WO2025014745A2

WO2025014745A2 - Composite hydrogel wound dressings and methods of use thereof

Info

Publication number: WO2025014745A2
Application number: PCT/US2024/036722
Authority: WO
Inventors: Joseph TANFANI; Sriramakamal Jonnalagadda; Jasmin Monpara
Original assignee: Saint Joseph's University
Current assignee: Saint Joseph's University
Priority date: 2023-07-07
Filing date: 2024-07-03
Publication date: 2025-01-16
Anticipated expiration: 2026-01-07
Also published as: WO2025014745A3

Abstract

The disclosure relates to wound dressings comprising a hydrogel composition, methods of preparing the same, and methods of use thereof for treating and/or ameliorating wounds. In certain embodiments, the wound dressing comprising a hydrogel composition is arranged in a honeycomb pattern. In certain embodiments, the wound dressing comprising a hydrogel composition comprises one or more antibiotic agents.

Description

TITLE OF THE INVENTION

Composite Hydrogel Wound Dressings and Methods of Use Thereof

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims priority to U.S. Provisional Patent Application No. 63/525.399, fded July 7, 2023. which is hereby incorporated by reference herein in its entirety.

BACKGROUND

Skin, comprising up to 16% of a person's body weight, is the body's largest organ and serves many important functions. Injuries to skin are extremely common. While many injuries can be treated by available over-the-counter treatments, more serious injuries require clinical attention.

Wound dressings are a common, first-line treatment for traumatic skin injuries such as bum wounds. An ideal wound dressing should provide complete coverage over the wound area, possess several characteristics, including but not limited to breathability. sufficient porosity, and/or biocompatibility. Many currently used wound dressings demonstrate deficiencies in one or more criteria, and accordingly do not represent ideal wound dressings. For example, during healing, applied dressings (e.g, gauze and/or adhesive bandages) can adhere to the wound, making removal painful for the patient. Additionally, many wound dressings fail to provide adequate moisture to the wound bed, which may delay the overall healing process.

There is thus a need in the art for improved wound dressing compositions and methods of use thereof. The present disclosure addresses this need.

BRIEF SUMMARY

In one aspect, the disclosure provides a wound dressing comprising a hydrogel composition. In certain embodiments, the hydrogel composition comprises at least one poloxamer and at least one cross-linked alginate. In certain embodiments, the least one poloxamer and the at least one cross-linked alginate have a mass ratio ranging from about 40:1 (w/w) to about 70: 1 (w/w) in the hydrogel composition. In certain embodiments, the hydrogel composition is arranged in a repeating hexagonal pattern.

In another aspect, the disclosure provides a method of preparing the wound dressing of the disclosure. In certain embodiments, the method comprises preparing a first mixture comprising a poloxamer solution and an alginate solution. In certain embodiments, the poloxamer solution comprises at least one poloxamer having a concentration of about 20% to about 30% (w/v) poloxamer. In certain embodiments, the alginate solution comprises at least one alginate having a concentration of about 0.1% (w/v) to about 5% (w/v) alginate. In certain embodiments, the method comprises arranging the mixture in a repeating hexagonal pattern to provide an arranged mixture. In certain embodiments, the mixture is arranged in a repeating hexagonal pattern via extrusion using 3D printer. In certain embodiments, the method comprises contacting the arranged mixture with an aqueous solution comprising at least one cationic salt.

In another aspect, the disclosure provides a wound dressing prepared according to a method of the disclosure.

BRIEF DESCRIPTION OF THE FIGURES

The drawings illustrate generally, by way of example, but not by way of limitation, various embodiments of the present application.

FIGs. 1 A-1C illustrate PATCH grid scaffold Computer-Aided Design (CAD) and preparation. FIG. 1A: open-source BLENDER® software is used to prepare the grid design and the method is exported as a . stl file for bioprinting. FIG. IB: PATCH grid scaffold directly after bioprinting. FIG. 1C: PATCH grid scaffold after crosslinking in 1.5% (w/v) calcium chloride (CaCb) for 15 min.

FIGs. 2A-2C illustrate PATCH honeycomb scaffold Computer-Aided Design (CAD) and preparation. FIG. 2A: open-source Blender® software is used to prepare the honeycomb design and the model is exported as a stl file for bioprinting. FIG. 2B: PATCH honeycomb scaffold, colored by FD&C Blue No. 2 (i.e.. indigotine), directly after bioprinting. FIG. 2C: PATCH honeycomb scaffold after crosslinking in 1.5% (w/v) calcium chloride (CaCb) for 15 mins.

FIGs. 3A-3B illustrate PATCH bioprinting process. FIG. 3A: PATCH honeycomb scaffold is bioprinted onto a 60 mm Petri dish utilizing an extrusion-based bioprinter. FIG. 3B: PATCH honeycomb scaffold directly after the bioprinting process.

FIG. 4 illustrates neomycin sulfate dissolution from honeycomb PATCH. Neomycin sulfate dissolution was quantified using absorbance at 356 nm after Hantzsch reaction. Samples were collected and reacted with the assay reagent to produce colored dihydrolutidine derivatives a previously prepared calibration curve was used to measure cumulative neomycin release. FIGs. 5A-5C illustrate dried and rehydrated PATCH honeycomb scaffold. FIG. 5A: PATCH honeycomb scaffold after drying for several weeks at room temperature. FIG. 5B: water is added to dried PATCH scaffold and the scaffold regains its swollen state after several minutes. FIG. 5C: reconstituted PATCH scaffold retains its physical integrity when picked up with metal spatula.

FIG. 6 illustrates results of a swelling study of exemplary dried honeycomb PATCH scaffolds of the disclosure, without neomycin sulfate, after swelling for a period of 24. 48. 120, and 168 hours.

FIG. 7 illustrates empirical results of a porosity study of exemplary PATCH scaffolds of the disclosure, either with or without neomycin, using an ethanol displacement method.

FIG. 8 provides an overlay of X-ray powder diffraction (XRPD) patterns of different of certain exemplary PATCH scaffolds of the disclosure. Samples tested: (a) PATCH scaffold crosslinked with CaCh and then soaked in 3% (w/v) neomycin sulfate for 15 minutes; (b) PATCH scaffold crosslinked with CaCh (not soaked in neomycin); (c) PATCH scaffold not crosslinked with CaCh; (d) Poloxamer 407 27% gel (PATCH ingredient; allowed to dry); (e) Poloxamer 407 powder from stock bottle; (f) sodium alginate 2% (w/v) gel (PATCH ingredient) dried film; (g) sodium alginate from stock bottle; (h) neomycin 3% (w/v) solution, allowed to dry; and (i) neomycin sulfate active pharmaceutical ingredient (API) powder from stock bottle.

DETAILED DESCRIPTION OF THE INVENTION

Reference will now be made in detail to certain embodiments of the disclosed subject matter, examples of which are illustrated in part in the accompanying drawings. While the disclosed subject matter will be described in conjunction with the enumerated claims, it will be understood that the exemplified subject matter is not intended to limit the claims to the disclosed subject matter.

Throughout this document, values expressed in a range format should be interpreted in a flexible manner to include not only the numerical values explicitly recited as the limits of the range, but also to include all the individual numerical values or sub-ranges encompassed within that range as if each numerical value and sub-range is explicitly recited. For example, a range of “about 0.1% to about 5%” or “about 0.1% to 5%” should be interpreted to include notjust about 0.1% to about 5%, but also the individual values (e.g., 1%, 2%, 3%, and 4%) and the sub-ranges (e.g., 0.1% to 0.5%, 1.1% to 2.2%, 3.3% to 4.4%) within the indicated range. The statement “about X to Y” has the same meaning as “about X to about Y,” unless indicated otherwise. Likewise, the statement ‘‘about X, Y, or about Z” has the same meaning as “about X, about Y, or about Z,” unless indicated otherwise.

In this document, the terms “a,” “an,” or “the” are used to include one or more than one unless the context clearly dictates otherwise. The term “or” is used to refer to a nonexclusive “or” unless otherwise indicated. The statement “at least one of A and B” or “at least one of A or B” has the same meaning as “A, B, or A and B.” In addition, it is to be understood that the phraseology or terminology employed herein, and not otherwise defined, is for the purpose of description only and not of limitation. Any use of section headings is intended to aid reading of the document and is not to be interpreted as limiting; information that is relevant to a section heading may occur within or outside of that particular section. All publications, patents, and patent documents referred to in this document are incorporated by reference herein in their entirety, as though individually incorporated by reference.

In the methods described herein, the acts can be carried out in any order, except when a temporal or operational sequence is explicitly recited. Furthermore, specified acts can be carried out concurrently unless explicit claim language recites that they be carried out separately. For example, a claimed act of doing X and a claimed act of doing Y can be conducted simultaneously within a single operation, and the resulting process will fall within the literal scope of the claimed process.

Description

The present disclosure relates in part to bioprinted, honey comb/grid scaffolds comprising a composite hydrogel, which is useful for promoting wound healing. In certain embodiments, the porous nature of the biocompatible hydrogels permit and/or promotes neovascularization and facilitate incorporation into a wound bed. In certain embodiments, the hydrogel composition provides a scaffold onto which cells can attach and repopulate the wound bed. In certain embodiments, the hydrogel composition releases a topical antibiotic to prevent further infection.

In certain embodiments, the hydrogel compositions are referred to herein as Poloxamer Alginate Therapeutic Composite Hydrogels (PATCHs). In certain embodiments, the PATCH is prepared by bioprinting, and thus the PATCH system can be prepared in according to any of a number of distinct shapes as necessary, to optimally accommodate irregular wound shapes. In certain embodiments, the PATCH scaffolds are painless to remove and/or apply, and may provide relief upon application.

Further, PATCHs can be prepared according to any of a number of distinct patterns. including but not limited to repeating hexagonal (z'.e., honeycomb) or square (z'.e., grid) designs. In certain embodiments, the PATCH is prepared according to a hexagonal, honeycomb design. In certain embodiments, the honeycomb design permits maximum mechanical strength of the printed PATCH compositions while using minimal material.

In certain embodiments, this design ensures that the scaffold persists in the wound bed long enough to allow for cell attachment to the scaffold and subsequent proliferation, helping to deter bacterial infection with antibiotic release. Further, the porous design of the PATCH gels permits and/or promotes neovascularization, an integral part of wound healing.

The PATCH composition swells upon contact with an aqueous solution, including but not limited to wound exudate. Thus, in certain embodiments, the PATCH composition may be applied to a wound while in a dry, shrunken state, whereby the PATCH composition swells upon release of exudate from the wound. Alternatively, the PATCH composition may be applied to the wound in a hydrated, swollen state. In certain embodiments, the PATCH composition can be adhered to an ordinary' fabric or gauze bandage and applied to a wound.

Further, in certain embodiments, the PATCH compositions described herein possess thermoreversibility owing to the dual-polymeric composition. Without wishing to be bound by any theory, it has been proposed that, on the molecular level, poloxamer micelle aggregation as a function of raised temperature causes the sol-gel transition of the PATCH composition before chemical cross-linking of the alginate by CaCh. Thus, the PATCH compositions described herein are cross-linked by two methods (z.e., physical cross-linking of the poloxamer and chemical cross-linking of the alginate polymer network).

This dual cross-linking provides improved structural integrity⁷ compared to compositions comprising the components in isolation and contributes to a robust structure. That is. compositions consisting essentially of poloxamer gels at the concentrations described herein for exemplary compositions (z.e., about 20%-26% w/v) revert from a gel state at elevated temperature to a liquid-like (sol state) at room temperature. Further, similar compositions consisting essentially of poloxamer gels (e.g., 35% w/v P407 poloxamer) lack integrity in an aqueous environment, readily dissolving under conditions common to the wound interface (e.g., 2 hours at 37 °C in aqueous media).

Thus, in one aspect, the present disclosure provides PATCH formulations which permit a stable gel state to be maintained at non-elevated temperatures (e.g., room temperature) at a wound interface.

The present disclosure describes the design, preparation, and selected characterization of non-limiting exemplary PATCH hydrogel compositions of the present invention. Definitions

The term '‘3D print” or “3D object” refers to a three-dimensional object obtained by 3D printing (additive manufacturing process), such as an object having a height, width and length.

The term “about” as used herein can allow for a degree of variability in a value or range, for example, within 10%. within 5%. or within 1% of a stated value or of a stated limit of a range, and includes the exact stated value or range.

The term absorb as used herein refers to the penetration of one substance into the mass of another substance (e.g., intake and/or impregnation of a liquid, or a solute dissolved therein, into a matrix in such as way as to not be readily removed) and/or onto the surface thereof (e.g., adsorption). The term “adsorbed” refers to the process by which a substance (e.g, small molecule) adheres to the surface of a solid or liquid material. The substance being adsorbed is know n as the “adsorbate,” while the material onto which it is adhering is called the “adsorbent.” This process can occur through various mechanisms, such as physical adsorption (e.g., physisorption), in which the adsorbate is attracted to the surface of the adsorbent through weak intermolecular forces, or chemical adsorption (also known as chemisorption), in which the adsorbate forms chemical bonds w ith the adsorbent surface.

The term “alginate” is meant to refer to the sodium salt of alginic acid. In certain embodiments, alginic acid refers to a linear copolymer with homopolymeric blocks of (1-4)- linked P-D-mannuronate (M) and its C-5 epimer a-L-guluronate (G) residues, respectively, which are covalently linked together in different sequences or blocks.

The term “antibiotic agent” refers to naturally occurring, semisynthetic, or fully synthetic agents which inhibit the growth of microbes (i.e., bacteria, fungi, viruses, parasites and microbial spores) thereby preventing their development and microbial or pathogenic action. An antibiotic agent can be selected from the group consisting of small organic or inorganic molecules; saccharides; oligosaccharides; polysaccharides; biological macromolecules, e g., peptides, proteins, and peptide analogs and derivatives; pepti do mimetics; antibodies and antigen binding fragments thereof; nucleic acids; nucleic acid analogs and derivatives; glycogens or other sugars; immunogens; antigens; an extract made from biological materials such as bacteria, plants, fungi, or animal cells; animal tissues; naturally occurring or synthetic compositions; and any combinations thereof. As used herein, the term “antibiotic agent” is intended to embrace antibacterial agents or antimicrobial agents, antifungal agents, antiprotozoal agents, antiviral agents and mixtures thereof. Non-limiting, exemplary antibiotic agents include, but are not limited to, acrosoxacin, amifioxacin, amikacin, amoxycillin, ampicillin, aspoxicillin, azidocillin, azithromycin, aztreonam, balofloxacin, biapenem, brodimoprim, cefaclor, cefadroxil, cefatrizine, cefcapene, cefdinir, cefetamet, ceftmetazole, cefoxitin, cefprozil, cefroxadine, ceftarolin, ceftazidime, ceftibuten, ceftobiprole, cefuroxime, cephalexin, cephalonium, cephaloridine, cephamandole, cephazolin, cephradine, chlorquinaldol, chlortetracycline, ciclacillin, cinoxacin, ciprofloxacin, clarithromycin, clavulanic acid, clindamycin, clofazimine, cloxacillin. colistin, danofloxacin, dapsone, daptomycin, demeclocycline, dicloxacillin, difloxacin, doripenem, doxycycline, enoxacin, enrofloxacin, erythromycin, fleroxacin, flomoxef, flucloxacillin, flumequine, fosfomycin, gentamycin, isoniazid, imipenem, kanamycin, levofloxacin, linezolid. mandelic acid, mecillinam, meropenem. metronidazole, minocycline, moxalactam, mupirocin, nadifloxacin, nalidixic acid, netilmycin, netromycin, nifuirtoinol, nitrofurantoin, nitroxoline, norfloxacin, ofloxacin, oxytetracycline, panipenem, pefloxacin, phenoxymethylpenicillin, pipemidic acid, piromidic acid, pivampicillin, pivmecillinam, prulifloxacin, rufloxacin, sparfloxacin, sulbactam, sulfabenzamide, sulfacytine, sulfametopyrazine. sulphacetamide. sulphadiazine, sulphadimidine. sulphamethizole, sulphamethoxazole, sulphanilamide, sulphasomidine, sulphathiazole, teicoplanin, temafioxacin, tetracycline, tetroxoprim, tigecyclin, tinidazole, tobramycin, tosufloxacin, trimethoprim, vancomycin, and pharmaceutically acceptable salts or esters thereof.

The term “contacting” as used herein refers to bringing at least two different compounds and/or compositions in physical proximity as to allow physical and/or chemical interaction of said compounds. A non-limiting example of facilitating contact include immersion.

The term “cross-link” or “cross-linked” as used herein in the context of alginate polymers refers to the process of forming bonding interactions (e.g, ionic and/or electrostatic interactions) between alginate polymer chains upon contact with certain divalent cations, including but not limited to calcium ions (z.e., Ca²⁺). Upon exposure of alginate to a calcium rich solution, calcium ions form cross-links between polymer chains, generating a three- dimensional network or “gel” matrix.

The term “extrusion” as used herein refer to a process of creating a 3D object, layer by layer, whereby a composition (e.g., hydrogel formulation) is forced through a small nozzle or opening (e.g., via application of pressure) in a controlled manner, following a predetermined path that is generated by a 3D modeling software or computer-aided design (CAD) file. In certain embodiments, the composition (e.g, hydrogel formulation) is warmed to a temperature of about 30 °C to about 40 °C prior to extrusion so as to induce gelation via thermoreversible gelation. In certain embodiments, the nozzle or opening has a temperature of about 37 °C.

The term “honeycomb pattern” or “honeycomb scaffold” as used herein refers to an arrangement of a material, wherein the material comprises a plurality of interconnected cell walls that define a plurality’ of cells, wherein the cells comprise regular or irregular hexagonal cells. The terms “regular hexagon” or “regular hexagonal” as used herein refer to a six-sided, two-dimensional geometric shape or polygon. It is a closed shape with six straight sides and six angles, each measuring 120 degrees. All of the sides of a hexagon are equal in length, and all of the angles are also equal. A regular hexagon has six equal sides and six equal angles, while an irregular hexagon has sides and angles of varying lengths and sizes. The term “hexagonal” as used herein, may include both irregular hexagons and regular hexagons while, the terms “irregular hexagon” and “regular hexagon” excludes a regular hexagon and irregular hexagon, respectively

The term “hydrogel” as used herein refers to a crosslinked polymeric material which is not water-soluble and can contains at least 10% by weight of water within its polymer matrix when fully hydrated.

The terms “patient,” “subject,” or “individual” are used interchangeably herein, and refer to any animal, or cells thereof whether in vitro or in situ, amenable to the methods described herein. In a non-limiting embodiment, the patient, subject or individual is a human.

The term “poloxamer” denotes non-ionic triblock copolymers composed of a central hydrophobic chain of polypropylene oxide) (PPO) flanked by two hydrophilic chains of poly(ethylene oxide) (PEO), each PPO or PEO chain can be of different molecular weights. Poloxamers are also known by the trade name Synperonics. Pluronics and Kolliphor. Poloxamers are available in wide range of molecular weights, melting points and hydrophilicity and are commonly used in the pharmaceutical formulations as wetting agents to improve the bioavailability. For the generic term “poloxamer”, these copolymers are commonly named with the letter “P” (for poloxamer) followed by three digits, the first two digits x 100 give the approximate molecular mass of the polyoxypropylene core, and the last digit x 10 gives the percentage polyoxyethylene content (e g., P407 = poloxamer with a poly oxypropylene molecular mass of 4,000 g/mol and a 70% polyoxyethylene content; Pl 88 = poloxamer with a polyoxypropylene molecular mass of 1.800 g/mol and a 80% polyoxyethylene content). For the Pluronic trade name, coding of these copolymers starts with a letter to define its physical form at room temperature (L = liquid, P = paste, F = flake (solid)) followed by two or three digits. The first digit (two digits in a three-digit number) in the numerical designation, multiplied by 300, indicates the approximate molecular weight of the hydrophobe; and the last digit x 10 gives the percentage polyoxyethylene content (e.g., L61 = Pluronic with a poly oxypropylene molecular mass of 1,800 g/mol and a 10% polyoxyethylene content). In the example given, poloxamer 181 (P181) = Pluronic L61, poloxamer 407 = Pluronic F127. For example, Poloxamer 188 is a poloxamer wherein the PPO chain has a molecular mass of 1800 g/mol and a PEO content of 80% (w/w).

The term “polymer” may include, according to some embodiments, any molecule comprising repeating structural units connected to each other, typically, by covalent chemical bonds. The term “polymer” may include, according to some embodiments, a homopolymer (which is a polymer derived from one monomer species), a copolymer (which is a polymer derived from two (or more) monomeric species) or a combination thereof. A polymer, as referred to herein, may include a mixture of polymers. A polymer, as referred to herein, may include linear and/branched poly mers which consist of a single main chain with one or more polymeric side chains.

The terms “treating” and “ameliorating” as used herein refer to the various steps involved in wound healing. Wound treatment is not limited to skin treatment, but also includes tissue repair of other ty pes of wounds as described above. Further, treatment of a wound may comprise reduction and/or amelioration of pain and/or infections associated with the wound.

The term “wound” as used herein refers to bums, ulcers (e.g, diabetic ulcers, vascular deficiencies, swelling and bum-derived ulcers) exfoliated skin, or other skin problems e.g., allergies), scratches, cuts, abrasions, and damage to tissues or skin caused by surgical procedures (e.g, those caused by minimally invasive surgery, laparoscopic surgery, robotic surgery, incision biopsy, general surgery' and cosmetic surgery), inter alia. Wounds can range from superficial (e.g., affecting only the epidermis) to deeper trauma (e.g., lesions affecting a layer of tissue under the skin or epidermis). The wound may be of any length or shape, for example, in certain embodiments, the wound may be straight, jagged or curved.

The term “wound dressing” applies to all compositions and/or materials, including gel compositions, fabrics, textiles, or any combination thereof, which is directly placed upon a wound for protection of the wound against infection and mechanical damage and for absorbing exudate and debris from the wound.

Wound Dressing Compositions In one aspect, the present disclosure provide a wound dressing comprising a hydrogel composition. In certain embodiments, the hydrogel composition comprises (a) at least one poloxamer. In certain embodiments, the hydrogel composition comprises (b) at least one cross-linked alginate. In certain embodiments, the least one poloxamer and the at least one cross-linked alginate have a mass ratio ranging from about 40: 1 (w/w) to about 70: 1 (w/w) in the hydrogel composition. In certain embodiments, the hydrogel composition is arranged in a repeating hexagonal pattern (z.e., honeycomb lattice).

In certain embodiments, the at least one poloxamer is a compound of formula (I):

wherein: x and z are each independently an integer ranging from 2 to 130; and y is an integer ranging from 15 to 67.

In certain embodiments, x is selected from the group consisting of 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22. 23. 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34,

35. 36. 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47. 48. 49. 50. 51. 52. 53, 54, 55, 56, 57, 58, 59,

60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84,

85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106,

107, 108. 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120. 121, 122, 123, 124, 125, 126. 127, 128, 129, and 130.

In certain embodiments, y is selected from the group consisting of 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, and 67.

In certain embodiments, z is selected from the group consisting of 2, 3, 4. 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34,

35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59,

85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97. 98, 99, 100, 101, 102, 103, 104, 105, 106.

107, 108. 109, 110, 111. 112, 113. 114, 115, 116, 117, 118. 119, 120. 121, 122, 123. 124, 125, 126, 127, 128, 129, and 130.

In certain embodiments, x is 101, y is 56, and z is 101.

In certain embodiments, the at least one poloxamer is P407. In certain embodiments, the at least one poloxamer is Pl 88. In certain embodiments, the at least one poloxamer is P288. In certain embodiments, the at least one poloxamer is P335. In certain embodiments, the at least one poloxamer is P338. In certain embodiments, the at least one poloxamer comprises Pl 88, and the hydrogel composition further comprises at least one additional poloxamer.

In certain embodiments, the at least one poloxamer comprises about 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or about 30% (w/v) to about 30% (w/v) of the hydrogel composition. In certain embodiments, the at least one poloxamer comprises more than about 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or about 30% (w/v) to about 30% (w/v) of the hydrogel composition. In certain embodiments, the at least one poloxamer comprises less than about 15, 16, 17. 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or about 30% (w/v) to about 30% (w/v) of the hydrogel composition. In certain embodiments, the at least one poloxamer comprises about 22% (w/v) of the hydrogel composition.

In certain embodiments, the at least one cross-linked alginate comprises about 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9. or about 1.0% (w/v) of the hydrogel composition. In certain embodiments, the at least one cross-linked alginate comprises about 0.4% (w/v) of the hydrogel composition.

In certain embodiments, the at least one poloxamer and the cross-linked alginate have amass ratio of about 40:1, 41:1, 42:1, 43:1, 44:1, 45:1, 46:1, 47:1, 48:1, 49:1, 50:1, 51:1, 52:1.53:1, 54:1, 55:1, 56:1, 57:1, 58:1, 59:1, 60:1, 61:1, 62:1, 63:1, 64:1, 65:1.66:1.67:1. 68:1, 69:1, or about 70:1 (poloxamer: alginate; w/w or w:w) in the hydrogel composition. In certain embodiments, the at least one poloxamer and the cross-linked alginate have a mass ratio of more than about 40:1, 41:1, 42:1, 43:1, 44:1, 45:1, 46:1, 47:1, 48:1, 49:1.50:1, 51:1, 52:1, 53:1, 54:1, 55:1, 56:1, 57:1, 58:1, 59:1, 60:1, 61:1, 62:1, 63:1, 64:1, 65:1.66:1.67:1. 68: 1, 69: 1, or about 70: 1 (poloxamer: alginate; w/w or w:w) in the hydrogel composition. In certain embodiments, the at least one poloxamer and the cross-linked alginate have a mass ratio of less than about 40:1, 41:1, 42:1, 43:1, 44:1, 45:1, 46:1, 47:1, 48:1, 49:1, 50:1, 51:1, 52:1, 53:1, 54:1, 55:1, 56:1, 57:1, 58:1, 59:1, 60:1, 61:1, 62:1, 63:1, 64:1, 65:1.66:1.67:1, 68: 1.69: 1. or about 70: 1 (poloxamer: alginate; w/w or w:w) in the hydrogel composition. In certain embodiments, the at least one poloxamer and the cross-linked alginate have a mass ratio of about 54: 1 (w/w) in the hydrogel composition.

In certain embodiments, the at least one alginate is cross-linked with an aqueous solution of CaCh, optionally wherein the aqueous solution has a CaCh concentration of about 1.5% (w/v). In certain embodiments, the hydrogel composition further comprises at least one adsorbed and/or absorbed antibiotic agent.

In certain embodiments, the antibiotic agent is neomycin. In certain embodiments, the antibiotic agent is polymyxin B. In certain embodiments, the antibiotic agent is bacitracin. In certain embodiments, the antibiotic agent is gentamicin. In certain embodiments, the antibiotic agent is silver. In certain embodiments, the antibiotic agent is clindamycin. In certain embodiments, the antibiotic agent is erythromycin. In certain embodiments, the antibiotic agent is tetracycline. In certain embodiments, the antibiotic agent is metronidazole. In certain embodiments, the antibiotic agent is rifampin. In certain embodiments, the antibiotic agent is ciprofloxacin. In certain embodiments, the antibiotic agent is ofloxacin. In certain embodiments, the antibiotic agent is levofloxacin. In certain embodiments, the antibiotic agent is doxycycline. In certain embodiments, the antibiotic agent is mupirocin. In certain embodiments, the antibiotic agent is silver sulfadiazine. In certain embodiments, the antibiotic agent is mafenide acetate. In certain embodiments, the antibiotic agent is fusidic acid. In certain embodiments, the antibiotic agent is nadifloxacin. In certain embodiments, the antibiotic agent is retapamulin. In certain embodiments, the antibiotic agent is ozenoxacin,

In certain embodiments, the neomycin is adsorbed and/or absorbed by contacting the hydrogel composition with an aqueous solution of neomycin sulfate, optionally wherein the neomycin sulfate solution has a concentration of about 3% (w/v). and optionally wherein the contacting occurs for a period of about 15 minutes.

In certain embodiments, the repeating hexagonal pattern (i. e. , honeycomb lattice) comprises a continuous layer comprising the hydrogel composition, wherein continuous layer is characterized by evenly distributed hexagonal pores.

In certain embodiments, the continuous layer has a thickness of about 1.0, 1.5, 2.0, 2.5, 3.0, 3.5, 4.0, 4.5, or about 5.0 mm.

In certain embodiments, the hexagonal pores are evenly distributed at a distance of about 1.0, 1.5, 2.0, 2.5, 3.0, 3.5, 4.0, 4.5, or about 5.0 mm.

In certain embodiments, parallel sides of each hexagonal pore have a distance of about 1, 2, 3, 4, 5, 6, 7, 8, 9, or about 10 mm.

In certain embodiments, the hydrogel composition is arranged in a repeating hexagonal pattern (i.e.. honeycomb lattice) by 3D printing.

Methods In one aspect, the present disclosure provides a method of preparing a wound dressing comprising a hydrogel composition. In certain embodiments, the hydrogel composition comprises (a) at least one poloxamer. In certain embodiments, the hydrogel composition comprises (b) at least one cross-linked alginate. In certain embodiments, the least one poloxamer and the at least one cross-linked alginate have a mass ratio ranging from about 40: 1 (w/w) to about 70: 1 (w/w) in the hydrogel composition. In certain embodiments, the hydrogel composition is arranged in a repeating hexagonal pattern (/.e.. honeycomb lattice).

In certain embodiments, the method comprises (a) preparing a first mixture comprising a poloxamer solution and an alginate solution. In certain embodiments, the poloxamer solution comprises at least one poloxamer having a concentration of about 20% to about 30% (w/v) poloxamer. In certain embodiments, the alginate solution comprises at least one alginate having a concentration of about 0.1% (w/v) to about 5% (w/v) alginate.

In certain embodiments, the method comprises (b) arranging the mixture in a repeating hexagonal pattern (z.e., honeycomb lattice) to provide an arranged mixture, optionally wherein the mixture is arranged in a repeating hexagon pattern via extrusion using 3D printer (e.g, bioprinting).

In certain embodiments, the method comprises (c) contacting the arranged mixture with an aqueous solution comprising at least one cationic salt.

In certain embodiments, the aqueous solution comprising at least one cationic salt is an aqueous solution of CaCh. In certain embodiments, the aqueous CaCh solution has a CaCh concentration of about 1 .5% (w/v).

In certain embodiments, the contacting comprises immersion.

In certain embodiments, the contacting occurs for a period of about 15 minutes.

In certain embodiments, the method further comprises drying the wound dressing, optionally wherein the wound dressing is dried by air-exposure and/or freeze-drying.

In certain embodiments, x is selected from the group consisting of 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22. 23. 24. 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59,

60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72. 73. 74. 75, 76, 77, 78, 79, 80, 81, 82, 83, 84,

107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124,

125, 126, 127, 128, 129, and 130.

In certain embodiments, y is selected from the group consisting of 15, 16. 17. 18, 19, 20. 21. 22. 23, 24, 25, 26, 27, 28, 29, 30, 31, 32. 33. 34. 35. 36. 37, 38, 39, 40, 41, 42, 43, 44,

45, 46, 47, 48, 49, 50, 51, 52, 53, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, and 67.

In certain embodiments, z is selected from the group consisting of 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22. 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34,

35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47. 48. 49. 50. 51, 52, 53, 54, 55, 56, 57, 58, 59,

107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124, 125, 126. 127, 128, 129, and 130.

In certain embodiments, x is 101, v is 56, and z is 101.

In certain embodiments, the at least one poloxamer is P407. In certain embodiments, the at least one poloxamer is Pl 88. In certain embodiments, the at least one poloxamer is P288. In certain embodiments, the at least one poloxamer is P335. In certain embodiments, the at least one poloxamer is P338.

In certain embodiments, the at least one poloxamer comprises about 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or about 30% (w/v) to about 30% (w/v) of the hydrogel composition. In certain embodiments, the at least one poloxamer comprises more than about 15, 16, 17, 18, 19, 20, 21, 22, 23. 24. 25. 26, 27, 28, 29, or about 30% (w/v) to about 30% (w/v) of the hydrogel composition. In certain embodiments, the at least one poloxamer comprises less than about 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or about 30% (w/v) to about 30% (w/v) of the hydrogel composition. In certain embodiments, the at least one poloxamer comprises about 22% (w/v) of the hydrogel composition.

In certain embodiments, the at least one cross-linked alginate comprises about 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, or about 1.0% (w/v) of the hydrogel composition. In certain embodiments, the at least one cross-linked alginate comprises about 0.4% (w/v) of the hydrogel composition.

In certain embodiments, the at least one poloxamer and the cross-linked alginate have amass ratio of about 40:1, 41:1, 42:1, 43:1, 44:1, 45:1, 46:1, 47:1, 48:1, 49:1, 50:1, 51:1, 52:1, 53:1, 54:1, 55:1, 56:1, 57:1, 58:1, 59:1, 60:1, 61:1, 62:1, 63:1, 64:1, 65:1.66:1.67:1. 68: 1, 69: 1, or about 70: 1 (poloxamer: alginate; w/w or w:w) in the hydrogel composition. In certain embodiments, the at least one poloxamer and the cross-linked alginate have a mass ratio of more than about 40:1, 41:1, 42:1, 43:1, 44:1, 45:1, 46:1, 47:1, 48:1, 49:1, 50:1, 51:1, 52:1, 53:1, 54:1, 55:1, 56:1, 57:1, 58:1, 59:1, 60:1, 61:1, 62:1, 63:1, 64:1, 65:1.66:1.67:1, 68: 1.69: 1. or about 70: 1 (poloxamer: alginate; w/w or w:w) in the hydrogel composition. In certain embodiments, the at least one poloxamer and the cross-linked alginate have a mass ratio of less than about 40:1, 41:1, 42:1, 43:1, 44:1, 45:1, 46:1, 47:1, 48:1, 49:1, 50:1, 51:1, 52:1, 53:1, 54:1, 55:1, 56:1, 57:1, 58:1, 59:1, 60:1, 61:1, 62:1, 63:1, 64:1, 65:1.66:1, 67:1, 68: 1, 69: 1, or about 70: 1 (poloxamer: alginate; w/w or w:w) in the hydrogel composition. In certain embodiments, the at least one poloxamer and the cross-linked alginate have a mass ratio of about 54: 1 (w/w) in the hydrogel composition.

In certain embodiments, the at least one alginate is cross-linked with an aqueous solution of CaCh, optionally wherein the aqueous solution has a CaCh concentration of about 1.5% (w/v).

In certain embodiments, the hydrogel composition further comprises at least one adsorbed and/or absorbed antibiotic agent.

In certain embodiments, the antibiotic agent is neomycin. In certain embodiments, the antibiotic agent is polymyxin B. In certain embodiments, the antibiotic agent is bacitracin. In certain embodiments, the antibiotic agent is gentamicin. In certain embodiments, the antibiotic agent is silver. In certain embodiments, the antibiotic agent is clindamycin. In certain embodiments, the antibiotic agent is erythromycin. In certain embodiments, the antibiotic agent is tetracycline. In certain embodiments, the antibiotic agent is metronidazole. In certain embodiments, the antibiotic agent is rifampin. In certain embodiments, the antibiotic agent is ciprofloxacin. In certain embodiments, the antibiotic agent is ofloxacin. In certain embodiments, the antibiotic agent is levofloxacin. In certain embodiments, the antibiotic agent is doxycycline. In certain embodiments, the antibiotic agent is mupirocin. In certain embodiments, the antibiotic agent is silver sulfadiazine. In certain embodiments, the antibiotic agent is mafenide acetate. In certain embodiments, the antibiotic agent is fusidic acid. In certain embodiments, the antibiotic agent is nadifloxacin. In certain embodiments, the antibiotic agent is retapamulin. In certain embodiments, the antibiotic agent is ozenoxacin.

In certain embodiments, the continuous layer has a thickness of about 1.0, 1.5, 2.0. 2.5, 3.0, 3.5, 4.0, 4.5, or about 5.0 mm.

In certain embodiments, the hydrogel composition is arranged in a repeating hexagonal pattern (i.e., honeycomb lattice) by 3D printing.

In another aspect, the present disclosure provides a method of treating or ameliorating a wound of a subject, the method comprising applying to the wound the wound dressing of the present disclosure.

In certain embodiments, the wound is a bum. In certain embodiments, the wound is an ulcer. In certain embodiments, the wound is a laceration. In certain embodiments, the wound is an incision. In certain embodiments, the wound is a puncture. In certain embodiments, the wound is an abrasion. In certain embodiments, the wound is an avulsion. In certain embodiments, the wound is a sting. In certain embodiments, the wound is a bite.

In certain embodiments, the bum is a chemical bum. In certain embodiments, the bum is an electrical bum. In certain embodiments, the bum is a radiation bum.

In certain embodiments, the ulcer is a diabetic ulcer. In certain embodiments, the ulcer is a venous ulcer. In certain embodiments, the ulcer is an arterial ulcer. In certain embodiments, the ulcer is a pressure ulcer.

In certain embodiments, the wound dressing is applied and/or removed from the wound with minimal pain. In certain embodiments, the dressing is applied and/or removed from the wound without pain.

In certain embodiments, the wound dressing does not impede formation of new vasculature. In certain embodiments, the wound dressing promotes vasculature growth.

EXAMPLES Various embodiments of the present application can be better understood by reference to the following Examples which are offered by way of illustration. The scope of the present application is not limited to the Examples given herein.

Example 1: Scaffold design

BLENDER®, an open-source program used for animation, effects, and modeling, was used to prepare 3D models for the scaffolds described herein. Square lattice (FIGs. 1A-1C) and honeycomb designs (FIGs. 2A-2C) were prepared. Lattice scaffolds in the program were created by subdividing a 3D shape mesh, and applying a wireframe modifier to the vertices. Z-axis thickness was applied to a flat honeycomb mesh available in the program to create the honeycomb 3D scaffold model. The .blend file created by the program was exported as a .stl (standard triangle language) file to be used by SunP® Biomaker® software.

Example 2: Hydrogel composition design

As described elsewhere herein, in one aspect, the present disclosure describes the evaluation and physical characterization of exemplary poloxamer/alginate hydrogels, herein referred to as PATCHs (Poloxamer Alginate Therapeutic Composite Hydrogels). Whereas gels comprising poloxamer alone quickly dissolve into a solution state at room temperature, the incorporation of alginate, and subsequent crosslinking utilizing CaCh, allows for the gels to persist for significant time after 3D printing. Maintenance of a gel state upon application to a wound allows the scaffold to persist in the wound bed long enough to provide architecture on which cells can begin to grow.

In certain embodiments, the hydrogel compositions described herein comprise poloxamer P407. Although poloxamer P407 has been the most highly utilized poloxamer for tissue engineering, the present invention is not limited to compositions comprising P407. In certain embodiments, the poloxamer may comprise one or more distinct poloxamers. Nonlimiting examples of poloxamers suitable for use in the compositions described herein include Pl 88, P288, P335, and P338, inter alia. Further, compositions comprising each of poloxamers P407, Pl 88. P288. P335, and P338 were prepared and assessed for potential as tissue scaffolds. P407 was selected for further study because of its easy printability, its low toxicity, and its accepted use by the FDA as an excipient in pharmaceutical products.

In certain embodiments, the hydrogel compositions described herein comprise an antibiotic agent. In certain embodiments, the antibiotic agent is neomycin. Neomycin is an aminoglycoside antibiotic, often administered topically, which is useful against Gram- negative bacteria and certain Gram-positive bacteria. In certain embodiments, the neomycin is incorporated into the hydrogel compositions described herein as the trisulfate salt . In certain embodiments, a solution containing 3% (w/v) neomycin sulfate and 1.5% (w/v) CaCh is added to the scaffolds during the crosslinking step.

Example 3: Hydrogel composition preparation and in vitro neomycin release

A mixture suitable for use in bioprinting the compositions described herein was prepared by combining of P407 (27% w/v) and alginate (2% w/v) at a ratio of 4: 1 (P407: alginate). Next, Poloxamer Alginate Therapeutic Composite Hydrogel (PATCH) scaffolds of two shapes (z.e., square lattice and honeycomb, n=3 for each) were first printed in 60 mm petri dishes using a SunP® BioMaker® extrusion 3D printer (FIGs. 1A-1C, FIGs. 2A-2C, and FIGs. 3A-3B). The dimensions of each scaffold upon printing were 25 mm by 25 mm by 2.5 mm (x by y by z). 8 mL of 1.5% (w/v) CaCh solution was first added to each scaffold-containing petri dish to crosslink the alginate in the hydrogels for 5 minutes. After 5 minutes, 10 mL of 3% (w/v) neomycin sulfate solution was added to each petri dish to allow the hydrogels to adsorb and/or absorb neomycin sulfate. After more 15 minutes. CaCh/neomycin sulfate solution was drained away and the scaffolds were carefully washed with PBS and placed in scintillation vials.

The scintillation vials were then filled with 20 mL of PBS at pH 7.4 and placed within a shaker-incubator shallow water bath set to 37 °C and 50 RPM. 1 mL PBS was extracted from each vial at 15 minutes, 30 minutes, 1 hour, 2 hours, 4 hours, 8 hours, and 12 hours. After each 1 mL sample collection, the vials w ere replenished with 1 mL fresh PBS.

Neomycin sulfate was then quantified using a spectrophotometric method described in the literature and known to those of ordinary skill in the art. Briefly, 1 mL of PBS was first added to each collected 1 mL sample and vortexed. Then, 2 mL of prepared assay reagent, comprising 0.6 M phosphoric acid, 37.5 % (w/v) formaldehyde, and acetylacetone, was added to the collected dissolution samples. The mixtures were then vortexed, wrapped in foil, and bathed in boiling water for 20 minutes to facilitate a Hantzsch reaction between neomycin sulfate and the assay solution. The resulting colored complexes, a result of the dihydrolutidine derivatives produced during the Hantzsch reaction, w ere measured for absorbance at 356 nm. Neomycin sulfate concentration in the samples was then determined using a previously prepared calibration curve (FIG. 4).

Example 4: Dehydration and rehydration of exemplary hydrogel compositions In certain embodiments, the hydrogel compositions described herein can be dehydrated using various dehydration methods known to those of ordinary skill in the art (e.g., air-drying and/or freeze drying). In certain embodiments, dehydrate hydrogel compositions may be reconstituted (e.g., swelled) by contact with an aqueous solution. In certain embodiments, the dehydrated hydrogel composition is reconstituted and/or swelled by contact with an exudate, such that application of the dehydrated hydrogel composition directly to a wound results in reconstitution and/or swelling of the hydrogel composition (z.e., gelling). In other embodiments, the dehydrated hydrogel composition is reconstituted by contact with an aqueous solution which is not a biological fluid.

In a non-limiting experiment, an exemplary hydrogel composition of the present disclosure was dehydrated by exposure to ambient conditions for a period of several weeks (i.e., air-drying) (FIG. 5 A). In certain embodiments, the dried-composition was contacted with water, resulting in reconstitution and/or swelling of the dehydrated hydrogel composition (FIG. 5B). Further, the reconstituted hydrogel composition retained its physical integrity (FIG. 5C).

Example 5: Surface area and porosity of exemplary hydrogel compositions

The program Blender® was used to analyze the 3D meshes of honeycomb scaffolds to investigate the theoretical surface area of the scaffold model. The surface area of 25 mm by 25 mm by 2.5 mm honeycomb 3D model was calculated to be 1581.45 mm². PATCH scaffolds were printed in dimensions of 25 mm x 25 mm x 2.5 mm (x by y by z). After printing, scaffolds were immersed in 1.5% CaCh solution for 10 minutes to facilitate chemical crosslinking via cation exchange. Subsequently, CaCh solution was drained away. One set of PATCH scaffolds (n=3) were immersed in 10 mL of 3% (w/v) solution of neomycin sulfate to facilitate uptake of the antibiotic into the hydrogel. A second set of PATCH scaffolds (n=3) were not exposed to neomycin sulfate solution. All scaffolds were carefully blotted with KimWipes® to remove excess solution clinging to the scaffolds and allowed to dry at room temperature for 3 days. After drying, a 20 mL scintillation vial was filled with 10 mL of 97% ethanol (v/v) and weighed as Mo. This was done separately for each replicate. Scaffolds were then weighed as Mi. The dried scaffolds were then immersed in the ethanol within the scintillation vials and the filled vials were weighed as M2. The scaffolds were allowed to soak in the ethanol for a 1 hour period. Scaffolds were then removed and the scintillation vials with leftover ethanol were weighed as M3. Percentage porosity was determined using the following equation:

The percentage porosity (cpp) of PATCH honeycomb scaffolds with and without neomycin sulfate was investigated using the ethanol displacement method. FIG. 7 shows empirical results of the percentage of porosity of PATCH formulated either with or without neomycin. In one aspect, results show surprising consistency between formulations with or without neomycin, and the PATCH scaffolds also both demonstrate appropriate porosity as wound healing materials.

Example 6: Swelling study of exemplary hydrogel compositions

The swelling ability' of dried honeycomb PATCH scaffolds was assessed to understand their suitability as wound healing materials. PATCH scaffolds were initially printed as 25 mm x 25 mm x 2.5 mm (x by y by z). The scaffolds (n=3) were allowed to dry for 3 days at room temperature after printing. Their dry weight (wa) was then recorded. They were then immersed in an excess amount of MilliQ® water and incubated at 37° C. At the 24 hours, 2 days, 5 days, and 7 days, the scaffolds were taken out and carefully blotted with a KimWipe® to remove excess water clinging to the scaffolds. Their swollen weight (w_s) was then taken. Swelling percentage was calculated by the following equation:

Swelling Percentage = (^— — x 100

FIG. 6 shows the results from an experiment studying the swelling ability’ of dried honeycomb PATCH scaffolds (without neomycin sulfate). The gradual swelling effect is surprisingly steady and suitable for helping stabilize wounds and absorbing excess wound exudate, thereby helping to heal a subject.

Example 7: X-ray powder diffraction of exemplary hydrogel compositions

X-ray powder diffraction (XRPD) was performed to analyze the crystallinity or noncrystallinity of certain exemplary PATCH samples of the disclosure, individual components thereof, and certain mixtures thereof. Diffractograms were obtained by scanning for 26 from 0° to 40° on a Rigaku® MiniFlex. Samples were spread as a single layer onto diffraction plates prior to scanning. Samples prepared include: (a) PATCH scaffold crosslinked with CaCh and then soaked in 3% (w/v) neomycin sulfate for 15 minutes; (b) PATCH scaffold crosslinked with CaCh but not soaked in neomycin; (c) PATCH scaffold not crosslinked with CaCh; (d) Poloxamer 407 27% gel (PATCH ingredient) allowed to dry; (e) Poloxamer 407 powder from stock bottle; (f) sodium alginate 2% (w/v) gel (PATCH ingredient) dried film; (g) sodium alginate from stock bottle; (h) neomycin 3% (w/v) solution allowed to dry, and (i) neomycin sulfate active pharmaceutical ingredient (API) powder from stock bottle (FIG. 8).

Sodium alginate 2% (w/v) dried film, sodium alginate, neomycin sulfate 3% (w/v), and neomycin sulfate samples all exhibited a completely amorphous nature over the 0° to 40° scanning period. This can be ascertained from the complete absence of any peaks in the diffractogram curves. Pure poloxamer 407 showed moderate crystalline peaks at roughly 20° 29 and 24° 20, and much smaller peaks at roughly 27° 20 and 36.5° 20. These poloxamer 407 crystalline peaks share a similar intensity across the following samples: pure poloxamer 407, poloxamer 407 27% (w/v) dried hydrogel, and PATCH not crosslinked with CaCh. It can be seen from the remaining diffractograms that these characteristic poloxamer 407 peaks shrink in intensity' after the crosslinking process with CaCh, indicating that the alginate crosslinking process interrupts some of the inherent crystallinity in poloxamer 407 within the PATCH formulation.

Enumerated Embodiments

The following exemplary embodiments are provided, the numbering of which is not to be construed as designating levels of importance:

Embodiment 1 provides a wound dressing comprising a hydrogel composition comprising: (a) at least one poloxamer; and (b) at least one cross-linked alginate, wherein the least one poloxamer and the at least one cross-linked alginate have a mass ratio ranging from about 40: 1 (w/w) to about 70: 1 (w/w) in the hydrogel composition; and wherein the hydrogel composition is arranged in a repeating hexagonal pattern (/.e.. honeycomb lattice).

Embodiment 2 provides the wound dressing of Embodiment 1, wherein the at least one poloxamer is a compound of formula (I):

(I), wherein: x and z are each independently an integer ranging from 2 to 130; and y is an integer ranging from 15 to 67.

Embodiment 3 provides the wound dressing of Embodiment 1 or 2, wherein the at least one poloxamer is selected from the group consisting of P407, Pl 88, P288, P335, and

P338.

Embodiment 4 provides the wound dressing of Embodiment 2 or 3, wherein x and z are each 101, and wherein is 56 (i.e. P407).

Embodiment 5 provides the wound dressing of any one of Embodiments 1-4, wherein the at least one poloxamer comprises about 15% (w/v) to about 30% (w/v) of the hydrogel composition, optionally wherein the at least one poloxamer comprises about 22% (w/v) of the hydrogel composition.

Embodiment 6 provides the wound dressing of any one of Embodiments 1-5, wherein the at least one cross-linked alginate comprises about 0. 1% (w/v) to about 1.0% (w/v) of the hydrogel composition, optionally wherein the at least one cross-linked alginate comprises about 0.4% (w/v) of the hydrogel composition.

Embodiment 7 provides the wound dressing of any one of Embodiments 1 -6, wherein the at least one poloxamer and the cross-linked alginate have a mass ratio of about 54: 1 (w/w) in the hydrogel composition.

Embodiment 8 provides the wound dressing of any one of Embodiments 1-7, wherein the at least one alginate is cross-linked with an aqueous solution of CaCh, optionally wherein the aqueous solution has a CaCh concentration of about 1.5% (w/v).

Embodiment 9 provides the wound dressing of any one of Embodiments 1-8, wherein the hydrogel composition further comprises at least one adsorbed and/or absorbed antibiotic agent.

Embodiment 10 provides the wound dressing of Embodiment 9, wherein the antibiotic agent is at least one selected from the group consisting of neomycin, polymyxin B. bacitracin, gentamicin, silver, clindamycin, erythromycin, tetracycline, metronidazole, rifampin, ciprofloxacin, ofloxacin, levofloxacin, doxycycline, silver sulfadiazine, mafenide acetate, fusidic acid, nadifloxacin, retapamulin, ozenoxacin, and mupirocin.

Embodiment 11 provides the wound dressing of Embodiment 9 or 10, wherein the at least one antibiotic agent is neomycin.

Embodiment 12 provides the wound dressing of Embodiment 11, w herein the neomycin is adsorbed and/or absorbed by contacting the hydrogel composition with an aqueous solution of neomycin sulfate, optionally wherein the neomycin sulfate solution has a concentration of about 3% (w/v), and optionally wherein the contacting occurs for a period of about 15 minutes.

Embodiment 13 provides the wound dressing of any one of Embodiments 1-12, wherein the repeating hexagonal pattern (z.e., honeycomb lattice) comprises a continuous layer comprising the hydrogel composition, wherein continuous layer is characterized by evenly distributed hexagonal pores. Embodiment 14 provides the wound dressing of Embodiment 13, wherein the continuous layer has a thickness of about 1 to about 5 mm, optionally the continuous layer has a thickness of about 2.5 mm.

Embodiment 15 provides the wound dressing of Embodiment 13 or 14, wherein the hexagonal pores are evenly distributed at a distance of about 1 to about 5 mm, optionally wherein the hexagonal pores are evenly distributed at a distance of about 2.5 mm.

Embodiment 16 provides the wound dressing of any one of Embodiments 13-15. wherein parallel sides of each hexagonal pore have a distance of about 1 to about 10 mm, optionally wherein parallel sides of each hexagonal pore have a distance of about 5 mm.

Embodiment 17 provides the wound dressing of any one of Embodiments 1-16, wherein the hydrogel composition is arranged in a repeating hexagonal pattern (z.e., honeycomb lattice) by 3D printing.

Embodiment 18 provides a method of preparing the wound dressing of any one of Embodiments 1-17, the method comprising:

(a) preparing a first mixture comprising a poloxamer solution and an alginate solution, wherein the poloxamer solution comprises at least one poloxamer having a concentration of about 20% to about 30% (w/v) poloxamer, and wherein the alginate solution comprises at least one alginate having a concentration of about 0.1% (w/v) to about 5% (w/v) alginate;

(b) arranging the mixture in a repeating hexagonal pattern (/.e., honeycomb lattice) to provide an arranged mixture, optionally wherein the mixture is arranged in a repeating hexagon pattern via extrusion using 3D printer (e.g., bioprinting); and

(c) contacting the arranged mixture with an aqueous solution comprising at least one cationic salt.

Embodiment 19 provides the method of Embodiment 18, wherein the aqueous solution comprising at least one cationic salt is an aqueous solution of CaCh, optionally wherein the aqueous CaCh solution has a CaCh concentration of about 1.5% (w/v).

Embodiment 20 provides the method of Embodiment 18 or 19, wherein the contacting comprises immersion.

Embodiment 21 provides the method of any one of Embodiments 18-20, wherein the contacting occurs for a period of about 15 minutes.

Embodiment 22 provides the method of any one of Embodiments 18-21, further comprising contacting arranged mixture with a solution comprising at least one antibiotic agent.

Embodiment 23 provides the method of Embodiment 22, wherein the antibiotic agent is at least one selected from the group consisting of neomycin, polymyxin B, bacitracin, gentamicin, silver, clindamycin, erythromycin, tetracycline, metronidazole, rifampin, ciprofloxacin, ofloxacin, levofloxacin, doxycycline, silver sulfadiazine, mafenide acetate, fusidic acid, nadifloxacin, retapamulin, ozenoxacin, and mupirocin.

Embodiment 24 provides the method of Embodiment 23, wherein the antibiotic agent is neomycin.

Embodiment 25 provides the method of Embodiment 24, wherein the neomycin is neomycin sulfate.

Embodiment 26 provides the method of any one of Embodiments 18-25, wherein the solution comprising the at least one antibiotic agent has a concentration of the antibiotic agent of about 3% (w/v).

Embodiment 27 provides the method of any one of Embodiments 18-26, wherein the contacting of the arranged mixture with a solution comprising at least one antibiotic agent occurs for a period of 15 minutes.

Embodiment 28 provides the method of any one of Embodiments 18-27, wherein the method further comprises drying the wound dressing, optionally wherein the wound dressing is dried by air-exposure and/or freeze-drying.

Embodiment 29 provides a wound dressing prepared according to the method of Embodiment 28.

Embodiment 30 provides a method of treating or ameliorating a wound of a subject, the method comprising applying to the wound the wound dressing of any one of Embodiments 1-17 and 29.

Embodiment 31 provides the method of Embodiment 30, wherein the wound is at least one selected from the group consisting of a bum, ulcer, laceration, incision, abrasion, avulsion, sting, bite, and puncture.

Embodiment 32 provides the method of Embodiment 31, wherein the wound is a bum, optionally wherein the bum is a chemical bum, electrical bum, or radiation bum.

Embodiment 33 provides the method of Embodiment 32, wherein the wound is an ulcer, optionally wherein the ulcer is a diabetic ulcer, venous ulcer, arterial ulcer, or pressure ulcer.

Embodiment 34 provides the method of any one of Embodiments 30-33, wherein the wound dressing is applied and/or removed from the wound with minimal pain.

Embodiment 35 provides the method of any one of Embodiments 30-34, wherein the wound dressing does not impede formation of new vasculature, optionally wherein the wound dressing promotes vasculature growth.

The terms and expressions employed herein are used as terms of description and not of limitation, and there is no intention in the use of such terms and expressions of excluding any equivalents of the features shown and described or portions thereof, but it is recognized that various modifications are possible within the scope of the embodiments of the present application. Thus, it should be understood that although the present application describes specific embodiments and optional features, modification and variation of the compositions, methods, and concepts herein disclosed may be resorted to by those of or inary skill in the art, and that such modifications and variations are considered to be within the scope of embodiments of the present application.

Claims

CLAIMS What is claimed is:

1. A wound dressing comprising a hydrogel composition comprising:

(a) at least one poloxamer; and

(b) at least one cross-linked alginate, wherein the least one poloxamer and the at least one cross-linked alginate have a mass ratio ranging from about 40: 1 (w/w) to about 70: 1 (w/w) in the hydrogel composition; and wherein the hydrogel composition is arranged in a repeating hexagonal pattern (z.e., honeycomb lattice).

2. The wound dressing of claim 1, wherein the at least one poloxamer is a compound of formula (I):

3. The wound dressing of claim 1 or 2, wherein the at least one poloxamer is selected from the group consisting of P407, P188, P288, P335, and P338.

4. The wound dressing of claim 2 or 3, wherein x and z are each 101, and wherein y is 56 (i.e. P407).

5. The wound dressing of any one of claims 1-4. wherein the at least one poloxamer comprises about 15% (w/v) to about 30% (w/v) of the hydrogel composition, optionally wherein the at least one poloxamer comprises about 22% (w/v) of the hydrogel composition.

6. The wound dressing of any one of claims 1-5, wherein the at least one cross-linked alginate comprises about 0.1% (w/v) to about 1.0% (w/v) of the hydrogel composition, optionally wherein the at least one cross-linked alginate comprises about 0.4% (w/v) of the hydrogel composition.

7. The wound dressing of any one of claims 1-6, wherein the at least one poloxamer and the cross-linked alginate have a mass ratio of about 54: 1 (w/w) in the hydrogel composition.

8. The wound dressing of any one of claims 1-7, wherein the at least one alginate is cross-linked with an aqueous solution of CaCh. optionally wherein the aqueous solution has a CaCh concentration of about 1.5% (w/v).

9. The wound dressing of any one of claims 1-8, wherein the hydrogel composition further comprises at least one adsorbed and/or absorbed antibiotic agent.

10. The wound dressing of claim 9, wherein the antibiotic agent is at least one selected from the group consisting of neomycin, polymyxin B, bacitracin, gentamicin, silver, clindamycin, erythromycin, tetracycline, metronidazole, rifampin, ciprofloxacin, ofloxacin, levofloxacin, doxycycline, silver sulfadiazine, mafenide acetate, fusidic acid, nadifloxacin. retapamulin, ozenoxacin, and mupirocin.

11. The wound dressing of claim 9 or 10, wherein the at least one antibiotic agent is neomycin.

12. The wound dressing of claim 11, wherein the neomycin is adsorbed and/or absorbed by contacting the hydrogel composition with an aqueous solution of neomy cin sulfate, optionally wherein the neomycin sulfate solution has a concentration of about 3% (w/v), and optionally wherein the contacting occurs for a period of about 15 minutes.

13. The wound dressing of any one of claims 1-12, wherein the repeating hexagonal pattern (z.e., honeycomb lattice) comprises a continuous layer comprising the hydrogel composition, wherein continuous layer is characterized by evenly distributed hexagonal pores.

14. The wound dressing of claim 13. wherein the continuous layer has a thickness of about 1 to about 5 mm, optionally the continuous layer has a thickness of about 2.5 mm.

15. The wound dressing of claim 13 or 14. wherein the hexagonal pores are evenly distributed at a distance of about 1 to about 5 mm. optionally wherein the hexagonal pores are evenly distributed at a distance of about 2.5 mm.

16. The wound dressing of any one of claims 13-15, wherein parallel sides of each hexagonal pore have a distance of about 1 to about 10 mm, optionally wherein parallel sides of each hexagonal pore have a distance of about 5 mm.

17. The wound dressing of any one of claims 1-16, wherein the hydrogel composition is arranged in a repeating hexagonal pattern (z.e., honeycomb lattice) by 3D printing.

18. A method of preparing the wound dressing of any one of claims 1-17, the method comprising:

(b) arranging the mixture in a repeating hexagonal pattern (i.e., honeycomb lattice) to provide an arranged mixture, optionally wherein the mixture is arranged in a repeating hexagon pattern via extrusion using 3D printer (e.g, bioprinting); and

19. The method of claim 18, wherein the aqueous solution comprising at least one cationic salt is an aqueous solution of CaCb. optionally wherein the aqueous CaCh solution has a CaCh concentration of about 1.5% (w/v).

20. The method of claim 18 or 19, wherein the contacting comprises immersion.

21. The method of any one of claims 18-20, wherein the contacting occurs for a period of about 15 minutes.

22. The method of any one of claims 18-21, further comprising contacting arranged mixture with a solution comprising at least one antibiotic agent.

23. The method of claim 22, wherein the antibiotic agent is at least one selected from the group consisting of neomycin, polymyxin B, bacitracin, gentamicin, silver, clindamycin, erythromycin, tetracycline, metronidazole, rifampin, ciprofloxacin, ofloxacin, levofloxacin, doxycycline, silver sulfadiazine, mafenide acetate, fusidic acid, nadifloxacin, retapamulin, ozenoxacin, and mupirocin.

24. The method of claim 23, wherein the antibiotic agent is neomycin.

25. The method of claim 24, wherein the neomycin is neomycin sulfate.

26. The method of any one of claims 18-25. wherein the solution comprising the at least one antibiotic agent has a concentration of the antibiotic agent of about 3% (w/v).

27. The method of any one of claims 18-26, wherein the contacting of the arranged mixture with a solution comprising at least one antibiotic agent occurs for a period of 15 minutes.

28. The method of any one of claims 18-27, wherein the method further comprises drying the wound dressing, optionally wherein the wound dressing is dried by air-exposure and/or freeze-drying.

29. A wound dressing prepared according to the method of claim 28.

30. A method of treating or ameliorating a wound of a subject, the method comprising applying to the wound the wound dressing of any one of claims 1-17 and 29.

31. The method of claim 30, wherein the w ound is at least one selected from the group consisting of a bum, ulcer, laceration, incision, abrasion, avulsion, sting, bite, and puncture.

32. The method of claim 31, wherein the wound is a bum, optionally wherein the bum is a chemical bum, electrical bum, or radiation bum.

33. The method of claim 32, wherein the wound is an ulcer, optionally wherein the ulcer is a diabetic ulcer, venous ulcer, arterial ulcer, or pressure ulcer.

34. The method of any one of claims 30-33. wherein the wound dressing is applied and/or removed from the wound with minimal pain.

35. The method of any one of claims 30-34, wherein the wound dressing does not impede formation of new vasculature, optionally wherein the wound dressing promotes vasculature growth.