Hemostatic sealant powder for sealing leaks in tissues

Abstract

Disclosed herein are powdered compositions comprising fibrinogen, thrombin, a glycerol-aqueous solution, and optionally oxidized cellulose (OC), wherein glycerol is present at a concentration ranging from about 0.7% to about 6% by weight, and OC is present at or absent at a concentration of less than about 9% by weight. Methods for preparing the compositions, as well as the use of the compositions in methods for treating hemorrhagic tissue and sealing leakage in tissue, are further disclosed.

Description

Technical Field

[0001] The present invention relates, inter alia, to a hemostatic sealant powder comprising a blend of fibrinogen and thrombin for use, for example, in sealing leaks in tissue.

Background Art

[0002] In various situations during surgery, therapeutic agents are spread in the human body to prevent postoperative bleeding. Existing drugs have problems. For example, it is difficult to place a sheet in minimal invasive surgeries (MIS).

[0003] The selection of an appropriate method or product for controlling bleeding depends on many factors, including, but not limited to, the severity of bleeding, the anatomical location of the bleeding source and the proximity of adjacent important structures, whether the bleeding is from a discrete bleeding source or from a larger surface area, the visibility and accurate identification of the bleeding source, and access to the bleeding source.

[0004] In efforts to address the above problems, substances for controlling excessive bleeding have been developed. Topical Absorbable Hemostat (TAH) is widely used in surgical applications. TAH includes products based on oxidized cellulose (OC), gelatin, collagen, chitin, chitosan, etc. To improve hemostatic performance, scaffolds based on the above substances can be combined with biologically derived coagulation factors such as thrombin and fibrinogen.

[0005] Fibrin sealants, also known as fibrin glue, have been used in hospitals for decades. Fibrin sealants often consist of two liquid components, a fibrinogen-containing component and a thrombin-containing component, which are stored frozen due to their inherent instability. Occasionally, fibrin sealant products consist of two lyophilized components, which require reconstitution immediately before use and delivery via a combined syringe or other dual-component delivery device.

[0006] Dry sealant powders, when mixed with fluids, are useful as flowable, extrudeable, and injectable hemostatic agents, forming pastes or slurries made from plasma-derived components, such as (a) fibrinogen mixtures mainly composed of fibrinogen, along with optionally catalytic amounts of factor XIII, and (b) high-potency thrombin. Due to their biodegradability, as well as their bactericidal and hemostatic properties, OC-based materials such as oxidized regenerated cellulose (ORC) have long been used as topical hemostatic agents. OC and ORC-based materials are also used as adhesion barriers. ORC-based products are used in a variety of surgical procedures, including neurosurgery, abdominal surgery, cardiovascular surgery, thoracic surgery, head and neck surgery, pelvic surgery, and skin and subcutaneous tissue procedures. Several methods are known for forming various types of hemostatic agents based on OC materials, whether made in powder, woven, nonwoven, knitted, or other forms. Hemostatic agents currently in use include powders or fabrics containing ORC.

[0007] For example, the effectiveness of fibrin sealants in sealing defects in constantly moving tissues such as gastrointestinal and lung tissues has been reported. However, there are inconsistencies in these reports. For instance, one approach to reducing postoperative defects such as leakage at staple or suture lines after gastrointestinal resection is to use staple / suture line reinforcement materials. Various staple / suture line reinforcement products are commercially available. Another approach is to use fibrin as an absorbable staple line reinforcement to prevent postoperative leakage.

[0008] U.S. Patent No. 10111980(B2) discloses a dry composition comprising one or more polyols that forms a substantially homogeneous paste suitable for use in hemostatic procedures when an aqueous medium is added. The paste spontaneously reconstitutes when a liquid is added.

[0009] U.S. Patent Publication No. 20210060204(A1) describes a biocompatible hemostatic composition and a biocompatible tissue sealant, which, when used together, provide a safe and effective method for achieving hemostasis. The biocompatible composition and sealant can be applied either to the surface of a patient's body or inside a body cavity.

[0010] International Publication No. 2021128050(A1) discloses a non-flowing and deformable hemostatic composition comprising xerogel crosslinked powdered polysaccharide dispersed in a substantially anhydrous blend of glycerol and polyethylene glycol (PEG), wherein PEG and glycerol are present in a weight ratio of greater than 1:1.3 to less than 1:2.7, and the powder content in the composition is greater than 50% by weight.

[0011] U.S. Patent No. 1,1413335 discloses a method for preparing a hemostatic non-aqueous solvent in a high shear mixing (HSM) apparatus, while allowing the volatile non-aqueous solvent to evaporate through any suitable port, for example, through the air seal of a mixing blade / impeller and / or through a vacuum tube, and for performing high shear mixing and regular mixing and / or stirring. [Overview of the project] [Means for solving the problem]

[0012] The present invention relates, in particular, to a powdered blend of fibrinogen, thrombin, and a small amount of glycerol-aqueous solution.

[0013] The object of the present invention is to provide a dry powder composition that can rapidly dissolve in an aqueous medium to form a gel layer with significantly increased gel strength and adhesive strength. The sealant powder is suitable for both hemostasis and sealing and is produced by a novel scale-up granulation process.

[0014] The granulation process for producing the powder involves utilizing the reaction between the biological materials themselves (fibrinogen and thrombin) and water to form a small amount of fibrin gel as a binder, without the need to introduce a new polymer binder, thereby reducing safety and biocompatibility concerns. Briefly, the novel high-shear mixer granulation process of the present invention involves raw materials of biological components (dry powder), then mixing, then spraying a specific amount of water for injection (WFI), then mixing with a specific ratio of glycerol, and then granulating. It is also advantageous to use glycerol (instead of a low-boiling solvent) because it may be difficult to reduce the low-boiling solvent in the final product to a safe level.

[0015] The composition can be easily applied to the required areas, including hard-to-reach areas of the body. The disclosed fibrin sealant powder containing glycerol has better gel strength and, for example in a dog model, can enhance adhesive strength with the possibility of reducing the amount of ORC, and the solubility appears to be equivalent to or better than that in the absence of ORC. The water content of the fibrin sealant powder containing glycerol may be less than 2.5%, which has a significant effect on powder stability.

[0016] Therefore, the present invention is based, in particular, on the remarkable finding that a microaggregate composition comprising specific amounts of fibrinogen, thrombin, glycerol, fibrin, and optionally ORC, produced by a novel granulation process (e.g., high-shear mixing (HSM)), can rapidly dissolve to form a gel layer, sealing tissue and stopping bleeding, and can form a gel with sufficient strength and adhesion to be used to seal leakage from damaged tissue without tamponade.

[0017] Therefore, the formed powder can be used immediately and sprayed directly onto the target area.

[0018] According to aspects of the present disclosure, there is a powdered composition comprising fibrinogen, thrombin, a glycerol-aqueous solution, and optionally oxidized cellulose (OC), wherein (i) glycerol is present in a concentration ranging from about 0.7% by weight to about 6.5% by weight, (ii) water is present in a concentration of less than 3% by weight, and (iii) OC is present in a concentration of less than 9% by weight or is not present.

[0019] In some embodiments, the composition further comprises fibrin.

[0020] In some embodiments, OC includes oxidized regenerated cellulose (ORC).

[0021] In some embodiments, the composition does not contain OC and / or ORC.

[0022] In some embodiments, the composition does not contain a non-aqueous low-boiling solvent.

[0023] In some embodiments, the composition does not contain hydrofluoroethers.

[0024] In some embodiments, the composition is in a rapidly soluble solid form in an aqueous medium, for example, dissolving in less than 3 minutes at 25°C.

[0025] In some embodiments, the powder comprises particles characterized by a D90 size distribution of up to 500 μm.

[0026] In some embodiments, the composition is in a state ready for immediate use for hemostatic and / or sealing applications.

[0027] In some embodiments, the composition is characterized by forming a gel in an aqueous medium, and the gel has a high gel strength in the range of, for example, 85 kPa to 160 kPa.

[0028] In some embodiments, the composition lacks additional polymer binders other than fibrin.

[0029] In some embodiments, the composition is for use, for example, in minimally invasive surgery to treat bleeding tissue and / or to prevent air leakage in organs that require prevention of leakage, such as air leakage.

[0030] In some embodiments, the composition further comprises a calcium salt.

[0031] In some embodiments, the composition is deposited on an article (e.g., a patch).

[0032] According to an aspect of the present invention, there is provided a method for producing a powdered hemostatic composition, comprising: (a) mixing solid fibrinogen, thrombin, and optionally ORC fibers to form a mixture; (b) inserting the mixture into a high-shear mixing reactor; (c) adding a solution of glycerol and water to the mixture in the reactor; and (d) drying and sieving the composition obtained in step (c) to thereby form a powdered hemostatic composition.

[0033] In some embodiments, glycerol is present in the solution at a concentration in the range of greater than about 10% by weight to less than about 40% by weight.

[0034] In some embodiments, water is added in an amount that prevents complete coagulation of fibrinogen.

[0035] In some embodiments, the mixture is substantially free of OC and / or ORC.

[0036] According to aspects of the present invention, in any embodiment, a powder composition is provided which can be obtained by the method disclosed herein.

[0037] According to aspects of the present disclosure, a composition is provided comprising a mixture of solid fibrinogen, thrombin, and optionally ORC fibers in a solution of water and glycerol, wherein the glycerol is present in the solution at a concentration ranging from more than about 10% by weight to less than about 40% by weight. In some embodiments, the composition further comprises fibrin.

[0038] Unless otherwise defined, all technical and / or scientific terms used herein have the same meaning as those generally understood by those skilled in the art in which the present invention pertains. Similar or equivalent methods and materials may be used to perform or test embodiments of the present invention, but exemplary methods and / or materials are described below. In case of any inconsistency, this specification shall prevail, including definitions. The materials, methods, and examples are illustrative and not necessarily intended to be limiting. [Brief explanation of the drawing]

[0039] To better understand the subject matter disclosed herein and to illustrate how it can actually be put into practice, embodiments are described here by non-limiting examples with reference to the accompanying drawings. [Figure 1A] These are images of the agglomerated powder (sample 15-02, see Table 6) prepared according to the embodiments of this disclosure, namely a photographic image (Figure 1A) and a scanning electron microscopic (SEM) image (Figure 1B). [Figure 1B] These are images of the agglomerated powder (sample 15-02, see Table 6) prepared according to the embodiments of this disclosure, namely a photographic image (Figure 1A) and a scanning electron microscopic (SEM) image (Figure 1B). [Figure 2A] These graphs show the tensile stress (Figure 2A) and tensile strain (Figure 2B) of a fibrin gel prepared from agglomerated powder prepared according to embodiments of this disclosure, compared to a gel prepared from powder generated by hydrofluoroether (HFE) (based on Tables 4 and 5, respectively). [Figure 2B] These graphs show the tensile stress (Figure 2A) and tensile strain (Figure 2B) of a fibrin gel prepared from agglomerated powder prepared according to embodiments of this disclosure, compared to a gel prepared from powder generated by hydrofluoroether (HFE) (based on Tables 4 and 5, respectively). [Figure 3A] This graph shows the tensile stress (Figure 3A) and tensile strain (Figure 3B) of a fibrin gel prepared from agglomerated powder prepared according to embodiments of this disclosure (based on Table 6). [Figure 3B] This graph shows the tensile stress (Figure 3A) and tensile strain (Figure 3B) of a fibrin gel prepared from agglomerated powder prepared according to embodiments of this disclosure (based on Table 6). [Figure 4]These are photographic images showing the results of solubilization tests of comparative compositions prepared using various amounts of glycerol in a water-glycerol dispersant: top figure - 10% glycerol (v / v%); middle figure - 20% glycerol (v / v%); bottom figure - 40% glycerol (v / v%). Each figure for glycerol volume is divided into various amounts of oxidized regenerated cellulose (ORC) - "10:1" means that 100g of the biological component corresponds to 10g of the ORC component, "20:1" means that 100g of the biological component corresponds to 5g of the ORC component, and "0" means that no ORC is added to the formulation (from top to bottom in each figure for glycerol volume). Solubility was tested at 0 min, 1 min, 2 min, and 3 min (from left to right in each figure for glycerol volume); see also Table 7. [Figure 5] This graph shows the comparative results of the solubilization tests of the comparative compositions based on Figure 4 and Table 7 below. [Figure 6] This graph shows the water content % (by weight) of powders prepared according to several embodiments, as measured by the Karl Fischer titration method (based on Table 8). [Figure 7] This photographic image shows that the solubility of HFE granular samples is relatively low. [Figure 8A] These are photographic images showing the adhesion properties of glycerol granule powder to porcine lung tissue in this ex vivo test (Figure 8A - before powder application, Figure 8B - after powder application and sealing). [Figure 8B] These are photographic images showing the adhesion properties of glycerol granule powder to porcine lung tissue in this ex vivo test (Figure 8A - before powder application, Figure 8B - after powder application and sealing). [Figure 9A] These are photographic images showing steps 1, 2, 4, and 5, respectively, of the liver resection model described in the Examples chapter. [Figure 9B] These are photographic images showing steps 1, 2, 4, and 5, respectively, of the liver resection model described in the Examples chapter. [Figure 9C] These are photographic images showing steps 1, 2, 4, and 5, respectively, of the liver resection model described in the Examples chapter. [Figure 9D] These are photographic images showing steps 1, 2, 4, and 5, respectively, of the liver resection model described in the Examples chapter. [Modes for carrying out the invention]

[0040] Commercial formulations of thrombin and fibrinogen blends exist in powder form. Application of powder to limited, specific sites can be difficult to achieve. The object of the present invention is to provide a powder composition that can rapidly dissolve in an aqueous medium to form a gel layer with significantly increased gel strength (also called "tensile strength"), and that can simultaneously address hemostasis and tissue leakage.

[0041] The disclosed powder composition is characterized in that its components are uniformly distributed therein.

[0042] According to aspects of the present disclosure, there are provided powder compositions comprising fibrinogen, thrombin, a glycerol-aqueous solution, and optionally oxidized cellulose (OC), wherein (i) glycerol is present at a concentration in the range of about 0.7% to about 6.5% by weight, and (ii) OC is present at a concentration of less than about 8.50% by weight, or is absent in some embodiments.

[0043] The absence of water in such a suspension is counterintuitive, but without being bound by any particular theory or mechanism, we can assume that after mixing the powder in a dispersant, the uniformly dispersed biopharmaceutical forms fibrin in a glycerol / water system. When applied to a bleeding site, the fibrin matrix acts as a binder, ensuring uniform distribution of the powder components.

[0044] The amount of water present is small because it is added in an amount that prevents complete coagulation of fibrinogen. That is, in some embodiments, the amount of water in the powder is less than 3% by weight. In some embodiments, the amount of water in the powder is less than 2.5% by weight. In some embodiments, the amount of water in the powder is less than 2% by weight. In some embodiments, the amount of water in the powder is in the range of about 1.5% to about 2.5% by weight (see, for example, Figure 6).

[0045] Therefore, in some embodiments, the composition is homogeneous. As used herein, “homogeneous” means that the composition and texture are uniform throughout, i.e., the powder and small amounts of dispersant are substantially uniformly dispersed throughout them. In this specification, “substantially uniformly dispersed” means that the differences in the concentrations of the components in the powder vary throughout them within a range of less than ±30% by weight, less than ±20% by weight, less than ±10% by weight, or less than ±5% by weight, i.e., there is no significant solid-liquid phase separation. Typically, the term “homogeneous” refers to a product that appears uniform and withstands liquid / solid phase separation at 8–40°C, 10–30°C, or 20–30°C (e.g., less than 4% by weight).

[0046] As used herein, the terms “powder” or “powdered composition” generally refer to a solid having fine particles. The powder may also be a granular material.

[0047] In some embodiments, the powder is in the form of aggregates or granules.

[0048] The term "aggregate" describes particles formed from combined components. Aggregates may optionally be produced by one of the following steps: humidifying the powder composition; forming aggregates by compressing the powder, for example, with rollers and / or striking the powder; dehumidifying; grinding; sieving the aggregates; and optionally, placing the resulting aggregates into a storage container or delivery device. The terms "granules" or "granular material" may particularly mean aggregates of discrete solid particles, typically less than 550, less than 500 micrometers, less than 400 micrometers, or less than 350 micrometers. Aggregates are also called "microaggregates."

[0049] In exemplary embodiments, a high-shear mixture is used to construct aggregates from fibrinogen, thrombin, and optionally CaCl2 particles, buffers such as lysine, albumin (present in the source from which the fibrinogen is collected), and / or ORC. In other embodiments, the composition is in the form of a mixture. The mixture may comprise, in exemplary embodiments, a combination of spray-dried first fine particles containing fibrinogen and spray-dried second fine particles containing thrombin, such as porcine thrombin, which together form particles.

[0050] The term “solid” refers to the state of a compound or composition at room temperature (e.g., 25°C) and atmospheric pressure (760 mmHg), i.e., a highly viscous compound or composition that retains its form during storage. In this application, this term also refers to non-fluid particles or dissolved substances. In contrast to “liquid” compounds and compositions, solids do not flow under their own weight.

[0051] In some embodiments, the powder is spray-dried and / or high-shear mixed.

[0052] Embodiments of spray drying and high-shear mixing are described throughout this specification.

[0053] The term “spray drying” is used herein in a broad sense and includes, but is not limited to, processes for converting solids dissolved or suspended in liquids into powder form. Typically, spray drying is a drying method used to produce powders from solutions, suspensions, or emulsions by spraying them through a spray nozzle into a stream of hot air and drying them immediately. A spray drying process for producing thrombin and fibrinogen powders that retain their activity can be controlled by several process parameters, including, but is not limited to, column airflow and temperature, nozzle size and spray air velocity, and material flow rate. Non-limiting exemplary parameters used to establish a robust process for producing a blend of thrombin and fibrinogen (e.g., in powder form) that retains their activity are provided in the following Examples chapter.

[0054] The term "high-shear mixing" generally refers to turbulent mixing. In some embodiments, high-shear mixing is characterized by mixing blade speeds and / or cutting blade speeds in the range of 150 to 500 rpm.

[0055] Additionally or alternatively, other methods may be used to obtain the dried composition, such as freeze-drying and then micronizing the fibrinogen solution and / or thrombin solution, for example. Typically, after freeze-drying the solution, a porous, spongy solid material called a "cake" (which may also be referred to herein as a "solid composition") is obtained.

[0056] The term "micronization" is used, for example, when the diameter of particles produced from a solid composition is only a few micrometers. Micronization can be achieved by processes including, but not limited to, jet milling, pearl ball milling, high-pressure homogenization, RESS (rapid expansion of supercritical solution) process, SAS (supercritical poor solvent) method, or PGSS (particles from gas-saturated solution) method. Typically, but not limited to, micronization results in a reduction of the size of protein powder by 30 to 400 times from its original size.

[0057] In some embodiments, the composition is stable.

[0058] The terms “stable” and “stability,” when referring to the disclosed powder, mean, for example, that at least 70% of the activity of the active ingredient in thrombin remains after a specific period of time at a specific temperature, i.e., that it can form fibrin clumps.

[0059] The powder can be compressed by roller compression or slugging compression, and then subjected to the steps of preliminary crushing, dehumidification, and subsequently final grinding. The resulting aggregates can be separated into a target hemostatic aggregate fraction by, for example, sieving for desired size and / or distribution separation (e.g., less than 500 μm or less than 250 μm, within the specified ranges of D50 and D90).

[0060] The terms "D50" and "D90" refer to the fact that 50% and 90% (by number or volume) of the powder have a size equal to or less than the corresponding value.

[0061] In some embodiments, the aggregates are characterized by a size distribution of D90 less than 500 μm and D50 less than 180 μm. In some embodiments, the aggregates are characterized by a D50 in the range of approximately 150 μm to approximately 180 μm. Refer to Figure 1B, which shows SEM images of powder produced according to embodiments of this disclosure.

[0062] The term "glycerol" refers to 1,2,3-propanetriol, also known as "glycerin," and can also include its derivatives. The term "derivative" refers to a compound that contains essential elements of the parent substance.

[0063] In some embodiments, glycerol is present at concentrations ranging from about 0.7% by weight to about 6.5% by weight. In some embodiments, glycerol is present at concentrations ranging from about 1% by weight to about 6% by weight. In some embodiments, glycerol is present at concentrations of about 0.5% by weight, 1% by weight, 1.5% by weight, 2% by weight, 2.5% by weight, 3% by weight, 3.5% by weight, 4% by weight, 4.5% by weight, 5% by weight, 5.5% by weight, 6% by weight, or 6.5% by weight (including any values ​​and ranges between them).

[0064] As used herein, “thrombin” refers to the activating enzyme resulting from the proteolytic cleavage of prothrombin (factor II). Thrombin can be produced by various methods known in the art, and includes, but is not limited to, recombinant thrombin and plasma-derived thrombin. Human thrombin is a 295-amino acid protein composed of two polypeptide chains joined by disulfide bonds. Both human thrombin and non-human (e.g., bovine and porcine) thrombin may be used within the scope of this disclosure.

[0065] The origin of the thrombin used in the present invention may be derived from one or more sources, including but not limited to plasma (e.g., porcine plasma), recombinant bacteria and / or cells (Vu et al., 2016, J. Viet. Env. 8(1):21-25), whole blood (which may or may not be pooled from several transfusions), and / or blood fractions (which may be pooled from multiple transfusions, e.g., plasma). Thrombin is available from manufacturers such as Johnson and Johnson, Baxter, and CSL Behring as a standalone product, e.g., EVITHROM®, or as a component of a product, e.g., EVICEL®, TISEEL®, Beriplast®, etc.

[0066] In some embodiments, thrombin is present in the powder at a concentration of 3% to 10% by weight or 4% to 5% by weight. In some embodiments, thrombin is present in the powder at a concentration of 3%, 3.5% by weight, 4% by weight, 4.5% by weight, 5% by weight, 5.5% by weight, 6% by weight, 6.5%, or 7% by weight (including any value and range between these). The activity may range from 100 to 1500 IU per dose, or in some embodiments, about 300 to 1200 IU of thrombin powder.

[0067] The term "fibrinogen" is intended to include any type of fibrinogen, even if not specified further. "Fibrinogen" refers to monomeric and dimeric fibrinogen molecules, hybrid molecules, and their variants, whether naturally occurring, modified, or synthetic, possessing the monomeric structure (AαBβγ). The term "fibrinogen" may refer to human-derived fibrinogen, but may also include fibrinogen from any species, particularly mammalian species, such as fibrinogen produced from porcine plasma.

[0068] Throughout this specification, any combination of thrombin, fibrinogen, and an amino acid, such as lysine, will also be referred to as a “biological component” when present in a composition or powder.

[0069] In some embodiments, fibrinogen is present in concentrations of 60% to 95% or 70% to 90% by weight of the blend. In some embodiments, thrombin is present in concentrations of 60%, 61%, 62%, 63%, 64%, 65%, 66%, 67%, 68%, 69%, 70%, 71%, 72%, 73%, 74%, 75%, 76%, 77%, 78%, 79%, 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, or 95% by weight of the blend (including any values ​​and ranges between them).

[0070] The term "oxidized cellulose" (or "OC") refers to a cellulose derivative in which at least a portion of a primary alcohol group, for example, the carbon at position 6 of an anhydrous glucose unit, is oxidized to a carboxylic acid and optionally functionalized. OC may constitute materials, products, articles, or compositions that contain or essentially consist of OC, such as bandages, fibrin glue, synthetic glue, pads, matrices, powders, tabs, pills, sutures, fibers, stents, implants, scaffolds, solutions, gels, waxes, gelatin, and the like.

[0071] In some embodiments, OC (e.g., ORC) is in the form of multiple fibers. In some embodiments, OC (e.g., ORC) is in the form of multiple fibers having a median diameter of less than 80 μm. In some embodiments, OC (e.g., ORC) is in the form of multiple fibers having a median diameter in the range of 50 to 100 μm or 60 to 80 μm, for example, 50, 60, 70, 80, 90, 100 μm (including any values ​​and ranges in between).

[0072] Oxidized cellulose (OC) can be produced by applying an oxidizing agent to cellulose. The oxidizing agent can be selected from, but is not limited to, chlorine, hydrogen peroxide, peracetic acid, chlorine dioxide, nitrogen dioxide, persulfates, permanganates, dichromates-sulfuric acids, hypochlorous acid, hypohalites, periodates, or any combination thereof, and / or various metal catalysts. Depending on the properties of the oxidizing agent and the reaction conditions, the oxidized cellulose may contain carboxylic acid groups, aldehyde groups, and / or ketone groups in place of or in addition to the original hydroxyl groups of the starting material cellulose.

[0073] The term "contact" or any grammatical variation thereof is used in its broadest sense to refer to any type of binding action that brings, for example, a hemostatic composition into close proximity with water, an aqueous solution, or blood, such that a blood clot or gel may be formed.

[0074] In some embodiments, the composition is substantially devoid of OC or ORC, partly because the addition of ORC may reduce the gel strength properties (see, for example, Table 6 and Figure 3A in the Examples section below).

[0075] In some embodiments, the water absorbent (e.g., ORC) is present in concentrations ranging from 3 to 15% by weight, for example, in concentrations of up to 3% by weight, up to 8% by weight, up to 10% by weight, up to 15% by weight, up to 20% by weight, or up to 25% by weight of the blend (i.e., the powdered composition), if it is not present. In some embodiments, the water absorbent is present in concentrations of about 9% by weight of the powder.

[0076] In some embodiments of any part of the compositions disclosed herein, the powder may contain additives, such as calcium salts, and / or one or more excipients selected from, for example, one or more amino acids, albumin, sugars, and / or sugar derivatives.

[0077] The term "additive" is understood to mean any substance that can be added to a composition or that originates from the raw materials, and may include active additives such as calcium salts as described below.

[0078] As used herein, the term “excipient” means, for example, an inactive or non-therapeutic agent added to a pharmaceutical composition to provide a desired viscosity or stabilizing effect.

[0079] Calcium is an important element in the coagulation cascade and may be necessary to activate factor XIII, which cross-links and stabilizes fibrin to form factor XIIIa, generating an insoluble mass.

[0080] Accordingly, in some embodiments of any part of the disclosed composition, the blend further comprises additives such as calcium, for example, but is not limited to these. The calcium used in the present invention may be in the form of a salt, such as a calcium chloride salt. Alternatively, additional salts such as calcium acetate and / or calcium citrate may be used.

[0081] In some embodiments, the disclosed compositions are provided in a kit, in any of its embodiments.

[0082] In the kit, the calcium salt may be provided in a composition containing the blend. Alternatively, the excipient and / or calcium salt may be provided in a separate container in the kit, or the excipient and / or calcium salt may be provided in the kit in the same container as the blend components.

[0083] In some embodiments of the kit or composition, a calcium salt, such as calcium chloride (CaCl2), is present in the blend at a concentration of 0.5% to 4% or 2.5% to 3.5% by weight. In some embodiments, a calcium salt, such as calcium chloride, is present in the blend at a concentration of 0.5%, 1%, 1.5%, 2%, 2.5%, 3%, 3.5%, or 4% by weight (including any values ​​and ranges between these).

[0084] OC is known to be acidic, and this acidity immediately denatures plasma proteins such as thrombin and fibrinogen. Therefore, simultaneous administration of oxidized cellulose with thrombin and / or fibrinogen may negate the effects of plasma proteins.

[0085] In some embodiments, a buffer is added to the composition to adjust the pH. The buffer allows for pH neutralization and provides increased adhesion and sealing properties compared to a control composition containing the same aggregate with components that do not contain a buffer.

[0086] In some such embodiments, a buffer, e.g., Tris, or Tris(hydroxymethyl)aminomethane buffer, or lysine, is added in powder form to the fibrinogen, thrombin, and ORC blend / mixture / powder to adjust the pH (for example, to about pH 7). In some embodiments, the buffer, e.g., Tris or lysine, is present in the blend / mixture / powder at concentrations of 0–25% by weight, 1%–8% by weight, or 1%–5% by weight. In some embodiments, the buffer, e.g., Tris or lysine, is present at concentrations (%) of 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, or about 25% by weight (including any values ​​and ranges between these).

[0087] In some embodiments of any part of the composition, the blend further comprises an excipient selected from an amino acid, such as lysine. In some embodiments of any part of the kit and / or composition, the blend comprises OC and lysine.

[0088] In some embodiments, the composition does not contain a non-aqueous low-boiling solvent. The term "non-aqueous low-boiling solvent" refers to an organic solvent with a boiling point of 150°C or less at atmospheric pressure. In exemplary embodiments, the composition is substantially free of hydrofluoroethers (HFEs).

[0089] In some embodiments, the composition further comprises fibrin.

[0090] The term “fibrin” refers not only to fully solidified fibrinogen, but also to any mixture of fibrin and fibrin that may result during the formation of fibrin from fibrinogen using thrombin, and therefore includes any conceivable ratio of fibrinogen / fibrin and any degree of gelation and / or solidification, as long as it does not adversely affect the final paste-like texture of the composition. Furthermore, the term “fibrin” further includes any partially or fully crosslinked, gelled, or solidified form.

[0091] Therefore, it is understood that fibrin itself exists as a high-molecular-weight, insoluble matrix containing fibrils, and that each fibril contains many fibrin molecules.

[0092] In some embodiments, the composition lacks additional polymer binders other than fibrin.

[0093] In some embodiments, the composition further comprises additional pharmaceutically active agents contained within or on the surface of the composition, in any embodiment thereof.

[0094] In some embodiments, the pharmaceutically active agent is selected from the group consisting of therapeutic agents and labeling agents.

[0095] In some embodiments, the therapeutic activator is selected from the group consisting of stem cells, growth factors, bone morphogenetic proteins, cells, cytokines, hormones, pharmaceuticals, minerals, therapeutic plasmids, and combinations thereof. As described herein, in some embodiments, the composition may further contain at least one biological activator. Non-limiting biological activators that may be included in the composition include calcium, as well as therapeutic agents such as antibiotics, anti-inflammatory agents, growth factors, or coagulation factors. Specifically, the disclosed composition may also contain components that promote mass formation, such as calcium salts, factor VIII, factor XIII, fibronectin, vitronectin, von Willebrand factor (vWF), etc., and these components may be provided as separate components or combined with powders.

[0096] In some embodiments, the composition is in a rapidly soluble solid form in an aqueous medium.

[0097] The term "aqueous solution" refers to a solution containing water as a solvent. This term may include water, saline solution, or a bleeding site. In some embodiments, the aqueous medium is selected from water, saline solution, calcium chloride solution, or a buffered aqueous medium.

[0098] In exemplary embodiments, the aqueous solution includes physiological saline. As used herein, the term “physiological saline” means water containing a salt concentration greater than about 0.5% by weight and less than about 20% by weight, for example, a solution of about 0.9 w / w% sodium chloride in desalinated water.

[0099] For the purposes of this disclosure, the term “rapidly soluble solid” means that after the powder is placed in water, the powder dissolves in less than approximately 350 seconds, preferably less than approximately 200 seconds (see, for example, Figures 4 and 5).

[0100] The object of the present invention is to provide a composition that can be easily applied to the necessary bleeding site, particularly in areas that are difficult to reach on the body. A further advantage of the compositions of the present invention is that they are bioabsorbable and therefore can be left in place postoperatively without causing any side effects.

[0101] "Applied to the necessary bleeding site" means, for example, applying the composition locally to the surgical site where tissue is bleeding or to its vicinity.

[0102] The compositions disclosed herein may be used for any therapeutic purpose. The term “any therapeutic purpose” means any curative or preventive treatment in a subject. Exemplary therapeutic purposes include, but are not limited to, sealing perforations formed in tissues or organs (e.g., bone), anastomoses in blood vessels, joining of tissues (e.g., soft tissues), treatment or prevention of dura mater abnormalities (e.g., post-injection rupture and leakage, cracks or tears), treatment or prevention of bleeding, treatment or prevention of air leakage such as after lung resection, treatment or prevention of abnormalities after bowel perforation, treatment or prevention of abnormalities after anastomotic procedures performed in any tissue (e.g., uterus, esophagus, stomach, pancreas, pancreatic duct, gallbladder, bile duct, intestines (including small and large intestines), and rectum), and treatment of any tissue (e.g., uterus, esophagus, stomach, pancreas, pancreatic duct, gallbladder, bile duct, intestines (including small and large intestines), and rectum). The treatment or prevention of postoperative leakage in surgical procedures; prevention or reduction of postoperative leakage in staples or sutures (e.g., by applying the powdered composition according to the present invention alone or in combination with a matrix, e.g., a patch or pad, to at least a portion of the defects in staples / sutures, etc.); for secure attachment of prostheses (e.g., during hernia surgery); for strengthening staples / sutures; prevention or reduction of air leakage (e.g., alveolar air leakage); treatment or prevention of kidney defects; treatment or prevention of fistulas; treatment or prevention of cardiac defects (e.g., penetrating cardiac trauma); strengthening of artificial vascular prostheses; and treatment or prevention of cerebrospinal fluid leakage. In some embodiments, the composition or matrix according to any embodiment disclosed herein is for use in providing hemostasis, sealing leakage, and / or joining structures. In some embodiments, a method is provided for providing hemostasis, sealing leakage, and / or joining structures in subjects requiring hemostasis, sealing leakage, and / or joining structures, the method comprising the use of the composition or matrix according to any embodiment disclosed herein.

[0103] As used herein, the term “bleeding” refers to the spillage of blood from any component of the circulatory system. Thus, “bleeding” encompasses undesirable, uncontrolled, and often excessive bleeding associated with surgery, trauma, or other forms of tissue injury, as well as undesirable bleeding in patients with hemorrhagic disorders, and severe postpartum bleeding, including massive bleeding associated with cesarean section. The term “blood” or any grammatical variation thereof also includes blood fractions such as plasma.

[0104] As used herein, “wound” means any injury to any tissue of a patient that results in the loss of blood from the circulatory system and / or any other fluid from the patient’s body. The injury may be caused by any agent or source, including trauma, infection, or surgical intervention. Wounds may be located in soft tissue, such as organs, or in hard tissue, such as bone. Tissue may be internal, such as organs or blood vessels, or external, such as skin. The loss of blood may be internal, such as from a ruptured organ, or external, such as from a laceration.

[0105] In some embodiments, the composition further comprises additional pharmaceutically active agents contained within or on the surface of the composition, in any embodiment thereof.

[0106] In some embodiments, the pharmaceutically active agent is selected from the group consisting of therapeutic agents and labeling agents.

[0107] In some embodiments, the therapeutic activator is selected from the group consisting of stem cells, growth factors, bone morphogenetic proteins, cells, cytokines, hormones, pharmaceuticals, minerals, therapeutic plasmids, and combinations thereof. As described herein, in some embodiments, the composition may further contain at least one biological activator. Non-limiting biological activators that may be included in the composition include calcium, as well as therapeutic agents such as antibiotics, anti-inflammatory agents, growth factors, or coagulation factors. Specifically, the disclosed composition may also contain components that promote mass formation, such as calcium salts, factor VIII, factor XIII, fibronectin, vitronectin, von Willebrand factor (vWF), etc., and these components may be provided as separate components or combined in a blend.

[0108] Optionally, a pharmaceutically active agent is conjugated to a protein (i.e., fibrinogen or fibrin) matrix (e.g., covalently to the protein). This agent may be conjugated to the matrix after the matrix has been prepared, and / or to a component (e.g., a protein and / or reducing sugar), which is then used to prepare a protein matrix, such as cross-linked fibrin.

[0109] Alternatively or additionally, the agent is absorbed by a crosslinked protein matrix. Absorption can be achieved, for example, by contacting a crosslinked fibrinogen or fibrin matrix or a non-crosslinked fibrin matrix with a solution containing the agent (e.g., by immersion, soaking, or washing), or by adding the agent before crosslinking (e.g., by including the agent in a crosslinking solution, or by including the agent in a solution of fibrinogen and thrombin).

[0110] The compositions described herein, and the contents of the kits described above, may, if desired, be provided in packs or dispenser devices that can contain one or more unit dosage forms containing the compositions or components (e.g., fibrinogen) and / or reagents (e.g., aqueous solutions) for preparing the compositions. Packs such as blister packs may contain, for example, metal or plastic foil. The packs or dispenser devices may be accompanied by instructions for administration. The packs or dispenser devices may be accompanied by notices in the form prescribed by government agencies that regulate the manufacture, use, or sale of pharmaceuticals, which reflect the approval of such government agencies of the form of the composition for administration to humans or animals. Such notices may include, for example, the labeling approved by the U.S. Food and Drug Administration (FDA) for prescription drugs, or approved product inserts. Compositions containing the preparations of the present invention, formulated in a pharmaceutically acceptable carrier, may also be prepared for use for specified uses and / or for the treatment of specified conditions, as further detailed herein, placed in appropriate containers, and labeled. The term “preparation” means one that is physiologically suitable for therapeutic use.

[0111] It is understood that the compositions of the embodiments of this disclosure may be attached to or included in a medical device, for example, to promote wound healing.

[0112] In another aspect of this disclosure, any composition disclosed in any aspect or embodiment thereof is intended for use in a method of preparing a fibrin sealant in / on a target damaged tissue by, for example, applying the composition disclosed in any aspect and / or embodiment thereof to or near the surface of the tissue.

[0113] Another aspect of the present disclosure provides a method for preparing a fibrin sealant in / on a target damaged tissue by, for example, applying a composition disclosed in any aspect and / or embodiment thereof to the surface of the tissue as a hemostatic agent.

[0114] Another aspect of the present disclosure provides a method for preparing a composition according to that embodiment, comprising combining OC or ORC fibers with fibrinogen powder and thrombin powder under conditions that enable the formation of aggregated forms of ORC fibers, fibrinogen, and thrombin when the combined powder is combined with a dispersant comprising glycerol and an aqueous solution.

[0115] In some embodiments, such conditions allow for the conversion of fibrinogen to fibrin. In some embodiments, these conditions allow for the partial polymerization of fibrin, for example, by adding a specific amount of aqueous solution to a dispersant, as will be detailed throughout this specification.

[0116] In some embodiments, “partial conversion” refers to the conversion of 0.1% to 25% of fibrinogen to fibrin so as to form aggregated particles containing fibrinogen, thrombin, and optionally ORC, while preventing complete coagulation of fibrinogen. In some embodiments, “partial conversion” refers to the conversion of 0.5% to 10% of fibrinogen to fibrin so as to form aggregated particles containing fibrinogen, thrombin, and optionally ORC, while preventing complete coagulation of fibrinogen.

[0117] Conditions that enable the formation of powder / aggregate forms of ORC fibers (if present), fibrinogen, and thrombin include, for example, pulverization of the powder and OC fibers by a high-shear process, a pH of the dispersant such as 4-6, and, but are not limited to, a temperature range of 10-50°C, 45-50°C, or 20-30°C.

[0118] In the context of the present invention, the term “sealant” is also known as “fibrin sealant” and “biological glue,” and should be understood as an adhesive, glue, or hemostatic agent having components that, upon contact with or proximity to tissue and / or blood, react to subsequently form a mass, and further act as a tissue adhesive, thereby preventing, reducing, or stopping bleeding, joining structures, and / or sealing physiological leakage of, for example, cerebrospinal fluids (CSF), lymph, bile, gastrointestinal (GI) contents, air leakage from the lungs, etc., such as originating from or derived from the disclosed compositions (see, for example, Figure 8B and the following Examples chapter). In some embodiments, the sealant formulation also includes one or more therapeutic agents, which are released as a result when the mass formed in the body spontaneously decomposes. The therapeutic agent may be a drug such as an antibiotic, analgesic, anti-inflammatory drug, or anticancer drug, or a cell of any type derived from human or other sources, such as embryonic stem (ES) cells, adult stem cells, or induced pluripotent stem cells (iPSCs), but is not limited to these.

[0119] The term "hemostasis" refers to the ability to prevent, reduce, or stop blood loss from a wound, such as a surgical wound or trauma, by, for example, promoting the formation of a blood clot.

[0120] "Hemostasis (or haemostasis)" refers to the first stage of wound healing. This is the process of stopping bleeding. "To assist in hemostasis" means to help reduce or stop bleeding. "To apply to bleeding tissue" means to apply a composition topically to the bleeding site, such as a surgical site, in order to control bleeding. Controlling bleeding is necessary in a variety of situations, including wound treatment.

[0121] As used herein, the terms “control,” “prevent,” or “reduce” may be used interchangeably herein with respect to bleeding (including any grammatical variations thereof), meaning to reduce the rate of hemorrhage by 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or even 100% of the initial bleeding rate compared to a situation in which the disclosed composition is not in contact with the bleeding site or on the bleeding site. Methods for determining the level of bleeding occurrence are known in the art.

[0122] Furthermore, in some embodiments, with respect to bleeding, the terms “control,” “prevent,” or “reduce” also mean, for example, at least partially sealing the blood vessels at the site of bleeding in soft tissue.

[0123] As used herein, the term “subject” means any animal, but is not limited to humans, mice, rats, rabbits, non-human primates, or any other mammal. In some embodiments, the subject is a human (e.g., a human patient). The subject may be male or female.

[0124] As used herein, the term “injured tissue” (or “damaged tissue”) means the disruption of the normal continuity of a structure caused by physical (e.g., mechanical) force, such as surgical incision, biological means (e.g., thermal or chemical radiation), or chemical means. The term “injured tissue” also encompasses bruised tissue, and tissue that has been cut, punctured, or lacerated, as well as injuries caused by opening, puncture, and, for example, tearing, scratching, pressure, and biting. The term “injured tissue” also encompasses wounds and bleeding sites as described throughout this specification.

[0125] The term "tissue" may refer to tissue selected from soft tissue and bone tissue.

[0126] The term "soft tissue," as used in this specification, refers to body tissue that is not solidified or calcified. This term refers to soft tissue that is angiogenic and therefore a source of bleeding. Examples of such tissues include, but are not limited to, connective tissue (tendons, ligaments, fascia, skin, fibrous tissue, fat, and synovial membrane), muscle, and viscera. Generally, soft tissue excludes bone tissue.

[0127] Therefore, in some embodiments, the disclosed compositions may be used, in any embodiment, by being applied to hard-to-reach bleeding sites during minimally invasive surgery (MIS), such as endoscopic surgery. One common form of endoscopic surgery is laparoscopic surgery, which is minimally invasive examination and surgery performed inside the abdominal cavity.

[0128] The term “surgery” also encompasses the time during or after a surgical or diagnostic procedure. Further non-limiting examples of procedures include neurological surgery, abdominal surgery, cardiovascular surgery, thoracic surgery, head and neck surgery, pelvic surgery, and skin and subcutaneous tissue procedures. For at least one of these situations, the compositions of the present invention may function as suitable sealants that can adhere to bleeding wounds and thus address hemostasis regardless of the condition of the wound tissue, e.g., the severity of the bleeding. The formation of the sealant occurs within a relatively short coagulation time after applying the composition disclosed in any aspect and / or its embodiments to the injured tissue. Typically, use after 1-3 minutes of tamponade may result in the formation of an incomplete mass, which is possible to stop bleeding, although a longer time may be required for the coagulation reaction to reach the paste surface and form a complete mass.

[0129] Another aspect of the present invention provides a method for treating a wound, comprising the step of applying (e.g., making contact) a composition disclosed in any aspect and / or embodiment onto and / or into the wound of an object requiring treatment of the wound.

[0130] "To treat a wound" further means, for example, reducing blood loss at the site of tissue bleeding (in vivo) in a patient undergoing surgery. Refer to the Examples chapter, for example, Figure 8B, which shows the strong adhesion properties of the glycerol granular powder to porcine lung tissue in this ex vivo test of the test specimen disclosed herein.

[0131] Accordingly, in some embodiments, the method is for, for example, reducing blood loss at a site of tissue bleeding in a patient undergoing surgery, and / or forming a sealant layer in / on such tissue, and the method includes bringing the composition disclosed in that embodiment into contact with the site of bleeding and / or its vicinity. Optionally, the method includes first applying an aqueous solution, such as physiological saline, to such tissue to promote a rapid coagulation time, and then applying the disclosed composition onto the aqueous solution. Accordingly, in some such embodiments, the composition is intended to be used as a hemostatic agent at a site of tissue bleeding in any embodiment.

[0132] Throughout this specification, “substantially lacking” or “substantially not containing” means that a composition contains less than 8%, less than 5%, less than 3%, less than 1%, or even completely absent any relevant components (e.g., hydrophilic dispersants), such as, but not limited to, low-boiling solvents. For example, a composition may substantially lack polyols. In some embodiments, a composition lacks polyethylene glycol (PEG) (i.e., less than 1% by weight, or sometimes less than 0.5% by weight).

[0133] In some embodiments, fibrin gels prepared from the disclosed powders are characterized by high tensile strength. As used herein, the term “tensile strength” is defined as the ultimate tensile strength of a material (e.g., a gel), which is the maximum tensile force that the material can withstand without breaking. The term “high tensile strength” as used herein refers to a tensile strength of at least about 80 kPa, at least about 90 kPa, at least about 100 kPa, at least about 110 kPa, at least about 120 kPa, or at least about 130 kPa.

[0134] In some embodiments, the fibrin gel is characterized by a high tensile strength in the range of about 70 kPa to about 170 kPa, or in some embodiments, 80 kPa to about 160 kPa, or 85 kPa to about 160 kPa, when measured according to a method such as that described in International Application No. CN2023 / 082645.

[0135] Refer to Figures 2A-2B to demonstrate that the gel strength of fibrin gels prepared from aggregated powders prepared according to embodiments of this disclosure is greater than that of gels prepared from powders generated by hydrofluoroethers (HFEs).

[0136] In some embodiments, fibrin gels prepared from the disclosed powders are characterized by a tensile strain (%) in the range of about 120% to about 190% when measured according to a method such as that described in International Application No. CN2023 / 082645.

[0137] As used herein, the term "tensile strain" refers to the elongation of a material subjected to a tensile stress corresponding to the point of fracture.

[0138] According to some embodiments of the present invention, a process for preparing the powder described herein is provided in any one of the embodiments.

[0139] In exemplary embodiments, the general method used is: (a) Mixing solid fibrinogen, thrombin, and optionally OC or ORC fibers to form a mixture, (b) Inserting the mixture into a high-shear mixing reactor, (c) Adding a glycerol and water solution to the mixture in the reactor, (d) The composition obtained in step (c) is dried and sieved to form a powdered hemostatic composition, Includes.

[0140] In some embodiments, the parameters for mixing in step b (also called “pre-mixing”) are an impeller speed in the range of 100–300 rpm, a chopper speed of 100–800 rpm, and pre-mixing for 1–10 minutes.

[0141] In some embodiments, the parameters for granulation in a high-shear reactor in step (c) (after pre-mixing) are an impeller speed in the range of 200–600 rpm, a chopper speed in the range of 300–1200 rpm, and an atomization pressure in the range of 0.005–0.02. For example, a syringe pump is used to spray a glycerol-to-water aqueous solution having a glycerol-to-water ratio in the range of about 10% to less than 40% by weight (e.g., 10–25 mL) into a high-shear bowl at a flow rate of, for example, 2–6 mL / min to bind the particles. The granulation step lasts for, for example, 2–10 minutes, then the chopper speed is changed to, for example, 800–1800 rpm, and the post-granulation step is optionally continued for, for example, 0–120 seconds. In exemplary embodiments, approximately 100 g of biological powder (fibrinogen, thrombin, and optionally lysine) is combined with 10–25 mL of a dispersant, the dispersant containing glycerol / water in a specific ratio (v / v): 10% glycerol, 20% glycerol, or 40% glycerol. In exemplary embodiments, the components in the composite powder include approximately 80–96% fibrinogen powder, 300–1200 IU / dose of thrombin powder, 0–8.77% ORC powder, 2–3% by weight of CaCl2 powder, and 0–4% of lysine powder (as specified herein, percentages are by weight).

[0142] In some embodiments, such conditions allow for at least partial conversion of fibrinogen to fibrin. In some embodiments, these conditions allow for at least partial polymerization of fibrin, for example, by adding a specific amount of aqueous solution to a dispersant, as will be detailed throughout this specification.

[0143] In some embodiments, the composition substantially lacks OC or ORC.

[0144] In some embodiments, the mixture substantially lacks additional polymer binders other than fibrin.

[0145] In some embodiments, the mixture is substantially free of non-aqueous low-boiling solvents. In exemplary embodiments, the mixture is substantially free of hydrofluoroethers (HFEs).

[0146] As described above, in some embodiments, at least a portion of the powder is in the form of aggregates or granules. The term “granules” or “granular material” may, in particular, mean aggregates of particles typically less than 900 micrometers, less than 800 micrometers, less than 700 micrometers, or even less than 500 micrometers. That is, at least a portion of the particles are not in the form of discrete solids. In some embodiments, high-shear mixing is utilized to construct aggregates from particles of fibrinogen, thrombin, and optionally CaCl2, lysine, and ORC.

[0147] In some embodiments, powder compositions are provided that can be obtained by the methods disclosed herein in any embodiment. In some embodiments, the mixture in the method comprises solid fibrinogen, thrombin, and optionally ORC fibers in a solution of water and glycerol, wherein the glycerol is present in the solution (also called the “dispersant”) at a concentration ranging from more than about 10% by volume to less than about 40% by volume. In some embodiments, the glycerol is present in the solution at a concentration ranging from more than about 15% by volume to less than about 25% by volume. In some embodiments, the composition is substantially devoid of OC or ORC. In some embodiments, the powder composition is devoid of additional polymer binders other than fibrin. In some embodiments, the mixture is substantially devoid of non-aqueous low-boiling point solvents. In exemplary embodiments, the mixture is substantially devoid of hydrofluoroethers (HFEs). In some embodiments, the ratio of dispersant to powder in the granulation step is at least about 0.05 mL / g. In some embodiments, the ratio of dispersant to powder in the granulation step is in the range of about 0.05 to about 0.35 mL / g. In some embodiments, the ratio of dispersant to powder is in the range of about 0.1 to about 0.25 mL / g.

[0148] According to another embodiment of the present invention, a kit for producing the compositions described herein is provided. The consistency of the composition is understood to be such that it can be applied, for example, by spreading or adhering it directly onto injured tissue and / or bleeding sites. Therefore, the composition does not need to be further spread or applied onto a solid surface, object, or other solid medium such as a strip or film in order to be in a form suitable for application to injured tissue or bleeding sites. Nevertheless, a suitable applicator, such as a syringe, may be used for easy access and handling in order to apply, spread, or adhere the composition onto bleeding sites.

[0149] In further embodiments and / or other embodiments, the present invention provides a kit comprising: a) a container for containing the composition of the present invention described above; b) an applicator for applying the composition to tissue; and c) optionally instructions for use.

[0150] In another embodiment, the present invention further provides a hemostatic kit comprising a container for containing the composition disclosed herein.

[0151] Any aspect and embodiment of the compositions disclosed throughout this specification, including the composition, dispersant, and / or powder embodiment, may be incorporated into the kit aspect and embodiment.

[0152] Optionally, fibrinogen and thrombin are packaged separately in the kit (e.g., in dry form or solution). Therefore, each of the two components is packaged in separate packaging materials in addition to the overall kit packaging materials.

[0153] Alternatively, fibrinogen, thrombin, and optionally oxidized cellulose are packaged together in the kit (e.g., as dry powder) in the same packaging material, and optionally, a dispersant (e.g., physiological saline) is provided in an additional container. In some embodiments, the kit further includes instructions on how to combine the components of the kit and / or how to combine the components of the kit with additional components (e.g., dispersants) to produce a desired composition.

[0154] Optionally, the kit further includes a dispersant. The dispersant may be, for example, water, saline solution, or an aqueous buffer solution. Optionally, the dispersant is packaged separately from the powder. Alternatively, the dispersant is packaged together with the powder, for example, as a ready-to-use composition as described herein. In such embodiments, the kit may further include measuring means for measuring the volume of the dispersant or its components, for example, a measuring cylinder.

[0155] Additionally or alternatively, the hemostatic kit may include a syringe containing a blend, mixture, or powder, and another syringe containing a dispersant. For example, a dual-syringe mixer can combine liquids and powders that were initially separate, and then produce a substantially homogeneous paste mixture by moving the blended contents back and forth between two connected syringes through interconnected outlets. Therefore, a low pressure force for dispensing the paste from the syringe may be preferable to facilitate mixing and, ultimately, to spread the resulting paste. The desired pressure force may be less than 1.51 lbf.

[0156] In some embodiments, at least one of the containers in the kit is a pre-filled syringe. In some embodiments, a syringe is provided in addition to the containers in the kit. In some embodiments, the container is of a specific type, such as a vial or an applicator such as a syringe.

[0157] In some embodiments, at least one of the containers in the kit is a pre-filled syringe. In some embodiments, syringes are provided in addition to the containers in the kit. The term “container” can refer to any general structure, such as a container or vial, that can hold a paste.

[0158] The kit may be applied using an applicator device that can be used to administer the composition injectable solution several times in succession. In one embodiment, the applicator device allows for multiple injections of a fixed dose of the mixed component onto a 2D surface of tissue while moving the device. In one embodiment, the applicator has a syringe equipped with a needle, which optionally automatically retracts from the patient's skin after the injection is complete, without requiring the administerer to lift the device above the injection surface. In one embodiment, the kit may be used to administer a sealant.

[0159] The hemostatic kit of the present invention may be, for example, a kit for use in reducing, blocking, or stopping blood flow in an open wound, and it can be used to reduce, block, or stop blood flow during, before, or after surgical procedures such as laparoscopic surgery, neurosurgery, abdominal surgery, cardiovascular surgery, thoracic surgery, head and neck surgery, pelvic surgery, and skin and subcutaneous tissue surgery. The kit can be used to reduce or block blood flow from the skin or internal organs.

[0160] In some embodiments of any part of the kit and / or composition, the composition is sterile. Sterility, specifically viral inactivation, is a critical issue, especially when handling blood products. Generally, viral inactivation can be carried out by any number of methods, including solvent detergents, thermal inactivation, irradiation, and nanofiltration. Typically, standards for viral inactivation require the use of two different methods. Furthermore, FDA standards for sterility require filtration.

[0161] As used herein, the term “sterile” means having a low degree of biological contamination, effectively free from pathogens, for example, essentially or absolutely free from microorganisms, such as bacteria and viruses. Sterilization is the process of reducing the degree of biological contamination to a level that is effectively free from pathogens. A sterile liquid or solid is generally defined as a liquid or solid that has undergone sterile filtration.

[0162] The terms “comprises,” “comprising,” “includes,” “including,” and “having,” and their conjugations, mean “including but not limited to.” The term “consisting of” means “including and limited to.” The term “consisting essentially of” means that a composition, method, or structure may include additional components, steps, and / or parts, provided that the additional components, steps, and / or parts do not substantially alter the basic and novel features of the claimed composition, method, or structure.

[0163] The term “exemplary” in this specification means “serving as an example, case, or illustration.” Any embodiment described as “exemplary” is not necessarily construed to be preferable or advantageous to other embodiments, and / or does not preclude the incorporation of features from other embodiments.

[0164] In this specification, the term “optionally” means “provided in some embodiments but not in other embodiments.” Any particular embodiment of the present invention may include multiple “optional” features, provided that such features do not conflict.

[0165] As used herein, unless otherwise explicitly indicated in the context, the singular forms "a," "an," and "the" refer to multiple objects. For example, the terms "compound" or "at least one compound" may refer to multiple compounds, including mixtures thereof.

[0166] Throughout this application, various embodiments of the invention may be presented in range form. It should be understood that the range form description is for convenience and brevity only and should not be interpreted as a firm limitation on the scope of the invention. Accordingly, the range description should be considered to specifically disclose all possible subranges and the individual numbers within those ranges. For example, a range description such as 1 to 6 should be considered to have specifically disclosed subranges such as 1 to 3, 1 to 4, 1 to 5, 2 to 4, 2 to 6, 3 to 6, and the individual numbers within those ranges, e.g., 1, 2, 3, 4, 5, and 6. This applies regardless of the breadth of the range.

[0167] Whenever a numerical range is shown herein, it means that any cited number (fraction or integer) within the indicated range is included. The terms “ranging / ranges” between the first and second indicated numbers and “ranging / ranges from” the first indicated number to the second indicated number are used interchangeably herein, meaning that the first and second indicated numbers and all fractions and integers between them are included.

[0168] As used in the present invention, the term “method” means a method, means, technique, and procedure for accomplishing a given task, and includes, but is not limited to, methods, means, techniques, and procedures known or readily developed by practitioners in the fields of chemistry, pharmacology, biology, biochemistry, and medicine.

[0169] As used in the present invention, the term "treating" includes suppressing, substantially inhibiting, slowing, or reversing the progression of a disease, substantially improving the clinical or aesthetic symptoms of a condition, or substantially preventing the appearance of the clinical or aesthetic symptoms of a condition.

[0170] Where expressions similar to "at least one of A, B, and C" are used, such expressions are generally intended to be understood in a way that a person skilled in the art would understand (for example, "a composition having at least one of A, B, and C" is not limiting, but includes compositions having only A, only B, only C, both A and B, both A and C, both B and C and / or all of A, B and C). A person skilled in the art will further understand that virtually all separators and / or phrases presenting two or more alternative terms in the specification, claims, or drawings should be understood to take into account the possibility of including one of the terms, either of the terms, or both of the terms. For example, the phrase "A or B" would be understood to include the possibilities of "A" or "B" or "A and B".

[0171] Certain features of the present invention are described in the context of separate embodiments for clarity, but it is understood that these may also be presented in combination in a single embodiment. Conversely, various features of the present invention are described in the context of a single embodiment for brevity, but these may also be provided separately, in any preferred partial combination, or in any other described embodiment of the present invention. Certain features described in the context of various embodiments should not be considered essential features of those embodiments unless the embodiment would be unable to operate without those elements.

[0172] As used herein, the term “about” means that values ​​that are 10% higher or lower than the specified value are intended to be included. Unless otherwise specified, all numbers, such as those representing ratios, weights, moles / moles, quantities, viscosity, temperature, etc., should be understood in all cases as being modified by the term “about.” Therefore, unless otherwise specified, the numerical parameters described herein and in the appended claims are approximations that may vary by up to ±10% depending on the desired properties that the invention seeks to achieve.

[0173] Various embodiments and aspects of the present invention, as detailed above in this specification and claimed in the following claims, will be experimentally supported in the following examples. [Examples]

[0174] Herein, in conjunction with the above description, the following examples illustrate some embodiments of the present invention in a non-limiting manner.

[0175] Example 1: Preparation of a microaggregate composition using high-shear mixing (HSM) with glycerol Materials and methods: The sources of fibrinogen, thrombin, and albumin were porcine plasma supplied by Guangzhou Bioseal Biotech CO., LTD., located in Guangzhou, China, which was fractionated to obtain fibrinogen, thrombin, and albumin; fibrinogen and thrombin powders were produced by a spray-drying process. L-lysine-ALADDIN, L103479-500g; Oxidized regenerated cellulose (ORC) (if present) - ORC powder was obtained by a grinding process onto SURGICEL® original fabric; Calcium chloride (CaCl2), Taishan Xinning Pharmaceutical Co., Ltd. Glycerol, Guangzhou Chemical Reagent Factory; Purified water supplied by Guangzhou Bioseal Biotech Co., Ltd., located in Guangzhou, China; Oxidized regenerated cellulose (ORC) (if present) - Characteristics: particle size <70 μm, fibrous form. Briefly, ORC powder was obtained by processing SURGICEL® original fabric in the following steps: 1) the fabric was divided and cut into approximately 2-inch x 8-inch pieces; 2) the fabric was ground to a powder particle size (typically D50 less than 94 micrometers, but not limited to these) using a known grinding method; approximately 100 grams of fabric was placed in a 500 mL zirconia jar, then 12-13 pieces of 20 mm zirconia balls (marbles) were placed in the same jar, the jar was covered, and fixed in a Retsch planetary ball mill (model PM100), and the fabric was ground at 450 rpm for 20 minutes; the ground powder was transferred to an 8-inch diameter, 300 micrometer mesh sieve, and the marbles and powder were separated by slight shaking, and finally the powder was collected.

[0176] In an exemplary procedure, the hemostatic composition of the present invention was prepared as follows, particularly by using a spray-drying method.

[0177] Spray drying is a drying method used to produce powders from solutions, suspensions, or emulsions by spraying them into a high-temperature airflow through a spray nozzle and drying them immediately. The spray drying process is controlled by several process parameters, including column airflow and temperature, nozzle size and spray air velocity, and material flow rate.

[0178] In the example procedure, thrombin and fibrinogen powders were produced (one at a time) using spray drying (using a 4M8-TriX spray dryer (with ProCepT NV, Zelzate, Belgium)) as follows.

[0179] The prepared thrombin and fibrinogen butafibrinogen solution was drawn into a syringe and supplied in varying amounts to the syringe pump of the spray dryer. The feed valve was closed and the syringe pump was set to the desired flow rate. While the feed valve of the syringe pump was closed, the spray dryer was started and the desired spray gas flow rate, drying gas flow rate, drying gas temperature, cooling gas flow rate, and cyclone gas flow rate were set. The cooling gas flow rate and temperature were selected so as not to disturb the laminar flow of the drying column, but to lower the gas flow temperature below the glass transition temperature of the composition to prevent the powder from sticking to the glass portion.

[0180] The spray dryer was operated until the measured parameter values ​​reached the set levels and remained constant, achieving a steady state. The feed valve was then opened, and the thrombin or fibrinogen solution was flowed through the feed inlet to the spray nozzle, atomized into small droplets by the atomizing gas stream, and then dried in a drying column. The spray-dried powder was then collected at the powder outlet of the spray dryer's cyclone.

[0181] The spray-dried powder recovered from the cyclone was weighed into a dehumidifier with a relative humidity of less than 30% and divided into 100-200 mg samples. Each sample was individually sealed in a stoppered test tube and sealed with Parafilm® (Bemis, Oshkosh, Wisconsin, USA) until evaluation. Spray parameters: fan pressure 0.3 bar, spray pressure 3 bar, flow rate 130 mL / min, liquid pressure 16 kPa, nozzle diameter 0.7 mm, stirring speed 90 rpm / min.

[0182] In the exemplary procedure, the powder was further granulated using a high-shear process. Equipment details: Balance: SS1000, High-shear mixer: Mini-CG 1L, Syringe pump: LSP01-1C, Vacuum dryer: Vacucell 111, Vibrating screen: Fritsch analyzette 3. Environment: Temperature: 22-26°C, Relative humidity: 50-70%.

[0183] A method for forming a powdered hemostatic composition included the following steps: Pre-mixing: The preparation containing fibrinogen, thrombin, oxidized regenerated cellulose (ORC) fibers, CaCl2, and lysine was weighed to a total solid weight of approximately 100 g. The materials were then placed in a high-shear mixing reactor, with key parameters being an impeller speed of 100-300 rpm, a chopper speed of 100-800 rpm, and a pre-mixing time of 1-10 minutes. Granulation: After pre-mixing, the parameters of the high-shear mixing reactor were changed to an impeller speed of 200-600 rpm, a chopper speed of 300-1200 rpm, and an atomization pressure of 0.005-0.02. Using a syringe pump, 10-25 mL of a glycerol-water mixture was sprayed into a bowl at a flow rate of 2-6 mL / min to bind the particles. In the exemplary procedure, approximately 100 g of biological powder (fibrinogen, thrombin, and optionally lysine) was combined with 10-25 mL of dispersant, which contained glycerol / water in a specific ratio (v / v): 10% glycerol, 20% glycerol, or 40% glycerol. Granulation was continued for 2-10 minutes, then the chopper speed was changed to 800-1800 rpm for 0-120 seconds after granulation.

[0184] Next, the first vacuum drying method was applied: the obtained composition was transferred to a tray and placed in a vacuum drying box. The vacuum pressure was set to approximately 0 Pa over 0.5 to 1 hour. Next, the powder was sieved: the powder was transferred to a vibrating sieve tray with sieve sizes of 500 / 355 micrometers in appropriate order. The amplitude was set to 1.5 mm (sieving time: 10 minutes). Next, the second vacuum drying method was applied: the powder was transferred to a vacuum drying box and maintained at a vacuum pressure of approximately 0 Pa for 3 hours to allow it to dry. Finally, the product was collected.

[0185] Materials in the resulting powder: 80-96% by weight of fibrinogen powder, 300-1200 IU / dose of thrombin powder, 0-8.77% of ORC powder if present, 2-3% by weight of CaCl2 powder, 0-4% by weight of lysine powder, and glycerol ranging from 0.7% to 6% by weight (may depend on HSM conditions).

[0186] Design of Experiments (DOE) was used to optimize the formulation, and detailed information is provided in the following tables. For example, please refer to Tables 1A-1B for the components and percentages of the corresponding powders produced.

[0187] [Table 1] * In grams ** Fibrinogen + Thrombin + Lysine (w / w)

[0188] [Table 2]

[0189] Example 2: Characterization The following data was generated from the Department of Engineering (DOE) for formulation optimization.

[0190] form The dried powder (see Figure 1A) was received and uniformly blended while maintaining protein activity to obtain aggregates with a controlled particle size range, as shown in Figure 1B, which displays an exemplary scanning electron microscope (SEM) image of the aggregates. The glycerol weight range in the final product was 0.7–6.4 g / 100 g. Particle size distribution in the final product: D 50 ≦Approx. 172mm, D 90 ≤355 mm, see Table 2; the upper limit of ORC of the final product (lower limit is 0%) was 8.77%.

[0191] [Table 3]

[0192] Tensile stress / strain The obtained powder was compared with that prepared using a hydrofluoroether (HFE) suspension (see Table 3) using a similar HSM method (as described in U.S. Patent No. 11413335). The results of the comparative tests showed that the hemostatic powder prepared using glycerol possessed superior physiological and chemical properties, as well as functional properties such as tensile strength and tensile strain.

[0193] [Table 4]

[0194] Tensile Test Procedure In the exemplary procedure, layers of powder were sprayed onto an aqueous solution in a mold, as described in International Application No. CN2023 / 082645. Briefly, atomized saline solution was sprayed between the different layers of powder to promote dissolution and gelation. After applying a pre-weighed amount of powder and waiting for a predetermined gelation time, the gel sample was ready for testing. Next, the dimensions of the formed gel sample were measured, and the gel was carefully fitted into a fixture of an Instron 5944 equipped with a 100N load cell. The sample was stretched, and the maximum force and corresponding displacement data were electronically recorded, and tensile stress and tensile strain were calculated based on the sample dimensions and the data collected by the instrument.

[0195] The results for tensile stress and tensile strength are shown in Tables 4 and 5 below, respectively.

[0196] [Table 5]

[0197] [Table 6]

[0198] The results are further shown in Figures 2A and 2B.

[0199] It can be concluded that fibrin gels prepared from powders generated by glycerol suspension using the HSM method exhibit improved tensile properties compared to fibrin gels prepared from powders generated by HFE suspension instead of glycerol.

[0200] Effect of ORC on tensile stress / strain In an exemplary procedure, the effect of the presence of ORC on the tensile stress / strain properties of the gel was investigated.

[0201] The samples tested are shown in Table 6 below. Various amounts of glycerol were used (glycerol concentrations of 10%, 20%, and 40%).

[0202] [Table 7] * The ORC ratio refers to the ratio of the biological component to the ORC in the formulation used to produce the powder. "10:1" means that 100g of the biological component is equivalent to 10g of the ORC component. "20:1" means that 100g of the biological component is equivalent to 5g of the ORC component. "0" means that no ORC was added to the formulation. ** This relates to the glycerol concentration in the formulation from which the corresponding powder is manufactured.

[0203] The results are further shown in Figures 3A and 3B. The results are presented as average values ​​(for example, the 10:1 tensile stress data are the average values ​​of 128.76, 85.83, and 97.90 (104 kPa), and these data were obtained from 20%, 10%, and 40% glycerol solutions, respectively). It can be concluded that formulations without ORC provide a powder that forms a gel with the highest tensile strength, and that 20% glycerol in the formulation provides the highest tensile strength.

[0204] Solubility test In an exemplary procedure, the effect of the presence of ORC on the solubility of the powder was investigated.

[0205] The samples tested are shown in Table 6 above and Table 7 below. Various amounts of glycerol were used (glycerol concentrations of 10%, 20%, and 40% were used in the preparation process).

[0206] In the example procedure, 1 g of powder was spread evenly over a 9 cm x 9 cm area of ​​gauze, which was the top of the sponge. This set was then immersed halfway in a container of saline solution, allowing the sponge to absorb the saline solution and moisten the powder, and the time it took for the powder to completely dissolve was then measured.

[0207] Referring to Figure 4, a comparison image with the corresponding powder is shown.

[0208] The comparison results are further summarized in Table 7 and presented in Figures 5A and 5B.

[0209] [Table 8]

[0210] As described herein, the dried powder formulation of the present invention enables the agglomerated powder to rapidly dissolve in an aqueous medium, forming a gel layer with significantly increased gel strength and adhesive strength.

[0211] Furthermore, it can be concluded that better solubility is obtained when the amounts of ORC and glycerol are small, but the solubility of powders containing a specific amount of glycerol (e.g., 20%) is likely to be similar even if the ratio of ORC is different.

[0212] water content The water content (by weight) significantly affects powder stability. The water content of the powder was measured by Karl Fischer titration. Table 8 (see also Figure 6) shows that the water content of all samples from these batches was less than 2.5%.

[0213] [Table 9]

[0214] Example 3: Evaluation of ex vivo and in vivo models In the exemplary procedure, selected samples were tested to evaluate the hemostatic and occlusive efficacy of the powdered composition using various models under heparinization conditions.

[0215] Ex vivo trial protocol for porcine lung Fresh pig lung plucks were obtained from the slaughterhouse. The lungs were ventilated before application of the sample product.

[0216] In the exemplary procedure, the lungs were ventilated before application of the sample product. Ventilation was initiated in stages: First, the ventilator was set to pressure control mode and the respiratory rate to 10 bpm. Next, the expiratory positive airway pressure (EPAP) was fixed at 5 cm H2O. In the exemplary procedure, the inspiratory positive airway pressure (IPAP) was gradually increased from 5 to 45 cm H2O, generating pressure support (PS) at 5 cm H2O, 10 cm H2O, 20 cm H2O, 30 cm H2O, and 40 cm H2O. Then, the lung was observed to be fully inflated before initiating the application of the sealant.

[0217] Next, the lung was removed from the ventilator before applying the sealant. A 1 cm x 0.5 cm incision was then made on the lung surface to expose a linear defect. The tested sealant was then applied to the defect while the lung was fully deflated.

[0218] After the sealant hardened, the lung was reconnected to the ventilator and gradually inflated to achieve 5 cm H2O, 10 cm H2O, 20 cm H2O, 30 cm H2O, and 40 cm H2O.

[0219] Air leakage was observed and qualitative evaluation of adhesion to the lung surface was recorded at each pressure level.

[0220] The results are summarized in Table 9 below.

[0221] [Table 10]

[0222] We can conclude that all tested formulations demonstrated good sealing efficacy in the Exvivota lung model, and the 10:1, 40% glycerol granular sample showed no leakage or bulging whatsoever.

[0223] In another experimental setup, the ex vivo solubility was assessed using an ex vivo model, comparing the resulting powder (prepared using a biologic containing glycerol versus an ORC ratio of 10:1) with that prepared using an HFE suspension (10:1 sample).

[0224] To demonstrate the better solubility of powders granulated with glycerol solution, photographs were used herein. As shown in Figure 7, the HFE granular sample was characterized by lower solubility compared to the glycerol granular sample (see Figure 8B; N=2, no air leakage under 45 cm hydrostatic pressure for test samples containing 4.8 g of glycerol per 100 g of powder, with a bioagent to ORC ratio of 10:1). Furthermore, slight swelling was observed in the HFE granular sample (10:1 bioagent to ORC) under 45 cm hydrostatic pressure.

[0225] Figures 8A and 8B demonstrate the strong adhesion properties of glycerol granules to porcine lung tissue in this ex vivo test.

[0226] Conclusion: All powders prepared from glycerol formulations showed good sealing efficacy against the exvivota lung model.

[0227] Liver resection model In an additional exemplary procedure, a liver resection model was used to evaluate powder hemostasis as follows (representative procedures and results for glycerol granule powder are shown in Figures 9A-9C to demonstrate that hemostasis was achieved). 1. A portion of the liver was removed with a scalpel or scissors (3 cm from the edge), and the surface was then dried with gauze before applying the powder (see Figure 9A). 2. Next, the device was used to spray the sample over the entire excision surface, followed by spraying an appropriate amount of saline solution (see Figure 9B). 3. Step 2 may be repeated 3-4 times as needed. 4. Wait 3 minutes to allow the powder to form a gel (see Figure 9C). 5. Next, the immediate hemostatic effect was evaluated, and then the bleeding site was rinsed with 10 mL of physiological saline solution and observed for 1 minute to evaluate the continuous hemostatic effect (see Figure 9D showing the achieved hemostasis).

[0228] Canine lung model In the exemplary procedure, sealing efficacy tests were performed in a canine lung model.

[0229] Protocol for an in vivo trial in canine lung The procedure was as follows: 1. The lungs were exposed using appropriate surgical procedures. 2. A linear air leakage defect (approximately L 12 mm × D 5 mm) was created in the resected lung lobe. 3. The severity of air leakage from the defect was tested using bubble testing techniques under a pressure of approximately 20 cmH2O. 4. The articles were tested for their defects, and then cured to achieve the best results. 5. Sealing effectiveness was tested by inflating the lungs from a pressure of 20 cmH2O to 30 cmH2O to check for air bubbles. 6. The sealing effect was checked again 3.5 hours post-surgery (the surgical site was reopened). 7. Number of animals: 7.

[0230] The results showed a 100% (4 / 4) sealing success rate at 0 hours and 3.5 hours postoperatively for all test samples of glycerol granulated powder, as well as for ventilator pressures (cm H2O) tested at 20, 30, and 40.

[0231] While the present invention has been described in conjunction with its specific embodiments, it is evident that many alternative, modified, and variant forms will be obvious to those skilled in the art. Therefore, this is intended to encompass all such alternative, modified, and variant forms that fall within the spirit and broad scope of the appended claims.

[0232] [Implementation Method] (1) A powdered composition comprising fibrinogen, thrombin, glycerol aqueous solution, and optionally oxidized cellulose (OC), (i) The glycerol is present in a concentration ranging from about 0.7% by weight to about 6.5% by weight, (ii) A powdered composition in which the OC is present or absent at a concentration of less than about 9% by weight. (2) The composition according to Embodiment 1, wherein the OC comprises oxidized regenerated cellulose (ORC). (3) The composition according to Embodiment 1 or 2, which is substantially free of OC and / or ORC. (4) The composition according to any one of embodiments 1 to 3, further comprising fibrin. (5) A composition according to any one of Embodiments 1 to 4, which is substantially free of hydrofluoroethers.

[0233] (6) The composition according to any one of Embodiments 1 to 5, which is substantially free of a non-aqueous low-boiling point solvent. (7) A composition according to any one of Embodiments 1 to 6, having solubility in an aqueous medium at 25°C for less than 3 minutes. (8) The composition according to any one of Embodiments 1 to 7, wherein the powder comprises particles characterized by a D90 size distribution of up to 500 μm. (9) A composition according to any one of embodiments 1 to 8, which is ready for use for hemostasis and / or sealing. (10) The composition according to any one of Embodiments 1 to 9, characterized in that it forms a gel in an aqueous medium, wherein the gel is characterized by high gel strength and / or adhesive strength.

[0234] (11) The composition according to any one of embodiments 1 to 10, which substantially lacks additional polymer binders other than fibrin. (12) A composition according to any one of Embodiments 1 to 11 for use in a method of preventing air leakage in organs that require treatment of bleeding tissue and / or prevention of air leakage. (13) The composition according to any one of embodiments 1 to 12, further comprising lysine or tris. (14) The composition according to any one of embodiments 1 to 13, further comprising a calcium salt. (15) A composition according to any one of embodiments 1 to 14, deposited on an article (e.g., a patch).

[0235] (16) The composition according to any one of embodiments 1 to 15, wherein the water is present at a maximum concentration of 2.5% by weight. (17) A method for preparing a powdered hemostatic composition, (a) A step of mixing solid fibrinogen, thrombin, and optionally ORC fibers to form a mixture, (b) The step of inserting the mixture into a high-shear mixing reactor, (c) Adding a glycerol and water solution to the mixture in the reactor, (d) The step of drying and sieving the composition obtained in step (c), thereby forming the powdered hemostatic composition, Methods that include... (18) The method according to Embodiment 17, wherein the glycerol is present in the solution at a concentration in the range of more than about 10% by volume and less than about 40% by volume. (19) The method according to embodiment 17 or 18, wherein the water is added in an amount that prevents complete coagulation of the fibrinogen. (20) The method according to any one of embodiments 17 to 19, wherein the mixture does not contain OC and / or ORC.

[0236] (21) The method according to any one of embodiments 17 to 20, which does not involve the step of adding a non-aqueous low-boiling solvent. (22) A powdered composition that can be obtained by any of the methods described in Embodiments 17 to 21. (23) A composition comprising a mixture of solid fibrinogen, thrombin, and optionally oxidized cellulose (OC) fibers in a solution of water and glycerol, wherein the glycerol is present in the solution at a concentration in the range of more than about 10% by volume and less than about 40% by volume. (24) The composition according to embodiment 23, further comprising fibrin. (25) The composition according to Embodiment 23 or 24, which substantially lacks additional polymer binders other than fibrin.

[0237] (26) The composition according to any one of embodiments 23 to 25, wherein the OC comprises oxidized regenerated cellulose (ORC). (27) A composition according to any of embodiments 23 to 26, which is substantially free of OC and / or ORC. (28) A composition according to any one of embodiments 23 to 27, which is substantially free of hydrofluoroethers. (29) The composition according to any one of embodiments 23 to 28, which is substantially free of non-aqueous low-boiling solvents. (30) The composition according to any one of embodiments 23 to 29, wherein the solid fibrinogen, the thrombin, and optionally the oxidized cellulose (OC) fibers are in the form of a highly shear aggregated powder.

Claims

1. A powdered composition comprising fibrinogen, thrombin, glycerol-aqueous solution, and optionally oxidized cellulose (OC), (i) The glycerol is present in a concentration ranging from about 0.7% by weight to about 6.5% by weight, (ii) A powdered composition in which the OC is present at a concentration of less than about 9% by weight or is not present.

2. The composition according to claim 1, wherein the OC comprises oxidized regenerated cellulose (ORC).

3. The composition according to claim 1 or 2, which is substantially free of OC and / or ORC.

4. The composition according to claim 1, further comprising fibrin.

5. The composition according to claim 1, which is substantially free of hydrofluoroethers.

6. The composition according to claim 1, which is substantially free of non-aqueous low-boiling point solvents.

7. The composition according to claim 1, having solubility in an aqueous medium at 25°C for less than 3 minutes.

8. The composition according to claim 1, wherein the powder comprises particles characterized by a D90 size distribution of up to 500 μm.

9. The composition according to claim 1, which is ready for immediate use for hemostasis and / or sealing.

10. The composition according to claim 1, characterized by forming a gel in an aqueous medium, wherein the gel is characterized by high gel strength and / or adhesive strength.

11. The composition according to claim 1, substantially lacking additional polymer binders other than fibrin.

12. The composition according to claim 1, for use in a method of preventing air leakage in organs that require treatment of bleeding tissue and / or prevention of air leakage.

13. The composition according to claim 1, further comprising lysine or tris.

14. The composition according to claim 1, further comprising a calcium salt.

15. The composition according to claim 1, deposited on an article (e.g., a patch).

16. The composition according to claim 1, wherein the water is present at a maximum concentration of 2.5% by weight.

17. A method for preparing a powdered hemostatic composition, (a) A step of mixing solid fibrinogen, thrombin, and optionally ORC fibers to form a mixture, (b) The step of inserting the mixture into a high-shear mixing reactor, (c) Adding a glycerol and water solution to the mixture in the reactor, (d) A step of drying and sieving the composition obtained in step (c) to form the powdered hemostatic composition, Methods that include...

18. The method according to claim 17, wherein the glycerol is present in the solution at a concentration in the range of more than about 10% by volume and less than about 40% by volume.

19. The method according to claim 17 or 18, wherein the water is added in an amount that prevents complete coagulation of the fibrinogen.

20. The method according to claim 17, wherein the mixture does not contain OC and / or ORC.

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