Stabilized protein preparations for a tissue adhesive

Details Stabilized protein preparations for a tissue adhesive Stabilized protein preparations for a tissue adhesive

2001-07-13

2002-09-10

Tate, Christopher R. (Department: 1651)

Drug, bio-affecting and body treating compositions

Enzyme or coenzyme containing

Hydrolases

C424S529000, C514S002600, C530S381000, C530S382000, C530S383000

Reexamination Certificate

active

06447774

ABSTRACT:

The subject matter of the invention is a tissue adhesive as well as stabilized protein preparations for a tissue adhesive or preparations for parenteral application, which have no significant loss of effect after being stored in a liquid state or after being defrosted following storage in a frozen state and during further storage in a liquid state over a period of several months or years.
It is already known that tissue adhesives can be used to connect human or animal tissue, and that the main components of such tissue adhesives include fibrinogen, Factor XIII and thrombin. These protein preparations require careful stabilization to retain their full effect and their application properties until they are used for gluing tissue during surgery.
Tissue gluing is a method that has already been described at the turn of the century and repeatedly thereafter (S. Bergel: On the effects of fibrin.
Dt. med. Wschr.
35:663-5 (1909); E. P. Cronkite, E. L. Lozner, J. M. Deaver: Use of thrombin and fibrinogen in skin grafting. JAMA 124:976-8 (1944); H. Matras; H. P. Dinges, H. Lassmann, B. Mammoli,
Wr. Med. Wschr.
37:517 (1972); H. Matras, H. P. Dinges, H. Lassmann, B. Mammoli:
J. max. fac. Surg.
1:37 (1973); H. Matras, F. Braun, H. Lassmann, H. P. Ammrer, B. Mammoli: Plasma clot welding of nerves (experimental report),
J. max. fac. Surg.
1:2364 (1973); H. Kudema, H. Matras,
Wiener klin. Wschr.
87:495-496 (1975). Initially, plasma or fibrinogen was still used as powder; later on, purified fibrinogen was used, for example, in the form of a cryoprecipitate.
With the commercial production of fibrinogen-/Factor XIII- and thrombin concentrates since the Seventies, the importance of tissue gluing has increased significantly. It is now used, for example, to support sutures, local haemostasis, the sealing of body cavities to avoid loss of fluid, and for wound care. Tissue gluing with fibrin adhesives is a physiological method and therefore has advantages compared to the synthetic adhesives with respect to compatibility and biodegradability of the adhesive components.
Tissue adhesives are commercially available either as lyophilizates or as frozen preparations. However, after reconstitution or after defrosting, the products are stable in solution for only a few days, because with the highly concentrated fibrinogen-/Factor XIII-solutions, an aggregation and therefore an (for example, proteolytic) inactivation occurs, which makes any further use impossible.
The tissue adhesives that have been described in the literature until now are not yet commercially available and are generally comprised of frozen or freeze-dried components that must be defrosted or dissolved prior to use. To improve the processing, the solubility, the defrosting or the stability of the fibrinogen concentrate, European patent 0,085,923, German patent application 196,17,369 and European patent application 0,856,317 describe the use of chaotropic agents or additives such as arginine or urea or their derivatives or derivatives of benzene, imidazole, pyrozol, furan, thiazole and purine, which generally improve the solubility of proteins. Chaotropic agents in this context are agents that reduce or destabilize the reciprocal effect between proteins or parts thereof and therefore reduce their tendency towards aggregation. It is important to guarantee the stability of the components such as fibrinogen and Factor XIII even in the presence of said chaotropic agents and under the selected conditions. So far, this has not been successful with frozen fibrinogen-/Factor XIII-concentrations that had to be stored for several weeks or months in liquid form after defrosting, or with fibrinogen-/Factor XIII-concentrations that could be stored only in liquid form.
With liquid storage, but especially also with storage in frozen state, the loss of F XIII-activity in the formulations described thus far was so high that in the presence of effective quantities of chaotropic agents, the F XIII-content often clearly drops after only a few weeks or months, often even below the detection limit.
With the formulations in accordance with European patent application 0,856,317, it was shown that tranexaminic acid (AMCA), especially in the presence of chaotropic agents such as arginine and inorganic salts, clearly reduced the F XIII content in the course of storage at −20° (Table 1 b, Batch 1). Storage at 4° Celsius leads to an increase in viscosity in this formulation (Table 1 a, Batch 1), which also rules out a long-term term storage. Thus, these formulations must be considered non-stable in view of the simultaneous stability of fibrinogen and F XIII. Formulations in accordance with DE 196,17,369 also indicate problems in maintaining F XIII-activity (see Table 1, Batch 2 and 2).
Another biological adhesive for human or animal tissue is known from the European patent specification 0,487,713. Said adhesive is stabilized in liquid form at low temperatures. This is supposed to be achieved in that the preparation containing fibrinogen comprises at least one chaotropic agent in a concentration between approximately 0.3 M and 1 M and in that the adhesive is liquid at the storage temperature.
Such fibrinogen concentrate typically comprised about 4 mmol tri-sodium citrate, 240 mmol NaCl, 80 mmol—amino caproic acid (EACA), 240 mmol glycine, 1% polysorbate, 0.6 grams/liter sodium caprolate, 0.5 mol urea, 2,000 KIE/ml aprotinin, if necessary, and a pH of 7.5. The stability was evaluated after only one month, which is very short for a therapeutic preparation. The F XIII-activity was not analyzed. J. Chabbat et al. reported about a fibrinogen concentrate that remains stable in liquid state at 4° Celsius over a period of six months (J. Chabbat, M. Tellier, P. Porte and M. Steinbuch: Properties of a new fibrin glue stable in liquid state. Thromb. Res. 76: 525-533 (1994)). In addition to other formulation components, typically 60 mmol/liter NaCl, 20 mmol/liter EACA and 60 mmol/liter glycine, this concentration comprised 0.5 mol urea or 5% arginine (=0.29 mol). However, the F XIII-strength of this concentration was also not tested.
These liquid formulations, which were described in the European patent specification 0,487,713 and in the literature, are characterized in that the aggregation (polymerization) and thus the increase in viscosity of the concentrated fibrinogen component, is prevented or reduced at refrigeration temperatures. However, Factor XIII, an essential component of fibrinogen concentrates for fibrin glues, is inactivated to a greater or lesser degree under these conditions. In the formulations provided for storage in cooled state in accordance with European patent specification 0,487,713 or the related publication by Chabbat et al. (J. Chabbat, M. Tellier, P. Porte and M. Steinbuch: Properties of a new fibrin glue stable in liquid state. Thromb. Res. 76:525-533 (1994)), the instability of F XIII is therefore a significant problem that is not solved by the proposed formulations (see Table 1, Batches 4-5). Furthermore, the strength of chaotropic agents is relatively high at 0.3 to 1.0 mol/liter, which makes lesser concentrations of chaotropic agents appear desirable (<0.3 mol/liter).
Thus, it can be noted that it was found in the analysis of the stability of fibrinogen/Factor XIII preparations as well as the viscosity of various known fibrinogen/Factor XIII-preparations in refrigerated state (0 to 10° Celsius) or frozen state with subsequent storage in refrigerated state (0 to 10° Celsius) that the previously described formulations do not lead to stable protein preparations. Either the fibrinogen or Factor XIII show a significant reduction in activity during the storage time, or the aggregation of fibrinogen leads to a viscous material that can no longer be applied (see Table 1, Batches 1 to 5).
Thus, the problem to be solved was to develop protein preparations that are liquid and stable over several months, or frozen and stable over several months following defrosting, in which the fibrinogen and/or Factor XIII are stabilized over months o

Affiliated with

Also associated with

Rating

Stabilized protein preparations for a tissue adhesive does not yet have a rating. At this time, there are no reviews or comments for this patent.

If you have personal experience with Stabilized protein preparations for a tissue adhesive, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Stabilized protein preparations for a tissue adhesive will most certainly appreciate the feedback.

Rate now

Comments { 0 }

Profile ID: LFUS-PAI-O-2829182