Fibrinogen-containing preparations and uses thereof

Abstract

The present invention relates to a composition comprised of albumin and clotting proteins, including fibrinogen, and its use, for example, to treat bleeding. Specifically, the composition comprises albumin and one or more clotting proteins, wherein the albumin and one or more clotting proteins are present in a total concentration of at least 90% based on total protein weight, the clotting protein fibronectin is present in an amount less than about 0.5% based on total protein weight or is absent, and the weight ratio of albumin to the clotting protein fibrinogen is at least 1:15, respectively.

Description

[Technical field]

[0001] The present disclosure relates to compositions comprising fibrinogen. In particular, the present invention relates to fibrinogen preparations that contain albumin and very small amounts of fibronectin.

[0002] (References) References that are deemed relevant as background to the subject matter of this disclosure are listed below. -U.S. Patent No. 7,241,603 -U.S. Patent Application No. 2016 / 0015792 -U.S. Patent No. 9,084,728 -U.S. Patent No. 8,962,025 -U.S. Patent No. 10,111,980 -U.S. Patent No. 9,056,092 - International Publication No. 2000 / 009018 - International Publication No. 1992 / 013495 -U.S. Patent Application No. 2020 / 0054720 -Bar, L., et al. The binding of fibrin sealant to collagen is influenced by the method of purification and the cross-linked fibrinogen-fibronectin (heteronectin) content of the "fibrinogen" component, Blood Coagulation & Fibrinolysis:2005,16(2),111-117.

[0003] The acknowledgement of the above references herein does not imply that they are to be construed as being in any way relevant to the patentability of the subject matter of this disclosure. [Background technology]

[0004] Control of bleeding is essential and important in surgical procedures to minimize blood loss. Biological glues / sealants used as tissue adhesives or for controlling hemostasis have been described in the art.

[0005] Fibrinogen plays a key role in the coagulation cascade and is believed to be a key component in a variety of hemostatic and sealing products, including fibrin patches and fibrin sealants, among others. Current fibrinogen products contain, in addition to fibrinogen, other components that are purified from plasma along with fibrinogen. These compounds include, for example, clotting factors such as factor VIII, factor XIII, von Willebrand factor (vWF), and fibronectin, the latter of which has previously been shown to be essential for promoting tissue adhesion (Bar, L., et al., Blood Coagulation & Fibrinolysis: 2005, 16(2), 111-117).

[0006] US Patent No. 7,241,603 describes a method for producing a protein composition comprising fibrinogen and fibronectin, in which a starting solution comprising fibrinogen and fibronectin is treated with a precipitating composition, whereby a precipitate comprising fibrinogen and fibronectin is formed in a single precipitation step, the precipitate formed being optionally further treated by methods known per se.

[0007] US Patent Application No. 2016 / 0015792 relates to a sterile powder composition suitable for medical use comprising thrombin and fibrinogen, and a method for producing the same.

[0008] US Pat. No. 9,084,728 describes a process for making a dry stable hemostatic composition.

[0009] US Pat. No. 8,962,025 describes compositions, methods, and kits for sealing applications.

[0010] US Pat. No. 10,111,980 describes a dry composition containing one or more polyols which upon addition of an aqueous medium forms a substantially homogeneous paste suitable for use in hemostatic treatments.

[0011] U.S. Patent No. 9,056,092 describes a hemostatic pad that includes a bioabsorbable scaffolding material, a lyophilized thrombin powder, a lyophilized fibrinogen powder, and a meltable binder powder, all of the powders disposed on the bioabsorbable scaffolding material.

[0012] WO 2000 / 009018 describes polymerized type I and / or type III collagen-based compositions for medical use as vascular sealants and wound dressings, and their preparation.

[0013] WO 1992 / 013495 describes a fibrinogen-based composition obtained from bovine plasma using polyethylene glycol ("PEG") and glycine precipitation techniques.

[0014] US Patent Application Publication No. 2020 / 0054720 provides methods and kits for large-scale preparation of a sterile stable liquid thrombin composition comprised of glycerol, a stable liquid thrombin composition, and a hemostatic composition.

[0015] Bar, L. et al. (2005) describe fibrinogen preparations and, using experiments with highly purified protein, show that fibronectin is essential to promote the progressive binding of fibrinogen to collagen under the action of activated factor XIII (transglutaminase). Summary of the Invention [Means for solving the problem]

[0016] According to some aspects, the disclosure provides compositions comprising one or more coagulation proteins and albumin, wherein the one or more coagulation proteins and albumin are present at a combined concentration of at least 90% based on total protein weight, the one or more coagulation proteins comprise fibrinogen and fibronectin, wherein fibronectin is at a concentration of less than about 0.5% based on total protein weight or is absent, and the weight ratio of albumin to fibrinogen is at least 1:15, respectively.

[0017] According to some aspects, the present disclosure provides compositions comprising one or more coagulation proteins, albumin, and thrombin, wherein the one or more coagulation proteins and albumin are present at a combined concentration of at least 90% based on total protein weight, the one or more coagulation proteins comprise fibrinogen and fibronectin, wherein fibronectin is at a concentration of less than about 0.5% based on total protein weight or is absent, and the weight ratio of albumin to fibrinogen is at least 1:15, respectively.

[0018] According to some aspects, the disclosure provides an article of manufacture comprising a composition comprising one or more coagulation proteins and albumin, wherein the one or more coagulation proteins and albumin are present at a total concentration of at least 90% based on total protein weight, the one or more coagulation proteins comprise fibrinogen and fibronectin, wherein the fibronectin is at a concentration of less than about 0.5% based on total protein weight or is absent, and the weight ratio of albumin to fibrinogen is at least 1:15, respectively, and the composition is in powder form.

[0019] According to some embodiments, the article of manufacture is a wound dressing.

[0020] According to some further aspects, the present disclosure further provides a composition comprising one or more coagulation proteins and albumin, wherein the one or more coagulation proteins and albumin are present at a total concentration of at least 90% based on total protein weight, the one or more coagulation proteins comprise fibrinogen and fibronectin, the fibronectin being at a concentration of less than about 0.5% based on total protein weight or absent, and the weight ratio of albumin to fibrinogen is at least 1:15, respectively. The present disclosure further provides an article comprising the composition, e.g., for use in a tissue adhesive.

[0021] According to some further aspects, the present disclosure further provides a composition comprising one or more coagulation proteins and albumin, wherein the one or more coagulation proteins and albumin are present at a total concentration of at least 90% based on total protein weight, the one or more coagulation proteins comprise fibrinogen and fibronectin, the fibronectin being present or at a concentration of less than about 0.5% based on total protein weight, and the weight ratio of albumin to fibrinogen is at least 1:15, respectively. The present disclosure further provides an article comprising the composition for use in treating, inhibiting, arresting, or slowing bleeding or any disorder related thereto.

[0022] According to some other aspects, the present disclosure provides a method for treating, inhibiting, preventing or slowing bleeding or any disorder related thereto, comprising topically administering to a wound tissue of a subject in need thereof an effective amount of a composition comprising one or more coagulation proteins and albumin, wherein the one or more coagulation proteins and albumin are present in a total concentration of at least 90% based on total protein weight, the one or more coagulation proteins comprise fibrinogen and fibronectin, the fibronectin is present in an amount of less than about 0.5% based on total protein weight or absent, and the weight ratio of albumin to fibrinogen is at least 1:15, respectively. The present disclosure further provides an article comprising the composition, thereby treating, inhibiting, preventing or slowing bleeding or any disorder related thereto. [Brief description of the drawings]

[0023] In order to better understand the subject matter disclosed herein, and to illustrate how it may be carried out in practice, embodiments will now be described, by way of non-limiting example only, with reference to the accompanying drawings, in which: [Figure 1] FIG. 1 is a graph showing tissue adhesion of exemplary samples as a function of albumin % relative to total protein using a tissue peel force test, with error bars representing standard deviation and p-values ​​calculated by pooled t-test. [Diagram 2] 1 is a bar graph showing tissue peel force as a function of % fibrinogen relative to total protein using the tissue peel force test, with error bars representing standard deviation. [Diagram 3] 1 is a bar graph showing tissue peel force as a function of % Albumin, where the Y-axis shows the average tissue peel force (N / meter), the X-axis shows % Albumin, and the error bars represent the standard deviation. [Figure 4] 1 is an image showing an exemplary wound dressing comprising a fibrinogen composition applied to a resected kidney. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0024] Control of bleeding is essential and important in, for example, surgical procedures to minimize blood loss, reduce post-operative complications, and reduce operative time in the operating room.

[0025] The inventors have surprisingly found that pure fibrinogen preparations containing high amounts of fibrinogen and very low amounts of fibronectin, and sometimes essentially free of fibronectin, when combined with albumin, are highly effective, achieving strong tissue adhesion and also initiating hemostatic activity. This low amount of fibronectin (if present), among other features, distinguishes the fibrinogen preparations of the present disclosure from known fibrinogen preparations.

[0026] Thus, the present disclosure relates to compositions that contain fibrinogen, albumin, and small amounts of fibronectin, and sometimes are essentially free of fibronectin, which may sometimes be considered pure fibrinogen compositions.

[0027] According to some aspects, the present disclosure provides compositions comprising one or more coagulation proteins and albumin, wherein the one or more coagulation proteins and albumin are at a combined concentration of at least 90 w / w% based on total protein weight, the one or more coagulation proteins comprise fibrinogen and fibronectin, wherein fibronectin is present in an amount less than about 0.5 w / w% based on total protein weight or is absent, and the weight ratio of albumin to fibrinogen is at least 1:15, respectively.

[0028] This composition is sometimes referred to herein as a "fibrinogen-comprising composition."

[0029] In the following text, reference to the composition should be understood as also referring to the articles of manufacture, methods and kits disclosed herein, and therefore whenever a feature is provided with respect to the composition, it should be understood as defining the same feature with respect to the articles of manufacture, methods and kits.

[0030] As shown below in Example 1, pure fibrinogen preparations containing small amounts of fibronectin achieved strong tissue adhesion upon addition of albumin, as shown in Example 2, complete hemostasis was achieved following administration of a patch containing a composition described herein.

[0031] It has therefore been suggested that pure fibrinogen preparations containing albumin and small amounts of fibronectin, or sometimes no fibronectin, are highly effective in adhering well to tissues and achieving homeostasis.

[0032] As described herein, the composition includes a coagulation protein. In the context of this disclosure, a "coagulation protein" that is part of the formed clot includes one or more of the plasma proteins.

[0033] Typically, the term "clotting protein" should be understood as not encompassing all enzymes that are activated as part of clot formation. Non-limiting examples of clotting proteins include fibrinogen, fibronectin, alpha 2 antiplasmin, and vitronectin.

[0034] The content of coagulable proteins in the composition can be calculated by any method known in the art. For example, the content of coagulable proteins can be calculated from the measurement of coagulable proteins and total proteins. "Coagulable proteins" can be measured by clotting a diluted sample with thrombin in the presence of calcium ions. The clot is washed and the remaining protein content is measured. Measurements can be made gravimetrically or by dissolving the clot in alkaline urea or other reagents followed by a spectrophotometric protein assay. "Total protein" can be measured by various analytical techniques including, inter alia, ultraviolet spectrophotometry, dye-binding assays such as Coomassie or Bradford, copper-based assays such as biuret, Lowry, and bicinchoninic acid (BCA) assays, and immunological methods.

[0035] The composition also comprises albumin.Albumin content can be calculated by any method known in the art.For example, albumin content can be calculated by dye-binding method, such as bromocresol green and bromocresol purple, electrophoresis, and immunological method, such as immunoturbidimetry.

[0036] In some embodiments, the composition comprises one or more coagulation proteins and albumin at a total concentration of at least about 91% based on total protein dry matter weight, sometimes at least about 92%, sometimes at least about 93%, sometimes at least about 94%, sometimes at least about 95%, sometimes at least about 96%, sometimes at least about 97%, sometimes at least about 97.5%, sometimes at least about 98%, sometimes at least about 98.5%, sometimes at least about 99%, sometimes at least about 99.2%, sometimes at least about 99.4%, sometimes at least about 99.6%, and sometimes at least about 99.8% based on total protein dry matter weight. In some embodiments, the one or more coagulation proteins and albumin are at a total concentration of at least about 95% based on total protein dry matter weight.

[0037] In some embodiments, a composition comprising fibrinogen comprises albumin and one or more coagulation proteins at a total concentration of about 90% to about 100% based on total protein dry matter weight.In some embodiments, a composition comprising fibrinogen comprises albumin and one or more coagulation proteins at a total concentration of about 90% to about 99.5% based on total protein dry matter weight.

[0038] In some other embodiments, the compositions comprising fibrinogen include one or more coagulation proteins and albumin at a total concentration of about 90% to about 99.9% based on total protein dry matter weight, sometimes about 92% to about 99.9%, about 94% to about 99.9%, about 95% to about 99.9%, about 97% to about 99.9%, about 98% to about 99.9%, about 99% to about 99.9%, about 99.2% to about 99.9%, about 99.4% to about 99.9%, or about 99.6% to about 99.9% based on total protein dry matter weight.

[0039] In some embodiments, compositions comprising fibrinogen comprise one or more coagulation proteins and albumin in a total concentration of about 90% based on total protein dry matter weight, sometimes about 92%, sometimes about 93%, sometimes about 94%, sometimes about 95%, sometimes about 96%, sometimes about 97%, sometimes about 97.5%, sometimes about 98%, sometimes about 98.5%, sometimes about 99%, sometimes about 99.2%, sometimes about 99.4%, sometimes about 99.5%, sometimes about 99.6%, sometimes about 99.7%, sometimes about 99.8%, and sometimes about 99.9% based on total protein dry matter weight.

[0040] In some embodiments, compositions comprising fibrinogen comprise one or more coagulation proteins and albumin at a total concentration of 99.5% based on total protein dry matter weight, sometimes at a total concentration of about 99.6%, sometimes at about 99.7%, sometimes at about 99.8%, and sometimes at about 99.9% based on total protein dry matter weight.

[0041] In some embodiments, the one or more coagulation proteins include at least one of fibrinogen, fibronectin, alpha 2 antiplasmin, vitronectin, or any combination thereof.

[0042] In some embodiments, the one or more coagulation proteins comprise fibrinogen, fibronectin, or a combination thereof.

[0043] In some embodiments, the one or more coagulation proteins comprises or is fibrinogen.

[0044] In some embodiments, the fibrinogen-containing composition comprises fibrinogen and albumin at a total concentration of at least about 91% based on total protein dry matter weight, sometimes at least about 92%, sometimes at least about 93%, sometimes at least about 94%, sometimes at least about 95%, sometimes at least about 96%, sometimes at least about 97%, sometimes at least about 97.5%, sometimes at least about 98%, sometimes at least about 98.5%, sometimes at least about 99%, sometimes at least about 99.2%, sometimes at least about 99.4%, sometimes at least about 99.6%, and sometimes at least about 99.8% based on total protein dry matter weight. In some embodiments, fibrinogen and albumin are at a total concentration of at least about 95% based on total protein dry matter weight.

[0045] In some embodiments, the fibrinogen-containing composition comprises fibrinogen and albumin at a combined concentration of about 90% to about 100% based on total protein dry matter weight, hi some embodiments, the fibrinogen-containing composition comprises fibrinogen and albumin at a combined concentration of about 90% to about 99.5% based on total protein dry matter weight.

[0046] In some other embodiments, the compositions comprising fibrinogen contain fibrinogen and albumin at a total concentration of about 90% to about 99.9% based on total protein dry matter weight, sometimes about 92% to about 99.9%, about 94% to about 99.9%, about 95% to about 99.9%, about 97% to about 99.9%, about 98% to about 99.9%, about 99% to about 99.9%, about 99.2% to about 99.9%, about 99.4% to about 99.9%, or about 99.6% to about 99.9% based on total protein dry matter weight.

[0047] In some embodiments, compositions comprising fibrinogen comprise fibrinogen and albumin at a total concentration of about 90% based on total protein dry matter weight, sometimes about 92%, sometimes about 93%, sometimes about 94%, sometimes about 95%, sometimes about 96%, sometimes about 97%, sometimes about 97.5%, sometimes about 98%, sometimes about 98.5%, sometimes about 99%, sometimes about 99.2%, sometimes about 99.4%, sometimes about 99.5%, sometimes about 99.6%, sometimes about 99.7%, sometimes about 99.8%, and sometimes about 99.9% based on total protein dry matter weight.

[0048] In some embodiments, compositions comprising fibrinogen comprise fibrinogen and albumin at a total concentration of about 99.5%, sometimes about 99.6%, sometimes about 99.7%, sometimes about 99.8%, and sometimes about 99.9% based on total protein dry matter weight.

[0049] As described herein, the weight ratio of albumin to fibrinogen in a composition comprising fibrinogen is at least about 1 to 15 (0.067), respectively.

[0050] In some embodiments that may be considered aspects of the present disclosure, there is provided a fibrinogen-containing composition comprising one or more coagulation proteins and albumin, wherein the one or more coagulation proteins comprise fibrinogen, and the weight ratio of albumin to fibrinogen is at least about 1:15, respectively.

[0051] In some embodiments, the compositions comprising fibrinogen comprise fibrinogen and albumin, and the weight ratio of each of the albumin to fibrinogen is at least about 1:14 (0.07), sometimes at least about 1:13 (0.076), sometimes at least about 1:12 (0.083), sometimes at least about 1:11 (0.09), sometimes at least about 1:10 (0.1), sometimes at least about 1:9 (0.11), sometimes at least about 1:8 (0.125), sometimes at least about 1:7 (0.14), and sometimes is at least about 1:6 (0.167), sometimes at least about 1:5 (0.2), sometimes at least about 1:4 (0.25), sometimes at least about 1:3.5 (0.28), sometimes at least about 1:3 (0.31), sometimes at least about 1:2.5 (0.4), sometimes at least 1:2 (0.5), sometimes about 1:1.6 (0.625), sometimes about 1:1.5 (0.66), sometimes at least about 1:1.4 (0.71), sometimes at least about 1:1.3 (0.77), and sometimes at least about 1:1.2 (0.83).

[0052] In some embodiments, the composition comprising fibrinogen comprises fibrinogen and albumin, wherein the weight ratio of each of the albumin to fibrinogen is about 1:0.8 (1.25) to about 1:15 (0.067).

[0053] In some embodiments, a composition comprising fibrinogen comprises fibrinogen and albumin, wherein the weight ratio of albumin to fibrinogen is about 1:0.9 to about 1:15 (0.067), sometimes about 1:1 to about 1:15, sometimes about 1:1 to about 1:14, sometimes about 1:1 to about 1:13, sometimes about 1:1 to about 1:12, sometimes about 1:1 to about 1:11, sometimes about 1:1 to about 1:10, sometimes about 1:1 to about 1:9, sometimes about 1:1 to about 1:8, sometimes about 1:1 to about 1:7, and sometimes about 1:2 to about 1:7.

[0054] In some embodiments, the composition comprising fibrinogen comprises fibrinogen and albumin, wherein the weight ratio of albumin to fibrinogen is from about 1:1 to about 1:10, sometimes from about 1:1.6 to about 1:3.

[0055] In some embodiments, a composition comprising fibrinogen comprises fibrinogen and albumin, wherein the weight ratio of albumin to fibrinogen is about 1:0.8, sometimes about 1:0.9, sometimes about 1:1, sometimes about 1:1.1, sometimes about 1:1.2, sometimes about 1:1.3, sometimes about 1:1.4, sometimes about 1:1.5, sometimes about 1:1.6, sometimes about 1:1.7, sometimes about 1:1.8, sometimes about 1:1.9, sometimes about 1:2, sometimes about 1:3, sometimes about 1:4, sometimes about 1:5, sometimes about 1:6, sometimes about 1:7, sometimes about 1:8, sometimes about 1:9, sometimes about 1:10, sometimes about 1:11, sometimes about 1:12, sometimes about 1:13, sometimes about 1:14, and sometimes about 1:15.

[0056] As described herein, the addition of albumin enhanced adhesion. In some embodiments, the fibrinogen-containing composition comprises about 0.1% to about 55% albumin based on total protein dry matter weight.

[0057] In some embodiments, compositions comprising fibrinogen comprise from about 0.5% to about 55% albumin based on total protein dry weight, sometimes from about 0.5% to about 53%, sometimes from about 0.5% to about 50%, sometimes from about 0.5% to about 48%, sometimes from about 0.5% to about 45%, and sometimes from about 0.5% to about 43% albumin based on the total protein weight in the composition on a dry basis.

[0058] In some embodiments, compositions containing fibrinogen contain from about 1% to about 55% albumin based on total protein dry matter weight, sometimes from about 3% to about 55%, sometimes from about 4.5% to about 55%, sometimes from about 5% to about 53%, sometimes from about 5% to about 50%, sometimes from about 5% to about 48%, sometimes from about 5% to about 45%, and sometimes from about 5% to about 43% albumin based on the total protein in the composition on a dry basis.

[0059] In some embodiments, compositions containing fibrinogen contain from about 10% to about 55% albumin based on total protein dry matter weight, sometimes from about 10% to about 53%, sometimes from about 10% to about 50%, sometimes from about 10% to about 48%, sometimes from about 10% to about 45%, and sometimes from about 10% to about 43% albumin based on the total protein in the composition on a dry basis.

[0060] In some embodiments, compositions containing fibrinogen contain from about 10% to about 55% albumin based on total protein dry matter weight, sometimes from about 12% to about 55%, sometimes from about 15% to about 55%, sometimes from about 18% to about 55%, and sometimes from about 20% to about 55% albumin based on the total protein in the composition on a dry basis.

[0061] In some embodiments, compositions containing fibrinogen contain from about 10% to about 55% albumin based on total protein dry matter weight, sometimes from about 12% to about 53%, sometimes from about 14% to about 50%, sometimes from about 16% to about 48%, sometimes from about 18% to about 45%, sometimes from about 20% to about 40%, and sometimes from about 23% to about 40% albumin based on the total protein in the composition on a dry basis.

[0062] In some embodiments, compositions comprising fibrinogen comprise about 1% (1 mg / ml) albumin based on the total protein in the composition on a dry basis, sometimes about 5%, sometimes about 10%, sometimes about 12%, sometimes about 15%, sometimes about 17%, sometimes about 20%, sometimes about 23%, sometimes about 25%, sometimes about 27%, sometimes about 30%, sometimes about 32%, sometimes about 35%, sometimes about 38%, sometimes about 40%, sometimes about 43%, sometimes about 45%, sometimes about 47%, sometimes about 50%, and sometimes about 53% albumin.

[0063] In some embodiments, compositions that contain fibrinogen contain about 20%, sometimes about 23%, sometimes about 25%, sometimes about 27%, sometimes about 30%, sometimes about 32%, sometimes about 35%, sometimes about 38%, sometimes about 39%, and sometimes about 40% albumin based on the total protein in the composition.

[0064] In some embodiments, the albumin is human serum albumin.

[0065] While compositions containing fibrinogen may contain other blood-derived proteins such as fibronectin, immunoglobulins, plasminogen, vWF, factor VIII, antithrombin III, and serpin proteins, these, if present, are in small amounts and therefore the compositions described herein are considered pure fibrinogen compositions.

[0066] When referring to "minor amounts" of other blood-borne proteins, this should be understood as being less than about 10% based on the total protein amount in the composition, sometimes less than about 8%, sometimes less than about 6%, sometimes less than about 5%, sometimes less than about 3%, sometimes less than about 1%, and sometimes less than about 0.6% based on the total protein amount in the composition.

[0067] Typically, the term "pure fibrinogen" refers to a composition that contains at least about 90 w / w% clottable fibrinogen based on total clottable protein dry matter. In other words, a composition that contains fibrinogen contains at least 90% fibrinogen.

[0068] In some embodiments, at least about 91% fibrinogen based on total coagulable protein dry matter, sometimes at least about 92% fibrinogen based on total coagulable protein dry matter, sometimes at least about 93% fibrinogen, sometimes at least about 94% fibrinogen, sometimes at least about 95% fibrinogen, sometimes at least about 96% fibrinogen, sometimes at least about 97% fibrinogen, sometimes at least about 98% fibrinogen, sometimes at least about 99% fibrinogen, sometimes at least about 99.1% fibrinogen, sometimes at least about 99.2% fibrinogen, sometimes at least about 99.3% fibrinogen, sometimes at least about 99.4% fibrinogen, sometimes at least about 99.5% fibrinogen, sometimes at least about 99.6% fibrinogen, sometimes at least about 99.7% fibrinogen, sometimes at least about 99.8% fibrinogen, sometimes at least about 99.9% fibrinogen. In some embodiments, the coagulable protein in the fibrinogen-containing composition comprises about 100% fibrinogen based on total coagulable protein dry matter.

[0069] In some embodiments, the coagulant protein in a composition comprising fibrinogen comprises about 91%-100% fibrinogen based on total coagulant protein dry matter.

[0070] In some embodiments, the coagulant protein in the composition comprises about 90% to about 99.9% fibrinogen based on total coagulant protein dry matter, sometimes about 91% to about 99.9% fibrinogen based on total coagulant protein dry matter, sometimes about 92% to about 99.9% fibrinogen, sometimes about 93% to about 99.9% fibrinogen, sometimes about 94% to about 99.9% fibrinogen, sometimes about 95% to about 99.9% fibrinogen, sometimes about 96% to about 99.9% fibrinogen, sometimes about 97% to about 99.9% fibrinogen, sometimes about 97.5% to about 99.9% fibrinogen, sometimes about 98% to about 99.9% fibrinogen, sometimes about 98.5% to about 99.9% fibrinogen, sometimes about 99% to about 99.9% fibrinogen.

[0071] In some embodiments, the coagulant protein in the composition comprises from about 99% to about 99.9% fibrinogen based on total coagulant protein dry matter, sometimes from about 99.1% to about 99.9%, sometimes from about 99.2% to about 99.9%, sometimes from about 99.3% to about 99.9%, sometimes from about 99.4% to about 99.9%, and sometimes from about 99.5% to about 99.9% based on total coagulant protein dry matter.

[0072] In some embodiments, the coagulant protein in the composition comprises about 91% fibrinogen based on total coagulant protein dry matter, sometimes about 92% fibrinogen based on total coagulant protein dry matter, sometimes about 93% fibrinogen, sometimes about 94% fibrinogen, sometimes about 95% fibrinogen, sometimes about 96% fibrinogen, sometimes about 97% fibrinogen, sometimes about 98% fibrinogen, sometimes about 99% fibrinogen, sometimes about 99.1% fibrinogen, sometimes about 99.2% fibrinogen, sometimes about 99.3% fibrinogen, sometimes about 99.4% fibrinogen, about 99.5% fibrinogen, sometimes about 99.6% fibrinogen, sometimes about 99.7% fibrinogen, sometimes about 99.8% fibrinogen, and sometimes about 99.9% fibrinogen.

[0073] Examples of fibrinogen sources include, but are not limited to, recombinant fibrinogen and fibrinogen purified from plasma. Non-limiting examples of plasma sources of fibrinogen include human, porcine, bovine, and salmon.

[0074] The amount of fibrinogen can be measured by any method known in the art. For example, fibrinogen can be measured by one or more of the following: Claus kinetic clotting method, clottable fibrinogen method, immunological method, or electrophoretic method. The Claus method involves measuring the rate of clot formation of a fibrinogen-containing solution. The clottable fibrinogen method involves washing the formed clot and measuring the protein content. The Claus method measures fibrinogen that can clot in a few seconds (e.g., 5-60 seconds). The clottable protein method (which involves allowing fibrinogen to completely clot, followed by washing, and then measuring the remaining protein) allows 20-60 minutes for clot formation. Sometimes this type of assay is called gravimetric fibrinogen because in the past clots were measured by weighing the clot. In current methods, the clotted protein is usually dissolved and measured by spectroscopy or various other protein measurement methods. Determination of clot formation endpoints can be done by several methods, including, for example, visual clot detection (when performed with plasma, this is called the tube tilt method), viscosity-based methods (the Stago analyzer is one example), and optical methods that identify clot formation based on changes in absorbance (such as most hospital coagulation analyzers). Details of such methods can be found, for example, in British Journal of Haematology, 2003, 121, 396-404, the contents of which are incorporated by reference.

[0075] As described herein, it has surprisingly been found that compositions of the present disclosure are highly effective, as demonstrated, for example, in abrasion tests, despite the fact that they contain small amounts of fibronectin, or sometimes essentially no fibronectin. As noted above, this is surprising, especially in light of prior publications (e.g., Bar et al.).

[0076] The amount of fibronectin can be measured by any method known in the art. For example, fibronectin can be measured by electrophoretic, turbidimetric, or immunological methods. Specifically, such methods can include Enzyme-Linked Immunosorbent Assay (ELISA), radial immunodiffusion, Ouchterlony immunodiffusion, and turbidimetric techniques. It should be noted that nephelometric methods involve the use of antibodies (sometimes coated on particles) that aggregate in the presence of the protein of interest and scatter light.

[0077] In some embodiments, compositions comprising fibrinogen contain less than about 0.5% fibronectin, sometimes less than about 0.4%, sometimes less than about 0.3%, sometimes less than about 0.2%, sometimes less than about 0.1%, sometimes less than about 0.05% fibronectin, sometimes less than about 0.04%, sometimes less than about 0.02%, sometimes even less than about 0.01, sometimes less than about 0.005% fibronectin, sometimes less than about 0.004%, sometimes less than about 0.002%, and sometimes even less than about 0.001% fibronectin on a total protein dry matter basis.

[0078] In some embodiments, the composition comprising fibrinogen comprises about 0.001% to about 0.5% fibronectin on a total protein dry matter basis.

[0079] In some embodiments, the composition comprising fibrinogen comprises about 0.004% to about 0.5% fibronectin on a total protein dry matter basis.

[0080] In some embodiments, compositions containing fibrinogen contain from about 0.01% to about 0.4% fibronectin on a total protein dry matter basis, sometimes from about 0.01% to about 0.3%, and sometimes from about 0.01% to about 0.2% fibronectin.

[0081] In some embodiments, the composition comprises about 0.01% fibronectin, sometimes about 0.05%, sometimes about 0.1%, sometimes about 0.15%, sometimes about 0.2%, sometimes about 0.25%, sometimes about 0.3%, sometimes about 0.35%, and sometimes about 0.4% fibronectin based on total protein dry matter.

[0082] In some embodiments, the composition is essentially free of fibronectin.

[0083] It should be noted that when referring to a composition comprising fibrinogen without fibronectin, the fibrinogen-containing composition encompasses compositions that contain fibronectin in an amount that is undetectable by methods used to measure fibronectin, such as the methods described herein.

[0084] It should be noted that recombinant fibrinogen or compositions thereof may sometimes be considered to be free of fibronectin. It should be further noted that sometimes recombinant fibrinogen compositions may contain fibronectin, but the amount of fibronectin is below detectable levels.

[0085] In some embodiments that can be considered aspects of the present disclosure, a composition comprising fibrinogen comprises fibrinogen and albumin in a total concentration of at least 90 w / w% based on total protein weight, fibronectin in an amount of less than about 0.5 w / w% based on total protein weight, and the weight ratio of albumin to fibrinogen is at least 1:15, respectively.

[0086] In some embodiments that can be considered aspects of the present disclosure, a composition comprising fibrinogen comprises fibrinogen and albumin in a total concentration of at least about 90 w / w% based on total protein weight, fibronectin in an amount of less than about 0.5 w / w% based on total protein weight, and the weight ratio of albumin to fibrinogen is from 1:1 to 1:15, respectively.

[0087] In some embodiments that can be considered aspects of the present disclosure, a composition comprising fibrinogen comprises fibrinogen and albumin in a total concentration of at least about 90 w / w% based on total protein weight, fibronectin in an amount of less than about 0.5 w / w% based on total protein weight, and the weight ratio of albumin to fibrinogen is from 1:1 to 1:10, respectively.

[0088] In some embodiments that can be considered aspects of the present disclosure, a composition comprising fibrinogen comprises fibrinogen and albumin in a total concentration of at least about 90 w / w% based on total protein weight, fibronectin in an amount of less than about 0.5 w / w% based on total protein weight, and the weight ratio of albumin to fibrinogen is about 1:1.6 to about 1:3, respectively.

[0089] In some embodiments that can be considered aspects of the present disclosure, a composition comprising fibrinogen comprises fibrinogen and albumin in a total concentration of at least about 95 w / w% based on total protein weight, fibronectin in an amount of less than about 0.5 w / w% based on total protein weight, and the weight ratio of albumin to fibrinogen is from 1:1 to 1:10, respectively.

[0090] In some embodiments that can be considered aspects of the present disclosure, a composition comprising fibrinogen comprises fibrinogen and albumin in a total concentration of at least about 95 w / w% based on total protein weight, fibronectin in an amount of less than about 0.5 w / w% based on total protein weight, and the weight ratio of albumin to fibrinogen is about 1:1.6 to about 1:3, respectively.

[0091] In one embodiment, the composition comprising fibrinogen comprises a cryoprecipitate comprising fibrinogen and albumin, wherein fibronectin is present in an amount of less than about 0.5% or absent based on total protein weight, hi one embodiment, the albumin is present in a weight ratio of albumin to fibrinogen of at least 1:15, respectively.

[0092] The composition comprising fibrinogen can be in any form. According to some embodiments, the composition comprising fibrinogen is in liquid form. When referring to liquid form, it should be understood as being liquid at room temperature. When in liquid form, according to some embodiments, the liquid carrier is a buffer having an essentially neutral pH, for example, a pH in the range of about 6.0 to about 8.0, sometimes in the range of about 6.5 to about 8.0, and sometimes a pH of about 7.0±0.5, and sometimes a pH of 7.25±0.5.

[0093] In some embodiments, the composition comprising fibrinogen is an aqueous formulation. Reference to an "aqueous formulation" should be understood to encompass a blend of components in liquid or frozen form that contain water molecules.

[0094] In some embodiments, the fibrinogen-containing composition is frozen. Prior to use, the fibrinogen-containing composition can be thawed, thereby resulting in a liquid form at room temperature.

[0095] According to some embodiments, the fibrinogen-containing composition is in powder or lyophilized form. If in powder or lyophilized form, the fibrinogen-containing composition can be mixed with a suitable aqueous formulation so that the formulation is in liquid form when used.

[0096] In some other embodiments, the composition comprising fibrinogen is in dry form.Typically, when referring to a composition in dry form, this refers to a composition that comprises less than 3% aqueous medium, sometimes less than 2% aqueous medium, and sometimes less than 1% aqueous medium.Dry form is, in some examples, essentially free of aqueous medium.

[0097] In some further embodiments, the composition is in powder form.

[0098] In some embodiments, the composition is in powder form characterized by a D50 (median particle diameter (also called "median particle size")) of 13 μm or more (i.e., 50% of the particles in a sample are 13 μm or more) and / or a D90 of less than 100 μm (i.e., 90% of the particles in a sample are 100 μm or less).

[0099] The present disclosure also refers to the combination of a thrombin component with a composition comprising fibrinogen. Such combinations with thrombin components can be used to improve performance, such as, for example, tissue adhesion.

[0100] Thus, according to some aspects, the present disclosure provides compositions comprising one or more coagulation proteins, albumin, and thrombin, wherein the one or more coagulation proteins and albumin are at a combined concentration of at least 90% based on total protein weight, the one or more coagulation proteins comprise fibrinogen and fibronectin, wherein fibronectin is present in an amount of less than about 0.5% based on total protein weight or is absent, and the weight ratio of albumin to fibrinogen is at least 1:15, respectively.

[0101] Thrombin can be obtained from a variety of sources. In some embodiments, the thrombin is human thrombin. In some other embodiments, the thrombin is bovine thrombin. In some embodiments, the thrombin is recombinant thrombin.

[0102] The thrombin component, according to some embodiments, comprises, in addition to thrombin, at least one of albumin, calcium, sodium chloride, glycine, acetate, or combinations thereof, hi some embodiments, the thrombin component comprises about 40 mM calcium.

[0103] In some embodiments, the thrombin component comprises at least one vitamin K-dependent clotting factor in addition to thrombin or in the absence of thrombin. In some embodiments, the thrombin component comprises, in addition to thrombin, an immunoglobulin, ceruloplasmin, or complement C4. The vitamin K-dependent clotting factor comprises at least one of factor II, factor VII, factor IX, factor X, protein C, protein S, or a combination thereof.

[0104] A composition comprising fibrinogen and a composition comprising thrombin are referred to herein as thrombin-containing compositions / compositions comprising thrombin, and are sometimes referred to as "fibrin sealant" or "fibrin glue."

[0105] According to some embodiments, the fibrinogen-containing composition and thrombin are separated from each other and combined before use. According to such embodiments, the fibrinogen-containing composition and thrombin are each present as separate formulations, e.g., aqueous formulations. It is noted that the fibrinogen-containing composition and thrombin can be frozen and thawed one by one, and then the two are combined for use.

[0106] As described herein, the compositions according to the present disclosure are hemostatic clot forming compositions. It is noted that, for example, when in contact with tissue and / or blood directly on or in close proximity to a wound (e.g., a bleeding wound), the compositions react to form a clot that then acts as a tissue adhesive, thereby stopping bleeding.

[0107] According to some embodiments, thrombin can be added to the fibrinogen-containing composition after lyophilization and grinding of the fibrinogen-containing composition.

[0108] In other words, a composition comprising fibrinogen in powder form is combined with thrombin, in some embodiments, the thrombin is ground into a powder before being combined with a composition comprising fibrinogen in powder form.

[0109] In some embodiments, the thrombin composition powder is characterized by a D50 of 5 μm or more (i.e., 50% of the particles in a sample are 5 μm or more) and / or a D90 of 39 μm or less (i.e., 90% of the particles in a sample are 39 μm or less).

[0110] In some embodiments, the thrombin powder is added to a composition powder that includes fibrinogen, and thus the powder includes a composition that includes thrombin and fibrinogen that have been separately lyophilized and ground into a powder (herein a combined powder).

[0111] This powder may further be used to be disposed on an article of manufacture, for example, as described below.

[0112] According to some aspects, the present disclosure provides an article of manufacture comprising a composition described herein.

[0113] In some embodiments, the article of manufacture comprises a composition comprising fibrinogen. In some embodiments, a powder comprising a composition comprising fibrinogen is disposed on the article of manufacture.

[0114] In some embodiments, the article of manufacture comprises both a composition comprising fibrinogen and a composition comprising thrombin.

[0115] In some embodiments, the article of manufacture includes a powder comprising fibrinogen, the powder comprising the composition and thrombin, or in other words, a powder comprising the composition comprising fibrinogen and thrombin (i.e., a combined powder) is disposed on the article of manufacture.

[0116] According to embodiments in which the article of manufacture comprises a composition comprising fibrinogen in a dry form, thrombin can be applied separately to the wound prior to application of the article of manufacture. Thrombin can be applied by any method known in the art, particularly by spraying.

[0117] As used herein, the article of manufacture should be understood to be a sterile dressing suitable for placement over tissue, particularly wound tissue.

[0118] It should be noted that upon application of the article of manufacture onto wound tissue, the dry powder should be hydrated so that a clot forms. In other words, the article of manufacture can be placed onto and / or adjacent to wound tissue, provided that any fluid, e.g., blood, is present on the surface of the tissue to allow hydration of the powder.

[0119] In some embodiments, the article of manufacture is a wound dressing comprising a composition of the invention (with or without thrombin) and is configured to be placed adjacent to or directly on a wound, e.g., a bleeding wound.

[0120] As used herein, the term "dressing" relates to a wound covering.

[0121] It should be noted that an article of manufacture comprising a composition of the present disclosure, specifically a wound dressing, refers to the composition in combination with a wound dressing.

[0122] It should be noted that "combined" should be understood as any form of association / contact between the composition and the coating, which may include, but is not limited to, any one of spraying, painting, dipping, immersion, brushing, casting, printing, pouring, coating, or any combination thereof, as further described below.

[0123] According to embodiments in which the composition is in powder form, e.g., lyophilized, the powder can be moistened with an aqueous solution and combined with a wound dressing prior to use.

[0124] The term "dressing," as used herein, is intended to have the same meaning as a patch or pad, all of which may be used interchangeably herein.

[0125] In some embodiments, the wound dressing comprises at least one layer, hi some embodiments, the wound dressing is a multi-layer dressing.

[0126] In some embodiments, the wound dressing comprises at least one layer that is a backing layer.

[0127] "Backing" as used herein refers to the physical structure that provides support to the composition. It is noted that the reinforcing fabric, and thus the backing layer, provides the support / strength that allows the wound dressing to be placed onto tissue (e.g., wound) to achieve hemostasis and / or tissue adhesion.

[0128] In some embodiments, the backing layer is or includes a reinforcing fabric.

[0129] In some embodiments, the reinforcing fabric comprises a woven or knitted fabric. In some embodiments, the woven or knitted fabric is a biocompatible material. In some embodiments, the woven or knitted fabric is a bioincompatible material.

[0130] In some embodiments, the woven or knitted fabric is an absorbent material.

[0131] In some embodiments, the wound dressing comprises a layer of absorbent woven or knitted fabric.

[0132] In some other embodiments, the woven or knitted fabric (forming the backing layer) comprises an oxidized polysaccharide.

[0133] In some other embodiments, the woven or knitted fabric comprises cellulose and / or its derivatives.

[0134] In some embodiments, the woven or knitted fabric comprises oxidized cellulose (OC), oxidized regenerated cellulose (ORC), collagen, or a synthetic polymer (eg, polyglactin 910, "PG910").

[0135] In some embodiments, the woven or knitted fabric comprises OC.

[0136] In some other embodiments, the woven or knitted fabric comprises ORC.

[0137] The backing layer can be any backing material known in the art, including, for example, those known under the trade names INTERCEED® and SURGICEL®, among others.

[0138] The SURGICEL® family of absorbable hemostatic agents consists of four main product groups, all of which are hemostatic wound dressings commercially available from Ethicon, Inc. (A Johnson & Johnson Company, Somerville, NJ). These products include SURGICEL® (original), SURGICEL® NU-KNIT®, SURGICEL® FIBRILLAR™, and SURGICEL® SNoW™.

[0139] In some embodiments, the multi-layer wound dressing comprises at least one layer of absorbent nonwoven in addition to the backing layer.

[0140] In some embodiments, the multi-layer wound dressing comprises at least one layer of reinforcing fabric and at least one layer of absorbent nonwoven fabric.

[0141] The term "nonwoven" as used herein also includes, but is not limited to, bonded, formed, or engineered fabrics, typically produced by processes other than, for example, spinning, weaving, or knitting.

[0142] In some embodiments, the nonwoven fabric comprises fibers comprised of an aliphatic polyester polymer or copolymer of one or more monomers selected from the group consisting of lactic acid, lactide (including L-, D-, meso, and D,L mixtures), glycolic acid, glycolide, e-caprolactone, p-dioxanone, and trimethylene carbonate.

[0143] In some embodiments, the nonwoven fabric comprises a glycolide / lactide copolymer.

[0144] In some embodiments, the multi-layer dressing comprises at least one layer comprising a nonwoven fabric comprising a glycolide / lactide copolymer and at least one layer comprising a woven or knitted fabric comprising ORC.

[0145] The absorbent nonwoven may be attached either directly or indirectly to an absorbent woven or knitted fabric, for example, the nonwoven may be incorporated into the absorbent woven or knitted fabric by needle punching, calendaring, embossing, or hydroentanglement, or by chemical or thermal bonding.

[0146] In some embodiments, the fibrinogen-containing composition or the thrombin-containing composition is substantially uniformly dispersed throughout the nonwoven fabric and / or disposed on a surface of the nonwoven fabric.

[0147] As described herein, the article of manufacture, e.g., a wound dressing, is placed onto the tissue of a subject in need thereof, hi some embodiments, the article of manufacture or the composition is for use in topical administration.

[0148] As shown herein, application of the wound dressings described herein has been found to be highly effective at adhering and initiating hemostasis.

[0149] Thus, according to some aspects, the present disclosure provides a composition comprising fibrinogen, a thrombin-containing composition (all collectively referred to herein as "compositions"), an article of manufacture comprising a composition comprising fibrinogen, or an article of manufacture comprising a thrombin-containing composition (all collectively referred to herein as articles of manufacture) for use in a tissue adhesive.

[0150] Tissue adhesion can be measured by any method known in the art. For example, tissue adhesion can be measured by peel test. In some examples, tissue adhesion is examined at a value of at least 40 Newtons / meter (N / m) measured by peel force test. In some examples, tissue adhesion is examined at a value of about 40 N / m to about 90 N / m measured by peel force test.

[0151] Additionally, according to some aspects, the present disclosure provides a composition comprising fibrinogen, a composition comprising thrombin (all of which are collectively referred to herein as compositions), an article of manufacture comprising a composition comprising fibrinogen, or an article of manufacture comprising a thrombin-containing composition (all of which are collectively referred to herein as articles of manufacture) for use in controlling bleeding.

[0152] Additionally, according to some aspects, the present disclosure provides a composition comprising fibrinogen, a composition comprising thrombin (all collectively referred to herein as compositions), an article of manufacture comprising a composition comprising fibrinogen, or an article of manufacture comprising a thrombin-containing composition (all collectively referred to herein as articles of manufacture) for use in treating, inhibiting, arresting, or slowing bleeding or any disorder related thereto.

[0153] The present disclosure also provides, according to some embodiments, a method for adhering tissue, comprising topically administering to a subject in need thereof an effective amount of a composition or article of manufacture, wherein the composition is a fibrinogen-containing composition or a thrombin-containing composition. It should be noted that when referring to a tissue adhesive, it is for stopping bleeding and / or sealing physiological leaks.

[0154] In some embodiments, the compositions and articles of manufacture are for use in sealing physiological leaks, including cerebrospinal fluid (CSF) leaks, lymphatic leaks, bile leaks, gastrointestinal (GI) content leaks, pancreatic leaks, pulmonary air leaks, or any combination thereof.

[0155] The present disclosure also provides, according to some aspects, methods for controlling bleeding. Controlling bleeding, as used herein, refers to treating, suppressing, arresting, or slowing bleeding or any disorder associated therewith.

[0156] The present disclosure also provides, according to some aspects, a method for treating, inhibiting, arresting, or slowing bleeding or any disorder related thereto, comprising locally administering to a subject in need thereof an effective amount of a composition or article of manufacture, wherein the composition is a fibrinogen-containing composition or a thrombin-containing composition.

[0157] The methods of the present invention comprise the step of topically administering to a subject in need thereof an effective amount of a composition or article of manufacture described herein.

[0158] In some embodiments, the methods of the invention comprise locally administering to a tissue of a subject in need thereof a composition comprising fibrinogen or an article of manufacture comprising fibrinogen, wherein the composition comprises one or more coagulation proteins and albumin, wherein the one or more coagulation proteins and albumin are at a total concentration of at least 90 w / w% based on total protein weight, the one or more coagulation proteins comprise fibronectin in an amount of less than about 0.5 w / w% based on total protein weight, and the one or more coagulation proteins comprise fibrinogen, wherein the weight ratio of albumin to fibrinogen is at least 1:15, respectively.

[0159] In some embodiments, the methods of the present invention comprise locally administering to a tissue of a subject in need thereof a composition comprising fibrinogen or an article of manufacture comprising a composition comprising fibrinogen, the composition comprising fibronectin and albumin at a combined concentration of at least 90 w / w% based on total protein weight, the fibronectin being in an amount of less than about 0.5 w / w% based on total protein weight, and the weight ratio of albumin to fibrinogen being at least 1:15, respectively.

[0160] In some embodiments, the methods of the invention comprise locally administering to a tissue of a subject in need thereof a composition comprising fibrinogen or an article of manufacture comprising a composition comprising fibrinogen, the composition comprising fibrinogen and albumin in a total concentration of at least 90 w / w% based on total protein weight, fibronectin in an amount of less than about 0.5 w / w% based on total protein weight, and the weight ratio of albumin to fibrinogen is between 1:1 and 1:15, respectively.

[0161] In some embodiments, the methods of the invention comprise locally administering to a tissue of a subject in need thereof a composition comprising fibrinogen or an article of manufacture comprising a composition comprising fibrinogen, the composition comprising fibrinogen and albumin in a total concentration of at least 90 w / w% based on total protein weight, fibronectin in an amount of less than about 0.5 w / w% based on total protein weight, and a weight ratio of albumin to fibrinogen of about 1:1.6 to about 1:3, respectively.

[0162] In some embodiments, the methods of the present invention comprise locally administering to a tissue of a subject in need thereof a thrombin-containing composition or an article of manufacture comprising a thrombin-containing composition described herein.

[0163] As used herein, the term "bleeding" may refer to at least one hemostatic deficiency that results in an increased susceptibility to bleeding, for example, hemophilia, a platelet disorder, or medication that affects the hemostatic system.

[0164] In some embodiments, the bleeding is caused by an external stimulus. The bleeding can be for a variety of reasons, such as due to personal care, due to trauma, or a bleeding wound during a surgical procedure.

[0165] In some embodiments, the bleeding can be at least one of injury-induced bleeding, surgery-induced bleeding, or trauma-induced bleeding.

[0166] In some embodiments, the bleeding is during a surgical procedure.

[0167] In some embodiments, the methods of the invention comprise administering the composition or article of manufacture during a surgical procedure.

[0168] In some embodiments, the methods of the present invention may be applicable to subjects at high risk of developing bleeding.

[0169] In some embodiments, the methods of the present invention may be applicable to subjects suffering from bleeding during a surgical procedure.

[0170] There are many surgical procedures in which the wound dressings of the present disclosure may be used, in some embodiments, the surgical procedures include abdominal surgery, cardiovascular surgery, thoracic surgery, head and neck surgery, dural surgery, spinal surgery, pelvic surgery, skin procedures, and / or subcutaneous tissue procedures.

[0171] The wound dressings of the present disclosure are suitable for application (administration) to a variety of wound tissues.

[0172] In some embodiments, the wound tissue is or includes a bleeding artery. In some embodiments, the wound tissue is or includes a bleeding vein. The vein may be a vena cava.

[0173] In some embodiments, the wound tissue is or comprises a capillary bed.

[0174] In some embodiments, the methods of the present invention comprise pressing / compressing the wound dressing for a period of time sufficient to allow adhesion of the wound dressing to at least a portion of the wound tissue.

[0175] In some embodiments, the wound dressing is for use as an adjunct to hemostasis by application of pressure, optionally by manual application.

[0176] In some embodiments, the method comprises pressing (compressing) the wound dressing against the wound tissue for a period of time sufficient to allow adhesion of the wound dressing to at least a portion of the wound tissue.

[0177] In some embodiments, the method includes pressing (pressing) the wound dressing against the wound tissue until bleeding has essentially stopped.

[0178] In some embodiments, the method includes applying (pressing) the wound dressing against the wound tissue for a period of time sufficient to allow for hydration, tissue adhesion, and hemostasis.

[0179] As will be appreciated, the time sufficient to allow for any of hydration, tissue adhesion, and hemostasis will depend on the wound being treated and the amount of bleeding. This time can be adjusted to achieve complete hemostasis. In some embodiments, the wound dressing is pressed against at least a portion of the wound for about 1 minute, and sometimes for about 3 minutes.

[0180] The present disclosure further provides a kit (package). In some embodiments, the kit comprises a composition comprising fibrinogen. In some embodiments, the kit comprises a thrombin-containing composition.

[0181] In some embodiments, the kit comprises an article of manufacture.

[0182] According to some embodiments, the kit of the present invention is for use in a subject suffering from bleeding or any disorder related thereto, as described herein above. It should be further understood that the kit of the present invention may also be used to prevent bleeding in subjects at high risk of bleeding.

[0183] In some embodiments, the kit further comprises instructions for using the formulation to treat, inhibit, or slow bleeding or any disorder related thereto.

[0184] The fibrinogen-containing composition and / or the thrombin-containing composition may be stored in liquid or frozen form and thawed immediately prior to use.

[0185] In some embodiments, the kit provides an article of manufacture, specifically a wound dressing, in combination with a fibrinogen-containing composition and / or a thrombin-containing composition for preparation of the wound dressing and its application to a wound, and instructions for use.

[0186] As indicated above, the method of the present invention comprises administering a therapeutically effective amount of the composition of the present invention. For purposes disclosed herein, "effective amount" or "therapeutically effective" refers to an amount of the composition effective to treat, inhibit, or delay one or more symptoms of the diseases described herein, specifically inhibit or stop bleeding.

[0187] As used herein, the term "subject" refers to any organism to be treated with the formulations described herein, including, but not limited to, any mammal, such as a human.

[0188] The term "about" as used herein refers to values ​​that may deviate by up to 1%, more specifically 5%, more specifically 10%, more specifically 15%, and in some cases up to 20% or more or less than the stated value, the deviation range including integer values ​​and, where applicable, non-integer values, constituting the continuous ranges disclosed and described.

[0189] It is to be understood that the present invention is not limited to the specific examples, method steps, and compositions disclosed herein, and as such, such methods, steps, and compositions may vary somewhat. It is also to be understood that the terminology used herein is used only for the purpose of describing particular embodiments, and is not intended to be limiting, since the scope of the present disclosure will be limited only by the appended claims and equivalents thereof.

[0190] It must be noted that as used in this specification and the appended claims, the singular forms "a," "an," and "the" include plural referents unless the content clearly dictates otherwise.

[0191] Throughout this specification and the examples, as well as the claims that follow, unless the context otherwise requires, the word "comprise" and variations such as "comprises" and "comprising" will be understood to imply the inclusion of a stated integer or step or group of integers or steps, but not the exclusion of any other integer or step or group of integers or steps.

[0192] The following examples are representative of techniques employed by the inventors in carrying out aspects of the present invention. It should be understood that while these techniques are exemplary of preferred embodiments for carrying out the invention, those skilled in the art will recognize that, in light of this disclosure, numerous modifications can be made without departing from the spirit and intended scope of the invention.

[0193] Non-limiting examples Example 1: Tissue Adhesion Test - Albumin Dose Study The following samples shown in Table 1 were prepared by adding albumin at the indicated percentage relative to fibrinogen. The amount of fibronectin was less than about 0.4 w / w% of the total protein weight. These are referred to herein as "compositions containing fibrinogen."

[0194] [Table 1]

[0195] Fibrinogen-containing compositions with various amounts of albumin and fibrinogen were prepared in the ratios detailed in Table 1. The fibrinogen-containing compositions were lyophilized and ground to a powder. Thrombin was lyophilized and ground separately from the fibrinogen composition.

[0196] Composition powders containing fibrinogen were tested such that the amount of clottable fibrinogen and thrombin activity was constant in all samples (but the amount of albumin varied). The amounts in Table 2 represent the amount of powder weighed for each peel test replicate.

[0197] [Table 2]

[0198] The combined powders (fibrinogen / albumin+thrombin--"thrombin-containing composition") were tested for tissue adhesion in a bovine dermal peel test in which the powder was sandwiched between two pieces of dermis. Specifically, dermis from Lampire Biological Laboratories, Inc. (Pipersville, PA 18947) was mounted in a fixture / frame with approximately 4" x 0.75" of tissue exposed, and then various amounts of powder were applied to the moistened tissue mounted in the frame. A second moistened dermal piece was applied on top of the mounted tissue, and a weight was applied to the back of the tissue for 3 minutes at 37°C. A 3 minute hold time was used for all samples. After 3 minutes, the edges of the dermis were gripped and a 90° peel test was performed down the length of the tissue. The tissue adhesion results are summarized in Table 3.

[0199] [Table 3]

[0200] Figure 1 shows the results for the first five compositions in Table 3 along with their corresponding standard deviation values. As can be seen, the mean peel force is significantly increased for compositions containing 23.8% and 38.5% albumin compared to compositions with less than 0.5% albumin. This is evident from the low p-values ​​of the t-test shown in Figure 1.

[0201] Figure 1 further shows tissue detachment force as a function of albumin% relative to total protein in a composition containing fibrinogen. In this connection, it should be noted that although albumin may be present in a thrombin-containing composition, its amount is relatively small, and in addition, the amount of thrombin (if present) is small compared to the amount of fibrinogen, resulting in a relatively small amount (trace amount) of albumin in the final composition. In other words, although Figure 1 shows albumin% relative to total protein in a composition containing fibrinogen, it can also be considered as albumin% relative to total protein (even after the addition of thrombin).

[0202] As can be seen in Figure 1, tissue adhesion was dependent on the % albumin in the sample. An increase in peel force was observed in samples containing about 23.8% albumin and about 38.5% albumin, indicating increased tissue adhesion. Increasing the amount of albumin to 55.6% reduced the peel force, indicating reduced tissue adhesion.

[0203] These results surprisingly suggest that albumin affects tissue adhesion and that the addition of albumin significantly improves adhesion.

[0204] Figure 2 shows the tissue detachment force as a function of fibrinogen % relative to total protein. Surprisingly, the results in Figure 2 show that in samples #2 and #3 (see Table 1), where the fibrinogen was less than 99.5% or 100%, the tissue detachment force was higher than the results obtained with sample #1, which contained 99.5% fibrinogen.

[0205] Taken together, these results indicate that the addition of albumin to the extent of 38.5% can enhance tissue adhesion. Further dilution of fibrinogen resulted in reduced tissue detachment.

[0206] All compositions tested contained low amounts of fibronectin, i.e., less than 0.5% fibronectin. The results of this example unexpectedly show that tissue adhesion occurs even when low amounts of fibronectin are present.

[0207] Example 2: Porcine partial nephrectomy model and tissue adhesion studies This example was designed to test hemostasis in a difficult bleeding model (see Cornum RL et al., The Journal of Urology 164, 864-867, 2000).

[0208] A powder containing fibrinogen, albumin, and thrombin was applied onto a matrix containing oxidized regenerated cellulose (ORC) and polyglactin 910. The properties of the matrix, also referred to herein as a fibrin patch, are summarized in Table 4.

[0209] [Table 4]

[0210] For adhesion testing, strips of fibrin patches (4 inches long x 0.75 inches wide) were adhered to moistened tissue surfaces with one end of the patch attached to the load cell of an Instron tensiometer. After the fibrin patch was allowed to adhere and solidify for 3 minutes, the patch was removed from the tissue by raising the load cell at a 90° angle from the tissue surface. The force required to separate the fibrin patch from the tissue matrix was recorded as force per unit width (N / m).

[0211] As shown, the coated matrix was able to adhere to a surface upon contact with a liquid.

[0212] [Table 5] * The amount of fibronectin was less than 0.4 w / w% of the total protein weight.

[0213] As can be seen in Table 5 and FIG. 3, increased adhesion was observed in the matrix coated samples containing albumin in amounts greater than 10% of the total protein, using tissue adhesion testing as in Example 1.

[0214] In addition, the coated matrices demonstrated hemostatic effects, specifically, the matrix coated with the formulation containing 38.5% albumin (corresponding to sample 3 in Table 1 above) demonstrated complete hemostasis in a porcine partial nephrectomy model, as shown in Table 6.

[0215] Technical details: (1) Mean arterial pressure (MAP) was maintained above 60 mmHg and systolic pressure was maintained above 90 mmHg during the evaluation, (2) the mean renal resection from the caudal pole (with capsule intact) was approximately 3.0 cm in length and 1.0 cm in depth. Bleeding at this site was graded as severe, and (3) complete hemostasis (4 out of 4) was achieved with a 100% success rate at both 3 and 15 minutes after application. No rebleeding was observed.

[0216] [Table 6] * A "4" indicates severe bleeding.

[0217] A representative image is shown in Figure 4, which shows a pad wound dressing containing the fibrinogen composition being applied to a bleeding surface in a porcine partial nephrectomy model. The results show that the composition was able to adhere and stop the bleeding. This indicates that the inclusion of albumin in the fibrinogen formulation produces a patch that is able to adhere well to tissue surfaces and also achieve hemostasis in the partial nephrectomy model.

[0218] [Embodiment] (1) A composition comprising one or more coagulation proteins and albumin, wherein the one or more coagulation proteins and the albumin are at a combined concentration of at least 90% based on total protein weight, the one or more coagulation proteins comprise fibrinogen and fibronectin, the fibronectin being in an amount of less than about 0.5% based on total protein weight or absent, and the weight ratio of the albumin to the fibrinogen, respectively, is at least 1:15. (2) The composition of embodiment 1, wherein the one or more coagulation proteins and the albumin are in a total concentration of about 90% to about 99.5%. (3) The composition of any one of the preceding claims, wherein the one or more coagulation proteins and the albumin are at a combined concentration of at least 95% based on total protein weight. (4) A composition according to any one of embodiments 1 to 3, wherein the coagulation protein comprises at least 90% fibrinogen. (5) A composition according to any one of the preceding claims, wherein the coagulation protein comprises at least 95% fibrinogen.

[0219] (6) The composition described in any one of embodiments 1 to 5, wherein the fibronectin is present in an amount of less than about 0.3% by weight. (7) The composition according to any one of embodiments 1 to 6, wherein the weight ratio of the albumin to the fibrinogen is from 1:1 to 1:15. (8) The composition according to any one of embodiments 1 to 7, wherein the weight ratio of the albumin to the fibrinogen is from 1:1 to 1:10. (9) The composition according to any one of embodiments 1 to 8, wherein the weight ratio of the albumin to the fibrinogen is 1:1.6 to 1:3. (10) The composition according to any one of embodiments 1 to 9, wherein the albumin content is from about 0.5% by weight to about 55% by weight.

[0220] (11) The composition according to any one of embodiments 1 to 10, wherein the albumin content is about 4.5% by weight to about 55% by weight. (12) The composition according to any one of embodiments 1 to 11, wherein the albumin content is about 5% by weight to about 43% by weight. (13) The composition according to any one of embodiments 1 and 12, which is in a dry form. (14) The composition according to any one of the preceding embodiments, which is in powder form. (15) The composition according to any one of the preceding embodiments, characterized by a tissue adhesion of at least 40 N / m as measured by a peel force test.

[0221] (16) The composition according to any one of embodiments 1 to 15, comprising thrombin. (17) The composition according to embodiment 16, wherein the albumin content is about 5% by weight to about 43% by weight. (18) The composition of embodiment 16 or 17, comprising less than about 0.5% fibronectin. (19) A composition comprising one or more coagulation proteins, albumin, and thrombin, wherein the one or more coagulation proteins and the albumin are at a combined concentration of at least 90% based on total protein weight, the one or more coagulation proteins comprise fibrinogen and fibronectin, wherein the fibronectin is in an amount of less than about 0.5% based on total protein weight or is absent, the weight ratio of the albumin to the fibrinogen, respectively, is at least 1:15, and the composition is in powder form. (20) An article of manufacture comprising the composition of any one of embodiments 1 to 19.

[0222] (21) The article of manufacture of embodiment 20, which is a wound dressing. (22) The article of any one of claims 20 to 21, comprising at least one layer of absorbent woven or knitted fabric. 23. The article of manufacture of claim 22, wherein the absorbent woven or knitted fabric comprises one or more oxidized polysaccharides. 24. The article of manufacture of claim 22 or 23, wherein the absorbent woven or knitted fabric comprises oxidized cellulose (OC), oxidized regenerated cellulose (ORC), or a combination thereof. (25) The article of manufacture of any one of embodiments 20 to 24, comprising at least one layer of absorbent nonwoven fabric.

[0223] (26) The article of manufacture of claim 25, wherein the absorbent nonwoven fabric is comprised of an aliphatic polyester polymer or copolymer of one or more monomers selected from the group consisting of lactic acid, lactide (including L-, D-, meso, and D,L mixtures), glycolic acid, glycolide, e-caprolactone, p-dioxanone, and trimethylene carbonate. 27. The article of manufacture of claim 25 or 26, wherein the composition is substantially uniformly dispersed throughout the nonwoven fabric and / or disposed on a surface of the nonwoven fabric. (28) A composition according to any one of embodiments 1 to 19 or an article of manufacture according to any one of embodiments 20 to 27 for use in topical administration. (29) The composition according to any one of embodiments 1 to 19 or the article of manufacture according to any one of embodiments 20 to 27 for use in a tissue adhesive. (30) The composition according to any one of embodiments 1 to 19 or the article of manufacture according to any one of embodiments 20 to 27 for use in treating, inhibiting, arresting or slowing bleeding or any disorder related thereto.

[0224] (31) A method for treating, suppressing, arresting or slowing bleeding or any disorder related thereto, comprising topically administering to a wound tissue of a subject in need thereof an effective amount of a composition according to any one of embodiments 1 to 19 or an article of manufacture according to any one of embodiments 20 to 27. (32) The method of embodiment 31, wherein the bleeding is at least one of injury-induced bleeding, surgery-induced bleeding, or trauma-induced bleeding. (33) The method of embodiment 31 or 32, wherein the bleeding occurs during a surgical procedure. (34) The method of embodiment 33, wherein the surgical procedure comprises abdominal surgery, cardiovascular surgery, thoracic surgery, head and neck surgery, dural surgery, spinal surgery, pelvic surgery, a skin procedure, or a subcutaneous tissue procedure. (35) The method according to any one of embodiments 31 to 34, comprising applying the composition or the article of manufacture to the wound tissue.

[0225] (36) The method of embodiment 35, wherein the pressing is for a time sufficient to allow adhesion of the composition or article to at least a portion of the wound tissue.

Claims

1. 1. A composition comprising one or more coagulation proteins and albumin, wherein the one or more coagulation proteins and the albumin are at a combined concentration of at least 90% based on total protein weight, the one or more coagulation proteins comprise fibrinogen and fibronectin, the fibronectin being in an amount of less than about 0.5% based on total protein weight or absent, and the weight ratio of the albumin to the fibrinogen is at least 1:15, respectively.

2. 10. The composition of claim 1, wherein the one or more coagulation proteins and the albumin are at a total concentration of about 90% to about 99.5%.

3. 2. The composition of claim 1, wherein the one or more coagulation proteins and the albumin are at a combined concentration of at least 95% based on total protein weight.

4. The composition of claim 1 , wherein the coagulation protein comprises at least 90% fibrinogen.

5. The composition of claim 1 , wherein the coagulation protein comprises at least 95% fibrinogen.

6. The composition of claim 1 , wherein the fibronectin is in an amount of less than about 0.3% by weight.

7. 2. The composition of claim 1, wherein the weight ratio of said albumin to said fibrinogen is from 1:1 to 1:

15.

8. 2. The composition of claim 1, wherein the weight ratio of said albumin to said fibrinogen is from 1:1 to 1:

10.

9. 2. The composition of claim 1, wherein the weight ratio of said albumin to said fibrinogen is from 1:1.6 to 1:

3.

10. 2. The composition of claim 1, wherein the albumin content is from about 0.5% to about 55% by weight.

11. 2. The composition of claim 1, wherein the albumin content is from about 4.5% to about 55% by weight.

12. 2. The composition of claim 1, wherein the albumin content is from about 5% to about 43% by weight.

13. The composition of claim 1 in a dry form.

14. The composition of claim 1 in powder form.

15. 10. The composition of claim 1, characterized by a tissue adhesion of at least 40 N / m as measured by a peel force test.

16. The composition of claim 1 comprising thrombin.

17. 17. The composition of claim 16, wherein the albumin content is from about 5% to about 43% by weight.

18. 17. The composition of claim 16, comprising less than about 0.5% fibronectin.

19. 1. A composition comprising one or more coagulation proteins, albumin, and thrombin, wherein the one or more coagulation proteins and the albumin are at a combined concentration of at least 90% based on total protein weight, the one or more coagulation proteins comprise fibrinogen and fibronectin, the fibronectin being in an amount of less than about 0.5% based on total protein weight or absent, the weight ratio of the albumin to the fibrinogen is at least 1:15, respectively, and the composition is in powder form.

20. An article of manufacture comprising the composition of claim 1.

21. 21. The article of manufacture of claim 20, which is a wound dressing.

22. 21. The article of claim 20 comprising at least one layer of absorbent woven or knitted fabric.

23. 23. The article of manufacture of claim 22, wherein the absorbent woven or knitted fabric comprises one or more oxidized polysaccharides.

24. 23. The article of manufacture of claim 22, wherein the absorbent woven or knitted fabric comprises oxidized cellulose (OC), oxidized regenerated cellulose (ORC), or a combination thereof.

25. 21. The article of manufacture of claim 20 comprising at least one layer of absorbent nonwoven.

26. 26. The article of manufacture of claim 25, wherein the absorbent nonwoven fabric is comprised of an aliphatic polyester polymer or copolymer of one or more monomers selected from the group consisting of lactic acid, lactide (including L-, D-, meso, and D,L mixtures), glycolic acid, glycolide, e-caprolactone, p-dioxanone, and trimethylene carbonate.

27. 26. The article of manufacture of claim 25, wherein the composition is substantially uniformly dispersed throughout the nonwoven fabric and / or disposed on a surface of the nonwoven fabric.

28. 21. The composition of claim 1 or the article of manufacture of claim 20 for use in topical administration.

29. 21. The composition of claim 1 or the article of manufacture of claim 20 for use in a tissue adhesive.

30. 21. The composition of claim 1 or the article of manufacture of claim 20 for use in treating, inhibiting, arresting or slowing bleeding or any disorder related thereto.

31. A method for treating, inhibiting, arresting or slowing bleeding or any disorder related thereto, comprising topically administering to wound tissue of a subject in need thereof an effective amount of a composition described in claim 1 or 19 or 20.

32. 32. The method of claim 31, wherein the bleeding is at least one of injury-induced bleeding, surgery-induced bleeding, or trauma-induced bleeding.

33. 32. The method of claim 31, wherein the bleeding is during a surgical procedure.

34. 34. The method of claim 33, wherein the surgical procedure comprises abdominal surgery, cardiovascular surgery, thoracic surgery, head and neck surgery, dura surgery, spinal surgery, pelvic surgery, skin procedure, or subcutaneous tissue procedure.

35. 32. The method of claim 31, comprising applying the composition or article of manufacture to the wound tissue.

36. 36. The method of claim 35, wherein the pressing is for a time sufficient to allow adhesion of the composition or article to at least a portion of the wound tissue.

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