Scaffold material for wound for suppressing scarring of biological tissue

Abstract

Disclosed is an artificial scaffold material for a wound applied to the wound in order to cure the wound while suppressing scarring in a living body. The scaffold material includes: a biomaterial having a property to which progenitor cells of myofibroblasts of the living body adhere; and an anti-fibrotic agent contained and retained in the biomaterial, the anti-fibrotic agent being gradually released from the biomaterial as the biomaterial dissolves in the living body. The wound includes at least one of an open wound and a closed wound in the living body. The anti-fibrotic agent has a low molecular weight, and inhibits the progenitor cells from differentiating into the myofibroblasts in the living body. The scaffold material for a wound is placed on a target surface of at least one of the wound surface of the open wound, the wound surface of the closed wound, and a portion of the surface of the living body where the closed wound is exposed, thereby allowing the anti-fibrotic agent to be locally administered to the target surface.

Description

Wound scaffold for inhibiting scarring of biological tissue

[0001] The present invention relates to a technique for inhibiting excessive scarring of biological tissue during the wound healing process and / or inhibiting adhesions between the abdominal wall and target organs after intraperitoneal laparotomy.

[0002] When a living body is locally injured, a wound is formed. Here, the term "wound" has various definitions, but it can refer to, for example, damage to the skin or soft tissue caused by an external force.

[0003] Wounds are classified into traumatic injuries caused by accidents and surgical wounds caused by surgery, including incision wounds (e.g., wounds made during surgery) and organ injuries (e.g., wounds to intraperitoneal organs or the peritoneum caused by abdominal surgery).

[0004] Furthermore, "wounds" are classified based on the degree of damage into defect wounds that involve tissue loss and non-defect wounds that do not involve tissue loss, and further into shallow skin wounds where the damage only reaches the dermis and deep wounds where the damage reaches the subcutaneous tissue.

[0005] The body is born with the ability to heal itself. "Natural healing" has various definitions, but it can mean, for example, that damaged tissues are repaired over time by regeneration (i.e., scarless healing) or scar healing.

[0006] In the mechanism of wound healing, i.e., the natural healing process of wounds in the body, soft granulation tissue containing fibroblasts and abundant blood vessels is first formed within the wound. Over time, the granulation tissue loses blood vessels and is replaced by hard scar tissue as fibroblasts produce collagen fibers. This phenomenon is called fibrosis, and in parallel with this, the epithelium is regenerated.

[0007] Initially, multiple collagen fibers exist sparsely within the granulation tissue, but over time, the collagen fibers become dense and homogenous. This natural healing process accompanied by fibrosis is called scar healing, and the tissue formed by this process is called scar tissue.

[0008] When scar tissue forms in the dermis of the skin of a living body and contracts or thickens, it can cause problems such as restricted skin movement, impeding the function of motor organs, pain, and mental distress associated with poor appearance, regardless of the severity.

[0009] Furthermore, when scar tissue forms within the body's organs and tissues (e.g., the eyes, lungs, heart, abdominal organs (liver, kidneys, etc.), and body cavities (peritoneal, thoracic, mediastinal, etc.)), it can cause symptoms of dysfunction or failure.

[0010] Therefore, although scarring in biological tissues is an unavoidable phenomenon that occurs during the wound repair process with the aim of natural wound healing, it is desirable to suppress excessive scarring to a level that does not inhibit the body's inherent natural healing function. For this reason, it is desirable to develop new technologies for suppressing scarring.

[0011] Japanese Patent Publication No. 2022-517510 discloses a technique for inhibiting scarring in the human eye caused by accidental eye damage, surgical incision on the eye, etc., i.e., an anti-scarring treatment method.

[0012] The publication further discloses an anti-scarring treatment method in which an extracellular matrix (hereinafter also referred to as "ECM" or "extracellular matrix") such as decorin is locally administered to the ocular surface of a human patient in the hope that it will act as an anti-fibrotic agent.

[0013] The publication also discloses several parameters that should be noted for assessing the occurrence of scarring, namely, several evaluation parameters, such as ECM components that increase with scarring, myofibroblasts or their marker protein α-smooth muscle actin (α-SMA) that increase with scarring, and TGF-β1, a growth factor that promotes the transformation of fibroblasts, that also increase with scarring.

[0014] The present inventors have conducted extensive research into treatment methods for inhibiting scarring in biological tissues, and as a result have discovered that it is important to administer an anti-fibrotic agent intra-wound or intraperitoneally rather than onto the surface of a wound, locally rather than systemically, and at an early stage of wound development in order to inhibit scarring.

[0015] Furthermore, the present inventors have also found that while inhibition of scarring in biological tissues has the advantage of inhibiting, preventing, or treating fibrosis or fibrotic diseases, it also has the disadvantage of delaying wound healing (e.g., in situations where the target wound is a defect wound or a deep wound), and therefore it is important to develop the treatment method in an individual-adaptive manner (e.g., adapted to the location, degree, and characteristics of the wound in each individual, as well as to the physiological characteristics such as the immune strength of each individual) so that the advantages more than compensate for the disadvantages.

[0016] Based on these findings, the present invention aims to provide a technique for suppressing excessive scarring of biological tissue during the wound healing process and / or suppressing adhesion between the abdominal wall and target organs after intraperitoneal laparotomy.

[0017] In order to solve this problem, according to one aspect of the present invention, there is provided an artificial wound scaffold that is applied to a wound in a living organism to heal the wound in a scar-inhibiting manner, the artificial wound scaffold comprising: a biomaterial that decomposes in the living organism and is absorbed by the living organism, and is configured to function as an artificial extracellular matrix to which precursor cells of myofibroblasts in the living organism have the property of adhering; and an anti-fibrotic agent contained in and retained by the biomaterial, the anti-fibrotic agent being gradually released from the biomaterial as the biomaterial dissolves in the living organism; the wound comprising at least one of an open wound and a closed wound in the living organism; the anti-fibrotic agent having a molecular weight of less than 10,000 and inhibiting the differentiation of the precursor cells into myofibroblasts in the living organism; and the wound scaffold is placed on at least one of the surface of the open wound and the surface of a local surrounding area of ​​the living organism including the closed wound, thereby enabling the anti-fibrotic agent to be administered locally to the target surface.

[0018] The present invention provides the following aspects. Each aspect is divided into paragraphs, each numbered, and described by citing the numbers of other paragraphs as necessary. This is to facilitate understanding of some of the technical features and combinations thereof that may be employed by the present invention, and should not be construed as limiting the technical features and combinations thereof that may be employed by the present invention to the following aspects. In other words, it should be understood that technical features that are not described in the following aspects but are described in this specification or drawings may be appropriately extracted and employed as technical features of the present invention.

[0019] Furthermore, describing each paragraph in a format that refers to the number of other paragraphs does not necessarily mean that the technical features described in each paragraph cannot be separated and made independent from the technical features described in other paragraphs, and it should be interpreted that the technical features described in each paragraph can be made independent as appropriate depending on their nature.

[0020] (Aspect 1) An artificial wound scaffold to be applied to a wound of a living organism, comprising a bioabsorbable biomaterial and an anti-fibrotic agent as main components, the biomaterial being configured to contain the anti-fibrotic agent at least during use of the wound scaffold, and the biomaterial being configured to function as an artificial extracellular matrix within the wound during use, the artificial extracellular matrix having a property to which myofibroblast precursor cells of the living organism adhere, The wound scaffold is placed within the wound during use, and / or placed within the abdominal cavity so as to make at least one of a first contact where the scaffold locally contacts an area of ​​the inner surface of the abdominal wall of the living body, the area having a first wound site on the abdominal wall as the wound, and a second contact where the scaffold locally contacts an area of ​​the outer surface of a target organ in the abdominal cavity, the area having a second wound site on the target organ as the wound, thereby allowing the anti-fibrotic agent to be locally administered within the wound, thereby enabling anti-scarring treatment to be performed during the healing process of the wound.

[0021] Here, the technical matter of "the wound scaffold is placed in the wound" and the technical matter of "the wound scaffold is placed in the abdominal cavity so as to make at least one of a first contact in which the wound scaffold locally comes into contact with a region of the inner surface of the abdominal wall of the living body, the region having a first wound site on the abdominal wall as the wound, and a second contact in which the wound scaffold locally comes into contact with a region of the outer surface of a target organ in the abdominal cavity, the region having a second wound site on the target organ as the wound" have in common that, in the end, the anti-fibrotic agent and the biomaterial, which are components of the wound scaffold, migrate from an initial placement position of the wound scaffold into the wound and are placed there. Therefore, these technical matters can be said to be the same technical feature, or if not, at least corresponding technical matters.

[0022] (Aspect 2) The wound scaffold according to aspect 1, wherein the biomaterial contains, as a main material, a protein, a polysaccharide, or a glycoprotein complex to which the progenitor cells have the property of adhering.

[0023] (Aspect 3) The wound scaffold according to Aspect 1 or 2, wherein the biomaterial is in the form of a hydrogel, powder, or sponge, or in the form of a film or sheet, and is flexible.

[0024] (Aspect 4) The wound scaffold of any of Aspects 1 to 3, wherein when the wound scaffold is placed in the living body, the wound scaffold defines a space in the living body that is isolated from tissue surrounding the wound, into which the progenitor cells can invade, and upon invading the space, the progenitor cells are allowed to react with the anti-fibrotic agent present in the wound scaffold, thereby inhibiting transformation of the progenitor cells into myofibroblasts, thereby enabling the anti-scarring treatment to be performed.

[0025] (Aspect 5) A wound scaffold according to any one of Aspects 1 to 4, wherein the scaffold has the functions of defining an administration route for the antifibrotic agent in the living body, providing a scaffold for the progenitor cells from outside the living body, and replenishing the wound from outside the living body with the artificial extracellular matrix to replace the original extracellular matrix that has been disrupted by the wound.

[0026] (Aspect 6) The wound scaffold according to any one of Aspects 1 to 5, wherein the scaffold is configured to retain the anti-fibrotic agent for at least a period from the time outside the body until the time when the scaffold is placed inside the body.

[0027] (Aspect 7) The wound scaffold according to any one of Aspects 1 to 6, wherein during the period in which the wound scaffold is present in the living body, the anti-fibrotic agent remains within the wound scaffold, while the progenitor cells from tissues surrounding the wound in the living body permeate into the wound scaffold, thereby causing the progenitor cells to react with the anti-fibrotic agent within the wound scaffold and inhibiting their transformation into myofibroblasts.

[0028] (Aspect 8) The wound scaffold according to any one of Aspects 1 to 7, wherein the progenitor cells are a cell population including at least fibroblasts or mesothelial cells from among fibroblasts, mesothelial cells, mesenchymal stem cells, bone marrow-derived stem cells, endothelial cells, vascular endothelial cells, smooth muscle cells, and epithelial cells, and differentiate into the myofibroblasts upon stimulation of the wound.

[0029] (Aspect 9) The wound scaffold according to any one of Aspects 1 to 8, wherein the wound is present in soft tissues of the body surface including the dermis and subcutaneous tissue, in a portion of subcutaneous or submuscular tissue surrounding an implant, or in the abdominal cavity of the living organism.

[0030] (Aspect 10) The wound scaffold according to any one of Aspects 1 to 9, wherein the wound scaffold is administered immediately after the wound is injured or in the early stage of wound healing.

[0031] (Aspect 11) The wound scaffold of any of Aspects 1 to 10, wherein the wound scaffold is administered before scar tissue forms in the wound and before or after the wound is sutured, whereby the wound scaffold is used to pre-treat the anti-scarring treatment.

[0032] (Aspect 12) The wound scaffold according to any one of Aspects 1 to 11, wherein the wound scaffold is administered to the surface of a new wound formed by the scar tissue excised from the living body by a surgical operation after the wound has naturally healed with the formation of scar tissue, and the new wound is then sutured, thereby allowing the wound scaffold to be used to subsequently perform the anti-scarring treatment.

[0033] (Aspect 13) The wound scaffold according to any one of Aspects 1 to 12, wherein the wound scaffold is a first multi-drug type prepared by mixing the biomaterial and the anti-fibrotic agent, at least either of which is in a liquid state, at the site where the anti-scarring treatment is to be performed.

[0034] (Aspect 14) The wound scaffold according to Aspect 13, wherein the biomaterial is a solid agent, and the anti-fibrotic agent is a liquid agent.

[0035] (Aspect 15) The wound scaffold according to any one of Aspects 1 to 14, wherein the wound scaffold is a second multi-drug type in which the biomaterial and the anti-fibrotic agent are both solid agents, and a liquid is added as a solvent, and the two are mixed at the site where the anti-scarring treatment is to be performed.

[0036] (Aspect 16) The wound scaffold according to any one of Aspects 1 to 15, comprising a scaffold in which the biomaterial and the anti-fibrotic agent are premixed to form a single liquid or solid agent.

[0037] (Aspect 17) The wound scaffold according to any one of Aspects 1 to 16, comprising a scaffold configured as an injectable agent to be injected into a gap between opposing wound surfaces in the wound.

[0038] (Aspect 18) The wound scaffold according to Aspect 17, wherein the injector is injected into the gap of the wound using an injector, the injector including: a storage section capable of storing the wound scaffold, the storage section being responsive to an external force to eject a required amount of the wound scaffold from the storage section; and a discharge section capable of ejecting the portion of the wound scaffold stored in the storage section that has ejected from the storage section.

[0039] (Aspect 19) A wound scaffold according to Aspect 18, wherein the storage section includes: a first portion capable of storing the biomaterial; a second portion capable of storing the anti-fibrotic agent; and a function of responding to a first external force that is the same as the external force or a second external force that is different from the first external force, to mix the biomaterial in the first portion with the anti-fibrotic agent in the second portion, thereby creating the wound scaffold.

[0040] (Aspect 20) The wound scaffold according to any one of Aspects 1 to 19, comprising a scaffold configured as a liquid, liniment, patch, or aerosol to be topically applied to the wound surface, the inner surface of the abdominal wall, or the outer surface of the target organ.

[0041] (Aspect 21) The wound scaffold according to Aspect 20, comprising the scaffold configured as a patch, the patch having the form of a flexible film or sheet, and configured to be inserted into a gap in the wound in a position extending along the wound surface, or inserted into the abdominal cavity so as to locally cover the inner surface of the abdominal wall and / or the outer surface of the target organ.

[0042] (Aspect 22) The wound scaffold of Aspect 20, wherein the wound scaffold is configured as a patch, the patch being in a sheet-like shape in a flexible state, the patch being for use during intraperitoneal surgery on the living body, the intraperitoneal surgery including: an incision step of making an incision in the abdominal wall to form a first wound site at the incision; and a surgical step of introducing a surgical instrument from outside the body through the incision into the abdominal cavity to perform a surgical operation on a target organ in the abdominal cavity to form a second wound site on an outer surface of the target organ, the patch being inserted into the abdominal cavity from outside the body through the incision during the surgical step, and the patch being retained in the abdominal cavity so as to make at least one of a first contact where the abdominal wall locally contacts a region of the inner surface of the abdominal wall including the first wound site and a second contact where the target organ locally contacts a region of the outer surface of the target organ including the second wound site.

[0043] (Aspect 23) The wound scaffold according to Aspect 22, wherein the anti-fibrotic agent contained in the patch migrates to the wound surfaces of the first wound site and the second wound site when the patch is placed in the abdominal cavity, thereby performing an anti-scarring treatment on the first and second wound sites.

[0044] (Aspect 24) The wound scaffold according to any one of Aspects 1 to 23, wherein administration of the wound scaffold is contraindicated when the wound is a defect wound accompanied by a tissue defect, reconstruction of the defect wound is not performed, and the wound surface of the wound is exposed.

[0045] (Aspect 25) The wound scaffold according to any one of Aspects 1 to 24, wherein the wound scaffold is prepared by simply mixing the biomaterial and the anti-fibrotic agent, each of which is a commercially available drug whose safety and efficacy have been established for other uses.

[0046] (Aspect 31) An artificial wound scaffold to be applied to a wound of a living organism, comprising a bioabsorbable biomaterial and an anti-fibrotic agent as main components, and configured so that the biomaterial contains the anti-fibrotic agent at least during use of the wound scaffold, and configured so that the biomaterial functions in the living organism during use as a substance that retains the anti-fibrotic agent and also functions as an artificial extracellular matrix to which myofibroblast precursor cells of the living organism adhere and which has the property of supporting the proliferation of these precursor cells, and the wound scaffold is placed in an incision formed in the skin as the wound during use, thereby allowing the anti-fibrotic agent and the biomaterial to be locally administered into the wound, thereby enabling anti-scarring treatment to be performed in a state that inhibits wound healing delay during the wound healing process, and the wound scaffold is further characterized in that: the wound does not include a wound formed by eye surgery in the living organism; and the wound does not include a defect wound. wherein the wound scaffold is placed in the wound so that the anti-fibrotic agent is administered locally to a location isolated from the epithelium of the skin, and the wound scaffold is placed in the wound during an early stage of a healing process of the wound.

[0047] (Aspect 32) An artificial wound scaffold to be applied to a wound of a living organism, the wound including a skin incision formed as a complete wound in the skin of the living organism, the wound scaffold having a bioabsorbable biomaterial and an anti-fibrotic agent as main components, and configured such that the biomaterial contains the anti-fibrotic agent at least during use of the wound scaffold, the wound scaffold being placed within the skin incision during use, thereby locally administering the anti-fibrotic agent and the biomaterial into the skin incision, thereby enabling anti-scarring treatment of the skin incision to be performed in a state that inhibits wound healing retardation during the healing process of the skin incision.

[0048] (Aspect 33) The wound scaffold according to Aspect 32, wherein the biomaterial is configured to function in the living body during the use as a substance that retains the antifibrotic agent and as an artificial extracellular matrix having properties to which myofibroblast precursor cells of the living body adhere and support the proliferation of the precursor cells.

[0049] (Aspect 34) The wound scaffold of Aspect 32 or 33, wherein the wound scaffold is placed within the skin incision such that the anti-fibrotic agent is administered locally to a location isolated from the epithelium of the skin.

[0050] (Aspect 35) The wound scaffold according to any one of Aspects 32 to 34, wherein the wound scaffold is placed in the skin incision during an early stage of the healing process of the skin incision.

[0051] (Aspect 41) An artificial wound scaffold to be applied to a wound of a living organism, comprising a bioabsorbable biomaterial and an anti-fibrotic agent as main components, and configured so that the biomaterial contains the anti-fibrotic agent at least during use of the wound scaffold, and the biomaterial is configured to function as an artificial extracellular matrix in the wound during use, having the property of allowing precursor cells of myofibroblasts of the living organism to adhere thereto, and the wound scaffold is placed in local contact with a wound formed on the inner surface of a closed cavity within the living organism during use, thereby locally administering the anti-fibrotic agent and the biomaterial to the wound, thereby enabling anti-scarring treatment to be carried out during the healing process of the wound in a state that inhibits wound healing retardation.

[0052] (Aspect 42) The wound scaffold according to Aspect 41, wherein the closed cavity includes an abdominal cavity of the living body.

[0053] (Aspect 43) The wound scaffold of Aspect 41, wherein the closed cavity comprises an artificially formed dissection cavity in the living body for breast reconstruction or breast enlargement, into which an implant is placed.

[0054] (Aspect 44) A wound scaffold according to any one of Aspects 41 to 43, which has the functions of defining an administration route for the antifibrotic agent in the living body, providing a scaffold for the progenitor cells from outside the living body, and replenishing the wound from outside the living body with the artificial extracellular matrix to replace the original extracellular matrix that has been disrupted by the wound.

[0055] (Aspect 45) The wound scaffold of any one of Aspects 41 to 44, comprising a scaffold configured as a liquid, liniment, patch, or aerosol to be topically applied to the inner surface of the closed cavity.

[0056] Aspect 46: The wound scaffold of Aspect 45, wherein the wound scaffold comprises a scaffold configured as a patch, the patch having the form of a flexible film or sheet, and configured to locally cover the inner surface of the closed cavity during use.

[0057] (Aspect 47) The wound scaffold according to any one of Aspects 41 to 46, wherein the wound scaffold is placed in the wound early in the healing process of the wound.

[0058] (Aspect 48) The wound scaffold of any one of Aspects 41 to 47, wherein the wound comprises an intact wound.

[0059] (Aspect 51) An artificial wound scaffold applied to a wound in a living organism to heal the wound in a scarring-inhibited manner, comprising: a biomaterial that is degraded in the living organism and absorbed by the living organism, and is configured to function as an artificial extracellular matrix to which precursor cells of myofibroblasts in the living organism have the property of adhering; and an anti-fibrotic agent contained and retained in the biomaterial, which is gradually released from the biomaterial as the biomaterial dissolves in the living organism; the wound comprises at least one of an open wound and a closed wound in the living organism; and the anti-fibrotic agent has a molecular weight of less than 10,000 and inhibits the differentiation of the precursor cells into the myofibroblasts in the living organism; The wound scaffold is placed on at least one of the target surfaces, which are the surface of the open wound, the surface of the closed wound, and a portion of the surface of tissue in the abdominal cavity of the living body where the closed wound is exposed, thereby enabling the anti-fibrotic agent to be administered locally to the target surface.

[0060] (Aspect 52) ​​The wound scaffold according to Aspect 51, wherein the biomaterial is configured to have shape retention.

[0061] (Aspect 53) The wound scaffold according to Aspect 52, wherein the biomaterial uses a hydrogel as a base material.

[0062] (Aspect 54) The wound scaffold according to Aspect 53, wherein the hydrogel comprises gelatin.

[0063] (Aspect 55) The wound scaffold according to Aspect 51, wherein the anti-fibrotic agent comprises fasudil, ripasudil, pirfenidone, and nintedanib.

[0064] (Aspect 56) The wound scaffold according to aspect 51, wherein the anti-fibrotic agent has a molecular weight of 400 or less.

[0065] (Aspect 57) The wound scaffold according to Aspect 56, wherein the anti-fibrotic agent comprises fasudil, ripasudil, and pirfenidone.

[0066] (Aspect 58) The wound scaffold according to any one of Aspects 51 to 58, wherein the open wound includes a skin-side surgical wound formed in the skin of the living body by surgery, and the wound scaffold is administered locally to the target surface, with the wound surface of the skin-side surgical wound being the target surface.

[0067] (Aspect 59) The wound scaffold according to Aspect 58, wherein the wound scaffold is placed within the skin-side surgical wound so that the anti-fibrotic agent is administered locally to a location isolated from the epithelium of the skin.

[0068] (Aspect 60) The wound scaffold according to any of Aspects 51 to 59, wherein the closed wound includes a peritoneal-side surgical wound formed by surgery in the peritoneum on the abdominal wall side within the abdominal cavity, and an organ-side surgical wound formed by surgery in a target organ within the abdominal cavity, and the wound scaffold is administered locally to both of the target surfaces, which are the surface of the peritoneal-side local peripheral region of the inner surface of the abdominal wall where the peritoneal-side surgical wound is exposed, and the surface of the organ-side local peripheral region of the outer surface of the target organ where the organ-side surgical wound is exposed.

[0069] (Aspect 61) The wound scaffold according to Aspect 60, wherein the wound scaffold is in the form of a sheet and is placed in the abdominal cavity between the peritoneum and the target organ, and is in surface contact with the peritoneum-side local peripheral region and the organ-side local peripheral region on both sides of the wound scaffold, respectively, thereby inhibiting scarring of each surgical wound and making it possible to inhibit adhesion between the abdominal wall and the target organ after intraperitoneal laparotomy.

[0070] (Aspect 71) An artificial wound scaffold that is applied to a wound in the skin of a living organism to heal the wound in a scarring-inhibited manner, comprising: a biomaterial that decomposes in the living organism and is absorbed by the living organism, and is configured to function as an artificial extracellular matrix to which precursor cells of myofibroblasts in the living organism have the property of adhering; and an anti-fibrotic agent contained and retained in the biomaterial, which is gradually released from the biomaterial as the biomaterial dissolves in the living organism, wherein the wound comprises a skin-side surgical wound formed in the skin of the living organism by surgery, and the wound scaffold is placed in a portion of the skin-side surgical wound that is isolated from the epithelium of the skin, thereby enabling the anti-fibrotic agent to be administered locally to a portion of the skin-side surgical wound that is isolated from the epithelium.

[0071] (Aspect 72) An artificial wound scaffold to be placed in the abdominal cavity after intraperitoneal laparotomy to suppress adhesions in the abdominal cavity, comprising: a biomaterial that is degraded in the living body and absorbed by the living body, and is configured to function as an artificial extracellular matrix to which precursor cells of myofibroblasts in the living body have the property of adhering; and an anti-fibrotic agent contained in and retained in the biomaterial, and that is gradually released from the biomaterial as the biomaterial dissolves in the living body; wherein the wounds include a peritoneal-side surgical wound formed in the peritoneum on the abdominal wall side in the abdominal cavity by surgery, and an organ-side surgical wound formed in a target organ in the abdominal cavity by surgery; The wound scaffold is in the form of a sheet and is placed in the abdominal cavity between the peritoneum and the target organ so that it is in surface contact with both target surfaces, namely, the surface of the peritoneal side local peripheral region including the peritoneal side surgical wound on the inner surface of the abdominal wall and the surface of the organ side local peripheral region including the organ side surgical wound on the outer surface of the target organ, respectively, thereby allowing the anti-fibrotic agent to be locally administered to the peritoneal side surgical wound and the organ side surgical wound, thereby inhibiting scarring of each surgical wound and suppressing the adhesions.

[0072] (Aspect 73) An artificial wound scaffold applied to a wound in a living organism to heal the wound in a scar-inhibiting manner, comprising: a biomaterial that decomposes in the living organism and is absorbed by the living organism, and is configured to function as an artificial extracellular matrix to which precursor cells of myofibroblasts in the living organism have the property of adhering; and an anti-fibrotic agent contained in and retained by the biomaterial, which is gradually released from the biomaterial as the biomaterial dissolves in the living organism; the wound includes a wound formed on the surface of a detachment cavity artificially formed by detaching one tissue from another tissue in the living organism for breast reconstruction or breast enlargement, into which an implant will be placed; the wound scaffold is applied to the surface of the implant so as to cover the surface of the implant prior to placement in the detachment cavity; and the implant is inserted and placed in the detachment cavity with the wound scaffold applied to the surface of the implant, thereby administering the anti-fibrotic agent to the surface of the detachment cavity.

[0073] Figure 1 is a cross-sectional view and a system diagram showing the basic wound healing process in a time series and conceptually. Figure 2 is a cross-sectional view and a system diagram showing the wound healing process using a wound scaffold according to some embodiments of the present invention in a time series and conceptually. Figure 3 is a perspective view conceptually showing a method of administering a wound scaffold according to some embodiments of the present invention into a wound by liquid injection. Figures 4(a) and 4(b) are both diagrams for explaining a method of inserting and leaving a wound scaffold according to some embodiments of the present invention as a sheet agent into a wound gap, as a first sheet agent insertion type. Specifically, Figure 4(a) is a perspective view conceptually showing a first example in which the sheet agent is inserted into a wound gap exposed at the body surface, while Figure 4(b) is a cross-sectional view conceptually showing a second example in which the sheet agent is inserted into a dissection cavity formed as a wound gap under the muscle layer (or subcutaneously) when placing an implant in the dissection cavity.

[0033] Figure 5(e) is a diagram for explaining a method of inserting and leaving the sheet agent in the abdominal cavity of a living body as a second sheet agent insertion type. Specifically, Figure 5(c) is a top perspective view for explaining the relative positional relationship between the exposed incision wound (an example of a wound) and the sheet agent left in the abdominal cavity during laparotomy. Figure 5(d) is a cross-sectional view conceptually showing a third example in which the sheet agent is left in the abdominal cavity of a living body during abdominal closure. Figure 5(e) is a perspective view conceptually showing a fourth example in which the sheet agent is left across two target organs in the abdominal cavity. Figure 5 is a diagram showing in table form several types of two-component mixed wound scaffolds according to some embodiments of the present invention. Figure 6 is a partial cross-sectional perspective view illustratively showing a timeline of an anti-scarring treatment process using a wound scaffold according to a first embodiment of the present invention. Figure 7 is a cross-sectional view illustrating an exemplary timeline of an anti-scarring treatment process using the wound scaffold according to the first embodiment. Figure 8 is a diagram illustrating, in a table format, two control groups and two treatment groups (hereinafter referred to as "four groups") in an experiment conducted to evaluate the effectiveness of the wound scaffold according to the first embodiment. Figure 9 is a plan view illustrating several types of procedures performed on mice as experimental animals in an experiment conducted to evaluate the effectiveness of the wound scaffold according to the first embodiment.Figure 10 shows multiple photographs taken on day 3 after skin incision for the four groups in the experiment. Figure 11 shows multiple photographs taken on day 7 after skin incision for the four groups in the experiment. Figure 12 shows a table showing the presence or absence of epithelialization at the wound site for each of the four groups in a chronological order in the form of a timeline. Figure 13 shows representative micrographs of scar tissue specimens taken on day 7 after skin incision for the control group 1 in the experiment. Figure 14 shows representative micrographs of scar tissue specimens taken on day 7 after skin incision for the control group 2 in the experiment. Figure 15 shows representative micrographs of scar tissue specimens taken on day 7 after skin incision for the treatment group 1 in the experiment. Figure 16 shows representative micrographs of scar tissue specimens taken on day 7 after skin incision for the treatment group 2 in the experiment. Figure 17 shows the boundary lines of the scar regions in representative micrographs of each of the four groups in the experiment. Figure 18 is a table showing the scar cross-sectional areas calculated for three mice in each of the four groups in the experiment and their average values. Figure 19 is a table showing the results of an ANOVA test for the results of the experiment. Figure 20 is a table showing multiple comparisons for the results of the experiment. Figure 21 is a table showing descriptive statistics for standard multiple statistics for the results of the experiment. Figure 22 is a graph showing the average and standard deviation of the scar cross-sectional areas for each of the four groups in the results of the experiment. Figure 23 is an exemplary partial cross-sectional side view showing an injector used for intrawound administration of a wound scaffold according to a second embodiment of the present invention.

[0074] Hereinafter, several exemplary embodiments of the present invention will be described in detail with reference to the accompanying drawings.

[0075] <Overview>

[0076] First, we will provide a technical overview common to these embodiments of the wound scaffold and the anti-scarring treatment method using the same, which is a treatment or therapy method (hereinafter also referred to as the "treatment method") that suppresses scarring of tissue during the wound healing process.

[0077] The purpose of the present inventors in proposing this treatment method is to achieve scarless wound healing (scarless wound healing, scarless healing, wound healing without leaving a scar, scar-free healing, healing with only a slight scar, scar formation inhibition therapy, etc.) by suppressing the differentiation (transformation) of fibroblasts, which are exemplary precursor cells, into myofibroblasts.

[0078] According to conventional wisdom, during the wound healing process, fibroblasts differentiate into myofibroblasts, primarily due to stimulation by TGF-β1 (a growth factor that promotes the transformation of fibroblasts), and these myofibroblasts then secrete excess ECM (such as collagen), resulting in scar formation. However, conventional anti-scarring treatments (such as taping and compression therapy) have limited effectiveness in inhibiting scar formation, making it inevitable that scars will remain after surgery or trauma.

[0079] On the other hand, several drugs that suppress differentiation into myofibroblasts have already been approved in Japan under names such as antifibrotic agents and ROCK inhibitors (Rho kinase inhibitors), and their efficacy and safety have been established for, for example, pulmonary fibrosis and glaucoma.

[0080] In response to this, the present inventors have considered that a safe and effective anti-scarring treatment method can be realized by combining anti-fibrotic agents, including but not limited to such approved drugs, with biomaterials and applying them to the field of medical treatment related to wound healing.

[0081] <General remarks>

[0082] 1. background

[0083] In humans, as an example of a living organism, skin injury after birth always leaves a scar at the site of injury, and the wound healing process often results in unsightly scars such as hypertrophic scars and keloids. Although various suturing techniques and aftercare methods have been developed to make scars less noticeable, the effectiveness of conventional anti-scarring treatments in inhibiting scar formation is limited, and new anti-scarring treatments are needed. In particular, unsightly scars exposed on the face or hands cause significant psychological distress and social stigma, significantly reducing patients' quality of life (QOL). "Scarless wound healing" is the wish of many patients who have undergone trauma or surgery, and its realization is eagerly awaited.

[0084] As shown conceptually in Figure 1, the basic wound healing process is characterized in that fibroblasts, an example of precursor cells, differentiate into α-SMA-positive myofibroblasts primarily through stimulation with TGF-β1 during wound healing, and these myofibroblasts then secrete excess ECM, resulting in scar formation.

[0085] Myofibroblasts contract granulation tissue, which can lead to scar contracture. Myofibroblasts are also abundant in the histological features of abnormal scars such as hypertrophic scars and keloids, and recent single-cell analysis suggests that a subpopulation of fibroblasts with myofibroblastic characteristics may contribute to keloid development.

[0086] Meanwhile, pirfenidone (which regulates the production of cytokines and growth factors) and nintedanib (a tyrosine kinase inhibitor) have already been approved in Japan as drugs that inhibit the differentiation of fibroblasts into myofibroblasts for the treatment of pulmonary fibrosis, and their efficacy and safety have been established.

[0087] ROCK inhibitors can also suppress differentiation into myofibroblasts through the control of the cytoskeleton, and as drugs classified as ROCK inhibitors, ripasudil for glaucoma and fasudil for cerebral vasospasm have been approved.

[0088] 2. Issues

[0089] The present inventors considered that these existing drugs that suppress differentiation into myofibroblasts, as well as other existing and generic drugs that can be substituted (hereinafter collectively referred to as "anti-fibrotic agents"), could be applied to scar treatment.

[0090] Furthermore, the present inventors have considered that by combining an antifibrotic agent with a biomaterial and devising a local administration form, side effects can be reduced compared to systemic administration of each agent, and have therefore set the following "objectives."

[0091] (1) What kind of biomaterial is optimal for delivering anti-fibrotic agents to wounds?

[0092] (2) Is scarless wound healing possible by using anti-fibrotic agents on wounds?

[0093] In order to solve these "problems," the present inventors conducted extensive research prior to completing the present invention.

[0094] 3. Purpose and effects of this treatment method

[0095] The goal of this treatment is to achieve scarless wound healing through a unique approach that uses anti-fibrotic agents to inhibit differentiation of fibroblasts into myofibroblasts.

[0096] To achieve this goal, the treatment method is characterized by the use of an artificial scaffold, as conceptually shown in Figure 2, which is designed to provide a dosage form in which an anti-fibrotic agent is administered locally to the wound to be healed, and which comprises a suitably constructed biomaterial containing a suitably selected anti-fibrotic agent, thereby inhibiting excessive scarring of tissue during the wound healing process.

[0097] This treatment method not only realizes scarless wound healing, but also has the potential to significantly contribute to the advancement of wound treatment as a fundamental treatment and preventative agent for keloids and hypertrophic scars.

[0098] Furthermore, when an existing drug whose safety has already been verified is used as an antifibrotic agent according to this treatment method, if the usefulness or effectiveness of the existing drug is further verified, it is expected that this treatment method will be clinically applicable soon.

[0099] Therefore, this treatment method is expected to produce an artificial scaffold with anti-fibrotic properties that will be useful in preventing adhesions after abdominal surgery and breast implant contracture, and may have broad application in the surgical field.

[0100] 4. How the idea for this treatment method came about

[0101] As a physician, the present inventor sees many patients in his daily practice in the field of plastic surgery who wish to receive treatment for scars that cause functional disorders, pain, and mental distress due to trauma or post-surgical scars (e.g., scars triggered by trauma or scars triggered by surgical incisions), and he feels that realizing scarless wound healing is an important issue in the field of plastic surgery.

[0102] Furthermore, the present inventors feel that, since the effectiveness of conventional symptomatic treatments for keloids and hypertrophic scars is limited, new treatments based on molecular mechanisms are needed.

[0103] Therefore, in a preliminary experiment, the inventors used immunohistochemistry (IHC) to observe the distribution of α-SMA-positive myofibroblasts in histological images of mature and immature scars. As a result, the inventors found that myofibroblasts are localized deep within immature scars and are completely absent in mature scars.

[0104] Based on this finding, the present inventors hypothesized that myofibroblasts appear in the initial stage of wound healing and secrete ECM, which then forms a scar, and then disappear as the scar matures due to apoptosis or other factors.

[0105] The present inventors then conceived the idea that if differentiation into myofibroblasts could be suppressed at the early stage of wound healing, excessive production of ECM would not occur and scarless wound healing would be possible.

[0106] Furthermore, the present inventors observed α-SMA-positive myofibroblasts in keloid tissue by immunostaining and found that myofibroblasts were distributed over a wide area of ​​the keloid lesion (e.g., an area covering more than half of the entire area).

[0107] Based on this finding, the inventors considered that keloids are a pathological condition in which differentiation into myofibroblasts occurs constantly due to continuous stimulation by mechanical stress and TGF-β1, and that inhibition of differentiation into myofibroblasts could also be applied to the treatment of keloids.

[0108] However, since side effects caused by systemic administration of anti-fibrotic agents have been reported, it is practically difficult to administer them systemically solely for the purpose of cosmetic improvement.

[0109] However, the present inventors considered that it may be possible to use the anti-fibrotic agent with the risk of side effects reduced as much as possible by combining it with a biomaterial that functions as an artificial ECM that contributes to the proliferation of the progenitor cells and locally administering it to the wound, as conceptually shown in Figure 2.

[0110] 5. Structure of scaffolding material

[0111] (1) Overall structure

[0112] The scaffold material has a biomaterial that functions as an artificial ECM for the progenitor cells and an anti-fibrotic agent as its main components, and the anti-fibrotic agent is contained in the biomaterial.

[0113] The scaffold may contain other components such as auxiliary components or additives, for example, stabilizers, preservatives, solvents, thickeners, etc. The scaffold may be a solid or a liquid having a high viscosity, or a liquid having a low viscosity such as water.

[0114] The scaffold is configured so that the anti-fibrotic agent and the biomaterial can act simultaneously, i.e., in parallel, without any substantial time lag, without taking measures to prevent time-delayed elution, such as encapsulating each component in microspheres to make the elution timing different from each other.

[0115] (2) Structural types

[0116] a. liquid injection mold

[0117] In the liquid injection type, as conceptually shown in Figure 3, the scaffold material is configured as a liquid (including a gel) that is injected into the gap between the wound surfaces (each of a pair of surfaces facing each other across a gap in the wound) using an injector.

[0118] Examples of the injector include a syringe in which a pressure piston is pushed into a cylinder to extrude a required amount of liquid from the cylinder, and a syringe in which a flexible container that contains the liquid is manually compressed to reduce the volume and extrude a required amount of liquid from the container.

[0119] b. Sheet insertion type

[0120] In the sheet-insertion type, as conceptually shown in Figure 4, the scaffold is configured as a flexible sheet (a solid agent including a bioabsorbable film, tape, cloth, etc.) inserted into the wound gap or other site, for example, into the abdominal cavity, and is impregnated, glued, coated, or attached with an antifibrotic agent. The sheet is configured by impregnating an antifibrotic agent into a flexible substrate (support base) made of a biomaterial. Methods for incorporating the antifibrotic agent into the biomaterial include, for example, impregnation, glueing, coating, attachment, or other methods that ultimately achieve incorporation.

[0121] In one example of a scaffold as a sheet, the biomaterial has shape-retaining properties and acts as a substrate for the scaffold, and in this example, the substrate also has shape-retaining properties, thereby allowing the scaffold as a whole to have shape-retaining properties even though the anti-fibrotic agent is a liquid and therefore does not have shape-retaining properties by itself.

[0122] (3) Liquid injection type

[0123] Liquid injection scaffolding materials can be one-component or two-component (an example of a multi-component) type.

[0124] In the one-component type, the scaffold is composed of a biomaterial and an anti-fibrotic agent mixed together in one component prior to arrival at the site of the anti-scarring procedure, and in this case the scaffold can be sold commercially as a finished product.

[0125] In contrast, the two-component mixed type can be classified into types 1-3, in which at least one of the biomaterial and the anti-fibrotic agent is a liquid, and type 4, in which both are powders, as shown in the table format of Figure 5. In this case, the scaffold can be sold commercially as a semi-finished product or kit product, with or without the injector.

[0126] Among the two-component mixed types, in types 1 to 3, the biomaterial and the anti-fibrotic agent are mixed together to form a single liquid at the site where the anti-scarring treatment is performed.

[0127] In contrast, in Type 4, at the site where the anti-scarring treatment is performed, a liquid (such as physiological saline or purified water) is added to at least one of the biomaterial and the anti-fibrotic agent as a solvent common to both the biomaterial and the anti-fibrotic agent, and the biomaterial and the anti-fibrotic agent are then mixed to form a single liquid.

[0128] (4) Sheet insertion type

[0129] An example of the sheet-insertion type is a first sheet-insertion type in which a sheet as a scaffold is inserted into the wound gap and left therein, as conceptually shown in Figures 4(a) and 4(b). In this type, the scaffold is inserted into the wound gap and left therein so that it does not substantially have any protrusions from the body surface.

[0130] Fig. 1(a) is a perspective view conceptually showing a first example in which the sheet agent is inserted into a wound gap exposed on the body surface, while Fig. 1(b) is a cross-sectional view conceptually showing a second example in which the sheet agent is inserted into a dissection cavity (described in detail later) formed as a wound gap under the muscle layer (or may be subcutaneously) when placing an implant in the dissection cavity.

[0131] Here, the second example will be described in detail.

[0132] In this example, as shown in the cross-sectional view of FIG. 1(b), the sheet agent is used when inserting and placing the implant subcutaneously or submuscularly in a human body.

[0133] One example of such an implant is an artificial breast implant, which is generally partially spherical and is inserted and placed within an artificial cavity in the human breast (also called a "dissection cavity," meaning a cavity formed between one tissue by dissecting another), thereby performing breast reconstruction or breast augmentation.

[0134] In its use for breast reconstruction or augmentation, the implant is a soft, capsule-like medical device filled with a silicone gel or the like.

[0135] As shown in the figure, an example of the dissection cavity is one formed under the human pectoral muscle (e.g., under the pectoralis major muscle, i.e., between the pectoral muscle and the chest wall), and in this example, the implant is inserted under the pectoralis major muscle.

[0136] Another example of a dissection cavity, not shown, is one formed under the mammary gland, and in this example, an implant is inserted under the mammary gland.

[0137] In either case, the tissue into which the implant is placed and into contact with the surface of the implant (hereinafter referred to as the "target tissue") is damaged due to detachment from other tissues, and a wound is formed across the detached surface of the target tissue. Since this wound can cause scarring, it is desirable to perform an anti-scarring treatment.

[0138] Therefore, in this example, the implant is inserted into the dissection cavity with its surface (ideally, the entire surface) covered with the above-mentioned sheet and left there. In this state, the antifibrotic agent contained in the sheet covering the implant migrates to the surface of the target tissue that forms the dissection cavity and is applied over substantially the entire surface (ideally, over the entire surface). As a result, scarring in the dissection cavity is suppressed.

[0139] In this example, coating the surface of the implant with a sheet material corresponds to an example of inserting the scaffold material into a gap in a wound (e.g., the gap between the surface of the pectoralis major muscle damaged by avulsion and the surface of the chest wall damaged by avulsion) in a position extending along the wound surface of the wound (e.g., the surface of the pectoralis major muscle and the surface of the chest wall), and also corresponds to an example of administering the scaffold material locally into the wound (e.g., rather than to the entire breast area).

[0140] The first sheet preparation insertion type has been described in detail above. In contrast to this, as another example, there is a second sheet preparation insertion type, which is an intraperitoneal placement in which a sheet preparation as a scaffold is inserted and placed in the abdominal cavity, as conceptually shown in Figures (c) to (e).

[0141] FIG. 1C is a top perspective view showing the relative positional relationship between the incision wound exposed at the time of abdominal incision and the sheet agent placed in the abdominal cavity.

[0142] Fig. 1(d) is a cross-sectional view conceptually showing a third example in which the sheet agent is placed in the abdominal cavity of a living body when the abdomen is closed, and Fig. 1(e) is a perspective view conceptually showing a fourth example in which the sheet agent is placed across two target organs in the abdominal cavity.

[0143] In the third example shown in Figure 1(d) and the fourth example shown in Figure 1(e), intraperitoneal laparotomy surgery is configured to include an incision process in which the patient's abdominal wall is incised, thereby forming a first wound site (e.g., a skin-side wound site) at the incision (incision wound), and a surgical process in which a surgical instrument is introduced from outside the body through the incision in the abdominal wall into the abdominal cavity to perform a surgical operation on a target organ, thereby forming a second wound site (e.g., an intraperitoneal organ-side wound site) on the outer surface of the target organ.

[0144] In the fourth example, the first wound site is a surgical wound formed on the inner surface of the peritoneum (e.g., the surface on the target organ side) so as to have a portion exposed toward the target organ side, and the second wound site is a surgical wound formed on the outer surface of the target organ so as to have a portion exposed toward the peritoneum side.

[0145] The sheet agent is inserted into the abdominal cavity from outside the body through an incision during the surgical procedure.

[0146] In a third example shown in Fig. 1(d), the sheet is placed in the abdominal cavity so as to make a first contact, that is, local contact (e.g., surface contact) with a region of the peritoneum that includes the first wound site. In this example, when the sheet is placed in the abdominal cavity and the surface of the sheet is in contact with the peritoneum, the anti-fibrotic agent contained in the sheet migrates from the sheet to the wound surface of the first wound site and performs an anti-scarring treatment on the first wound site.

[0147] In the fourth example shown in Figure 1 (e), the sheet is placed in the abdominal cavity so as to make both a first contact (e.g., surface contact) with the area of ​​the peritoneum that includes the first wound site, and a second contact (e.g., surface contact) with the outer surface of the target organ that includes the second wound site.

[0148] When the sheet is placed in the abdominal cavity and the surface of the sheet is in contact with the peritoneum, the anti-fibrotic agent contained in the sheet migrates from the sheet to the wound surface of the first wound site to perform anti-scarring treatment on the first wound site, while when the back surface of the sheet is in contact with the outer surface of the target organ, the anti-fibrotic agent migrates from the sheet to the wound surface of the second wound site to perform anti-scarring treatment on the second wound site.

[0149] More specifically, as shown in Fig. 1(e), the sheet is placed across two target organs in the abdominal cavity and on the two surfaces of the target organs, which is a perspective view showing the state in which the scaffold is placed across two target organs in the abdominal cavity and on the surfaces of the organs.

[0150] In this example, when the sheet is configured as a single sheet member, the sheet is configured to comprehensively perform the first contact for anti-scarring treatment on an incision (surgical wound) in the abdominal wall, i.e., a first wound site, the second contact for anti-scarring treatment on the outer surface of a first target organ within the abdominal cavity, and the second contact for anti-scarring treatment on the outer surface of a second target organ within the abdominal cavity.

[0151] In addition, FIG. 1(e) shows how two target organs are sutured together. Examples of the two organs that can be sutured together include the stomach and small intestine, and the liver and intestinal tract.

[0152] Furthermore, in some of the above examples, intraperitoneal local administration using the sheet is performed at least at the first wound site for anti-scarring treatment, but in addition to this, for example, for the first wound site, the aforementioned liquid injection type may be additionally adopted and the liquid as a scaffold material may be locally administered into the wound at the first wound site using an injector.

[0153] In summary, the sheet agent shown in Figure 1(e) constitutes an example of a concentrated anti-scarring treatment type patch in which both the first contact and the second contact are performed using a single sheet member.

[0154] In the third and fourth examples, the sheet agent is made of at least a biomaterial, which may suggest the possibility that the biomaterial may promote adhesion between the abdominal wall and the intraperitoneal organs.

[0155] However, what is noteworthy is that rather than having such a healing effect, the sheet material acts as a physical spacer that physically separates the abdominal wall from the intraperitoneal organs, and as a routing (pathway definition) that guides the anti-fibrotic agent to the target site (in the abdominal wall, the opening on the abdominal cavity side of the first wound site, and in the intraperitoneal organs, the opening on the abdominal cavity side of the second wound site).

[0156] Therefore, by placing the sheet in the abdominal cavity, particularly in a position between the abdominal wall and the intraperitoneal organs, not only is it possible to inhibit scarring of the abdominal wall and the intraperitoneal organs, but it is also possible to prevent adhesion between the abdominal wall and the intraperitoneal organs after surgery.

[0157] In this specification, the "target organ" may be defined as an intraperitoneal organ on which a surgical procedure is performed, or as an intraperitoneal organ on which a surgical procedure is not performed but which comes into contact with a scaffold.

[0158] Hereinafter, some other examples of the second sheet material insertion type will be described.

[0159] In one example, although not shown, the sheet agent has a first sheet member and a second sheet member that are independent of each other. The first sheet member is configured to locally contact, on its front surface, an area in the abdominal wall that includes a first wound site. The second sheet member is configured to locally contact, on its back surface, an area in the abdominal cavity that includes a second wound site.

[0160] In this example, when the sheet is placed in the abdominal cavity and the surface of the first sheet member contacts the inner surface of the abdominal wall, the anti-fibrotic agent contained in the sheet member migrates from the first sheet member to the wound surface of the first wound site and performs anti-scarring treatment on the first wound site, while when the back surface of the second sheet member contacts the outer surface of the target organ, the anti-fibrotic agent migrates from the second sheet member to the wound surface of the second wound site and performs anti-scarring treatment on the second wound site.

[0161] Just to be clear, the biomaterial contained in the same sheet material also migrates from the first sheet member to the wound surface of the first wound site and from the second sheet member to the wound surface of the second wound site, just like the anti-fibrotic agent, and therefore has material migration properties similar to those of the anti-fibrotic agent.

[0162] In summary, in this example, the sheet agent constitutes an example of a dispersed anti-scarring treatment type patch in which the first contact and the second contact are performed at discrete locations using separate sheet members (multiple sheet members that are independent of each other).

[0163] In the multiple examples described above for the first and second sheet agent insertion types, the sheet agents are all flexible but have shape-retaining properties (their shape does not change without external force), and therefore can be left in contact with curved anti-scarring treatment surfaces such as the inner surface (inner surface) of the abdominal wall and the outer surface (outer surface) of an organ.

[0164] In this way, an advantage of using a sheet for anti-scarring treatment is that its shape has the property of easily conforming to the surface of the place where it is to be placed, and so it can be placed in a state of surface contact with the curved anti-scarring treatment surface.

[0165] Generally, when localized damage, such as a surgical wound, occurs on the surface of an organ in the abdominal cavity, scarring begins in the organ from that location, and the scarring has the characteristic of spreading in a planar fashion on the surface of the organ.

[0166] Therefore, an advantage of using a sheet for anti-scarring treatment is that the sheet can be applied locally to the surface of the organ so as to cover not only the damaged area but also the surrounding area, making it easier to effectively suppress the spread of scarring on the surface of the organ.

[0167] (5) Materials that can be used as components of biomaterials

[0168] Biomaterials contain proteins (e.g., collagen, elastin, gelatin), polysaccharides (e.g., hyaluronic acid, chondroitin sulfate, cellulose), or glycoprotein complexes (e.g., aggrecan, versican) as their main components. They are in the form of hydrogels, powders, sponges, or sheets, and are flexible and shape-retaining.

[0169] There are no particular limitations on the materials that make up the biomaterials, as long as they are materials that can be dissolved, decomposed, metabolized, or absorbed by cells, microorganisms, or the natural life activities of living organisms.

[0170] Furthermore, examples of commercially available drugs (medical products, etc.) that constitute biomaterials include collagen sponges (e.g., Pelnac (registered trademark), Terdermis (registered trademark)), collagen sheets (e.g., Integra (registered trademark)), gelatin hydrogels (e.g., Genocel (registered trademark)), and gelatin sponges (e.g., Spongel (registered trademark)).

[0171] (6) Drugs that can be used as antifibrotic agents

[0172] The antifibrotic agent may be a low molecular weight agent such as fasudil, ripasudil, pirfenidone, or nintedanib. The antifibrotic agent may be in liquid, powder, or particulate form. The concentration of the antifibrotic agent may vary depending on the type of antifibrotic agent, the type of wound, the severity of the wound, and the characteristics of the individual (organism, human) to be administered.

[0173] 6. Basic guidelines for experimental methods

[0174] (1) Purpose

[0175] In order to confirm the efficacy of an anti-fibrotic agent, which is commercially available and whose efficacy and safety as a treatment method for living organisms, for example, human subjects has been established in relation to other uses (e.g., treatment of other diseases), when applied together with a biomaterial to the skin of a living organism for anti-scarring treatment in this treatment method (its safety is naturally confirmed without the need for further experiments, as it is commercially available), an animal experiment using the scaffold material of the invention is carried out as an in vivo test, thereby confirming the anti-scarring effect of the anti-fibrotic agent.

[0176] (2) Experiment overview

[0177] Treatment experiments in a mouse wound model

[0178] An incision is made on the back of a mouse, reaching from the epidermis to the fascia, and the incision is sutured. A mouse wound model is then prepared in such a state that an antifibrotic agent-containing scaffold is locally administered and placed in the intrawound gap (e.g., the gap between a pair of opposing wound surfaces separated by a gap in the incision, the dermal gap, etc.).

[0179] Therefore, it is expected that myofibroblasts will be formed in the region from the boundary layer between the epidermis and dermis to the fascia at the wound surface.

[0180] (3) Setting up a control group

[0181] Two control groups were set up: Control Group 1: The wound was simply sutured without administration of either the biomaterial or the antifibrotic agent, and Control Group 2: A biomaterial containing no antifibrotic agent, but containing purified water, was administered into the wound as a gel scaffold (without the antifibrotic agent). Three mice were used in each control group.

[0182] Here, the significance of each control group will be explained. Control group 2 has the significance of making it possible, by comparing it with control group 1, to confirm the advantages and disadvantages that can be obtained solely from the biomaterial in the scaffold when the scaffold of the invention is applied to a wound.

[0183] (4) Treatment group selection

[0184] Two treatment groups were established: Treatment group 1: A biomaterial containing a first antifibrotic agent (e.g., fasudil) at a predetermined concentration was administered into the wound as a gel scaffold (with the antifibrotic agent); and Treatment group 2: A biomaterial containing a second antifibrotic agent (e.g., ripasudil) at a predetermined concentration was administered into the wound as a gel scaffold (with the antifibrotic agent). Three mice were used in each treatment group.

[0185] It should be noted here that the scaffold of the invention contains an anti-fibrotic agent, whereas the scaffold used in the experiment does not contain an anti-fibrotic agent for the control group, but contains an anti-fibrotic agent only for the treatment group.

[0186] Furthermore, to explain the significance of each treatment group, treatment group 1 has the following significance: by comparing it with control group 1, it makes it possible to confirm the effects of a scaffold material as an invention having a biomaterial and a first antifibrotic agent; by comparing it with control group 2, it makes it possible to confirm the advantages and disadvantages obtained solely by the first antifibrotic agent in the scaffold material as an invention; and by comparing it with treatment group 2, it makes it possible to confirm the advantages and disadvantages obtained solely by the first antifibrotic agent in the scaffold material as an invention relative to the advantages and disadvantages obtained solely by the second antifibrotic agent in the scaffold material as an invention.

[0187] Furthermore, treatment group 2 is significant in that it makes it possible to confirm the effects of the inventive scaffold material having a biomaterial and a second antifibrotic agent by comparison with control group 1, it makes it possible to confirm the advantages and disadvantages obtained solely by the second antifibrotic agent in the inventive scaffold material by comparison with control group 2, and it makes it possible to confirm the advantages and disadvantages obtained solely by the second antifibrotic agent in the inventive scaffold material by comparison with treatment group 1 relative to the advantages and disadvantages obtained solely by the first antifibrotic agent in the inventive scaffold material.

[0188] (5) Incision and suture

[0189] Two skin incisions are made per mouse to ultimately form "skin suture wounds." Specifically, two incisions are made per mouse on the shaved back, each 10 mm long and full-thickness deep. The incisions are then sutured with two stitches (e.g., 6-0 nylon) in the epidermis only.

[0190] (6) Local administration of scaffold material

[0191] Immediately after suturing, a single intrawound administration was performed using the gel scaffold, for example, with the injector for each of the control group 2 and the treatment groups 1 and 2. This completes one treatment for each mouse. In each single intrawound administration, the scaffold was locally administered and placed at a location isolated from the epidermis, i.e., the epithelium, at the beginning of the healing process of each sutured skin wound (e.g., immediately after the mouse's skin was incised), as shown in the two right-hand figures in Figure 7.

[0192] (7) Appearance evaluation and scar tissue collection

[0193] After surgery, the appearance of the incision surface (exposed surface) was evaluated and scar tissue was harvested from each mouse. The caudal wound of each mouse was harvested on day 3 after surgery, and the cranial wound on day 7 after surgery.

[0194] (8) Specimen collection and scar tissue evaluation

[0195] For each group, one tissue specimen per scar tissue was taken, formalin-fixed, and paraffin-embedded. Each tissue specimen was evaluated for scar tissue.

[0196] The evaluation method involved measuring the cross-sectional area of ​​the scar for each tissue specimen, and performing immunohistochemical staining using myofibroblast markers such as α-SMA and SM22 to confirm the presence or absence of myofibroblasts.

[0197] <Specifics>

[0198] Definition of Terms

[0199] 1. "wound"

[0200] As used herein, the term "wound" refers to any tissue injury (including, for example, acute, subacute, delayed, or difficult-to-heal wounds, and chronic wounds). "Wound" may also include open wounds and closed wounds. Tissues in which wounds occur may include the skin and subcutaneous tissue, muscle tissue, peritoneum, digestive organs, digestive tract, etc.

[0201] The wound healing process is generally divided into four steps: hemostasis, inflammation, proliferation, and tissue remodeling. During the proliferation phase, myofibroblasts and capillaries infiltrate the wound site, promoting myofibroblast proliferation and collagen production. As a result, granulation tissue is formed at the wound site during the proliferation phase. The abundant blood vessels in the granulation tissue formed during the proliferation phase eventually regress during the subsequent tissue remodeling phase and are ultimately replaced by scar tissue primarily composed of collagen. It is known that excessive collagen deposition during this series of wound healing processes can lead to the formation of hypertrophic scars and keloids.

[0202] Here, we will classify wounds into defect wounds and non-defect wounds and explain the wound healing process in detail. Both defect and non-defect wounds sequentially experience the hemostasis, inflammation, proliferation, and tissue remodeling phases. However, defect wounds have tissue defects that must be filled during the inflammation and proliferation phases, whereas non-defect wounds do not have tissue defects that must be filled during the inflammation and proliferation phases.

[0203] Therefore, for defect wounds, administering a scaffold containing an antifibrotic agent into the wound in the early stage of the wound healing process, even if the purpose is to inhibit scarring, may delay wound healing.In contrast, for non-defect wounds, administering a scaffold containing an antifibrotic agent into the wound in the early stage of the wound healing process does not or, if there is any, is only a small risk of delaying wound healing.

[0204] Based on this finding, the present inventors have devised a technique in which, when the healing target is a non-defective wound, the scaffold is placed in the non-defective wound at an early stage of the healing process.

[0205] 2. "Scarring," "inhibition of scarring," etc.

[0206] As used herein, the term "scar" refers to fibrous connective tissue that forms at the site of injury to any bodily tissue. Scar tissue is typically composed of the same proteins (i.e., collagen) as the tissue it replaces. However, the fibrous composition of scar tissue differs significantly from the fibrous composition of non-scar tissue. Types of scars may include, but are not limited to, atrophic scars, skin graft scars, hypertrophic scars, keloids, and the like. Scar sites may also include, but are not limited to, scars of the skin and subcutaneous tissue, scars of the peritoneum, abdominal cavity, and intra-abdominal organs, scars of muscles, tendons, or joints, and the like.

[0207] As used herein, the term "hypertrophic scar" refers to a raised scar formed by excessive production of ECM in an attempt to repair a wound (hereinafter sometimes referred to as "wound site") after trauma. Hypertrophic scars that spread to normal skin are specifically called "keloids."

[0208] As used herein, "scarring (also referred to as "scar formation")" refers to the replacement of damaged areas of biological tissue with ECM (mainly collagen).

[0209] In the case of wounds on the human body surface, with a few exceptions such as fingertips, injuries that do not reach deeper than the outer layer of the skin (papillary dermis) result in little or no scarring (i.e., regeneration). However, injuries that reach the reticular dermis result in scarring and tissue remodeling, leading to healing (i.e., scar healing).

[0210] As used herein, the terms "inhibition of scarring," "inhibition of scar formation," "anti-scarring," and "inhibition of scar formation" refer to the inhibition of excessive proliferation of granulation tissue or excessive production of collagen during the proliferation and tissue remodeling phases of the wound healing process.

[0211] 3. "Treatment"

[0212] As used herein, the term "treatment" refers not only to treatment in the usual sense administered to a patient after the onset of the target disease, but also to curing, ameliorating, or at least partially ameliorating the disorder, and also to preventive treatment administered in advance to prevent the occurrence and recurrence of hypertrophic scars and / or keloids.

[0213] 4. "Extracellular matrix (ECM)"

[0214] As used herein, the term "extracellular matrix (ECM)" refers to a substance that plays a role in cellular organization in living organisms. The ECM provides structural scaffolding for cells and is a major component of physical support in the formation of tissues, organs, and tissues. The ECM is primarily composed of three major classes of biomolecules, including fibrous proteins such as collagen (e.g., types I and III) and elastin, glycoproteins such as fibrillin, fibronectin, and laminin, and glycoprotein complexes such as aggrecan and versican.

[0215] 5. Collagen

[0216] As used herein, the term "collagen" refers to an abundant protein found in the ECM.

[0217] 6. "Fibrosis"

[0218] As used herein, the term "fibrosis" refers to the deposition of ECM proteins (mainly collagen) in biological tissues.

[0219] 7. "Anti-fibrosis" etc.

[0220] As used herein, the terms "anti-fibrotic," "fibrosis suppression," and "fibrosis inhibition" refer to the suppression of the progression of fibrosis in tissue.

[0221] 8. "Anti-fibrotic agents" etc.

[0222] As used herein, the terms "antifibrotic agent," "fibrosis suppressing agent," or "fibrosis inhibitor" refer to a pharmaceutical agent that has a preventive or therapeutic effect against fibrosis in tissue. For example, an antifibrotic agent prevents or treats fibrosis by suppressing the differentiation of fibroblasts into myofibroblasts.

[0223] Antifibrotic agents are also classified according to their molecular weight, such as those with low molecular weights (molecular weights of about 300 to about 400, e.g., pirfenidone) and those with high molecular weights (molecular weights of about 36,000 to about 40,000, e.g., decorin).

[0224] 9. Biomaterials

[0225] In this specification, the term "biomaterial" refers to a product with a specific use and a specific function, as opposed to the broader term "biomaterial" which simply means a specific material.

[0226] Specifically, in this embodiment, the "biomaterial" is made using natural or synthetic biocompatible materials that maintain selected biologically active cells in a viable state and are suitable for introduction into living tissue.

[0227] Furthermore, in this embodiment, the "biomaterial" is prepared using a material among the biocompatible materials that functions as an artificial ECM that functions as a scaffold for the progenitor cells and is suitable for supporting the proliferation of the progenitor cells.

[0228] The biocompatible material can be of the indwelling type, which is left in the body after being introduced into biological tissue, or the biodegradable type, which is decomposed and absorbed in the body after being introduced into biological tissue. In this embodiment, the "biomaterial" is made using a biodegradable biocompatible material. Therefore, the "biomaterial" is of the in vivo degradable type, and is replaced by the body's own tissue over time, resulting in its disappearance in the body.

[0229] 10. "Scaffolding" and "Scaffolding Materials"

[0230] As used herein, the term "scaffold" is defined as a material that fulfills the functions of, for example, securing space for tissue regeneration and assisting the regeneration of damaged tissue while maintaining the shape of the regenerated tissue. The term may also be defined as a material that fulfills the functions of, for example, providing space for the regeneration of target cells while distributing them three-dimensionally and imparting a specific shape to them.

[0231] The actual object or substance administered into the body to realize this “scaffold” function is an artificial “scaffolding material.” Therefore, in this embodiment, the “scaffolding material” has, for example, good adhesiveness to target cells (e.g., the progenitor cells), bioaffinity or biocompatibility as a property that does not adversely affect the living body, low immunogenicity, is easily absorbed by the living body after being implanted in the body and is no longer needed, and has sufficient strength to withstand damage even when the body moves.

[0232] Next, the above-mentioned several embodiments will be described individually.

[0233] First Embodiment

[0234] 1. Structure of scaffolding material

[0235] (1) Selection of antifibrotic agents

[0236] Two options for antifibrotic agents were selected: ripasudil hydrochloride hydrate (Ripasudil) and fasudil hydrochloride hydrate (Fasudil), both of which are commercially available drugs. Both ripasudil and fasudil are classified as ROCK inhibitors. Both of these antifibrotic agents are liquid formulations.

[0237] (2) Selection of biomaterials

[0238] Gelatin hydrogel (Genocel®) was selected as the biomaterial scaffold material. Genocel® is a cell culture scaffold material using gelatin with a nonwoven structure, manufactured by Kyoto Medical Design Co., Ltd.

[0239] This gelatin hydrogel is available in sheet, block, and powder types. The biomaterial used in the scaffold according to this embodiment is a powder type, and is configured as a powder. Therefore, this biomaterial is a solid agent.

[0240] (3) Specific composition of scaffolding materials

[0241] a. Ingredients

[0242] The scaffold is constructed using a biomaterial and an anti-fibrotic agent as a pharmacologically active substance.

[0243] b. composition

[0244] The biomaterial acts as a cell culture scaffold, for example, using a gelatin-containing hydrogel with a nonwoven structure as a substrate.

[0245] The structure of the scaffold, including the nonwoven fabric structure, is porous, and the interstices within the structure are filled with an aqueous phase containing a pharmacologically active substance. The pharmacologically active substance is present uniformly within the aqueous phase and also on the surface of the aqueous phase.

[0246] The scaffold is constructed by incorporating an anti-fibrotic agent as a pharmacologically active substance into a biomaterial, i.e., the scaffold is constructed from an anti-fibrotic agent-containing hydrogel.

[0247] Hydrogel refers to a gel that does not dissolve in water and contains water. Hydrogel refers to a gel obtained by forming chemical or physical crosslinks between polymers such as gelatin using various chemical or physical crosslinking methods for polymer compounds such as gelatin. Hydrogels named Genocel (registered trademark) have only physical crosslinks.

[0248] c. Gel-sol transformation and molecular weight reduction of hydrogel

[0249] As hydrogels degrade in vivo, for example, due to the loss of gel crosslinks caused by the breakdown of the three-dimensional network structure, polymers such as gelatin transform from a solid (gel) to a fluid phase (sol) (solization), or due to molecular degradation, the polymers are decomposed into small molecules such as peptides (lowering of molecular weight).

[0250] d. Water solubility and release characteristics of each ingredient

[0251] In the initial state of the scaffold, the biomaterial does not dissolve and therefore releases very little of the antifibrotic agent. However, as the biomaterial dissolves in the body, the antifibrotic agent is released from the biomaterial. In the initial state of the scaffold, the antifibrotic agent is immobilized in the hydrogel and therefore is hardly released from the hydrogel.

[0252] Specifically, in hydrogels, a three-dimensional network structure is formed by intermolecular cross-linking, and thanks to this structure, the antifibrotic agent is immobilized between molecules, thereby allowing the antifibrotic agent to be held by the hydrogel.

[0253] However, the constituent molecules of the hydrogel transition from high molecular weight to low molecular weight, and in the high molecular weight state, the gel cross-links disappear in the hydrogel, the three-dimensional network structure is lost, and the hydrogel is not solid. Therefore, when the constituent molecules of the hydrogel are in a fluid state, the hydrogel cannot retain the anti-fibrotic agent.

[0254] 2. Manufacturing method of scaffold material

[0255] (1) Two-drug combination

[0256] The scaffold according to this embodiment is Type 1 in FIG. 5, and is manufactured by mixing a powdered biomaterial with a liquid anti-fibrotic agent.

[0257] (2) On-site manufacturing type

[0258] To manufacture the scaffold, the biomaterial and anti-fibrotic agent are mixed at the site where the anti-scarring procedure will be performed (eg, in the operating room).

[0259] The mixing process is carried out in a two-component injector as shown in FIG.

[0260] Specifically, the injector has a main body with two chambers and a nozzle located at the tip of the main body. The injector also has a switching function that normally keeps the two chambers isolated from each other but switches them to a connected state when an external force is applied. Before mixing, the biomaterial and the anti-fibrotic agent are contained separately and isolated in the two chambers.

[0261] When the external force is applied to mix the scaffold material at the site where the anti-scarring treatment is to be performed, the two chambers are brought into communication with each other, thereby initiating mixing. Then, when a first external force identical to the first external force or a second external force different from the first external force is applied to mix the scaffold material, the scaffold material moves from the container to the nozzle, and the required amount of scaffold material is discharged from the nozzle.

[0262] The injector has a mixing function for mixing the two components in addition to an injection function for injecting the mixed liquid, but instead, an injector having an injection function but no mixing function may be used. In this case, prior to using the injector, an operator may mix the two components on-site or at a different location, and then fill the injector with the mixed liquid thus prepared.

[0263] 3. Scaffold administration method

[0264] liquid injection mold

[0265] The scaffold is configured as a liquid injectable material that is injected into the gap between opposing wound surfaces in a wound.

[0266] The scaffold is administered topically, e.g., intradermally, to the wound at the site of the anti-scarring treatment. The scaffold is administered to the wound early in the wound healing process, e.g., immediately after suturing or within 5 minutes of completing suturing. The administration can be after completing suturing the entire wound or can be administered in parallel with the suturing process.

[0267] As shown in Figures 6 and 7, an incisional wound is sutured with sutures or staples, and in some cases is closed and fixed (wound closure) without sutures. Incisional wounds are sutured with dermal sutures and / or epidermal sutures. Dermal sutures are a suture method in which a needle is inserted into the second layer of the dermis without penetrating the first layer of the epidermis. Epidermal sutures are a general term for various suture methods that penetrate the first layer of the epidermis. Here, an "incisional wound" is referred to as a "skin suture wound" when it has been sutured, as mentioned above. An "incisional wound" is both a surgical wound and a non-defective wound.

[0268] After suturing, the scaffold is locally administered into the wound. As shown in Figures 6 and 7, the scaffold is injected into the wound using the injector with its nozzle inserted into the wound gap. Alternatively, the scaffold can be injected into the wound using, for example, a needle or needleless device, a syringe, a bottle, a dropper, a pipette, etc. As is clear from the first and second right figures in Figure 7, the scaffold is placed in a position isolated from the epidermis or epithelium shown in Figure 2.

[0269] The scaffold material is injected into at least one of a plurality of regions separated by a plurality of sutures along the axial direction of the wound after suturing, and the scaffold material can be injected into the regions one after the other in the axial direction, or every other region or every other region.

[0270] For ease of explanation, the scaffold material is injected into the wound gap, as shown in an exaggerated manner in Figure 7. Because the scaffold material is bioabsorbable, it will eventually be replaced by the body's own tissue and disappear.

[0271] 4. Mechanism of action of anti-scarring using scaffolds containing anti-fibrotic agents

[0272] As shown in FIG. 2, when the scaffold is administered into the wound, fibroblasts in the living tissue migrate into the scaffold.

[0273] The fibroblasts adhere to the scaffold and react with the anti-fibrotic agent in the scaffold, which inhibits the differentiation of fibroblasts into myofibroblasts and the proliferation of myofibroblasts. As a result, collagen production from myofibroblasts is suppressed, thereby inhibiting excessive scarring of biological tissue during the wound healing process. The biomaterial then degrades and disappears.

[0274] Specifically, when the scaffold is placed in the body, it defines a space in the body that is isolated from the tissue surrounding the wound and into which the progenitor cells can invade. Once the progenitor cells invade the space, they react with the anti-fibrotic agent present in the scaffold, thereby inhibiting the transformation of the progenitor cells into myofibroblasts and thereby providing an anti-scarring treatment.

[0275] Furthermore, the scaffold material has the functions of defining the administration route of antifibrotic agents in the body, of providing a scaffold for progenitor cells from outside the body, and of replenishing the wound from outside the body as the artificial ECM to replace the original ECM that has been disrupted by the wound.

[0276] 5. Experimental Method

[0277] Based on the above basic principles, experiments were conducted to confirm the effectiveness of the scaffold according to this embodiment in relation to anti-scarring treatment in the wound healing process in living skin.

[0278] The purpose of this experiment was to evaluate the effectiveness of the scaffold using an animal model. Specifically, we used an 8-week-old mouse incision wound model to evaluate its effect of inhibiting scar formation during the wound healing process.

[0279] 5-1. Animals used in the experiment

[0280] Twelve C57BL / 6JJcl mice (male, 8 weeks old, CLEA Japan, Inc.) were used as experimental animals. Mice were housed in clean S cages and mouse M2 cages, with three mice per cage, in a standard environment (temperature: 20-26°C, humidity: 40-70%, 12-hour light-dark cycle). Food and water were available ad libitum. After the treatment described below, mice were housed one per cage to avoid the risk of other mice damaging the wound.

[0281] 5-2. Grouping of multiple mice

[0282] Twelve mice were randomly assigned to four groups: control group 1 (n=3), control group 2 (n=3), treatment group 1 (n=3), and treatment group 2 (n=3), as tabulated in FIG. 8.

[0283] 5-3. Treatment of mice

[0284] Each mouse was anesthetized by inhalation of an anesthetic (e.g., isoflurane inhalation anesthetic "VTRS" (Viatris Pharmaceutical Co., Ltd.)). The back of each mouse was shaved. The skin at the incision site on the back of each mouse was disinfected with a disinfectant (e.g., povidone-iodine gel).

[0285] Furthermore, two separate longitudinal (body length) incisions were made in each mouse, each approximately 10 mm long and reaching the full skin depth. Each incision was then closed with two epidermal sutures (e.g., 6-0 nylon thread) for each mouse. The "incisions" referred to here are surgical wounds, and therefore are complete wounds.

[0286] Two wound sites are created in each mouse, one of which is harvested for scar tissue on postoperative day 3, and the other is harvested for scar tissue on postoperative day 7.

[0287] 5-4. Setting the control group

[0288] As shown in the table in Figure 8, two control groups were set: Control group 1: the wound was simply sutured without administering any biomaterial or antifibrotic agent; and Control group 2: a biomaterial containing no antifibrotic agent but containing purified water was administered into the wound as a gel scaffold.

[0289] Three mice were used in each control group. The three mice in control group 1, numbered 101, 102, and 103, and the three mice in control group 2, numbered 201, 202, and 203, are shown in the figure.

[0290] More specifically, the control groups 1 and 2 are as follows.

[0291] Administration group: Control group 1 Dose of antifibrotic agent: 0 Dose volume of scaffold: 0 Administration group: Control group 2 Dose of antifibrotic agent: 0 Dose volume of scaffold: 20 μL / wound site (20 μL was administered to each of two wound sites of each mouse.) (That is, a total of 20 μL of scaffold was administered to each wound site so that it was distributed approximately evenly among the three areas, i.e., administration positions, described below; this was the same for the other groups.) Preparation method: Using a 200 μL pipette tip, 200 μL of purified water was added to 800 μg of Genocel® powder, and the mixture was suspended by pipetting or inversion. Administration was performed immediately after preparation.

[0292] 5-5. Setting of treatment groups

[0293] As shown in the table in Figure 8, two treatment groups were established: Treatment group 1: A biomaterial containing fasudil was administered into the wound as a gel scaffold; and Treatment group 2: A biomaterial containing ripasudil was administered into the wound as a gel scaffold.

[0294] Three mice were used in each treatment group. The three mice in treatment group 1, numbered 301, 302, and 303, and the three mice in treatment group 2, numbered 401, 402, and 403, are shown in the figure.

[0295] More specifically for treatment groups 1 and 2:

[0296] Administration group: Treatment group 1 Antifibrotic agent administration concentration: 30 mg / mL Scaffold administration volume: 20 μL / wound site (20 μL was administered to each of two wound sites on each mouse.) Scaffold administration amount: Approximately 53.1 mg / kg (weight of scaffold administered to each mouse per kg of mouse) Preparation method: Using a 200 μL pipette tip, 200 μL of fasudil (30 mg of Eril® intravenous infusion solution) was added to 800 μg of Genocel® powder, and the mixture was suspended by pipetting or inversion. The mixture was administered immediately after preparation.

[0297] Administration group: Treatment group 2 Antifibrotic agent administration concentration: 4 mg / mL Scaffold administration volume: 20 μL / wound site (20 μL administered to each of two wound sites on each mouse) Scaffold administration amount: approximately 7.1 mg / kg (weight of scaffold administered to each mouse per kg of mouse) Preparation method: Using a 200 μL pipette tip, 200 μL of Ripasudil (Glanatec® eye drops 0.4%) was added to 800 μg of Genocel® powder and suspended by pipetting or inversion. Administration was performed immediately after preparation.

[0298] In treatment group 1, the concentration of fasudil was 30 mg / mL, but in the scaffold according to this embodiment, the concentration of fasudil can be, for example, within the range of about 28 mg / mL to about 32 mg / mL, within the range of about 25 mg / mL to about 35 mg / mL, or within the range of about 20 mg / mL to about 40 mg / mL.

[0299] Similarly, in treatment group 2, the concentration of Ripasudil was 4 mg / mL, but in the scaffold according to this embodiment, the concentration of Ripasudil can be, for example, within the range of about 3.5 mg / mL to about 4.5 mg / mL, or within the range of about 3 mg / mL to about 5 mg / mL, or within the range of about 2.5 mg / mL to about 5.5 mg / mL.

[0300] 5-6. Administration method

[0301] For control group 2, the scaffold (without antifibrotic agent) was administered to each mouse immediately after fabrication, and for treatment groups 1 and 2, the scaffold (with antifibrotic agent) was administered to each mouse immediately after fabrication.

[0302] Specifically, as shown in Figure 9, after the suture treatment, mice in control group 2, treatment group 1, and treatment group 2 were anesthetized and 20 μL of scaffold material was evenly administered into the wound using a 200 μL pipette tip into three areas created by suturing each incision (each wound site) in two places.

[0303] Therefore, a total volume of 20 μL of scaffold material was administered per wound site. The scaffold material was administered locally using the pipette tip to a location isolated from the epithelium within the internal space (gap) of each wound site, as described above with reference to Figure 7. After administration, each mouse was kept warm on a heat-retaining mat. After confirming that each mouse had recovered from anesthesia, the mouse was allowed to freely consume an analgesic (carprofen).

[0304] 6. Experimental results

[0305] 6-1. Observation of the wound site

[0306] The day of surgery (wound creation) was designated as Day 0, and the wound site of each mouse was photographed with a scale on Day 0, Day 3, and Day 7. Of the photographed images, those on Day 3 (Day 3) are shown in Figure 10 and those on Day 7 (Day 7) are shown in Figure 11.

[0307] In each figure, the label "Sham" refers to control group 1, the label "Genocel®" refers to control group 2, the label "Genocel® + Fasudil" refers to treatment group 1, and the label "Genocel® + Ripasudil" refers to treatment group 2.

[0308] Furthermore, in each figure, the label "HE" indicates that the photograph is of a specimen stained with hematoxylin and eosin (hereinafter referred to as "HE staining"), and the label "α-SMA" indicates that the photograph is of a specimen stained immunohistochemically using an anti-α-SMA antibody (or referred to as "α-SMA immunostaining").

[0309] 12 shows in tabular form for each group whether the wound site had epithelialized on days 3 and 7. Here, "epithelialization" means that the epithelium at the wound site had become completely continuous and the wound had healed.

[0310] 6-2. staining

[0311] On days 3 and 7, scar tissue was excised from each wound site from each mouse, and HE staining and α-SMA immunostaining were performed.

[0312] Specifically, the following tasks were carried out in sequence:

[0313] (1) On the third and seventh days, after observation of each wound site was completed, each mouse was anesthetized by inhaling the isoflurane inhalation anesthetic "VTRS" (manufactured by Viatris Pharmaceutical Co., Ltd.).

[0314] (2) After the sutures were removed from each mouse, a teardrop-shaped portion (e.g., a plate-like portion that is generally circular or oval in plan view) with a major axis of approximately 15 mm was excised from the skin of each mouse for each wound site, with each wound site positioned approximately in the center. The excised portion was then trimmed into a rectangular shape to prepare a scar specimen.

[0315] (3) After extracting scar specimens from each wound site from each mouse in this manner, the skin of each mouse was sutured at each wound site, and each mouse was given an analgesic (carprofen) in drinking water ad libitum.

[0316] (4) Each excised scar specimen was fixed in formalin and then embedded in paraffin.

[0317] (5) Multiple sections were cut from each paraffin-embedded scar specimen, and each section was prepared as a paraffin section.

[0318] (6) A first subset of the paraffin sections was stained with HE staining, and a second subset was immunohistochemically stained using an anti-α-SMA antibody (M0858, 1:300, Dako) (hereinafter also referred to as "α-SAM immunostaining").

[0319] For each paraffin-embedded scar specimen, one HE-stained specimen and one α-SMA immunostained specimen were prepared as histological staining samples.

[0320] 6-3. Evaluation of scarring

[0321] (1) Measurement of scar cross-sectional area

[0322] For each group, three HE-stained specimens were observed and photographed under an optical microscope on day 7. In each photomicrograph, the boundary between the scarred and non-scarred areas was drawn using NDP.view2 software (Hamamatsu Photonics Co., Ltd.), and the area of ​​the enclosed scarred area was quantified as the cross-sectional area of ​​the scar.

[0323] 13-16 show photomicrographs of one HE-stained specimen on day 7 for control groups 1 and 2 and treatment groups 1 and 2, respectively.

[0324] In the micrographs shown in each figure, the boundary line between the scarred area and the other areas was extracted as a single closed line, focusing on the difference in collagen fiber properties. Specifically, in the dermis layer, areas with a higher cell count, denser collagen fibers, and randomly arranged collagen fiber bundles compared to the surrounding areas were distinguished from other areas and determined to be areas where scarring had occurred.

[0325] For ease of illustration, lines have been added to the micrographs shown in each figure to highlight the boundaries of the scar regions extracted as described above.

[0326] In FIG. 17, the four boundary lines extracted as described above for the four groups are shown aligned in a horizontal row.

[0327] The calculated scar cross-sectional area for each group and for each mouse for the corresponding scar area is tabulated in Figure 18. The figure also tabulates the mean scar cross-sectional area for each group.

[0328] (2) Significance of experimental data

[0329] To evaluate the statistical reliability and significance of the multiple scar cross-sectional areas as experimental data, one-way analysis of variance (ANOVA) was performed using GraphPad Prism 9 (GraphPad Software, LLC), followed by a Tukey test. The experimental data were statistically analyzed based on the results of these tests. The results are shown in tables in Figures 19-22.

[0330] Specifically, FIG. 19 shows the ANOVA test results (labeled "ANOVA Results") in a tabular format.

[0331] FIG. 20 also shows the multiple comparisons (labeled "Multiple Comparisons (Tukey test results)") in a table format.

[0332] FIG. 21 also shows, in table form, descriptive statistics (labeled "Descriptive Statistics") for several standard statistical values ​​for the experimental data used in ANOVA tests and multiple comparisons.

[0333] Figure 22 also graphically depicts the mean and standard deviation of scar cross-sectional area for each group, and further demonstrates that there is a significant difference in scar cross-sectional area between control group 2 and treatment group 2 (P=0.03).

[0334] In these figures, the definitions of the main symbols are as follows:

[0335] A: Control group 1 B: Control group 2 C: Treatment group 1 D: Treatment group 2 ns: No significant difference *: Significant difference SS: Sum of squares, variation DF: Degrees of freedom, degrees of freedom MS: Mean square, mean square SD: Standard deviation F: F value Cl: Confidence interval P-value: P value

[0336] When P<0.05 was established for the difference between experimental data groups, the difference between the groups was considered to be statistically significant, and the contrasted groups were considered to have a significant difference.

[0337] (3) Evaluation of myofibroblast quantity

[0338] The α-SMA immunostained sections on days 3 and 7 were observed and photographed under a microscope, and the presence or absence of myofibroblasts was evaluated visually from the images of each micrograph. The micrographs of each section on day 3 are shown in Figure 10, and the micrographs of each section on day 7 are shown in Figure 11.

[0339] 6-4. Results and Discussion

[0340] (1) Visual inspection of the wound site

[0341] On day 3, the wound sites of the mice in none of the groups were epithelialized, whereas on day 7, the wound sites of the mice in all groups were epithelialized.

[0342] (2) Statistical analysis

[0343] a. Effectiveness of Ripasudil as an antifibrotic agent

[0344] As shown in Figures 20 and 22, treatment group 2 showed a decrease in scar cross-sectional area compared to control group 2, and there was a statistically significant difference between the two (p=0.03).

[0345] From this, it was considered that Ripasudil alone, which is one of the scaffold materials according to this embodiment, has an anti-scarring effect.

[0346] b. The potential of fasudil as an antifibrotic agent

[0347] As shown in Figures 20 and 22, treatment group 1 had a reduction in scar cross-sectional area relative to control group 2.

[0348] In this case, there was no statistically significant difference between the two, but it was considered that fasudil, one of the scaffold materials according to this embodiment, may have an anti-scarring effect by itself.

[0349] It is estimated that if the concentration and / or dosage of fasudil in treatment group 1 had been set higher than the values ​​in this experiment, the anti-scarring effect for this treatment group 1 would have been greater than that shown in Figure 22.

[0350] Furthermore, if the same concentration and dosage of fasudil as in this experiment were used in treatment group 1 and more individuals were treated, there is a possibility that the scar area would be significantly reduced compared to control group 2.

[0351] Therefore, it is concluded that not only ripasudil but also fasudil are anti-fibrotic agents that can exert anti-scarring effects in the presence of biomaterials.

[0352] c. The significance of biomaterials

[0353] As shown in Figures 20 and 22, Control Group 2 was larger than Control Group 1 in terms of scar cross-sectional area.

[0354] The reason for this is thought to be that in control group 2, a biomaterial that did not contain an antifibrotic agent was placed in the gap between the sutured wounds, which allowed the biomaterial to function as a space where myofibroblasts could adhere, resulting in active collagen production by the myofibroblasts within, resulting in a larger cross-sectional scar area than in control group 1.

[0355] On the other hand, the scar cross-sectional area was not larger in treatment groups 1 and 2 compared to control group 1, which indicates that the anti-fibrotic agent was functioning sufficiently to suppress collagen production.

[0356] (3) Visual inspection of micrographs of HE-stained specimens

[0357] Macroscopic observation of HE-stained micrographs confirmed scar tissue at the wound site in mice of all groups on day 7, as shown in FIGS. 13-16.

[0358] Furthermore, as shown in Figure 17, treatment groups 1 and 2 were smaller in terms of scar cross-sectional area relative to control groups 1 and 2.

[0359] From these findings, it was considered that ripasudil and fasudil, as antifibrotic agents, have anti-scarring effects.

[0360] (4) Visual inspection of micrographs of α-SMA immunostained specimens

[0361] As shown in FIG. 11, macroscopic observation of the α-SMA immunostained specimens confirmed that scar tissue containing myofibroblasts had formed in the wound.

[0362] Furthermore, as shown in the figure, the number of α-SMA-positive myofibroblasts was lower in treatment groups 1 and 2 compared to control groups 1 and 2.

[0363] The number of myofibroblasts in control group 2 was higher than that in control group 1. The reason for this has been explained above.

[0364] 6-5. Supplementary information

[0365] The results and discussion of the above-described experiment demonstrate that the scaffold according to the present embodiment has a scarring inhibitory effect. Although this experiment was conducted on mouse skin, common technical knowledge and experience suggest that similar experiments conducted on human skin would have a similar mechanism of action and effect.

[0366] 7. Effects

[0367] (1) Measures to prevent delayed wound healing as a side effect of scar inhibition

[0368] According to this embodiment, tissue scarring is suppressed during the natural healing process of deep wounds (e.g., surgical wounds) in which the wound surface reaches the subcutaneous tissue without causing tissue loss, making it easier to relieve patients from inconveniences such as functional impairment, pain, and mental distress caused by scarring.

[0369] In this embodiment, scarring of tissues is suppressed during the wound healing process, which may delay wound healing. However, the biomaterial in the scaffold has the function of supporting the migration and adhesion of progenitor cells such as fibroblasts in the body.

[0370] Therefore, according to this embodiment, unless early wound healing is particularly desired due to tissue loss, it is expected that epithelialization will not be prolonged due to delayed healing caused by the inhibition of scarring. This was verified by the experimental results showing that epithelialization was achieved in all treatment groups on Day 7 (see FIG. 12).

[0371] In other words, according to this embodiment, the scaffold material can be considered to have two bases with antagonistic effects: an anti-fibrotic agent that suppresses tissue scarring in the wound healing process and, as a secondary effect, delays wound healing, and a biomaterial that attenuates the wound healing delay effect by assisting the migration and adhesion of the progenitor cells.

[0372] Therefore, according to this embodiment, the delayed wound healing can be attenuated not only by adjusting the concentration (concentration and dilution) and / or the dose (increase and decrease) of the antifibrotic agent, but also by adjusting the concentration and / or the dose of the biomaterial instead, and in addition to the above adjustments, the delayed wound healing can also be attenuated by adjusting the concentration and / or the dose of the biomaterial.

[0373] As a result, according to this embodiment, the operator has multiple prescription options to choose from in order to attenuate the side effect of delaying wound healing, making it easier to select and optimize these multiple options in an individualized manner to suit the characteristics of the patient and the wound.

[0374] (2) Simplifying the procurement of scaffolding materials

[0375] According to this embodiment, several types of commercially available medicines whose safety and efficacy have been established for other uses can be converted into biomaterials and anti-fibrotic agents as the main components of the scaffold, respectively, and can be obtained by simply mixing the two.

[0376] Therefore, according to this embodiment, it becomes easy to procure a scaffold material for performing anti-scarring treatment during the wound healing process simply, at low cost, and safely.

[0377] (3) High permeability due to low molecular weight anti-fibrotic agents

[0378] The wound scaffold according to this embodiment uses an antifibrotic agent with a relatively low molecular weight, such as one with a molecular weight of 400 or less. Therefore, such low-molecular-weight antifibrotic agents are cheaper and easier to chemically synthesize than high-molecular-weight antifibrotic agents, such as those with a molecular weight of 10,000 or more, and in addition to having a stable structure, such low-molecular-weight antifibrotic agents act intracellularly, making it possible to directly and effectively inhibit the signaling pathway that differentiates the precursor cells into myofibroblasts.

[0379] Reducing the molecular weight of the antifibrotic agent used in the scaffold in this manner improves the cell permeability of the antifibrotic agent, thereby contributing to enhancing the effect of the antifibrotic agent in inhibiting the differentiation of the progenitor cells into myofibroblasts.

[0380] As mentioned above, the molecular weight of an antifibrotic agent having such an effect is preferably 400 or less, but as long as the molecular weight is less than 10,000, the differentiation inhibitory effect of the antifibrotic agent is improved compared to when the molecular weight is 10,000 or more. Furthermore, even an antifibrotic agent with a molecular weight of more than 400 exhibits good cell permeability as long as its molecular weight is 500 or less, and in this case, the differentiation inhibitory effect of the antifibrotic agent does not result in a significant difference in cell membrane permeability compared to antifibrotic agents with a molecular weight of 400 or less.

[0381] Furthermore, even compounds with a molecular weight of more than 500 can exhibit high cell permeability as long as their molecular weight is 1,000 or less due to specific molecular properties (e.g., optimal lipid solubility, low polar surface area, shielding of polar groups by intramolecular hydrogen bonds, or the use of specific carrier transport). For example, macrocyclic peptides such as cyclosporin A (molecular weight approximately 1,202) are known to exhibit high cell membrane permeability due to their ability to form intramolecular hydrogen bonds. Therefore, while it is desirable for the molecular weight of an antifibrotic agent to be 400 or less, by designing the molecule to have appropriate physicochemical properties, it is possible to ensure sufficient cell permeability to reach intracellular targets even in molecular weights of more than 500, or even in the range of molecular weights of 1,000 or less.

[0382] As described above, low-molecular-weight drugs with high cell permeability are generally distributed widely throughout the body when administered orally, increasing the risk of off-target effects and systemic side effects. However, by incorporating an anti-fibrotic agent into a wound scaffold and administering it locally to the body, as in the present embodiment, it is possible to reduce the risk of off-target effects and systemic side effects and improve the therapeutic effect.

[0383] (4) Long-term storage of drugs

[0384] According to this embodiment, the biomaterial and anti-fibrotic agent are stored as solid agents rather than liquid agents, for example, in a storage location or in an injector, prior to administering the scaffold into the wound using the injector shown in Fig. 3. Therefore, according to this embodiment, deterioration of the agent is suppressed compared to when the biomaterial is a liquid agent, and long-term storage of the agent is facilitated.

[0385] (5) Improved quality stability of scaffolding materials

[0386] According to this embodiment, at the site where anti-scarring treatment is being performed, an operator or assistant can use an injector to mix the biomaterial and the anti-fibrotic agent to prepare a scaffold at the very stage when the treatment is about to be performed, so even if there is a risk of some kind of denaturation occurring in the mixed solution after mixing, the period during which such risk occurs is short.

[0387] As a result, according to this embodiment, it becomes easier to use the scaffolding material with the desired performance, and the quality stability of the scaffolding material is improved.

[0388] (6) Possible application in the field of cosmetic medicine

[0389] The wound scaffold according to this embodiment can suppress or prevent the occurrence of scars and marks, and therefore can be applied not only in the medical field associated with disease treatment but also in the field of cosmetic surgery.

[0390] Second Embodiment

[0391] In this embodiment, an injector 10 shown in FIG. 23 is used as a specific example of the two-component mixed injector shown in FIG.

[0392] 1. Structure

[0393] As shown in the figure, this injector 10 has a front end and a rear end, and a main body 30 having a first chamber 20 on the front end side and a second chamber 22 on the rear end side. Before preparing the scaffold material (mixed liquid) 100, i.e., before mixing the biomaterial (powder) 102 and the anti-fibrotic agent (liquid) 104, the biomaterial 102 and the anti-fibrotic agent 104 are separately contained as drugs in the first and second chambers 20, 22, respectively. The biomaterial 102 and the anti-fibrotic agent 104 may be contained in either chamber 20, 22.

[0394] In the example shown in the figure, a powdered biomaterial 102 is contained in the presence of air in the first chamber 20. In the example shown in the figure, the second chamber 22 is formed in a glass ampoule, and a liquid or gel anti-fibrotic agent 104 is sealed in the ampoule.

[0395] The nozzle 40 extends from the tip of the tip side portion 60. The tip side portion 60 has a characteristic of being at least partially flexible. The nozzle 40 also has a characteristic of being at least partially flexible. The cross-sectional shape of the nozzle 40 may be circular or flat. If the nozzle 40 has a flattened cross-sectional shape, movement resistance may be reduced when moving the nozzle 40 in its longitudinal direction through a narrow wound gap, improving convenience. An outlet for discharging the scaffold material 100 (mixed liquid) is opened at the tip of the nozzle 40.

[0396] The main body 30 is divided into two parts: a tip side part 60 that forms the first chamber 20 and a rear side part 62 that forms the second chamber 22, and the two parts are connected at their opposing ends so that they can rotate relative to each other.

[0397] The injector 10 further includes a selective communication mechanism 72 that normally does not break a dividing membrane functioning portion 70 that functions as a dividing membrane (or partition) between the first and second chambers 20, 22, thereby keeping the first and second chambers 20, 22 separated from each other, but breaks the dividing membrane functioning portion 70 and brings the first and second chambers 20, 22 into communication with each other when a relative rotational force is applied to the front end portion 60 and the rear end portion 62, causing them to rotate relative to each other.

[0398] The selective communication mechanism 72 can be configured to include, for example, a screw mechanism that converts the relative rotational force into an axial force acting in the axial direction of the injector 10, and an engaging protrusion that is moved in the axial direction by the axial force, and to break the diaphragm function part 70, which has a portion extending in a direction intersecting the axial direction, by engaging with the engaging protrusion.

[0399] An example of the diaphragm function 70 is a portion of the wall of the glass ampoule. When the wall portion engages with the engaging protrusion (a rigid body such as metal), the portion breaks, and the broken portion then functions as a passage that allows the anti-fibrotic agent 104 to move from the second chamber 22 to the first chamber 20.

[0400] 2. action

[0401] With the first and second chambers 20, 22 in communication, for example, the anti-fibrotic agent in the second chamber 22 flows into the first chamber 20, and the anti-fibrotic agent that has flowed in and the powdered biomaterial that is already present in the first chamber 20 are mixed together to form a single liquid agent in the first chamber 20. In this state, the operator agitates the mixed liquid by, for example, shaking the injector 10, and uniformly mixes the two agents.

[0402] During the mixing and stirring process (e.g., in a standby state), a locking member 80 may be used that can selectively block at least locally the nozzle 40 so that the scaffolding material in the injector 10 does not leak out of the nozzle 40 against the operator's will. The locking member 80 is, for example, an elastic clip formed by a pair of arms elastically connected to each other at a hinge.

[0403] 3. Effects

[0404] Thanks to the property that the tip side portion 60 is at least partially flexible, when a scaffolding material is present as a mixed liquid in the first chamber 20, the operator can press the tip side portion 60 to reduce the volume of the first chamber 20, thereby ejecting the scaffolding material from the nozzle 40 in the required amount according to the pressing force.

[0405] Thanks to the at least partially flexible nature of the nozzle 40, when the operator injects the scaffolding material into the wound, for example, when the nozzle 40 is inserted into the wound gap, if the nozzle 40 tends to tilt relative to the wound surface, the operator can easily maintain a state in which the nozzle 40 follows the wound surface tightly without applying excessive force to the wound surface.

[0406] <Some other embodiments>

[0407] In some of the preceding embodiments, the biomaterial has a powder form, but may instead be in a hydrogel or sponge form. Alternatively, instead of a dispersion form, the biomaterial may be in the form of a continuous body, such as a film or sheet (e.g., flexible or non-flexible). Furthermore, the biomaterial may be in the form of a sheet, such as a mesh sheet, in which the material is processed to have a specific shape (e.g., an external shape, an overall shape, a three-dimensional shape, a shape with holes penetrating in the thickness direction, a porous shape, etc.).

[0408] In some of the preceding embodiments, the scaffold is configured to retain the antifibrotic agent at least until it is placed in the body from outside the body (e.g., until it is transferred from outside the body to inside the body), but it may be configured to retain the antifibrotic agent for a longer period, for example, while it is placed in the body (until the scaffold is absorbed and decomposed and disappears). Just to be clear, the scaffold is configured to function as, for example, a retainer, which is an object that retains the antifibrotic agent.

[0409] In the preceding embodiments, fibroblasts have been considered as the precursor cells, but mesothelial cells may alternatively or additionally be considered.

[0410] Furthermore, in addition to fibroblasts or mesothelial cells, cell populations including at least one of mesenchymal stem cells, bone marrow-derived stem cells, endothelial cells, vascular endothelial cells, smooth muscle cells, and epithelial cells may also be considered as precursor cells.

[0411] In some of the preceding embodiments, the scaffold is administered into the wound immediately after wound injury (e.g., when scarring begins in the tissue) or during the early stages of wound healing, but alternatively or additionally, the scaffold may be administered into the wound at a later time and stage.

[0412] In some of the preceding embodiments, the scaffold is administered to the wound surface before scar tissue forms in the wound and after (or optionally before) the wound is sutured, thereby using the scaffold to pre-treat the anti-scarring treatment.

[0413] In contrast, a scaffold material may be administered to the surface of a new wound formed by the excision after the wound has healed naturally with scar tissue formed in the wound being surgically excised from the living body and the new wound formed by the excision is sutured, thereby allowing the scaffold material to be used to perform the anti-scarring treatment after the fact.

[0414] In some of the preceding embodiments, the scaffold is configured as a single liquid formulation in which the biomaterial and the anti-fibrotic agent are premixed, but may alternatively be configured as a single solid formulation.

[0415] In some previous embodiments, the scaffold is configured as an injectable or patch that is injected into the wound space, but may alternatively be configured as a paint or aerosol.

[0416] <Additional explanation regarding the multiple scaffold placement method mentioned above>

[0417] Here, if we classify and organize the multiple scaffold placement methods disclosed in this application document without taking into consideration the material flow characteristics of the two types of components in the scaffold, namely the biomaterial and the anti-fibrotic agent, the scaffold placement methods can be broadly divided into an "intra-wound placement method" in which the scaffold is placed within a wound, and an "intraperitoneal placement method" in which the scaffold is placed within the abdominal cavity.

[0418] The "intra-wound placement method" is further classified into a "skin suture wound placement method" in which the scaffold is placed in a skin suture wound, and an "implant avulsion cavity placement method" in which the scaffold is placed in a wound formed on the inner surface of the avulsion cavity as a closed cavity surrounding the implant (for example, a wound that unfolds into a spherical shape).

[0419] To associate these three individual placement methods with Figures 3 and 4(a)-(e), the "skin suture wound placement method" is associated with Figures 3 and 4(a) , the "intraperitoneal cavity placement method" is associated with Figures 4(c)-(e) , and the "implant dissection cavity placement method" is associated with Figure 4(b) .

[0420] The experimental results shown in Figures 8 to 22 were obtained when experiments were conducted using mice using the "skin suture wound placement method."

[0421] In contrast to the "skin suture intrawound placement method," the "intraperitoneal placement method" and the "implant intra-ablation cavity placement method" have in common that they use scaffolds with the same attributes and that the scaffolds are placed in the wound early in the wound healing process. Therefore, if experiments were conducted using mice using the latter two placement methods, it is expected that experimental results equivalent to those described above would be obtained, or at least experimental results would be obtained that show that wound healing took place in a state where scar formation and healing delay were inhibited.

[0422] Furthermore, as mentioned above, the "skin suture wound placement method" is characterized by "isolated placement of the antifibrotic agent on the skin epithelium."

[0423] In contrast, with the "implant placement in the detached cavity method," there is no need to promote epithelialization during the wound healing process because there is no skin suture wound as part of the skin incision.

[0424] Furthermore, as an example of the "intraperitoneal placement method," there is a mode in which an abdominal incision is made and surgical procedures are performed through the resulting incision.

[0425] In this exemplary embodiment, the portion of the incision wound that appears on the inner surface of the abdominal cavity is the healing site (more precisely, the site at the initial placement position) where the scaffold is placed. Therefore, in this example, once the scaffold is placed at the healing site, the scaffold is automatically isolated from the skin epithelium.

[0426] Therefore, when implemented in this manner, the "intraperitoneal placement method" is characterized by "isolated placement of the antifibrotic agent on the skin epithelium," similar to the "skin sutured wound placement method."

[0427] <Supplementary explanation about the "skin suture wound placement method">

[0428] The following provides a detailed explanation of the problems that the "skin suture placement method" must solve and the technical concepts employed to solve these problems.

[0429] (1) Attributes of wounds as healing targets

[0430] In the "Skin Suture Intra-Wound Placement Method," the term "wound" is defined as not including "wounds formed by surgical operations on the eye of a living body" and not including "defect wounds."

[0431] Here, "skin suture wounds" does not include "wounds formed by surgical operations on the eye of a living body."

[0432] In the "skin sutured wound placement method," the scaffold material composed of an anti-fibrotic agent and a biomaterial acts within the space within the skin sutured wound, and since the space within the skin sutured wound is exposed to the body surface and communicates with the outside space, the space within the skin sutured wound does not qualify as a closed cavity.

[0433] In contrast to this, in the "intraperitoneal placement method" and the "implant intra-dissected cavity placement method," the scaffold acts in the abdominal cavity and the dissected cavity, respectively, as closed cavities.

[0434] However, these placement methods have in common that an intact wound is the target for healing, and the scaffold is placed in the intact wound at the early stage of the wound healing process.

[0435] (2) Summary of the "Skin suture wound placement method"

[0436] As a result of clinical research in which the inventor was involved as a surgeon, he discovered that the wound healing mechanism in the living body (particularly the mechanism of epithelialization in a wound, or the mechanism of epithelial regeneration in a wound) differs between a defective wound in which the skin epithelium is completely discontinuous and a non-defective wound in which the continuity of the skin epithelium is maintained, while developing a wound scaffold that combines an anti-fibrotic agent and a biomaterial to support wound healing so as to simultaneously inhibit scarring and inhibit delayed healing.

[0437] Specifically, the findings are as follows:

[0438] For defect wounds, early epithelialization of the epithelial defect, i.e., early filling of the epithelial defect, is important, so it is necessary to apply a biomaterial from the early stage of wound healing, while it is also necessary to isolate the antifibrotic agent from the skin defect. Therefore, when a wound scaffold containing both an antifibrotic agent and a biomaterial is introduced into the epithelial defect from the early stage of wound healing, it is necessary to devise a way to temporarily isolate the antifibrotic agent from the skin defect (to temporarily block the drug effect) (for example, a technique for adjusting the drug elution rate for each drug).

[0439] In contrast, for non-defective wounds that do not have tissue defects, wound healing is possible even if scarring is suppressed throughout the entire wound healing process and without the support of biomaterials.

[0440] Therefore, the inventor realized that for non-defective wounds, if a scaffold material is placed in the wound from the early stage of wound healing in a state in which the anti-fibrotic agent, acting as a brake on wound healing, and the biomaterial, acting as an accelerator on wound healing, can act simultaneously, it is possible to achieve both inhibition of scarring and inhibition of delayed healing.

[0441] Furthermore, the present inventors have realized that if the antifibrotic agent is placed in the wound so as to be isolated from the skin epithelium, it is possible to more effectively inhibit the delay in healing caused by the antifibrotic agent.

[0442] Based on these findings, the present inventors have proposed a wound scaffold that selects an intact wound as the target for healing and that contains both an antifibrotic agent and a biomaterial in a state that allows them to act without a time lag, and that the wound scaffold be placed in the wound at a position isolated from the skin epithelium of the intact wound so that the antifibrotic agent does not delay epithelialization. The details of this proposal are disclosed in this application.

[0443] (3) The first issue and technical concept of the "skin suture wound placement method"

[0444] The disclosure of this application documents states that the first problem is to "inhibit scarring at wound sites in living organisms where natural healing should occur, while inhibiting the delay of natural healing at the wound site," and to solve this first problem, the first technical concept is adopted, which is to "simultaneously activate the phenomenon of 'inhibition of scarring by anti-fibrotic agents → delay of wound healing,' which can be likened to stepping on the brakes in the wound healing process, and the phenomenon of 'support of proliferation of myofibroblast precursor cells by biomaterials → promotion of wound healing,' which can be likened to stepping on the accelerator."

[0445] This first technical idea is realized by configuring the wound scaffold as a combination (compound) containing an anti-fibrotic agent as a first base or active ingredient and a biomaterial as a second base or active ingredient, thereby enabling the anti-fibrotic agent and the biomaterial to act simultaneously and in parallel.

[0446] (4) Second issue and technical concept of the "skin suture wound placement method"

[0447] The disclosure of this application further states that a second objective is to "suppress the delay of natural healing by suppressing the inhibition of skin epithelialization by antifibrotic agents, which are used as a brake to suppress the differentiation of myofibroblast precursor cells during the wound healing process," and to achieve this second objective, a second technical concept is adopted in which the wound scaffold is placed within the wound so that the antifibrotic agent is administered locally at a position isolated from the skin epithelium.

[0448] According to this second technical concept, the anti-fibrotic agent is locally administered in vivo to a location of the wound that is isolated from the skin epithelium, thereby preventing the skin epithelium from being exposed to the anti-fibrotic agent.

[0449] As a result, according to the disclosure of this application, although the anti-fibrotic agent is used as a brake to suppress the differentiation of myofibroblast precursor cells during the wound healing process, the area in which the anti-fibrotic agent acts as a brake to prevent the anti-fibrotic agent from inhibiting skin epithelialization and slowing natural healing is limited in location so that it does not reach the skin epithelium.

[0450] To elaborate further, according to the disclosure of this application, the scaffold is placed in a position of the skin that is isolated from the epidermis, as is self-evident from Figure 7 etc. Furthermore, when the scaffold is injected as a liquid agent into the gap of the sutured skin wound using the injector exemplarily shown in Figure 23 or an equivalent (e.g., a pipette tip), it is self-evident that the tip of the injector is inserted deep into the gap of the sutured skin wound, and as a result, the liquid agent is prevented from moving to the skin epithelium.

[0451] As a result, according to the disclosure of this application, the effect of "suppressing the delay of natural healing in the presence of an anti-fibrotic agent" is obtained.

[0452] This is also supported by the experimental results comparison table shown in Figure 12. Specifically, in both the control group and the treatment group, skin epithelialization was not observed until day 3 after the start of the experiment, but skin epithelialization was observed on day 7. This confirms that epithelialization of the wound site occurred in all groups, regardless of whether an anti-fibrotic agent was administered or not.

[0453] This means that even in situations where scarring is inhibited by an antifibrotic agent during the wound healing process, wound healing occurs without any relative delay or inferiority compared to situations in which the antifibrotic agent is not present.

[0454] That is, according to the disclosure of this application, an anti-fibrotic agent is locally administered inside (within the gap) of a wound such as a sutured skin wound to a location of the wound that is isolated from the skin epithelium, thereby preventing the skin epithelium from being exposed to the anti-fibrotic agent.

[0455] (5) The third issue and technical concept of the "skin suture wound placement method"

[0456] The disclosure of this application further states that a third objective is to "achieve both the inhibition of scar formation and the inhibition of delayed wound healing in intact wounds," and to achieve this third objective, a third technical concept is adopted in which a scaffold containing both a biomaterial and an anti-fibrotic agent is placed in the body at the early stage of the wound healing process.

[0457] Further expanding on this, if the wound is an intact wound and a scaffold is placed in the wound during the early stages of the wound healing process, differentiation into myofibroblasts is suppressed during the early stages of the wound healing process, resulting in no excessive collagen production and ultimately in suppression of excessive scarring.

[0458] (6) Regarding the medical contribution of the disclosure of this application in enabling harmless use of ROCK inhibitors

[0459] Several papers have already reported that ROCK inhibitors, including fasudil and ripasudil, have various effects on keratinocytes, which make up the epithelial layer of the skin. In particular, it is well known that the ROCK inhibitor ripasudil, which is widely used as an eye drop, causes blepharitis, a type of skin disease, as a side effect. This fact has also been reported in several papers.

[0460] As such, ROCK inhibitors are drugs that are highly likely to cause skin disorders when administered into a wound as an anti-fibrotic agent, and therefore trials of them on human skin wounds have been avoided.

[0461] In contrast, according to the disclosure of this application, the anti-fibrotic agent is not placed in the wound alone, but is placed in the wound while being held or captured in a biomaterial that functions as a support, and the biomaterial is placed in a limited location within the wound that is isolated from the skin epithelium.

[0462] As a result, the anti-fibrotic agent is prevented from unintentionally migrating within the wound and reaching the skin epithelium, thereby preventing exposure of the anti-fibrotic agent to the skin epithelium, a site where early regeneration is important in wound healing.

[0463] Therefore, the disclosure of this application makes it possible to use a ROCK inhibitor as an anti-fibrotic agent in wounds while suppressing its side effects, and in this respect makes a medical contribution.

[0464] Disclosure of laparotomy method and surgical set for wound healing

[0465] The application further discloses, as exemplary embodiments, an open surgical procedure and a surgical wound healing set including the instruments and medications necessary to perform the open surgical procedure, as related to wound healing.

[0466] The open surgical method may be, for example, partially or completely performed by a surgeon and / or robotically.

[0467] This laparotomy method is, for example, a laparotomy method in which the abdominal wall of a patient is incised and a treatment is performed on a target organ in the abdominal cavity of the patient.

[0468] This laparotomy surgery method is configured to include an incision step of cutting the abdominal wall to form a skin incision, a surgical step of introducing a surgical instrument from outside the body into the abdominal cavity through the skin incision in the abdominal wall to perform a surgical operation on the target organ, a suturing step of suturing the skin incision after completion of the surgical operation to form a skin sutured wound, and an introduction step of introducing a wound scaffold into the formed skin sutured wound.

[0469] The wound scaffold material is configured so that a bioabsorbable biomaterial and an anti-fibrotic agent are each the main components, and the biomaterial contains the anti-fibrotic agent at least during use of the wound scaffold material.

[0470] The biomaterial is configured to function, in use, in the skin wound as an artificial extracellular matrix having properties to which the patient's myofibroblast precursor cells can adhere.

[0471] The wound scaffold illustrated in Figure 4(a) has a dosage form of a sheet, whereas in the examples shown in Figures 6 and 7, the wound scaffold is a liquid, and the introducing step in the laparotomy surgery method includes a step of injecting the wound scaffold into the sutured skin wound using an injector configured to have a main body for containing the liquid and a nozzle for discharging the contained liquid, with at least a tip of the nozzle inserted into the gap in the sutured skin wound.

[0472] An example of the wound healing surgical set is configured to include the wound scaffold and the injector.

[0473] In this example, the wound scaffold and the injector may be configured as a single finished product (e.g., a product in which the wound scaffold is pre-loaded into the injector), or may be configured as a kit product in which each part is provided as a separate component and assembled by a worker on-site.

[0474] Furthermore, in some aspects of the aforementioned embodiments, for ease of explanation, attention has been focused on a method in which the wound scaffold is placed in an incision made in the skin as the wound after the incision has been sutured, as an example of a scaffold placement method in which the wound scaffold is placed in the incision.

[0475] However, instead of this method, a scaffold placement method can be adopted in which the scaffold is placed in the incision before it is sutured.

[0476] A scaffold for placement in the abdominal cavity according to one embodiment and a method for placing the same in the abdominal cavity have already been described in this specification with reference to FIGS. 4( c) to 4(e). Here, several features of this embodiment will be described in more detail.

[0477] The scaffold according to this embodiment is an artificial wound scaffold for inhibiting adhesions within the abdominal cavity after intraperitoneal laparotomy. This scaffold has the following configurations: first configuration: a bioabsorbable biomaterial containing an anti-fibrotic agent; second configuration: the biomaterial is configured to function as an artificial extracellular matrix to which precursor cells of myofibroblasts in the body have the property of adhering, and acts as a cell culture scaffold using a hydrogel as a substrate; third configuration: the anti-fibrotic agent has a molecular weight of 400 or less, and inhibits the differentiation of the precursor cells into myofibroblasts in the body, thereby providing anti-fibrosis to a peritoneal side surgical wound, which is a surgical wound formed in the peritoneum on the abdominal wall side, and an organ side surgical wound, which is a surgical wound formed in a target organ; and fourth configuration: the wound scaffold is in the form of a sheet, and is placed in the abdominal cavity between the peritoneum and the target organ, and on both sides of the wound scaffold, in surface contact with the local surrounding area of ​​the peritoneum including the peritoneal side surgical wound and the local surrounding area of ​​the target organ including the organ side surgical wound, respectively.

[0478] As described above, this embodiment employs a "surgical wound-targeting local administration method of an antifibrotic agent" as a method of administering the antifibrotic agent, in which the antifibrotic agent is administered locally to the surfaces of the peritoneum and the organ, targeting both the peritoneal surgical wound and the organ surgical wound, so as to cover both targets.

[0479] In this embodiment, as described above, the molecular weight of the antifibrotic agent is further reduced to 400 or less, thereby improving the cell permeability of the antifibrotic agent and thereby enhancing the effect of the antifibrotic agent in inhibiting the differentiation of the progenitor cells into myofibroblasts.

[0480] This embodiment employs both a "surgical wound-targeting local administration method of an antifibrotic agent" and "low molecular weight administration of an antifibrotic agent," so that both the peritoneal surgical wound and the organ surgical wound are targeted by the antifibrotic agent within the abdominal cavity, and the antifibrotic agent is administered with high penetration to both targets.

[0481] As a result, according to this embodiment, in both targets, differentiation into myofibroblasts is effectively inhibited, resulting in a synergistic effect in which a good anti-scarring effect is exerted, and ultimately a good adhesion-inhibiting effect is exerted.

[0482] According to this embodiment, furthermore, the safety of ripasudil and fasudil as antifibrotic agents can be improved by site-specific administration. This will be specifically explained below.

[0483] As mentioned above, the inventors knew at the time of the priority date of this application that ripasudil and fasudil (hereinafter, any one of these drugs will be referred to as the "antifibrotic agent of interest") may cause side effects when administered to target tissues in the body near the skin.

[0484] Therefore, the present inventors have come up with the idea that, rather than administering the antifibrotic agent of interest directly to the target tissue, the antifibrotic agent of interest can be trapped in a biomaterial and the rate of release or elution from the biomaterial can be limited, for example, by gradually releasing the antifibrotic agent of interest from the biomaterial, thereby positioning the antifibrotic agent of interest at an appropriate location in the body, and thereby administering the antifibrotic agent of interest to a site in the body where it will not cause side effects, thereby allowing the pharmacological effects of the antifibrotic agent of interest to be expressed without side effects.

[0485] Therefore, according to this embodiment, by administering the antifibrotic agent of interest into the peritoneal cavity while it is entrapped in a biomaterial, it becomes possible to use ripasudil and fasudil as antifibrotic agents that do not cause side effects, thereby achieving the effect of expanding the range of safe antifibrotic agent options for inhibiting intraperitoneal adhesions.

[0486] The details of some of the above-described embodiments or examples are provided for the purpose of interpreting the claims and should not be construed as limiting the scope of the present invention. Although only a few specific examples of the present invention have been described in detail in text in the above description, those skilled in the art will readily understand that many variations exist in those specific examples without substantially departing from the novel teachings and advantages of the present invention. For example, multiple features described in connection with one specific example may be incorporated, in whole or in part, into any other specific example of the present invention.

[0487] Therefore, all such modifications are intended to be encompassed within the scope of the present invention, which is defined in the following claims and all equivalents thereto. Furthermore, it is contemplated that many embodiments will not achieve all of the advantages of some embodiments, particularly the preferred embodiments described above, and the absence of a particular advantage does not necessarily mean that the embodiment in question is not within the scope of the present invention. Because various modifications can be made within the foregoing scope without departing from the scope of the present invention, all matter contained in the detailed description of the invention should be interpreted as interpreting the scope of the claims, and not as limiting them.

[0488] The wound scaffold according to the present invention is useful as a drug for inhibiting scar formation, adhesion, etc., and has industrial applicability.

Claims

1. An artificial wound scaffold applied to a wound in a living organism to heal the wound in a scar-inhibiting manner, comprising: a biomaterial that degrades in the living organism and is absorbed by the living organism, and is configured to function as an artificial extracellular matrix to which myofibroblast precursor cells of the living organism have the property of adhering; and an anti-fibrotic agent contained and retained in the biomaterial, which is gradually released from the biomaterial as the biomaterial dissolves in the living organism; the wound comprises at least one of an open wound and a closed wound in the living organism; and the anti-fibrotic agent has a molecular weight of less than 10,000 and inhibits the differentiation of the precursor cells into myofibroblasts in the living organism; The wound scaffold is placed on at least one of the target surfaces, which are the surface of the open wound, the surface of the closed wound, and a portion of the surface of tissue in the abdominal cavity of the living body where the closed wound is exposed, thereby enabling the anti-fibrotic agent to be administered locally to the target surface.

2. The wound scaffold according to claim 1, wherein the biomaterial is configured to have shape retention.

3. The wound scaffold according to claim 2, wherein the biomaterial uses a hydrogel as a base material.

4. The wound scaffold according to claim 3, wherein the hydrogel contains gelatin.

5. The wound scaffold of claim 1, wherein the antifibrotic agent comprises fasudil, ripasudil, pirfenidone, and nintedanib.

6. The wound scaffold of claim 1, wherein the anti-fibrotic agent has a molecular weight of 400 or less.

7. The wound scaffold of claim 6, wherein the anti-fibrotic agent comprises fasudil, ripasudil, and pirfenidone.

8. A wound scaffold according to claim 1, wherein the open wound includes a skin-side surgical wound formed in the skin of the living body by surgery, and the wound scaffold is administered locally to the target surface, with the wound surface of the skin-side surgical wound being the target surface.

9. The wound scaffold according to claim 8, wherein the wound scaffold is placed within the skin-side surgical wound so that the anti-fibrotic agent is administered locally to a location isolated from the epithelium of the skin.

10. A wound scaffold as claimed in claim 1, wherein the closed wound comprises a peritoneal-side surgical wound formed by surgery in the peritoneum on the abdominal wall side within the abdominal cavity, and an organ-side surgical wound formed by surgery in a target organ within the abdominal cavity, and the wound scaffold is administered locally to both of the target surfaces, which are the surface of the peritoneal-side local peripheral region on the inner surface of the abdominal wall where the peritoneal-side surgical wound is exposed, and the surface of the organ-side local peripheral region on the outer surface of the target organ where the organ-side surgical wound is exposed.

11. The wound scaffold according to claim 10, wherein the wound scaffold is in the form of a sheet and is placed in the abdominal cavity between the peritoneum and the target organ, and on both sides of the wound scaffold, in surface contact with the local peripheral area on the peritoneum side and the local peripheral area on the organ side, respectively, thereby inhibiting scarring of each surgical wound and making it possible to inhibit adhesion between the abdominal wall and the target organ after intraperitoneal laparotomy.

Images