Gelma polymer composition comprising cells

Abstract

The present disclosure provides improved polymer compositions, such as GelMA polymer compositions. In certain embodiments, the improved polymer compositions may be used to deliver one or more therapeutic agents, such as cells, to a target treatment area, such as the eye of a subject. In certain embodiments, the improved polymer composition is a hydrogel comprising a methacrylated gelatin (i.e., GelMA) or a polymeric cross-linked derivative thereof.

Description

[0001] Related Applications

[0002] This application claims priority to U.S. Provisional Patent Application No. 63 / 332,963, filed on April 20, 2022, and U.S. Provisional Patent Application No. 63 / 410,041, filed on September 26, 2022. The entire contents of these applications are incorporated herein by reference. Technical Field

[0003] The present disclosure provides improved polymer compositions, such as GelMA polymer compositions. In certain embodiments, the improved polymer compositions can be used for delivering one or more therapeutic agents such as cells to a target treatment area, such as an eye of a subject. In certain embodiments, the improved polymer compositions are hydrogels comprising methacryloyl gelatin (i.e., GelMA) or a polymerized cross-linked derivative thereof.

[0004] introduction

[0005] GelMA polymer compositions have become effective materials for sealing, repairing and / or treating injuries, defects or diseases in the soft tissue of a subject. The design and production of improved GelMA polymer compositions for this purpose is an active area of ​​research.

[0006] There remains a need for improved GelMA polymer compositions, methods of producing GelMA polymer compositions, and therapeutic uses of GelMA polymer compositions. Summary of the invention

[0007] The details of various embodiments of the disclosure are set forth in the description below.

[0008] In certain embodiments, the present disclosure provides a polymer composition comprising at least one chemically modified gelatin. In certain embodiments, the polymer composition comprises at least one acrylated gelatin. In certain embodiments, the polymer composition comprises at least one methacrylated gelatin (GelMA). In certain embodiments, the polymer composition comprises about 0.5% to about 5.0% w / v of chemically modified gelatin. In certain embodiments, the polymer composition comprises about 0.5% to about 5.0% w / v of acrylated gelatin. In certain embodiments, the polymer composition comprises about 0.5% to about 5.0% w / v of GelMA.

[0009] In certain embodiments, the polymer composition comprises at least one chemically modified gelatin (optionally acrylated gelatin, such as GelMA) and at least one polymer cross-linking initiator (e.g., a photoinitiator). In certain embodiments, the polymer composition comprises: (i) at least one chemically modified gelatin (optionally acrylated gelatin); (ii) at least one polymer cross-linking initiator; and (iii) at least one therapeutic agent (e.g., a cell). In certain embodiments, the polymer composition comprises: (i) at least one chemically modified gelatin (optionally methacrylated gelatin, such as GelMA); (ii) at least one polymer cross-linking initiator; and (iii) at least one cell. In certain embodiments, the polymer composition is a precursor polymer composition. In certain embodiments, the polymer composition is a gel polymer composition.

[0010] In certain embodiments, the polymer composition comprises at least one crosslinking initiator. In certain embodiments, the crosslinking initiator comprises one or more photoactivated photoinitiators, optionally one or more photoinitiators activated by visible light. In certain embodiments, the polymer crosslinking initiator comprises Eosin Y, N-vinyl caprolactam, triethanolamine, or any combination thereof. In certain embodiments, the polymer crosslinking initiator comprises Eosin Y disodium salt (EYDS), N-vinyl caprolactam (NVC), triethanolamine, or any combination thereof. In certain embodiments, the polymer crosslinking initiator comprises Eosin Y disodium salt (EYDS), N-vinyl pyrrolidone (NVP), triethanolamine, or any combination thereof. In certain embodiments, the polymer crosslinking initiator comprises: (i) about 50 μM of Eosin Y or Eosin Y disodium salt (EYDS); (ii) about 3.5 to about 5.0 μL / mL of N-vinyl caprolactam (NVC) or N-vinyl pyrrolidone (NVP); and (iii) triethanolamine. In certain embodiments, the polymer cross-linking initiator comprises: (i) about 50 μM eosin Y disodium salt (EYDS); (ii) about 5.0 μL / mL N-vinyl pyrrolidone (NVP); and (iii) about 1.5% v / v triethanolamine.

[0011] In certain embodiments, the chemically modified gelatin is acrylated gelatin. In certain embodiments, the acrylated gelatin has a degree of acrylate of about 5-40%. In certain embodiments, the acrylated gelatin has a degree of acrylate of 5-20%. In certain embodiments, the acrylated gelatin has a degree of acrylate of about 5%, about 10%, or about 15%.

[0012] In certain embodiments, the chemically modified gelatin is methacrylated gelatin (GelMA). In certain embodiments, the GelMA has a degree of methacrylate of about 5-40%. In certain embodiments, the GelMA has a degree of methacrylate of 5-20%. In certain embodiments, the GelMA has a degree of methacrylate of about 5%, about 10%, or about 15%.

[0013] In certain embodiments, the polymer composition comprises about 2% to about 5% w / v of chemically modified gelatin (e.g., GelMA). In certain embodiments, the polymer composition comprises about 3% to about 5% w / v of chemically modified gelatin (e.g., GelMA). In certain embodiments, the polymer composition comprises about 3% to about 4% w / v of chemically modified gelatin (e.g., GelMA). In certain embodiments, the polymer composition comprises about 3.5% w / v of chemically modified gelatin (e.g., GelMA). In certain embodiments, the polymer composition comprises about 3% to about 4% w / v of GelMA. In certain embodiments, the polymer composition comprises about 3.5% w / v of GelMA.

[0014] In certain embodiments, the polymer composition comprises a combination of a first GelMA mixture and a second GelMA mixture. In certain embodiments, the polymer composition comprises a combination of a first GelMA mixture and a second GelMA mixture, wherein the total GelMA in the polymer composition is about 0.5% to about 5% w / v. In certain embodiments, the total GelMA in the polymer composition is about 2% to about 5% w / v. In certain embodiments, the total GelMA in the polymer composition is about 2% to about 4% w / v. In certain embodiments, the total GelMA in the polymer composition is about 3% to about 5% w / v. In certain embodiments, the total GelMA in the polymer composition is about 3% to about 4% w / v. In certain embodiments, the total GelMA in the polymer composition is about 3.5% w / v.

[0015] In certain embodiments, the polymer composition comprises about 0.5% to about 3% of the first GelMA mixture and about 0.5% to about 3% of the second GelMA mixture. In certain embodiments, the polymer composition comprises about 0.5% to about 1.5% of the first GelMA mixture and about 2% to about 3% of the second GelMA mixture. In certain embodiments, the polymer composition comprises about 1% of the first GelMA mixture and about 2.5% of the second GelMA mixture.

[0016] In certain embodiments, the first GelMA mixture includes a GelMA having a high average molecular weight and a low degree of methacrylate (DOM). In certain embodiments, the second GelMA mixture includes a GelMA having a low average molecular weight and a high degree of methacrylate (DOM). In certain embodiments, the first GelMA mixture includes a GelMA having a high average molecular weight and a low DOM; and the second GelMA mixture includes a GelMA having a low average molecular weight and a high DOM. In certain embodiments, the first GelMA mixture includes a GelMA having an average molecular weight of 140-180 kDa and a DOM of 5% to 40%; and the second GelMA mixture includes a GelMA having an average molecular weight of 75-115 kDa and a DOM of 50% to 80%. In certain embodiments, the first GelMA mixture includes a GelMA having an average molecular weight of 140-180 kDa and a DOM of 5% to 20%; and the second GelMA mixture includes a GelMA having an average molecular weight of 80-100 kDa and a DOM of 50% to 70%. In certain embodiments, the first GelMA mixture includes GelMA having an average molecular weight of about 160 kDa and a DOM of about 10%; and the second GelMA mixture includes GelMA having an average molecular weight of about 90 kDa and a DOM of about 60%.

[0017] In certain embodiments, the polymer composition comprises: about 0.5% to about 3% w / v of a first GelMA mixture comprising a GelMA having a high average molecular weight and a low DOM; and about 0.5% to about 3% of a second GelMA mixture comprising a GelMA having a low average molecular weight and a high DOM. In certain embodiments, the polymer composition comprises: about 0.5% to about 3% w / v of a first GelMA mixture comprising a GelMA having an average molecular weight of 140-180 kDa and a DOM of 5% to 40%; and about 0.5% to about 3% w / v of a second GelMA mixture comprising a GelMA having an average molecular weight of 75-115 kDa and a DOM of 50% to 80%. In certain embodiments, the polymer composition comprises: about 0.5% to about 1.5% w / v of a first GelMA mixture comprising a GelMA having an average molecular weight of 140-180 kDa and a DOM of 5% to 40%; and about 2% to about 3% w / v of a second GelMA mixture comprising a GelMA having an average molecular weight of 75-115 kDa and a DOM of 50% to 80%. In certain embodiments, the polymer composition comprises: about 0.5% to about 1.5% w / v of a first GelMA mixture comprising a GelMA having an average molecular weight of 140-180 kDa and a DOM of 5% to 20%; and about 2% to about 3% w / v of a second GelMA mixture comprising a GelMA having an average molecular weight of 85-100 kDa and a DOM of 50% to 70%.

[0018] In certain embodiments, the polymer composition comprises: about 1% w / v of a first GelMA mixture comprising GelMA having an average molecular weight of about 160 kDa and about 10% DOM; and about 2.5% w / v of a second GelMA mixture comprising GelMA having an average molecular weight of about 90 kDa and about 60% DOM.

[0019] In certain embodiments, at least one cell includes endothelial cells. In certain embodiments, at least one cell includes human umbilical vein endothelial cells (HUVEC). In certain embodiments, at least one cell includes epithelial cells. In certain embodiments, at least one cell includes human retinal pigment epithelial cells (HRPEC). In certain embodiments, at least one cell includes human retinal pigment epithelial cells (HRPEC) derived from induced pluripotent stem cells (iPSC). In certain embodiments, at least one cell includes eye cells, or eye cells derived from pluripotent or embryonic stem cells.

[0020] In certain embodiments, the polymer composition further comprises at least 0.1% (w / v) of a hydrophilic nonionic surfactant. In certain embodiments, the hydrophilic nonionic surfactant comprises at least one poloxamer surfactant, such as poloxamer 407. In certain embodiments, the composition comprises about 0.2% (w / v) of a poloxamer surfactant, such as poloxamer 407.

[0021] In certain embodiments, the present disclosure describes a precursor polymer composition comprising a polymer composition of the present disclosure. In certain embodiments, the present disclosure describes a gel polymer composition formed by photocrosslinking a precursor polymer composition of the present disclosure. In certain embodiments, the present disclosure describes a hydrogel polymer composition formed by photocrosslinking a precursor polymer composition of the present disclosure.

[0022] In certain embodiments, the present disclosure provides a method for treating and / or repairing a defect, injury and / or disease in a target soft tissue of a subject. In certain embodiments, the present disclosure provides a method for treating and / or repairing a defect, injury and / or disease in a target soft tissue of a subject, the method comprising: providing a precursor polymer composition of the present disclosure; applying the precursor polymer composition to the surface or below the surface of the target soft tissue of the subject, optionally to the location of the soft tissue defect, injury and / or disease; and crosslinking the precursor polymer composition by exposing the polymer crosslinking initiator in the polymer composition to crosslinking conditions, wherein the crosslinking of the precursor polymer composition produces a gel polymer composition.

[0023] In certain embodiments, the present disclosure provides a method for treating a defect, injury and / or disease in a target soft tissue of a subject, the method comprising: providing a gel polymer composition of the present disclosure; and applying the gel polymer composition to the surface, subsurface or near the surface of the target soft tissue of the subject. In certain embodiments, the gel polymer composition is applied at the location of the soft tissue defect, injury and / or disease.

[0024] In certain embodiments, the target soft tissue is ocular tissue. In certain embodiments, the target soft tissue is subconjunctival ocular tissue or retinal ocular tissue. In certain embodiments, the polymer composition is applied to the surface or subsurface of ocular tissue by subconjunctival injection, subretinal injection or suprachoroidal injection.

[0025] In certain embodiments, the defect, injury and / or disease of the target soft tissue comprises an eye defect, injury and / or disease; optionally an eye ulcer, such as a corneal ulcer caused by infection, injury, perforation or other defect. In certain embodiments, the eye defect, injury and / or disease comprises a retinal degenerative disease. In certain embodiments, the eye defect, injury and / or disease comprises age-related macular degeneration (AMD). In certain embodiments, the eye defect, injury and / or disease comprises retinitis pigmentosa. BRIEF DESCRIPTION OF THE DRAWINGS

[0026] The foregoing and other objects, features and advantages will be apparent from the following description of certain embodiments of the present disclosure, as illustrated in the accompanying drawings. The drawings are not necessarily drawn to scale or to full length, emphasis instead being placed upon illustrating the principles of various embodiments of the present disclosure.

[0027] Figure 1A An example of a reaction in which gelatin is modified with methacrylic anhydride (MA) to form methacryloyl-substituted gelatin (GelMA) is provided. Figure 1B An example of a reaction in which hyaluronic acid is modified with glycidyl methacrylate to form methacrylated hyaluronic acid (MeHA) is provided. Figure 1C An example of a reaction in which poly(ethylene glycol) (PEG) is modified with acryloyl chloride to form poly(ethylene glycol) diacrylate (PEGDA) is provided.

[0028] Figure 2 A method 100 for producing a gel polymer composition of the present disclosure is provided.

[0029] Figure 3 An example of a series of reactions for producing a GelMA hydrogel polymer composition from a methacrylated gelatin polymer precursor using a photoinitiator component and light energy is provided.

[0030] Figure 4A and Figure 4B Results of a study on the correlation between the degree of crosslinking and photopolymerization time within the hydrogels of the present disclosure are presented. Figure 4A The degree of cross-linking (%) of the hydrogel containing only HAMA is shown; Figure 4B The [ME methyl group of lysine CH 2 group] ratio.

[0031] Figure 5A , Figure 5B , Figure 5C and Figure 5D Results of a study on the swelling ratios of hydrogels of the present disclosure with varying GelMA, HAMA, and PEGDA concentrations are presented. Figure 5A and Figure 5B Shown are the swelling ratio measurements of four hydrogel formulations of the present disclosure; Figure 5C The swelling ratio measurement results of four hydrogel formulations disclosed in the present invention under re-swelling conditions are shown; Figure 5D Swelling ratio measurements of seven GelMA, PEGDA, and GelMA+PEGDA hydrogel formulations of the present disclosure are shown.

[0032] Fig. 6A and Figure 6B Shown are the results of a study on the swelling ratios of hydrogels of the present disclosure prepared with active agents and having varying concentrations of GelMA, HAMA, and PEGDA. Fig. 6A Shown are the swelling ratio measurements of six hydrogel formulations of the present disclosure (with and without active agents); Figure 6B Shown are the swelling ratio measurements of six hydrogel formulations of the present disclosure (with and without active agent) under reswelling conditions.

[0033] Fig. 7A , Figure 7B , Figure 7C and Fig.7D Results of a study on the drug release properties of the hydrogels of the present disclosure with varying GelMA, HAMA, and PEGDA concentrations are presented. Fig. 7A The G4-H of the present disclosure is shown M 1-P1 and G4-H G The drug release properties of 3-P1 hydrogel formulations up to 10-13 days; Figure 7B and Figure 7C G4-H M 1-P1 up to 35 days ( Figure 7B ) and 65 days ( Figure 7C )’s long-term drug release properties; Fig.7D The G4-H of the present disclosure is shown M Drug release properties of 1-P1, G4-P1, and G7-P1 hydrogel formulations.

[0034] Fig. 8A and Figure 8B Results of a study on the effect of vacuum drying on the drug release properties of hydrogels of the present disclosure prepared with active agents and having varying concentrations of GelMA, HAMA, and PEGDA are presented. Fig. 8A G4-H of the present disclosure is shown in "wet" and "vacuum dried" forms. M Drug release properties of 1-P1 hydrogel formulation; Figure 8B The drug release properties of the G7-P1 and G4-P1 hydrogel formulations of the present disclosure in "wet" and "vacuum dried" forms are shown.

[0035] Fig. 9A and Fig. 9B Results of a study on the effects of hydrogel shape and hydration state on drug release properties of hydrogels of the present disclosure prepared with active agents and having varying concentrations of GelMA, HAMA, and PEGDA are presented. Fig. 9A G4-H of the present disclosure is shown in "stick" and "disc" forms (including wet, vacuum-dried and freeze-dried stick forms). M 1-Total drug release properties of P1 hydrogel formulation; Fig. 9B G4-H of the present disclosure is shown in "stick" and "disc" forms (including wet, vacuum-dried and freeze-dried stick forms). M 1-Percent drug release properties of P1 hydrogel formulations.

[0036] Fig.10 Results of a study on the correlation between the release properties of the GelMA+PEGDA hydrogels of the present disclosure and the degree of methacrylateization of the GelMA within the hydrogels are provided.

[0037] Fig.11A and Fig. 11B Results of cell aggregation and viability studies performed using certain embodiments of the disclosed hydrogels are presented. Fig.11A Calcein AM+-based imaging of certain hydrogel samples is provided; Fig. 11B Calcein AM+ percentage (alive / alive+dead) quantitative data for certain hydrogel samples are shown.

[0038] Fig. 12A , Fig. 12B , Fig. 12C , Fig.12D , Fig.12E , Fig.12F , Figure 12G , Fig.12H and Fig.12I Presented are GFP+ cell imaging results from transwell underside cell growth studies using human umbilical vein endothelial cells (HUVECs) and certain embodiments of the disclosed hydrogels.

[0039] Fig.13A , Fig. 13B , Fig. 13C , Fig.13D and Fig.13E Confocal imaging results of cell growth studies on the underside of permeable cell culture inserts using human umbilical vein endothelial cells (HUVECs) and certain embodiments of the disclosed hydrogels are presented.

[0040] Fig.14A , Fig. 14B , Fig. 14C , Fig.14D , Fig.14E , Fig.14F , Figure 14G , Fig.14H , Fig.14I and Fig.14J Presented are the results of GFP+ cell imaging on the underside of permeable cell culture inserts for cell growth studies using human umbilical vein endothelial cells (HUVECs) and certain embodiments of the disclosed hydrogels.

[0041] Fig.15A , Fig. 15B , Fig. 15C , Fig.15D , Fig.15E , Fig.15F and Figure 15G Results of Calcein AM+-based imaging of cell growth studies on the underside of permeable cell culture inserts using human retinal pigment epithelial cells (HRPECs) and certain embodiments of the disclosed hydrogels are presented.

[0042] Fig.16A , Fig. 16B and Fig. 16C Results of Calcein AM+-based imaging of cell growth studies on the apical side of permeable cell culture inserts using human retinal pigment epithelial cells (HRPECs) and certain embodiments of the disclosed hydrogels are presented.

[0043] Fig.17A Live cell Calcein AM imaging results are provided that correlate with in vitro retinal cell growth studies of the hydrogel cell delivery formulations of the present disclosure (-20 million cells / mL). Fig. 17B Measurements of fluid shear rates associated with extrusion of GelMA precursor polymer formulations through small diameter needles are provided. Fig. 17C Live cell Calcein AM imaging results are provided that correlate with in vitro retinal cell growth studies of the disclosed hydrogel cell delivery formulations (-1 million cells / mL). Fig.17D Live cell imaging results (anti-CD73 and anti-rhodopsin) associated with in vitro retinal cell growth studies of the hydrogel cell delivery formulations of the present disclosure are provided.

[0044] Fig.18A The results of studies on cell localization and migration of the hydrogel formulations of the present disclosure are provided. Fig.18B Study results of hydrogel degradation studies of the hydrogel formulations of the present disclosure are provided. Fig. 18C and Fig.18D Images showing retinal detachment associated with the hydrogel formulations of the present disclosure. Fig.18E Image of human RPE cells deposited onto a native porcine RPE layer.

[0045] Fig.19 The results of a GelMA propagator study of the hydrogel formulations of the present disclosure are provided.

[0046] Fig. 20 Normalized cell viability study results associated with the hydrogel formulations of the present disclosure are provided.

[0047] Fig.21 Results of cell growth studies related to the hydrogel formulations of the present disclosure are provided. DETAILED DESCRIPTION

[0048] I. Polymer Composition

[0049] General

[0050] The present disclosure provides a polymer composition (e.g., a GelMA polymer composition) having one or more advantages over current commercial use or compositions known in the art. In certain embodiments, relative to one or more compositions currently used in business or known in the art, the polymer composition has one or more of the following advantages: (i) lower cost; (ii) easier to produce; (iii) improved biocompatibility; (iv) faster and / or stronger cross-linking and stabilization; (v) easier and / or more stable application; (vi) stronger adhesion and / or retention to the target surface; (vii) degradation characteristics that can be engineered and adjusted; (viii) once applied, the surface is smooth; and / or (ix) higher cell viability or improved encapsulated cell delivery. In certain embodiments, the polymer composition of the present disclosure allows one or more therapeutic agents or cells to be controlled and continuously released over a period of time. Therefore, the polymer composition of the present disclosure presents significant improvements relative to current commercial use and compositions currently known in the art.

[0051] As used herein, the term "polymer composition" may refer to a precursor polymer composition (e.g., a polymer composition before cross-linking polymerization) and / or a gel polymer composition (e.g., a polymer composition after cross-linking polymerization), as provided in the corresponding context of the present disclosure. Examples of gel polymer compositions include hydrogels and polymer compositions having increased viscosity due to cross-linking polymerization in the polymer composition (e.g., soft gels).

[0052] In general, reference to a polymer component (e.g., GelMA, MeHA, PEGDA) in the present disclosure may refer to a polymer precursor component (e.g., a monomer or a precursor oligomer), a cross-linked form of the polymer component in an oligomer (e.g., a cross-linked oligomer), and / or a polymerized form of the polymer component in a gel polymer composition (e.g., a hydrogel polymer), depending on the context within the present disclosure.

[0053] In certain embodiments, the polymer composition comprises chemically modified gelatin, such as methacrylated gelatin (i.e., GelMA). In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA) and one or more cross-linking agents. In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA) and one or more polymer cross-linking initiators, such as photoactivated photoinitiator components. In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA), one or more cross-linking agents and one or more polymer cross-linking initiators, such as photoactivated photoinitiator components.

[0054] In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA) and chemically modified hyaluronic acid (e.g., MeHA). In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA), chemically modified hyaluronic acid (e.g., MeHA) and one or more cross-linking agents. In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA), chemically modified hyaluronic acid (e.g., MeHA) and one or more polymer cross-linking initiators, such as photoactivated photoinitiator ingredients. In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA), chemically modified hyaluronic acid (e.g., MeHA), one or more cross-linking agents and one or more polymer cross-linking initiators, such as photoactivated photoinitiator ingredients. In certain embodiments, the polymer composition comprises unmodified HA. In certain embodiments, the polymer composition comprises unmodified HA and chemically modified HA (e.g., MeHA).

[0055] In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA) and chemically modified poly (ethylene glycol) (PEG) (e.g., PEGDA). In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA), chemically modified PEG (e.g., PEGDA) and one or more cross-linking agents. In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA), chemically modified PEG (e.g., PEGDA) and one or more polymer cross-linking initiators, such as a photoactivated photoinitiator component. In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA), chemically modified PEG (e.g., PEGDA), one or more cross-linking agents and one or more polymer cross-linking initiators, such as a photoactivated photoinitiator component. In certain embodiments, the polymer composition comprises unmodified PEG. In certain embodiments, the polymer composition comprises unmodified PEG and chemically modified PEG (e.g., PEGDA).

[0056] In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA), chemically modified hyaluronic acid (e.g., MeHA), and chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA), chemically modified hyaluronic acid (e.g., MeHA), chemically modified PEG (e.g., PEGDA) and one or more cross-linking agents. In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA), chemically modified hyaluronic acid (e.g., MeHA), chemically modified PEG (e.g., PEGDA) and one or more polymer cross-linking initiators, such as photoactivated photoinitiator ingredients. In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA), chemically modified hyaluronic acid (e.g., MeHA), chemically modified PEG (e.g., PEGDA), one or more cross-linking agents and one or more polymer cross-linking initiators, such as photoactivated photoinitiator ingredients. In certain embodiments, the polymer composition comprises unmodified HA and / or unmodified PEG.

[0057] In certain embodiments, the polymer composition does not comprise a hydrolase. In certain embodiments, the polymer composition does not comprise a glycosidase hydrolase.

[0058] In certain embodiments, the gel polymer composition is a hydrogel. A hydrogel generally comprises a cross-linked polymer framework that encompasses a pore network filled with an interstitial solvent (e.g., fluid) including water. In certain embodiments, the water content of the hydrogel polymer composition is about 80% or more. In certain embodiments, the water content of the hydrogel polymer composition is greater than about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98% or greater than about 99%.

[0059] In certain embodiments, the polymer composition (e.g., hydrogel or hydrogel precursor) (e.g., GelMA polymer composition) of the present disclosure is a primary polymer composition and may include one of a plurality of secondary hydrogel-forming polymer components (i.e., polymer or its precursor) or in combination with one of a plurality of secondary hydrogel-forming polymer components (i.e., polymer or its precursor). In certain embodiments, the polymer composition (e.g., hydrogel or hydrogel precursor) of the present disclosure (e.g., GelMA polymer composition) is a primary polymer composition and may include one of a plurality of secondary hydrogel-forming polymer components selected from the following or in combination with one of a plurality of secondary hydrogel-forming polymer components selected from the following: acrylamide, acrylic acid, alginate, alginate methacrylate, cellulose, chitosan, chitosan methacrylate, dimethylacrylamide, methylenebisacrylamide, fibronectin, alum, Glue, gelatin methacrylate, ethylene glycol chitosan, ethylene glycol chitosan methacrylate, hexyl methacrylate, hyaluronic acid, hyaluronic acid methacrylate, hydroxyethyl methacrylate, hydroxyethyl acrylate, isopropyl acrylamide, isopropyl methacrylamide, laminin, methacrylamide, methacrylic acid, polyamide, polycaprolactone, polyethylene glycol (PEG), polyethylene terephthalate, polylactic acid, polyurethane, polyvinyl alcohol, polyethylene oxide dimethacrylate, siloxane, polysiloxane or their oligomers, polymers and / or combinations. In certain embodiments, the secondary hydrogel polymer (formed by the secondary hydrogel-forming polymer precursor) is covalently crosslinked with the primary gel polymer composition (e.g., GelMA polymer composition). In certain embodiments, the secondary hydrogel polymer (formed from the secondary hydrogel-forming polymer precursor) is not covalently cross-linked with the primary gel polymer composition (e.g., the GelMA polymer composition), e.g., the secondary hydrogel polymer forms a polymer network that is interwoven with the polymer network of the primary gel polymer composition.

[0060] In certain embodiments, polymer compositions of the present disclosure include one or more biocompatible polymer components or polysaccharides. In certain embodiments, polymer compositions of the present disclosure include one or more biocompatible polymer components or polysaccharides selected from the following: agarose, alginate, pullulan, amylose, carrageenan, cellulose, chitin, chitosan, chondroitin sulfate, collagen, dermatan sulfate, dextran, elastin, elastin-like polypeptide (ELP), tropoelastin, fibrin, fibrinogen, fibronectin, gelatin, glycogen, heparan, heparan sulfate, heparin, heparan sulfate, hyaluronan, hyaluronic acid, keratan sulfate, laminin, pectin, polysebacic acid glyceride (PGS), polyethylene glycol (PEG), polylactic acid (PLA), polylysine, starch, thrombin, their derivatives or their combinations.

[0061] In certain embodiments, the polymer compositions of the present disclosure comprise one or more cell adhesion agents selected from the group consisting of fibronectin, laminin, vitronectin, RGD, vixapatin, derivatives thereof, or combinations thereof.

[0062] In certain embodiments, polymer compositions of the present disclosure include one or more synthetic polymer components, such as biocompatible synthetic polymer components. In certain embodiments, polymer compositions include one or more synthetic polymer components selected from the following: polyurethane, polysiloxane, silicone, polyethylene, polyvinyl pyrrolidone, polyhydroxyethyl methacrylate (poly-HEMA), polymethyl methacrylate, polyvinyl alcohol, polyacrylic acid, polyacrylamide, polyethylene-to-vinyl acetate, polyethylene glycol, polymethacrylic acid, polylactic acid, polyglycolic acid, polylactide-to-glycolide, nylon, polyamide, polyanhydride, polyethylene-to-vinyl alcohol, polycaprolactone, polyvinyl acetate, polyethylene hydroxide, polyethylene oxide, polyorthoester, polyallylamine, polyethyleneimine, polylysine, polyarginine, their derivatives or their combinations and / or copolymers.

[0063] In certain embodiments, the polymer composition of the present disclosure comprises one or more polymer components (e.g., monomers, precursors, polymers) comprising crosslinkable groups. In certain embodiments, the polymer composition of the present disclosure comprises one or more polymer components comprising crosslinkable groups selected from (or formed by reacting) anhydrides, acyl halides, carboxylic acids, glycols, acrylic anhydride, methacrylic anhydride, acryloyl chloride, acryloyl bromide, methacryloyl chloride, methacryloyl bromide, acrylic acid, glycidyl methacrylate, methacrylic acid, dopamine, derivatives thereof, or combinations thereof.

[0064] In certain embodiments, hydroxyethyl methacrylate (HEMA) or a polymer thereof may be present in the polymer composition at a concentration of about 1% to about 60% weight / volume (w / v).

[0065] In certain embodiments, polymer compositions of the present disclosure include one or more stabilizers and / or reinforcing agents. In certain embodiments, polymer compositions of the present disclosure include one or more stabilizers and / or reinforcing agents selected from the following: polar amino acids (e.g., tyrosine, cysteine, serine, threonine, asparagine, glutamine, aspartic acid, glutamic acid, arginine, lysine and histidine), amino acid analogs, amino acid derivatives, collagen, divalent cation chelators (e.g., ethylenediaminetetraacetic acid (EDTA) or its salt) or combinations thereof.

[0066] In certain embodiments, the polymer compositions of the present disclosure may be transparent and / or translucent. In certain embodiments, the polymer compositions may be partially translucent or partially opaque. In certain embodiments, the polymer compositions may be opaque.

[0067] In certain embodiments, the polymer compositions of the present disclosure may include a polymeric component or a therapeutic component and may be produced, analyzed, or used as disclosed in US20140377326, US20150274805, US20160175488, US 20170232138, US20190022280 A1, WO 2020051133, and WO 2020081673, to the extent that each patent describes the composition, production, analysis, and use of acrylated gelatin polymeric compositions such as GelMA hydrogels, each of which is incorporated herein by reference in its entirety.

[0068] preparation

[0069] In certain embodiments, the polymer composition of the present disclosure comprises chemically modified gelatin (e.g., GelMA), chemically modified hyaluronic acid (e.g., MeHA), chemically modified PEG (e.g., PEGDA); or any combination thereof. In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA). In certain embodiments, the polymer composition comprises chemically modified hyaluronic acid (e.g., MeHA). In certain embodiments, the polymer composition comprises chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA) and chemically modified hyaluronic acid (e.g., MeHA). In certain embodiments, the polymer composition comprises chemically modified gelatin (e.g., GelMA) and chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises chemically modified hyaluronic acid (e.g., MeHA) and chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises chemically modified gelatin (eg, GelMA), chemically modified hyaluronic acid (eg, MeHA), and chemically modified PEG (eg, PEGDA).

[0070] In certain embodiments, the polymer compositions of the present disclosure comprise a combination of precursor polymer components according to Table 1 (percentages are w / v concentrations in the total precursor polymer formulation). Unless otherwise specified, the GelMA materials in Table 1 are 160 / 80 (i.e., having a molecular weight (MW) of 160 kDa and a degree of methacrylate (DoM) of 80%). Unless otherwise specified, the HAMA materials in Table 1 are 500 / 30 (i.e., having a molecular weight (MW) of 500 kDa and a degree of methacrylate (DoM) of 30%). Unless otherwise specified, the PEGDA materials in Table 1 are formed from 35 kDa PEG materials. Poloxamer 407=Px 407.

[0071] Table 1 - Examples of Precursor Polymer Compositions

[0072]

[0073]

[0074]

[0075]

[0076] In certain embodiments, the polymer composition comprises about 4-20% w / v of chemically modified gelatin (e.g., GelMA); about 0-1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0-5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 4-10% w / v of chemically modified gelatin (e.g., GelMA); about 1-1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1-5% w / v of chemically modified PEG (e.g., PEGDA).

[0077] In certain embodiments, the polymer composition comprises a GelMA having a molecular weight (MW) of about 160 kDa. In certain embodiments, the polymer composition comprises a GelMA having a molecular weight (MW) of about 160 kDa. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 80% to about 90%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 85%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 80%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 75% to about 85%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 70% to about 80%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 75%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 70%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 65% to about 75%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 60% to about 70%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 65%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 60%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 60%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 55% to about 65%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 50% to about 60%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 55%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 50%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 45% to about 55%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 40% to about 50%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 45%. In certain embodiments, the polymer composition comprises a GelMA having a DoM of about 40%.In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 35% to about 45%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 30% to about 40%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 35%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 30%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 25% to about 35%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 20% to about 30%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 25%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 20%. In certain embodiments, the polymer composition comprises a GelMA having a DoM of about 20%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 15% to about 25%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate (DoM) of about 10% to about 20%. In certain embodiments, the polymer composition comprises a GelMA having a DoM of about 15%. In certain embodiments, the polymer composition comprises a GelMA having a DoM of about 10%. In certain embodiments, the polymer composition comprises a GelMA having a DoM of about 5%. In certain embodiments, the polymer composition comprises a GelMA having a DoM of about 10-40%. In certain embodiments, the polymer composition comprises a GelMA having a DoM of about 50-80%. In certain embodiments, the polymer composition comprises a GelMA having a DoM of about 10-20%. In certain embodiments, the polymer composition comprises a GelMA having a DoM of about 7%. In certain embodiments, the polymer composition comprises a GelMA having a DoM of about 5%. In certain embodiments, the polymer composition comprises GelMA having a DoM of about 5-40%. In certain embodiments, the polymer composition comprises GelMA having a DoM of about 5-20%. In certain embodiments, the polymer composition comprises GelMA having a DoM of about 1-10%.

[0078] In certain embodiments, the polymer composition comprises a combination of a first GelMA mixture and a second GelMA mixture. In certain embodiments, the polymer composition comprises a combination of a first GelMA mixture and a second GelMA mixture, wherein the total GelMA in the polymer composition is about 0.5% to about 5%. In certain embodiments, the polymer composition comprises a combination of a first GelMA mixture and a second GelMA mixture, wherein the total GelMA in the polymer composition is about 2% to about 5%. In certain embodiments, the polymer composition comprises a combination of a first GelMA mixture and a second GelMA mixture, wherein the total GelMA in the polymer composition is about 2% to about 4%. In certain embodiments, the polymer composition comprises a combination of a first GelMA mixture and a second GelMA mixture, wherein the total GelMA in the polymer composition is about 3% to about 5%. In certain embodiments, the polymer composition comprises a combination of a first GelMA mixture and a second GelMA mixture, wherein the total GelMA in the polymer composition is about 3% to about 4%.

[0079] In certain embodiments, the polymer composition comprises about 0.5% to about 3% of the first GelMA mixture. In certain embodiments, the polymer composition comprises about 0.5% to about 3% of the second GelMA mixture. In certain embodiments, the polymer composition comprises about 0.5% to about 3% of the first GelMA mixture and about 0.5% to about 3% of the second GelMA mixture. In certain embodiments, the polymer composition comprises about 0.5% to about 1.5% of the first GelMA mixture and about 2% to about 3% of the second GelMA mixture. In certain embodiments, the polymer composition comprises about 2% to about 3% of the first GelMA mixture and about 0.5% to about 1.5% of the second GelMA mixture. In certain embodiments, the polymer composition comprises about 1% of the first GelMA mixture and about 2.5% of the second GelMA mixture.

[0080] In certain embodiments, the first GelMA mixture includes a GelMA having a high average molecular weight (e.g., 140-180 kDa, or about 160 kDa). In certain embodiments, the first GelMA mixture includes a GelMA having a low degree of methacrylate (DOM) (e.g., 5% to 40%, or about 10%). In certain embodiments, the first GelMA mixture includes a GelMA having a high average molecular weight (e.g., 140-180 kDa, or about 160 kDa) and a low (DOM) (e.g., 5% to 40%, or about 10%). In certain embodiments, the first GelMA mixture includes a GelMA having an average molecular weight of 140-180 kDa and a DOM of 5% to 40%. In certain embodiments, the first GelMA mixture includes a GelMA having an average molecular weight of 140-180 kDa and a DOM of 5% to 20%. In certain embodiments, the first GelMA mixture includes a GelMA having an average molecular weight of about 160 kDa and a DOM of about 10%.

[0081] In certain embodiments, the second GelMA mixture includes a GelMA with a low average molecular weight (e.g., 75-115 kDa, or about 90 kDa). In certain embodiments, the second GelMA mixture includes a GelMA with a high degree of methacrylate (DOM) (e.g., 50% to 80%, or about 60%). In certain embodiments, the second GelMA mixture includes a GelMA with a low average molecular weight (e.g., 75-115 kDa, or about 90 kDa) and a high (DOM) (e.g., 50% to 80%, or about 60%). In certain embodiments, the second GelMA mixture includes a GelMA with an average molecular weight of 75-115 kDa and a DOM of 50% to 80%. In certain embodiments, the second GelMA mixture includes a GelMA with an average molecular weight of 80-115 kDa and a DOM of 50% to 70%. In certain embodiments, the second GelMA mixture includes a GelMA with an average molecular weight of about 90 kDa and a DOM of about 60%.

[0082] In certain embodiments, the first GelMA mixture includes a GelMA having a high average molecular weight (e.g., 140-180 kDa, or about 160 kDa) and a low (DOM) (e.g., 5% to 40%, or about 10%); and the second GelMA mixture includes a GelMA having a low average molecular weight (e.g., 75-115 kDa, or about 90 kDa) and a high (DOM) (e.g., 50% to 80%, or about 60%). In certain embodiments, the first GelMA mixture includes a GelMA having an average molecular weight of 140-180 kDa and a DOM of 5% to 40%; and the second GelMA mixture includes a GelMA having an average molecular weight of 75-115 kDa and a DOM of 50% to 80%. In some embodiments, the first GelMA mixture comprises a GelMA having an average molecular weight of 140-180 kDa and a DOM of 5% to 20%; and the second GelMA mixture comprises a GelMA having an average molecular weight of 80-100 kDa and a DOM of 50% to 70%. In some embodiments, the first GelMA mixture comprises a GelMA having an average molecular weight of about 160 kDa and a DOM of about 10%; and the second GelMA mixture comprises a GelMA having an average molecular weight of about 90 kDa and a DOM of about 60%.

[0083] In certain embodiments, the polymer composition comprises about 0.5% to about 3% w / v of a first GelMA mixture comprising a GelMA having a high average molecular weight (e.g., 140-180 kDa, or about 160 kDa) and a low (DOM) (e.g., 5% to 40%, or about 10%). In certain embodiments, the polymer composition comprises about 0.5% to about 3% w / v of a second GelMA mixture comprising a GelMA having a low average molecular weight (e.g., 75-115 kDa, or about 90 kDa) and a high (DOM) (e.g., 50% to 80%, or about 60%). In certain embodiments, the polymer composition comprises: about 0.5% to about 3% w / v of a first GelMA mixture comprising a GelMA having a high average molecular weight (e.g., 140-180 kDa, or about 160 kDa) and a low (DOM) (e.g., 5% to 40%, or about 10%); and about 0.5% to about 3% w / v of a second GelMA mixture comprising a GelMA having a low average molecular weight (e.g., 75-115 kDa, or about 90 kDa) and a high (DOM) (e.g., 50% to 80%, or about 60%).

[0084] In certain embodiments, the polymer composition comprises: about 0.5% to about 1.5% w / v of a first GelMA mixture comprising a GelMA having an average molecular weight of 140-180 kDa and a DOM of 5% to 40%; and about 2% to about 3% w / v of a second GelMA mixture comprising a GelMA having an average molecular weight of 75-115 kDa and a DOM of 50% to 80%. In certain embodiments, the polymer composition comprises: about 0.5% to about 1.5% w / v of a first GelMA mixture comprising a GelMA having an average molecular weight of 140-180 kDa and a DOM of 5% to 20%; and about 2% to about 3% w / v of a second GelMA mixture comprising a GelMA having an average molecular weight of 85-100 kDa and a DOM of 50% to 70%.

[0085] In certain embodiments, the polymer composition comprises: about 1% w / v of a first GelMA mixture comprising GelMA having an average molecular weight of about 160 kDa and about 10% DOM; and about 2.5% w / v of a second GelMA mixture comprising GelMA having an average molecular weight of about 90 kDa and about 60% DOM.

[0086] In certain embodiments, the polymer composition comprises a combination of a GelMA having a DoM of about 10% (e.g., GelMA 160 / 10) and a GelMA having a DoM of about 80% (e.g., GelMA 160 / 80). In certain embodiments, the polymer composition comprises a combination of a GelMA having a DoM of about 10% and a GelMA having a DoM of about 80%, wherein the polymer composition comprises about 4-10% w / v GelMA. In certain embodiments, the polymer composition comprises a combination of a GelMA having a DoM of about 10% (e.g., GelMA 160 / 10) and a GelMA having a DoM of about 80% (e.g., GelMA 160 / 80), wherein the ratio of 10% DoM GelMA to 80% DoM GelMA is about 1:9 to about 9:1. In certain embodiments, the ratio of 10% DoM GelMA to 80% DoM GelMA is about 1:9. In certain embodiments, the ratio of 10% DoM GelMA to 80% DoM GelMA is about 2:8 (i.e., about 1:4). In certain embodiments, the ratio of 10% DoM GelMA to 80% DoM GelMA is about 3:7. In certain embodiments, the ratio of 10% DoM GelMA to 80% DoM GelMA is about 4:6 (i.e., about 2:3). In certain embodiments, the ratio of 10% DoM GelMA to 80% DoM GelMA is about 5:5 (i.e., about 1:1). In certain embodiments, the ratio of 10% DoM GelMA to 80% DoM GelMA is about 4:6 (i.e., about 2:3). In certain embodiments, the ratio of 10% DoM GelMA to 80% DoM GelMA is about 3:7. In certain embodiments, the ratio of 10% DoM GelMA to 80% DoM GelMA is about 2:8 (i.e., about 1:4). In certain embodiments, the ratio of 10% DoM GelMA to 80% DoM GelMA is about 1:9.

[0087] In certain embodiments, the polymer composition comprises a glycidyl methacrylate functionalized GelMA having a DoM of about 45% (e.g., GelMA 160 / 45). In certain embodiments, the polymer composition comprises a glycidyl methacrylate functionalized GelMA having a DoM of about 45% and 160 kDa (GelMA 160 / 45). In certain embodiments, the polymer composition comprises a glycidyl methacrylate functionalized GelMA having a DoM of about 45% and 90 kDa (GelMA 90 / 45).

[0088] In certain embodiments, the polymer composition comprises MeHA having a molecular weight (MW) of about 500 kDa. In certain embodiments, the polymer composition comprises MeHA having a degree of methacrylate (DoM) of about 30%. In certain embodiments, the polymer composition comprises PEGDA formed from a PEG material of about 35 kDa. In certain embodiments, the polymer composition comprises PEGDA formed from a PEG material of about 2 kDa.

[0089] In certain embodiments, the polymer composition comprises a poloxamer surfactant (e.g., poloxamer 407). In certain embodiments, the polymer composition comprises about 0.1-0.5% w / v (e.g., about 0.2% w / v) of a poloxamer surfactant (e.g., poloxamer 407). In certain embodiments, the polymer composition comprises a tyloxapol surfactant. In certain embodiments, the polymer composition comprises about 0.1-0.5% w / v (e.g., about 0.1% w / v) of a tyloxapol surfactant.

[0090] In certain embodiments, the polymer composition comprises about 4% w / v of chemically modified gelatin (e.g., GelMA). In certain embodiments, the polymer composition comprises about 4% w / v of chemically modified gelatin (e.g., GelMA); about 1-1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1-5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 4% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1-5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 4% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 4% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 4% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.67% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 4% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 1.0% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 4% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1-5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 4% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 4% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.5% w / v of chemically modified PEG (e.g., PEGDA).In certain embodiments, the polymer composition comprises about 4% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.67% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 4% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 1.0% w / v of chemically modified PEG (e.g., PEGDA).

[0091] In certain embodiments, the polymer composition comprises about 5% w / v of chemically modified gelatin (e.g., GelMA). In certain embodiments, the polymer composition comprises about 5% w / v of chemically modified gelatin (e.g., GelMA); about 1-1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1-5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 5% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1-5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 5% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 5% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 5% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.67% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 5% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 1.0% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 5% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1-5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 5% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 5% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.5% w / v of chemically modified PEG (e.g., PEGDA).In certain embodiments, the polymer composition comprises about 5% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.67% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 5% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 1.0% w / v of chemically modified PEG (e.g., PEGDA).

[0092] In certain embodiments, the polymer composition comprises about 10% w / v of chemically modified gelatin (e.g., GelMA). In certain embodiments, the polymer composition comprises about 10% w / v of chemically modified gelatin (e.g., GelMA); about 1-1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1-5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 10% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1-5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 10% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 10% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 10% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.67% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 10% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 1.0% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 10% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1-5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 10% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 10% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.5% w / v of chemically modified PEG (e.g., PEGDA).In certain embodiments, the polymer composition comprises about 10% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.67% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 10% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 1.0% w / v of chemically modified PEG (e.g., PEGDA).

[0093] In certain embodiments, the polymer composition comprises about 20% w / v of chemically modified gelatin (e.g., GelMA). In certain embodiments, the polymer composition comprises about 20% w / v of chemically modified gelatin (e.g., GelMA); about 1-1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1-5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 20% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1-5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 20% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 20% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 20% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.67% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 20% w / v of chemically modified gelatin (e.g., GelMA); about 1% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 1.0% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 20% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1-5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 20% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.1% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 20% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.5% w / v of chemically modified PEG (e.g., PEGDA).In certain embodiments, the polymer composition comprises about 20% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 0.67% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 20% w / v of chemically modified gelatin (e.g., GelMA); about 1.5% w / v of chemically modified hyaluronic acid (e.g., MeHA); and about 1.0% w / v of chemically modified PEG (e.g., PEGDA).

[0094] In certain embodiments, the polymer composition comprises about 4-20% w / v of chemically modified gelatin (e.g., GelMA); and about 0-5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 4-10% w / v of chemically modified gelatin (e.g., GelMA); and about 0.1-5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 4% w / v of chemically modified gelatin (e.g., GelMA); and about 0.1-5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 4% w / v of chemically modified gelatin (e.g., GelMA); and about 0.1% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 4% w / v of chemically modified gelatin (e.g., GelMA); and about 0.5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 4% w / v of chemically modified gelatin (e.g., GelMA); and about 0.67% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 4% w / v of chemically modified gelatin (e.g., GelMA); and about 1.0% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 5% w / v of chemically modified gelatin (e.g., GelMA); and about 0.1% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 5% w / v of chemically modified gelatin (e.g., GelMA); and about 0.5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 5% w / v of chemically modified gelatin (e.g., GelMA); and about 0.67% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 5% w / v of chemically modified gelatin (e.g., GelMA); and about 1.0% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 7% w / v of chemically modified gelatin (e.g., GelMA); and about 0.1% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 7% w / v of chemically modified gelatin (e.g., GelMA); and about 0.5% w / v of chemically modified PEG (e.g., PEGDA).In certain embodiments, the polymer composition comprises about 7% w / v of chemically modified gelatin (e.g., GelMA); and about 0.67% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 7% w / v of chemically modified gelatin (e.g., GelMA); and about 1.0% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 10% w / v of chemically modified gelatin (e.g., GelMA); and about 0.1% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 10% w / v of chemically modified gelatin (e.g., GelMA); and about 0.5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 10% w / v of chemically modified gelatin (e.g., GelMA); and about 0.67% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 10% w / v of chemically modified gelatin (e.g., GelMA); and about 1.0% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 20% w / v of chemically modified gelatin (e.g., GelMA); and about 0.1% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 20% w / v of chemically modified gelatin (e.g., GelMA); and about 0.5% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 20% w / v of chemically modified gelatin (e.g., GelMA); and about 0.67% w / v of chemically modified PEG (e.g., PEGDA). In certain embodiments, the polymer composition comprises about 20% w / v chemically modified gelatin (eg, GelMA); and about 1.0% w / v chemically modified PEG (eg, PEGDA).

[0095] In certain embodiments, the polymer composition comprises: about 4% GelMA (10-40% DoM); and about 1% PEGDA (35 kDa). In certain embodiments, the polymer composition comprises: about 2% GelMA (10-40% DoM); about 2% gelatin; and about 1% PEGDA (35 kDa). In certain embodiments, the polymer composition comprises: about 4% gelatin acrylate (10-40% DoM); and about 1% PEGDA (35 kDa). In certain embodiments, the polymer composition comprises: about 2% gelatin acrylate (10-40% DoM); about 2% gelatin; and about 1% PEGDA (35 kDa). In certain embodiments, the polymer composition comprises: about 4% GelMA (10-40% DoM); about 1% PEGDA (35 kDa); and about 1-20% PEG methyl ether acrylate (35 kDa). In certain embodiments, the polymer composition comprises: about 4% GelMA (10-40% DoM); about 1% HAMA (500 kDa, 5-40% DoM); and about 1% PEGDA (35 kDa). In certain embodiments, the polymer composition comprises: about 2% GelMA (10-40% DoM); about 2% gelatin; about 1% HAMA (500 kDa, 5-40% DoM); and about 1% PEGDA (35 kDa). In certain embodiments, the polymer composition comprises: about 4% gelatin acrylate (10-40% DoM); about 1% HAMA (500 kDa, 5-40% DoM); and about 1% PEGDA (35 kDa). In certain embodiments, the polymer composition comprises: about 2% gelatin acrylate (10-40% DoM); about 2% gelatin; about 1% HAMA (500 kDa, 5-40% DoM); and about 1% PEGDA (35 kDa). In certain embodiments, the polymer composition comprises: about 4% GelMA (10-40% DoM); about 1% HAMA (500 kDa, 5-40% DoM); about 1% PEGDA (35 kDa); and about 1-20% PEG methyl ether acrylate (35 kDa). In certain embodiments, the polymer composition comprises: about 5-20% GelMA (10-40% DoM). In certain embodiments, the polymer composition comprises: about 5-20% GelMA (10-40% DoM); and about 1% HAMA (500 kDa, 5-40% DoM).

[0096] In certain embodiments, the polymer composition comprises: about 4% GelMA (80% DoM); about 1% PEGDA (2 kDa); and about 0.2% (w / v) of a poloxamer surfactant (e.g., poloxamer 407); optionally, an active agent (e.g., a corticosteroid). In certain embodiments, the polymer composition comprises: about 4% GelMA (40% DoM); about 1% PEGDA (35 kDa); and about 0.2% (w / v) of a poloxamer surfactant (e.g., poloxamer 407); optionally, an active agent (e.g., a corticosteroid). In certain embodiments, the polymer composition comprises: about 4% GelMA (10% DoM); about 1% PEGDA (35 kDa); and about 0.2% (w / v) of a poloxamer surfactant (e.g., poloxamer 407); optionally, an active agent (e.g., a corticosteroid). In certain embodiments, the polymer composition comprises: about 20% GelMA (40% DoM); and about 0.2% (w / v) poloxamer surfactant (eg, poloxamer 407); and optionally an active agent (eg, a corticosteroid).

[0097] Chemically modified gelatin

[0098] Gelatin is a biocompatible mixture of natural sources of peptides and proteins derived from collagen, and collagen is the main structural component of animal tissue (including eye tissue, bone and skin). Natural matrix peptides and proteins (e.g., denatured collagen) that can be used to produce gelatin materials of the present disclosure may include gelatin components derived from animals, and the animals include but are not limited to pigs, cattle, horses, chickens and fish. In certain embodiments, gelatin materials may be derived from connective tissue proteins, such as collagen. In certain embodiments, gelatin materials may be derived from bone, skin or eye tissue. In certain embodiments, gelatin materials may be prepared by acid hydrolysis and / or alkaline hydrolysis of connective tissue proteins (e.g., collagen).

[0099] In certain embodiments, the polymer composition of the present disclosure comprises chemically modified gelatin. In certain embodiments, the polymer composition comprises acrylated gelatin. In certain embodiments, the polymer composition comprises methacrylated gelatin (i.e., GelMA). In certain embodiments, chemically modified gelatin may be included in the precursor polymer composition of the present disclosure. In certain embodiments, chemically modified gelatin comprises a photocrosslinkable derivative of gelatin. In certain embodiments, chemically modified gelatin can be modified with acrylic anhydride or acryloyl chloride (substituted or unsubstituted) to form a acryloyl-substituted gelatin. In certain embodiments, chemically modified gelatin can be modified with one or more crosslinkable groups selected from the following: methyl acrylate, ethyl acrylate, propyl acrylate, methyl methacrylate, ethyl methacrylate, methacryloyl, catechol, ethylene oxide or propylene oxide. In certain embodiments, chemically modified gelatin can be modified with methacrylic anhydride (MA) (also referred to as methacrylic anhydride) to form a methacryloyl-substituted gelatin (commonly referred to as methacrylated gelatin or GelMA). Figure 1A An example of a reaction in which gelatin is modified with methacrylic anhydride to form methacryloyl-substituted gelatin (GelMA) is provided.

[0100] In certain embodiments, the acryloyl modification of gelatin can be carried out by a synthetic reaction of gelatin with a functionalized compound containing an acrylic acid group. In certain embodiments, the methacryloyl modification of gelatin can be carried out by a synthetic reaction of gelatin with the following substances: methacrylic anhydride, methacryloyl chloride, 2-isocyanatoethyl methacrylate, 2-hydroxyethyl methacrylate, glycidyl methacrylate, N-hydroxysuccinimide methacrylate, allyl methacrylate, vinyl methacrylate, di(2-methacryloyl)oxyethyl disulfide, 2-hydroxy-5-N-methacrylamidobenzoic acid, or a combination thereof.

[0101] As used herein, the terms "acryloyl-substituted gelatin" and "acrylated gelatin" may describe gelatin having free amines (e.g., lysine, arginine, asparagine, or glutamine side chains) and / or free hydroxyls (e.g., serine, threonine, aspartic acid, or glutamic acid side chains) that have been replaced by at least one acryloyl group. Typically, the acryloyl group is a substituted acryloyl group having the formula H 2 C=CR'-C(=O)-R represents an a,b-unsaturated carbonyl compound, wherein R' can be, but is not limited to, hydrogen, halogen, hydroxyl, C 1 -C 5 Alkoxy, C 1 -C 5 Alkyl, C 3 -C 8 Cycloalkyl, C1 -C 5 Heteroalkyl, C 3 -C 8 Heterocycloalkyl, aryl, heteroaryl or amino groups, each optionally substituted by halogen, C 1 -C 5 Alkoxy, C 1 -C 5 Alkyl, C 3 -C 8 Cycloalkyl, C 1 -C 5 Heteroalkyl, C 3 -C 8 Substitution with heterocycloalkyl, aryl, heteroaryl or amino groups. For the acryl-substituted gelatin of the present disclosure, the R group represents the terminal amine and / or hydroxyl groups on the gelatin that are acryl-functionalized.

[0102] In certain embodiments, the R' group of the acryloyl moiety is a methyl group, commonly referred to as a methacryloyl group. As used herein, the terms "methacryloyl-substituted gelatin," "methacrylylated gelatin," and "GelMA" can describe gelatin having free amines (e.g., lysine, arginine, asparagine, or glutamine side chains) and / or free hydroxyls (e.g., serine, threonine, aspartic acid, or glutamic acid side chains) replaced by at least one methacryloyl group, such as a methacrylamide group (from a free amine on the gelatin) and / or a methacrylic acid group (from a free hydroxyl on the gelatin).

[0103] In certain embodiments, the chemically modified gelatin (e.g., GelMA) can be present in the polymer composition at a concentration of about 1% to about 60% weight / volume (w / v). In certain embodiments, the chemically modified gelatin (e.g., GelMA) can be present in the polymer composition at a concentration of about 0.5%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about , about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59% or about 60% weight / volume (w / v) concentration is present in the polymer composition. In certain embodiments, chemically modified gelatin (e.g., GelMA) can be present in an amount of about 1-3%, about 3-6%, about 6-10%, about 1-5%, about 1-10%, about 5-10%, about 11-13%, about 13-16%, about 16-20%, about 10-20%, about 10-15%, about 15-20%, about 21-23%, about 23-26%, about 26-30%, about 20-30%, about 20-25%, about 25-30%, about 3 1-33%, about 33-36%, about 36-40%, about 30-40%, about 30-35%, about 35-40%, about 41-43%, about 43-46%, about 46-50%, about 40-50%, about 40-45%, about 45-50%, about 51-53%, about 53-56%, about 56-60%, about 50-60%, about 50-55% or about 55-60% weight / volume concentration (w / v) is present in the polymer composition.

[0104] In certain embodiments, polymer compositions include acrylated gelatin (i.e., GelMA) with a certain degree of acryloyl substitution (i.e., methacryloyl functionalization). As used herein, the term "acryloyl substitution degree" can describe the percentage of free amines and hydroxyls substituted by acryloyl groups in gelatin. As used herein, the term "methacryloyl substitution degree" can describe the percentage of free amines and hydroxyls substituted by methacryloyl groups in gelatin. In certain embodiments, polymer compositions include acrylated gelatin with an acryloyl substitution degree of at least about 10%, at least about 15%, at least about 20%, at least about 25%, at least about 30%, at least about 35%, at least about 40%, at least about 45%, at least about 50%, at least about 55%, at least about 60%, at least about 65%, at least about 70%, at least about 75%, at least about 80%, at least about 85% or at least about 90%. In certain embodiments, the polymer composition comprises acrylated gelatin having an acryl substitution degree of about 10-99%. In certain embodiments, the acryl substitution degree is about 1-5%, about 5-10%, about 10-15%, about 15-20%, about 20-25%, about 25-30%, about 30-35%, about 35-40%, about 40-45%, about 45-50%, about 50-55%, about 55-60%, about 60-65%, about 65-70%, about 70-75%, about 75-80%, about 80-85%, about 85-90%, about 90-95%, or about 95-99%. In certain embodiments, the polymer composition comprises a GelMA having a degree of methacryloyl substitution of about 1-5%, about 5-10%, about 10-15%, about 15-20%, about 20-25%, about 25-30%, about 30-35%, about 35-40%, about 40-45%, about 45-50%, about 50-55%, about 55-60%, about 60-65%, about 65-70%, about 70-75%, about 75-80%, about 80-85%, about 85-90%, about 90-95%, or about 95-99%.

[0105] In certain embodiments, the polymer composition comprises a GelMA having a certain degree of methacrylamide substitution (i.e., methacrylamide functionalization). In certain embodiments, the polymer composition comprises a GelMA having a methacrylamide substitution of at least about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or at least about 90%. In certain embodiments, the polymer composition comprises a GelMA having a methacrylamide substitution of about 20-90%. In certain embodiments, the methacrylamide degree of substitution is about 1-5%, about 5-10%, about 10-15%, about 15-20%, about 20-25%, about 25-30%, about 30-35%, about 35-40%, about 40-45%, about 45-50%, about 50-55%, about 55-60%, about 60-65%, about 65-70%, about 70-75%, about 75-80%, about 80-85% or about 85-90%. In certain embodiments, the methacrylamide degree of substitution can be measured using proton nuclear magnetic resonance. In certain embodiments, the methacrylamide degree of substitution can be measured using fluorinated aldehyde assay.

[0106] In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylate substitution (i.e., methacrylate functionalization). In certain embodiments, the polymer composition comprises a GelMA having a methacrylate substitution of at least about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, about 80%, about 85%, or at least about 90%. In certain embodiments, the polymer composition comprises a GelMA having a methacrylate substitution of about 20-90%. In certain embodiments, the methacrylate degree of substitution is about 1-5%, about 5-10%, about 10-15%, about 15-20%, about 20-25%, about 25-30%, about 30-35%, about 35-40%, about 40-45%, about 45-50%, about 50-55%, about 55-60%, about 60-65%, about 65-70%, about 70-75%, about 75-80%, about 80-85% or about 85-90%. In certain embodiments, the methacrylic acid degree of substitution can be measured using proton nuclear magnetic resonance. In certain embodiments, the methacrylic acid degree of substitution can be measured using an assay based on Fe (III)-hydroxamic acid. In certain embodiments, the measurement of the methacrylic acid degree of substitution can include the aminolysis reaction (e.g., by exposure to a hydroxylamine solution) of converting a methacrylic acid group into an N-hydroxymethyl acrylamide group.

[0107] In certain embodiments, the polymer composition comprises a GelMA having a degree of methacrylamide substitution and a degree of methacrylate substitution. In certain embodiments, the ratio of methacrylamide substitution to methacrylate substitution in GelMA is about 1:1 to 99:1. In some embodiments, the ratio of methacrylamide substitution to methacrylate substitution is about 1:1 to 2:1, about 2:1 to 3:1, about 3:1 to 4:1, about 4:1 to 5:1, about 1:1 to 5:1, about 5:1 to 10:1, about 10:1 to 15:1, about 15:1 to 20:1, about 20:1 to 25:1, about 25:1 to 30:1, about 30:1 to 35:1, about 1 to 99: 1. In certain embodiments, the ratio of methacrylate substitution to methacrylamide substitution in GelMA is about 1: 1 to 99: 1. In some embodiments, the ratio of methacrylate substitution to methacrylamide substitution is about 1:1 to 2:1, about 2:1 to 3:1, about 3:1 to 4:1, about 4:1 to 5:1, about 1:1 to 5:1, about 5:1 to 10:1, about 10:1 to 15:1, about 15:1 to 20:1, about 20:1 to 25:1, about 25:1 to 30:1, about 30:1 to 35:1, about 1, about 35:1 to 40:1, about 40:1 to 45:1, about 45:1 to 50:1, about 50:1 to 55:1, about 55:1 to 60:1, about 60:1 to 65:1, about 65:1 to 70:1, about 70:1 to 75:1, about 75:1 to 80:1, about 80:1 to 85:1, about 85:1 to 90:1, about 90:1 to 95:1, or about 95:1 to 99:1.

[0108] In certain embodiments, the polymer composition comprises a GelMA having a methacryloyl modification of gelatin by reaction of gelatin with methacrylic anhydride. In certain embodiments, the polymer composition comprises a GelMA having a methacryloyl modification of gelatin by reaction of gelatin with glycidyl methacrylate.

[0109] In certain embodiments, gelatin can be functionalized with anchoring integrins and / or proteins (e.g., proteins that bind to surface proteins of a target surface). The functionalization can be performed with poly(ethylene glycol) (PEG) or other polymer linkers between gelatin and integrins and / or proteins.

[0110] Chemically modified hyaluronic acid

[0111] Hyaluronic acid (HA) is a viscoelastic and biocompatible glycosaminoglycan naturally present in the cornea and other tissues. In certain embodiments, the polymer composition of the present disclosure comprises chemically modified hyaluronic acid (HA). In certain embodiments, the polymer composition comprises acryloyl-substituted hyaluronic acid. In certain embodiments, the polymer composition comprises methacrylated hyaluronic acid (MeHA). In certain embodiments, the chemically modified HA may be included in the precursor polymer composition of the present disclosure. In certain embodiments, the chemically modified HA comprises a photocrosslinkable derivative of HA. In certain embodiments, the chemically modified HA comprises methacrylated hyaluronic acid (MeHA). In certain embodiments, the chemically modified HA comprises methacrylated hyaluronic acid (MeHA), which comprises methacrylic anhydride-hyaluronic acid (HAMA); i.e., MeHA formed by reacting methacrylic anhydride with hyaluronic acid. In certain embodiments, the chemically modified HA comprises methacrylated hyaluronic acid (MeHA), which comprises glycidyl methacrylate-hyaluronic acid (HAGM); i.e., MeHA formed by reacting glycidyl methacrylate with hyaluronic acid. In certain embodiments, methacrylation of HA can be performed via a ring-opening reaction of the HA backbone coupled with a reversible transesterification reaction. Figure 1B An example of a reaction in which hyaluronic acid is modified with glycidyl methacrylate to form the HAGM form of methacrylated hyaluronic acid (MeHA) is provided.

[0112] In certain embodiments, the chemically modified HA (e.g., MeHA) can be present in the polymer composition at a concentration of about 1% to about 60% weight / volume (w / v). In certain embodiments, the chemically modified HA (e.g., MeHA) can be present in the polymer composition at a concentration of about 0.5%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, , about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59% or about 60% weight / volume concentration (w / v) is present in the polymer composition. In certain embodiments, the chemically modified HA (e.g., MeHA) can be present in an amount of about 1-3%, about 3-6%, about 6-10%, about 1-5%, about 1-10%, about 5-10%, about 11-13%, about 13-16%, about 16-20%, about 10-20%, about 10-15%, about 15-20%, about 21-23%, about 23-26%, about 26-30%, about 20-30%, about 20-25%, about 25-30%, about 3 1-33%, about 33-36%, about 36-40%, about 30-40%, about 30-35%, about 35-40%, about 41-43%, about 43-46%, about 46-50%, about 40-50%, about 40-45%, about 45-50%, about 51-53%, about 53-56%, about 56-60%, about 50-60%, about 50-55% or about 55-60% weight / volume concentration (w / v) is present in the polymer composition.

[0113] In certain embodiments, the polymer compositions of the present disclosure comprise acryl-substituted gelatin (e.g., GelMA) and acryl-substituted hyaluronic acid (e.g., MeHA) in a ratio of about 30:1 to about 1:30 w / w. In certain embodiments, the polymer compositions of the present disclosure comprise acryl-substituted gelatin and acryl-substituted hyaluronic acid in a ratio (w / w) of about 30:1, about 29:1, about 28:1, about 27:1, about 26:1, about 25:1, about 24:1, about 23:1, about 22:1, about 21:1, about 20:1, about 19:1, about 18:1, about 17:1, about 16:1, about 15:1, about 14:1, about 13:1, about 12:1, about 11:1, about 10:1, about 9:1, about 8:1, about 7:1, about 6:1 :1, about 5:1, about 4:1, about 3:1, about 2:1, about 1:1, about 1:2, about 1:3, about 1:4, about 1:5, about 1:6, about 1:7, about 1:8, about 1:9, about 1:10, about 1:11, about 1:12, about 1:13, about 1:14, about 1:15, about 1:16, about 1:17, about 1:18, about 1:19, about 1:20, about 1:21, about 1:22, about 1:23, about 1:24, about 1:25, about 1:26, about 1:27, about 1:28, about 1:29 or about 1:30.

[0114] In certain embodiments, an acryl-substituted hyaluronic acid (e.g., MeHA) can be synthesized as taught in Bencherif et al., Biomaterials 29, 1739–1749 (2008); or Prata et al., Biomacromolecules 11, 769–775 (2010); each of which is incorporated herein by reference in its entirety to the extent that each describes the composition, production, analysis, and use of acryl-substituted hyaluronic acid polymer compositions such as MeHA.

[0115] Chemically modified poly(ethylene glycol)

[0116] Poly (ethylene glycol) (PEG) is a synthetic linear polymer, known to have high biocompatibility and immunotolerance in the human body, and is soluble in many aqueous and organic solvents. In certain embodiments, polymer compositions of the present disclosure include chemically modified PEG. In certain embodiments, polymer compositions include acryloyl-substituted PEG. In certain embodiments, polymer compositions include one or more acryloyl-substituted PEGs selected from the following: PEG diacrylate (PEGDA), PEG monoacrylate, PEG dimethacrylate, PEG monomethacrylate, methoxy PEG acrylate, methoxy PEG methacrylate, ethoxy PEG acrylate, ethoxy PEG methacrylate, propoxy PEG acrylate or propoxy PEG methacrylate.

[0117] In certain embodiments, the polymer composition comprises poly (ethylene glycol) diacrylate (PEGDA). In certain embodiments, chemically modified PEG may be included in the precursor polymer composition of the present disclosure. In certain embodiments, the chemically modified PEG comprises a photocrosslinkable derivative of PEG. In certain embodiments, the chemically modified PEG comprises poly (ethylene glycol) diacrylate (PEGDA). In certain embodiments, the chemical modification of PEG can be carried out by reacting PEG with acryloyl chloride or a functionally similar acrylated compound. Figure 1C An example of a reaction in which poly(ethylene glycol) (PEG) is modified with acryloyl chloride to form poly(ethylene glycol) diacrylate (PEGDA) is provided.

[0118] In certain embodiments, the chemically modified PEG has a molecular weight of about 5 kDa to about 200 kDa. In certain embodiments, the chemically modified PEG can have a molecular weight of about 5-10 kDa, about 10-15 kDa, about 15-20 kDa, about 20-25 kDa, about 25-30 kDa, about 30-35 kDa, about 35-40 kDa, about 40-45 kDa, about 45-50 kDa, about 50-55 kDa, about 55-60 kDa, about 60-65 kDa, about 65-70 kDa, about 70-75 kDa, about 75-80 kDa, about 80-85 kDa, about 85-90 kDa, about 90-95 kDa, about 95-100 kDa, about 100-105 kDa, about 105 -110 kDa, about 110-115 kDa, about 115-120 kDa, about 120-125 kDa, about 125-130 kDa, about 130-135 kDa, about 135-140 kDa, about 140-145 kDa, about 145-150 kDa, about 150-155 kDa, about 155-160 kDa, about 160-165 kDa, about 165-170 kDa, about 170-175 kDa, about 175-180 kDa, about 180-185 kDa, about 185-190 kDa, about 190-195 kDa, or about 195-200 kDa.

[0119] In certain embodiments, the chemically modified PEG (e.g., PEGDA) can be present in the polymer composition at a concentration of about 1% to about 60% weight / volume (w / v). In certain embodiments, the chemically modified PEG (e.g., PEGDA) can be present in the polymer composition at a concentration of about 0.5%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 8 , about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59% or about 60% weight / volume (w / v) concentration is present in the polymer composition. In certain embodiments, chemically modified PEG (e.g., PEGDA) can be present in an amount of about 1-3%, about 3-6%, about 6-10%, about 1-5%, about 1-10%, about 5-10%, about 11-13%, about 13-16%, about 16-20%, about 10-20%, about 10-15%, about 15-20%, about 21-23%, about 23-26%, about 26-30%, about 20-30%, about 20-25%, about 25-30%, about 3 1-33%, about 33-36%, about 36-40%, about 30-40%, about 30-35%, about 35-40%, about 41-43%, about 43-46%, about 46-50%, about 40-50%, about 40-45%, about 45-50%, about 51-53%, about 53-56%, about 56-60%, about 50-60%, about 50-55% or about 55-60% weight / volume concentration (w / v) is present in the polymer composition.

[0120] In certain embodiments, the polymer compositions of the present disclosure comprise acryl-substituted gelatin (e.g., GelMA) and acryl-substituted PEG (e.g., PEGDA) in a ratio of about 30:1 to about 1:30 w / w. In certain embodiments, the polymer compositions of the present disclosure comprise acryl-substituted gelatin and acryl-substituted PEG in a ratio (w / w) of about 30:1, about 29:1, about 28:1, about 27:1, about 26:1, about 25:1, about 24:1, about 23:1, about 22:1, about 21:1, about 20:1, about 19:1, about 18:1, about 17:1, about 16:1, about 15:1, about 14:1, about 13:1, about 12:1, about 11:1, about 10:1, about 9:1, about 8:1, about 7:1, about 8:1 :1, about 5:1, about 4:1, about 3:1, about 2:1, about 1:1, about 1:2, about 1:3, about 1:4, about 1:5, about 1:6, about 1:7, about 1:8, about 1:9, about 1:10, about 1:11, about 1:12, about 1:13, about 1:14, about 1:15, about 1:16, about 1:17, about 1:18, about 1:19, about 1:20, about 1:21, about 1:22, about 1:23, about 1:24, about 1:25, about 1:26, about 1:27, about 1:28, about 1:29 or about 1:30.

[0121] In certain embodiments, the polymer compositions of the present disclosure comprise acryl-substituted PEG (e.g., PEGDA) and acryl-substituted hyaluronic acid (e.g., MeHA) in a ratio of about 30: 1 to about 1:30 w / w. In certain embodiments, the polymer compositions of the present disclosure comprise acryl-substituted PEG and acryl-substituted hyaluronic acid in a ratio (w / w) of about 30: 1, about 29: 1, about 28: 1, about 27: 1, about 26: 1, about 25: 1, about 24: 1, about 23: 1, about 22: 1, about 21: 1, about 20: 1, about 19: 1, about 18: 1, about 17: 1, about 16: 1, about 15: 1, about 14: 1, about 13: 1, about 12: 1, about 11: 1, about 10: 1, about 9: 1, about 8: 1, about 7: 1, about about 1:24, about 1:25, about 1:26, about 1:27, about 1:28, about 1:29, or about 1:30.

[0122] In certain embodiments, the polymer compositions of the present disclosure comprise one or more synthetic polymer components (i.e., polymers or precursors) selected from the group consisting of methacrylate-oligolactide-PEO-oligolactide-methacrylate, polyethylene glycol (PEG), polyglycerol sebacate (PGS), polylactic acid (PLA), polypropylene glycol (PPO), PEG-PPO-PEG copolymers (e.g., pluronics), polyphosphazenes, polymethacrylates, poly(N-vinylpyrrolidone), and polyethyleneimine.

[0123] Crosslinking agent

[0124] In certain embodiments, the polymer composition of the present disclosure includes a crosslinking agent. As used herein, the phrase "crosslinking agent" can describe a substance that forms, promotes or regulates intermolecular bonding (covalent bonding, ionic bonding, hydrogen bonding) between polymer units or chains to produce a polymer chain network. Crosslinking agents typically exhibit one or more, optionally two or more bonding functionalities that can produce chemical bonds between two or more polymer chains. Crosslinking agents can include, for example, two vinyl bonds (four functionalities) or three amines (three functionalities).

[0125] In certain embodiments, the polymer composition comprises a cross-linking agent that can be used to activate or promote the polymerization, gelation and curing of the polymer composition from the precursor polymer composition into a gel polymer composition. In certain embodiments, exposing the polymer composition of the present disclosure (e.g., a precursor polymer composition) to cross-linking conditions (e.g., exposure to visible light in the presence of a photoinitiator) can cause one or more acryloyl groups in the polymer composition (e.g., acryloyl-substituted gelatin, acryloyl-substituted HA, acryloyl-substituted PEG and other acryloyl-based cross-linkers) to react with other acryloyl groups to cross-link the polymer composition and form a gel polymer composition (e.g., GelMA hydrogel).

[0126] In certain embodiments, the polymer composition of the present disclosure (e.g., a precursor polymer composition) comprises from about 1% to about 50% (w / v) of one or more cross-linking agents. In certain embodiments, the polymer composition comprises a concentration (w / v) of at least about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, or about 40% of one or more cross-linking agents. In certain embodiments, the polymer composition comprises a concentration (w / v) of no more than about 50%, about 45%, about 40%, about 35%, or about 30% of one or more cross-linking agents. In certain embodiments, the polymer composition comprises a concentration (w / v) of about 1-3%, about 3-6%, about 6-10%, about 1-5%, about 1-10%, about 5-10%, about 11-13%, about 13-16%, about 16-20%, about 10-20%, about 10-15%, about 15-20%, about 21-23%, about 23-26%, about 26 -30%, about 20-30%, about 20-25%, about 25-30%, about 31-33%, about 33-36%, about 36-40%, about 30-40%, about 30-35%, about 35-40%, about 41-43%, about 43-46%, about 46-50%, about 40-50%, about 40-45% or about 45-50% of one or more cross-linking agents.

[0127] In certain embodiments, the polymer compositions of the present disclosure comprise one or more cross-linking agents selected from the group consisting of glutaraldehyde, epoxides (e.g., diethylene oxide), oxidized dextran, p-azidobenzoyl hydrazide, N-(a-maleimidoacetoxy)succinimide ester, p-azidophenylglyoxal monohydrate, bis-((4-azidosalicylamido)ethyl)disulfide, bis(sulfosuccinimidyl)suberate, dithiobis(succinimidylpropionate), disuccinimidyl suberate, 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC), ethoxylated trimethylpropane triacrylate, N-hydroxysuccinimide (NHS), derivatives thereof, or combinations thereof.

[0128] In certain embodiments, the polymer composition of the present invention comprises one or more crosslinking agents selected from the group consisting of polyethylene oxide dimethacrylate, methylene bisacrylamide, methylene bis(2-methacrylamide), methylene diacrylate, methylene bis(2-methacrylate), diethylene glycol diacrylate, hexamethylene diacrylate, hexamethylene diisocyanate, oxybis(methylene)bis(2-methacrylate), oxybis(ethane-2,1-diyl)bis(2-methacrylate), trimethylolpropane triacrylate, pentaerythritol triacrylate, tris(2-hydroxyethyl)isocyanurate triacrylate, isocyanurate tris(2-acryloyloxyethyl), ethoxylated trimethylolpropane triacrylate, pentaerythritol triacrylate, and glycerol triacrylate, oxyphosphinyl tris(oxyethylene) triacrylate, derivatives thereof, or combinations thereof.

[0129] Polymer crosslinking initiator / photoinitiator

[0130] In certain embodiments, the polymer composition comprises one or more polymer crosslinking initiators, such as a photoinitiator component. In certain embodiments, the polymer crosslinking initiator forms free radicals when exposed to specific polymer crosslinking conditions (e.g., acidic conditions, alkaline conditions, high salt conditions, low salt conditions, high temperature, stirring, dissolution conditions, exposure), wherein the free radicals can cause bond formation between reactive groups in the composition, such as vinyl bond crosslinking between methacrylate groups in the GelMA polymer composition.

[0131] In certain embodiments, the polymer composition comprises one or more photoinitiator components (i.e., a cross-linking initiator that is initiated or activated by absorbing light of a certain wavelength). In certain embodiments, the precursor polymer composition of the present disclosure comprises one or more photoinitiator components. In certain embodiments, the photoinitiator component can be activated by exposure. In certain embodiments, exposure can activate the photoinitiator to form free radicals, wherein the free radicals can cause bond formation between reactive groups in the composition, such as vinyl bond cross-linking between methacrylic acid groups in the GelMA polymer composition.

[0132] In certain embodiments, the photoinitiator composition can be activated by being exposed to one or more light sources selected from the following: a visible light source (e.g., white light or blue light), an ultraviolet (UV) light source, a near infrared (NIR) light source, and a fluorescent light source. In certain embodiments, the photoinitiator composition includes a visible light activated photoinitiator, such as a visible light activated photoinitiator activated when exposed to a wavelength of about 380nm to about 740nm. In certain embodiments, the visible light activated photoinitiator can be activated when exposed to a wavelength of about 380-435nm (i.e., purple light), about 435-500nm (i.e., blue light), about 500-565nm (i.e., green light), about 565-600nm (i.e., yellow light), about 600-650nm (i.e., orange light) or about 650-740nm (i.e., red light). In certain embodiments, the photoinitiator composition includes a UV-activated photoinitiator. In certain embodiments, the photoinitiator composition includes a near infrared (NIR) light activated photoinitiator. In certain embodiments, the photoinitiator component comprises a white light activated photoinitiator. In certain embodiments, the photoinitiator component comprises a blue light activated photoinitiator.

[0133] In certain embodiments, the polymer composition comprises one or more photoinitiator components selected from the group consisting of triethanolamine; 1-vinyl-2-pyrrolidone (NVP); N-vinyl caprolactam (NVC); ethylene glycol diacrylate (EGDA); riboflavin; azobisisobutyronitrile; benzoyl peroxide; 1-benzoylcyclohexanol; di-tert-butyl peroxide; eosin Y (e.g., disodium salt), (2-(2,4,5,7-tetrabromo-6-oxido-3-oxo-3H-xanthene-9-yl)benzoate); 4,6-trimethylbenzoylphosphite; triethanolamine; 2,3-diketo-1,7,7-trimethylnorbornane; 1-phenyl-1,2-propanedione; 2,4,6-trimethylbenzoyl-diphenyl phosphine oxide; bis(2,6-dichlorobenzoyl)-(4-propylphenyl)phosphine oxide; 4,4'-bis(dimethylamino)benzophenone; 4,4'-bis(diethylamino)benzophenone; 2-chlorothioxanthene-9-one; 4-(dimethylamino)benzophenone; phenanthrenequinone; ferrocene; 2-hydroxy-4'-(2-hydroxyethoxy)-2-methylpropiophenone; 2-hydroxy-2-methylpropiophenone; diphenyl(2,4,6-trimethylbenzoyl)phosphine oxide / 2-hydroxy-2-methylpropiophenone (blend); benzoin methyl ether; benzoin isopropyl ether; 2,2-diethoxyacetophenone; dibutoxyacetophenone; 2,2-dimethoxy-2-phenyl-1-phenylethanone; 2,2-dimethoxy-2-phenylacetophenone; dibenzosuberyl enone; (Benzene) tricarbonyl chromium; resazurin; resorufin; benzoyl trimethylgermanium; lithium phenyl-2,4,6-trimethylbenzoylphosphonate; camphorquinone; 2-methyl-1-(4-(methylthio)phenyl)-2-morpholinopropan-2-one; 2-benzyl-2-(dimethylamino)-4'-morpholinobutyrophenone; 2-benzyl-2-dimethylamino-1-(4-morpholinophenyl)butan-1-one; methyl benzoylformate; bis(2,4,6-trimethylbenzoyl)-phenylphosphine oxide; bis(2,4-cyclopentadien-1-yl)-bis(2,6-difluoro-3-(1H-pyrrol-1-yl)-phenyl)titanium; 5,7-diiodo-3-butoxy-6-fluorone; 2,4,5,7-tetraiodo-3-hydroxy-6-fluorone ; 2,4,5,7-tetraiodo-3-hydroxy-9-cyano-6-fluorone; dimethoxyhydroxy-acetophenone; 2-naphthalene-sulfonyl chloride; 1-phenyl-1,2-propanedione-2-(O-ethoxy-carbonyl)oxime; 2-ethylthioxanthone; 2-isopropylthioxanthone; 2,4-diethylthioxanthone; 2-tert-butylthioxanthone; 2-chlorothioxanthone; 2-propoxythioxanthone; methylphenylglyoxylate; phenyl 2-hydroxy-2-propyl ketone; 4-isopropylphenyl 2-hydroxy-2-propyl ketone; 4-n-dodecylphenyl 2-hydroxy-2-propyl ketone; 4-(2-hydroxyethoxy)phenyl 2-hydroxy-2-propyl ketone; 1-[4-(2-hydroxyethoxy)-phenyl]-2-hydroxy-2-methyl-1-propane-1-one);4-(2-acryloyloxyethoxy)phenyl 2-hydroxy-2-propyl ketone; derivatives thereof, or combinations thereof. In certain embodiments, the polymer composition comprises a combination of Eosin Y, triethanolamine, and / or vinyl caprolactam. In certain embodiments, the polymer crosslinking initiator comprises Eosin Y disodium salt (EYDS), N-vinyl caprolactam (NVC), triethanolamine, or any combination thereof. In certain embodiments, the polymer crosslinking initiator comprises Eosin Y disodium salt (EYDS), N-vinyl pyrrolidone (NVP), triethanolamine, or any combination thereof. ;

[0134] In certain embodiments, the polymer cross-linking initiator comprises: (i) about 50 μM Eosin Y or Eosin Y disodium salt (EYDS), optionally about 50 μM Eosin Y disodium salt (EYDS); (ii) about 3.5 to about 5.0 μL / mL N-vinyl caprolactam (NVC) or N-vinyl pyrrolidone (NVP), optionally about 3.5 to about 5.0 μL / mL NVP, optionally about 5.0 μL / mL NVP; and (iii) triethanolamine, optionally about 1.5% v / v triethanolamine.

[0135] In certain embodiments, the polymer composition comprises one or more photoinitiator components selected from the group consisting of acetophenone; anisole; anthraquinone; anthraquinone-2-sulfonic acid sodium salt monohydrate; (benzene) tricarbonyl chromium; 4-(boc-aminomethyl) phenyl isothiocyanate; benzin; benzoin; benzoin ethyl ether; benzoin isobutyl ether; benzoin methyl ether; benzoic acid; benzophenyl-hydroxycyclohexyl phenyl ketone; 3,3',4,4'-benzophenone tetracarboxylic acid dianhydride; 4-benzoylbiphenyl; 2-benzyl-2-(dimethylamino)-4'-morpholinobutyrophenone; 4,4'-bis(diethylamino)benzophenone; 4,4'-bis(dimethylamino)benzophenone; Michler's ketone ketone); camphorquinone; 2-chlorothioxanthene-9-one; 5-dibenzocycloheptenone; (cumene) cyclopentadienyl iron (II) hexafluorophosphate; dibenzocycloheptenone; 2,2-diethoxyacetophenone; 4,4'-dihydroxybenzophenone; 2,2-dimethoxy-2-phenylacetophenone; 4-(dimethylamino)benzophenone; 4,4'-dimethylbenzyl; 2,5-dimethylbenzophenone; 3,4-dimethyl benzophenone; diphenyl (2,4,6-trimethylbenzoyl) phosphine oxide; 2-hydroxy-2-methylpropiophenone; 4'-ethoxyacetophenone; 2-ethylanthraquinone; ferrocene; 3'-hydroxyacetophenone; 4'-hydroxyacetophenone; 3-hydroxybenzophenone; 4-hydroxybenzophenone; 1-hydroxycyclohexyl phenyl ketone; 2-hydroxy-2-methylpropiophenone; 2-methylbenzophenone; 3-methylbenzophenone; benzoylmethyl methyl ester; 2-methyl-4'-(methylthio)-2-morpholinopropiophenone; 9,10-phenanthrenequinone; 4'-phenoxyacetophenone; thioxanthen-9-one; triarylsulfonium hexafluoroantimonate; triarylsulfonium hexafluorophosphate; 3-mercapto-1-propanol; mercapto-1-undecanol; 1-mercapto-2-propanol; 3-mercapto-2-butanol; hydrogen peroxide; benzoyl peroxide; 4,4'-dimethoxybenzoin; 2,2-dimethoxybenzoin 2-phenylacetophenone; dibenzoyl disulfide; diphenyl dithiocarbonate; 2,2'-azobisisobutyronitrile; 2,2'-azobis[2-methyl-N-(2-hydroxyethyl)propionamide; camphorquinone; eosin; dimethylaminobenzoate; dimethoxy-2-phenyl-acetophenone; quantum cure ITX photosensitizers; Irgacures (e.g., 907, 2959, 651); Darocur 2959; 4-N,N-dimethylaminobenzoic acid ethyl ester; 1-[-(4-benzoylphenylthio)phenyl]-2-methyl-2-(4-methylphenylsulfonyl)propan-1-one; 1-hydroxy-cyclohexyl-phenyl-ketone; 2,4,6-trimethylbenzoyldiphenylphosphine oxide; diphenyl (2,4,6-trimethylbenzoyl) phosphine; 2-ethylhexyl-4-dimethylaminobenzoate; 2-hydroxy-2-methyl-1-phenyl-1-propanone;Oligo[2-hydroxy-2-methyl-1-[4-(methylvinyl)phenyl]propanone] and propoxylated glyceryl triacrylate; benzodimethyl ketal; benzophenone; blends of benzophenone and α-hydroxy-cyclohexyl-phenyl ketone; blends of Esacure KIP 150 and Esacure TZT; blends of Esacure KIP150 and Esacure TZT; blends of Esacure KIP150 and TPGDA; phosphine oxide, Esacure KIP 150 and Esacure Blends of TZT; difunctional α-hydroxy ketones; ethyl 4-dimethylaminobenzoate; isopropyl thioxanthone; 2-hydroxy-2-methyl-phenylacetone; 2,4,6-trimethylbenzoyldiphenylphosphine oxide; 2,4,6-trimethylbenzophenone; blends of 4-methylbenzophenone and benzophenone; oligomeric (2-hydroxy-2-methyl-1-(4(1-methylvinyl)phenyl)acetone; oligomeric (2-hydroxy-2-methyl-1-4(1-methylvinyl)phenylacetone and 2-hydroxy-2-methyl-1-phenyl-1-propanone; 4-methylbenzophenone; trimethylbenzophenone and methylbenzophenone; and aqueous emulsions of 2,4,6-trimethylbenzoylphosphine oxide, α-hydroxy ketone, trimethylbenzophenone and 4-methylbenzophenone.

[0136] In certain embodiments, the polymer composition comprises one or more cationic and / or anionic photoinitiator components selected from the group consisting of titanium tetrachloride, vanadium tetrachloride, bis(cyclopentadienyl)titanium dichloride, ferrocene, cyclopentadienyl manganese tricarbonyl, manganese decacarbonyl, diazonium salts, diaryliodonium salts (e.g., 3,3'-dinitrodiphenyliodonium hexafluoroarsenate, diphenyliodonium fluoroborate, 4-methoxydiphenyliodonium fluoroborate), and triarylsulfonium salts.

[0137] Photoinitiated polymerization and photoinitiators are discussed in detail in Rabek, Mechanisms of Photophysical Processes and Photochemical Reactions in Polymers, New York: Wiley & Sons, 1987; and Fouassier, Photoinitiation, Photopolymerization, and Photocuring, Cincinnati, Ohio: Hanser / Gardner; Fisher et al., 2001, Annu. Rev. Mater. Res., 31:171; each is incorporated herein by reference in its entirety to the extent that each describes the use of polymerization and photoinitiators in the production of polymer compositions, including acryl gelatin such as GelMA hydrogels.

[0138] In certain embodiments, the polymer composition comprises a crosslinking agent or initiator comprising one or more metal 2+ Ions and / or metals 3+ In certain embodiments, the polymer composition comprises a crosslinking agent comprising one or more metals selected from the group consisting of 2+ Ions and / or metals 3+ Ion: Fe 2+ , Fe 3+ 、Ni 2+ 、Zn 2+ , Cu 2+ 、Ag 2+ 、Au 3+ 、Co 2+ 、Co 3+ Cr 2+ Cr 3+ 、Cd 2+ , Mn 2+ Mg 2+ , Pd 2+ , Pt 2+ 、Al 3+ In certain embodiments, the precursor polymer composition of the present disclosure comprises one or more photoinitiator components and one or more metal 2+ / 3+ ions both.

[0139] In certain embodiments, the polymer composition comprises a cross-linker or initiator that is polymerized cross-linked using click bioconjugation chemistry. In certain embodiments, the polymer composition comprises a cross-linker or initiator that is polymerized cross-linked using click bioconjugation chemistry selected from the following: metal-catalyzed azide-alkyne cycloaddition, strain-promoted azide-alkyne cycloaddition, strain-promoted alkyne-nitrone cycloaddition (e.g., olefin / azide [3+2] cycloaddition, olefin / tetrazine reverse demand Diels-Alder, olefin / tetrazolyl photoclick reaction), or a combination thereof.

[0140] II. Physical, Mechanical and Structural Characteristics

[0141] Viscosity, shear strength and shear resistance

[0142] The viscosity of a material is a measure of its resistance to deformation at a given rate. The viscosity of a fluid material is often related to the thickness and / or density of the material.

[0143] In certain embodiments, the polymer composition of the present disclosure may have a therapeutically effective viscosity. In certain embodiments, the polymer composition may have a viscosity that provides strong adhesion and high polymer composition retention on the target tissue of the subject. In certain embodiments, the precursor polymer composition of the present disclosure may have a viscosity that provides strong adhesion and high polymer composition retention on the target tissue of the subject. In certain embodiments, the precursor polymer composition may have a viscosity greater than water. In certain embodiments, the precursor polymer composition may have a viscosity comparable to a paste. In certain embodiments, the gel polymer composition of the present disclosure may have a viscosity that provides strong adhesion and high polymer composition retention on the target tissue of the subject. In certain embodiments, the precursor polymer composition may have a viscosity comparable to water. In certain embodiments, the gel polymer composition may maintain its shape and / or consistency on the surface of the target tissue for one or more hours, one or more days, or one or more weeks.

[0144] In certain embodiments, the polymer composition is subjected to a low shear rate (e.g., at about 0.001 s -1 To about 1s -1 In some embodiments, the polymer composition may have a viscosity of about 0.5 Pa-second (Pa·s) to about 300 Pa·s at a shear rate of about 1.5-2.0. In some embodiments, the polymer composition may have a viscosity of about 0.5-100 Pa·s at a low shear rate. In some embodiments, the polymer composition may have a viscosity of about 0.5-5 Pa·s, about 5-10 Pa·s, about 10-15 Pa·s, about 15-20 Pa·s, about 20-25 Pa·s, about 25-30 Pa·s, about 30-35 Pa·s, about 35-40 Pa·s, about 40-45 Pa·s, about 45-50 Pa·s, about 50-55 Pa·s, about 55-60 Pa·s, about 60-65 Pa·s, about 65-70 Pa·s at a low shear rate. ·s, about 70-75 Pa·s, about 75-80 Pa·s, about 80-85 Pa·s, about 85-90 Pa·s, about 90-95 Pa·s, about 95-100 Pa·s, about 100-125 Pa·s, about 125-150 Pa·s, about 150-175 Pa·s, about 175-200 Pa·s, about 200-225 Pa·s, about 225-250 Pa·s, about 250-275 Pa·s or about 275-300 Pa·s.

[0145] Shear strength and / or shear resistance are measurements of the ability of a material to resist external shear stress (i.e., shear load) without failure (i.e., loss of adhesion or integrity). In certain embodiments, the polymer composition of the present disclosure may have a therapeutically effective shear strength. In certain embodiments, the polymer composition may have a shear strength that provides a lasting adhesion and a high polymer composition retention force on the target tissue of the subject. In certain embodiments, the gel polymer composition of the present disclosure may have a shear strength that provides a lasting adhesion and a high polymer composition retention force on the target tissue of the subject. In certain embodiments, the gel polymer composition may have a shear strength that allows the polymer composition to maintain its shape, adhesion, connectivity and / or consistency on the surface of the target tissue for one or more hours, one or more days, or one or more weeks.

[0146] In certain embodiments, the polymer composition can have a shear strength of about 1 to about 360 kPa. In certain embodiments, the polymer composition can have a shear strength of about 100-360 kPa. In certain embodiments, the polymer composition can have a shear strength of about 200-360 kPa. In certain embodiments, the polymer composition can have a shear strength of about 1-20 kPa, about 20-40 kPa, about 40-60 kPa, about 60-80 kPa, about 80-100 kPa, 100-120 kPa, about 120-140 kPa, about 140-160 kPa, about 160-180 kPa, about 180-200 kPa, 200-220 kPa, about 220-240 kPa, about 240-260 kPa, about 260-280 kPa, about 280-300 kPa, 300-320 kPa, about 320-340 kPa, or about 340-360 kPa.

[0147] In certain embodiments, the shear strength of the polymer composition may be measured using ASTM F2255-05 or a modified lap shear test variation thereof.

[0148] Swelling and water content

[0149] In certain embodiments, the polymer composition comprises a gel. The gel generally comprises a cross-linked polymeric framework that encompasses a network of pores filled with an interstitial solvent (e.g., a fluid). In certain embodiments, the polymer composition comprises a hydrogel, wherein the interstitial fluid comprises water. In certain embodiments, the polymer composition comprises an alcohol gel, wherein the interstitial fluid comprises an alcohol (e.g., methanol, ethanol).

[0150] Swelling (i.e., increase in volume) can occur in a gel as the gel material absorbs and holds additional interstitial fluid in the pore network of the gel. Similarly, shrinkage (i.e., decrease in volume) can occur in a gel as the gel material expels interstitial fluid from the pore network of the gel. The ability and / or tendency of a gel material to swell and / or shrink in certain solvent environments will depend on the chemical properties of the polymer and solvent (e.g., solubility, hydrophobicity, pore structure, affinity) and the elasticity of the polymer network of the gel. The swelling ratio of a gel is a measure of the weight fraction increase in a gel due to fluid absorption (e.g., the weight gain of a hydrogel due to water absorption).

[0151] In certain embodiments, the polymer compositions of the present disclosure may have a therapeutically effective swelling ratio and / or water content. In certain embodiments, the polymer compositions may have a swelling ratio and / or water content that provides strong adhesion and high polymer composition retention on the target tissue of the subject. . In certain embodiments, the gel polymer compositions of the present disclosure may have a swelling ratio and / or water content that provides strong adhesion and high polymer composition retention on the target tissue of the subject. In certain embodiments, the gel polymer compositions may have a swelling ratio and / or water content that allows the polymer composition to maintain its shape, adhesion, connectivity and / or consistency on the surface of the target tissue for one or more hours, one or more days, or one or more weeks.

[0152] In certain embodiments, polymer composition can have a swelling ratio of about 5% to about 50%. In certain embodiments, polymer composition can have a swelling ratio of at least about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35% or about 40%. In certain embodiments, polymer composition can have a swelling ratio of no more than about 50%, about 45%, about 40%, about 35%, about 30%, about 25%, about 20%, about 15% or about 10%. In certain embodiments, polymer composition has a swelling ratio of about 25% or less, about 20% or less, about 15% or less, or about 10% or less. In certain embodiments, the polymer composition can have a swelling ratio of about 1-3%, about 3-6%, about 6-10%, about 1-5%, about 1-10%, about 5-10%, about 11-13%, about 13-16%, about 16-20%, about 10-20%, about 10-15%, about 15-20%, about 21-23%, about 23-26%, about 26-30%, about 20-30%, about 20-25%, about 25-30%, about 31-33%, about 33-36%, about 36-40%, about 30-40%, about 30-35%, about 35-40%, about 41-43%, about 43-46%, about 46-50%, about 40-50%, about 40-45%, or about 45-50%. In certain embodiments, the polymer composition may have about 1-3%, about 3-6%, about 6-10%, about 1-5%, about 1-10%, about 5-10%, about 11-13%, about 13-16%, about 16-20%, about 10-20%, about 10-15%, about 15-20%, about 21-23%, about 23-26%, about 26-30%, about 20-30 %, about 20-25%, about 25-30%, about 31-33%, about 33-36%, about 36-40%, about 30-40%, about 30-35%, about 35-40%, about 41-43%, about 43-46%, about 46-50%, about 40-50%, about 40-45% or about 45-50% short-term swelling ratio (i.e., the swelling ratio measured for about 1 to 24 hours).In certain embodiments, the polymer composition may have about 1-3%, about 3-6%, about 6-10%, about 1-5%, about 1-10%, about 5-10%, about 11-13%, about 13-16%, about 16-20%, about 10-20%, about 10-15%, about 15-20%, about 21-23%, about 23-26%, about 26-30%, about 20-3 0%, about 20-25%, about 25-30%, about 31-33%, about 33-36%, about 36-40%, about 30-40%, about 30-35%, about 35-40%, about 41-43%, about 43-46%, about 46-50%, about 40-50%, about 40-45% or about 45-50% mid-term swelling ratio (i.e., the swelling ratio is measured for about 1 to 7 days). In certain embodiments, the polymer composition can have about 1-3%, about 3-6%, about 6-10%, about 1-5%, about 1-10%, about 5-10%, about 11-13%, about 13-16%, about 16-20%, about 10-20%, about 10-15%, about 15-20%, about 21-23%, about 23-26%, about 26-30%, about 20-3 ...15%, about 15-20%, about 10-20%, about 10-15%, about 15-20%, about 21-23%, about A long-term swelling ratio (i.e., a swelling ratio measured for about 1 to 4 weeks or longer) of about 20-25%, about 25-30%, about 31-33%, about 33-36%, about 36-40%, about 30-40%, about 30-35%, about 35-40%, about 41-43%, about 43-46%, about 46-50%, about 40-50%, about 40-45%, or about 45-50%.

[0153] In certain embodiments, the hydrogel polymer composition may have a water content of about 5% to about 99%. In certain embodiments, the hydrogel polymer composition may have a water content of about 50% to about 99%. In certain embodiments, the hydrogel polymer composition may have a water content of about 65% to about 85%. In certain embodiments, the polymer composition may have a water content of at least about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, about 75%, or about 80%. In certain embodiments, the polymer composition can have a swelling ratio of about 99% or less, about 95% or less, about 90% or less, about 85% or less, about 80% or less, about 75% or less, about 70% or less, about 65% or less, about 60% or less, about 55% or less, about 50% or less, about 45% or less, about 40% or less, about 35% or less, or about 30% or less. In certain embodiments, the polymer composition can have about 1-3%, about 3-6%, about 6-10%, about 1-5%, about 5-10%, about 1-10%, about 11-13%, about 13-16%, about 16-20%, about 10-15%, about 15-20%, about 10-20%, about 21-23%, about 23-26%, about 26-30%, about 20-25%, about 25-30%, about 20-30%, about 31-33%, about 33-36%, about 36-40%, about 30-35%, about 35-40%, about 30-40%, about 41-43%, about 43-46%, about 46-50%, about 40-45%, about 45-50%, about 40-5 0%, about 51-53%, about 53-56%, about 56-60%, about 50-55%, about 55-60%, about 50-60%, about 61-63%, about 63-66%, about 66-70%, about 60-65%, about 65-70%, about 60-70%, about 71-73%, about 73-76%, about 76-80%, About 70-75%, about 75-80%, about 70-80%, about 81-83%, about 83-86%, about 86-90%, about 80-85%, about 85-90%, about 80-90%, about 91-93%, about 93-96%, about 96-99%, about 90-95%, about 95-99% or about 90-99% water content.

[0154] In certain embodiments, the hydrogel polymer compositions of the present disclosure allow for controlled and sustained release of one or more therapeutic agents over a period of time. In certain embodiments, the hydrogel polymer compositions allow for release of at least 1 μg / day, at least 2 μg / day, at least 3 μg / day, at least 4 μg / day, at least 5 μg / day, at least 6 μg / day, at least 7 μg / day, at least 8 μg / day, at least 9 μg / day, at least 10 μg / day, at least 11 μg / day, at least 12 μg / day, at least 13 μg / day, at least 14 μg / day, at least 15 μg / day, at least 16 μg / day, at least 17 μg / day, at least 18 μg / day, at least 19 μg / day, at least 20 μg / day, at least 25 μg / day, at least 30 μg / day, at least 31 μg / day, at least 32 μg / day, at least 33 μg / day, at least 34 μg / day, at least 35 μg / day, at least 36 μg / day, at least 37 μg / day, at least 38 μg / day, at least 39 μg / day, at least 40 μg / day, at least 41 μg / day, at least 42 μg / day, at least 43 μg / day, at least 44 μg / day, at least 45 μg / day, at least 46 μg / day, at least 47 μg / day, at least 48 μg / day, at least 49 μg / day, at least 50 μg / day, at least 51 μg / day, at least 52 μg / day, at least 53 μg / day, at least 54 μg / day, at least 55 μg / day, at least 56 μ In some embodiments, the hydrogel polymer composition allows release of at least 10 μg / day of the therapeutic agent.

[0155] Durability and degradation

[0156] In certain embodiments, the polymer compositions of the present disclosure may have a therapeutically effective polymer degradation rate (i.e., degradation rate). In certain embodiments, the polymer compositions may have a degradation rate that provides sustained adhesion and high polymer composition retention on the target tissue of the subject. In certain embodiments, the gel polymer compositions of the present disclosure may have a degradation rate that provides sustained adhesion and high polymer composition retention on the target tissue of the subject. In certain embodiments, the gel polymer compositions may have a degradation rate that allows the polymer composition to maintain its shape, adhesion, connectivity, and / or consistency on the surface of the target tissue for one or more hours, one or more days, or one or more weeks.

[0157] In certain embodiments, the polymer composition may have a degradation rate of 1 to 50 days. In certain embodiments, the polymer composition can have a degradation rate of about 1-3 days, about 3-6 days, about 6-10 days, about 1-5 days, about 1-10 days, about 5-10 days, about 11-13 days, about 13-16 days, about 16-20 days, about 10-20 days, about 10-15 days, about 15-20 days, about 21-23 days, about 23-26 days, about 26-30 days, about 20-30 days, about 20-25 days, about 25-30 days, about 31-33 days, about 33-36 days, about 36-40 days, about 30-40 days, about 30-35 days, about 35-40 days, about 41-43 days, about 43-46 days, about 46-50 days, about 40-50 days, about 40-45 days, or about 45-50 days.

[0158] Biocompatibility

[0159] In certain embodiments, the polymer compositions of the present disclosure are biocompatible with a target tissue of a subject. In certain embodiments, the biomechanical properties of the polymer compositions are similar to and / or biocompatible with the biomechanical properties of a target tissue of a subject (e.g., a cornea of ​​a subject).

[0160] In certain embodiments, the biocompatibility of the polymer composition can be demonstrated by a low inflammatory response in a target tissue or subject. In certain embodiments, the biocompatibility of the polymer composition can be demonstrated by the viability of cells from the target tissue that are implanted or incorporated into a portion of the polymer composition.

[0161] shape

[0162] In certain embodiments, the polymer compositions of the present disclosure can be formed into molded, stamped or shaped gel compositions. Molded, stamped or shaped hydrogels can be prepared using methods such as those described in US 20050008675 or US20040258729, each of which is incorporated herein by reference in its entirety to the extent that each document describes the composition, production (including molding), analysis and use of hydrogels (including acrylated gelatin polymeric compositions such as GelMA hydrogels).

[0163] In certain embodiments, the polymer compositions (e.g., hydrogel polymer compositions) of the present disclosure can be formed into cylinders, each cylinder having a length and a diameter. In certain embodiments, the polymer compositions (e.g., hydrogel polymer compositions) of the present disclosure can be conformable to the shape of a target surface. In certain embodiments, the polymer composition conforms to a convex, concave, or curved shape of a target surface.

[0164] In certain embodiments, the polymer composition can form a cylindrical rod. As used herein, "cylindrical rod" or "rod" describes a cylinder whose cylinder length is at least 3 times (3x) of the cylinder diameter. As a non-limiting example, a cylindrical rod can have: a length of about 3mm and a diameter of about 0.75mm; or a length of about 2.5mm and a diameter of about 0.75mm. In certain embodiments, the hydrogel rod of the present disclosure can be a length of about 3mm and a diameter of about 0.75mm. In certain embodiments, the hydrogel rod of the present disclosure can be a length of about 6mm and a diameter of about 0.75mm.

[0165] In certain embodiments, the polymer composition can be formed into a cylindrical disc. As used herein, a "cylindrical disc" or "disc" describes a cylinder having a cylinder diameter that is at least 2 times (2x) the length of the cylinder. As a non-limiting example, a cylindrical disc can have: a length of about 2.5 mm and a diameter of about 6 mm; or a length of about 2 mm and a diameter of about 6 mm.

[0166] III. Gel production

[0167] In certain embodiments, the polymer compositions of the present disclosure (e.g., GelMA polymer compositions) can be produced as described in the art, including Nichol et al., Biomaterials, 2010 Jul, 31(21):5536-44; Assmann et al., Biomaterials, 2017, 140:115-127; Noshadi et al., Biomater. Sci., 2017, 5:2093-2105; to the extent that each document describes the production of polymer compositions (including acryl gelatin polymer compositions such as GelMA hydrogels), each document is incorporated herein by reference in its entirety.

[0168] In certain embodiments, polymer composition of the present disclosure can be formed by two or more chemically modified gelatin components in the cross-linked precursor polymer composition to form a gel polymer composition. In certain embodiments, polymer composition of the present disclosure can be cross-linked, polymerized and / or gelled to form a gel polymer composition under moistening, aqueous and / or biological conditions. In certain embodiments, when exposed to specific cross-linking conditions (for example, acidic conditions, alkaline conditions, high salt conditions, low salt conditions, high temperature, stirring, dissolution conditions), the cross-linking of the two or more chemically modified gelatin components is initiated, promoted or realized. In certain embodiments, the cross-linking of the two or more chemically modified gelatin components is initiated, promoted or realized by a cross-linking agent. In certain embodiments, the cross-linking of the two or more chemically modified gelatin components is initiated, promoted or realized by a cross-linking agent under specific cross-linking conditions.

[0169] In certain embodiments, the present disclosure provides methods for producing a gel polymer composition, such as a hydrogel polymer composition. In certain embodiments, the present disclosure provides methods for producing a GelMA hydrogel polymer composition. Figure 2 A method 100 for producing a gel polymer composition is provided. In step 110, a precursor polymer composition is provided, which comprises a chemically modified gelatin (e.g., acryl-substituted gelatin, GelMA) having a crosslinkable group. In an optional step 115, one or more additional chemically modified polymer precursors (e.g., MeHA, PEGDA) having a crosslinkable group are added to the precursor polymer composition. In certain embodiments, the polymer composition comprises unmodified HA and / or unmodified PEG and / or unmodified tropoelastin. In step 120, a solution comprising one or more crosslinking agents and / or photoinitiators is added to the precursor polymer composition. In an optional step 125, a therapeutic agent, cells and / or particles (i.e., microparticles or nanoparticles) are added to the precursor polymer composition. In step 130, the precursor polymer composition is polymerized / crosslinked to produce a gel polymer composition.

[0170] In certain embodiments, the method for producing a gel polymer composition may include providing a precursor polymer composition comprising a chemically modified gelatin (e.g., acryloyl-substituted gelatin, GelMA) having a crosslinkable group. In certain embodiments, the chemically modified gelatin comprises acrylated gelatin. In certain embodiments, the chemically modified gelatin comprises methacrylated gelatin (i.e., GelMA).

[0171] In certain embodiments, the precursor polymer composition comprises one or more solvents or liquid vehicles, diluents, dispersion media, dispersants, granulating agents, binders, disintegrants, suspending agents, surfactants, emulsions or emulsifiers, isotonic agents, thickening agents, preservatives, solid binders, buffers, lubricants, colorants, coating agents, sweeteners, flavoring agents, fragrances, or combinations thereof.

[0172] In certain embodiments, the precursor polymer composition comprises one or more solvents. In certain embodiments, the solvent comprises an aqueous solvent. Examples of aqueous solvents include, but are not limited to, distilled water, deionized water, saline, Dulbecco's phosphate-buffered saline (DPBS) and Ringer's solution. In certain embodiments, the solvent comprises DPBS. In certain embodiments, the solvent comprises an organic solvent. Examples of organic solvents include, but are not limited to, hexane, benzene, toluene, acetone, ether, chloroform, methylene chloride, isopropanol, methanol, ethanol, n-propanol and n-butanol.

[0173] In certain embodiments, the precursor polymer composition can be a sprayable form. In certain embodiments, the precursor polymer composition can be a high viscosity form (e.g., a paste-like viscosity). In certain embodiments, the precursor polymer composition can be a low viscosity form (e.g., a liquid-like viscosity).

[0174] In certain embodiments, the method for producing a gel polymer composition may include the step of adding one or more additional chemically modified polymer precursors with crosslinkable groups to the precursor polymer composition. In certain embodiments, the method for producing a gel polymer composition may include: (i) providing a precursor polymer composition comprising a chemically modified gelatin (e.g., acryl-substituted gelatin, GelMA) with a crosslinkable group; and (ii) adding one or more additional chemically modified polymer precursors with crosslinkable groups to the precursor polymer composition. In certain embodiments, the one or more additional chemically modified polymer precursors include chemically modified hyaluronic acid, such as acryl-substituted hyaluronic acid. In certain embodiments, the chemically modified hyaluronic acid includes methacrylated hyaluronic acid (MeHA). In certain embodiments, the one or more additional chemically modified polymer precursors include chemically modified poly (ethylene glycol) (PEG), such as acryl-substituted PEG. In certain embodiments, the chemically modified hyaluronic acid includes poly (ethylene glycol) diacrylate (PEGDA).

[0175] In certain embodiments, the method for producing a gel polymer composition may include the step of adding one or more crosslinking agents and / or polymer crosslinking initiators (e.g., photoinitiators) to a precursor polymer composition. In certain embodiments, the method for producing a gel polymer composition may include: (i) providing a precursor polymer composition comprising a chemically modified gelatin having a crosslinkable group (e.g., acryl-substituted gelatin, GelMA); and (ii) adding one or more crosslinking agents and / or polymer crosslinking initiators (e.g., photoinitiators) to the precursor polymer composition. In certain embodiments, the method for producing a gel polymer composition may include: (i) providing a precursor polymer composition comprising a chemically modified gelatin having a crosslinkable group (e.g., acryl-substituted gelatin, GelMA); (ii) adding one or more additional chemically modified polymer precursors having a crosslinkable group to the precursor polymer composition, and (iii) adding one or more crosslinking agents and / or polymer crosslinking initiators (e.g., photoinitiators) to the precursor polymer.

[0176] In certain embodiments, one or more crosslinking agents and / or polymer crosslinking initiators (e.g., photoinitiators) may be added to the precursor polymer prior to adding one or more additional chemically modified polymer precursors having crosslinkable groups to the precursor polymer composition. In certain embodiments, a method for producing a gel polymer composition may include: (i) providing a precursor polymer composition comprising a chemically modified gelatin (e.g., acryl-substituted gelatin, GelMA) having crosslinkable groups; (ii) adding one or more crosslinking agents and / or polymer crosslinking initiators (e.g., photoinitiators) to the precursor polymer; and (iii) adding one or more additional chemically modified polymer precursors having crosslinkable groups to the precursor polymer composition.

[0177] In certain embodiments, the polymer composition comprises one or more polymer crosslinking initiators (e.g., a crosslinking initiator that forms free radicals when exposed to specific polymer crosslinking conditions, such as acidic conditions, alkaline conditions, high salt conditions, low salt conditions, high temperature, stirring, dissolution conditions, and exposure). In certain embodiments, the polymer composition comprises one or more photoinitiator components (i.e., a crosslinking initiator that is initiated or activated by absorbing light of a certain wavelength). In certain embodiments, the precursor polymer composition of the present disclosure comprises one or more photoinitiator components (i.e., a crosslinking initiator that is initiated or activated by visible light). In certain embodiments, the photoinitiator component can be activated by exposure. In certain embodiments, exposure can activate the photoinitiator to form free radicals, wherein the free radicals can cause bond formation between reactive groups in the composition, such as crosslinking of vinyl bonds between methacrylic acid groups in the GelMA polymer composition. Figure 3 An example of a series of reactions for producing a GelMA hydrogel polymer composition is provided, wherein: (i) a photoinitiator component is activated by light energy (hv) to form free radicals (R*), which then initiate bond formation between reactive groups on separate methacryloyl gelatin polymer precursors, thereby forming a crosslinked GelMA polymer network. The sequential reactions between reactive groups on the methacryloyl gelatin components will result in the formation of a broader GelMA hydrogel polymer composition.

[0178] In certain embodiments, the photoinitiator composition can be activated by being exposed to one or more light sources selected from the following: a visible light source (e.g., white light or blue light), an ultraviolet (UV) light source, a near infrared (NIR) light source, and a fluorescent light source. In certain embodiments, the photoinitiator composition includes a visible light activated photoinitiator, such as a visible light activated photoinitiator activated when exposed to a wavelength of about 380nm to about 740nm. In certain embodiments, the visible light activated photoinitiator can be activated when exposed to a wavelength of about 380-435nm (i.e., purple light), about 435-500nm (i.e., blue light), about 500-565nm (i.e., green light), about 565-600nm (i.e., yellow light), about 600-650nm (i.e., orange light) or about 650-740nm (i.e., red light). In certain embodiments, the photoinitiator composition includes a UV-activated photoinitiator. In certain embodiments, the photoinitiator composition includes a near infrared (NIR) light activated photoinitiator. In certain embodiments, the photoinitiator component comprises a white light activated photoinitiator. In certain embodiments, the photoinitiator component comprises a blue light activated photoinitiator.

[0179] In certain embodiments, the method for producing a gel polymer composition may include the step of adding one or more of a therapeutic agent and / or a particle (i.e., a microparticle or a nanoparticle) to a precursor polymer composition. In certain embodiments, one or more of the therapeutic agent and / or the particle may be added to the precursor polymer before one or more additional chemically modified polymer precursors having crosslinkable groups are added to the precursor polymer composition. In certain embodiments, one or more of the therapeutic agent and / or the particle may be added to the precursor polymer before one or more crosslinking agents and / or polymer crosslinking initiators (e.g., a photoinitiator) are added to the precursor polymer composition. In certain embodiments, the method for producing a gel polymer composition may include: (i) providing a precursor polymer composition comprising a chemically modified gelatin having a crosslinkable group (e.g., an acryl-substituted gelatin, GelMA); (ii) optionally adding one or more additional chemically modified polymer precursors having crosslinkable groups to the precursor polymer composition, (iii) adding one or more crosslinking agents and / or polymer crosslinking initiators (e.g., a photoinitiator) to the precursor polymer; and (iv) optionally adding one or more therapeutic agents and / or particles.

[0180] In certain embodiments, the precursor polymer composition can be clarified, purified or processed for quality and / or purity before any polymerization / cross-linking step. In certain embodiments, the precursor polymer composition can be filtered. In certain embodiments, the precursor polymer composition can be lyophilized. In certain embodiments, the precursor polymer composition can be stored frozen.

[0181] In certain embodiments, the method for producing a gel polymer composition may include the step of polymerizing / crosslinking a precursor polymer composition to produce a gel polymer composition. In certain embodiments, the method for producing a gel polymer composition may include: (i) providing a precursor polymer composition comprising a chemically modified gelatin (e.g., acryl-substituted gelatin, GelMA) having a crosslinkable group; (ii) optionally adding one or more additional chemically modified polymer precursors having a crosslinkable group to the precursor polymer composition, (iii) adding one or more crosslinking agents and / or polymer crosslinking initiators (e.g., photoinitiators) to the precursor polymer; (iv) optionally adding one or more therapeutic agents and / or particles; and (v) polymerizing / crosslinking the precursor polymer composition to produce a gel polymer composition.

[0182] In certain embodiments, crosslinking of the chemically modified gelatin component and any additional chemically modified polymer precursors (e.g., MeHA, PEGDA) is initiated, promoted, or achieved by exposure to UV or visible light in the presence of a photoinitiator component. In certain embodiments, exposure to UV or visible light in the presence of a photoinitiator causes the acryloyl groups on one chemically modified gelatin molecule to react with the acryloyl groups on other chemically modified gelatin molecules to crosslink the acryloyl-substituted gelatin component and produce a gel (e.g., a hydrogel). In certain embodiments, exposure to visible light in the presence of a photoinitiator causes the methacryloyl groups on one methacryloyl gelatin molecule to react with the methacryloyl groups on other methacryloyl gelatin molecules to crosslink the methacryloyl-substituted gelatin component and produce a methacrylylated gelatin (GelMA) hydrogel.

[0183] In certain embodiments, the polymer composition is exposed to the light source for a duration of 1-60 minutes. In certain embodiments, the polymer composition is exposed to the light source for a duration of 1 minute or longer, 5 minutes or longer, 10 minutes or longer, 15 minutes or longer, 20 minutes or longer, 25 minutes or longer, or 30 minutes or longer. In certain embodiments, the polymer composition is exposed to the light source for a duration of 1 minute or less, 5 minutes or less, 10 minutes or less, 15 minutes or less, 20 minutes or less, 25 minutes or less, or 30 minutes or less, 35 minutes or less, or 40 minutes or less. In certain embodiments, the polymer composition is exposed to the light source for a duration of about 5 seconds, about 10 seconds, about 15 seconds, about 20 seconds, about 25 seconds, about 30 seconds, about 35 seconds, about 40 seconds, about 45 seconds, about 50 seconds, about 55 seconds, about 60 seconds, about 65 seconds, about 70 seconds, about 75 seconds, about 80 seconds, about 85 seconds, about 90 seconds, about 95 seconds, about 100 seconds, about 105 seconds, about 110 seconds, about 115 seconds, about 120 seconds, about 3 minutes, about 4 minutes, about 5 minutes, about 6 minutes, about 7 minutes, about 8 minutes, about 9 minutes, about 10 minutes, about 11 minutes, about 12 minutes, about 13 minutes, about 14 minutes, about 15 minutes, about 16 minutes, about 17 minutes, about 18 minutes, about 19 minutes , about 20 minutes, about 21 minutes, about 22 minutes, about 23 minutes, about 24 minutes, about 25 minutes, about 26 minutes, about 27 minutes, about 28 minutes, about 29 minutes, about 30 minutes, about 31 minutes, about 32 minutes, about 33 minutes, about 34 minutes, about 35 minutes, about 36 minutes, about 37 minutes, about 38 minutes, about 39 minutes, about 40 minutes, about 41 minutes, about 42 minutes, about 43 minutes, about 44 minutes, about 45 minutes, about 46 minutes, about 47 minutes, about 48 minutes, about 49 minutes, about 50 minutes, about 51 minutes, about 52 minutes, about 53 minutes, about 54 minutes, about 55 minutes, about 56 minutes, about 57 minutes, about 58 minutes, about 59 minutes or about 60 minutes.In certain embodiments, the polymer composition is exposed to the light source for a duration of about 1-3 minutes, about 3-6 minutes, about 6-10 minutes, about 1-5 minutes, about 1-10 minutes, about 5-10 minutes, about 11-13 minutes, about 13-16 minutes, about 16-20 minutes, about 10-20 minutes, about 10-15 minutes, about 15-20 minutes, about 21-23 minutes, about 23-26 minutes, about 26-30 minutes, about 20-30 minutes, about 20-25 minutes. , about 25-30 minutes, about 31-33 minutes, about 33-36 minutes, about 36-40 minutes, about 30-40 minutes, about 30-35 minutes, about 35-40 minutes, about 41-43 minutes, about 43-46 minutes, about 46-50 minutes, about 40-50 minutes, about 40-45 minutes, about 45-50 minutes, about 51-53 minutes, about 53-56 minutes, about 56-60 minutes, about 50-60 minutes, about 50-55 minutes or about 55-60 minutes.

[0184] In certain embodiments, the polymer composition may have a thickness of about 1 μm to about 10000 μm. In certain embodiments, the polymer composition may have a thickness of about 1-50 μm, about 50-100 μm, about 100-150 μm, about 150-200 μm, about 200-250 μm, about 250-300 μm, about 300-350 μm, about 350-400 μm, about 400-450 μm, about 450-400 μm, about 400- 450μm, about 450-500μm, about 500-550μm, about 550-600μm, about 600-650μm, about 650-700μm, about 700-750μm, about 750-800μm, about 800-850μm, about 850-900μm, about 900-950μm, about 950-1000μm, about 10 00-1500μm, about 1500-2000μm, about 2000-2500μm, about 2500-3000μm, about 3000-3500μm, about 3500-4000μm, about 4000-4500μm, about 4500-4000μm, about 4000-4500μm, about 4500-5000μm, about 50 The thickness of the film may be about 100-5500 μm, about 1000-6000 μm, about 1000-6500 μm, about 10000-7000 μm, about 10000-7500 μm, about 10000-8000 μm, about 10000-8500 μm, about 10000-9000 μm, about 10000-9500 μm or about 10000-10000 μm.

[0185] In certain embodiments, the precursor polymer composition may be cooled before or during the cross-linking reaction. In certain embodiments, the precursor polymer composition may be cooled to a temperature of about 0°C to about 30°C before or during the cross-linking reaction. In certain embodiments, the precursor polymer composition may be cooled to a temperature of about 0-5°C, about 5-10°C, about 0-10°C, about 10-15°C, about 15-20°C, about 10-20°C, about 20-25°C, about 25-30°C, or about 20-30°C. In certain embodiments, the precursor polymer composition may be heated before or during the cross-linking reaction. In certain embodiments, the precursor polymer composition may be heated to a temperature of about 30°C to about 150°C before or during the cross-linking reaction. In certain embodiments, the precursor polymer composition can be heated to about 30-35°C, about 35-40°C, about 30-40°C, about 40-45°C, about 45-50°C, about 40-50°C, about 50-55°C, about 55-60°C, about 50-60°C, about 60-65°C, about 65-70°C, about 60-70°C, about 70-75°C, about 75-80°C, about 70-80°C, about 80-85°C, about 85-90°C. , a temperature of about 80-90°C, about 90-95°C, about 95-100°C, about 90-100°C, about 100-105°C, about 105-110°C, about 100-110°C, about 110-115°C, about 115-120°C, about 110-120°C, about 130-135°C, about 135-140°C, about 130-140°C, about 140-145°C, about 145-150°C or about 140-150°C.

[0186] Once the cross-linking reaction is completed or stopped, the resulting gel polymer material can be clarified, purified or processed for quality, purity and / or therapeutic activity. In certain embodiments, the gel polymer composition can be dialyzed to remove any unreacted compounds from the gel mixture or structure. In certain embodiments, the gel polymer composition can be dialyzed with a dialysis buffer comprising deionized water. In certain embodiments, the gel polymer composition can be filtered. In certain embodiments, the gel polymer composition can be dried. In certain embodiments, the gel polymer composition can be lyophilized. In certain embodiments, the gel polymer composition can be stored frozen.

[0187] In certain embodiments, the polymer compositions of the present disclosure can be formed, molded, extruded, woven, or otherwise produced or processed into fibers, films, discs, fabrics, tubes, conduits, rods, rings, nets, or any other form or shape of polymer or gel materials known in the art. In certain embodiments, the polymer compositions of the present disclosure can be formed, molded, extruded, woven, or otherwise produced or processed into single-layer structures or multilayer structures (e.g., two layers, three layers, four layers, etc.).

[0188] In certain embodiments, the polymer composition of the present disclosure is included in the macromolecular polymer and / or fiber component that is interwoven or entangled in the interstitial porous network of the polymer composition but is not chemically connected to the core cross-linked polymer network. Non-limiting examples of such macromolecules include polycaprolactone, gelatin, gelatin methacrylate, alginate, alginate methacrylate, chitosan, chitosan methacrylate, ethylene glycol chitosan, ethylene glycol chitosan methacrylate, hyaluronic acid, hyaluronic acid methacrylate and other non-crosslinked natural or synthetic polymer chains. The gel material including the interwoven macromolecular structure can be referred to as a composite structure or composite gel. Examples of hydrogel / fiber composite materials are described, for example, in Moutos et al. Nat. Mater., 2007, 6 (2), pp. 162-7; to the extent that the document describes the composition, production, analysis and use of composite gel materials, the document is incorporated herein by reference as a whole. In certain embodiments, the precursor polymer composition can be in a high viscosity form (e.g., paste-like viscosity) and incorporated into a macromolecular polymer matrix (e.g., a fiber mat or a tissue matrix). In certain embodiments, the precursor polymer composition can be in a low viscosity form (eg, liquid-like viscosity) and incorporated into a macromolecular polymer matrix (eg, a fiber mat or a tissue matrix).

[0189] In certain embodiments, the crosslinked polymer composition may have a substantially covalent matrix form. In certain embodiments, the crosslinked polymer composition may have an amorphous matrix form (ie, a matrix formed primarily by ionic bonding and / or hydrogen bonding).

[0190] In certain embodiments, the polymer compositions of the present disclosure can be formed into patterned gel compositions (e.g., micropatterned hydrogels). Micropatterned hydrogels can be prepared using methods such as those described in US 6,423,252, which is incorporated herein by reference in its entirety to the extent that it describes the composition, production (including micropatterning), analysis, and use of hydrogels (including acrylated gelatin polymer compositions such as GelMA hydrogels). For example, the method includes: (i) contacting a precursor polymer composition with a mold or surface comprising a three-dimensional negative configuration (i.e., a template) of a micropattern; and (ii) crosslinking and / or polymerizing the precursor polymer composition to produce a crosslinked gel polymer composition (e.g., GelMA hydrogel) that includes a micropattern at least on the surface of the hydrogel.

[0191] In certain embodiments, the polymer compositions of the present disclosure can be formed into molded, stamped or shaped gel compositions. Molded, stamped or shaped hydrogels can be prepared using methods such as those described in US 20050008675 or US20040258729, each of which is incorporated herein by reference in its entirety to the extent that each document describes the composition, production (including molding), analysis and use of hydrogels (including acrylated gelatin polymeric compositions such as GelMA hydrogels).

[0192] IV. Administration and Treatment

[0193] General

[0194] Suturing, tissue grafting, and the use of tissue adhesives are common treatments for soft tissue (such as corneal or scleral tissue) defects and / or trauma. However, each treatment carries risks and complications: (i) suturing requires advanced surgical techniques and early treatment, it often produces irregular scars, and can often lead to microbial entrapment and infection; (ii) tissue implantation and transplantation require donor tissue (with high costs), advanced surgical techniques, and present a high risk of immune response or complete rejection of the implanted tissue; (iii) tissue adhesives (such as cyanoacrylate glues, fibrin glues, or polyethylene glycol (PEG)-based sealants) have limited effectiveness and adhesion (especially in aqueous and physiological environments), limited durability, can be difficult to apply and control texture, have a high potential for leakage, lack biocompatibility (such as inflammatory) and can be toxic, lack translucency / transparency, have a high risk of infection (including risks associated with high porosity), and are generally not FDA-approved for safety to reduce corneal defects or repair significant corneal incisions, perforations, or trauma.

[0195] There remains a need for improved polymer compositions that are effective in treating and / or sealing injuries, defects, and diseases of soft tissue (ie, body tissue other than bone) of a subject.

[0196] In certain embodiments, the polymer compositions of the present disclosure can be used as sealant compositions for treating or repairing soft tissue of a subject. In certain embodiments, the polymer compositions of the present disclosure can be used as delivery vehicles for administering therapeutic agents to treat or repair soft tissue of a subject. In certain embodiments, the polymer compositions of the present disclosure can be used as sealant compositions for treating or repairing soft tissue of a subject, and as delivery vehicles for administering therapeutic agents to treat or repair soft tissue of a subject.

[0197] In certain embodiments, the method and composition of the present disclosure can be used for adhesion, sealing or treating the target soft tissue of subject.In certain embodiments, the method and composition of the present disclosure can be used for adhesion, sealing or treating one or more target soft tissues selected from the following: adipose tissue, bladder tissue, bone marrow, cardiovascular tissue (for example, heart tissue), dura mater, endocrine gland, gastrointestinal tissue, hair follicle, kidney tissue, liver tissue, lung tissue, lymph node, muscle tissue, nervous system / nerve tissue (for example, peripheral nervous system), eye tissue (for example, corneal tissue), oral tissue (for example, craniofacial tissue, tooth tissue, periodontal tissue), pancreatic tissue, kidney tissue, skin tissue (for example, for treating local ulcer, such as diabetic ulcer), urethral tissue, vascular tissue.In certain embodiments, the method and composition of the present disclosure can be used for adhesion, sealing or treating one or more target soft tissues in stress and / or physiological environment, or need elasticity and / or similar application of adhesion composition.

[0198] The polymer compositions of the present disclosure (eg, GelMA polymer compositions) can be administered by any route that produces a therapeutically effective outcome.

[0199] In certain embodiments, the method includes applying a pregelled polymer composition to an applicator; placing an applicator containing the pregelled polymer composition on the surface of a subject's target tissue; and crosslinking (e.g., photocrosslinking) the polymer composition by exposing the pregelled polymer composition to crosslinking conditions (e.g., visible light and a photoinitiator). In certain embodiments, the pregelled polymer composition is applied directly to the surface of the target tissue without an applicator. In certain embodiments, application to the surface of the target tissue includes application to the outer surface of the target tissue (e.g., topical application). In certain embodiments, application to the surface of the target tissue includes application / injection into the space just below the surface of the target tissue (e.g., subconjunctival application to eye tissue, subretinal application to eye tissue).

[0200] In certain embodiments, the target soft tissue can be treated or closed by applying a first layer comprising a first polymer composition of the present invention, the first polymer composition being engineered to have specific physical, mechanical, structural, chemical and / or biological properties (e.g., elasticity, biodegradability, porosity); and then applying a second layer comprising a second polymer composition, the second polymer composition being engineered to have different physical, mechanical, structural, chemical and / or biological properties (e.g., elasticity, biodegradability, porosity). In certain embodiments, the method can include applying one or more additional layers (e.g., a third layer, a fourth layer, etc.), each layer comprising a polymer composition of the present disclosure, the polymer composition being engineered to have specific physical, mechanical, structural, chemical and / or biological properties (e.g., elasticity, biodegradability, porosity).

[0201] In certain embodiments, the target soft tissue can be treated by: (i) forming a preformed polymer composition by polymerizing the polymer composition of the present disclosure; and (ii) applying the preformed polymer composition to the surface or subsurface (e.g., subconjunctival, subretinal) of the target tissue of the subject. In certain embodiments, application to the surface of the target tissue (e.g., on the surface or subsurface) includes application / injection to the space just below the surface of the target tissue (e.g., subconjunctival application to eye tissue, subretinal application to eye tissue). In certain embodiments, the preformed polymer composition can be engineered to have specific physical, mechanical, structural, chemical and / or biological properties (e.g., elasticity, biodegradability, porosity).

[0202] In certain embodiments, the target soft tissue can be treated by: (i) forming a preformed hydrogel polymer composition by polymerizing a polymer composition of the present disclosure; (ii) drying the hydrogel polymer by removing a majority of the interstitial fluid from the hydrogel (e.g., at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% of the interstitial fluid); (iii) applying the preformed polymer composition to the surface or subsurface of the target tissue of the subject (e.g., subconjunctival, subretinal); and (iv) optionally rehydrating the dried hydrogel polymer to a substantially hydrated form (e.g., at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% of the volume of the interstitial fluid). In certain embodiments, applying to the surface of the target tissue (e.g., on the surface or subsurface) includes applying / injecting into the space just below the surface of the target tissue (e.g., subconjunctival application to ocular tissue, subretinal application to ocular tissue). In certain embodiments, preformed polymer compositions can be engineered to have specific physical, mechanical, structural, chemical, and / or biological properties (eg, elasticity, biodegradability, porosity).

[0203] Therapeutic Compositions

[0204] In certain embodiments, the polymer composition of the present disclosure can be prepared into a therapeutic composition or included in a therapeutic composition. In certain embodiments, the hydrogel polymer composition of the present disclosure can be prepared into a therapeutic composition or included in a therapeutic composition. In certain embodiments, the GelMA hydrogel polymer composition of the present disclosure can be prepared into a therapeutic composition or included in a therapeutic composition. Such compositions include one or more polymer compositions of the present disclosure (optionally including one or more therapeutic agents or active ingredients) and one or more therapeutically acceptable excipients (e.g., carriers, solvents or delivery vehicles).

[0205] According to the relative amounts of the polymer composition (e.g., GelMA hydrogel polymer composition), the therapeutically acceptable excipient and / or any additional ingredients in the therapeutic composition of the present disclosure, it can vary, depending on the characteristics, size and / or condition of the subject or tissue being treated, and further depending on the route of administration or application of the composition. In certain embodiments, the therapeutic composition comprises 0.1% to 99% (w / v) of the polymer composition of the present disclosure by volume of the therapeutic composition. In certain embodiments, the therapeutic composition comprises a polymer composition of the present disclosure at a weight / volume concentration (w / v) of about 0.5%, about 1%, about 2%, about 3%, about 4%, about 5%, about 6%, about 7%, about 8%, about 9%, about 10%, about 11%, about 12%, about 13%, about 14%, about 15%, about 16%, about 17%, about 18%, about 19%, about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about about 20%, about 21%, about 22%, about 23%, about 24%, about 25%, about 26%, about 27%, about 28%, about 29%, about 30%, about 31%, about 32%, about 33%, about 34%, about 35%, about 36%, about 37%, about 38%, about 39%, about 40%, about 41%, about 42%, about 43%, about 44%, about 45%, about 46%, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 78%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about %, about 47%, about 48%, about 49%, about 50%, about 51%, about 52%, about 53%, about 54%, about 55%, about 56%, about 57%, about 58%, about 59%, about 60%, about 61%, about 62%, about 63%, about 64%, about 65%, about 66%, about 67%, about 68%, about 69%, about 70%, about 71%, about 72%, about 73%, about 74%, about 75%, about 76%, about 77%, about 88%, about 79%, about 80%, about 81%, about 82%, about 83%, about 84%, about 85%, about 86%, about 87%, about 88%, about 89%, about 90%, about 91%, about 92%, about 93%, about 94%, about 95%, about 96%, about 97%, about 98% or about 99%.In certain embodiments, the therapeutic composition comprises a polymer composition of the present disclosure at a weight / volume concentration (w / v) of about 1-3%, about 3-6%, about 6-10%, about 1-5%, about 5-10%, about 1-10%, about 11-13%, about 13-16%, about 16-20%, about 10-15%, about 15-20%, about 10-20%, about 21-23%, about 23-26%, about 26-30%, about 20-25%, about 25-30%, about 20-30%, about 31-33%, about 33-36%, about 36-40%, about 30-35%, about 35-40%, about 30-40%, about 41-43%, about 43-46%, about 46-50%, about 40-45%, about about 45-50%, about 40-50%, about 51-53%, about 53-56%, about 56-60%, about 50-55%, about 55-60%, about 50-60%, about 61-63%, about 63-66%, about 66-70%, about 60-65%, about 65-70%, about 60-70%, about 71-73%, about 73-76%, About 76-80%, about 70-75%, about 75-80%, about 70-80%, about 81-83%, about 83-86%, about 86-90%, about 80-85%, about 85-90%, about 80-90%, about 91-93%, about 93-96%, about 96-99%, about 90-95%, about 95-99% or about 90-99%.

[0206] In certain embodiments, the therapeutic compositions and formulations of the present disclosure include, but are not limited to, saline, liposomes (e.g., unilamellar vesicles, multilamellar vesicles), lipid particles (including microparticles and nanoparticles), and / or polymer particles (including microparticles and nanoparticles). In certain embodiments, the therapeutic compositions and formulations of the present disclosure include polymer compositions of the present disclosure incorporated with, but are not limited to, saline, liposomes, lipid particles (including microparticles and nanoparticles), polymer particles (including microparticles and nanoparticles), or combinations thereof.

[0207] In certain embodiments, the therapeutic compositions and formulations of the present disclosure are aqueous formulations (ie, formulations comprising water). In certain embodiments, the therapeutic compositions and formulations of the present disclosure comprise water, sterile water, or water for injection (WFI).

[0208] In certain embodiments, the therapeutic compositions and formulations of the present disclosure include one or more of the following: pH buffer solutions (e.g., phosphate buffered saline (PBS), HEPES, TES, MOPS), isotonic saline, Ringer's solution, polyols (e.g., glycerol, propylene glycol, liquid polyethylene glycol), alginic acid, ethanol, and therapeutically acceptable mixtures thereof. In certain embodiments, the therapeutic compositions and formulations of the present disclosure include phosphate buffered saline (PBS).

[0209] The formulations of the present disclosure may be used in any step of producing, processing, preparing, storing, extending or applying the polymer compositions of the present disclosure.

[0210] In certain embodiments, the therapeutic compositions of the present disclosure include one or more therapeutically acceptable excipients (e.g., vehicles capable of suspending or dissolving polymer compounds). Excipients may include, for example, antiadherents, antioxidants, binders, coatings, compression aids, disintegrants, dyes (pigments), emollients, emulsifiers, fillers (diluents), film formers or coatings, flavoring agents, fragrances, glidants (flow enhancers), lubricants, preservatives, printing inks, adsorbents, suspending agents or dispersants, sweeteners, and hydration water. Exemplary excipients include, but are not limited to, acetic acid, aluminum stearate, butylated hydroxytoluene (BHT), calcium carbonate, calcium chloride, calcium phosphate (dibasic), calcium stearate, carboxymethylcellulose, cross-linked carboxymethylcellulose, cross-linked polyvinyl pyrrolidone, citric acid, cross-linked polyvinylpyrrolidone, cysteine, ethylcellulose, gelatin, glucose, glucuronic acid, gluconic acid, hydroxypropylcellulose, hydroxypropylmethylcellulose, hydroxysuccinic acid, inositol, lactose, magnesium chloride, magnesium stearate, maltitol, mannitol, methionine, methylcellulose, Vitamin C, methylparaben, microcrystalline cellulose, phosphoric acid, polyethylene glycol, polyvinyl pyrrolidone, povidone, pregelatinized starch, propylparaben, retinyl palmitate, sucrose, shellac, silicon dioxide, sodium acetate, sodium carbonate, sodium bicarbonate, sodium carboxymethylcellulose, sodium chloride, sodium citrate, sodium hydroxide, sodium phosphate, sodium starch glycolate, sorbitol, starch (corn), stearic acid, sucrose, talc, titanium dioxide, vitamin A, vitamin E, vitamin C, xylitol, zinc stearate, and combinations thereof.

[0211] Therapeutic agents

[0212] In certain embodiments, the polymeric compositions of the present disclosure may comprise a therapeutic agent. In certain embodiments, the polymeric compositions of the present disclosure may comprise a therapeutic agent as a delivery payload.

[0213] In certain embodiments, the polymer compositions of the present disclosure may include a therapeutic agent at a concentration (w / v) of about 0% to about 40%. In certain embodiments, the precursor polymer compositions of the present disclosure may include a therapeutic agent at a concentration (w / v) of about 0% to about 40%. In certain embodiments, the gel polymer compositions of the present disclosure may include a therapeutic agent at a concentration (w / v) of about 0% to about 40%. In certain embodiments, the polymer compositions of the present disclosure may include a therapeutic agent at a concentration (w / v) of about 1-2%, about 2-4%, about 4-6%, about 6-8%, about 8-10%, about 1-5%, about 5-10%, about 1-10%, 10-12%, about 12-14%, about 14-16%, about 16-18%, about 18-20%, about 10-15%, about 15-20%, about 10-20%, about 20-22%, about 22-24%, about 24-26%, about 26-28%, about 28-30%, about 20-25%, about 25-30%, about 20-30%, about 30-32%, about 32-34%, about 34-36%, about 36-38%, about 38-40%, about 30-35%, about 35-40% or about 30-40% of the therapeutic agent.

[0214] In certain embodiments, the precursor polymer composition of the present disclosure may include a therapeutic agent at a concentration of about 0.1 mg / mL to about 500 mg / mL. In certain embodiments, the polymer composition of the present disclosure may include a therapeutic agent at a concentration of about 0.1-0.5 mg / mL, about 0.5-1.0 mg / mL, about 1.0-2.5 mg / mL, about 2.5-5.0 mg / mL, about 5.0-10.0 mg / mL, about 10.0-25.0 mg / mL, about 25.0-50.0 mg / mL, about 50.0-100.0 mg / mL, about 100-150 mg / mL, about 150-200 mg / mL, about 200-250 mg / mL, about 250-300 mg / mL , about 300-350 mg / mL, about 350-400 mg / mL, about 400-450 mg / mL, about 450-500 mg / mL, about 500-550 mg / mL, about 550-600 mg / mL, about 600-650 mg / mL, about 650-700 mg / mL, about 700-750 mg / mL, about 750-800 mg / mL, about 800-850 mg / mL, about 850-900 mg / mL, about 900-950 mg / mL or about 950-1000 mg / mL of a therapeutic agent.

[0215] In certain embodiments, polymer composition can deliver therapeutic agent in less than 1 hour and reach peak concentration.In certain embodiments, polymer composition can deliver therapeutic agent in less than 1 day and reach peak concentration.In certain embodiments, polymer composition can deliver therapeutic agent in less than about 0-2 hour, about 2-4 hour, about 4-6 hour, about 6-8 hour, about 8-10 hour, about 10-12 hour, about 12-16 hour, about 16-20 hour, about 20-24 hour, about 24-30 hour, about 30-36 hour, about 36-42 hour or about 42-48 hour and reach peak concentration.In certain embodiments, polymer composition can deliver therapeutic agent in less than 1 week and reach peak concentration. In certain embodiments, polymer composition can deliver therapeutic agent to reach peak concentration in about 0-2 days, about 2-4 days, about 4-6 days, about 6-8 days, about 8-10 days, about 10-12 days, about 12-16 days, about 16-20 days, about 20-24 days, about 24-30 days, about 30-35 days, about 35-40 days, about 40-45 days, about 45-50 days, about 50-55 days, about 55-60 days. In certain embodiments, polymer composition can deliver therapeutic agent to reach peak concentration in less than 1 month. In certain embodiments, polymer composition can deliver therapeutic agent to reach peak concentration in less than 12 months. In certain embodiments, the polymer composition can deliver a therapeutic agent to peak concentration within about 0-1 month, about 1-2 months, about 2-3 months, about 3-4 months, about 4-5 months, about 5-6 months, about 6-7 months, about 7-8 months, about 8-9 months, about 9-10 months, about 10-11 months, or about 11-12 months.

[0216] In certain embodiments, the therapeutic agent comprises one or more of the following: growth factors, hemostats, analgesics, anesthetics, antifungals, antibiotics, antibacterials, anti-inflammatory agents, antimicrobials, anthelmintics, antidotes, antiemetics, antihistamines, antihypertensives, antimalarials, antimicrobials, antipsychotics, antipyretics, antiseptics, antiarthritics, antituberculosis agents, antitussives, antivirals, cardiovascular active drugs, laxatives, chemotherapeutic agents, colored or fluorescent imaging agents, corticosteroids (such as steroids), antidepressants, sedatives, diagnostic aids, diuretics, enzymes, expectorants, hormones, hypnotics, immunosuppressants, minerals, nutritional supplements, parasympathomimetics, potassium supplements, radiation Sensitizers, radioisotopes, sedatives, sulfonamides, stimulants, sympathomimetics, tranquilizers, urinary tract anti-infectives, vasoconstrictors, vasodilators, vitamins, xanthine derivatives, organic molecules, small molecule inhibitors, glycosaminoglycans, organometallic agents, chelated metals or metal salts, peptide-based drugs, vitamins, nutritional supplements, glycoproteins (e.g., collagen), extracellular matrix proteins or fragments thereof, fibronectin, peptides and / or proteins, polysaccharides, carbohydrates (simple and / or complex), proteoglycans, antigens, oligonucleotides (sense and / or antisense DNA and / or RNA), antibodies, nucleic acid sequences, gene therapy agents, triamcinolone acetonide, ovalbumin, or combinations thereof.

[0217] In certain embodiments, the therapeutic agent comprises one or more anti-acanthamoeba agents, antiviral agents and / or antibacterial agents. In certain embodiments, the therapeutic agent comprises one or more agents selected from the following: acyclovir, valacyclovir, famciclovir, penciclovir, trifluridine, adenosine, hydroxychloroquine, gatifloxacin, daptomycin, tigecycline, telavancin, chloramphenicol, fusidic acid, chlorhexidine, polyhexamethylene biguanide, propamidine, hexamidine, bacitracin, metronidazole, rifampicin, ethambutol, streptomycin, isoniazid, silver nanoparticles, copper oxide nanoparticles, glicopeptide (e.g., teicoplanin, vancomycin), aminoglycosides (e.g., gentamycin, tobramycin, amikacin, netilmicin, imicin), cephalosporins (e.g., cefazolin, cefoxitin, cefotaxime, cefuroxime, moxalactam), macrolides (e.g., erythromycin), oxazolidinones (e.g., linezolid), quinolones, polymyxins, sulfonamides, tetracyclines, penicillins, carbapenems, monobactams, lincosides, spectinomycin, clindamycin, ansamycin, daptomycin, nitrofurans, trimethoprim-sulfamethoxazole, chitosan, penicillins, ciprofloxacin, or a combination thereof.

[0218] In certain embodiments, the therapeutic agent comprises one or more antifungal agents. In certain embodiments, the therapeutic agent comprises one or more agents selected from the following: amphotericin B, natamycin, candicin, filipin, hamycin, nystatin, rimocidin, voriconazole, imidazoles, triazoles, thiazoles, allylamines, echinocandins, benzoic acid, ciclopirox, flucytosine, griseofulvin, haloprogin, tolnaftate, undecylenic acid and povidone iodine or a combination thereof.

[0219] In certain embodiments, the therapeutic agent comprises one or more antimicrobial agents. In certain embodiments, the therapeutic agent comprises one or more antimicrobial agents selected from the group consisting of polymyxin B, vancomycin, cholera toxin, diphtheria toxin, lysostaphin, hemolysin, bacitracin, boceprevir, albavancin, daptomycin, enfuvirtide, oritavancin, teicoplanin, telaprevir, telavancin, guavanin 2, Maximin H5, dermcidin, cecropin, andropin, moricin, keratin toxin, bee venom, magainin, dermaseptin, brevinin-1, esculentin, buforin II, CAP18, LL37, baecin, apidaecin, prophenin, indolicidin, antimicrobial peptides (AMPs) (e.g., Tet213), chlorhexidine, chlorhexidine salts, triclosan n), polymyxins, tetracyclines, aminoglycosides (e.g., gentamicin, tobramycin), rifampin, erythromycin, neomycin, chloramphenicol, miconazole, quinolones, penicillins, fusidic acid, cephalosporins, mupirocin, metronidazole, secropin, protegrins, bacteriolcins, defensins, nitrofurazone, mafenide, aracyclovir, clindamycin, lincomycin, sulfonamides, norfloxacin, pefloxacin, nalidizic acidacid), cinnamycin, anti-DEFA5, duramycin, nisin, pediocin, Abaecin, Ct-AMP1, Apidaecin IA, Apidaecin IB, Bactenecin, Bactenecin 5, Bactenecin 7, Bactenecin B-2, Aurein family, SMAP-29, Temporin B, Pleurocidin, Tachyplesin III, LL-37, Citropin 1.1, BMAP-27, BMAP-28, Agelaia-MP, Temporin 1Ola, NA-CATH, Histatin, Latarcin, Halocidin, Bombinin, Cathelicidin, Malacidin, MP196, MS100a7a15, Murepavadin, Myticin, Mytilin, Paenibacterin, Pardaxin, Peptaibol, SAAP-148, Sarcotoxin, Stomoxyn, Tachyplesin, Thioether-containing protein 1, Thionin, Acetoxin Alamethicin, Arenicin, dermorphin, deltorphin, dermaseptin, pseudon, bombesin, maculatin, LEAP2, Efrapeptin, Arylomycin, Capreomycin, Gramicidin B, Antiamoebin, Bacillomycin, Teixobactin, Tyrothricin, Viomycin, oxalic acid, or a combination thereof.

[0220] In certain embodiments, the therapeutic agent comprises one or more anti-inflammatory agents. In certain embodiments, the therapeutic agent comprises one or more anti-inflammatory agents selected from the group consisting of steroidal anti-inflammatory drugs (e.g., prednisolone), corticosteroids (e.g., loteprednol etabonate), salicylates, nonsteroidal anti-inflammatory drugs (e.g., bromfenac), mTOR inhibitors, calcineurin inhibitors, synthetic or natural anti-inflammatory proteins, dexamethasone, 5-fluorouracil, daunorubicin, paclitaxel, curcumin, resveratrol, mitomycin, methylprednisolone, prednisolone, hydrocortisone, fludrocortisone, prednisone, celecoxib, ketorolac, piroxicam, diclofenac, ibuprofen and ketoprofen, rapamycin, cyclosporin, tacrolimus / FK-506, or a combination thereof.

[0221] In certain embodiments, the therapeutic agent comprises one or more growth factors. In certain embodiments, the therapeutic agent comprises a growth factor, and the growth factor comprises recombinant hepatocyte growth factor or recombinant nerve growth factor. In certain embodiments, the therapeutic agent comprises one or more growth factors selected from the following: activin (e.g., activin A, activin B, activin AB), adrenomedullin (AM), albumin, α-2 macroglobulin, annexin, angiopoietin (Ang), Artemin (Artemin), autocrine motility factor, bone morphogenetic protein (BMP) (e.g., BMP-1, BMP-2, BMP-3, BMP-4, BMP-5, BMP-6, BMP-7, BMP-8, BMP-9), brain-derived neurotrophic factor (BDNF), ciliary neurotrophic factor family, ciliary neurotrophic factor (CNTF ), connective tissue activating peptide (CTAP), epidermal growth factor (EGF), ephrins (e.g., ephrin A1, ephrin A2, ephrin A3, ephrin A4, ephrin A5, ephrin B1, ephrin B2, ephrin B3), erythropoietin (EPO), fibroblast growth factor (FGF) (e.g., FGF1, FGF2, FGF3, FGF4, FGF5, FGF6, FGF7, FGF8, FGF9, FGF10, FGF11, FGF12, FGF13, FGF14, FGF15, FGF16, FGF17, FGF18, FGF19, FGF20, FGF21, FGF22, FGF23, FGF24, FGF25, FGF26, FGF27, FGF28, FGF29, FGF30, FGF31, FGF32, FGF33, FGF34, FGF35, FGF36, FGF37, FGF38, FGF39, FGF40, FGF41, FGF42, FGF43, FGF44, FGF45, FGF46, FGF47, FGF48, FGF49, FGF50, FGF51, FGF52, FGF53, FGF54, FGF55, FGF56, FGF57, FGF58, FGF59, FGF60, FGF61, FGF62, FGF63, FGF64, FGF65, FGF66, FGF67, FGF68, FGF69, FGF70, FGF71, FGF72, FGF73, FGF74, FGF75, FGF76, FGF77, FGF78, FGF79, FGF8 F20, FGF21, FGF22, FGF23), basic fibroblast growth factor (bFGF), acidic fibroblast growth factor (aFGF), fetal bovine growth hormone (FBS), glial cell line-derived neurotrophic factor (GDNF), granulocyte colony-stimulating factor (G-CSF), granulocyte macrophage colony-stimulating factor (GM-CSF), growth differentiation factor (GDF) (e.g., GDF1, GDF9), heparin-binding growth factor, hepatocyte growth factor (HGF), hepatocyte growth factor-like protein (HGFLP), hepatoma-derived growth factor (HDGF), inhibins (e.g., inhibin A, inhibin B) , insulin, insulin-like growth factor (IGF) (e.g., IGF-1, IGF-2), interleukins (IL) (e.g., IL-1, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-11 and IL-13), keratinocyte growth factor (KGF), leukemia inhibitory factor (LIF), macrophage colony stimulating factor (M-CSF), macrophage stimulating protein (MSP), migration stimulating factor (MSF), myostatin, neuregulin (NRG) (e.g., NRG1, NRG2, NRG3, NRG4), neurotrophins (NT) (e.g.,NT-1, NT-2, NT-3, NT-4), neurotrophic factors (Neurturin), nerve growth factor (NGF), osteogenic factors, Persephin, placental growth factor (PGF), platelet-derived growth factor (PDGF), renal enzyme (RNLS), stromal cell-derived factor-1, T cell growth factor (TCGF), thrombopoietin (TPO), transforming growth factor alpha (TGF-α), transforming growth factor beta (TGF-β), tumor necrosis factor alpha (TNF-α) and vascular endothelial growth factor (VEGF), anti-vascular endothelial growth factor (anti-VEGF) (e.g., bevacizumab, ranibizumab, aflibercept) and biologically active analogs, fragments, derivatives or combinations thereof of such growth factors. ,

[0222] In certain embodiments, the therapeutic agent comprises one or more hormones. In certain embodiments, the therapeutic agent comprises one or more hormones selected from the following: anti-mullerian hormone, mullerian inhibitory factor or hormone, adiponectin, adrenocorticotropic hormone, corticotropin, angiotensinogen, angiotensin, antidiuretic hormone, vasopressin, arginine vasopressin, atrial natriuretic peptide, atrial peptide, calcitonin, cholecystokinin, cholecystokinin, corticotropin-releasing hormone, erythropoietin, follicle stimulating hormone, gastrin, ghrelin, glucagon, gonadotropin-releasing hormone, growth hormone-releasing hormone, human chorionic gonadotropin, human placental lactogen, growth hormone, somatomedin, leptin, luteinizing hormone, melanocyte stimulating hormone, orexin, oxytocin, parathyroid hormone, prolactin, relaxin, secretin, somatostatin, thrombopoietin, thyroid stimulating hormone, thyrotropin, thyrotropin-releasing hormone, or a combination thereof.

[0223] In certain embodiments, polymer compositions of the present disclosure may include one or more growth factors having a concentration (w / v) of about 0.001 μg / mL to about 2g / mL. In certain embodiments, polymer compositions of the present disclosure may include one or more growth factors having a concentration (w / v) of about 0.001 μg / mL to about 1000 μg / mL. In certain embodiments, polymer compositions may include one or more growth factors having a concentration (w / v) of about 0.01 μg / mL to about 500 μg / mL. In certain embodiments, polymer compositions may include one or more growth factors having a concentration (w / v) of about 0.1 μg / mL to about 200 μg / mL.In certain embodiments, the polymer composition may comprise a concentration (w / v) of about 0.1-0.5 μg / mL, about 0.5-1.0 μg / mL, about 1-2 μg / mL, about 2-4 μg / mL, about 4-6 μg / mL, about 6-8 μg / mL, about 8-10 μg / mL, about 10-12 μg / mL, about 12-14 μg / mL, about 14-16 μg / mL, about 16-18 μg / mL, about 18-20 μg / mL, about 20-22 μg / mL, about 22-24 μg / mL, about 24-26 μg / mL, about 26-28 μg / mL, about 28-30 μg / mL, about 30-35 μg / mL, about 35-40μg / mL, about 40-45μg / mL, about 45-50μg / mL, about 50-55μg / mL, about 55-60μg / mL, about 60-65μg / mL, about 65-70μg / mL, about 70-75μg / mL, about 75-80μg / mL, about 80-85μg / mL, about 85-90μg / mL, about 90-95μg / mL, about 95-100μg / mL, about 100-125μg / mL, about 125-150μg / mL, about 150-175μg / mL, about 175-200μg / mL, about 200-225μg / mL, about 225-250 μg / mL, about 250-275 μg / mL, about 275-300 μg / mL, about 300-325 μg / mL, about 325-350 μg / mL, about 350-375 μg / mL, about 375-400 μg / mL, about 400-425 μg / mL, about 425-450 μg / mL, about 450-475 μg / mL, about 475-500 μg / mL, about 500-550 μg / mL, about 550-600 μg / mL, about 600-650 μg / mL, about 650-700 μg / mL, about 700-750 μg / mL, about 750-800

[0047] In the present invention, the amount of one or more growth factors in the composition may be about 100 μg / mL, about 800-850 μg / mL, about 850-900 μg / mL, about 900-950 μg / mL, about 950-1000 μg / mL, about 1000-1100 μg / mL, about 1100-1200 μg / mL, about 1200-1300 μg / mL, about 1300-1400 μg / mL, about 1400-1500 μg / mL, about 1500-1600 μg / mL, about 1600-1700 μg / mL, about 1700-1800 μg / mL, about 1800-1900 μg / mL or about 1900-2000 μg / mL.

[0224] In certain embodiments, the therapeutic agent comprises one or more hemostatic agents (i.e., substances that promote hemostasis) and / or immunosuppressants. In certain embodiments, the therapeutic agent comprises one or more reagents selected from the following: platelets, platelet-like nanoparticles (e.g., silicate nanoparticles), coagulation factors (e.g., thrombin, prothrombin), alkylating agents, antimetabolites, mycophenolate mofetil, cyclosporin, tacrolimus, rapamycin, or a combination thereof. In certain embodiments, the therapeutic agent comprises an anticoagulant or a blood thinner (e.g., heparin).

[0225] In certain embodiments, polymer compositions of the present disclosure can be incorporated into or coated with cells or cell precursors of target tissue. In certain embodiments, polymer compositions can be incorporated into or coated with cells or cell precursors of one or more target tissues selected from the following: nerve cells, muscle cells, myocytes, cardiomyocytes, hepatocytes, keratinocytes, melanocytes, ameloblasts, fibroblasts, preosteoblasts, osteoblasts, osteoclasts, endothelial cells, epithelial cells, mesenchymal stem cells, nerve sheath cells (i.e., Schwann cells), embryonic stem cells, adult stem cells, pluripotent stem cells, multipotent stem cells, hematopoietic stem cells, adipose-derived stem cells, bone marrow-derived stem cells, osteocytes, nerve cells or their combinations. In certain embodiments, polymer compositions can be incorporated into or coated with endothelial cells (e.g., corneal endothelial cells). In certain embodiments, polymer compositions can be incorporated into or coated with eye cells. In certain embodiments, polymer compositions can be incorporated into or coated with adhesion cell types (i.e., cells forming intercellular networks, 3D vasculature). In certain embodiments, polymer compositions can be incorporated into or coated with monolayer cell types (i.e., 2D). In certain embodiments, the polymer composition can be incorporated or coated with epithelial cells, endothelial cells, corneal cells and combinations thereof. In certain embodiments, the polymer composition can be incorporated or coated with human umbilical vein endothelial cells (HUVEC) or vascular endothelial cells. In certain embodiments, the polymer composition can be incorporated or coated with human retinal pigment epithelial cells (HRPEC), human neuroepithelial cells, human photoreceptor cells, human corneal endothelial cells, human neural crest cells, human retinal ganglion cells, human limbal cells, human cardiomyocytes, human hepatocytes, human dermal cells, human gastrointestinal epithelial cells, human neurons and human islet cells, human immune cells and other therapeutic human cells.

[0226] In certain embodiments, cells or cell precursors can be incorporated into or onto a polymer gel matrix by placing the polymer gel composition in a cell culture mixture for a period of time. The culture time may vary depending on the cells used, but may generally be 1 to 21 days. In certain embodiments, the polymer gel composition is repeatedly exposed to the cell culture to increase the cell density in or on the gel matrix.

[0227] In certain embodiments, the polymer compositions of the present disclosure can be incorporated into cells or cell precursors according to the procedures disclosed in WO 2013040559; or Loessner et al., Nature protocols. 2016 Apr; 11(4):727.A1; to the extent that each document describes incorporating cells or cell precursors into a gel matrix (such as a GelMA hydrogel) or into a gel matrix, each document is incorporated herein by reference in its entirety.

[0228] Therapeutic applications

[0229] In certain embodiments, the polymer compositions of the present disclosure can be used as sealants and / or therapeutic compositions for treating and / or repairing soft tissue of a subject. In certain embodiments, the polymer compositions of the present disclosure can be used as delivery vehicles for administering therapeutic agents to treat and / or repair soft tissue of a subject. In certain embodiments, the polymer compositions of the present disclosure can be used as sealants and / or therapeutic compositions for treating and / or repairing soft tissue of a subject, and as delivery vehicles for administering therapeutic agents to treat and / or repair soft tissue of a subject.

[0230] In certain embodiments, the methods and compositions of the present disclosure can be used to adhere, seal or treat one or more target soft tissues selected from the following: ocular tissue (i.e., eye), lung, cardiovascular, skin, kidney, bladder, urethra, dura mater, liver, gastrointestinal or oral (i.e., mouth) tissue. In certain embodiments, the methods and compositions of the present disclosure can be used to adhere, seal or treat one or more target soft tissues in stress and / or physiological environments, or similar applications requiring elasticity and / or adhesion compositions.

[0231] In certain embodiments, the present disclosure provides a method for treating and / or repairing a subject's soft tissue using a polymer composition of the present disclosure. In certain embodiments, the present disclosure provides a method for treating and / or repairing a defect, injury and / or disease in a subject's soft tissue using a polymer composition of the present disclosure. In certain embodiments, the method comprises: applying a pregelled polymer composition of the present disclosure (e.g., a polymer composition comprising acryl-substituted gelatin) to an applicator; placing an applicator containing the pregelled polymer composition on the surface of a subject's target soft tissue (e.g., the location of a soft tissue defect, injury and / or disease); and crosslinking (e.g., photocrosslinking) the polymer composition by exposing the pregelled polymer composition to a crosslinking initiator (e.g., a photoinitiator and visible light). In certain embodiments, the method comprises removing the applicator from the gel polymer composition and / or the soft tissue surface after the polymerization crosslinking and / or gelling of the polymer composition is completed. In certain embodiments, the pregelled polymer composition is applied directly to the surface of the target soft tissue without an applicator. In certain embodiments, the pregelled polymer composition is applied on or near the target soft tissue (e.g., on the same tissue or under the tissue). In certain embodiments, the pregelled polymer composition has strong, sustained adhesion and high retention on the target soft tissue of the subject. In certain embodiments, the gel polymer composition can have strong, sustained adhesion and high retention on the target soft tissue of the subject. In certain embodiments, the polymer composition is engineered to exhibit physical, mechanical, structural, chemical and / or biological properties (elasticity, water content) to match or resemble the target soft tissue. In certain embodiments, the polymer composition is engineered to distribute the therapeutic agent to the target soft tissue.

[0232] Eye injuries and diseases

[0233] In certain embodiments, the polymer compositions of the present disclosure can be used as sealants and / or therapeutic compositions for treating and / or repairing ocular soft tissues in the eyes of a subject. In certain embodiments, the polymer compositions of the present disclosure can be used as sealants and / or therapeutic compositions for treating and / or repairing ocular defects, ocular surface damage or ocular diseases in the eyes of a subject. In certain embodiments, the ocular defect, damage or disease is a corneal or scleral defect, damage or disease. In certain embodiments, the corneal or scleral damage is a laceration (partial or full thickness), a perforation, an incision (e.g., a surgical incision) or a similar surface trauma (such as a trauma caused by a foreign object or projectile). In certain embodiments, the ocular defect, damage or disease is an ocular ulcer, such as a corneal ulcer caused by a severe infection, injury, perforation or other defect. In certain embodiments, the target soft tissue is an ocular tissue; optionally subconjunctival ocular tissue or retinal ocular tissue.

[0234] In certain embodiments, the present disclosure provides a method for treating an eye defect, an ocular surface injury or an eye disease of a subject with a polymer composition of the present disclosure. In certain embodiments, the method comprises: applying a pregelled polymer composition of the present disclosure (e.g., a polymer composition comprising acryl-substituted gelatin) to an applicator; placing an applicator containing the pregelled polymer composition on the surface of the subject's eye; and crosslinking (e.g., photocrosslinking) the polymer composition by exposing the pregelled polymer composition to a crosslinking initiator (e.g., visible light). In certain embodiments, the method comprises removing the applicator from the gelled polymer composition and / or the ocular surface after the polymerization crosslinking and / or gelling of the polymer composition is completed. In certain embodiments, the pregelled polymer composition is directly applied to the surface of the target ocular tissue without an applicator. In certain embodiments, the pregelled polymer composition has strong, sustained adhesion and high retention on the ocular tissue of the subject. In certain embodiments, the gelled polymer composition may have strong, sustained adhesion and high retention on the ocular tissue of the subject. In certain embodiments, the polymer composition is engineered to exhibit physical, mechanical, structural, chemical, and / or biological properties (elasticity, water content) to match or resemble a target ocular tissue (eg, corneal tissue).

[0235] In certain embodiments, the applicator is a curved concave surface. In certain embodiments, the applicator is a curved lens (e.g., a contact lens). In certain embodiments, the curvature of the applicator is similar to the curvature of the target eye surface.

[0236] In certain embodiments, an ocular defect, ocular surface injury or ocular disease in a target ocular tissue can be treated by: (i) forming a preformed polymer composition by polymerizing a polymer composition of the present disclosure; and (ii) applying the preformed polymer composition to the surface or subsurface (e.g., subconjunctival, subretinal) of a target tissue of a subject. In certain embodiments, application to the surface of the target tissue includes application / injection to the space just below the surface of the target tissue (e.g., subconjunctival application to ocular tissue, subretinal application to ocular tissue). In certain embodiments, the preformed polymer composition can be engineered to have specific physical, mechanical, structural, chemical and / or biological properties (e.g., elasticity, biodegradability, porosity).

[0237] In certain embodiments, an ocular defect, ocular surface injury, or ocular disease in a target ocular tissue can be treated by: (i) forming a preformed hydrogel polymer composition by polymerizing a polymer composition of the present disclosure; (ii) drying the hydrogel polymer by removing a majority of the interstitial fluid from the hydrogel (e.g., at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% of the interstitial fluid); (iii) applying the preformed polymer composition to the surface or subsurface of a target tissue of a subject (e.g., subconjunctival, subretinal); and (iv) optionally rehydrating the dried hydrogel polymer to a substantially hydrated form (e.g., at least 50%, at least 60%, at least 70%, at least 80%, at least 90%, or at least 95% of the volume of the interstitial fluid). In certain embodiments, applying to the surface of the target tissue includes applying / injecting into the space just below the surface of the target tissue (e.g., subconjunctival application to the ocular tissue, subretinal application to the ocular tissue). In certain embodiments, preformed polymer compositions can be engineered to have specific physical, mechanical, structural, chemical, and / or biological properties (eg, elasticity, biodegradability, porosity).

[0238] Oral injuries and diseases

[0239] In certain embodiments, polymer compositions of the present disclosure can be used as sealants and / or therapeutic compositions for treating and / or repairing soft tissues in the subject's oral cavity. In certain embodiments, polymer compositions can be used for treating and / or repairing oral tissues with periodontal disease, damage or ailments. In certain embodiments, periodontal disease, damage or ailments may include those associated with periodontal implants, including peri-implant disease (PID), such as peri-implant mucositis (PIM) and peri-implantitis (PI). These ailments are often associated with inflammation of the soft tissues around periodontal implants (caused by bacterial accumulation and biofilm formation), causing bleeding, suppuration, erythema, swelling and infection of oral tissues, and progressive bone loss that may cause implant failure.

[0240] In certain embodiments, polymer compositions of the present disclosure can be used for closing the soft tissue area around periodontal implants. In certain embodiments, polymer compositions of the present disclosure can be used for delivering therapeutic agents (e.g., antimicrobial agents or anti-inflammatory agents) to the soft tissue area around periodontal implants. In certain embodiments, the polymer compositions include osteoinductive agents. In certain embodiments, the polymer compositions include one or more osteoinductive agents selected from silicate nanoparticles (SN), calcium salts, bioglass, hydroxyapatite, decalcified bone matrix (DBM) or their combination. In certain embodiments, the polymer compositions include one or more silicate nanoparticles, including SN containing one or more metals such as calcium, aluminum, silver, gold, platinum, palladium, lithium, magnesium, sodium, titanium, vanadium, chromium, manganese, iron, cobalt, nickel, copper, zinc, iridium or their combination. In certain embodiments, silicate nanoparticles include laponite nanoparticles. In certain embodiments, the polymer compositions include one or more calcium salts, such as calcium phosphate, calcium sulfate, calcium hydroxide, calcium bromide, calcium fluoride, calcium iodide, calcium hydride or their combination.

[0241] In certain embodiments, the present disclosure provides a method for treating a defect, injury or disease in a subject's oral soft tissue with a polymer composition of the present disclosure. In certain embodiments, the method includes: applying a pregelled polymer composition of the present disclosure (e.g., a polymer composition comprising acryl-substituted gelatin) to an applicator; placing an applicator containing the pregelled polymer composition on the surface of a subject's oral soft tissue (e.g., soft tissue around a periodontal implant); and crosslinking (e.g., photocrosslinking) the polymer composition by exposing the pregelled polymer composition to a crosslinking initiator (e.g., visible light). In certain embodiments, the method includes removing the applicator from the gelled polymer composition and / or the oral soft tissue surface after the polymerization crosslinking and / or gelling of the polymer composition is completed. In certain embodiments, the pregelled polymer composition is directly applied to the surface of the target oral soft tissue without an applicator. In certain embodiments, the pregelled polymer composition has strong, sustained adhesion and high retention on the oral soft tissue of the subject. In certain embodiments, the gelled polymer composition may have strong, sustained adhesion and high retention on the oral soft tissue of the subject. In certain embodiments, the polymer composition is engineered to exhibit physical, mechanical, structural, chemical, and / or biological properties (elasticity, water content) to match or resemble the target oral soft tissue (eg, soft tissue surrounding a periodontal implant).

[0242] Nerve damage and disease

[0243] In certain embodiments, the polymer compositions of the present disclosure can be used as sealants and / or therapeutic compositions for treating and / or repairing soft tissue in a subject's nervous system (e.g., central nervous system (CNS), peripheral nervous system (PNS)). In certain embodiments, the polymer compositions can be used to treat and / or repair neural tissue associated with traumatic injury or surgical damage, including peripheral nerve injury (PNI). Typical surgical interventions for these ailments, including suturing and / or commercial adhesives, are often associated with inflammation, increased foreign body response (FBR), scarring, slow nerve regeneration, or loss of nerve function (partial or complete).

[0244] In certain embodiments, neural tissue may be treated or sealed by applying the polymer composition of the present disclosure to the target neural tissue. In certain embodiments, neural tissue may be treated or sealed by applying the polymer composition of the present disclosure to the lumen of a nerve conduit at a site of neural injury.

[0245] In certain embodiments, the present disclosure provides a method for treating a defect, injury or disease in a nerve or CNS tissue of a subject with a polymer composition of the present disclosure. In certain embodiments, the method includes: applying a pregelled polymer composition of the present disclosure (e.g., a polymer composition comprising acryl-substituted gelatin) to an applicator; placing an applicator containing a pregelled polymer composition on the surface of a subject's nerve or CNS soft tissue (e.g., a nerve of the peripheral nervous system); and crosslinking (e.g., photocrosslinking) the polymer composition by exposing the pregelled polymer composition to a crosslinking initiator (e.g., visible light). In certain embodiments, the method includes removing the applicator from the gel polymer composition and / or nerve / CNS tissue surface after the polymerization crosslinking and / or gelling of the polymer composition is completed. In certain embodiments, the pregelled polymer composition is directly applied to the surface of the target nerve or CNS tissue without an applicator. In certain embodiments, the pregelled polymer composition has strong, persistent adhesion and high retention on the target nerve or CNS tissue of the subject. In certain embodiments, the gel polymer composition can have strong, persistent adhesion and high retention on the target nerve or CNS tissue of the subject. In certain embodiments, the polymer composition is engineered to exhibit physical, mechanical, structural, chemical and / or biological properties (elasticity, water content) to match or resemble the target nerve or CNS tissue (e.g., nerves of the peripheral nervous system).

[0246] In certain embodiments, the polymer compositions of the present disclosure may include a polymeric component or a therapeutic component, or may be produced, analyzed, or used by methods as disclosed in US20190070338 (including methods for treating nerve damage), which is incorporated herein by reference in its entirety to the extent that the patent describes the composition, production, analysis, and use of acrylated gelatin polymeric compositions such as GelMA hydrogels.

[0247] Cardiovascular injury and disease

[0248] In certain embodiments, the polymer compositions of the present disclosure can be used as sealants and / or therapeutic compositions for treating and / or repairing soft tissue in a subject's cardiovascular system (e.g., heart). In certain embodiments, the polymer compositions can be used to treat and / or repair cardiovascular tissue (including cardiac tissue) with traumatic injury or surgical damage. Typical surgical interventions for these ailments (including suturing and / or commercial adhesives) are often associated with inflammation and infection, scarring, slow tissue regeneration, or loss of function (partial or complete).

[0249] In certain embodiments, the polymer composition of the present invention can be applied to the target blood vessel / cardiovascular tissue to treat or close blood vessel / cardiovascular tissue. In certain embodiments, the cell-loaded hydrogel composition of the present invention can be applied to the target blood vessel / cardiovascular tissue to treat or close blood vessel / cardiovascular tissue. In certain embodiments, the cell-loaded hydrogel composition includes cells or cell precursors that stimulate or promote the repair, recovery, replacement or regeneration of blood vessel / cardiovascular tissue (e.g., cardiac tissue). In certain embodiments, the cell-loaded hydrogel composition includes one or more cells or cell precursors selected from the following: smooth muscle cells, cardiomyocytes, fibroblasts, mesenchymal stem cells, bone marrow stem cells or their combinations. In certain embodiments, the cell-loaded hydrogel composition is a pad, a fabric, a net or other shapes suitable for use as a covering or a graft.

[0250] In certain embodiments, the present disclosure provides a method for treating defects, injuries or diseases in cardiovascular tissue of a subject with a polymer composition of the present disclosure. In certain embodiments, the method comprises: applying a pregelled polymer composition of the present disclosure (e.g., a polymer composition comprising acryl-substituted gelatin) to an applicator; placing an applicator containing the pregelled polymer composition on the surface of the subject's cardiovascular tissue (e.g., heart tissue); and crosslinking (e.g., photocrosslinking) the polymer composition by exposing the pregelled polymer composition to a crosslinking initiator (e.g., visible light). In certain embodiments, the method comprises removing the applicator from the gelled polymer composition and / or the surface of the cardiovascular tissue after the polymerization crosslinking and / or gelling of the polymer composition is completed. In certain embodiments, the pregelled polymer composition is directly applied to the surface of the target cardiovascular tissue without an applicator. In certain embodiments, the pregelled polymer composition has strong, sustained adhesion and high retention on the subject's cardiovascular tissue. In certain embodiments, the gelled polymer composition may have strong, sustained adhesion and high retention on the subject's cardiovascular tissue. In certain embodiments, the polymer composition is engineered to exhibit physical, mechanical, structural, chemical, and / or biological properties (elasticity, water content) to match or resemble a target cardiovascular tissue (eg, cardiac tissue).

[0251] In certain embodiments, the polymer compositions of the present disclosure may include a polymeric component or a therapeutic component, or may be produced, analyzed, or used by methods as disclosed in WO2014063194 (including methods for treating cardiovascular damage), which is incorporated herein by reference in its entirety to the extent that the patent describes the composition, production, analysis, and use of acrylated gelatin polymeric compositions such as GelMA hydrogels.

[0252] Lung injury and disease

[0253] In certain embodiments, the polymer compositions of the present disclosure can be used as sealants and / or therapeutic compositions for treating and / or repairing soft tissue in a subject's lung. In certain embodiments, the polymer compositions can be used for treating and / or repairing lung tissue associated with traumatic injury or surgical damage. Typical surgical interventions for these ailments (including suturing and / or commercial adhesives) are often associated with inflammation and infection, scarring, slow tissue regeneration, or loss of function (partial or complete).

[0254] In certain embodiments, lung tissue can be treated or sealed by applying the polymer composition of the present disclosure to the target vascular / cardiovascular tissue. In certain embodiments, lung tissue can be treated or sealed by applying the cell-laden hydrogel composition of the present disclosure to the target lung tissue. In certain embodiments, the cell-laden hydrogel composition comprises cells or cell precursors that stimulate or promote the repair, restoration, replacement or regeneration of lung tissue. In certain embodiments, the cell-laden hydrogel composition is in the form of a pad, fabric, mesh or other shape suitable for use as a covering or graft.

[0255] In certain embodiments, the polymer composition comprises acryl-substituted gelatin (e.g., GelMA) and acryl-substituted PEG (e.g., PEGDA) in a ratio of about 30: 1 to about 1: 30 w / w. In certain embodiments, the polymer composition comprises acryl-substituted gelatin (e.g., GelMA) and acryl-substituted hyaluronic acid (e.g., MeHA) in a ratio of about 30: 1 to about 1: 30 w / w. In certain embodiments, the polymer composition comprises acryl-substituted gelatin (e.g., GelMA), acryl-substituted PEG (e.g., PEGDA) and acryl-substituted hyaluronic acid (e.g., MeHA).

[0256] In certain embodiments, the present disclosure provides a method for treating a defect, injury or disease in a lung tissue of a subject with a polymer composition of the present disclosure. In certain embodiments, the method comprises: applying a pregelled polymer composition of the present disclosure (e.g., a polymer composition comprising acryl-substituted gelatin) to an applicator; placing an applicator containing the pregelled polymer composition on the surface of the lung tissue of the subject; and crosslinking (e.g., photocrosslinking) the polymer composition by exposing the pregelled polymer composition to a crosslinking initiator (e.g., visible light). In certain embodiments, the method comprises removing the applicator from the gel polymer composition and / or the lung tissue surface after the polymerization crosslinking and / or gelling of the polymer composition is completed. In certain embodiments, the pregelled polymer composition is directly applied to the surface of the target lung tissue without an applicator. In certain embodiments, the pregelled polymer composition has strong, sustained adhesion and high retention on the lung tissue of the subject. In certain embodiments, the gel polymer composition may have strong, sustained adhesion and high retention on the lung tissue of the subject. In certain embodiments, the polymer composition is engineered to exhibit physical, mechanical, structural, chemical, and / or biological properties (elasticity, water content) to match or resemble the target lung tissue.

[0257] V. Definition

[0258] At various places in the present disclosure, substituents or properties of compounds of the present disclosure are disclosed in the form of groups or ranges. It is specifically intended that the present disclosure includes every individual or subcombination of the members of such groups and ranges.

[0259] Unless otherwise indicated, the following terms and phrases have the following meanings. These definitions are not intended to be limiting in nature, but are used to provide a clearer understanding of certain aspects of the disclosure.

[0260] GelMA polymer composition: As used herein, the term "GelMA polymer composition" refers to.

[0261] Administering: As used herein, the term "administering" refers to providing a composition to a subject.

[0262] Improvement: As used herein, the term "improvement" refers to a decrease in the severity of at least one indicator of a condition or disease.

[0263] Animal: As used herein, the term "animal" refers to any member of the animal kingdom. In certain embodiments, "animal" refers to a person at any stage of development. In certain embodiments, "animal" refers to a non-human animal at any stage of development. In certain embodiments, the non-human animal is a mammal (e.g., a rodent, a mouse, a rat, a rabbit, a monkey, a dog, a cat, a sheep, a cow, a primate, or a pig). In certain embodiments, animals include, but are not limited to, mammals, birds, reptiles, amphibians, fish, and worms. In certain embodiments, animals are transgenic animals, genetically engineered animals, or clones.

[0264] Approximately: As used herein, the term "approximately" or "about", as applicable to one or more target values, refers to a value similar to a specified reference value. The term may refer to + / -10% of the value recited. In certain embodiments, unless otherwise specified or otherwise apparent from the context (unless such numerals exceed 100% of possible values), the term refers to a series of values ​​falling within 25%, 20%, 19%, 18%, 17%, 16%, 15%, 14%, 13%, 12%, 11%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1% or less of a range of values ​​in either direction (greater than or less than) of a specified reference value.

[0265] Associating with: As used herein, the terms "associated with," "conjugated," "linked," "attached," and "tethered," when used with respect to two or more moieties, mean that the moieties are physically associated or linked to one another, either directly or via one or more additional moieties that act as linkers, to form a sufficiently stable structure such that the moieties remain physically associated under the conditions (e.g., physiological conditions) under which the structure is used. "Associated" need not strictly be by direct covalent chemical bonding. It may also imply ionic or hydrogen bonding or hybridization-based linkages that are sufficiently stable such that the "associated" entities remain physically associated.

[0266] Biocompatible: As used herein, the term "biocompatible" refers to materials that produce minimal or zero toxic, damaging, or immune response in living tissue.

[0267] Biodegradable: As used herein, the term "biodegradable" refers to a material that can be partially or completely decomposed into bioprocessable byproducts under physiological conditions. For example, if at least 10%, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, or at least 90% of the material can be decomposed under physiological conditions within a desired time period (e.g., minutes, hours, days, weeks, or months, depending on the nature of the material and the physiological application), the material can be considered biodegradable. The term "biodegradable" may encompass the term "bioresorbable," which describes a substance that decomposes under physiological conditions into products that undergo bioresorption into a host subject (e.g., as a metabolite in a biochemical system).

[0268] Bioactive: As used herein, the term "bioactive" refers to the characteristic of any substance or material that is active in a biological system and / or organism. For example, a material that has a biological effect on an organism when administered to the organism is considered bioactive.

[0269] Compounds: Compounds of the present disclosure include all isotopes of atoms present in the intermediates or final compounds. "Isotopes" refer to atoms having the same atomic number but having different mass numbers due to different numbers of neutrons in the nucleus. For example, isotopes of hydrogen include tritium and deuterium. Compounds and salts of the present disclosure can be prepared by conventional methods in combination with solvents or water molecules to form solvates and hydrates.

[0270] Cross-link: As used herein, the term "cross-link" or "cross-linking" refers to the formation of a bond (eg, covalent bond formation) that connects one polymer unit to another polymer unit.

[0271] Encapsulate: As used herein, the term "encapsulate" means to enclose, surround, or enclose.

[0272] Engineered: As used herein, embodiments of the present disclosure are "engineered" when they are designed to have structural or chemical features or properties that differ from the starting or naturally occurring molecule.

[0273] Effective amount: As used herein, the term "effective amount" of an agent is an amount sufficient to achieve a beneficial or desired result (e.g., a clinical result), and thus, the "effective amount" depends on the context in which it is used. For example, in the context of applying an agent to treat an ocular injury or condition, the effective amount of the agent is, for example, an amount sufficient to achieve treatment of the ocular injury or condition compared to the response obtained when the agent is not administered.

[0274] Characteristics: As used herein, "characteristics" refers to features, characteristics, or distinguishing elements.

[0275] In vitro: As used herein, the term "in vitro" refers to events that occur in an artificial environment, such as in a test tube or reaction vessel, in cell culture, in a Petri dish, etc., rather than within an organism (e.g., an animal, plant, or microorganism).

[0276] In vivo: As used herein, the term "in vivo" refers to events that occur within an organism (eg, an animal, plant, or microorganism, or a cell or tissue thereof).

[0277] Modified: As used herein, "modified" refers to an altered state or structure of a molecule of the present disclosure. Molecules can be modified in a variety of ways, including chemically, structurally, and functionally. As used herein, embodiments of the present disclosure are modified when they have or possess structural or chemical features or properties that are different from the starting or native molecule.

[0278] Non-human animals: As used herein, "non-human animals" include all animals (e.g., vertebrates) other than Homo sapiens, including wild and domesticated species. Examples of non-human vertebrates include, but are not limited to, mammals such as alpacas, wild cattle, bison, camels, cats, cattle, deer, dogs, donkeys, cattle, goats, guinea pigs, horses, llamas, mules, pigs, rabbits, reindeer, sheep, buffalo, and yaks. Non-human animals include non-human primates.

[0279] Pharmaceutically acceptable: The term "pharmaceutically acceptable" or "therapeutically acceptable" is used herein to refer to those compounds, materials, compositions and / or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and animals without excessive toxicity, irritation, allergic response, or other problems or complications, commensurate with a reasonable benefit / risk ratio.

[0280] Pharmaceutically acceptable excipient: As used herein, the term "pharmaceutically acceptable excipient" or "therapeutically acceptable excipient" refers to ingredients other than the polymer composition described herein and having the property of being substantially non-toxic and non-inflammatory in a subject (e.g., a vehicle capable of suspending or dissolving a polymer compound).

[0281] Pharmaceutically acceptable salts: The present disclosure also includes pharmaceutically acceptable salts of the compounds described herein. As used herein, "pharmaceutically acceptable salts" refers to derivatives of the disclosed compounds in which the parent compound is modified by converting an existing acid or base moiety into its salt form (e.g., by reacting a free base group with a suitable organic acid). Examples of pharmaceutically acceptable salts include, but are not limited to, inorganic or organic acid salts of basic residues such as amines; basic or organic salts of acidic residues such as carboxylic acids; and the like. Representative acid addition salts include acetate, acetic acid, adipate, alginate, ascorbate, aspartate, benzenesulfonate, benzenesulfonic acid, benzoate, bisulfate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, fumarate, glucoheptonate, glycerophosphate, hemisulfate, heptanoate, hexanoate, hydrobromide, hydrochloride, hydroiodide, 2-hydroxy-ethanesulfonate, 4-(2- The pharmaceutically acceptable salts of the present invention include, but are not limited to, ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, ethylamine, and the like. The pharmaceutically acceptable salts of the present invention include, but are not limited to, ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, ethylamine, and the like. The pharmaceutically acceptable salts of the present invention include, for example, conventional non-toxic salts of the parent compound formed from non-toxic inorganic or organic acids. Pharmaceutically acceptable salts of the present disclosure can be synthesized from parent compounds containing a basic or acidic moiety by conventional chemical methods. Typically, such salts can be prepared by reacting the free acid or base form of these compounds with a stoichiometric amount of an appropriate base or acid in water or in an organic solvent or in a mixture of the two; typically, a non-aqueous medium such as ether, ethyl acetate, ethanol, isopropanol or acetonitrile is used.

[0282] Subject: As used herein, the term "subject" refers to any organism to which a composition according to the present disclosure can be administered, e.g., for experimental, diagnostic, preventive and / or therapeutic purposes. Typical subjects include animals (e.g., mammals such as mice, rats, rabbits, non-human primates, and humans) and / or plants. A subject or patient may be seeking or in need of treatment for a particular disease or condition, requiring treatment, being treated, about to be treated, or being cared for by a trained professional.

[0283] Substantially: As used herein, the term "substantially" refers to the qualitative condition of exhibiting a characteristic or property of interest in full or near full extent or degree. It will be understood by those of ordinary skill in these fields that biological and chemical phenomena rarely, if ever, complete and / or proceed to completion or achieve or avoid an absolute result. Thus, the term "substantially" is used herein to explicitly capture the potential lack of completeness inherent in many biological and chemical phenomena. Likewise, excluding the term "substantially" does not exclude the possible lack of completeness inherent in many biological and chemical phenomena.

[0284] Synthetic: The term "synthetic" means artificially produced, prepared and / or manufactured. The synthesis of polynucleotides or polypeptides or other molecules of the present disclosure may be chemical or enzymatic.

[0285] Therapeutic agent: The term "therapeutic agent" refers to any agent that has a therapeutic, diagnostic and / or prophylactic effect and / or causes a desired biological and / or pharmacological effect when administered to a subject. Examples of therapeutic agents include, but are not limited to, oligonucleotides (e.g., sense and / or antisense DNA and / or RNA), proteins and polypeptides (e.g., hormones, growth factors), small molecules and drugs, and cells (e.g., stem cells, epithelial cells).

[0286] Treat: As used herein, the term "treat" may refer to partially or completely alleviating, ameliorating, modifying, relieving, preventing, delaying the onset of, inhibiting the progression of, reducing the severity of, and / or reducing the incidence of one or more symptoms or characteristics of a particular infection, disease, disorder, and / or condition. Treatment may be administered to a subject who does not exhibit signs of a disease, disorder, and / or condition, and / or to a subject who exhibits only early signs of a disease, disorder, and / or condition, in order to reduce the risk of developing pathology associated with the disease, disorder, and / or condition.

[0287] Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments of the disclosure described herein.The scope of the present disclosure is not intended to be limited to the above description.

[0288] In the claims, unless indicated to the contrary or obvious from the context, articles such as "a", "an", and "the" may mean one or more than one. Unless indicated to the contrary or otherwise obvious from the context, a claim or description containing "or" between one or more members of the group is considered to be satisfied if one, more than one, or all of the group members are present in, used in, or otherwise related to a given product or method. The disclosure may include embodiments in which exactly one member of the group is present in, used in, or otherwise related to a given product or method. The disclosure may include embodiments in which more than one member of the group or the entire group of members is present in, used in, or otherwise related to a given product or method.

[0289] It should also be noted that the term "comprising" is intended to be open ended, and permits but does not require the inclusion of additional elements or steps. When the term "comprising" is used herein, the term "consisting of" is also encompassed and disclosed thereby.

[0290] The abbreviation "eg" is derived from the Latin "exempli gratia" and is used herein to indicate a non-limiting example. Thus, the abbreviation "eg" is synonymous with the term "for example".

[0291] The abbreviation "ie" is derived from the Latin word id est and is used herein to indicate a non-limiting restatement or clarification. Thus, the abbreviation "ie" is synonymous with the term "that is to say".

[0292] When a range is given, the endpoints are included. In addition, it should be understood that unless otherwise indicated or otherwise obvious from the context of the present disclosure and the understanding of one of ordinary skill in the art, unless the context clearly indicates otherwise, the values ​​expressed as ranges can assume any specific value or sub-range within the range specified in the various embodiments of the present disclosure, to one-tenth of the unit of the lower limit of the range.

[0293] In addition, it should be understood that any specific embodiment of the present disclosure that falls within the prior art can be explicitly excluded from any one or more claims. Because such embodiments are considered to be known to those of ordinary skill in the art, they can be excluded even if the exclusion is not explicitly set forth herein. Any specific embodiment of the composition of the present disclosure (e.g., any antibiotic, therapeutic or active ingredient; any production method; any method of use; etc.) can be excluded from any one or more claims for any reason, whether or not related to the existence of prior art.

[0294] It is to be understood that the words which have been used are words of description rather than limitation and that variations may be made within the purview of the appended claims without departing from the true scope and spirit of the disclosure in its broader aspects.

[0295] While the present disclosure has been described in considerable detail and with a certain particularity with respect to several described embodiments, it is not intended that the present disclosure should be limited to any such details or embodiments or to any particular embodiment, but rather be interpreted with reference to the appended claims so as to provide the broadest possible interpretation of such claims in light of the art to effectively cover the intended scope of the present disclosure.

[0296] All publications, patent applications, patents, and other references mentioned herein are incorporated by reference in their entirety. In the event of a conflict, the present specification, including definitions, will prevail. In addition, section headings, materials, methods, and examples are illustrative only and are not intended to be limiting.

[0297] Example

[0298] Example 1. Preparation of Precursor Polymer Composition

[0299] (a) Preparation of Methacrylated Gelatin (GelMA) Precursor Polymer Composition

[0300] The GelMA precursor polymer composition can be synthesized as described in the art. For example, GelMA is synthesized by dissolving 10% (w / v) gelatin (e.g., porcine gelatin) in phosphate buffered saline (PBS) and then heating at 60°C for 20 minutes. After heating, 8% (v / v) methacrylic anhydride is added dropwise at 50°C for 3 hours (under continuous stirring), followed by dilution with PBS and dialysis at 40-50°C for about 7 days (using deionized water). The resulting mixture is filtered and lyophilized for 4 days. The resulting GelMA precursor polymer composition can be stored at -80°C until further use.

[0301] In an alternative, GelMA was synthesized by dissolving 10 g of fish skin gelatin in 100 ml of DPBS at 60°C for 30 minutes, then adding 8% (v / v) methacrylic anhydride dropwise to the solution at 60°C with stirring for 3 hours. Another 300 ml of DPBS was added to stop the reaction. The resulting mixture was dialyzed using a 50°C deionized water bath for about 5 days to remove unreacted methacrylic anhydride. The resulting solution was filtered and lyophilized for about 4 days.

[0302] (b) Preparation of Methacrylated Hyaluronic Acid (MeHA) Precursor Polymer Composition

[0303] MeHA precursor polymer compositions can be synthesized as described in the art, such as those presented in the following literature: Bencherif et al., Biomaterials 29, 1739-1749 (2008); Prata et al., Biomacromolecules 11, 769-775 (2010). For example, MeHA is synthesized by dissolving about 2 grams of hyaluronic acid sodium salt in 200 ml of deionized water, followed by the sequential addition of 8.0 mL of triethylamine, 8.0 mL of glycidyl methacrylate, and 4.0 grams of tetrabutylammonium bromide (stirring for 1 hour between each sequential addition). The resulting mixture is incubated at 55 ° C for 1 hour, then cooled (ice bath) and precipitated in acetone (4 L) to form a white solid precipitate. The precipitate is rinsed with fresh acetone, dissolved in purified water, dialyzed for 2 days, and then frozen and lyophilized for storage.

[0304] (c) Preparation of polyethylene glycol diacrylate (PEGDA) precursor polymer composition

[0305] The PEGDA precursor polymer composition can be synthesized as described in the art. For example, PEGDA is synthesized by reacting 10 grams of PEG in dichloromethane (10% w / v) with triethylamine and acryloyl chloride (1:4:4 molar ratio) at 4° C. under inert conditions (stirring overnight). The resulting mixture is filtered and then precipitated with ice-cold ether. The resulting precipitated product is filtered and dried in a vacuum desiccator overnight to remove residual material.

[0306] In an alternative, PEGDA is synthesized by dissolving PEG diol in benzene, followed by azeotropic distillation in toluene using a Dean-Stark water trap to remove water and ensure dry acrylation conditions. PEG acrylation is performed by dissolving PEG in a dichloromethane solution (under argon), followed by adding acryloyl chloride and triethylamine in a molar ratio of PEG: acryloyl chloride: OH-groups of triethylamine of 2:3:3. The resulting mixture is stirred overnight at room temperature (darkroom conditions). The resulting product is then precipitated with ether and cooled to 4°C, then filtered for recovery and vacuum oven drying.

[0307] Example 2: Preparation of hydrogel polymer composition

[0308] The hydrogel polymer composition can be synthesized as described in the art. For example, the freeze-dried GelMA precursor polymer composition produced according to Example 1(a) is dissolved in PBS or (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid) buffered saline at a concentration of 5-25% (w / v). 2-Hydroxy-4'-(2-hydroxyethoxy)-2-methylpropiophenone or Eosin Y or Eosin Y disodium salt is added as a photoinitiator, and the mixture is dissolved at 20-80°C. The resulting precursor polymer composition is photocrosslinked by visible light irradiation (e.g., blue light or white light) to form a GelMA hydrogel polymer composition. In an alternative, a target concentration of a MeHA precursor polymer composition [Example 1(b)] and / or a PEGDA precursor polymer composition [Example 1(c)] can be added to the precursor polymer solution, wherein the amount of each component is added based on the desired physical, mechanical, structural, chemical and / or biological properties of the hydrogel polymer composition.

[0309] In an alternative, the GelMA hydrogel polymer composition is synthesized by first dissolving 7-15% w / v of the methacrylated gelatin of Example 1 in a solution containing at least one photoinitiator component, such as a mixture of triethanolamine (about 2% w / v) and N-vinyl caprolactam (about 1.25% w / v) in distilled water at room temperature. Eosin Y disodium salt solution (0.5 mM) is then added to the methacrylated gelatin solution, and the resulting precursor polymer composition is then photocrosslinked under exposure to visible light (420-480 nm) for 120 seconds. In an alternative, a target concentration of a MeHA precursor polymer composition [Example 1(b)] and / or a PEGDA precursor polymer composition [Example 1(c)] can be added to the precursor polymer solution, wherein the amount of each component is added based on the desired physical, mechanical, structural, chemical and / or biological properties of the hydrogel polymer composition.

[0310] In one alternative, microparticles (eg, micelles) containing a therapeutic agent (eg, an ophthalmic antibiotic such as ciprofloxacin) are incorporated into the GelMA precursor polymer composition prior to photocrosslinking.

[0311] Porosity can be measured and analyzed by making freeze-dried, gold sputter-coated hydrogel samples, which can then be imaged using a scanning electron microscope (SEM).

[0312] Samples can also be subjected to a range of mechanical tests including elasticity, swelling, compression testing, texture and tensile testing.

[0313] In an alternative, a GelMA hydrogel polymeric composition is formed on the surface of a target tissue. The resulting samples can be subjected to a range of mechanical and therapeutic tests, including adhesion, burst pressure, wound closure strength, shear strength, and durability / degradation rate.

[0314] Example 3: Preparation of hydrogel polymer composition

[0315] The hydrogel polymer composition was prepared according to the following steps.

[0316] A photopolymerization initiator mixture was prepared containing: 0.35 mg / mL Eosin Y (20% v / v), 12.5 mg / mL N-vinylcaprolactam, and 18.75 mg / mL triethanolamine (80% v / v) in phosphate buffered saline (PBS; pH 7), adjusting the pH as needed using concentrated HCl.

[0317] Polymer precursors were obtained from the following sources: (1) GelMA-Rousselot Biomedical (160P80 or GelMA160P40 or GelMA 160P10); (2) HAMA-HTL Biotechnology (BLo-RD029-008); (3) HAGM-synthesized in-house according to methods known in the art (see, e.g., Example 1(b)); and (4) PEGDA-Jen Kem (ACLT-PEG35K-ACLT). The polymer precursors were allowed to reach room temperature (RT) before being incorporated into the hydrogel polymer precursor composition.

[0318] The PEGDA precursor material (when appropriate for the target formulation) is first added to the photopolymerization initiator mixture at the desired concentration (eg, 0.1-20% w / v) and allowed to dissolve at 37°C for about 5 minutes.

[0319] GelMA precursor material (when appropriate for the intended formulation) is then added to the hydrogel precursor mixture at a desired concentration (eg, 4-20% w / v) and allowed to dissolve at 60°C for approximately 2 hours with occasional vortexing.

[0320] MeHA (ie, HAMA or HAGM) precursor material (when appropriate for the intended formulation) is then added to the hydrogel precursor mixture at the desired concentration (eg, 1-3% w / v) and allowed to dissolve with stirring at 60°C overnight (to prevent any phase separation).

[0321] Once all precursor materials are fully dissolved into the hydrogel precursor mixture, the active agent (when applicable to the target formulation) is added at the desired concentration (eg, 1-350 mg / mL). The mixture is maintained at 37°C with stirring until ready for polymerization.

[0322] The hydrogel disc samples were prepared by pipetting about 100 μL of the hydrogel precursor mixture into a single poly(dimethylsiloxane) (PDMS) cylindrical mold located in a well of an untreated 24-well plate. The polymer composition was then photocrosslinked using a Dolan-Jenner high-intensity LED illuminator (MI-LED-US-B1) equipped with a two-arm gooseneck configuration (one arm above and one arm below, thus allowing dual exposure from top and bottom; incident light 90°). The average light power output per arm was ~100 mW / cm2 (λmax=450, 540 nm), and the exposure time varied from 5 seconds to 4 minutes.

[0323] The hydrogel rod sample was prepared by immersing a borosilicate glass capillary with an inner diameter of 0.75 mm into the hydrogel precursor mixture and then oscillating the capillary until it was filled to about 10 mm from the opening. The polymer composition was then photocrosslinked using a Dolan-Jenner high-intensity LED illuminator (MI-LED-US-B1) equipped with a double-arm gooseneck structure (one arm above and one arm below, thus allowing double exposure from the top and bottom; incident light 90°). The average light power output by each arm was ~100 mW / cm2 (λmax=450, 540 nm), and the exposure time was about 4 minutes. A quartz rod with a diameter of 0.5 mm was used to extrude the hydrogel rod from the capillary and then cut to size using calipers.

[0324] Example 4: Study on hydrogel properties

[0325] a) Crosslinking degree-photopolymerization time

[0326] Completed the study analyzing the correlation between the degree of cross-linking within the hydrogel and the photopolymerization time.

[0327] Hydrogels containing only HAMA were prepared according to the general procedure of Example 3, with photocrosslinking times of 15 seconds, 1 minute, 2 minutes, and 4 minutes. The resulting hydrogels were dried under vacuum, dissolved in deuterated DMSO, and then analyzed using proton NMR analysis (d-DMSO solvent). Other techniques, such as Fourier transform infrared spectroscopy (FTIR) and Raman spectroscopy, can also be used. For HAMA hydrogels, the change in the proton ratio between methacrylate methyl groups and HA carbonyl methyl groups was quantified according to the exposure time and normalized to the ratio present in uncrosslinked HAMA to express the degree of crosslinking (%). Figure 4A The results in show that the degree of cross-linking increases with increasing exposure time.

[0328] Hydrogels containing only GelMA were prepared according to the general procedure of Example 3, with photocrosslinking times of 30 seconds, 1 minute, 2 minutes, and 4 minutes. The resulting hydrogels were dried under vacuum, dissolved in deuterated DMSO, and then analyzed using proton NMR analysis (d-DMSO solvent). Other techniques such as Fourier transform infrared spectroscopy (FTIR) and Raman spectroscopy can also be used. For GelMA hydrogels, the binding of ME methyl groups to GelMA lysine CH 2 The ratio of groups. Figure 4B The results show that [ME methyl group and lysine CH 2 The ratio of the groups] decreases with the increase of exposure time.

[0329] b) Swelling ratio

[0330] A study analyzing the swelling ratio of hydrogels with different GelMA, HAMA, and PEGDA concentrations was completed.

[0331] G4-H was prepared according to the general procedure of Example 3 M 1-P1, G7-H M 1. G4-H M 1 and H M 1-P1 hydrogel (described in Table 1) with a photocrosslinking time of 4 minutes. The resulting hydrogel cylinder had a diameter of 6 mm and a volume of 75 μL.

[0332] To evaluate swelling, two methods were used. In the first method, the weight of the hydrogel just after cross-linking was used as the "dry" hydrogel weight (Wd-1), while in the second method, the dry polymer weight (vacuum dried hydrogel) was used as the dry hydrogel weight (Wd-2). In both cases, the "wet" hydrogel weight (Ws) refers to the hydrogel incubated in 1× PBS at 37°C for 48 hours. The swelling ratio was calculated as follows:

[0333] Swelling ratio = (Ws-Wd) / Wd

[0334] The results of the first measurement method are inconsistent, e.g. Figure 5A The results of the second measurement method are more consistent, as shown in Figure 5B As shown, increasing GelMA concentration plays a role in reducing hydrogel swelling.

[0335] Study G4-H M 1-P1, G7-H M 1. G4-H M 1 and H MSwelling / reswelling effect of 1-P1 hydrogel. The sample was dried and swelled using the second method, and then dried and re-swelled a second time. Figure 5C The results presented in show that the swelling ratio decreases significantly when the hydrogels are exposed to more than one drying / swelling cycle.

[0336] G4-P1, G4-P0.1, G20, G10, G5, P20 and P5 hydrogels (described in Table 1) were prepared according to the general procedure of Example 3, with a photocrosslinking time of 4 minutes. Swelling was assessed using the dry polymer weight (vacuum dried hydrogel) as the "dry" hydrogel weight (Wd), while the "wet" hydrogel weight (Ws) refers to the hydrogel incubated in 1× PBS at 37°C for 48 hours. Figure 5D The results presented in show that the swelling mass increased significantly with the inclusion of PEGDA, and increasing GelMA concentration also increased the swelling mass of the hydrogels.

[0337] c) Swelling ratio and active agent

[0338] A study analyzing the swelling ratio of hydrogels loaded with active agents and having different concentrations of GelMA, HAMA, and PEGDA was completed.

[0339] G4-H was prepared according to the general procedure of Example 3 M 1-P1, G4-H M 1. G7-H M 1. H M 1-P1, G4-P1, and G7-P1 hydrogels (described in Table 1) were photocrosslinked for 4 minutes. Samples of each hydrogel were also prepared with 13.2 mg / mL of corticosteroid active agent.

[0340] Swelling was assessed using the dry polymer weight (vacuum dried hydrogel) as the "dry" hydrogel weight (Wd), while the "wet" hydrogel weight (Ws) refers to the hydrogel incubated in 1× PBS at 37°C for 48 hours. The swelling ratio was calculated as follows:

[0341] Swelling ratio = (Ws-Wd) / Wd

[0342] Fig. 6A The results presented show that hydrogels loaded with active agents generally have higher swelling ratios, which may be due to the disruption of gel network crosslinks associated with the incorporation of active agents into the gel network and the lower crosslink density.

[0343] G4-H was also studied M 1-P1, G4-H M 1. G7-H M 1. H MSwelling / reswelling effects of 1-P1, G4-P1 and G7-P1 hydrogels (with active agent) The samples were dried and swelled, then dried and re-swelled a second time. Figure 6B The results presented in show that the swelling ratio of hydrogels containing MeHA decreased significantly when the hydrogels were exposed to more than one drying / swelling cycle, while hydrogels containing only GelMA+PEGDA showed minimal effect on re-swelling.

[0344] d) Enzymatic degradation

[0345] A study analyzing the enzymatic degradation stability of hydrogels with different GelMA, MeHA, and PEGDA concentrations was completed.

[0346] G4-H was prepared according to the general procedure of Example 3 G 3-P1, G4-H M 1-P0.67, G4-H G 3. G4-H M 1. G7-H G 3. G7-H M 1. H G 3-P1 and H M 1-P0.67 hydrogel (as described in Table 1) was photocrosslinked for 4 minutes. Then, 20 U / mL or 2 U / mL of hyaluronidase (Hy) and type I collagenase (C I ) or collagenase type II (C II ) were subjected to enzymatic digestion of the samples. The degradation times obtained are shown in Table 2.

[0347] Table 2 - Enzymatic degradation time

[0348]

[0349] e) Drug Release

[0350] A study analyzing the drug release rate of hydrogels with varying GelMA, MeHA, and PEGDA concentrations was completed.

[0351] G4-H was prepared according to the general procedure of Example 3 with 13.2 mg / mL of corticosteroid active agent. M 1-P1 and G4-H G 3-P1 hydrogel (described in Table 1) with a photocrosslinking time of 4 minutes. The resulting hydrogel cylinder had a diameter of 6 mm and a volume of 75 μL.

[0352] For release studies, the hydrogels were statically (without physical stirring) incubated at 37°C in 1 mL of 1×PBS supplemented with 2% Triton X-100 to simulate tear fluid. At each time point (after 10-13 days), the incubation solution was completely removed and replaced with fresh 1×PBS+2% Triton X-100. In order to quantify the release of corticosteroids, the samples were diluted 1:2 in acetonitrile and analyzed using reversed phase liquid chromatography. An Agilent Zorbax Eclipse (XDB-C18) 4.6x 250mm, 5μm analytical column was used on an Agilent 1290 HPLC system equipped with a diode array detector. The column was balanced with 70% acetonitrile and 30% water at 25°C. After injecting 20 μL of sample, the solvent gradient increased from 70% to 90% ACN in a time span of 10 minutes. When the ACN gradient reached approximately 80%, the corticosteroid eluted at nearly 5 minutes. This peak was integrated and the concentration was determined using the area under the curve by comparison to a standard curve of corticosteroids. Fig. 7A The results presented in show that hydrogels containing higher concentrations of MeHA provide faster release properties. These results correlate with the results of corresponding studies showing that higher concentrations of MeHA in hydrogels cause increased swelling of the hydrogels.

[0353] Based on the results of the swelling ratio study, higher concentrations of MeHA in the hydrogel may cause increased swelling of the hydrogel and thus a faster burst release of the active agent. Higher concentrations of MeHA may also lead to phase separation from GelMA in the precursor solution, which may result in gel network defects (i.e., regions of higher and lower cross-linking density), leading to a higher initial burst release.

[0354] G4-H M The release properties of 1-P1 last for 35 days ( Figure 7B ) and 65 days ( Figure 7C ). Also G4-H M The release properties of 1-P1 were compared with those of G4-P1 and G7-P1 ( Fig.7D ), again showing that the presence of MeHA in the hydrogel increases the release rate of the active agent from the hydrogel.

[0355] f) Vacuum drying

[0356] A study was completed to analyze the effect of vacuum drying on the drug release rate from hydrogels with varying GelMA, MeHA, and PEGDA concentrations.

[0357] G4-H was prepared according to the general procedure of Example 3 with 13.2 mg / mL of corticosteroid active agent. M1-P1, G4-P1 and G7-P1 hydrogels (as described in Table 1) were photocrosslinked for 4 minutes. Samples of each hydrogel were then vacuum dried. Release studies using wet and dry samples of each hydrogel were then completed according to the general study procedures of Example 3(e). G4-H M Results of 1-P1 hydrogel ( Fig. 8A ) showed that the release properties of hydrogels containing MeHA can be reduced by vacuum drying the hydrogels, such that the inclusion of MeHA in the hydrogel formulation can reduce the release properties of the dried samples while alternatively increasing the swelling and corresponding release properties of the undried samples. Results for G4-P1 and G7-P1 hydrogels ( Figure 8B ) showed that the release properties of GelMA+PEGDA hydrogels without MeHA were generally not affected by vacuum drying of the hydrogels.

[0358] g) Rods and discs

[0359] A study was completed analyzing the effect of hydrogel shape (ie, rods vs. disks) on drug release rates from hydrogels containing GelMA, MeHA, and PEGDA.

[0360] According to the general procedure of Example 3, G4-H was treated with 13.2 mg / mL of corticosteroid active agent. M 1-P1 hydrogels (as described in Table 1) were prepared in both disk and rod forms with a photocrosslinking time of 4 min.

[0361] G4-H M The diameter (D) of the 1-P1 hydrogel disk is 6 mm, the volume (V) is 75 μL, and the surface area (SA) is 107 mm 2 , and the SA:V ratio is 1.4.

[0362] G4-H M The diameter (D) of the 1-P1 hydrogel rod is 2 mm, the volume (V) is 25 μL, and the surface area (SA) is 56 mm 2 , and the SA:V ratio is 2.2.

[0363] The samples from the rod-shaped hydrogels were then vacuum dried or freeze dried (i.e., lyophilized). Release studies using the resulting wet and dry samples were then completed according to the general study procedures of Example 3(e). Results of total drug release ( Fig. 9A ) showed that cylindrical disks provided greater total release of active agent (possibly as a result of surface area), while rods 湿 ,Great 冻干 and stick 干 All had similar total release. Results for percentage drug release ( Fig. 9B) showed that wet hydrogels (cylindrical disks and rods) released a higher percentage of active agent than vacuum dried or freeze dried rod-shaped hydrogels. The results thus show that the swelling characteristics, surface area (i.e., shape), and hydration state of the hydrogel play a role in the drug release properties of the hydrogel composition.

[0364] h) Degree of crosslinking – degree of methacrylate

[0365] A study was completed to analyze the correlation between the release properties of GelMA+PEGDA hydrogels and the degree of methacrylate esterification of GelMA within the hydrogels.

[0366] G4(160P80)-P1(2K) and G4(160P40)-P1(35K) hydrogels (described in Table 1) were prepared according to the general procedure of Example 3 with 13.2 mg / mL corticosteroid active agent and a photocrosslinking time of 4 minutes. Release studies were then completed according to the general study procedure of Example 3(e), exposing each sample to collagenase II 0.5 U / mL conditions and non-enzymatic standard conditions. Results of total drug release ( Fig.10 ) showed that the lower 40% DoM in GelMA provided faster release properties than the higher 80% DoM GelMA hydrogel.

[0367] Example 5: Cell aggregation and viability studies

[0368] Human umbilical vein endothelial cell (HUVEC) populations were encapsulated in two separate hydrogel formulations: (i) G7(40)P1 [7% GelMA (160P40), 1% PEGDA (35 kDa)]; and (ii) P8(35 kDa) [8% PEGDA, 35 kDa]. After 16 days, the hydrogel samples were stained with calcein AM and ethidium homidmer-1 and imaged to observe cell aggregation and viability. The results are shown in Fig.11A In. Fig.11A As seen in , hydrogels comprising a combination of chemically modified gelatin (e.g., GelMA) and chemically modified PEG (e.g., PEGDA) reduced cell aggregation of 3D adherent cells (e.g., HUVEC cells) encapsulated within the hydrogel framework.

[0369] The two hydrogel formulations were also analyzed at day 3, day 8, and day 16 as percent Calcein AM (alive) / [percent Calcein AM (alive) + ethidium bromide dimer-1 (dead)] to quantify HUVEC cell viability in each hydrogel. The results are shown in Fig. 11B In. Fig. 11BAs seen in Figure 1, hydrogels containing a combination of chemically modified gelatin (e.g., GelMA) and chemically modified PEG (e.g., PEGDA) increased the cell viability of 3D adherent cells (e.g., HUVEC cells) encapsulated within the hydrogel framework. G7(40)P1 hydrogels maintained an average cell viability of 60-80% over a duration of 16 days, while P8(35kDa) provided an average cell viability of 20-40%.

[0370] Example 6: Study of cell growth on the underside of permeable cell culture chambers - HUVEC

[0371] Studies were completed to evaluate the ability of various hydrogel formulations to attach and deliver cells to membranes with challenging gravity requirements.

[0372] Populations of human umbilical vein endothelial cells (HUVECs) were added to two hydrogel precursor formulations: (i) G5(80) [5% GelMA, 80% DoM]; and (ii) P5(35kDa) [5% PEGDA, 35kDa], prepared in PBS as in Example 3. The cell concentration was approximately 10 million HUVECs / mL (GFP+).

[0373] Then the PET permeable cell culture chamber insert (pore size 0.4 μm) is incubated in serum containing endothelial growth medium for 15 minutes. The insert is wiped dry with an eye spear, and 13.2 μL of each hydrogel precursor solution (containing HUVEC) is added to the lower side of a separate permeable cell culture chamber and exposed to high-intensity white light for 1 minute (equipped with a Dolan-Jenner high-intensity LED illuminator MI-LED-US-B1 with a double-arm gooseneck structure). The permeable cell culture chamber is then inverted onto a well plate so that the hydrogel on the lower side of each permeable cell culture chamber is immersed in the endothelial growth medium supplemented with growth factors and serum. Free HUVEC (i.e., non-hydrogel encapsulated cells) is also added to the endothelial growth medium in the well plate as a control. After a few days (e.g., the 4th and 11th days), GFP+ imaging is used and confocal imaging is used to analyze the cell growth of each preparation on the 11th day.

[0374] a) GFP+ imaging results

[0375] The results of GFP+ imaging are shown in Figures 12A-12I middle.

[0376] For G5(80) - After 4 days, samples showed that HUVECs were still localized on the underside of the permeable cell culture insert ( Fig. 12A ), with strong endothelial cell viability and clear cell network formation (see Fig. 12AAfter 11 days, the G5(80) sample continued to show HUVEC localization on the underside of the permeable cell culture insert ( Fig. 12B ), with continued strong endothelial cell viability and clear cell network formation (see Fig. 12B ), and cell debris was found only on the bottom of the well plate.

[0377] For P5 (35 kDa) - after 4 days, the sample showed that part of the hydrogel had detached from the underside of the permeable cell culture insert, and some HUVECs were still localized on the underside of the permeable cell culture insert ( Fig. 12C ), other HUVEC migrated to the bottom of the well plate ( Fig.12D ). Overall cell density was low in both locations. After 11 days, the P5 (35 kDa) sample continued to show low cell density and minimal cell network formation on the underside of the permeable cell culture insert ( Fig.12E ), few viable cells remained, and most of the cell debris found was localized on the bottom of the well plate.

[0378] For free cell control samples (i.e., no hydrogel) - after 4 days, the control samples showed very little HUVEC attachment to the underside of the permeable cell culture insert ( Fig.12F ), most cells migrated to the bottom of the well ( Figure 12G The overall cell density was low. After 11 days, the control sample continued to show low cell density and minimal cell network formation on the underside of the permeable cell culture insert ( Fig.12H ), the remaining living cells localized on the bottom of the well plate ( Fig.12I ), the cells were elongated but little cell network formation was observed.

[0379] b) Confocal imaging results

[0380] On day 11, hydrogel samples of G5 (80) and P8 (35 kDa) from Example 6(a) were stained for: actin (cytoskeleton showing cell spreading and network formation); PECAM / CD31 (endothelial marker); and DAPI (nuclei). The stained samples were then confocal imaged; the results are shown in Figures 13A-13E middle.

[0381] The G5(80) sample showed high HUVEC deposition on the underside of the permeable cell culture insert ( Fig.13A and Fig. 13B ), including the formation of a dense cell network with well-distributed cells and no signs of cell clumping.

[0382] The P8 (35 kDa) sample showed limited HUVEC deposition on the underside of the permeable cell culture insert ( Fig. 13C and Fig.13D ), including low cell density, patchy distribution, high cellular debris content, limited cell spreading and network formation, absence of CD31 expression (possibly associated with poor endothelial cell function), and localization of actin filaments to the cell surface.

[0383] When compared to the G5(80) sample, the free cell (i.e., no hydrogel) control sample showed lower cell density and poor network formation ( Fig.13E ).

[0384] c) Testing of Additional Formulations

[0385] Additional hydrogel formulations were analyzed according to the procedure of Example 6(a). The results of imaging and analysis of the underside of the permeable cell culture insert are shown in Table 3.

[0386] Table 3- GFP+HUVEC analysis

[0387]

[0388]

[0389] d) Research Observations

[0390] The results showed that HUVECs in GelMA-based formulations spread more effectively, produced stronger cell networks, and aggregated less than PEGDA-based formulations. The results also showed that higher crosslinking density (e.g., higher polymer concentration, higher DOM) does not generally promote strong cell spreading and network formation. GelMA-based materials also provide stronger adhesion to the test surface (i.e., the lower surface of the permeable cell culture chamber) and more effectively deposit cells to form dense cell networks, even in challenging environments and growth conditions (e.g., against gravity).

[0391] Example 7: Study of cell growth on the underside of permeable cell culture chambers - HRPEC

[0392] Studies were completed to evaluate the ability of various hydrogel formulations to attach and deliver cells to membranes in a challenging gravity environment simulating the subretinal space.

[0393] Populations of human retinal pigment epithelial cells (HRPECs) were added to the following hydrogel precursor formulations: (i) G2(80)H2(500kDa,30)[2% GelMA,80% DOM; 2% HAMA,500kDa,~30% DOM]; (ii) G5(80)[5% GelMA,80% DoM]; (iii) G4(80)[5% GelMA,80% DoM]; (iv) H2(500kDa,30)[2% HAMA,500kDa,~30% DOM]; (v) P5(35kDa)[5% PEGDA,35kDa], prepared in PBS as in Example 3. The cell concentration was approximately 20 million HRPECs per mL.

[0394] The PET permeable cell culture inserts (pore size 0.4 μm) were then incubated in serum containing DMEM / F-12 medium (supplemented with 10% FBS) for 15 minutes. The inserts were wiped dry with an eye spear, and 13.2 μL of each hydrogel precursor solution (containing HRPEC) was added to the lower side of the individual permeable cell culture chamber and exposed to light for 1 minute. The permeable cell culture chambers were then inverted onto the well plate so that the hydrogel on the lower side of each permeable cell culture chamber was immersed in DMEM / F-12 medium (supplemented with 10% FBS). After several days (e.g., day 3, day 8, day 10, day 20), cell growth of each formulation was imaged and analyzed, including staining and imaging with calcein AM.

[0395] The results are shown in Figures 15A-15G middle.

[0396] For G2(80)H2(500 kDa, 30) - after 3 days, the samples showed that HRPECs were still localized within the hydrogel on the underside of the permeable cell culture insert and failed to form a cell monolayer on the surface of the permeable cell culture insert ( Fig.15A ). Similar localization results were observed after 10 days without HRPEC monolayer formation ( Fig. 15B ).

[0397] For G5(80) - after 8 days, the samples showed well-dispersed round cells, but also showed minimal cell spreading and no cell network formation ( Fig. 15C ).

[0398] For H2 (500 kDa, 30) - after 7 days, the samples showed well dispersed round cells, but also showed minimal cell spreading and no cell network formation ( Fig.15D ).

[0399] For G4(80) - after 8 days, the hydrogel showed rapid degradation / detachment but also appeared to provide HRPEC monolayers with good coverage and viability, including mature monolayer formation zones ( Fig.15E After 20 days, a mature HRPEC monolayer was shown to have formed over the entire permeable cell culture chamber membrane surface ( Fig.15F ).

[0400] For P5 (35 kDa) - after 8 days, the hydrogel showed rapid degradation / detachment and also appeared to provide HRPEC monolayers with incomplete coverage and inconsistent monolayer formation ( Figure 15G ).

[0401] a) Research Observations

[0402] The results of the study showed that HRPEC had minimal spreading and networking in hydrogel formulations with high methacrylation degree, high molecular weight and / or high hydrogel polymer concentration (e.g., G2(80)H2(500 kDa, 30). HRPEC had improved spreading and networking in hydrogel formulations with lower methacrylation degree and lower hydrogel polymer concentration, allowing efficient HRPEC deposition and monolayer formation.

[0403] In addition, PEG-based hydrogels such as PEGDA are generally shown to have poor bioadhesion and poor biodegradability (often detaching from underneath the permeable cell culture insert in as little as 24-48 hours). The remaining HRPECs on the surface of the permeable cell culture insert are believed to be those that were captured on the surface of the PEGDA hydrogel during the photopolymerization process. The cells are unable to attach or migrate through the PEGDA polymer network, but remain captured and can only attach to neighboring cells.

[0404] Example 8: Permeable Cell Culture Insert Apical Cell Growth Study - HRPEC

[0405] Studies were completed to evaluate the ability of various hydrogel formulations to degrade and deliver cells to the membrane.

[0406] Populations of human retinal pigment epithelial cells (HRPECs) were added to two hydrogel precursor formulations: (i) G5(10) [5% GelMA, 10% DoM]; and (ii) G5(80) [5% GelMA, 80% DoM], prepared in PBS as in Example 3. The cell concentration was approximately 20 million HRPECs per mL.

[0407] The PET permeable cell culture inserts (pore size 0.4 μm) were then incubated in serum containing DMEM / F-12 medium (supplemented with 10% FBS) for 15 minutes. The inserts were dried with an eye spear, and approximately 10 μL of each hydrogel precursor solution (containing HRPEC) was added to the top side of a separate permeable cell culture insert and exposed to light for 1 minute. The permeable cell culture inserts were then immersed in DMEM / F-12 medium (supplemented with 10% FBS). Free HRPEC (i.e., no hydrogel) was also added to the medium in the well plate as a control.

[0408] After 24 h, the hydrogel samples were stained with Calcein AM and imaged to observe cell aggregation and viability ( Fig.16A ). Also 3 days later ( Fig. 16B ) and 6 days later ( Fig. 16C )Analyze the samples.

[0409] In 24 hours ( Fig.16A )--The cell-only control sample showed uninhibited cell growth and monolayer formation. The G5(10) sample showed that the hydrogel began to degrade, deposited HRPEC, and began to form a monolayer network. The G5(80) sample showed no hydrogel degradation and no monolayer formation, with cells generally remaining round and suspended within the hydrogel.

[0410] At 3 days ( Fig. 16B )--The cell-only control sample continued to show uninhibited cell growth and monolayer formation, in which the cells showed dense packing. The G5(10) sample showed that the hydrogel was mostly degraded, and the deposited HRPEC formed a transparent monolayer network similar to the control sample at 24 hours, with a few round encapsulated cells remaining above the monolayer. The G5(80) sample continued to show little hydrogel degradation and little monolayer formation, in which the cells remained generally round and suspended within the hydrogel.

[0411] At 6 days ( Fig. 16C )--The cell-only control sample continued to show uninhibited cell growth and monolayer formation with packed cell density and networking. The G5(10) sample showed hydrogel degradation, and the deposited HRPEC monolayer network was similar in density and morphology to the control sample, with few round cells remaining above the monolayer. The G5(80) sample showed that the hydrogel began to degrade in some areas and formed an early monolayer ( Fig. 16C The upper right corner of the G5(80) image in Figure 5 ), while other regions remained undegraded and no monolayer was formed ( Fig. 16C (bottom left corner of the image of G5(80) in the middle), HRPECs remain round and suspended within the hydrogel.

[0412] Example 9: In vitro retinal cell growth study

[0413] Studies were completed to evaluate the G5(10) [5% GelMA, 10% DoM] hydrogel formulation to promote in vitro delivery and cell growth of stem cell-derived retinal cells, including retinal pigment epithelial (RPE) cells and rod photoreceptor cells.

[0414] a) Monolayer formation studies

[0415] A G5(10) [5% GelMA, 10% DoM] hydrogel precursor formulation was prepared according to the general procedure in Example 3, wherein the concentration of the photopolymerization initiator mixture was reduced. A population of human retinal pigment epithelial cells (HRPECs) derived from induced pluripotent stem cells (iPSCs) was then added to the G5(10) hydrogel precursor formulation (~20 million cells / mL). iPSC-derived RPE was also added to the saline solution as a control. Approximately 200 μL of each formulation was injected (in PBS) into a 6-well permeable cell culture insert plate with a 0.4 μm polyester (polyethylene terephthalate) membrane using a 38G (outer diameter) (41G inner diameter) subretinal cannula (using a standard 1 mL luer-lok plastic syringe) at a controlled rate of 200-300 μL / min (flow rate controlled with a syringe pump). The polyester membrane was included to mimic the natural Bruch's membrane; the membrane is permeable to nutrients and proteins, but not cells. The injected formulations were then photocrosslinked in 10 μL drops using white light.

[0416] One week later, live cell imaging was performed using Calcein AM. The results are shown in Fig.17A The results showed that RPE cells were able to slowly degrade the G5(10) hydrogel substrate after cross-linking and then migrate to the permeable cell culture chamber membrane. RPE cells formed a transparent cell monolayer after one week, similar to the target anatomical pattern of functional RPE cells (see Fig.17A ). RPE cells delivered in a saline (control) formulation failed to form an observable cell monolayer after one week despite being delivered at the same seeding density as cells in the hydrogel formulation.

[0417] Fluid shear rates associated with extrusion through a 38G (41G ID) small diameter needle were measured during sample delivery. Results (see Fig. 17B ) showed that the GelMA prepolymer can be shear-thinned in the precursor polymer solution, thereby enabling protective plug flow dynamics through a small diameter needle, which would protect the passenger cells.

[0418] b) iPSC dedifferentiation studies

[0419] The precursor formulation from Example 9a was prepared to ~1 million cells / mL and then injected (in PBS) into a 6-well permeable cell culture insert plate with a 0.4 μm polyester (polyethylene terephthalate) membrane using the same delivery system and conditions as Example 9A. The injected formulation was then photocrosslinked in 10 μL drops using white light. One week later, live cell imaging was completed using Calcein AM. The results are shown in Fig. 17C In. The results of the study show that RPE cells form a monolayer, in which cells remain as the cubic morphology and pigmentation pattern of the feature of standard healthy RPE morphology. The RPE cells delivered in saline (control) preparations become elongated and highly proliferated, and fail to maintain its cubic morphology and RPE pigmentation pattern indicating dedifferentiation.

[0420] Without being bound by theory, observations suggest that the hydrogel substrate provides a localized and concentrated environment for RPE cells, thereby enabling monolayer formation with characteristic RPE morphology by reducing RPE dedifferentiation due to environmental factors.

[0421] c) Rod photoreceptor delivery studies

[0422] G5 (10) [5% GelMA, 10% DoM] hydrogel precursor preparations were prepared according to the general procedure in Example 3, wherein the concentration of the photopolymerization initiator mixture was reduced. A population of rod photoreceptor cells (LakoLab, Newcastle University) derived from human retinal organoids was then added to the G5 (10) hydrogel precursor preparation. The preparation was then injected (in PBS) onto the RPE monolayer in a 6-well permeable cell culture chamber plate with a 0.4 μm polyester (polyethylene terephthalate) membrane using the same delivery system and conditions as in Example 9A. The injected preparation was then photocrosslinked using white light.

[0423] Two weeks later, live cell imaging was performed using anti-CD73 and anti-rhodopsin imaging. The results are shown in Fig.17D The results show that rod photoreceptor cells ( Fig.17D Upper left) localized in the RPE monolayer ( Fig.17D ) and remained viable at two weeks, with both rod photoreceptor cells and RPE cells expressing CD73. Only rod photoreceptor cells showed expression of rhodopsin ( Fig.17D on the right side of the ).

[0424] Example 10: In vivo retinal cell growth study

[0425] A study was performed to evaluate the cellular localization and viability of retinal pigment epithelial (RPE) cells following a single subretinal (SR) injection in pigs of one of three G5 [5% GelMA] hydrogel formulations photopolymerized in situ by intraocular surgery.

[0426] The four test groups consisted of 200 μl injections into 6 eyes (a total of 24 eyes and 12 animals for each group):

[0427] Group 1 (control): saline + 200k RPE cells; Group 2: G5 (10) [5% GelMA, 10% DoM] + 200k RPE cells; Group 3: G5 (80) [5% GelMA, 80% DoM] + 200k RPE cells; Group 4: [5% GelMA, 80% DoM] G5 (80) + 1m RPE cells.

[0428] Animals in groups 2-4 received a single SR injection of RPE with a specified hydrogel formulation. A single injection of RPE in saline was administered to group 1 as a control. Oral prednisone was given preoperatively and throughout the study. For injection, a 38G / 41G MedOne subretinal needle equipped with a 6-7 inch extension tube was used. Freshly prepared hydrogel precursor and cell mixtures were prepared before injection to each group of animals. Irradiation was performed using a Constellation 23G light tube, which emitted approximately 27mW of optical power (as measured at 517nm) at maximum power.

[0429] All animals underwent ophthalmological examination, color funduscopy, and ocular coherence tomography (OCT) after injection and on days 1, 3, 7, 14, 21, and 28. All eyes included were stained for STEM121 (transplanted human cells), Iba-1 (microglia / leukocytes), RPE65 (RPE cells), and DAPI (all cell nuclei).

[0430] Two eyes were also selected from each group for immunofluorescence staining. Eye blocks were sagittally sectioned (14 μm) at 5 levels throughout the eye: nasal periphery, midway between the nasal and optic nerve head (ONH), ONH, midway between the ONH and the temporal, and temporal periphery. At least 10 slides / level were collected. Slides were scanned during cell reservoir acquisition. A series of 8-10 slides / eye were selected for IHC.

[0431] Fig.18AObservations related to cell migration to the retinal surface are shown in FIG. Due to poor cell localization (i.e., cells outside the subretinal area), the saline control formulation showed more cell migration (over 80%) and corresponding adverse events in the outer retinal layer. This finding was confirmed by histological examination of the eyes after day 28, in which vitreous cells were found, while the subretinal hydrogel formulations showed very little cell migration of about 20% or less.

[0432] Fig.18B Observations related to hydrogel degradation after 28 days are shown in FIG. 1 . Group 1 (control) contained no hydrogel. For Group 2, only about 33% of the hydrogel remained after 28 days (i.e., a loss of about 67%). Groups 3 and 4 had about 100% of the hydrogel remaining after 28 days (i.e., a loss of about 0%). After 28 days, the hydrogels with 80% high methacrylate (Groups 3 and 4) showed detachment of the retina from the underlying layers, as shown in FIG. Fig. 18C and Fig.18D The staining results indeed showed that human stem cells were successfully transplanted onto the natural pig RPE layer, and the RPE layer was not damaged (see Fig.18E , where arrows indicate human cells).

[0433] Example 11: Free Radical Propagator Study

[0434] A study was completed to evaluate N-vinyl caprolactam (NVC), N-vinyl pyrrolidone (NVP), and ethylene glycol diacrylate (EGDA) in hydrogel formation.

[0435] Each of the three propagating agents (NVC, NVP, EGDA) was combined with G5(80) [5% GelMA, 80% DoM] at a propagating agent concentration of 10uL / mL and then gelled. The average Young's modulus (kPa) of the resulting hydrogels was as follows: EGDA [7.2494kPa]; NVP [6.4995kPa]; NVC [3.4744kPa]. The minimum concentrations of NVP and EGDA that still allowed hydrogel formation in G5(10) and G5(80) (crosslinking at ~0.43W for 1 minute) were also tested. The minimum propagating agent concentrations for hydrogel formation were as follows: G5(10) [NVP-5uL / mL; EGDA-1.5uL / mL]; G5(80) [NVP-0.3uL / mL; EGDA-0.1uL / mL]

[0436] Then G5 (10) -1.5uL / mL EGDA and G5 (10) -5uL / mL NVP were squeezed out by needle to study the encapsulation of RPE cells (with PBS as a control). After initial squeezing, all three samples showed that cells maintained pigmentation and morphology. After 24 hours, PBS cells lost pigmentation and showed early signs of epithelial-mesenchymal transition (EMT). Both hydrogel preparations showed that cells maintained pigmentation and morphology. The same results were observed at the 6th day.

[0437] The following cytotoxicity studies were also completed on RPE cells: NVC, NVP, EGDA, and phenylacetyl bromide (BAP), with 10% DMSO as a control. Fig.19 The cell viability results after cells were incubated in each of the four propagating agents for 18 hours are shown in . The results show that NVP and BAP have higher average cell viability than NVC at all relative concentrations, while EGDA has low cell viability results.

[0438] Example 12: G5 Formulation Study Using N-vinylpyrrolidone (NVP)

[0439] The gelation and cell encapsulation properties of G5 formulations with different average molecular weights [5% GelMA, different DoM and MW] were studied. Specifically, G5 (10) 160 kDa, G5 (60) 90 kDa and G5 (45) 160 kDa were mixed with standard photoinitiator concentrations (but using 1% NVP instead of NVC) and 5M ARPE-19 cells / mL formulation (a total of ~200k cells per gel). Samples (including squeezed saline with cells) were injected into the sample container using a MedOne 38 / 41G subretinal needle (300 μL / min injection rate) and a light pipe (set to 26-27 mW) was used to simulate the Constellation vitrectomy setting (exposure 30 seconds). Saline (non-squeezed) was used as a normalization control.

[0440] Fig. 20 Cell viability results using Calcein AM (normalized to non-extruded saline) are shown in Figure 3. All formulations showed at least 150% normalized cell viability, with G5(10)160kDa and G5(60)90kDa showing more than 200% normalized cell viability. After 48 hours, gel degradation was evident in all three hydrogel samples, and cells attached to the permeable cell culture insert surface and formed an early monolayer. Few cells remained round and unattached.

[0441] Further studies were completed to analyze the gelation properties of various G4 formulations [4% GelMA, different DoM and MW] and G5 formulations [5% GelMA, different DoM and MW] using NVP as a free radical propagator. Table 4 presents the study conditions and results for various formulations.

[0442] Table 4 - NVP gelation studies

[0443]

[0444] Additional polymer blends using GelMA concentrations ranging from 0.5% to 3.0% were also tested for gelation and cell encapsulation properties. Table 5 presents the study conditions and results for various formulations.

[0445] Table 5 - NVP Gelation Studies - Low GelMA %

[0446]

[0447]

[0448] Example 13: Combination Formulation Study

[0449] G1 (160 / 10) G2.5 (90 / 60) blends were studied with various photoinitiator formulations including 1.5% v / v triethanolamine (TEOA) and polymerization conditions. Table 6 presents the study conditions and results for various formulations.

[0450] Table 6 - Combination Formulation Studies

[0451]

[0452] Rxn 3 and Rxn 4 were further studied. 10M cells / mL were added to each of Rxn 3, Rxn 4, and PBS (as a control). Rxn 3 samples, Rxn 4 samples, and a portion of the PBS control were extruded onto the surface of the permeable cell culture insert, and unextruded PBS was applied to the surface of the permeable cell culture insert. After two weeks, both Rxn 3 and Rxn 4 showed healthy RPE monolayer growth, similar to the PBS control, with the Rxn 3 sample providing higher cell counts than either PBS control sample. Fig.21 The cell counting results are shown in .

Claims

1. A polymer composition, comprising: (i) from about 0.5% to about 5.0% w / v chemically modified gelatin; (ii) at least one polymer cross-linking initiator; and (iii) at least one cell.

2. The polymer composition of claim 1, wherein the polymer cross-linking initiator comprises one or more photoactivated photoinitiators; optionally one or more photoinitiators activated by visible light.

3. The polymer composition of claim 2, wherein the polymer cross-linking initiator comprises: (i) Eosin Y, N-vinyl caprolactam (NVC), triethanolamine, or any combination thereof; (ii) Eosin Y disodium salt (EYDS), N-vinyl caprolactam (NVC), triethanolamine, or any combination thereof; or (iii) Eosin Y disodium salt (EYDS), N-vinyl pyrrolidone (NVP), triethanolamine, or any combination thereof.

4. The polymer composition of claim 2, wherein the polymer cross-linking initiator comprises: (i) about 50 μM of Eosin Y or Eosin Y disodium salt (EYDS); (ii) about 3.5 to about 5.0 μL / mL of N-vinyl caprolactam (NVC) or N-vinyl pyrrolidone (NVP); and (iii) triethanolamine.

5. The polymer composition of claim 2, wherein the polymer cross-linking initiator comprises: (i) about 50 μM of eosin Y disodium salt (EYDS); (ii) about 5.0 μL / mL of N-vinyl pyrrolidone (NVP); and (iii) about 1.5% v / v of triethanolamine.

6. The polymer composition according to any one of claims 1 to 5, wherein the chemically modified gelatin is acrylated gelatin.

7. The polymer composition of claim 6, wherein the acrylated gelatin has a degree of acrylate of about 5-40%; optionally 5-20%; optionally about 5%, about 10% or about 15%.

8. The polymer composition according to any one of claims 1 to 5, wherein the chemically modified gelatin is methacrylated gelatin (GelMA).

9. The polymer composition of claim 8, wherein the GelMA has a degree of methacrylation of about 5-40%; optionally 5-20%; optionally about 5%, about 10% or about 15%.

10. The polymer composition of any one of claims 1-9, wherein the polymer composition comprises from about 2% to about 5% w / v of the chemically modified gelatin; optionally from about 3% to about 5% w / v of the chemically modified gelatin.

11. The polymer composition of claim 10, wherein the polymer composition comprises about 3% to about 4% w / v of the chemically modified gelatin; optionally about 3.5% w / v.

12. The polymer composition of claim 10, wherein the polymer composition comprises about 3% to about 4% w / v GelMA; optionally about 3.5% w / v GelMA.

13. The polymer composition of any one of claims 1-10, wherein the polymer composition comprises a combination of a first GelMA mixture and a second GelMA mixture.

14. The polymer composition of claim 13, wherein the polymer composition comprises about 0.5% to about 3% w / v of the first GelMA mixture and about 0.5% to about 3% w / v of the second GelMA mixture; optionally about 0.5% to about 1.5% w / v of the first GelMA mixture and about 1.5% to about 3% w / v of the second GelMA mixture; optionally about 1% w / v of the first GelMA mixture and about 2.5% w / v of the second GelMA mixture.

15. A polymer composition according to claim 13 or claim 14, wherein the first GelMA mixture comprises a GelMA having a high average molecular weight and a low degree of methacrylateization (DOM); and the second GelMA mixture comprises a GelMA having a low average molecular weight and a high (DOM); optionally, wherein the first GelMA mixture comprises a GelMA having an average molecular weight of 140-180 kDa and a DOM of 5% to 40%, and the second GelMA mixture comprises a GelMA having an average molecular weight of 75-115 kDa and a DOM of 50% to 80%; optionally, wherein the first GelMA mixture comprises a GelMA having an average molecular weight of 140-180 kDa and a DOM of 5% to 20%; and the second GelMA mixture comprises a GelMA having an average molecular weight of 80-100 kDa and a DOM of 50% to 70%.

16. The polymer composition of claim 13 or claim 14, wherein the first GelMA mixture comprises a GelMA having an average molecular weight of about 160 kDa and a DOM of about 10%; and the second GelMA mixture comprises a GelMA having an average molecular weight of about 90 kDa and a DOM of about 60%.

17. The polymer composition of claim 13, wherein the polymer composition comprises: about 1% w / v of a first GelMA mixture comprising GelMA having an average molecular weight of about 160 kDa and a DOM of about 10%; and about 2.5% w / v of a second GelMA mixture comprising GelMA having an average molecular weight of about 90 kDa and a DOM of about 60%.

18. The polymer composition of any one of claims 1-17, wherein the at least one cell comprises an endothelial cell; optionally a human umbilical vein endothelial cell (HUVEC).

19. A polymer composition according to any one of claims 1-17, wherein the at least one cell comprises an epithelial cell; optionally a human retinal pigment epithelial cell (HRPEC); optionally a HRPEC derived from an embryonic stem cell or an induced pluripotent stem cell (iPSC).

20. The polymer composition of any one of claims 1-17, wherein the at least one cell comprises an ocular cell; optionally an ocular cell derived from a pluripotent stem cell or an embryonic stem cell.

21. A polymer composition according to any one of claims 1-20, further comprising at least 0.1% (w / v) of a hydrophilic nonionic surfactant; optionally wherein the hydrophilic nonionic surfactant comprises at least one poloxamer surfactant, such as poloxamer 407; optionally wherein the composition comprises about 0.2% (w / v) of a poloxamer surfactant, such as poloxamer 407.

22. The polymer composition of any one of claims 1-21, further comprising one or more non-cellular therapeutic agents; optionally wherein the one or more non-cellular therapeutic agents comprise a small molecule or protein therapeutic.

23. A precursor polymer composition comprising the polymer composition according to any one of claims 1-22.

24. A gel polymer composition, wherein the gel polymer composition is formed by photocrosslinking the precursor polymer composition of claim 23; optionally wherein the gel polymer composition is a hydrogel.

25. The gel polymer composition of claim 24, wherein the shape of the polymer composition conforms to the shape of a target surface; optionally wherein the polymer composition conforms to a convex, concave or curved shape of the target surface.

26. A gel polymer composition according to claim 24, wherein the polymer composition is in the shape of a cylinder; optionally wherein the polymer composition is in the shape of a disc cylinder or a rod cylinder; optionally wherein the polymer composition is in the shape of a rod cylinder, the rod cylinder having a diameter of about 0.75 mm and a length of about 3 mm, or having a diameter of about 0.75 mm and a length of about 6 mm.

27. A method for treating and / or repairing a defect, injury and / or disease in a target soft tissue of a subject, the method comprising: Providing a precursor polymer composition according to claim 23; applying the precursor polymer composition onto or beneath the surface of the target soft tissue of the subject, optionally to the site of the soft tissue defect, injury and / or disease; as well as The precursor polymer composition is crosslinked by exposing the polymer crosslinking initiator in the polymer composition to crosslinking conditions, wherein the crosslinking of the precursor polymer composition produces a gel polymer composition.

28. A method for treating a defect, injury and / or disease in a target soft tissue of a subject, the method comprising: Providing a gel polymer composition according to any one of claims 24-26; as well as The gel polymer composition is applied to the subject on, below or near the surface of the target soft tissue, optionally at the site of the soft tissue defect, injury and / or disease.

29. The method of any one of claims 27-28, wherein the target soft tissue is ocular tissue; optionally subconjunctival ocular tissue or retinal ocular tissue.

30. The method of claim 29, wherein the polymer composition is applied onto or beneath the surface of the ocular tissue by subconjunctival injection, subretinal injection, or suprachoroidal injection.

31. The method of any one of claims 27-30, wherein the defect, injury and / or disease of the target soft tissue comprises an eye defect, injury and / or disease; optionally an eye ulcer; optionally a corneal ulcer caused by infection, injury, perforation or other defect.

32. The method of claims 27-30, wherein the ocular defect, injury and / or disease comprises a retinal degenerative disease; optionally age-related macular degeneration (AMD) or retinitis pigmentosa.

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