CA2302224C - Fibrinogen-based tissue adhesive - Google Patents
Fibrinogen-based tissue adhesive Download PDFInfo
- Publication number
- CA2302224C CA2302224C CA002302224A CA2302224A CA2302224C CA 2302224 C CA2302224 C CA 2302224C CA 002302224 A CA002302224 A CA 002302224A CA 2302224 A CA2302224 A CA 2302224A CA 2302224 C CA2302224 C CA 2302224C
- Authority
- CA
- Canada
- Prior art keywords
- tissue adhesive
- adhesive according
- fibrinogen
- tissue
- inhibitor
- 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.)
- Expired - Lifetime
Links
- 239000003106 tissue adhesive Substances 0.000 title claims abstract description 116
- 108010049003 Fibrinogen Proteins 0.000 title claims abstract description 38
- 102000008946 Fibrinogen Human genes 0.000 title claims abstract description 38
- 229940012952 fibrinogen Drugs 0.000 title claims abstract description 38
- 239000003602 elastase inhibitor Substances 0.000 claims abstract description 27
- 229940122858 Elastase inhibitor Drugs 0.000 claims abstract description 20
- 230000003480 fibrinolytic effect Effects 0.000 claims abstract description 18
- 238000002360 preparation method Methods 0.000 claims abstract description 4
- 239000003112 inhibitor Substances 0.000 claims description 29
- 230000001070 adhesive effect Effects 0.000 claims description 23
- 239000000853 adhesive Substances 0.000 claims description 22
- 229940012957 plasmin Drugs 0.000 claims description 22
- 108010088842 Fibrinolysin Proteins 0.000 claims description 21
- 108010039627 Aprotinin Proteins 0.000 claims description 20
- 230000009089 cytolysis Effects 0.000 claims description 20
- ZPNFWUPYTFPOJU-LPYSRVMUSA-N iniprol Chemical group C([C@H]1C(=O)NCC(=O)NCC(=O)N[C@H]2CSSC[C@H]3C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@H](C(N[C@H](C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC=4C=CC(O)=CC=4)C(=O)N[C@@H](CC=4C=CC=CC=4)C(=O)N[C@@H](CC=4C=CC(O)=CC=4)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](C)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](C)C(=O)NCC(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CSSC[C@H](NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](C)NC(=O)[C@H](CO)NC(=O)[C@H](CCCCN)NC(=O)[C@H](CC=4C=CC=CC=4)NC(=O)[C@H](CC(N)=O)NC(=O)[C@H](CC(N)=O)NC(=O)[C@H](CCCNC(N)=N)NC(=O)[C@H](CCCCN)NC(=O)[C@H](C)NC(=O)[C@H](CCCNC(N)=N)NC2=O)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CSSC[C@H](NC(=O)[C@H](CC=2C=CC=CC=2)NC(=O)[C@H](CC(O)=O)NC(=O)[C@H]2N(CCC2)C(=O)[C@@H](N)CCCNC(N)=N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(=O)N2[C@@H](CCC2)C(=O)N2[C@@H](CCC2)C(=O)N[C@@H](CC=2C=CC(O)=CC=2)C(=O)N[C@@H]([C@@H](C)O)C(=O)NCC(=O)N2[C@@H](CCC2)C(=O)N3)C(=O)NCC(=O)NCC(=O)N[C@@H](C)C(O)=O)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@H](C(=O)N[C@@H](CC=2C=CC=CC=2)C(=O)N[C@H](C(=O)N1)C(C)C)[C@@H](C)O)[C@@H](C)CC)=O)[C@@H](C)CC)C1=CC=C(O)C=C1 ZPNFWUPYTFPOJU-LPYSRVMUSA-N 0.000 claims description 20
- 229960004405 aprotinin Drugs 0.000 claims description 19
- 102000013566 Plasminogen Human genes 0.000 claims description 11
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- MDCUNMLZLNGCQA-HWOAGHQOSA-N elafin Chemical compound N([C@H](C(=O)N[C@@H](CCCCN)C(=O)NCC(=O)N1CCC[C@H]1C(=O)N[C@H](C(=O)N[C@@H](CO)C(=O)N[C@H](C(=O)N[C@@H](CCCCN)C(=O)N1CCC[C@H]1C(=O)NCC(=O)N[C@@H](CO)C(=O)N[C@@H]1C(=O)N2CCC[C@H]2C(=O)N[C@H](C(=O)N[C@H](C(=O)N[C@@H](CC(C)C)C(=O)N[C@H](C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@H]2CSSC[C@H]3C(=O)NCC(=O)N[C@@H](CCSC)C(=O)N[C@@H](C)C(=O)N[C@@H](CSSC[C@H]4C(=O)N5CCC[C@H]5C(=O)NCC(=O)N[C@H](C(N[C@@H](CCCCN)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CSSC[C@H](NC(=O)[C@H](CCCNC(N)=N)NC(=O)[C@H](CC(N)=O)NC(=O)[C@H]5N(CCC5)C(=O)[C@H]5N(CCC5)C(=O)[C@H](CC(N)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CCSC)NC(=O)[C@H](C)NC2=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(O)=O)C(=O)N4)C(=O)N[C@@H](CSSC1)C(=O)N[C@@H](CCC(O)=O)C(=O)NCC(=O)N[C@@H](CO)C(=O)N3)=O)[C@@H](C)CC)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](C(C)C)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CCC(N)=O)C(O)=O)[C@@H](C)CC)[C@@H](C)CC)[C@@H](C)CC)[C@@H](C)O)C(C)C)C(C)C)C(=O)[C@@H]1CCCN1C(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](C)N MDCUNMLZLNGCQA-HWOAGHQOSA-N 0.000 claims description 2
- YMDXZJFXQJVXBF-STHAYSLISA-N fosfomycin Chemical compound C[C@@H]1O[C@@H]1P(O)(O)=O YMDXZJFXQJVXBF-STHAYSLISA-N 0.000 claims description 2
- 229920000159 gelatin Polymers 0.000 claims description 2
- 239000008273 gelatin Substances 0.000 claims description 2
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- 235000011852 gelatine desserts Nutrition 0.000 claims description 2
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- 235000019364 tetracycline Nutrition 0.000 claims description 2
- 150000003522 tetracyclines Chemical class 0.000 claims description 2
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 claims 1
- 229940126575 aminoglycoside Drugs 0.000 claims 1
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- GYDJEQRTZSCIOI-LJGSYFOKSA-N tranexamic acid Chemical compound NC[C@H]1CC[C@H](C(O)=O)CC1 GYDJEQRTZSCIOI-LJGSYFOKSA-N 0.000 claims 1
- 229960000401 tranexamic acid Drugs 0.000 claims 1
- 150000003952 β-lactams Chemical class 0.000 claims 1
- 210000001519 tissue Anatomy 0.000 description 15
- BWGVNKXGVNDBDI-UHFFFAOYSA-N Fibrin monomer Chemical compound CNC(=O)CNC(=O)CN BWGVNKXGVNDBDI-UHFFFAOYSA-N 0.000 description 13
- 230000020764 fibrinolysis Effects 0.000 description 13
- 108010073385 Fibrin Proteins 0.000 description 12
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- 238000000034 method Methods 0.000 description 12
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- 108010080379 Fibrin Tissue Adhesive Proteins 0.000 description 10
- 210000004369 blood Anatomy 0.000 description 8
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- 206010052428 Wound Diseases 0.000 description 7
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- 230000009471 action Effects 0.000 description 7
- 229940075469 tissue adhesives Drugs 0.000 description 7
- 208000007536 Thrombosis Diseases 0.000 description 6
- 230000002401 inhibitory effect Effects 0.000 description 6
- 230000001965 increasing effect Effects 0.000 description 5
- 230000002028 premature Effects 0.000 description 5
- 238000007789 sealing Methods 0.000 description 5
- 239000000126 substance Substances 0.000 description 5
- 238000011282 treatment Methods 0.000 description 5
- 102000015081 Blood Coagulation Factors Human genes 0.000 description 4
- 108010039209 Blood Coagulation Factors Proteins 0.000 description 4
- 108060005987 Kallikrein Proteins 0.000 description 4
- 102000001399 Kallikrein Human genes 0.000 description 4
- 108010067372 Pancreatic elastase Proteins 0.000 description 4
- 102000016387 Pancreatic elastase Human genes 0.000 description 4
- 108010001014 Plasminogen Activators Proteins 0.000 description 4
- 102000001938 Plasminogen Activators Human genes 0.000 description 4
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 4
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- 229940127126 plasminogen activator Drugs 0.000 description 3
- 108010000196 Factor XIIIa Proteins 0.000 description 2
- 241000700605 Viruses Species 0.000 description 2
- 230000004913 activation Effects 0.000 description 2
- 238000004026 adhesive bonding Methods 0.000 description 2
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- 239000003527 fibrinolytic agent Substances 0.000 description 2
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- 125000004042 4-aminobutyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])N([H])[H] 0.000 description 1
- FDQGNLOWMMVRQL-UHFFFAOYSA-N Allobarbital Chemical class C=CCC1(CC=C)C(=O)NC(=O)NC1=O FDQGNLOWMMVRQL-UHFFFAOYSA-N 0.000 description 1
- 108090000317 Chymotrypsin Proteins 0.000 description 1
- 208000032544 Cicatrix Diseases 0.000 description 1
- 241000282412 Homo Species 0.000 description 1
- ZDXPYRJPNDTMRX-VKHMYHEASA-N L-glutamine Chemical compound OC(=O)[C@@H](N)CCC(N)=O ZDXPYRJPNDTMRX-VKHMYHEASA-N 0.000 description 1
- 101100281518 Neurospora crassa (strain ATCC 24698 / 74-OR23-1A / CBS 708.71 / DSM 1257 / FGSC 987) fox-2 gene Proteins 0.000 description 1
- 241000283973 Oryctolagus cuniculus Species 0.000 description 1
- 108091005804 Peptidases Proteins 0.000 description 1
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- 241000022563 Rema Species 0.000 description 1
- 102000004002 Secretory Leukocyte Peptidase Inhibitor Human genes 0.000 description 1
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- 108090000631 Trypsin Proteins 0.000 description 1
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- 102000003990 Urokinase-type plasminogen activator Human genes 0.000 description 1
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- ZDXPYRJPNDTMRX-UHFFFAOYSA-N glutamine Natural products OC(=O)C(N)CCC(N)=O ZDXPYRJPNDTMRX-UHFFFAOYSA-N 0.000 description 1
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Classifications
-
- 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
Abstract
A fibrinogen-based tissue adhesive is disclosed which comprises an elastase inhibitor. The fibrinogen-based tissue adhesive may be used for producing a preparation or an application device to be used in fields with high fibrinolytic activity, in particular in urology.
Description
Fibrinogen-Based Tissue Adhesive The invention relates to a fibrinogen-based tissue adhesive.
Fibrinogen-based tissue adhesives which can also be called fibrin adhesives, in their adhesive action imitate the final phase of blood coagulation. In this instance, fibrinogen is cleaved into fibrin monomers by the action of thrombin which mostly is added to the fibrinogen solution during the glueing procedure, which, however, is also present in every wound. The fibrin monomers agglomerate spontaneously to arranged fiber-type structures called fibrin. This fibrin monomer aggregate then is further stabilized under the action of factor XIIIa by covalent cross linking. At this, in a transamidizing reaction, peptide bonds form between specific glutamine and lysine side chains of the fibrin monomers. The factor XIIIa which likewise is cleaved by thrombin from inactive factor XIII is an active transamidase and, on account of its action, it is also called "fibrin-stabilizing factor".
Although, when applying a tissue adhesive, in principle the same processes occur as in "natural" blood coagulation, in a tissue adhesive the participating components and factors are more concentrated by a multiple than in blood. Due to this, the coagulation of blood also occurs much more rapidly, i RCV E~Y:900-~5 METCAt.FF . '?-'04_ p : 6:3,AM : +43 1 X12 98 t~5-. :# 3 and the achieved tissue sealing or the formed blood clot are much safer and also more stable.
A prerequisite for the breakthro~xgh of the fibrin adhesives at the end of the 70~s was the progress in the fractionation and purification of blood coagulation factors. Because of this it was possible to prepare the natural coagulation factors rwith such purity and at such a concez~tratzan as is required for an efficient tissue adhesion. The first commercially' available tissue adhesives were launched an the market at the end of the 70~s, and since then they have proven suitable in a large number of Fossible fields df application;
primarily in those fields in which the use of cvzzventianal surgi.ca7. technidues repeatedly have resulted i.n sevexe pxob~.ems, e.g. with .severe hemorrhages, When glueing nerves or when inner organs, such as the liver and spleen, were torn.
A further advantage of a fibrin adhesive in contrast to sutures using,needle az~d thread resides in that the tissue or organ to be treated is not addit~.onally damaged by the sewing procedure, and therefore, when using tissue adhesives based on fibrinogen, there are much fewer complications and less obtrusive scars than with Conventional surgical sutures. Bexides the optimum adhesi~~re effect which comprises a high load bearing capacity and a high inner strength~of the sealing as well as a c~aod adherence -a-il RCV BY:~00-56 METCALFE . Z-Z4.- p : F_;::3E~A!1~1 : +4.3 1 512 98 05-~ ;# 4.
o~ the adhesive to the wound or tissue surfaces, respectively, also the processes immediately following adhesion are essential for optimizing tissue adhesives (cf . A'T-B-359 652 arid 359 653 ) . Aznang them are the control of the durability of the sealant within the body as well as their capability of being absorned and the adhesive s properties of enhancing wound healing.
Therefore, for a tissue adhesive not only the rapid and secure sealing effect is of decisive importance but also that the sealant formed or the blood clot formed, respectively, dissolve again in the body within a certain period of tune arid the wound completely heals up as a consequence of the complete absorpt~.on of the formed clot.
~n this connection, it is necessary to also control the (endagenous) process of dissolving the formed blood clot, i.e. the fibrinolysis, by optimizing the tissue adhesive.
In fibrinolysis, the fibrin present in the blood clot formed is degraded and/or removed, and thereby the blood clot is dissolved. At .first, under the influence of intrinsic or extrinsic plasminogen activators, such as blood coagulation factors Xz and KzI, pz~ekallikrein, urokinase or t-PA, the fibrinoJ.ytically active plasmin is farmed fxom the inactive plasminogen, said plasmin also cleaving fibrinogen and the blood coagulation i'.~
RCV BY:900-55 '11ETCALFE . ~-24- O : 6:3EiA!~f : +43 1 W'> :~S 05-~ :#~ 5 factors v and viii in addition to fibrin.
Since the endogenous fibrinolysis prace$raes mostly start immediately after formation of a clot and thus there is a risk that an existing tissue sealant will not adhere good enough or that a clot farmed will become destabilized too early, it has become a rule in tissue glue~.ng to provide for the addition of a plasmin .inhibitor or a plasminogen activator .inhibitor so as to inhibit the action of plasmin directly or indirectly, to thus protect the sealant, primarily in its initial phase, from a premature tibrinolysis. With the concentration of the inhibitor also a targeted control of the dissolution times (lysis times? of the clot formed or of the sealant, respectively, is~ possible.
The larger the amount of inhibitor provided, the more stable the clot relative to fibrinolysis, i.e., the longer wi21 this clot rema,i.n stable and the longer will it take for the adhesive to be completely absorbed.
Thus, when using the fibrino3.ysis inhibitor, an optimum balance must be found between preventing premature fibrinolysis $nd an as rapid as possible wound healing process.
lri the commercia~.~.y available tissue adhesives, aprotinin is used as the plasrnin inhibitor, which is al;;o called bovine basic pancreatic trypsin inhibitor.
Aprotinin is a polyvalent proteinase (kallikrein) inhibitor axed inhibits coagulation factors ICIIa, XIa, ii RCV', BY :900-55 METC.~1LFE . ~~-fig.- 0 : 6 : 36AM : +~~.3 1 512 9F3 05~
vllza as we~.7. as, primarily, plasmin and plasmin activators, but also trypsin, chymotrypsin and kallikrein.
Previously, aprotinin I~a.s mainly been produced Pram cattle. Due to the problems involved in the use of bovine material in medicaments which ara used fox the treatment of humans, however, more and more frequently recombinantly prepared aprotinin is being used.
In tissue sealants, apxotinin is uses in an amount of fxom 20 to 3000'kallikrein inactivator units (KIU)/ml tissue adhesive as a rule, its optimum concentration depending on the fibrinolytic activity of the respective tissue. Howeverr it has been shown that mainly in tissues with high fibrinolytic activity, the fibrinolysis-inhibitory action of aprotinin can be eontr~ol.lerl to a very li.mzted extent anly, despite the use of high aprotinin concentrations, and thus undesired, early lytic processes may occur.
Thus, it is an object of the present invention to provide a tissue adhesive with which the disadvantages of the prior art are overcome and with which also primarily with tissue sealants in wounds exhibiting a high plasmin activity, a satisfactory and reliable protection against a~ premature fibrinolysis will be ensuxed, whexein the quality of.the adhaszon must not be negat~.vely affected.
According to the invention, this object is achieved ,n RCV, BY : 90,0-55 METC.ALFE . 2-24 - 0 : g ::3(=,A11 : +.~.3 1 12 9f3 Oa-~ : #
by a fibrinogen-based tissue adhesive which is characLcrized in that ~.t comprises az~ admixed elastase inhibitor. For, surprisingly, it has been shown that the fibrinolysis process cannot only be prevented by inhibiting plasmin or by inhibiting the activation of plasminogen to plasmin, but also by elastase inhibitors or by inhibitors whose fibr'inolysis-inhibitixt,g action is mainly based on a noz~-plasmin fibrinolysis mechanism, respectively. For reasons of simplicity, such non-plasminogen fibx~.n,olysis inhibitors are encompassed by the term "ele.stase inhibitor" for the purposes bf the present invention. xt has been speculated that besides the plasmin-mediated f:ibrinolysis, also further fibrinolytic processes might exist which are not based on plasmin (e. g_ a lysosomal process; cf. Siman et al., BLOOD ~2 (B) (2933), pp.
24,4-2422), and which cannot be substantially inhibited by aprotinin; yet, it has also been shown that this non-plasmin fibrinolytic pathway could not be inhibited by specific elastase~inhibiting peptides, such as N-metho~cy-swecinyl-L-alanyl-L-prolyl-v-valanyl chloromethyl ketone (AAPVCK), either (cf. Simon et al.). xt was the more surprising teat it could be found out in the course of the present invention that inhibitors which do not have any (substantial) p~.asmin-or plasminogen activator-inhibiting activities, i.e.
the elastase inY~~,b~.tvrs of the invention, can ensure a r ~i RC4' BY : 900 -55 ME'1'CALFF . ~-~>4~- 0 : 6 : ~37AM : -H4.3 1 ;1~ 9t3 (u;-~ :
~ B
very we7.1 controllable lytic praaess of the clot far~,ed both in vitro and in vivo. This proved particularly suitable in tissues with increased fibrinolytic activity in which they can prevent early lysis even at moderate concentrations.
Premature lysis also plays a role in tissues with higk~ fibrinolytic activity, primarily within the first' time after the sealing ha:s been made, since premature lysis may lead to a (partial.? detachment of the sealant and, thus, to rebleeding.
Furthexmoxe, it has been shown that the elastase inhibitor to be used according to the invention ~.n the tissue adhesive exhihited its fibrinolytic activity not only in combination with conventional inhibitors acting on p7.asmin, but that even the er_tire fibrinolysis inhibition can be ensured by the elastase inhibitor alone. One particular embodiment of the present invention thus consists in that the tissue adhes~.ve does not contain any further active components besides fibrinogen, the elastase inhbitor, and, optiona~.ly, faCtar XZZY.
zn the present adhesive, the fibrinogen concentration corresponds to that of known t~.ssue adhesives and, as a rule, should be at least more than 50 rng, in particular more than ?0-80 mg of fibrinogen/ml, i.e. at least approximately the 2o-fold of the fibrinogen concentration in blood (~-4 mg/ml}.
,i RCV BY : 9U0-55 METCALFE . 2-24- U : 6: 3 r A~1 : +43 1 t>1'~ 98 05-i : #y g Preferably, the fibrinogen is present in a further purif~.ed form as compared to the cryoprecipitate.
preferred elastase inhibitors within tha scope of the.present invention are selected from the group of eglin, elastase-al-proteinase inhibitor, a~-antiprotease, elafin, leukocyte protease inhibitor, in particular a leukocyte fraction, prefexab3.y a granulocyte-derived fraction, or human secretory leukoprotease inhibitor, or mixtures thereof. As the leukocyte fraction, e.g., a cell lysate, in particular one derived from human cells, may be used. Other elastase- or other fibrinolyeis inhibitors which do not act on plasmin may be assayed by the skilled artisan far their usefulness in the tissue adhesive of the invention in a simple manner by means of the assaying systems disclosed in the Examples or by applying the elastar5e inhibiting assays known from the prior art.
The preferred elastase inhibitors include aJ.so va~r~.ous derivatives of the elastase inhibitors of the invention, e.g, fragments or forms of these inhibitors which have been modified chemically or by (recombinant) protein design, wherein, however, it is, of course, always necessary that these derivatives have the qualitative elastase-inhibitor property of the basic inhibitor.
preferably, the tissue adhesive according to the invention is exclusively comprised of human proteins, _ g _ i RCV BY : 9O0-55 h1ETCA1_,F'E . 2-24- t~ : E; : 37Ah1 : +4:3 1 512 98 05-> : #
wherein also recombinantly prepared humain proteins are to be understood as being "human prote,ins~~. preferably, therefore, the proteins used in the tissue adhesive are prepared either from blood, plasma, cryoprecipitate, or from a recombinant ceJ.~, cu7.ture.
A particularly preferred tiggue adhesive is characterized in that it is exclusively cpmposed from human blood or plasma proteins.
The ratio of the amounts of elastase inhibitor to mg of fibrinogen preferably is from 2:100 to i:150,~00, preferably from 1:500 to 1:110,000. Expressed in units of inhibitor to g of fibrinogen, prefex'ably at least i.0-6 U/g fibrinogen are admixed to the tissue adhesive.
Particularly preferred As a range of between 10-3 and ~/g of fibrinogen. The amount of inhibitor admixed in the tissue adhesive of the invention, which inhibitor may also naturally be present in blood or plasma, rcspecti~rely, preferably is at least 2ox, in particular at least 50x higher than its physiological concentration in blood or plasma, respectively, the tissue adhesive of the invention may, e.g., be composed as follows: 75-115 mg/ml clottable protein, 50-110mg/ml, preferably 70-110 1~g/ml, thereof.being fibrinogen; optionally 1-50, preferably 10-50, TCT
factor xllz/ml,. .As the a.nhibitox', e.g. eglin may be admixed in an amount of between 1-100 ~Cg/ml Qr a~-antiprotease at 0.01-1 U/ml. As a. rule it will be .. g _ RCV BY : 90O - u5 ME:TC E1LFF . ~ _ ~oq. _ f> : f; : ~?A~1 : +4:3 1 0 l? 9r3 05~ : ~ 1 1 sufficient to admix the elastase inhibitor in an amount which corresponds to th~ fibrinolysis-inhibiting activity of aprotinin in known tassue adhesives.
Depending on the purpose of tha sealing, the adhesive according to the invention may contain plasminogen or may be free from pla~smir~ogen. Tf plasminogen is contained, it should be contained in an amount of at .east 0.0001 mg/mg of fibrinogen,, preferably more than 0.00., in particular mare than 0.01. with the presence of plasminogen in the tissue adhesive, based on ita activation to plasmin, it is also possible to def~.z~e the fibrinolysis properties of the tissue adhesive even more clearly.
On the other hand, the tissue adhe$ive, in a further preferred embodiment, does r_r~t contain any plasminogen at all or contains only a small amount thereof, respectively.
As has been mentioned, the presence of the elastase inhibitor as the only fibrinolysis z.xihibitor in a tissue adhesive surprisingly suffices for the functionality of the adhesir~-e according to the inver_tion. Preferably, however, also a plasmin iz~hxbitor or a plasmin activator inhibitor is used besides the elastase inhibitor, which also contributes to a better control of lysis, of absorption, and thus of wound-heal~.x~g. Preferred plasmin inhibitors or plasmi.nogen activator inhibitors are primarily RCS: Bl':900-:~;, ME'~CALFE . 2-24- 0 : 8:.'~BAM : +4-:3 1 512 ,)fl Ou-> :#12 aprotinin, but also a~-macroglobulin, cx1-antitrypsin, E-amino-caproic acid, tranexamic arid or mixtures of these substances. Although i.n same instances authors have also ascribed arl-antitrypsin, e.g., certain actions on elastase, the substances mentioned here are viewed as plasmin or plasminogen activators, respectively, since this is the primary activity exhibited by these substances in the present field.
Fibrinogen-based tissue adhesives which can also be called fibrin adhesives, in their adhesive action imitate the final phase of blood coagulation. In this instance, fibrinogen is cleaved into fibrin monomers by the action of thrombin which mostly is added to the fibrinogen solution during the glueing procedure, which, however, is also present in every wound. The fibrin monomers agglomerate spontaneously to arranged fiber-type structures called fibrin. This fibrin monomer aggregate then is further stabilized under the action of factor XIIIa by covalent cross linking. At this, in a transamidizing reaction, peptide bonds form between specific glutamine and lysine side chains of the fibrin monomers. The factor XIIIa which likewise is cleaved by thrombin from inactive factor XIII is an active transamidase and, on account of its action, it is also called "fibrin-stabilizing factor".
Although, when applying a tissue adhesive, in principle the same processes occur as in "natural" blood coagulation, in a tissue adhesive the participating components and factors are more concentrated by a multiple than in blood. Due to this, the coagulation of blood also occurs much more rapidly, i RCV E~Y:900-~5 METCAt.FF . '?-'04_ p : 6:3,AM : +43 1 X12 98 t~5-. :# 3 and the achieved tissue sealing or the formed blood clot are much safer and also more stable.
A prerequisite for the breakthro~xgh of the fibrin adhesives at the end of the 70~s was the progress in the fractionation and purification of blood coagulation factors. Because of this it was possible to prepare the natural coagulation factors rwith such purity and at such a concez~tratzan as is required for an efficient tissue adhesion. The first commercially' available tissue adhesives were launched an the market at the end of the 70~s, and since then they have proven suitable in a large number of Fossible fields df application;
primarily in those fields in which the use of cvzzventianal surgi.ca7. technidues repeatedly have resulted i.n sevexe pxob~.ems, e.g. with .severe hemorrhages, When glueing nerves or when inner organs, such as the liver and spleen, were torn.
A further advantage of a fibrin adhesive in contrast to sutures using,needle az~d thread resides in that the tissue or organ to be treated is not addit~.onally damaged by the sewing procedure, and therefore, when using tissue adhesives based on fibrinogen, there are much fewer complications and less obtrusive scars than with Conventional surgical sutures. Bexides the optimum adhesi~~re effect which comprises a high load bearing capacity and a high inner strength~of the sealing as well as a c~aod adherence -a-il RCV BY:~00-56 METCALFE . Z-Z4.- p : F_;::3E~A!1~1 : +4.3 1 512 98 05-~ ;# 4.
o~ the adhesive to the wound or tissue surfaces, respectively, also the processes immediately following adhesion are essential for optimizing tissue adhesives (cf . A'T-B-359 652 arid 359 653 ) . Aznang them are the control of the durability of the sealant within the body as well as their capability of being absorned and the adhesive s properties of enhancing wound healing.
Therefore, for a tissue adhesive not only the rapid and secure sealing effect is of decisive importance but also that the sealant formed or the blood clot formed, respectively, dissolve again in the body within a certain period of tune arid the wound completely heals up as a consequence of the complete absorpt~.on of the formed clot.
~n this connection, it is necessary to also control the (endagenous) process of dissolving the formed blood clot, i.e. the fibrinolysis, by optimizing the tissue adhesive.
In fibrinolysis, the fibrin present in the blood clot formed is degraded and/or removed, and thereby the blood clot is dissolved. At .first, under the influence of intrinsic or extrinsic plasminogen activators, such as blood coagulation factors Xz and KzI, pz~ekallikrein, urokinase or t-PA, the fibrinoJ.ytically active plasmin is farmed fxom the inactive plasminogen, said plasmin also cleaving fibrinogen and the blood coagulation i'.~
RCV BY:900-55 '11ETCALFE . ~-24- O : 6:3EiA!~f : +43 1 W'> :~S 05-~ :#~ 5 factors v and viii in addition to fibrin.
Since the endogenous fibrinolysis prace$raes mostly start immediately after formation of a clot and thus there is a risk that an existing tissue sealant will not adhere good enough or that a clot farmed will become destabilized too early, it has become a rule in tissue glue~.ng to provide for the addition of a plasmin .inhibitor or a plasminogen activator .inhibitor so as to inhibit the action of plasmin directly or indirectly, to thus protect the sealant, primarily in its initial phase, from a premature tibrinolysis. With the concentration of the inhibitor also a targeted control of the dissolution times (lysis times? of the clot formed or of the sealant, respectively, is~ possible.
The larger the amount of inhibitor provided, the more stable the clot relative to fibrinolysis, i.e., the longer wi21 this clot rema,i.n stable and the longer will it take for the adhesive to be completely absorbed.
Thus, when using the fibrino3.ysis inhibitor, an optimum balance must be found between preventing premature fibrinolysis $nd an as rapid as possible wound healing process.
lri the commercia~.~.y available tissue adhesives, aprotinin is used as the plasrnin inhibitor, which is al;;o called bovine basic pancreatic trypsin inhibitor.
Aprotinin is a polyvalent proteinase (kallikrein) inhibitor axed inhibits coagulation factors ICIIa, XIa, ii RCV', BY :900-55 METC.~1LFE . ~~-fig.- 0 : 6 : 36AM : +~~.3 1 512 9F3 05~
vllza as we~.7. as, primarily, plasmin and plasmin activators, but also trypsin, chymotrypsin and kallikrein.
Previously, aprotinin I~a.s mainly been produced Pram cattle. Due to the problems involved in the use of bovine material in medicaments which ara used fox the treatment of humans, however, more and more frequently recombinantly prepared aprotinin is being used.
In tissue sealants, apxotinin is uses in an amount of fxom 20 to 3000'kallikrein inactivator units (KIU)/ml tissue adhesive as a rule, its optimum concentration depending on the fibrinolytic activity of the respective tissue. Howeverr it has been shown that mainly in tissues with high fibrinolytic activity, the fibrinolysis-inhibitory action of aprotinin can be eontr~ol.lerl to a very li.mzted extent anly, despite the use of high aprotinin concentrations, and thus undesired, early lytic processes may occur.
Thus, it is an object of the present invention to provide a tissue adhesive with which the disadvantages of the prior art are overcome and with which also primarily with tissue sealants in wounds exhibiting a high plasmin activity, a satisfactory and reliable protection against a~ premature fibrinolysis will be ensuxed, whexein the quality of.the adhaszon must not be negat~.vely affected.
According to the invention, this object is achieved ,n RCV, BY : 90,0-55 METC.ALFE . 2-24 - 0 : g ::3(=,A11 : +.~.3 1 12 9f3 Oa-~ : #
by a fibrinogen-based tissue adhesive which is characLcrized in that ~.t comprises az~ admixed elastase inhibitor. For, surprisingly, it has been shown that the fibrinolysis process cannot only be prevented by inhibiting plasmin or by inhibiting the activation of plasminogen to plasmin, but also by elastase inhibitors or by inhibitors whose fibr'inolysis-inhibitixt,g action is mainly based on a noz~-plasmin fibrinolysis mechanism, respectively. For reasons of simplicity, such non-plasminogen fibx~.n,olysis inhibitors are encompassed by the term "ele.stase inhibitor" for the purposes bf the present invention. xt has been speculated that besides the plasmin-mediated f:ibrinolysis, also further fibrinolytic processes might exist which are not based on plasmin (e. g_ a lysosomal process; cf. Siman et al., BLOOD ~2 (B) (2933), pp.
24,4-2422), and which cannot be substantially inhibited by aprotinin; yet, it has also been shown that this non-plasmin fibrinolytic pathway could not be inhibited by specific elastase~inhibiting peptides, such as N-metho~cy-swecinyl-L-alanyl-L-prolyl-v-valanyl chloromethyl ketone (AAPVCK), either (cf. Simon et al.). xt was the more surprising teat it could be found out in the course of the present invention that inhibitors which do not have any (substantial) p~.asmin-or plasminogen activator-inhibiting activities, i.e.
the elastase inY~~,b~.tvrs of the invention, can ensure a r ~i RC4' BY : 900 -55 ME'1'CALFF . ~-~>4~- 0 : 6 : ~37AM : -H4.3 1 ;1~ 9t3 (u;-~ :
~ B
very we7.1 controllable lytic praaess of the clot far~,ed both in vitro and in vivo. This proved particularly suitable in tissues with increased fibrinolytic activity in which they can prevent early lysis even at moderate concentrations.
Premature lysis also plays a role in tissues with higk~ fibrinolytic activity, primarily within the first' time after the sealing ha:s been made, since premature lysis may lead to a (partial.? detachment of the sealant and, thus, to rebleeding.
Furthexmoxe, it has been shown that the elastase inhibitor to be used according to the invention ~.n the tissue adhesive exhihited its fibrinolytic activity not only in combination with conventional inhibitors acting on p7.asmin, but that even the er_tire fibrinolysis inhibition can be ensured by the elastase inhibitor alone. One particular embodiment of the present invention thus consists in that the tissue adhes~.ve does not contain any further active components besides fibrinogen, the elastase inhbitor, and, optiona~.ly, faCtar XZZY.
zn the present adhesive, the fibrinogen concentration corresponds to that of known t~.ssue adhesives and, as a rule, should be at least more than 50 rng, in particular more than ?0-80 mg of fibrinogen/ml, i.e. at least approximately the 2o-fold of the fibrinogen concentration in blood (~-4 mg/ml}.
,i RCV BY : 9U0-55 METCALFE . 2-24- U : 6: 3 r A~1 : +43 1 t>1'~ 98 05-i : #y g Preferably, the fibrinogen is present in a further purif~.ed form as compared to the cryoprecipitate.
preferred elastase inhibitors within tha scope of the.present invention are selected from the group of eglin, elastase-al-proteinase inhibitor, a~-antiprotease, elafin, leukocyte protease inhibitor, in particular a leukocyte fraction, prefexab3.y a granulocyte-derived fraction, or human secretory leukoprotease inhibitor, or mixtures thereof. As the leukocyte fraction, e.g., a cell lysate, in particular one derived from human cells, may be used. Other elastase- or other fibrinolyeis inhibitors which do not act on plasmin may be assayed by the skilled artisan far their usefulness in the tissue adhesive of the invention in a simple manner by means of the assaying systems disclosed in the Examples or by applying the elastar5e inhibiting assays known from the prior art.
The preferred elastase inhibitors include aJ.so va~r~.ous derivatives of the elastase inhibitors of the invention, e.g, fragments or forms of these inhibitors which have been modified chemically or by (recombinant) protein design, wherein, however, it is, of course, always necessary that these derivatives have the qualitative elastase-inhibitor property of the basic inhibitor.
preferably, the tissue adhesive according to the invention is exclusively comprised of human proteins, _ g _ i RCV BY : 9O0-55 h1ETCA1_,F'E . 2-24- t~ : E; : 37Ah1 : +4:3 1 512 98 05-> : #
wherein also recombinantly prepared humain proteins are to be understood as being "human prote,ins~~. preferably, therefore, the proteins used in the tissue adhesive are prepared either from blood, plasma, cryoprecipitate, or from a recombinant ceJ.~, cu7.ture.
A particularly preferred tiggue adhesive is characterized in that it is exclusively cpmposed from human blood or plasma proteins.
The ratio of the amounts of elastase inhibitor to mg of fibrinogen preferably is from 2:100 to i:150,~00, preferably from 1:500 to 1:110,000. Expressed in units of inhibitor to g of fibrinogen, prefex'ably at least i.0-6 U/g fibrinogen are admixed to the tissue adhesive.
Particularly preferred As a range of between 10-3 and ~/g of fibrinogen. The amount of inhibitor admixed in the tissue adhesive of the invention, which inhibitor may also naturally be present in blood or plasma, rcspecti~rely, preferably is at least 2ox, in particular at least 50x higher than its physiological concentration in blood or plasma, respectively, the tissue adhesive of the invention may, e.g., be composed as follows: 75-115 mg/ml clottable protein, 50-110mg/ml, preferably 70-110 1~g/ml, thereof.being fibrinogen; optionally 1-50, preferably 10-50, TCT
factor xllz/ml,. .As the a.nhibitox', e.g. eglin may be admixed in an amount of between 1-100 ~Cg/ml Qr a~-antiprotease at 0.01-1 U/ml. As a. rule it will be .. g _ RCV BY : 90O - u5 ME:TC E1LFF . ~ _ ~oq. _ f> : f; : ~?A~1 : +4:3 1 0 l? 9r3 05~ : ~ 1 1 sufficient to admix the elastase inhibitor in an amount which corresponds to th~ fibrinolysis-inhibiting activity of aprotinin in known tassue adhesives.
Depending on the purpose of tha sealing, the adhesive according to the invention may contain plasminogen or may be free from pla~smir~ogen. Tf plasminogen is contained, it should be contained in an amount of at .east 0.0001 mg/mg of fibrinogen,, preferably more than 0.00., in particular mare than 0.01. with the presence of plasminogen in the tissue adhesive, based on ita activation to plasmin, it is also possible to def~.z~e the fibrinolysis properties of the tissue adhesive even more clearly.
On the other hand, the tissue adhe$ive, in a further preferred embodiment, does r_r~t contain any plasminogen at all or contains only a small amount thereof, respectively.
As has been mentioned, the presence of the elastase inhibitor as the only fibrinolysis z.xihibitor in a tissue adhesive surprisingly suffices for the functionality of the adhesir~-e according to the inver_tion. Preferably, however, also a plasmin iz~hxbitor or a plasmin activator inhibitor is used besides the elastase inhibitor, which also contributes to a better control of lysis, of absorption, and thus of wound-heal~.x~g. Preferred plasmin inhibitors or plasmi.nogen activator inhibitors are primarily RCS: Bl':900-:~;, ME'~CALFE . 2-24- 0 : 8:.'~BAM : +4-:3 1 512 ,)fl Ou-> :#12 aprotinin, but also a~-macroglobulin, cx1-antitrypsin, E-amino-caproic acid, tranexamic arid or mixtures of these substances. Although i.n same instances authors have also ascribed arl-antitrypsin, e.g., certain actions on elastase, the substances mentioned here are viewed as plasmin or plasminogen activators, respectively, since this is the primary activity exhibited by these substances in the present field.
2'his, of course, also applies for the purposes of the present invention. Moreover, also anti-adhesive additives, e.g. hyaluroaic acid, many be Contained.
A further preferred embodiment of the tissue adhesive according to the invention consists in providing an antibiotic in the adhesive, as has already been suggested in AT-B-369,990. FartiGUlarly preferred antibiotics acre selected from the group of am~,noglycoeides, betalaotams, polypeptides, phosphomycin, tetracyclines or mixtures tk~.ereof. In a further preferred embodiment, the antibiotic is present in the form of a poorly soluble derivative.
Preferably, also factor :VIII is provided in the tissue adhesive according to the invention so that the inner strength of the clot and the strength and durability of the adhesion will be positively influenced. To this end, factor XIII is preferably used in an amount of from 1-50 unitsrml, preferably around LT/ml. Based on fibrinogen, factor xIII ia~ preferably - 17. -ri RCV BY:900-85 METCALPE . 2-24- 0 : 6:38A11 : +43 I u12 98 0.r5-~ :#13 present in a minimum concentration of Q.OO1 U/mg of fibrinogen, in particular at least 0.1 U/mg of fibrinogen, pepending an the type of adhesion or type of tissue, however, the optimum factor XIIz concentration may easily be optimized by any skilled artisan. If an antibiot~.c is present in the tissue adhesive, it is basically racoimnendable to provide somewhat more factor xIII (cf. RT-~-369,90).
Preferably, the tissue adhasive a.ccorc~ing to the invention is free of kininogenic proteins (such as, e.g., kallikrein, etc.), whereby interfering side reactions can be prevented from the beginni.n.g.
According to a preferred embodiment, the tissue adhesive according to the invention is present in combination with a solid surface as a fleece, whereby primarily for large-area wounds, an optimum wound closure and az~ optimum covexirzg will be attained.
Examples of such fleeces have been listed in AT-S 374,367. The solid surface of the fleece thus preferably is a collagen, gelatin o:r polyse.ccharide surface, wherein, of course, a7.so further medica~.ly suitable surfaces which optionally may also have been impregnated far the spe~:ifiC purpose of use may be employed.
It has been shown that according to the invention, with the tissue adhesive comprising an alastase inhibitor a resistance to lysis can be achieved for a II
RCV BY:900-v5 ME'fCALFE . 2-24- 0 : E;:3gA!~9 : +4:3 1 S1'O 9f3 U5-a :#1 l4 period of at least 10 hours, preferably 15 hours, even in an environment with high fibrinolytia activity, Resistance to lysis according to the present invention means that a respective fibrin clot gill not be degraded within a certain period of time, i.e. it remains in existence. Determination of the resistance to lysis is, e.g., effected by a photometric measurement in dependence on time. A preferred tissue adhesive according to the present invention thus.has a resistance to lysis of at least 10 hours, preferably at least Z5 hours, in an environment of high fibrinolytic activity. By "high fibrinolytic activity', e.g., a plasmin activity is understood which is above the physiological plasmin potential. The fibrinalytic potential may, e.g., be expressed by the plasrrinogen Concentration (Cf . , e.g. , Eieririksson et a7_. , Thrombosis Research 16: 301-312; 1979). This property of a.
resistance to lysis may be checked by any skilled artisan by a simple assay, as described in the Exz~mples .
When being app7.ied, the tissue adhesive of the invention preferably is presexxt in solution, yet for storage purposes, either deep treeaing of the solution so that the tissue adhesive is present in deep-frozen form, ox lyophilizing of the adhesive, i.e. pxoviding it in lyophilized form, is reeomrrlendable. By "lyophilized form", of course, only a tissue adhesive -_._~ I, i,l RCV BY:9~0-55 b9ETCALFE . 2-~~4- 0 : G:~;gAM : +43 1 >12 98 05-. :#15 preparation made storable by treeae-drying ie to be understood which in a subsequent reconstitution thereof can be reconstituted almost completely ( i.e. to at 1~ast 8o%j with3.n a Few minutes at 37nC.
The tissue adhesive according to the invention ad~cra.ntageously is present in virus--inactivated dorm.
This inactivation treatment preferably is ensured by a tenside and/or heat treatment, e.g. by a heat treatment in tha solid state, in particular by a vapour treatment according to .EP-0 159 311, or EP-0 519 901 or BP-0 674 531.
Further treatments for inactivation of viruses also comprise a treatment by chemical or cher~ical/physical methods, e.g. with chaotrapic substances according to Wd94/13329, DE 44 34 538 or EP 0 131 740 (solvent), or photoinactivation.
Nanofiltration also is a preferred method for depleting viruses within the scope of the present invention.
Accordi:~g to a preferred embadirnent, the elastase inhibitors admixed according to the invention may also be of recomb~.n.ant origin.
Furthermore, the present invention relates to a tissue adhesive system comprisitlg, as one eamponent thereof, a tissue adhesive according to the invention which comprises an elastase inhibitor.
As a rule, the tissue adhesive system of the --~-KCB; BY ~ 9Q0-55 NETC~ALf E ~ :Z-~4- 0 : Ei:3g.AM : +43 i. 51.1 98 05-~ : #16 invention comprises a thrombin component as a further component, in which thrombin is present either in liquid form or as a lyophilisate capable oi' being reconstituted, wherein the thrombin component may have different concentrations when used in the adhesion, depending on the field of application.
A tissue adhesive system which also falls within the scope of the present invention is characterized in that it comprisess a fi3~rinogen component and a component separate therefrom whi;h contains an elast~ase inhibitor. As a rule, however, it is suitable to provide the fibrinolysis inhibitor component in the fibrinogen component (cf. AT-B-359,652 e.nd 359,653}. By suitable application devices, the inhibitor component may, however, also be supplied separatel~r from the fibrinogen component. Preferably, that component which contains an elastase inhibitor at the same time also contains thrombin, it, again be~.ng passibl~! to provide thzs component anther as a lyaphilisate or as an taptionally deep-frozen) saltzt~.on.
The tissue adhesive systems according to the invention further comprise suitable application devices for the system componez~t(s), In particular, double syringe ssystems asp descri35ed izx EF 0 037 393, EP 0 210 160 or EP 0 292 472, or application devices as described in. EP 0 315 322 or ~P J 669 100 have proven suitable. With these special application devices, also in RCV BY '.900-55 11E'1'CALFE . 2-24-- 0 : 6:39AM : +43 1 522 98 05--those embodiments in which the inhibitor is applied in the thrombin component will yield reziable adhesive results.
The present adhesive is ssuitable far all the fields of known applications possible fox fibrin adhesives. It has, however, proven particularly suitable when providing adhesions in fields with high fibrinolytic activity. Thus, a subject matter of ,the present invention is also the use of the tissue adhesive of the invention or of a tissue adhesive system of the invention, respectively, ~ox producing a preparation or an application device, rcspective~y, to be used in fields with high fibrinolytic activity, i.n particular in urology. A subject matter of the present invention is, moreover, a method for using a tissue adhesive of the invention or a tissue adhesive system of the invention, respectively, in surgery in fields involving high fibrinolytic activity, in particular in urology.
The invention will now be explained in more detail by way of the following Examples and dzawing figures, to which, however, it shall not be restricte3.
Fig. 1 shows the decrease of extinction corresponding to an increasing lysis of the clot a) fibrin adhesive without aprotinin b) fibrin adhesive with aprotinin (1,000 U/ml) c) fibrin adhesive with alpha-1 pI (0.01 U/ml) d) fibrin adhesive with alpha-1 PI (0,001 U/ml) Y 16 _ RCS BY:900-56 METCALFE , 2-24- O : 6::39AM : +4:3 1 5l'.? 98 05~ :#18 e) fibrin adhesive with alpha-1 PI 10.0001 U/ml);
Fig. 2 shows the decrease of extincta.on corresponding to an increasing lysis of the clot a) fibrin adhesive with aprotinin (1,000 U/ml) b) fibrin adhesive with eglin (1 ~cg/tnJ.) c) fibrin adhesive without aprotinin;
Fig. 3 shows the extent of rebleeding, expressed by the inCreaee in the weight of the pre-weighed ps.ds, in a hyper-fiY~rinolytic environment, which had been induced by infusion of t-PA; ar~d Fig. 4 shows the extent of rebreeding, expressed by the increase in the weight of pxe-weighed pads, in an environment with norm*1 fibrinolytic activity, i.e.
without t-PA infusion.
Exam~oles 1. =a vitro-tests !or assaying tho fibriaolysis-fah3bititig *Ctiou of the tis~sv.o adhessvre *ccord3.ug to the iaveatioa (at present considered by applicant to be the best mode of carrying out the invention) In this Example, blocking of the lysis of a tissue adhesive clot by means of eglin or al-antiprotease is shown. In this ixxstance, the tissue adhesive sTIM3 ( IN~tilNO AGy Vienna, AT) ( comprising 70 mg of fibrinogen/ml) is dissolved .in water and subsequently diluted 1:6 with an, 0.9 M NaCl solution.
This tissue adhesive solution is mixed 1:1 With a thrombin solution that had been dissa~.ved in 40 mM
~,i RCV BY:9~U-o5 ;41ETCØ1.FE . '?-24- 0 : 6:39A;~9 : +43 1 X12 98 05-. :#19 CaCl.2 and subsequently had been diluted with a 40 mM
CaCl= /0 . 9 M NaCl (1: 5) solution to 0 _ 1 U/ml, arid pipetted on a micro plate, with 100 ~C1/well being provided.
Various concentrations of tk~e inhibitors were added to each 5 wl of tissue adhesive (Fglin Z-100 ~,g/ml, antiprotease 0.01-1 U/ml?.
For hardening of the adhesive, the microtiter plate was incubated at 37oC for approximately 1.5 hours. The corresponding lysis reagents (a: cell-free supernatant ~rom leukocyte homogenate (3x freezing/thawing) of 5DO,ODO leukocytes/~Cl; b: t-PA 2 mg/ml as positive control; NaCI 0.9% as negative control) were then pipetted onto the clots (100 ul/well). Subsequently, the microtiter wells were measured in the plate photometer at 37~C at a wave length of 405 nm kinetically over night 60x900 s in the Photometer ShT
340 ATTC. The results are illustrated in Figs. 1 and 2, the decrease in the extinction corresponding to the increasing lysis of the clots.
It has been shown that both, with ~ 1 ~tg of eglin/ml and with ~ D.O1 U~ of a~-antiprotease/ml it is possible to prevent the lysis of the fibrin clot which occurs in the a.s~say within 15 hours, which, on the one hand, is a hint to the central. role played by the leukocyte px'oteases for the degradation of the fibrin clot and, on th~ other hard, shows the excellent effect _ 1~ _ ;,i RCV BY:9Q0-55 11ETCAL.FE . '?-24- 0 : E;:.~.pp~t : +43 1 512 ~~8 ()5-~ :#20 of the in~rentive elastase inhibitors ,for preventing this lysis.
Z. In vfvo aeti~rity of the elaetaae inhibitors used accordia~Q to tha iavaatioa Ta determine the importance of leukocyte prateases, in particular elastase inhibitors, with~.n the scope of the present invention, the effect of the tissue adhesive of the invention both in hyper-fibrzz~olyt3c systems and in case of normal f~.brinolytic activity were tested to illustrate hemostasis by means of tissue mdhesive and compared with adhesives without inhibitors, and with an adhesive containing only aprotinin as plasmin inhibitor,. respectively.
2.a) 8yp~erfibrimolysi:
Anesthethised rabbits (2-3 kg) were hepa.rinized (4,OOp Q/kg). Half an hour later, a part of the right J.iver lobe was clamped anal partially resected distal from the clamp. Hemorrhages from larger vessels were stopped by electrocoagulation, and the residual d3,ffuse haemorrhage was sealed by applying tissue adhesive (max. 4 ml) within 200 seconds. ~0 minut~s later, infusion of t-pA (700 U/kg/h) was started, and, the extent of rebleediag was determined fox 2 haura by measuring the increase xz~ wøight of pre-weighed pads.
Iri do~.ng sc~, 3 different tissue adhesives were tested.
a) Tissue adhesive (STIM3) with aprotinin ii RCV BY : 9Q0-55 '.NETCAL:FE , '~-24- 0 : g: øpAh9 : +43 1 51'? 98 Op-~ : #21 (3,OO0 ~/ml> as negative control b? Tissue adhesive (sTIM3) without aprotinin as positive control c) Tissue adhesive (sTIM3) without aprotinin, with eglin (1D ~,g/ml); adhesive accvx'ding to the irve~ntion These adhesives were blind-tested and applied by means of a Duploject~ syringe ($rom TMMUNO, 'Vienna., AT ) .
The results obtained are illustrated in Fig. 3.
2.b) Norm*1 fibrinolytia *ativity In addition to the hyperfibrinolysis model, also the same assays were carried aut without t-PA infusion, yet with increased observation periods of ~ haure. The results obtained are illustrated in ~xg. 4;, It has been shown that both, in the hyperfibx~.nolysis model and with normal fibrinolysis, reduced rebleeding is achieved as compared to conventioz~a~. adhesives, with improved properties as compared to aprotinin, primarily in case of longer lysis periods.
These results prove the excellent effects of the tissue adhesives aCCOrding to the invention, with which an early lyszs of the fibrin adhesive can be prevezzGed, whereby rebleedings could be prevented also in fields with h~.gh fibrinolytie activity.
A further preferred embodiment of the tissue adhesive according to the invention consists in providing an antibiotic in the adhesive, as has already been suggested in AT-B-369,990. FartiGUlarly preferred antibiotics acre selected from the group of am~,noglycoeides, betalaotams, polypeptides, phosphomycin, tetracyclines or mixtures tk~.ereof. In a further preferred embodiment, the antibiotic is present in the form of a poorly soluble derivative.
Preferably, also factor :VIII is provided in the tissue adhesive according to the invention so that the inner strength of the clot and the strength and durability of the adhesion will be positively influenced. To this end, factor XIII is preferably used in an amount of from 1-50 unitsrml, preferably around LT/ml. Based on fibrinogen, factor xIII ia~ preferably - 17. -ri RCV BY:900-85 METCALPE . 2-24- 0 : 6:38A11 : +43 I u12 98 0.r5-~ :#13 present in a minimum concentration of Q.OO1 U/mg of fibrinogen, in particular at least 0.1 U/mg of fibrinogen, pepending an the type of adhesion or type of tissue, however, the optimum factor XIIz concentration may easily be optimized by any skilled artisan. If an antibiot~.c is present in the tissue adhesive, it is basically racoimnendable to provide somewhat more factor xIII (cf. RT-~-369,90).
Preferably, the tissue adhasive a.ccorc~ing to the invention is free of kininogenic proteins (such as, e.g., kallikrein, etc.), whereby interfering side reactions can be prevented from the beginni.n.g.
According to a preferred embodiment, the tissue adhesive according to the invention is present in combination with a solid surface as a fleece, whereby primarily for large-area wounds, an optimum wound closure and az~ optimum covexirzg will be attained.
Examples of such fleeces have been listed in AT-S 374,367. The solid surface of the fleece thus preferably is a collagen, gelatin o:r polyse.ccharide surface, wherein, of course, a7.so further medica~.ly suitable surfaces which optionally may also have been impregnated far the spe~:ifiC purpose of use may be employed.
It has been shown that according to the invention, with the tissue adhesive comprising an alastase inhibitor a resistance to lysis can be achieved for a II
RCV BY:900-v5 ME'fCALFE . 2-24- 0 : E;:3gA!~9 : +4:3 1 S1'O 9f3 U5-a :#1 l4 period of at least 10 hours, preferably 15 hours, even in an environment with high fibrinolytia activity, Resistance to lysis according to the present invention means that a respective fibrin clot gill not be degraded within a certain period of time, i.e. it remains in existence. Determination of the resistance to lysis is, e.g., effected by a photometric measurement in dependence on time. A preferred tissue adhesive according to the present invention thus.has a resistance to lysis of at least 10 hours, preferably at least Z5 hours, in an environment of high fibrinolytic activity. By "high fibrinolytic activity', e.g., a plasmin activity is understood which is above the physiological plasmin potential. The fibrinalytic potential may, e.g., be expressed by the plasrrinogen Concentration (Cf . , e.g. , Eieririksson et a7_. , Thrombosis Research 16: 301-312; 1979). This property of a.
resistance to lysis may be checked by any skilled artisan by a simple assay, as described in the Exz~mples .
When being app7.ied, the tissue adhesive of the invention preferably is presexxt in solution, yet for storage purposes, either deep treeaing of the solution so that the tissue adhesive is present in deep-frozen form, ox lyophilizing of the adhesive, i.e. pxoviding it in lyophilized form, is reeomrrlendable. By "lyophilized form", of course, only a tissue adhesive -_._~ I, i,l RCV BY:9~0-55 b9ETCALFE . 2-~~4- 0 : G:~;gAM : +43 1 >12 98 05-. :#15 preparation made storable by treeae-drying ie to be understood which in a subsequent reconstitution thereof can be reconstituted almost completely ( i.e. to at 1~ast 8o%j with3.n a Few minutes at 37nC.
The tissue adhesive according to the invention ad~cra.ntageously is present in virus--inactivated dorm.
This inactivation treatment preferably is ensured by a tenside and/or heat treatment, e.g. by a heat treatment in tha solid state, in particular by a vapour treatment according to .EP-0 159 311, or EP-0 519 901 or BP-0 674 531.
Further treatments for inactivation of viruses also comprise a treatment by chemical or cher~ical/physical methods, e.g. with chaotrapic substances according to Wd94/13329, DE 44 34 538 or EP 0 131 740 (solvent), or photoinactivation.
Nanofiltration also is a preferred method for depleting viruses within the scope of the present invention.
Accordi:~g to a preferred embadirnent, the elastase inhibitors admixed according to the invention may also be of recomb~.n.ant origin.
Furthermore, the present invention relates to a tissue adhesive system comprisitlg, as one eamponent thereof, a tissue adhesive according to the invention which comprises an elastase inhibitor.
As a rule, the tissue adhesive system of the --~-KCB; BY ~ 9Q0-55 NETC~ALf E ~ :Z-~4- 0 : Ei:3g.AM : +43 i. 51.1 98 05-~ : #16 invention comprises a thrombin component as a further component, in which thrombin is present either in liquid form or as a lyophilisate capable oi' being reconstituted, wherein the thrombin component may have different concentrations when used in the adhesion, depending on the field of application.
A tissue adhesive system which also falls within the scope of the present invention is characterized in that it comprisess a fi3~rinogen component and a component separate therefrom whi;h contains an elast~ase inhibitor. As a rule, however, it is suitable to provide the fibrinolysis inhibitor component in the fibrinogen component (cf. AT-B-359,652 e.nd 359,653}. By suitable application devices, the inhibitor component may, however, also be supplied separatel~r from the fibrinogen component. Preferably, that component which contains an elastase inhibitor at the same time also contains thrombin, it, again be~.ng passibl~! to provide thzs component anther as a lyaphilisate or as an taptionally deep-frozen) saltzt~.on.
The tissue adhesive systems according to the invention further comprise suitable application devices for the system componez~t(s), In particular, double syringe ssystems asp descri35ed izx EF 0 037 393, EP 0 210 160 or EP 0 292 472, or application devices as described in. EP 0 315 322 or ~P J 669 100 have proven suitable. With these special application devices, also in RCV BY '.900-55 11E'1'CALFE . 2-24-- 0 : 6:39AM : +43 1 522 98 05--those embodiments in which the inhibitor is applied in the thrombin component will yield reziable adhesive results.
The present adhesive is ssuitable far all the fields of known applications possible fox fibrin adhesives. It has, however, proven particularly suitable when providing adhesions in fields with high fibrinolytic activity. Thus, a subject matter of ,the present invention is also the use of the tissue adhesive of the invention or of a tissue adhesive system of the invention, respectively, ~ox producing a preparation or an application device, rcspective~y, to be used in fields with high fibrinolytic activity, i.n particular in urology. A subject matter of the present invention is, moreover, a method for using a tissue adhesive of the invention or a tissue adhesive system of the invention, respectively, in surgery in fields involving high fibrinolytic activity, in particular in urology.
The invention will now be explained in more detail by way of the following Examples and dzawing figures, to which, however, it shall not be restricte3.
Fig. 1 shows the decrease of extinction corresponding to an increasing lysis of the clot a) fibrin adhesive without aprotinin b) fibrin adhesive with aprotinin (1,000 U/ml) c) fibrin adhesive with alpha-1 pI (0.01 U/ml) d) fibrin adhesive with alpha-1 PI (0,001 U/ml) Y 16 _ RCS BY:900-56 METCALFE , 2-24- O : 6::39AM : +4:3 1 5l'.? 98 05~ :#18 e) fibrin adhesive with alpha-1 PI 10.0001 U/ml);
Fig. 2 shows the decrease of extincta.on corresponding to an increasing lysis of the clot a) fibrin adhesive with aprotinin (1,000 U/ml) b) fibrin adhesive with eglin (1 ~cg/tnJ.) c) fibrin adhesive without aprotinin;
Fig. 3 shows the extent of rebleeding, expressed by the inCreaee in the weight of the pre-weighed ps.ds, in a hyper-fiY~rinolytic environment, which had been induced by infusion of t-PA; ar~d Fig. 4 shows the extent of rebreeding, expressed by the increase in the weight of pxe-weighed pads, in an environment with norm*1 fibrinolytic activity, i.e.
without t-PA infusion.
Exam~oles 1. =a vitro-tests !or assaying tho fibriaolysis-fah3bititig *Ctiou of the tis~sv.o adhessvre *ccord3.ug to the iaveatioa (at present considered by applicant to be the best mode of carrying out the invention) In this Example, blocking of the lysis of a tissue adhesive clot by means of eglin or al-antiprotease is shown. In this ixxstance, the tissue adhesive sTIM3 ( IN~tilNO AGy Vienna, AT) ( comprising 70 mg of fibrinogen/ml) is dissolved .in water and subsequently diluted 1:6 with an, 0.9 M NaCl solution.
This tissue adhesive solution is mixed 1:1 With a thrombin solution that had been dissa~.ved in 40 mM
~,i RCV BY:9~U-o5 ;41ETCØ1.FE . '?-24- 0 : 6:39A;~9 : +43 1 X12 98 05-. :#19 CaCl.2 and subsequently had been diluted with a 40 mM
CaCl= /0 . 9 M NaCl (1: 5) solution to 0 _ 1 U/ml, arid pipetted on a micro plate, with 100 ~C1/well being provided.
Various concentrations of tk~e inhibitors were added to each 5 wl of tissue adhesive (Fglin Z-100 ~,g/ml, antiprotease 0.01-1 U/ml?.
For hardening of the adhesive, the microtiter plate was incubated at 37oC for approximately 1.5 hours. The corresponding lysis reagents (a: cell-free supernatant ~rom leukocyte homogenate (3x freezing/thawing) of 5DO,ODO leukocytes/~Cl; b: t-PA 2 mg/ml as positive control; NaCI 0.9% as negative control) were then pipetted onto the clots (100 ul/well). Subsequently, the microtiter wells were measured in the plate photometer at 37~C at a wave length of 405 nm kinetically over night 60x900 s in the Photometer ShT
340 ATTC. The results are illustrated in Figs. 1 and 2, the decrease in the extinction corresponding to the increasing lysis of the clots.
It has been shown that both, with ~ 1 ~tg of eglin/ml and with ~ D.O1 U~ of a~-antiprotease/ml it is possible to prevent the lysis of the fibrin clot which occurs in the a.s~say within 15 hours, which, on the one hand, is a hint to the central. role played by the leukocyte px'oteases for the degradation of the fibrin clot and, on th~ other hard, shows the excellent effect _ 1~ _ ;,i RCV BY:9Q0-55 11ETCAL.FE . '?-24- 0 : E;:.~.pp~t : +43 1 512 ~~8 ()5-~ :#20 of the in~rentive elastase inhibitors ,for preventing this lysis.
Z. In vfvo aeti~rity of the elaetaae inhibitors used accordia~Q to tha iavaatioa Ta determine the importance of leukocyte prateases, in particular elastase inhibitors, with~.n the scope of the present invention, the effect of the tissue adhesive of the invention both in hyper-fibrzz~olyt3c systems and in case of normal f~.brinolytic activity were tested to illustrate hemostasis by means of tissue mdhesive and compared with adhesives without inhibitors, and with an adhesive containing only aprotinin as plasmin inhibitor,. respectively.
2.a) 8yp~erfibrimolysi:
Anesthethised rabbits (2-3 kg) were hepa.rinized (4,OOp Q/kg). Half an hour later, a part of the right J.iver lobe was clamped anal partially resected distal from the clamp. Hemorrhages from larger vessels were stopped by electrocoagulation, and the residual d3,ffuse haemorrhage was sealed by applying tissue adhesive (max. 4 ml) within 200 seconds. ~0 minut~s later, infusion of t-pA (700 U/kg/h) was started, and, the extent of rebleediag was determined fox 2 haura by measuring the increase xz~ wøight of pre-weighed pads.
Iri do~.ng sc~, 3 different tissue adhesives were tested.
a) Tissue adhesive (STIM3) with aprotinin ii RCV BY : 9Q0-55 '.NETCAL:FE , '~-24- 0 : g: øpAh9 : +43 1 51'? 98 Op-~ : #21 (3,OO0 ~/ml> as negative control b? Tissue adhesive (sTIM3) without aprotinin as positive control c) Tissue adhesive (sTIM3) without aprotinin, with eglin (1D ~,g/ml); adhesive accvx'ding to the irve~ntion These adhesives were blind-tested and applied by means of a Duploject~ syringe ($rom TMMUNO, 'Vienna., AT ) .
The results obtained are illustrated in Fig. 3.
2.b) Norm*1 fibrinolytia *ativity In addition to the hyperfibrinolysis model, also the same assays were carried aut without t-PA infusion, yet with increased observation periods of ~ haure. The results obtained are illustrated in ~xg. 4;, It has been shown that both, in the hyperfibx~.nolysis model and with normal fibrinolysis, reduced rebleeding is achieved as compared to conventioz~a~. adhesives, with improved properties as compared to aprotinin, primarily in case of longer lysis periods.
These results prove the excellent effects of the tissue adhesives aCCOrding to the invention, with which an early lyszs of the fibrin adhesive can be prevezzGed, whereby rebleedings could be prevented also in fields with h~.gh fibrinolytie activity.
Claims (41)
1. A fibrinogen-based tissue adhesive, wherein said adhesive comprises an admixed elastase inhibitor.
2. The tissue adhesive according to claim 1, wherein the elastase inhibitor is selected from the group of eglin, elastase-.alpha.1-proteinase inhibitor, .alpha.1-antiprotease, leukocyte protease inhibitor, elafin, and mixtures thereof.
3. The tissue adhesive according to claim 2, wherein the leukocyte protease inhibitor is provided as a leukocyte fraction.
4. The tissue adhesive according to claim 3, wherein the leukocyte fraction is a granulocyte-derived fraction.
5. The tissue adhesive according to any one of claims 1 to 3, wherein said tissue adhesive is composed exclusively of human proteins.
6. The tissue adhesive according to any one of claims 1 to 5, wherein said tissue adhesive is composed exclusively of human blood- or plasma proteins.
7. The tissue adhesive according to any one of claims 1 to 6, wherein the elastase inhibitor is contained in an amount ratio of from 1:100 to 1:150,000, based on mg of fibrinogen.
8. The tissue adhesive according to claim 7, wherein the elastase inhibitor is contained in an amount ratio of from 1:500 to 1:110,000 based on mg of fibrinogen.
9. The tissue adhesive according to any one of claims 1 to 8, wherein at least 10 -6 U of elastase inhibitor are contained per g of fibrinogen.
10. The tissue adhesive according to claim 9, wherein between 10 -3 and 10 U of elastase inhibitor are contained per g of fibrinogen.
11. The tissue adhesive according to any one of claims 1 to 10, wherein said tissue adhesive contains plasminogen in an amount of at least 0.0001 mg/mg of fibrinogen.
12. The tissue adhesive according to claim 11, wherein said tissue adhesive contains plasminogen in an amount of at least 0.001 mg/mg of fibrinogen.
13. The tissue adhesive according to claim 12, wherein said tissue adhesive contains plasminogen in an amount of at least 0.01 mg/mg of fibrinogen.
14. The tissue adhesive according to any one of claims 1 to 9, wherein said tissue adhesive does not contain any plasminogen.
15. The tissue adhesive according to any one of claims 1 to 14, wherein said tissue adhesive further comprises a plasmin inhibitor or a plasmin activator inhibitor.
16. The tissue adhesive according to claim 15, wherein said plasmin inhibitor or plasmin activator inhibitor is selected from the group consisting of aprotinin, .alpha.2-macroglobulin, .alpha.1-antitrypsin, .epsilon.-aminocaproic acid, tranexamic acid and mixtures thereof.
17. The tissue adhesive according to any one of claims 1 to 16, wherein said tissue adhesive comprises an antibiotic.
18. The tissue adhesive according to claim 17, wherein said antibiotic is selected from the group consisting of aminoglycosides, betalactams, polypeptides, phosphomycin, tetracycline and mixtures thereof.
19. The tissue adhesive according to any one of claims 1 to 18, wherein said tissue adhesive comprises factor XIII.
20. The tissue adhesive according to claim 18, wherein said tissue adhesive comprises factor XIII in an amount of at least 0.001 U/mg of fibrinogen.
21. The tissue adhesive according to claim 20, wherein said tissue adhesive comprises factor XIII in an amount of at least 0.1 U/mg of fibrinogen.
22. The tissue adhesive according to any one of claims 1 to 21, wherein said tissue adhesive is free from kininogenic proteins.
23. The tissue adhesive according to any one of claims 1 to 22, wherein said tissue adhesive is present in combination with a solid surface as a fleece.
24. The tissue adhesive according to claim 23, wherein the solid surface is a collagen, gelatin or polysaccharide surface.
25. The tissue adhesive according to any one of claims 1 to 24, wherein said tissue adhesive, in an environment of high fibrinolytic activity is resistant to lysis for a period of time of at least 10 hours.
26. The tissue adhesive according to claim 25, wherein said tissue adhesive is resistant to lysis for a period of time of at least 15 hours.
27. The tissue adhesive according to any one of claims 1 to 26, wherein said tissue adhesive is lyophilized.
28. The tissue adhesive according to any one of claims 1 to 26, wherein it is present in solution.
29. The tissue adhesive according to claim 28, wherein the solution is deep-frozen.
30. The tissue adhesive according to any one of claims 1 to 29, wherein it is present in virus-inactivated form.
31. The tissue adhesive according to any one of claims 1 to 30, wherein the elastase inhibitor is of recombinant origin.
32. A tissue adhesive system, wherein said system comprises a tissue adhesive according to any one of claims 1 to 31 and a separate or admixed solvent.
33. The tissue adhesive system according to claim 32, wherein it further comprises a component which comprises thrombin.
34. The tissue adhesive according to claim 33, wherein it further comprises calcium.
35. A tissue adhesive system, wherein said system comprises a fibrinogen component and a component which comprises an elastase inhibitor.
36. The tissue adhesive system according to claim 35, wherein the component which comprises an elastase inhibitor comprises thrombin.
37. The tissue adhesive system according to any one of claims 32 to 36, wherein said system further comprises an application device for the component(s) of the system.
38. The tissue adhesive system according to claim 37, wherein said application device comprises a double-syringe system.
39. Use of the tissue adhesive according to any one of claims 1 to 29 for producing a preparation to be applied in fields with high fibrinolytic activity.
40. Use of the tissue adhesive system according to any one of claims 32 to 37 for producing an application device to be employed in fields with high fibrinolytic activity.
41. The use of the tissue adhesive system according to claim 40, wherein said field with high fibrinolytic activity is urology.
Applications Claiming Priority (3)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| ATA1449/97 | 1997-08-28 | ||
| AT0144997A AT406120B (en) | 1997-08-28 | 1997-08-28 | TISSUE ADHESIVE |
| PCT/AT1998/000202 WO1999011301A1 (en) | 1997-08-28 | 1998-08-26 | Fibrinogen-based tissue adhesive |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CA2302224A1 CA2302224A1 (en) | 1999-03-11 |
| CA2302224C true CA2302224C (en) | 2006-11-14 |
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| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CA002302224A Expired - Lifetime CA2302224C (en) | 1997-08-28 | 1998-08-26 | Fibrinogen-based tissue adhesive |
Country Status (16)
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| US (4) | US7091015B1 (en) |
| EP (1) | EP1007109B1 (en) |
| JP (1) | JP4932082B2 (en) |
| AT (2) | AT406120B (en) |
| AU (1) | AU743181B2 (en) |
| CA (1) | CA2302224C (en) |
| DE (1) | DE59807667D1 (en) |
| DK (1) | DK1007109T3 (en) |
| ES (1) | ES2195375T3 (en) |
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| HU (1) | HUP0104043A3 (en) |
| IL (2) | IL134728A0 (en) |
| NO (1) | NO20000921L (en) |
| PT (1) | PT1007109E (en) |
| SK (1) | SK2562000A3 (en) |
| WO (1) | WO1999011301A1 (en) |
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| AT407484B (en) | 1997-11-12 | 2001-03-26 | Bio Prod & Bio Eng Ag | MEDICINES FOR PROMOTING Wound Healing |
| US7276235B2 (en) | 1998-11-18 | 2007-10-02 | Zlb Behring Gmbh | Tissue glue with improved antiadhesive properties |
| DE10025001A1 (en) * | 2000-05-22 | 2001-11-29 | Aventis Behring Gmbh | Tissue glue with improved anti-adhesive properties |
| AT500670A1 (en) * | 1999-05-19 | 2006-02-15 | Bio & Bio Licensing Sa | DRUGS FOR LOCAL APPLICATION |
| GB9928882D0 (en) * | 1999-12-08 | 2000-02-02 | Zeneca Ltd | Assay method |
| WO2002030445A2 (en) * | 2000-10-13 | 2002-04-18 | Baxter International Inc. | Fibrinogen plus a non-plasmin-acting fibrinolysis inhibitor for the reduction or prevention of adhesion formation |
| CA2594396A1 (en) * | 2005-01-14 | 2006-07-20 | Arblast Co., Ltd. | Sheet-shaped composition utilizing amnion and method of preparing the same |
| KR101307722B1 (en) * | 2006-11-30 | 2013-09-11 | 가부시끼가이샤 비엠지 | Self-degradable adhesive for medical use of two-component reactant system comprising powder-liquid or powder-powder |
| EP2167114A2 (en) | 2007-06-19 | 2010-03-31 | Baxter International Inc. | Fibrin gel for controlled release of pdgf and uses thereof |
| WO2011025957A2 (en) * | 2009-08-28 | 2011-03-03 | Ecole Polytechnique Federal De Lausanne | Tg-aprotinin fusion proteins and matrices thereof |
| US20130202675A1 (en) | 2012-02-03 | 2013-08-08 | Dynasil Biomedical Corporation | Systems and methods for the fabrication of tissue patches |
| 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 |
| US9540548B1 (en) | 2015-08-07 | 2017-01-10 | Xcede Technologies, Inc. | Adhesive compositions and related methods |
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| AT359652B (en) | 1979-02-15 | 1980-11-25 | Immuno Ag | METHOD FOR PRODUCING A TISSUE ADHESIVE |
| 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 |
| AT369990B (en) * | 1981-07-28 | 1983-02-25 | Immuno Ag | METHOD FOR PRODUCING A TISSUE ADHESIVE |
| AT374367B (en) * | 1982-02-23 | 1984-04-10 | Immuno Ag | METHOD FOR PRODUCING A TISSUE ADHESIVE |
| DE3212412C2 (en) * | 1982-04-02 | 1986-01-02 | Dr. Ruhland Nachf. GmbH, 8425 Neustadt | Tissue-bondable collagen wound dressing |
| IL68218A (en) * | 1983-03-23 | 1985-12-31 | Univ Ramot | Compositions for cartilage repair comprising embryonal chondrocytes |
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| AT389815B (en) | 1984-03-09 | 1990-02-12 | Immuno Ag | METHOD FOR INACTIVATING VARIABLE FILTERABLE DISEASERS IN BLOOD PRODUCTS |
| AT382783B (en) | 1985-06-20 | 1987-04-10 | Immuno Ag | DEVICE FOR APPLICATING A TISSUE ADHESIVE |
| DD240334B5 (en) * | 1985-08-27 | 1995-12-14 | Bundesamt Fuer Wehrtechnik Und | Method for producing an autologous tissue adhesive |
| DE3622642A1 (en) * | 1986-07-05 | 1988-01-14 | Behringwerke Ag | ONE-COMPONENT TISSUE ADHESIVE AND METHOD FOR THE PRODUCTION THEREOF |
| SE454480B (en) * | 1986-09-25 | 1988-05-09 | Lars Hammarstrom | BINDING INDUCING COMPOSITION |
| AT388503B (en) | 1987-05-21 | 1989-07-25 | Immuno Ag | SET FOR PROVIDING AND APPLICATION OF A TISSUE ADHESIVE |
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| NL8900943A (en) * | 1989-04-14 | 1990-11-01 | Euro Biopharm Technology B V | PROTEASE INHIBITOR. |
| US5084006A (en) * | 1990-03-30 | 1992-01-28 | Alza Corporation | Iontopheretic delivery device |
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| SE9101853D0 (en) * | 1991-06-17 | 1991-06-17 | Jonas Wadstroem | IMPROVED TISSUE ASHESIVE |
| GB9113164D0 (en) * | 1991-06-18 | 1991-08-07 | Ici Plc | Pharmaceutical agent |
| AT402891B (en) | 1991-06-20 | 1997-09-25 | Immuno Ag | METHOD FOR PRODUCING AN INACTIVATED BLOOD PRODUCT |
| AU6653094A (en) | 1992-12-16 | 1994-07-04 | Immuno Aktiengesellschaft | Process for preparing a virus-safe biological composition |
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| AT400304B (en) | 1994-02-28 | 1995-12-27 | Immuno Ag | DEVICE FOR APPLICATING A MULTI-COMPONENT TISSUE ADHESIVE |
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-
1997
- 1997-08-28 AT AT0144997A patent/AT406120B/en not_active IP Right Cessation
-
1998
- 1998-08-26 AU AU89637/98A patent/AU743181B2/en not_active Expired
- 1998-08-26 AT AT98941134T patent/ATE235269T1/en active
- 1998-08-26 WO PCT/AT1998/000202 patent/WO1999011301A1/en not_active Application Discontinuation
- 1998-08-26 EP EP98941134A patent/EP1007109B1/en not_active Expired - Lifetime
- 1998-08-26 CA CA002302224A patent/CA2302224C/en not_active Expired - Lifetime
- 1998-08-26 IL IL13472898A patent/IL134728A0/en active IP Right Grant
- 1998-08-26 HU HU0104043A patent/HUP0104043A3/en unknown
- 1998-08-26 ES ES98941134T patent/ES2195375T3/en not_active Expired - Lifetime
- 1998-08-26 DK DK98941134T patent/DK1007109T3/en active
- 1998-08-26 JP JP2000508402A patent/JP4932082B2/en not_active Expired - Fee Related
- 1998-08-26 US US09/486,516 patent/US7091015B1/en not_active Expired - Fee Related
- 1998-08-26 PT PT98941134T patent/PT1007109E/en unknown
- 1998-08-26 DE DE59807667T patent/DE59807667D1/en not_active Expired - Lifetime
- 1998-08-26 SK SK256-2000A patent/SK2562000A3/en unknown
-
2000
- 2000-02-24 IL IL134728A patent/IL134728A/en not_active IP Right Cessation
- 2000-02-24 NO NO20000921A patent/NO20000921L/en not_active Application Discontinuation
- 2000-02-25 HR HR20000100A patent/HRP20000100B1/en not_active IP Right Cessation
-
2006
- 2006-05-30 US US11/443,925 patent/US7459295B2/en not_active Expired - Fee Related
-
2008
- 2008-10-16 US US12/252,988 patent/US7892802B2/en not_active Expired - Fee Related
-
2011
- 2011-02-04 US US13/021,551 patent/US8425947B2/en not_active Expired - Fee Related
Also Published As
| Publication number | Publication date |
|---|---|
| DE59807667D1 (en) | 2003-04-30 |
| IL134728A (en) | 2007-09-20 |
| ATE235269T1 (en) | 2003-04-15 |
| US20090041748A1 (en) | 2009-02-12 |
| HRP20000100B1 (en) | 2010-09-30 |
| ATA144997A (en) | 1999-07-15 |
| US7091015B1 (en) | 2006-08-15 |
| US7459295B2 (en) | 2008-12-02 |
| AU743181B2 (en) | 2002-01-17 |
| US20060216280A1 (en) | 2006-09-28 |
| US20110190812A1 (en) | 2011-08-04 |
| EP1007109A1 (en) | 2000-06-14 |
| JP2001514050A (en) | 2001-09-11 |
| NO20000921D0 (en) | 2000-02-24 |
| DK1007109T3 (en) | 2003-06-23 |
| NO20000921L (en) | 2000-04-26 |
| AU8963798A (en) | 1999-03-22 |
| ES2195375T3 (en) | 2003-12-01 |
| HUP0104043A1 (en) | 2002-02-28 |
| HRP20000100A2 (en) | 2000-10-31 |
| IL134728A0 (en) | 2001-04-30 |
| SK2562000A3 (en) | 2000-09-12 |
| AT406120B (en) | 2000-02-25 |
| WO1999011301A1 (en) | 1999-03-11 |
| HUP0104043A3 (en) | 2004-08-30 |
| JP4932082B2 (en) | 2012-05-16 |
| EP1007109B1 (en) | 2003-03-26 |
| US7892802B2 (en) | 2011-02-22 |
| US8425947B2 (en) | 2013-04-23 |
| PT1007109E (en) | 2003-08-29 |
| CA2302224A1 (en) | 1999-03-11 |
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