a level insufficient to effect fibrin sealant formation.
13. The method according to claim 11 wherein, in step (ii), said preparation resulting from step (i) is contacted with an amount of calcium ions sufficient to effect fibrin sealant formation.
14. A method of effecting the formation of fibrin sealant at a body site of a patient comprising:
i) forming a suspension comprising a first phase which comprises fibrinogen and Factor XIII and a second phase which comprises thrombin: and ii) applying said suspension to said body site under conditions such that mixing of said fibrinogen, Factor XIII and thrombin is effected so that said fibrin sealant is formed.
i) forming a suspension comprising a first phase which comprises fibrinogen and Factor XIII and a second phase which comprises thrombin: and ii) applying said suspension to said body site under conditions such that mixing of said fibrinogen, Factor XIII and thrombin is effected so that said fibrin sealant is formed.
15. The method according to claim 14 wherein, in step ii, said mixing occurs in a body fluid of said patient present at said body site.
16. The method according to claim 14 wherein, in step (ii), said suspension of step (i) is contacted with a phase transfer agent that effects mixing of said first and said second phase such that said fibrin sealant is formed.
17. A kit for use in the preparation of a fibrin sealant comprising an applicator that comprises:
i) a container means having disposed therein a solution comprising fibrinogen, Factor XIII and mature thrombin; and ii) an outlet means operably connected to said container means.
i) a container means having disposed therein a solution comprising fibrinogen, Factor XIII and mature thrombin; and ii) an outlet means operably connected to said container means.
18. The kit according to claim 17 wherein said solution further comprises a calcium chelator.
19. The kit according to claim 18 wherein said outlet means has a calcium salt disposed therein.
20. The kit according to claim 17 wherein said solution has a pH of less than about 5.5.
21. The kit according to claim 20 wherein said outlet means has a neutralizing buffer salt disposed therein.
22. The kit according to claim 17 wherein said solution further comprises a photosensitive inhibitor of thrombin clotting activity and wherein said container means and said outlet means are constructed of a material that does not transmit light of a wavelength to which said inhibitor is sensitive.
23. The kit according to claim 22 wherein said outlet means includes a source of light that, when activated, emits light at a wavelength that inactivates said photosensitive inhibitor.
24. The kit according to claim 17 wherein said solution further comprises a photosensitive inhibitor of thrombin clotting activity and a second inhibitor of thrombin clotting activity and wherein said container means and said outlet means are constructed of a material that does not transmit light of a wavelength to which said inhibitor is sensitive.
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US07/844,497 US5318524A (en) | 1990-01-03 | 1992-03-02 | Fibrin sealant delivery kit |
US07/844,497 | 1992-03-02 |
Publications (1)
Publication Number | Publication Date |
---|---|
CA2117495A1 true CA2117495A1 (en) | 1993-10-14 |
Family
ID=25292875
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CA002117495A Abandoned CA2117495A1 (en) | 1992-03-02 | 1993-03-02 | Fibrin sealant delivery method |
Country Status (6)
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US (1) | US5318524A (en) |
EP (1) | EP0630275A4 (en) |
JP (1) | JPH07506349A (en) |
AU (1) | AU675639B2 (en) |
CA (1) | CA2117495A1 (en) |
WO (1) | WO1993019805A1 (en) |
Families Citing this family (127)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US6559119B1 (en) * | 1990-11-27 | 2003-05-06 | Loyola University Of Chicago | Method of preparing a tissue sealant-treated biomedical material |
DK83092D0 (en) * | 1992-06-24 | 1992-06-24 | Unes As | PROCEDURE FOR THE EXTRACTION OF THROMBIN |
CN1091315A (en) * | 1992-10-08 | 1994-08-31 | E·R·斯奎布父子公司 | Fibrin sealant compositions and using method thereof |
US5443454A (en) * | 1992-12-09 | 1995-08-22 | Terumo Kabushiki Kaisha | Catheter for embolectomy |
US5795780A (en) * | 1993-06-23 | 1998-08-18 | Bristol-Myers Squibb Company | Method of use of autologous thrombin blood fraction in a cell culture with keratinocytes |
US5665114A (en) * | 1994-08-12 | 1997-09-09 | Meadox Medicals, Inc. | Tubular expanded polytetrafluoroethylene implantable prostheses |
ATE207374T1 (en) | 1994-08-17 | 2001-11-15 | Boston Scient Corp | IMPLANT AND APPLICATION DEVICE |
US5585007A (en) * | 1994-12-07 | 1996-12-17 | Plasmaseal Corporation | Plasma concentrate and tissue sealant methods and apparatuses for making concentrated plasma and/or tissue sealant |
US6965014B1 (en) | 1996-01-16 | 2005-11-15 | Baxter International Inc. | Fibrin material and method for producing and using the same |
US20020131933A1 (en) * | 1996-01-16 | 2002-09-19 | Yves Delmotte | Biopolymer membrane and methods for its preparation |
ATE244584T1 (en) * | 1995-01-16 | 2003-07-15 | Baxter Int | SELF-SUPPORTING SURFACES MADE OF CROSS-LINKED FIBRIN FOR INHIBITING POSTOPERATIVE ADHESIONS |
US6599515B1 (en) | 1995-01-16 | 2003-07-29 | Baxter International Inc. | Fibrin porous structure |
US5674394A (en) * | 1995-03-24 | 1997-10-07 | Johnson & Johnson Medical, Inc. | Single use system for preparation of autologous plasma |
US5707331A (en) * | 1995-05-05 | 1998-01-13 | John R. Wells | Automatic multiple-decanting centrifuge |
USRE38730E1 (en) * | 1995-05-05 | 2005-04-26 | Harvest Technologies Corporation | Automatic multiple-decanting centrifuge and method of treating physiological fluids |
ATE309006T1 (en) * | 1996-03-15 | 2005-11-15 | Chemo Sero Therapeut Res Inst | SPRAY DEVICE FOR APPLYING FABRIC ADHESIVE |
DE19781869T1 (en) * | 1996-04-30 | 2000-03-16 | Medtronic Inc | Process for the production of an autologous fibrin hemostatic agent |
WO2000062828A1 (en) * | 1996-04-30 | 2000-10-26 | Medtronic, Inc. | Autologous fibrin sealant and method for making the same |
AU4648697A (en) * | 1996-09-23 | 1998-04-14 | Chandrashekar Pathak | Methods and devices for preparing protein concentrates |
US8003705B2 (en) * | 1996-09-23 | 2011-08-23 | Incept Llc | Biocompatible hydrogels made with small molecule precursors |
US6566406B1 (en) * | 1998-12-04 | 2003-05-20 | Incept, Llc | Biocompatible crosslinked polymers |
US5975367A (en) * | 1996-09-27 | 1999-11-02 | Thermogenesis Corp. | Fibrin glue line and dot dispenser |
US5759171A (en) * | 1996-09-27 | 1998-06-02 | Thermogenesis Corp. | Sprayer for fibrin glue |
US5844087A (en) | 1996-11-05 | 1998-12-01 | Bayer Corporation | Method and device for delivering fibrin glue |
US6743248B2 (en) | 1996-12-18 | 2004-06-01 | Neomend, Inc. | Pretreatment method for enhancing tissue adhesion |
FR2757770B1 (en) * | 1996-12-30 | 1999-02-26 | Inoteb | PROCESS FOR THE PREPARATION OF A BIOLOGICAL GLUE CAPABLE OF COAGULATING BY SIMPLE ADDITION OF CALCIUM IONS |
US6177609B1 (en) | 1997-03-10 | 2001-01-23 | Meadox Medicals, Inc. | Self-aggregating protein compositions and use as sealants |
US7241309B2 (en) * | 1999-04-15 | 2007-07-10 | Scimed Life Systems, Inc. | Self-aggregating protein compositions and use as sealants |
US20040176801A1 (en) * | 1997-03-12 | 2004-09-09 | Neomend, Inc. | Pretreatment method for enhancing tissue adhesion |
US6371975B2 (en) | 1998-11-06 | 2002-04-16 | Neomend, Inc. | Compositions, systems, and methods for creating in situ, chemically cross-linked, mechanical barriers |
US20030191496A1 (en) * | 1997-03-12 | 2003-10-09 | Neomend, Inc. | Vascular sealing device with microwave antenna |
CA2284679C (en) | 1997-03-27 | 2006-06-13 | Bristol-Myers Squibb Company | Laparoscopic sealant applicator |
AU7977298A (en) | 1997-06-18 | 1999-01-04 | Cohesion Technologies, Inc. | Compositions containing thrombin and microfibrillar collagen, and methods for preparation and use thereof |
US6589199B1 (en) | 1997-08-28 | 2003-07-08 | Boston Scientific Corporation | System for implanting a cross-linked polysaccharide fiber and methods of forming and inserting the fiber |
WO1999011191A1 (en) | 1997-08-28 | 1999-03-11 | Boston Scientific Corporation | System for implanting a cross-linked polysaccharide fiber and methods of forming and inserting the fiber |
WO1999018931A1 (en) * | 1997-10-10 | 1999-04-22 | University Of Virginia | Use of fibrin sealant to maintain hemostasis, lymphostasis and prevent local accumulation of body fluids |
US6478808B2 (en) | 1997-12-17 | 2002-11-12 | Closys Corporation | Clotting cascade initiating apparatus and methods of use and methods of closing wounds |
US6159232A (en) * | 1997-12-16 | 2000-12-12 | Closys Corporation | Clotting cascade initiating apparatus and methods of use and methods of closing wounds |
US6033427A (en) * | 1998-01-07 | 2000-03-07 | Lee; Benjamin I. | Method and device for percutaneous sealing of internal puncture sites |
USD408527S (en) * | 1998-02-02 | 1999-04-20 | Thermogenesis Corp. | Medical dispensing gun |
US6274090B1 (en) | 1998-08-05 | 2001-08-14 | Thermogenesis Corp. | Apparatus and method of preparation of stable, long term thrombin from plasma and thrombin formed thereby |
US6994686B2 (en) | 1998-08-26 | 2006-02-07 | Neomend, Inc. | Systems for applying cross-linked mechanical barriers |
US6458147B1 (en) * | 1998-11-06 | 2002-10-01 | Neomend, Inc. | Compositions, systems, and methods for arresting or controlling bleeding or fluid leakage in body tissue |
US6899889B1 (en) | 1998-11-06 | 2005-05-31 | Neomend, Inc. | Biocompatible material composition adaptable to diverse therapeutic indications |
US6830756B2 (en) * | 1998-11-06 | 2004-12-14 | Neomend, Inc. | Systems, methods, and compositions for achieving closure of vascular puncture sites |
US6949114B2 (en) | 1998-11-06 | 2005-09-27 | Neomend, Inc. | Systems, methods, and compositions for achieving closure of vascular puncture sites |
US7279001B2 (en) * | 1998-11-06 | 2007-10-09 | Neomend, Inc. | Systems, methods, and compositions for achieving closure of vascular puncture sites |
DE10025001A1 (en) * | 2000-05-22 | 2001-11-29 | Aventis Behring Gmbh | Tissue glue with improved anti-adhesive properties |
US20030069601A1 (en) * | 1998-12-15 | 2003-04-10 | Closys Corporation | Clotting cascade initiating apparatus and methods of use |
WO2000061256A1 (en) | 1999-04-12 | 2000-10-19 | Harvest Technologies Corporation | Method and apparatus for producing platelet rich plasma and/or platelet concentrate |
US20030007954A1 (en) | 1999-04-12 | 2003-01-09 | Gail K. Naughton | Methods for using a three-dimensional stromal tissue to promote angiogenesis |
EP1185278A4 (en) * | 1999-06-01 | 2002-09-18 | Bristol Myers Squibb Co | Fibrin polymer structure |
WO2000074575A1 (en) * | 1999-06-01 | 2000-12-14 | Closys Corporation | Clotting cascade initiating apparatus and methods of use |
US6472162B1 (en) | 1999-06-04 | 2002-10-29 | Thermogenesis Corp. | Method for preparing thrombin for use in a biological glue |
JP2003520830A (en) | 2000-01-25 | 2003-07-08 | エドワーズ ライフサイエンシーズ コーポレイション | Delivery system for treatment of restenosis and anastomotic intimal hyperplasia |
EP1301245A4 (en) * | 2000-07-17 | 2007-02-28 | Haemacure Corp | Spray head for applying a multi-component mixture |
EP1359947A2 (en) * | 2001-01-25 | 2003-11-12 | Nycomed Pharma AS | A suspension comprising fibrinogen, thrombin and alcohol and a method of coating a carrier with the same |
US7098315B2 (en) | 2001-01-25 | 2006-08-29 | Nycomed Pharma As | Method of preparing a collagen sponge, a device for extracting a part of a collagen foam, and an elongated collagen sponge |
US7052713B2 (en) | 2001-02-13 | 2006-05-30 | Nycomed Pharma As | Carrier with solid fibrinogen and solid thrombin |
ES2274746T3 (en) * | 2001-05-09 | 2018-04-26 | Baxter International Inc. | Fibrin material and its production and use procedure |
DE60121826T2 (en) * | 2001-05-09 | 2007-02-22 | Baxter International Inc., Deerfield | FIBRIN MATERIAL AND METHOD FOR ITS MANUFACTURE AND USE |
US6623959B2 (en) | 2001-06-13 | 2003-09-23 | Ethicon, Inc. | Devices and methods for cell harvesting |
JP2005507926A (en) * | 2001-10-09 | 2005-03-24 | ザイモジェネティクス,インコーポレイティド | Method for inhibiting the formation of seromas using factor XIII |
AU2002336694A1 (en) * | 2001-11-01 | 2003-05-12 | Lawrence M. Boyd | Devices and methods for the restoration of a spinal disc |
JP4125234B2 (en) * | 2001-11-01 | 2008-07-30 | スパイン・ウェイブ・インコーポレーテッド | Apparatus and method for pretreatment of endplates between discs |
US20050118156A1 (en) * | 2001-12-04 | 2005-06-02 | Woolverton Christopher J. | Storage-stable fibrin sealant |
WO2003071986A2 (en) * | 2002-02-22 | 2003-09-04 | Control Delivery Systems, Inc. | Method for treating otic disorders |
US7832566B2 (en) | 2002-05-24 | 2010-11-16 | Biomet Biologics, Llc | Method and apparatus for separating and concentrating a component from a multi-component material including macroparticles |
US20030205538A1 (en) | 2002-05-03 | 2003-11-06 | Randel Dorian | Methods and apparatus for isolating platelets from blood |
US7992725B2 (en) | 2002-05-03 | 2011-08-09 | Biomet Biologics, Llc | Buoy suspension fractionation system |
DE10392686T5 (en) | 2002-05-24 | 2005-07-07 | Biomet Mfg. Corp., Warsaw | Apparatus and method for separating and concentrating liquids containing multiple components |
US20060278588A1 (en) | 2002-05-24 | 2006-12-14 | Woodell-May Jennifer E | Apparatus and method for separating and concentrating fluids containing multiple components |
US7845499B2 (en) | 2002-05-24 | 2010-12-07 | Biomet Biologics, Llc | Apparatus and method for separating and concentrating fluids containing multiple components |
US7135027B2 (en) | 2002-10-04 | 2006-11-14 | Baxter International, Inc. | Devices and methods for mixing and extruding medically useful compositions |
US20040243044A1 (en) * | 2003-06-02 | 2004-12-02 | Penegor Stephen A. | Hemostatic wound dressing |
WO2005070071A2 (en) * | 2004-01-08 | 2005-08-04 | Spine Wave Inc. | Apparatus and method for injecting fluent material at a distracted tissue site |
EP1768617A4 (en) * | 2004-06-29 | 2011-08-10 | Spine Wave Inc | Methods for treating defects and injuries of an intervertebral disc |
CA2574773A1 (en) * | 2004-07-22 | 2006-02-02 | Allan D. Pronovost | Compositions and methods for treating excessive bleeding |
ATE550423T1 (en) * | 2004-08-30 | 2012-04-15 | Theregen Inc | THREE-DIMENSIONAL CULTIVATED TISSUES AND THEIR USE |
US20080226604A9 (en) * | 2005-06-17 | 2008-09-18 | Theregen Company | Methods for treating ischemic tissue |
KR100689516B1 (en) * | 2004-09-15 | 2007-03-02 | 삼성전자주식회사 | Method and apparatus for indicating preferred layer information in multimedia broadcast/multicast system |
US20060094865A1 (en) * | 2004-10-29 | 2006-05-04 | Kapur Terri A | Intraoperative method for isolating and concentrating autologous growth factors and for forming residual autologous growth factor compositions |
US7866485B2 (en) | 2005-02-07 | 2011-01-11 | Hanuman, Llc | Apparatus and method for preparing platelet rich plasma and concentrates thereof |
WO2006086199A1 (en) | 2005-02-07 | 2006-08-17 | Hanuman Llc | Platelet rich plasma concentrate apparatus and method |
EP1848474B1 (en) | 2005-02-07 | 2013-06-12 | Hanuman LLC | Platelet rich plasma concentrate apparatus and method |
AU2006213822B2 (en) * | 2005-02-09 | 2011-05-26 | Covidien Lp | Synthetic sealants |
US7766900B2 (en) * | 2005-02-21 | 2010-08-03 | Biomet Manufacturing Corp. | Method and apparatus for application of a fluid |
US20070014780A1 (en) * | 2005-07-13 | 2007-01-18 | Statseal | Storage-stable human fibrinogen solutions |
US8567609B2 (en) | 2006-05-25 | 2013-10-29 | Biomet Biologics, Llc | Apparatus and method for separating and concentrating fluids containing multiple components |
US8357168B2 (en) * | 2006-09-08 | 2013-01-22 | Spine Wave, Inc. | Modular injection needle and seal assembly |
US20090227981A1 (en) * | 2007-03-05 | 2009-09-10 | Bennett Steven L | Low-Swelling Biocompatible Hydrogels |
US20090227689A1 (en) * | 2007-03-05 | 2009-09-10 | Bennett Steven L | Low-Swelling Biocompatible Hydrogels |
JP5479319B2 (en) | 2007-04-12 | 2014-04-23 | バイオメット・バイオロジックス・リミテッド・ライアビリティ・カンパニー | Buoy suspension fractionation system |
US8328024B2 (en) | 2007-04-12 | 2012-12-11 | Hanuman, Llc | Buoy suspension fractionation system |
US8067028B2 (en) * | 2007-08-13 | 2011-11-29 | Confluent Surgical Inc. | Drug delivery device |
EP2567692B1 (en) | 2008-02-27 | 2016-04-06 | Biomet Biologics, LLC | Use of a device for obtaining interleukin-1 receptor antagonist rich solutions |
US8337711B2 (en) | 2008-02-29 | 2012-12-25 | Biomet Biologics, Llc | System and process for separating a material |
US8753670B2 (en) * | 2008-03-26 | 2014-06-17 | Baxter International Inc. | Fibrin foam and process |
US8512740B2 (en) * | 2008-03-26 | 2013-08-20 | Baxter International Inc. | Fibrin foam and process for making |
US8518272B2 (en) | 2008-04-04 | 2013-08-27 | Biomet Biologics, Llc | Sterile blood separating system |
US8182769B2 (en) | 2008-04-04 | 2012-05-22 | Biomet Biologics, Llc | Clean transportation system |
US8012077B2 (en) | 2008-05-23 | 2011-09-06 | Biomet Biologics, Llc | Blood separating device |
US8187475B2 (en) | 2009-03-06 | 2012-05-29 | Biomet Biologics, Llc | Method and apparatus for producing autologous thrombin |
US8313954B2 (en) | 2009-04-03 | 2012-11-20 | Biomet Biologics, Llc | All-in-one means of separating blood components |
WO2010118352A1 (en) | 2009-04-09 | 2010-10-14 | Arizona Board Of Reagents On Behalf Of The University Of Arizona | Cellular seeding and co-culture of a three dimensional fibroblast construct |
US9011800B2 (en) | 2009-07-16 | 2015-04-21 | Biomet Biologics, Llc | Method and apparatus for separating biological materials |
US8591391B2 (en) | 2010-04-12 | 2013-11-26 | Biomet Biologics, Llc | Method and apparatus for separating a material |
US10231721B2 (en) | 2010-06-24 | 2019-03-19 | St. Croix Surgical Systems, Llc | Failsafe percutaneous wound barrier |
US20120065592A1 (en) * | 2010-09-10 | 2012-03-15 | E.I. Du Pont De Nemours And Company | Device for dispensing microliter quantities of a material into a puncture wound site |
US20130202675A1 (en) | 2012-02-03 | 2013-08-08 | Dynasil Biomedical Corporation | Systems and methods for the fabrication of tissue patches |
US9642956B2 (en) | 2012-08-27 | 2017-05-09 | Biomet Biologics, Llc | Apparatus and method for separating and concentrating fluids containing multiple components |
US10016527B2 (en) | 2012-10-23 | 2018-07-10 | Orthovita, Inc. | Materials and methods for repair of cartilage defects |
US9895418B2 (en) | 2013-03-15 | 2018-02-20 | Biomet Biologics, Llc | Treatment of peripheral vascular disease using protein solutions |
US10208095B2 (en) | 2013-03-15 | 2019-02-19 | Biomet Manufacturing, Llc | Methods for making cytokine compositions from tissues using non-centrifugal methods |
US9950035B2 (en) | 2013-03-15 | 2018-04-24 | Biomet Biologics, Llc | Methods and non-immunogenic compositions for treating inflammatory disorders |
US20140271589A1 (en) | 2013-03-15 | 2014-09-18 | Biomet Biologics, Llc | Treatment of collagen defects using protein solutions |
US10143725B2 (en) | 2013-03-15 | 2018-12-04 | Biomet Biologics, Llc | Treatment of pain using protein solutions |
IL230150A0 (en) | 2013-12-24 | 2014-09-30 | Omrix Biopharmaceuticals Ltd | One component fibrin glue comprising zymogens |
IL230151A0 (en) | 2013-12-24 | 2014-09-30 | Omrix Biopharmaceuticals Ltd | One component fibrin glue comprising a polymerization inhibitor |
IL231230A0 (en) | 2014-02-27 | 2014-08-31 | Omrix Biopharmaceuticals Ltd | Fibrinogen formulation |
IL231792A0 (en) | 2014-03-27 | 2014-08-31 | Omrix Biopharmaceuticals Ltd | Device and method for preparing and administering one-component fibrin sealant |
US9713810B2 (en) | 2015-03-30 | 2017-07-25 | Biomet Biologics, Llc | Cell washing plunger using centrifugal force |
US9757721B2 (en) | 2015-05-11 | 2017-09-12 | Biomet Biologics, Llc | Cell washing plunger using centrifugal force |
US9540548B1 (en) | 2015-08-07 | 2017-01-10 | Xcede Technologies, Inc. | Adhesive compositions and related methods |
US9833538B2 (en) | 2015-08-07 | 2017-12-05 | Xcede Technologies, Inc. | Adhesive compositions and related methods |
AU2016307447A1 (en) | 2015-08-07 | 2018-02-22 | Xcede Technologies, Inc. | Adhesive compositions and related methods |
WO2023102558A1 (en) * | 2021-12-03 | 2023-06-08 | The Board Of Trustees Of The Leland Stanford Junior University | Photo-enzymatic printing |
Family Cites Families (26)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
DE703476C (en) * | 1938-06-09 | 1941-03-10 | I G Farbenindustrie Akt Ges | s of slaughter animals |
US2533004A (en) * | 1943-10-27 | 1950-12-05 | John D Ferry | Fibrin clots and methods for preparing the same |
US2492458A (en) * | 1944-12-08 | 1949-12-27 | Jr Edgar A Bering | Fibrin foam |
US3723244A (en) * | 1971-01-18 | 1973-03-27 | Atomic Energy Commission | Fibrous fibrin sheet and method for producing same |
US4265233A (en) * | 1978-04-12 | 1981-05-05 | Unitika Ltd. | Material for wound healing |
AT359653B (en) * | 1979-02-15 | 1980-11-25 | Immuno Ag | METHOD FOR PRODUCING A TISSUE ADHESIVE |
AT366916B (en) * | 1980-04-02 | 1982-05-25 | Immuno Ag | DEVICE FOR APPLICATING A TISSUE ADHESIVE BASED ON HUMAN OR ANIMAL PROTEINS |
US4359463A (en) * | 1980-11-26 | 1982-11-16 | Rock Gail A | Stabilization of Factor VIII activity in whole blood or blood plasma |
DE3105624A1 (en) * | 1981-02-16 | 1982-09-02 | Hormon-Chemie München GmbH, 8000 München | MATERIAL FOR SEALING AND HEALING Wounds |
EP0068047B1 (en) * | 1981-06-25 | 1986-07-23 | Serapharm GmbH & Co. KG | Enriched plasma derivative for promoting wound sealing and wound healing |
US4442655A (en) * | 1981-06-25 | 1984-04-17 | Serapharm Michael Stroetmann | Fibrinogen-containing dry preparation, manufacture and use thereof |
AT379311B (en) * | 1984-03-29 | 1985-12-27 | Immuno Ag | DEVICE FOR APPLICATING A TISSUE ADHESIVE |
JPS6144825A (en) * | 1984-08-09 | 1986-03-04 | Unitika Ltd | Hemostatic agent |
US4627879A (en) * | 1984-09-07 | 1986-12-09 | The Trustees Of Columbia University In The City Of New York | Fibrin adhesive prepared as a concentrate from single donor fresh frozen plasma |
US4668621A (en) * | 1985-04-22 | 1987-05-26 | Wake Forest University | Detecting blood clotting factors with immobilized fibrinogen and labeled fibrinogen |
AT382783B (en) * | 1985-06-20 | 1987-04-10 | Immuno Ag | DEVICE FOR APPLICATING A TISSUE ADHESIVE |
US4696812A (en) * | 1985-10-28 | 1987-09-29 | Warner-Lambert Company | Thrombin preparations |
DE3622642A1 (en) * | 1986-07-05 | 1988-01-14 | Behringwerke Ag | ONE-COMPONENT TISSUE ADHESIVE AND METHOD FOR THE PRODUCTION THEREOF |
AT397203B (en) * | 1988-05-31 | 1994-02-25 | Immuno Ag | FABRIC ADHESIVE |
US4874368A (en) * | 1988-07-25 | 1989-10-17 | Micromedics, Inc. | Fibrin glue delivery system |
US4902281A (en) * | 1988-08-16 | 1990-02-20 | Corus Medical Corporation | Fibrinogen dispensing kit |
US5226877A (en) * | 1989-06-23 | 1993-07-13 | Epstein Gordon H | Method and apparatus for preparing fibrinogen adhesive from whole blood |
US5116315A (en) * | 1989-10-03 | 1992-05-26 | Hemaedics, Inc. | Biological syringe system |
US5104375A (en) * | 1989-10-16 | 1992-04-14 | Johnson & Johnson Medical, Inc. | Locking holder for a pair of syringes and method of use |
US5030215A (en) * | 1990-01-03 | 1991-07-09 | Cryolife, Inc. | Preparation of fibrinogen/factor XIII precipitate |
US5219328A (en) * | 1990-01-03 | 1993-06-15 | Cryolife, Inc. | Fibrin sealant delivery method |
-
1992
- 1992-03-02 US US07/844,497 patent/US5318524A/en not_active Expired - Lifetime
-
1993
- 1993-03-02 WO PCT/US1993/002228 patent/WO1993019805A1/en not_active Application Discontinuation
- 1993-03-02 AU AU38033/93A patent/AU675639B2/en not_active Ceased
- 1993-03-02 CA CA002117495A patent/CA2117495A1/en not_active Abandoned
- 1993-03-02 JP JP5517458A patent/JPH07506349A/en not_active Ceased
- 1993-03-02 EP EP93907428A patent/EP0630275A4/en not_active Withdrawn
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JPH07506349A (en) | 1995-07-13 |
EP0630275A1 (en) | 1994-12-28 |
US5318524A (en) | 1994-06-07 |
AU3803393A (en) | 1993-11-08 |
AU675639B2 (en) | 1997-02-13 |
WO1993019805A1 (en) | 1993-10-14 |
EP0630275A4 (en) | 1995-05-24 |
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EEER | Examination request | ||
FZDE | Discontinued |