WO2025106906A1

WO2025106906A1 - Supramolecular hydrogels and uses thereof

Info

Publication number: WO2025106906A1
Application number: PCT/US2024/056254
Authority: WO
Inventors: Alireza MOSHAVERINIA; Weihao YUAN
Original assignee: University of California Berkeley; University of California San Diego UCSD
Current assignee: University of California Berkeley; University of California San Diego UCSD
Priority date: 2023-11-16
Filing date: 2024-11-15
Publication date: 2025-05-22
Anticipated expiration: 2026-05-16

Abstract

Hydrogel compositions are described with dynamic and tunable properties, mimicking extracellular matrix, for use in enhancing tissue remodeling, tissue engineering or wound healing, among others.

Description

SUPRAMOLECULAR HYDROGELS AND USES THEREOF

CROSS-REFERENCE TO RELATED APPLICATIONS

[001] This application claims priority to U.S. Provisional Application serial no. 63/599,856, filed November 16, 2023, which is incorporated herein by reference in its entirety.

INCORPORATION BY REFERENCE OF SEQUENCE LISTING

[002] The instant application contains a Sequence Listing conforming the rules of WIPO Standard ST.26 which has been submitted electronically in XML format and is hereby incorporated by reference in its entirety. The XML copy, created on November 14, 2024, is named P-623311-PC_SQL_ST26_15NOV24.xml and is 24,603 bytes in size.

FIELD

[003] The disclosure is directed to supramolecular hydrogel compositions and uses, for example in enhancing tissue remodeling, tissue engineering and wound healing.

BACKGROUND

[004] Biomaterials-based scaffolds hold considerable promise with respect to enhancing the efficacy of tissue engineering and regenerative medicine, and hydrogels rank among the most ideal carriers for stem cell delivery. Hydrogels are highly hydrated polymer networks, and their architectures and properties can be finely tuned according to the specific application. Previous studies have shown that hydrogels, including hyaluronic acid (HA, poly(lactic-co-glycolic acid) (PLGA, poly (2-hydroxyethylmethacrylate) (HEMA), nanofiber hydrogel, and self-assembly peptide hydrogel, can be engineered to mimic the architecture of the lost extracellular matrix (ECM) in tissue defects and can structurally support cell growth and differentiation. However, most of the existing hydrogels cannot completely recapitulate the dynamic properties of natural ECM, which are critical for supporting various cellular activities essential for tissue regeneration, including migration, proliferation, differentiation, and morphogenesis. Hydrogels with dynamic networks would better support the expansion and differentiation of the encapsulated stem cells and recruitment of host immune cells than hydrogels with the encapsulated stem cells alone.

[005] The present disclosure is directed to improved hydrogels with enhanced dynamic properties.

SUMMARY

[006] In one aspect, a dynamic supramolecular hydrogel composition is provided comprising:

(a) a first polymer comprising first polymer subunits, wherein the first polymer further comprises covalently-bound cyclodextrin moieties; and

(b) a second polymer comprising second polymer subunits, wherein the second polymer further comprises covalently -bound hydrophobic moieties; wherein the first polymer and the second polymer are noncovalently bound by the cyclodextrin moieties non-covalendy bound to the hydrophobic moieties.

[007] In some embodiments, the first polymer and the second polymer are each independently selected from a peptide, a polypeptide, a protein, a polysaccharide, a nucleic acid, or repeating subunits of one or more monomers. In some embodiments, the first polymer subunits and the second polymer subunits are the same. In some embodiments, the ratio of first polymer subunits comprising non-covalently bound cyclodextrin moieties to the total first polymer subunits is about 0.01 to about 0.8, or about 0.17 to about 0.36, or about 0.17 to about 0.29. In some embodiments, the cyclodextrin moieties are a- cyclodextrin, 0- cyclodextrin, or y-cyclodextrin, or any combination thereof. In some embodiments, the cyclodextrin is covalently bound to the first polymer subunits by maleimide groups covalently bound to the first polymer subunits reacted with thiol groups on the cyclodextrin. In some embodiments, the first polymer is gelatin. [008] In some embodiments, the first polymer comprises a cyclodextrin covalently bound to the gelatin as represented in Formula II:

wherein represents a cyclodextrin moiety and

gelatin.

[009] In some embodiments, the first polymer is gelatin covalently bound to [3- cyclodextrin.

[010] In some embodiments, the first polymer further comprises one or more bioactive peptide, an antibody, or a bioactive protein. In some embodiments, the bioactive peptide is a neuron growth and differentiation-inductive peptide. In some embodiments, the one or more bioactive peptide, antibody or bioactive protein is selected from or comprises nerve growth factor, RGD or a peptide comprising RGD, an integrin binding peptide, or a fragment of any of the foregoing, or a mimetic of any of the foregoing. In some embodiments, the bioactive peptide comprises RGD, such as GCGYGRGDSPG (SEQ ID NO:1). In some embodiments, the bioactive peptide, antibody or bioactive protein comprises a thiol group and is bound to maleimide groups on the first polymer. In some embodiments, the dynamic supramolecular hydrogel composition further comprises a hydrophobic molecule bound to cyclodextrins that are not non-covalently bound to the second polymer. In some embodiments, the hydrophobic molecule is a hydrophobic drug.

[011] In some embodiments, the ratio of second polymer subunits comprising covalently- bound hydrophobic moieties to the total second polymer subunits is about 0.01 to about 0.8, or about 0.34. In some embodiments, the covalently-bound hydrophobic moieties are adamantane moieties. In some embodiments, the second polymer is a polysaccharide. In some embodiments, the polysaccharide is hyaluronic acid. In some embodiments, the hyaluronic acid has a molecular weight of about 10 kDa to about 400 kDa. In some embodiments, the hyaluronic acid has a molecular weight of about 60 kDa to about 400 kDa. In some embodiments, the hyaluronic acid has a molecular weight of between about 60 kDa and about 80 kDa.

[012] In some embodiments, the second polymer comprising covalently -bound adamantane moieties comprises a polymer of Formula I:

wherein y/(x + y) is between about 0. 01 and about 0.8.

[013] In some embodiments, the second polymer further comprises one or more bioactive peptide, antibody or bioactive protein. In some embodiments, the bioactive peptide is a neuron growth and differentiation-inductive peptide. In some embodiments, the bioactive peptide, an antibody, or a bioactive protein is selected from or comprises nerve growth factor, RGD or a peptide comprising RGD, an integrin binding peptide, a fragment of any of the foregoing, or a mimetic of any of the foregoing. In some embodiments, the bioactive peptide comprises RGD, such as GCGYGRGDSPG (SEQ ID NO:1). In some embodiments, the one or more bioactive peptide, antibody or bioactive protein is covalently bound to the second polymer.

[014] In one aspect, the dynamic supramolecular hydrogel composition as described herein remodels in response to a traction force during cell spreading. In some embodiments, the ratio of polymer subunits comprising covalently-bound hydrophobic moieties to the total polymer subunits provides for the dynamic property. In some embodiments, the dynamic property is stiffness, strength, toughness, self-healing, shear-thinning, fast stress relaxation, and easy matrix remodeling. In some embodiments, the composition mimics the dynamic properties of extracellular matrix. In some embodiments, the hydrogel is biodegradable.

[015] In some embodiments, the dynamic supramolecular hydrogel composition embodied herein further comprising cells. In some embodiments, the cells are stem cells. In some embodiments, the cells are immune cells. In some embodiments, the immune cells are T cells, macrophages or microglia.

[016] In some embodiments, the dynamic supramolecular hydrogel composition further comprises a non-covalently bound bioactive peptide, antibody, bioactive protein or drug.

[017] In one aspect, a wound dressing composition is provided comprising the dynamic supramolecular hydrogel composition described herein.

[018] In one aspect, a medical device is provided for in vivo application or implantation comprising the dynamic supramolecular hydrogel composition described herein.

[019] In one aspect, a method is provided for enhancing tissue remodeling, tissue engineering or wound healing, comprising applying to a tissue repair site or wound site the dynamic supramolecular hydrogel composition as described herein. In some embodiments, the dynamic supramolecular hydrogel composition further comprises cells. In some embodiments, the cells are stem cells, immune cells, or the combination thereof. In some embodiments, the immune cells are T cells, macrophages, or microglia. In some embodiments, the cells are obtained from a patient, a donor or a cell line. In some embodiments, the composition is applied to the site together with the cells.

[020] In one aspect, a method is provided for delivering one or more drugs to a bodily site comprising formulating the one or more drugs with the dynamic supramolecular hydrogel composition described herein, and administering the one or more drug-containing composition to a bodily site in a subject. In some embodiments, each of the one or more drugs is independently covalently bound to the composition, is non-covalendy bound to the composition, or the combination thereof. In some embodiments, the covalently bound is a labile covalent bond. In some embodiments, the administering is topical, parenteral, or to a specific bodily site.

[021] In one aspect, a method is provided for evaluating in vitro the activities of cells that interact in vivo with extracellular matrix comprising evaluating the cells in vitro in contact with the dynamic supramolecular hydrogel composition as described herein. In some embodiments, the activities include tissue regeneration, cell migration, cell proliferation, cell differentiation and cell morphogenesis. In some embodiments, the cells are stem cells.

[022] In one aspect, a method for making a dynamic supramolecular hydrogel composition disclosed herein is provided, comprising the steps of: a. preparing a hyaluronic acid covalently bound to adamantane moieties; b. preparing a P-cyclodextrin having a sulfhydryl group; c. preparing gelatin substituted with maleimidyl groups; d. combining the hyaluronic acid covalently bound to adamantane moieties with the P-cyclodextrin having a sulfhydryl group; e. combining the combination of step d with the gelatin substituted with maleimidyl groups; whereby the maleimidyl groups react with the sulfhydryl groups, thereby forming the composition. [023] In some embodiments, a portion of the maleimidyl groups on the gelatin are bound to a RGD containing peptide. In some embodiments, step (e) further comprises including cells. In some embodiments, the degree of substitution of the gelatin substituted with maleimidyl groups is about 0.01 to about 0.8. In some embodiments, the degree of substitution of the gelatin substituted with maleimidyl groups is about 0.17 to about 0.36. In some embodiments, the degree of substitution of the hyaluronic acid covalently conjugated to adamantane moieties is about 0.05 to about 0.5. In some embodiments, the degree of substitution of the hyaluronic acid covalently conjugated to adamantane moieties is about 0.34. In some embodiments, the hyaluronic acid has a molecular weight of about 10 kDa to about 400 kDa. In some embodiments, the hyaluronic acid has a molecular weight of about 60 kDa to about 400 kDa. In some embodiments, the hyaluronic acid has a molecular weight of about 60 kDa to about 80 kDa.

BRIEF DESCRIPTION OF THE FIGURES

[024] Figs. 1A-1B-1C-1D-1E depict the tunable dynamics and rational design of hydrogels. Fig. 1A is a schematic illustration of the tunable dynamics and corresponding cell morphologies in dynamic or non-dynamic hydrogels. Fig. IB shows the rational design of biomacromolecule crosslinking strategy to achieve the tunable dynamics without change of overall stiffness. Examples of hydrogel structures of dynamic and non-dynamic hydrogels are shown. HA is hyaluronic acid. Fig. 1C depicts the different cell morphologies in high dynamic and low dynamic hydrogels. Fig. ID shows the tunable stiffness of a dynamic hydrogel via modulating degree of substitution of the first polymer. The degree of substitution of the second polymer was adjusted based on the degree of substitution of first polymer, to provide a full match of binding moieties. Fig. IE shows the stiffness of several typical human organs and tissues. The stiffness of the dynamic hydrogel can be tuned to match a wide range of human tissues for various uses as described herein.

[025] Fig. 2 depicts SEM images of dynamic and non-dynamic hydrogels.

[026] Figs. 3A-3B-3C-3D show rheological tests of Dynamic and Non-dynamic hydrogels. Fig. 3 A depicts oscillatory frequency sweeps at 1% strain. Fig. 3B depicts oscillatory shear strain sweeps at 1 Hz. G’: Storage Modulus. G”: Loss Modulus. Fig. 3C shows the cyclic deformation of hydrogels by alternative 1% (low, unshaded areas) and 100% (high, shaded areas) shear strain at 1 Hz. Fig. 3D shows stress-relaxation after a fixed shear strain of 10%.

[027] Fig. 4 is a schematic illustration of culture of dental pulp stem cells (DPSCs) in Dynamic or Non-dynamic-hydrogel.

[028] Fig. 5 depicts immunofluorescence staining of alkaline phosphatase (ALP) and osteocalcin (OCN) of dental pulp stem cells (DPSCs) encapsulated in the dynamic supramolecular “Host-Guest”-based hydrogels for 7 days in osteogenic medium and growth medium.

[029] Figs. 6A-6B-6C-6D depict differentiation of dental pulp stem cells (DPSCs). Fig. 6A shows confocal images of Nestin and Sox2 to show neurogenic differentiation of DPSCs encapsulated in dynamic hydrogels. Fig. 6B depicts quantification results of the expression level of Nestin and Sox 2. Fig. 6C shows quantification results of cell area and

A TA shape factor. The shape factor (— ) is a dimensionless metric used to quantify cell morphology and spreading, where A is the cell's projected area and P is its perimeter. A shape factor of 1 indicates a perfectly circular cell, while values less than 1 reflect elongation or irregularity. It is a critical parameter in understanding cell-material interactions, as substrate properties like stiffness, topography, and ligand density significantly influence cell spreading and shape. Higher shape factors indicate isotropic spreading, while lower values are associated with polarized or elongated morphologies. Fig. 6D is a heat map illustration of the expression level of key genes related to neurogenic differentiation.

[030] Figs. 7A-7B-7C-7D-7E depict upregulation of markers by dental pulp stem cells (DPSCs) in dynamic hydrogels. Fig. 7A shows the pathway of neurogenic differentiation. Fig. 7B shows confocal images of Integrin-pi and Integrin-fG demonstrating enhanced integrin clustering in DPSCs encapsulated in dynamic hydrogels. Fig. 7C shows p-FAK staining which indicates induction of FAK activation. Fig. 7D shows quantification results of p-FAK expression from Fig. 7C. Fig. 7E shows mRNA expression level of Integrin-|31, Integrin-|33, YAP and FAK is significantly increased by DPSCs in dynamic vs. nondynamic hydrogels.

DETAILED DESCRIPTION

[031] Unless otherwise defined herein, scientific and technical terms used in connection with the present application shall have the meanings that are commonly understood by those of ordinary skill in the art. Further, unless otherwise required by context, singular terms shall include pluralities and plural terms shall include the singular.

[032] As employed above and throughout the disclosure, the following terms and abbreviations, unless otherwise indicated, shall be understood to have the following meanings.

[033] In the present disclosure, the singular forms “a,” “an,” and “the” include the plural reference, and reference to a particular numerical value includes at least that particular value, unless the context clearly indicates otherwise. Thus, for example, a reference to “a compound” is a reference to one or more of such compounds and equivalents thereof known to those skilled in the art, and so forth. The term “plurality”, as used herein, means more than one. When a range of values is expressed, another embodiment includes from the one particular and/or to the other particular value.

[034] Similarly, when values are expressed as approximations, by use of the antecedent “about,” it is understood that the particular value forms another embodiment. All ranges are inclusive and combinable. In the context of the present disclosure, by “about” a certain amount it is meant that the amount is within ± 20% of the stated amount, or preferably within ± 10% of the stated amount, or more preferably within ± 5% of the stated amount.

[035] Throughout this application, various embodiments of this invention may be presented in a range format. It should be understood that the description in range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the invention. Accordingly, the description of a range should be considered to have specifically disclosed all the possible sub ranges as well as individual numerical values within that range. For example, description of a range such as from 1 to 6 should be considered to have specifically disclosed sub ranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6 etc., as well as individual numbers within that range, for example, 1, 2, 3, 4, 5, and 6. This applies regardless of the breadth of the range.

[036] Whenever a numerical range is indicated herein, it is meant to include any cited numeral (fractional or integral) within the indicated range. The phrases “ranging/ranges between” a first indicate number and a second indicate number and “ranging/ranges from” a first indicate number “to” a second indicate number are used herein interchangeably and are meant to include the first and second indicated numbers and all the fractional and integral numerals there between.

[037] As used herein, the terms “treat”, “treatment”, or “therapy” (as well as different forms thereof) refer to therapeutic treatment, including prophylactic or preventative measures, wherein the object is to prevent or slow down (lessen) an undesired physiological change associated with a disease or condition. Beneficial or desired clinical results include, but are not limited to, alleviation of symptoms, diminishment of the extent of a disease or condition, stabilization of a disease or condition (i.e., where the disease or condition does not worsen), delay or slowing of the progression of a disease or condition, amelioration or palliation of the disease or condition, and remission (whether partial or total) of the disease or condition, whether detectable or undetectable. Those in need of treatment include those already with the disease or condition as well as those prone to having the disease or condition or those in which the disease or condition is to be prevented.

[038] As used herein, the terms “component,” “composition,” “formulation”, “composition of compounds,” “compound,” “drug,” “pharmacologically active agent,” “active agent,” “therapeutic,” “therapy,” “treatment,” or “medicament,” are used interchangeably herein, as context dictates, to refer to a compound or compounds or composition of matter which, when administered to a subject (human or animal) induces a desired pharmacological and/or physiologic effect by local and/or systemic action. A personalized composition or method refers to a product or use of the product in a regimen tailored or individualized to meet specific needs identified or contemplated in the subject.

[039] The terms "subject," "Individual," and "patient" are used interchangeably herein, and refer to an animal, for example a human, to whom treatment with a composition or formulation in accordance with the present invention, is provided. The term “subject” as used herein refers to human and non-human animals. The terms "non-human animal” and “non-human mammal” are used interchangeably herein and include all vertebrates, e.g., mammals, such as non-human primates, (particularly higher primates), sheep, dog, rodent, (e.g., mouse or rat), guinea pig, goat, pig, cat, rabbits, cows, horses and non-mammals such as reptiles, amphibians, chickens, and turkeys. The term “higher vertebrates” is used herein and includes avians (birds) and mammals. The compositions described herein can be used to treat any suitable mammal, including primates, such as monkeys and humans, horses, cows, cats, dogs, rabbits, sheep, goats, pigs, and rodents such as rats and mice. In one embodiment, the mammal to be treated is human. The human can be any human of any age. In an embodiment, the human is an adult. In another embodiment, the human is a child. The human can be male, female, pregnant, middle-aged, adolescent, or elderly. According to any of the methods of the present invention and in one embodiment, the subject is human. In another embodiment, the subject is a non-human primate. In another embodiment, the subject is murine, which in one embodiment is a mouse, and, in another embodiment is a rat. In another embodiment, the subject is canine, feline, bovine, equine, laprine, or porcine. In another embodiment, the subject is mammalian.

[040] Conditions and disorders in a subject for which a particular drug, compound, composition, formulation (or combination thereof) is said herein to be “indicate” are not restricted to conditions and disorders for which that drug or compound or composition or formulation has been expressly approved by a regulatory authority, but also include other conditions and disorders known or reasonably believed by a physician or other health or nutritional practitioner to be amenable to treatment with that drug or compound or composition or formulation or combination thereof. [041] In some embodiments, the localized site of a wound or site for enhancing tissue remodeling or tissue engineering comprises a muscular-skeletal injury, a neurological injury, an eye or ear injury, an internal or external wound, or a localized abscess, an area of mucosa that is affected (e.g., conjunctiva, sinuses, esophagus), or an area of skin that is affected (e.g., infection, autoimmunity), a surgical site or a transplant site. In some embodiments, the transplant or other surgical site includes, for example, but is not limited to, the site and/or its local environment or surroundings of an organ, corneal, skin, limb, face, or other transplant, or a surgical site and/or its local environment or surroundings, for, e.g., but not limited to, treatment of surgical trauma, treatment of a condition related to the transplant or surgery, or prevention of infection. In some embodiments, the site is at or adjacent to a blood clot causing or at risk for causing a myocardial infarction, an ischemic stroke, or a pulmonary embolism. In some embodiments, the methods disclosed herein treat one or more symptoms of a disease, reaction, infection, injury, transplant, surgery, or blood clot.

[042] In some embodiments, the methods disclosed herein treat regeneration of human tissues including, but not limited to spinal cord, peripheral nerves and dental pulp.

[043] As will be described in further detail herein, in one embodiment, supramolecular hydrogel compositions are disclosed that are based on the “Host-Guest” interactions between, but not limited to, [3-cyclodextrin and adamantane (Fig. IB). Such dynamic “Host-Guest” interactions in one embodiment allow the network remodeling of hydrogels in response to the cell traction force during cell spreading. Such supramolecular hydrogel compositions mimic, in one embodiment, the physicochemical properties of extracellular matrix, and thus are useful for various purposes including but not limited to enhancing tissue remodeling, tissue engineering and wound healing. The dynamic properties of the hydrogel are readily adjusted to meet the needs for various uses. As will be described herein, the stiffness and other mechanical properties of the hydrogel can be tuned to the particular organ or tissue for its use (Figs. ID and IE). In contrast to the dynamic hydrogel herein disclosed, a control, covalently-cross-linked hydrogel (also referred to as a non-dynamic hydrogel or static hydrogel) with similar polymeric network topology but with static chemical linkages (e.g., via “thiol-ene” click chemistry) can be compared to the dynamic hydrogels disclosed herein.

[044] As will be shown in the examples herein, dental pulp stem cells (DPSCs) reprogrammed to neuron stem-like cells (iNSCs) were encapsulated in hydrogels and used to study the properties of the dynamic supramolecular hydrogels disclosed herein. The neuron stem cell markers Sox 2 and Nestin were used to track the cells. The encapsulated iNSCs in dynamic hydrogels exhibited significant spreading morphology and more expression of key neuron-related markers, Nestin and Sox 2. The quantification also confirmed the larger cell area, higher spreading shape factor and higher expression level of Nestin and Sox 2 (Figs. 6A, 6B) compared to static hydrogels. In further studies, the enhanced neurogenesis in dynamic hydrogels was attributed to higher expression and clustering of integrin-pi in iNSCs encapsulated dynamic hydrogels, whereas no significant changes were observed for Integrin-P3, indicating that enhanced neurogenesis in dynamic hydrogels can be attributed to the enhanced expression and clustering of integrin-pi in iNSCs encapsulated dynamic hydrogels and the activation of downstream mechanotransduction-related signaling pathways (Fig. 7B).

[045] Successful dental pulp regeneration also requires cell spreading and osteogenic differentiation within local defects. In the Example, DPSCs encapsulated in the dynamic supramolecular “Host-Guest”-based hydrogels for 7 days in osteogenic medium and growth medium showed significant cell spreading and expression of osteogenic markers, such as alkaline phosphatase (ALP) and osteocalcin (OCN) (Fig. 5). Thus, such dynamic supramolecular “Host-Guest”-based hydrogels have several commercial applications, including use in biomedical and biotechnological fields, such as tissue engineering, drug delivery systems, and wound healing. They can also be used in industrial applications, such as water treatment, oil and gas recovery, and as thickeners and stabilizers in food and cosmetics. Dynamic supramolecular “Host-Guest’’-based hydrogels have several competitive advantages over traditional hydrogels. They can change their properties in response to changes in the environment, such as temperature, pH, and ionic strength. This allows for greater control over the delivery of drugs or other materials. They also have improved mechanical properties, such as increased strength and toughness, which makes them more suitable for use in biomedical and industrial applications. Additionally, dynamic hydrogels are often biocompatible and biodegradable, which makes them a more sustainable option for many applications.

[046] The present disclosure is generally directed to a dynamic supramolecular hydrogel composition comprising:

(b) a second polymer comprising second polymer subunits, wherein the second polymer further comprises covalently -bound hydrophobic moieties; wherein the first polymer and the second polymer are noncovalently bound by the cyclodextrin moieties non-covalently bound to the hydrophobic moieties.

[047] As described herein, the stiffness and other properties of the hydrogel can be tuned for a particular use, e.g., a particular organ or tissue, by selecting the degree of substitution of the cross-linking moieties. Each of the components therein, and non-limiting methods for preparing the hydrogel composition, are described herein, the disclosure not intended to be limiting in any way as one of skill in the art, guided by the principles described herein, may readily make variations with the same properties and useful for the same purposes.

[048] First polymer. The first polymer, to which cyclodextrin moieties are covalently bound, may comprise a peptide, a polypeptide, a protein, a polysaccharide, a nucleic acid, or repeating subunits of one or more monomers such as lactic acid, glycolic acid polymers or co-polymers, and the like. In one embodiment, the first polymer is a protein such as gelatin. In one embodiment the first polymer is a polysaccharide such as hyaluronic acid.

[049] In one embodiment, the first polymer is biodegradable.

[050] The first polymer is comprised of first polymer subunits, which may be amino acids, sugar monomers, dimers or oligosaccharides, nucleotides, an alpha-hydroxyacid, etc. [051] In some embodiments, the first polymer is hyaluronic acid. In some embodiments, the hyaluronic acid has a molecular weight of about 10 kDa to about 400 kDa. In some embodiments, the hyaluronic acid has a molecular weight of about 60 kDa to about 400 kDa. In some embodiments, the hyaluronic acid has a molecular weight of between about 60 kDa and about 80 kDa.

[052] In one embodiment, the degree of substitution (DS) of the first polymer with cyclodextrin moieties is between about 0.01 and 0.8, to provide a fully tunable system in which the stiffness of the dynamic hydrogel can be selected. Degree of substitution of the first polymer is based on the number of sites that can be bound to the cyclodextrin; for example, if the first polymer is gelatin and the cyclodextrins are bound via carboxylic acid moieties on the gelatin, the DS is the number of bound cyclodextrins to the total number of carboxylic acid moieties. The ratio of first polymer subunits comprising non-covalently bound cyclodextrin moieties to the total first polymer subunits is about 0.01 to about 0.8. In some embodiments, the first polymer is gelatin. Thus, in some embodiments, the modification of each cyclodextrin moiety will occupy one carboxyl functional group on the gelatin backbone. The amount of carboxyl functional group on the gelatin backbone will be quantified as x mol of -COOH/g of gelatin, which is varied based on different batches of gelatin from the supplier. For example, the degree of substitution of cyclodextrin of 0.8 denotes that 80% of carboxyl functional groups on gelatin backbone were modified with cyclodextrin. As noted herein, the DS of the first polymer is selected to create a hydrogel to match the stiffness of a particular organ or tissue (for example, Fig. IE) in or on which the hydrogel is used for the purposes herein. The relationship between DS of the first polymer and stiffness of the hydrogel is shown in Fig. ID; as described below, the DS of the second polymer is provided to match the cyclodextrins and hydrophobic moieties.

[053] As will be noted herein for both the first polymer and for the second polymer, each polymer comprises groups to which one or more further molecules may be covalently bound; in the case of the first polymer, a cyclodextrin; in the case of the second polymer, a hydrophobic moiety; and in the case of either or both polymers, independently an optional bound bioactive peptide, antibody or bioactive protein that is selected from or comprises nerve growth factor, RGD or a peptide comprising RGD, or a fragment of any of the foregoing, or a mimetic of any of the foregoing. Each of these bound or optionally bound molecules are described further below.

[054] In some embodiments, the first or second polymer subunits comprise a pendant reactive moiety to which the respective additional molecule can be bound, such as in the non-limiting example of hyaluronic acid, comprises carboxylic acid moieties to covalently bind, directly or through an intermediate spacer or linker molecule, the cyclodextrin or hydrophobic moiety. In one embodiment, hyaluronic acid is derivatized to comprise maleimide groups that can then react and covalently bind a thiol group on a thiol-modified cyclodextrin. In another embodiment, a hydrophobic group with a reactive moiety such as adamantanecarboxylic acid can be covalently coupled to hyaluronic acid carboxylic acid moieties by an one of a number of reactions, examples of which are provided herein. In another embodiment, a protein e.g., gelatin may have its carboxylic acid side chains used to bind to cyclodextrin.

[055] Second polymer. The second polymer, to which is covalently bound a hydrophobic moiety that non-covalently binds to the cyclodextrin on the first polymer, may be the same or different polymer than the first polymer, including but not limited to any of those described for the first polymer. The second polymer is comprised of second polymer subunits. In some embodiments, wherein the first and second polymers are the same (e.g., both hyaluronic acid), the lengths and/or molecular weights may be different. As will be noted below, fine tuning of the properties of the supramolecular hydrogel composition of the disclosure comprises adjusting the first and second polymers’ compositions, lengths, molecular weights, and degrees of substitution of the respective components.

[056] In one embodiment, the second polymer is biodegradable.

[057] In some embodiments, the second polymer is hyaluronic acid, as described above. In some embodiments, the second polymer is gelatin. [058] In some embodiments, the first polymer is the same as the second polymer. In some embodiments, the first polymer subunits are the same as the second polymer subunits.

[059] In one embodiment, the degree of substitution (DS) of the second polymer is provided to match the DS of the first polymer (which may be selected to provide a certain stiffness or other property for use with a particular organ or tissue) to provide a match between the cyclodextrins on the first polymer and hydrophobic groups on the second polymer. In one embodiment, the degree of substitution of the second polymer with hydrophobic moieties is between about 0.01 and 0.8. The degree of substitution of the second polymer is based on the number of bound hydrophobic moieties (e.g., adamantane) to the number of second polymer subunits (e.g., hyaluronic acid monomers, a disaccharide of D-glucuronic acid and N-acetyl-D-glucosamine).

[060] Thus, the ratio of second polymer subunits comprising non-covalently bound hydrophobic moieties to the total second polymer subunits is about 0.01 to about 0.8. In one embodiment, the second polymer is hyaluronic acid. As noted herein, the DS of the second polymer is selected to match the DS of the first polymer which is selected to provide a dynamic hydrogel of stiffness for the intended use, e.g., matching the stiffness of an organ or tissue. The stiffness of the dynamic hydrogel thus can be tuned to match a wide range of organs and tissues, among other uses.

[061] As noted above, the optional additional molecules on the second polymer and other features optionally shared by the first and second polymer are described above. It should be noted that any modification or derivatization described herein for the first or second polymer is equally applicable to the second or the first polymer.

[062] Cyclodextrin. The first polymer, in one embodiment, comprises cyclodextrin covalently bound thereto. The cyclodextrin may be any cyclodextrin or cyclodextrin derivative capable of binding to a hydrophobic moiety, such as but not limited to an adamantane moiety. The cyclodextrin may be a-cyclodextrin, |3-cyclodextrin, or y- cyclodextrin, or any combination thereof. After covalent binding to the first polymer, the bound cyclodextrin retains its ability to bind a hydrophobic moiety. [063] A cyclodextrin is covalently bound to the first polymer. Such covalent binding is provided by any means that allows the covalently-bound cyclodextrin to bind non- covalently to a hydrophobic moiety such as an adamantane moiety. In one embodiment, the carboxyl group on a cyclodextrin is used to covalently bind the cyclodextrin to the polymer. In one embodiment, a sulfhydryl group added to a cyclodextrin is used to covalently bind the cyclodextrin to the polymer. Non-limiting examples of cyclodextrins that are derivatized to allow for covalent binding to a reactive group on the first polymer include, but are not limited to, a thiol-modified cyclodextrin, an amino-modified cyclodextrin such as aminated a-cyclodextrin, and an carboxymethyl-modified cyclodextrin such as carboxymethylated y-cyclodextrin.

[064] First Polymer Bound to Cyclodextrin. As described above, in one embodiment, the first polymer is gelatin. The cyclodextrin may be covalently bound to gelatin by any of several methods, in one example, carboxyl groups on the gelatin are modified with maleimide groups by coupling N-2(2-aminoethyl)maleimide using 3- (ethyliminomethyleneamino)-N,N-dimethylpropan-l -amine (EDC) and hydroxybenzotriazole (HOBt) as known in the art. In one embodiment, the degree of substitution of maleimide groups is about 0.17. Subsequently, the maleimide-modified gelatin is reacted with thiol-modified cyclodextrin as described herein. In other embodiments, to provide a higher degree of substitution of the first polymer with maleimide groups, the coupling can be performed in an organic solvent such as DMSO, and benzotriazol- 1-yloxy tris(dimethylamino)phosphonium hexafluorophosphate (BOP) to couple N-2(2-aminoethyl)maleimide to the gelatin. In one embodiment, such a degree of substitution (DS) of the maleimide groups is about 0.36. As noted herein, the DS of the first polymer may be selected to provide a desired stiffness among other properties for a particular use, such as matching the stiffness of an organ or tissue in or on which the hydrogel will be used. Thus, the stiffness of the dynamic hydrogel may be tuned following the guidance herein.

[065] In one embodiment, the cyclodextrin covalently bound to the first polymer is shown below, wherein a portion of the gelatin amino acid sequence is depicted as Formula (II), a maleimide is bound to a carboxyl side chain of a gelatin amino acid, and a cyclodextrin (the truncated cone symbol) is covalently bound to the maleimide. In one embodiment, the cyclodextrin is P-cyclodextrin.

[066] Hydrophobic moieties. The hydrophobic moiety refers to any chemical moiety capable of binding to a cyclodextrin, when the chemical moiety is covalently bound to the second polymer. Non-limiting examples of hydrophobic moieties useful for the purposes disclosed herein include adamantane, cholesterol, phenyl groups, benzyl groups, aliphatic hydrocarbons such as octadecane, aromatic hydrocarbons such as naphthalene, steroids such as testosterone, fatty acid chains such as stearic acid, and hydrophobic drugs such as ibuprofen. In one embodiment a hydrophobic moiety with a reactive groups such as adamantanecarboxylic can be covalently coupled to a second polymer such as hyaluronic acid. Methods for coupling a hydrophobic moiety to a polymer are well known in the art. In such embodiments, the cyclodextrin-binding ability of the hydrophobic moiety is retained after covalently binding to the second polymer.

[067] In one embodiment, the degree of substitution (DS) of the second polymer with hydrophobic moieties is between about 0.01 and 0.8. Thus, the ratio of second polymer subunits comprising non-covalently bound hydrophobic moieties to the total second polymer subunits is about 0.01 to about 0.8. In one embodiment, the second polymer is hyaluronic acid. In one embodiment the hydrophobic moiety is adamantane. As noted herein, the DS of the second polymer may be provided to match that of the first polymer, which DS is selected to provide a stiffness and other properties for an intended use such as matching the stiffness of an organ or tissue it will be use in or on.

[068] Second Polymer Bound to Hydrophobic Moieties. As described above, in one embodiment, the second polymer is hyaluronic acid, and is covalently bound to a hydrophobic moiety such as adamantane. In one embodiment, 1 -adamantanecarboxylic acid is conjugated to hydroxyl groups on the hyaluronic acid using, by way of non-limiting example, the methods described in the examples herein. In one embodiment, a portion of the hyaluronic acid polymer with covalently bound adamantane groups is shown in Formula (I) below. In this example, the degree of substitution of the hyaluronic acid (the value of y/(x + y)) is between about 0. 01 and about 0.8.

(I)-

[069] Additional Covalently-Bound Molecules. Either the first polymer, the second polymer, or both, may further comprise one or more of the same or different other molecules covalently bound thereto. In one embodiment, such additional molecules are bound to the respective polymer independently of the binding of the cyclodextrin or the hydrophobic moiety; in other embodiments, the additional molecules may be bound similarly thereto, or use the same coupling chemistry as used for the cyclodextrin and/or hydrophobic moiety, and/or substitute in part for the cyclodextrin and/or hydrophobic moiety during the conjugation process. By way of non-limiting example, a RGD- containing peptide with a thiol group (e.g., a cysteine) may be covalently bound to a maleimide-conjugated gelatin along with the thiol-modified cyclodextrin, such that both cyclodextrin and RGD peptide are covalently bound to the first polymer, a ratio selected to provide desirable features to the supramolecular hydrogel composition. Similarly, the second polymer may comprise hydrophobic moieties and another molecule at a ratio to be determined to provide desirable properties.

[070] Non-limiting examples of such additional molecules include one or more bioactive peptides, antibodies, or bioactive proteins. In some embodiments, the bioactive peptide is a neuron growth and differentiation-inductive peptide. In some embodiments, the bioactive peptide, antibody or bioactive protein may be, or may comprise, nerve growth factor, an integrin binding peptide, RGD or a peptide comprising RGD, or a fragment of any of the foregoing, or a mimetic of any of the foregoing. In one embodiment, the bioactive peptide comprises RGD, such as GCGYGRGDSPG (SEQ ID NO:1). In some embodiments, a bioactive peptide has an extension of amino acids such as GCG or GGCGGC added to facilitate cross-linking, such as using a cysteine sulfhydryl group, Other examples of RGD containing peptides or other integrin binding peptides that may be incorporated the first or second polymer as described herein include YGRGDSPG (SEQ ID NO:2), NGEPRGDTYRAY (SEQ ID NOG), KGGPQVTRGDVFTMP (SEQ ID NO:4), RSTDLPGLKAATHYTITIRGV (SEQ ID NOG), VFDNFVLK (SEQ ID NOG), ESQEEVVSESRGDNPDPTTSY (SEQ ID NO:7), TVDVPDGRGDSLAYG (SEQ ID NOG), SVVYGLR (SEQ ID NO:9), GCGNGEPRGDTYRAY (SEQ ID NO: 10), GCGKGGPQVTRGDVFTMP (SEQ ID NO: 11),

GCGRSTDLPGLKAATHYTITIRGV (SEQ ID NO: 12), GCGVFDNFVLK (SEQ ID NO:13), GCGESQEEVVSESRGDNPDPTTSY (SEQ ID NO:14), GCGTVDVPDGRGDSLAYG (SEQ ID NO: 15), GCGSVVYGLR (SEQ ID NO: 16), GGCGGCYGRGDSPG (SEQ ID NO: 17), GGCGGCNGEPRGDTYRAY (SEQ ID NO: 18), GGCGGCKGGPQVTRGDVFTMP (SEQ ID NO: 19),

GGCGGCRSTDLPGLKAATHYTITIRGV (SEQ ID NO:20), GGCGGCVFDNFVLK (SEQ ID NO:21), GGCGGCESQEEVVSESRGDNPDPTTSY (SEQ ID NO:22), GGCGGCTVDVPDGRGDSLAYG (SEQ ID NO:23), or GGCGGCSVVYGLR (SEQ ID NO:24), GCGYGRSTDLPGLKAATHYTITIRGV (SEQ ID NO:25), GCGYGGGGNGEPRGDTYRAY (SEQ ID NO:26), and GCGYGTVDVPDGRGDSLAYG (SEQ ID NO:27). As noted above, in one embodiment, bioactive peptide, antibody or bioactive protein comprises a thiol group or is modified to comprise a thiol group, and is bound to maleimide groups on the first polymer. In another embodiment, the conjugation chemistry used to bind the hydrophobic moiety to the second polymer may also be used to bind additional molecule.

[071 ] The dynamic supramolecular hydrogel may comprise one or more bioactive peptides on either or both the first polymer and/or the second polymer. In one embodiment, both the first polymer and the second polymer comprise the same one or more bioactive peptides. In one embodiment, the first polymer comprises one or more bioactive peptides and the second polymer comprises a different one or more bioactive peptides.

[072] Additional Non-Covalently-Bound Molecules. In another embodiment, additional molecules may be included in the supramolecular hydrogel composition that are not covalently bound to either the first polymer or the second polymer or any pendant moieties therein. In one embodiment, the first polymer comprising cyclodextrin moieties may comprise hydrophobic molecules non-covalently bound to the cyclodextrin. Non-limiting examples include cholesterol, molecules with phenyl groups, molecules with benzyl groups, aliphatic hydrocarbons such as octadecane or molecules comprising them, aromatic hydrocarbons such as naphthalene or molecules comprising them, steroids such as testosterone, fatty acids such as stearic acid or molecules comprising them, and hydrophobic drugs. Such non-covalently bound hydrophobic molecules include, by way of non-limiting examples, an NSAID such as ibuprofen, a chemotherapy drug such as paclitaxel, a hydrophobic polyphenol such as curcumin, a vitamin such as vitamin D3; an immunosuppressive drug such as cyclosporine, a lipophilic antibiotic such as rifampicin and erythromycin; an antifungal drug such as ketoconazole and itraconazole; an antiviral drug such as saquinavir and ritonavir; a lipid-based hormone such as progesterone; and other hydrophobic compounds such as coenzyme Q10 and lutein. Additionally, lipid- soluble vitamins like vitamin A and vitamin E, as well as certain essential oils with therapeutic properties, can also be effectively encapsulated in cyclodextrin complexes. In some embodiments, the design of such a supramolecular hydrogel composition with regard to number of cyclodextrins on the first polymer available to non-covalently bind to hydrophobic moieties on the second polymer, and the number of cyclodextrins binding to an additional molecule, will be guided by the desirable properties of the supramolecular hydrogel composition.

[073] Additional Components. The supramolecular hydrogel compositions disclosed herein may be prepared to include other molecules (e.g., drugs), biomolecules, cells, or other components that provide certain features to the compositions for certain purposes. By way of non-limiting examples, any of the aforementioned bioactive peptides, drugs, antibodies, or bioactive proteins, and non-limiting examples thereof, may be admixed during the preparation of the composition to be released at a site of use for example to slowly elute from the site, providing particular properties. Cells of one type or any combination may be loaded into the hydrogel composition before, for example, depositing at a tissue repair site, wound or other location in a body, to enhance the property of the composition. By way of non-limiting examples, various types of stem cells such as mesenchymal stem cells, neural stem cells, induced pluripotent stem cells (iPSCs), and embryonic stem cells can be loaded for purposes such as but not limited to neuron regeneration. Additionally, immune cells such as T cells, macrophages, or microglia can also be incorporated. These cells may be autologous, derived from the subject or patient into whom the hydrogel will be used, or they may be allogeneic, coming from a donor or an established cell line.

[074] In the context of neuron regeneration, neural progenitor cells and oligodendrocyte progenitor cells are particularly relevant to the present disclosure, as they can differentiate into various types of neural cells, potentially aiding in the repair and regeneration of neural tissue. Furthermore, Schwann cells and astrocytes can also be included to support neuronal growth and myelination. The combination of these cell types within a hydrogel matrix can create a conducive environment for nerve regeneration, potentially accelerating the healing process in neural injuries or degenerative diseases. In some embodiments, the hydrogel may also be modified to release growth factors or other bioactive molecules to further support cell growth and tissue regeneration. This approach offers a versatile and potent strategy for regenerative medicine, particularly in the field of neurology.

[075] In another embodiment, as noted herein, a drug, such as a hydrophobic drag, may be non-covalently bound the cyclodextrin of the composition, such that the drag will be released from the composition and delivered to a selected bodily site where the composition is implanted, or delivered to distal sited therefrom. In another embodiment, a drug admixed in the composition may similarly be released from the composition and delivered to a selected bodily site where the composition is implanted, or delivered to distal sited therefrom. Any of such aforementioned drags, molecules, compounds, bioactive peptides, bioactive proteins, antibodies, described as being covalently bound to one or both polymers herein, or bound to the cyclodextrin, may independently be present in the hydrogel, as an admixture to or loaded into the polymers or hydrogel during or after preparation. In some embodiments, the same drag, molecule, compound, peptide, protein or antibody that is covalently bound to the hydrogel or bound to the cyclodextrin may also be admixed with the hydrogel, for example, to provide an early or bolus release when implanted, followed by a slower release by the cyclodextrin- or polymer-bound same aforementioned drug etc.

[076] Any of the foregoing embodiments are not necessarily separate, and any combination thereof may be applied to the use of the compositions herein. By way of non-limiting example, a composition may comprise a hydrophobic drug bound to some cyclodextrins, a population of stem cells, and a bioactive protein covalently bound to one or both polymers, providing an implantable extracellular matrix that delivers the incorporated drug as well as exhibits properties of extracellular matrix, including any of the other uses herein described.

[077] Other Features. In some embodiments, both the first polymer and the second polymer are biodegradable. As noted herein, in some embodiments, the first polymer and the second polymer may be comprised of the same polymer subunits (e.g., amino acids, monosaccharides, disaccharides) but in some embodiments, the first and second polymer lengths, degrees of substitution by the respective moieties, and other properties may be different. [078] Properties of the Supramolecular Hydrogel Compositions. The foregoing covalent and optional non-covalent modifications of the first polymer and second polymer provide a composition with properties that, in some embodiments, mimic the properties of extracellular matrix, and in some embodiments, providing a composition with utility in enhancing biological processes that utilize extracellular matrix. As noted herein (Figs. 3A- D), the compositions, in one embodiment mimic extracellular matrix, and are dynamic in that they can form and reform depending on the physical forces applied thereto. They can change their properties in response to changes in the environment, such as but not limited to shear force, temperature, pH, ionic strength, pressure, etc. In some embodiments, the properties of the supramolecular hydrogel compositions disclosed herein may be tuned for particular uses or applications, where such properties enhance certain biological processes such as but not limited to enhancing tissue remodeling, tissue engineering or wound healing. As noted above, the degree of substitution of the cyclodextrin and/or hydrophobic moieties on the respective first and second polymer, the selection of hydrophobic moiety, the selection of additional molecules covalently bound to one or both polymers, and any molecules non-covalently bound thereto, may be modified to provide supramolecular hydrogel compositions with particular properties suitable for the desired uses.

[079] As described herein, in one embodiment, the ratio of polymer subunits comprising covalently-bound hydrophobic moieties to the total polymer subunits provides for the dynamic properties, and can be tuned or adjusted, dependent on the dynamic properties desired. In one embodiment, the ratio of polymer subunits comprising covalently-bound cyclodextrin to the total polymer subunits provides for the dynamic properties, and can be adjusted dependent on the dynamic properties desired. In one embodiment, the ratio of polymer subunits comprising covalently -bound cyclodextrin to the total polymer subunits of the first polymer, and the ratio of polymer subunits comprising covalently-bound hydrophobic to the total polymer subunits of the second polymer, provides for the dynamic properties, and can be adjusted dependent on the dynamic property or properties desired. As noted herein, the ratio, also called degree of substitution (DS), of the first polymer may be selected to match the stiffness of a tissue or organ, and the DS of the second polymer adjusted such that the available cyclodextrins match the bound hydrophobic moieties. For example, the matching of functional groups of the first polymer and the second polymer is based on the amounts of functional groups by weight of each polymer. For example, if the DS of HA- Ada is 0.30, the number of functional groups per gram of hyaluronic acid is calculated (x mol of Ada per gram of HA). Then, the DS needed for the first polymer is calculated to reach the same molar number of Ada and cyclodextrin. Thus, the DS of the first polymer and that of the second polymer are independent, and as noted herein, the DS of the first polymer may be selected to provided a stiffness of for the intended use.

[080] As noted herein, for the first polymer, the ratio of cyclodextrin to polymer subunits may range from about 0.01 to about 0.8. Non-limiting ratios include about 0.01, 0.05, 0.1, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45, 0.5, 0.55, 0.6, 0.65, 0.7, 0.75 and 0.8. As noted herein, for the second polymer, the ratio of hydrophobic moieties to polymer subunits may range from about 0.01 to about 0.8. Non-limiting ratios include about 0.01, 0.05, 0.1, 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45, 0.5, 0.55, 0.6, 0.65, 0.7, 0.75 and 0.8. Ratios of up to 1 and approaching 0 are also embodied herein.

[081] In one embodiment, the supramolecular hydrogel composition is capable of remodeling in response to the traction force during cell spreading.

[082] In one embodiment, the supramolecular hydrogel composition, the dynamic property is stiffness, strength, toughness, self-healing, shear-thinning, fast stress relaxation, and easy matrix remodeling.

[083] As shown in Fig. ID, a degree of substitution of the first polymer may be selected to match a particular organ or tissue, and the second polymer’s DS can be selected to provide a match of the cyclodextrins to the hydrophobic moieties.

[084] Uses. The following are non-limiting examples of uses of the compositions disclosed herein, in addition to those mentioned elsewhere herein. In some embodiments, the compositions disclosed herein may be used for enhancing tissue remodeling, tissue engineering or wound healing, by applying the composition to a tissue repair site or wound site. In some embodiments, the composition comprises cells. Cells of one type, or any combination of cell types, may be loaded into the hydrogel composition before, in one nonlimiting example, depositing at a tissue repair site, wound or other location in a body, to enhance the property of the composition and its desired effects. By way of non-limiting examples, stem cells, immune cells (e.g., T cells, macrophages or microglia) or multiple types of cells, or any other type of cell, may be loaded therein. Such cells may be obtained from the subject or patient on whom the hydrogel will be used; in other embodiments, a donor or cell line may be the source or one of more cell types. In some embodiments, the cells comprise the subject’s cells in combination with one or more of the same or other cell type from a donor, cell line, or any other source. In some embodiments, the cells may be non-human cells. In one embodiment, the cells are induced neuron stem-like cells (iNSCs). In other embodiments, the cells are endothelial cells for vascular regeneration, cardiomyocytes for heart tissue repair, hepatocytes for liver regeneration, or pancreatic beta cells for diabetes therapy. Additionally, fibroblasts, keratinocytes, or melanocytes may be used for skin repair and regeneration. The versatility of the hydrogel composition allows for the inclusion of various types of cells, tailored to specific therapeutic goals and sitespecific regeneration, making it an adaptable tool for a wide range of regenerative medicine applications.

[085] The composition may be used in or applied to any location where the properties of extracellular matrix are desired, such properties including but not limited to enhancing cell regeneration, migration, proliferation, differentiation and morphogenesis. In another embodiment, the compositions disclosed herein provide expansion and/or differentiation of encapsulated stem cells and/or recruitment of host immune cells. In some embodiments, the compositions disclosed herein comprising encapsulated stem cells provide better expansion and/or differentiation of the stem cells that occurs with the encapsulated stem cells alone.

[086] Hydrogels disclosed herein may be administered or applied by any one of a number of methods at any location in the body, such as topically, parenterally, intradentally, into a joint, into a wound, into an organ such as the brain, liver or kidney, into the spinal column or spinal cord. Non-limiting examples include parenterally, paracancerally, transmucosally, transdermally, intramuscularly, intravenously, intradermally, subcutaneously, intraperitoneally, intraventricularly, intravaginally, intracranially and intratumorally.

[087] Dental Uses. In one embodiment, the compositions disclosed herein provide for enhanced dental pulp regeneration, where the hydrogel can be loaded with dental pulp stem cells or odontoblast-like cells to promote the repair and regeneration of damaged tooth structures.

[088] Bone Uses. The hydrogels can also be adapted for bone tissue engineering, where osteoblasts, mesenchymal stem cells, or bone marrow-derived cells can be incorporated to facilitate bone growth and healing. This application is particularly relevant in the context of fractures, bone defects, and osteoporosis treatment.

[089] Cardiac Uses. For cardiac tissue repair, especially after myocardial infarction, the hydrogels can be loaded with cardiomyocytes, endothelial cells, or cardiac stem cells to aid in the regeneration of heart muscle and the restoration of its function.

[090] Neurological Uses. In the field of neurology, hydrogels containing neural stem cells or induced neuron stem-like cells (iNSCs) can be used for the regeneration of neural tissue in diseases, not only in neurodegenerative diseases such as Parkinson's and Alzheimer's but also in injuries to the central nervous system (CNS) and peripheral nervous system (PNS).

[091] Skin and Wound Healing Uses. For skin regeneration and wound healing, the hydrogels can be embedded with fibroblasts, keratinocytes, or melanocytes to enhance the healing of bums, ulcers, and other skin injuries.

[092] Liver and Pancreatic Uses. In liver regeneration, hepatocytes can be used, whereas for diabetes treatment, pancreatic beta cells can be incorporated to restore insulin production and regulation. [093] Methods for Manufacture. The descriptions and examples herein provide nonlimiting examples of methods for making the supramolecular hydrogel compositions disclosed herein, but such descriptions are merely exemplary, and one of skill in the art following the teachings herein can readily prepare compositions disclosed herein for any of various purposes also described herein, among others.

[094] In one embodiment, the method for making a dynamic supramolecular hydrogel composition disclosed herein comprises the steps of: preparing a second polymer covalently bound to hydrophobic moieties; preparing a cyclodextrin having a reactive group; preparing first polymer substituted with groups that react with the reactive group; combining the second polymer covalently bound to hydrophobic moieties with the cyclodextrin having a reactive group; combining the combination of the foregoing step with a first polymer substituted with groups that react with the reactive groups on the cyclodextrin, thereby forming the composition.

[095] In some embodiments, an additional one or more molecules may be covalently bound to the first polymer, the second polymer, or both. In some embodiments, the same coupling methods for the cyclodextrin and/or hydrophobic moiety is used to couple the additional one or more molecules. In some embodiments, the additional one or more molecules are coupled by other methods. In some embodiments, the composition comprises non- covalently bound molecules, such as a drug non-covalently bound to cyclodextrins, or cells, peptides, proteins or other molecules entrapped or included in the hydrogel as it is being prepared.

[096] In one embodiment, the first polymer is gelatin, the second polymer is hyaluronic acid, the hydrophobic moieties are adamantane moieties, the cyclodextrin is |3-cyclodextrin, the reactive group on the P-cyclodextrin is a thiol group, and the first polymer is substituted with maleimide groups. To prepare such a hydrogel, the following steps are performed: preparing a hyaluronic acid covalently bound to adamantane moieties; preparing a P-cyclodextrin having a sulfhydryl group; preparing gelatin substituted with maleimidyl groups; combining the hyaluronic acid covalently bound to adamantane moieties with the P-cyclodextrin having a sulfhydryl group; combining the combination of step d with gelatin substituted with maleimidyl groups; whereby the maleimidyl groups react with the sulfhydryl groups, thereby forming the composition.

[097] In some embodiments, the hyaluronic acid has a molecular weight of about 10 kDa to about 400 kDa. In some embodiments, the hyaluronic acid has a molecular weight of about 60 kDa to about 400 kDa. In some embodiments, the hyaluronic acid has a molecular weight of about 60 kDa to about 80 kDa.

[098] In some embodiments, degree of substitution of the gelatin substituted with maleimidyl groups is about 0.01 to about 0.8. In some embodiments, the degree of substitution of the gelatin substituted with maleimidyl groups is about 0.17 to about 0.36. In some embodiments, the degree of substitution of the hyaluronic acid covalently conjugated to adamantane moieties is about 0.05 to about 0.5. In some embodiments, the degree of substitution of the hyaluronic acid covalently conjugated to adamantane moieties is about 0.34.

[099] In some embodiments, the foregoing composition comprises non-covalently bound molecules, such as a drug non-covalently bound to a portion of the cyclodextrins, or cells, peptides, proteins or other molecules entrapped or included in the hydrogel as it is being prepared. [100] In some embodiments, some of the reactive groups on the gelatin are bound to an additional molecule, such as a RGD containing peptide, such that in one embodiment, some maleimide groups on the gelatin bind to the thiol-modified cyclodextrin and others to a RGD peptide comprising a thiol group.

EXAMPLES

Example 1. Synthesis of Hyaluronic Acid - Adamantane (HA-Ada or HA- Adamantane)

[101] Dowex-50 resin was purified with a 2 M hydrochloric acid solution and subsequently thoroughly washed with deionized (DI) water until a neutral pH was achieved. Sodium hyaluronate (7.5 g, 18.75 mmol) was dissolved in DI water to a concentration of 2% and then subjected to ion exchange with Dowex-50 resin (22.5 g) in water, with gentle stirring overnight. Following filtration, the hyaluronic acid solution was neutralized to pH 7.0 using a 0.2 M tetrabutylammonium (TBA) hydroxide solution. After lyophilization, HA-TBA was produced.

[102] In an N2 atmosphere, vacuum-dried HA-TBA (1 g, 1.4 mmol), 1- adamantanecarboxylic acid, and 4-dimethylaminopyridine were combined in a 100 mL flask. Anhydrous DMSO (50 mL) was introduced to create a 2% HA-TBA solution. After complete dissolution, a varying quantity of di-tert-butyl dicarbonate (BOC2O) was injected using a syringe, and the reaction was conducted at 45 °C for 24 hours. Post cooling, the mixture was dialyzed (MWCO 7000) against DMSO for three days, followed by a gradual replacement of the dialysis medium with DI water, and further dialysis continued at room temperature for three more days. Following filtration, the solution was freeze-dried to yield the final product.

[103] By using 1H-NMR in D2O, the degree of substitution (DS) of the adamantane groups was determined by integrating the ethyl multiples of adamantane (6=1 .42-1 .70 ppm, 12H) in relation to the hyaluronic acid backbone (8=3.10-4.10 ppm, 10H). The DS was found to be 0.34. Example 2. Synthesis of Gelatin-Maleimide (Gel-Mal)

[104] A low degree of substitution Gel-Mal was created by attaching N-(2-aminoethyl) maleimide to the carboxyl groups of gelatin using the EDC/HOBt coupling method. To summarize the process, gelatin (1 g) was first dissolved in deionized water (100 mL). N- (2-aminoethyl) maleimide trifluoroacetate salt (317.5 mg, 1.25 mmol) was then added. Following this, hydroxybenzotriazole (HOBt; 337.5 mg, 2.5 mmol), which had been predissolved in 10 mL of a mixed solvent (water: acetonitrile = 1:1), was incorporated into the solution. The solution's initial pH was about 3.8 and was subsequently adjusted to 6.0 with a dilute sodium hydroxide solution.

[105] 3-(Ethyliminomethyleneamino)-N,N-dimethylpropan-l -amine (EDC) (479.5 mg, 2.5 mmol) was then introduced to the mixture, and the gradual pH increase signified the reaction. The mixture was stirred overnight and subsequently dialyzed against a 0.3 M sodium chloride solution in a dialysis bag (MWCO 7000 Dalton). The dialysis medium was refreshed several times a day and progressively changed to a 0.1 M sodium chloride solution over a period of 2 days. Following this, the mixture was dialyzed against distilled water for an extra two days. Throughout the entire dialysis process, the medium was cooled with ice to prevent maleimide hydrolysis. The final product was obtained through lyophilization, resulting in a yield of 90%.

[106] The degree of substitution (DS) of the maleimide groups was found to be 0.17 by using H-NMR in D2O.

[107] A high DS Gel-Mal was produced using an organic solvent by combining N-(2- aminoethyl) maleimide with the carboxyl groups of gelatin using the benzotriazol- 1-yloxy tris(dimethylantino)phosphonium hexafluorophosphate (BOP) coupling process. To improve its solubility in organic solvents, gelatin was converted into tetrabutylammonium salts (Gel-TBA).

[108] In anhydrous DMSO (60 mL), dried Gel-TBA (1.114 g), N-(2-aminoethyl) maleimide trifluoroacetate salt (0.408 g; 1.61 mmol), and benzotriazol- 1-yloxy) tris(dimethylamino)phosphonium hexafluorophosphate (BOP, 0.712 g, 1.61 mmol) were dissolved and stirred at room temperature to initiate the coupling reaction. After two hours, the mixture was cooled, and deionized (DI) water (50 mL) was introduced.

[109] For two days, the solution was dialyzed (MWCO 14000) against 1.6 L of a half-and- half mixture of water and DMSO, which was cooled with ice. The co-solvent medium was replaced three times a day. Following this, the mixture underwent further dialysis against an ice-cooled sodium chloride solution (0.3-0.1 M) and then DI water for 3-5 days. Throughout the entire dialysis process, ice was used to cool the medium to prevent the breakdown of the maleimide. The solution was then filtered through a 0.22 pm membrane and freeze-dried to get the final product.

[110] The degree of substitution (DS) of the maleimide groups was determined to be 0.36 by using 'H-NMR in D2O.

Example 3. Synthesis of Hyaluronic Acid-SH (HA-SH)

[111] HA-SH was synthesized by coupling 3,3'-dithiodipropionic acid dihydrazide to the carboxyl groups of hyaluronic acid (HA) via the EDC/HOBt coupling method. To summarize the procedure, hyaluronic acid (3 g, 7.5 mmol) was first dissolved in deionized (DI) water (300 mL). This was followed by the addition of 3,3'-dithiodipropionic acid dihydrazide (3.15 g, 15 mmol), and HOBt (0.69 g, 4.5 mmol), which had been predissolved in a 50 mL mixed solvent (water: acetonitrile = 1:1).

[112] The solution’s pH was then adjusted to 4.7. EDC (287.6 mg, 1.5 mmol) was added to the mixture, and a slow increase in pH was observed, indicating that the reaction was underway. After an overnight stirring, the pH was adjusted to 8.5. An excess amount of DTT (11.6 g, 75 mmol) was then added and the solution was left to stir overnight.

[113] The mixture was subsequently dialyzed against a water solution containing 0.3 M sodium chloride (NaCl) at pH 3.5. Gradually, the dialysis medium was changed to acidified water without NaCl and finally to neutralized, degassed water (pH 7.4). The dialysis was carried out under a nitrogen environment to prevent thiol group oxidation. The product was obtained through lyophilization.

[114] The degree of substitution (DS) of the thiol groups on the hyaluronic acid was calculated to be 0.15 by integrating the methylene protons (82.75 and 2.60 ppm, 4H, - CH2CH2SH) relative to the hyaluronic acid's methyl singlet (81.90 ppm, 3H) using ^JH- NMR in D2O. Different DS may be obtained by varying the synthetic procedure.

Example 4. Preparation of Hydrogels

[115] Dynamic supramolecular hydrogels were prepared using HA- Ada and Gel-Mal polymers described above with either high or low degrees of substitution (DS). A RGD- containing peptide GCGY GRGDSPG (SEQ ID NO: 1) was included, such that a portion of the Gel-Mal bound RGD peptide and a portion bound CD-SH. Human mesenchymal stem cells (HMSC) were also included. To prepare the hydrogel, a HA-Ada/CD-SH solution was mixed with the Gel-Mal/RGD peptide/cell suspension. The hydrogel encapsulated cells and was allowed to react for an additional 30 minutes for full crosslinking. For example, 1 mL of a 6% HA-Ada solution was combined with an equal molar amount of cyclodextrin-SH (CD-SH). After thorough mixing and sonicating, a clear solution formed through the complexation of adamantane guest molecules with host cyclodextrin groups on the polymer chain. The HA-Ada/CD-SH solution was then mixed with an equal volume of 6% Gel-Mal solution, resulting in a viscous gel formation within a minute. The supramolecular hydrogels produced were centrifuged to remove air bubbles and promptly used for rheological measurements.

[116] For example, a Gel-Mal of 0.36 DS was used to conjugate a RGD peptide thereto, before preparing the hydrogel. To conjugate peptide, 7.2% Gel-Mal (DS 0.36) solution (containing 0.0533 mmol of Mai groups per 1 mL) was added to 9.2 mg of GCGYGRGDSPG (SEQ ID NO:1) peptide (trifluoroacetate salt, 1025 g/mol, 0.009 mmol) for 30 minutes of coupling. Such in-situ conjugation resulted in an estimated 7% DS of RGD on the Gel-Mal backbone, thus leaving 0.29 DS maleimide groups for hydrogel crosslinking. The hydrogel using this Gel-Mal/RGD was then used for hydrogel preparation and cell incorporation as follows.

[117] The Gel-Mal/RGD Polymer Solution was filtered and diluted with a cell suspension containing 3 million cells/mL in phosphate-buffered saline. The final concentration of the Gel-VIal/RGD/cell suspension was 4.8% (Gel-Mal, DS 0.36, 0.0297 mmol/mL of remaining Mai groups, 3 mM of RGD, 1 million cells/mL).

[118] The HA-Ada/CD-SH Solution was prepared using a 4% HA- Ada solution (DS = 0.34) contained 0.0296 mmol of Ada groups. An equal molar amount of CD-SH (34 mg, 0.0296 mmol) was added to the HA-Ada solution, and the mixture was vigorously mixed and sonicated.

[119] The hydrogel was prepared using 25 pL of the above 4% HA-Ada/CD-SH solution (DS 0.34, 0.0296 mmol of Ada groups) that was mixed with 25 p L of the above 4.8% Gel- Mal/RGD peptide/cell suspension (DS 0.36, 0.0297 mmol/mL of remaining Mai groups).

[120] This resulted in a hydrogel encapsulated with cells (0.5 million cells/mL) that was allowed to react for an additional 30 minutes for full crosslinking.

[121] In some experiments, the RGD peptide concentration in the final hydrogels is fixed at 1.5 mM.

[122] In some experiments where human mesenchymal stem cells (HMSCs) cells were incorporated into the hydrogel, HMSCs were expanded to passage 5 in growth media. In other studies, neuron stem-like cells (iNSCs) or dental pulp stem cells (DPSCs) were studied.

[123] A control, covalently crosslinked hydrogel was also prepared with similar polymeric network topology. The crosslinking points are changed to static chemical linkages via “thiol-ene” click chemistry. [124] In some experiments, an equimolar amount of HA-SH is added to the gel-mal solution, mixed, and sonicated, for use as a control polymer with covalent, static bonds between the first and second polymers.

[125] Thus, a supramolecular hydrogel was prepared, following the guidance in the prior examples, based on the “Host-Guest” interactions between -cyclodextrin and adamantane (Fig. IB). SEM images are shown in Fig. 2. The dynamic “Host-Guest” interactions allow the network remodeling of hydrogels in response to the cell traction force during cell spreading. A control, covalently crosslinked hydrogel was prepared with similar polymeric network topology. The crosslinking points are changed to static chemical linkages via “thiol-ene” click chemistry.

Example 5. Incorporation of Induced Neuron Stem-like Cells into Supramolecular Hydrogels

[126] Dental pulp stem cells (DPSCs) were cultured (Fig. 4) and their reprogramming induced to neuron stem-like cells; Sox 2 and Nestin are two major markers of neuron stem cells. DPSCs were incubated with NeuronPlus medium (Catalog No. A3582901, Gibco) containing 2% B-27, 50 ng/mL EGF, 20 ng/mE NGF and 20 ng/mE GDNF. The cells exhibited significant expression of Sox 2 and Nestin after 7 days (Figs. 6A, 6B).

[127] Such induced neuron stem-like cells (iNSCs) were incorporated into the herein- described dynamic hydrogels and control, non-dynamic hydrogels. The encapsulated iNSCs in dynamic hydrogels exhibited significant spreading morphology and more expression of key neuron-related markers, Nestin and Sox 2. The quantification also confirmed the larger cell area, higher spreading shape factor (Fig. 6C) and higher expression level of Nestin and Sox 2 (Fig. 6B).

[128] The mechanism of enhanced neurogenesis in dynamic hydrogels was explored. Immunofluorescence staining of integrin-pi and integrin-[33 showed that there is more expression and clustering of integrin-pi in iNSCs encapsulated in the dynamic supramolecular hydrogels, whereas no significant changes were observed for integrin-P3 (Figs. 7A, 7B). The enhanced neurogenesis in dynamic hydrogels can be attributed to the enhanced expression and clustering of integrin-pi in iNSCs encapsulated dynamic hydrogels and the activation of downstream mechanotransduction-related signaling pathways.

[129] Fig. 6 shows enhanced neurogenic differentiation of DPSCs encapsulated in dynamic hydrogels. Neurogenic differentiation is vital in dental pulp regeneration, as it involves the development of cells into neuron-like cells, which can contribute to the restoration of the nerve functions within the dental pulp. This differentiation is particularly important for the restoration of sensory functions in the tooth, which is a critical aspect of dental pulp health

Example 6. Incorporation of Dental Pulp Stem Cells into Supramolecular Hydrogels

[130] Successful dental pulp regeneration requires cell spreading and osteogenic differentiation in the local defects. Local defects in this context refer to areas within the dental pulp or surrounding dentin that have been compromised due to decay, trauma, or disease, leading to a loss of viable tissue and structure. These defects create a need for regeneration to restore the tooth's integrity and functionality. Cultured DPSCs were encapsulated in the dynamic supramolecular “Host-Guest”-based hydrogels for 7 days in osteogenic medium and growth medium. Most of cells cultured in osteogenic medium showed significant cell spreading and expression of osteogenic markers, such as alkaline phosphatase (ALP) and osteocalcin (OCN) (Fig. 5).

Claims

WHAT IS CLAIMED IS:

1. A dynamic supramolecular hydrogel composition comprising:

2. The dynamic supramolecular hydrogel composition of claim 1 , wherein the first polymer and the second polymer are each independently selected from a peptide, a polypeptide, a protein, a polysaccharide, a nucleic acid, or repeating subunits of one or more monomers.

3. The dynamic supramolecular hydrogel of claim 1 or 2, wherein the first polymer subunits and the second polymer subunits are the same.

4. The composition of claim 1 , wherein the ratio of first polymer subunits comprising non-covalently bound cyclodextrin moieties to the total first polymer subunits is about 0.01 to about 0.8, or about 0.17 to about 0.36, or about 0.17 to about 0.29.

5. The dynamic supramolecular hydrogel composition of claim 1 , wherein the cyclodextrin moieties are a- cyclodextrin, 0- cyclodextrin, or y-cyclodextrin, or any combination thereof.

6. The dynamic supramolecular hydrogel composition of claim 1 , wherein the cyclodextrin is covalently bound to the first polymer subunits by maleimide groups covalently bound to the first polymer subunits reacted with thiol groups on the cyclodextrin.

7. The dynamic supramolecular hydrogel composition of claim 2, wherein the first polymer is gelatin.

8. The dynamic supramolecular hydrogel composition of claim 5, wherein the first polymer comprises a cyclodextrin covalently bound to the gelatin as represented in Formula II:

wherein represents a cyclodextrin moiety and

gelatin.

9. The dynamic supramolecular hydrogel composition of claim 5, wherein the first polymer is gelatin covalently bound to P-cyclodextrin.

10. The dynamic supramolecular hydrogel composition of claim 1 , wherein the first polymer further comprises one or more bioactive peptide, an antibody, or a bioactive protein.

11. The dynamic supramolecular hydrogel composition of claim 10, wherein the bioactive peptide is a neuron growth and differentiation-inductive peptide.

12. The dynamic supramolecular hydrogel composition of claim 10 wherein the one or more bioactive peptide, antibody or bioactive protein is selected from or comprises nerve growth factor, RGD or a peptide comprising RGD, an integrin binding peptide, or a fragment of any of the foregoing, or a mimetic of any of the foregoing.

13. The dynamic supramolecular hydrogel composition of claim 10 wherein the bioactive peptide comprises RGD, such as GCGYGRGDSPG (SEQ ID NO:1).

14. The dynamic supramolecular hydrogel composition of claim 10 wherein the bioactive peptide, antibody or bioactive protein comprises a thiol group and is bound to maleimide groups on the first polymer.

15. The dynamic supramolecular hydrogel composition of claim 1 , further comprising a hydrophobic molecule bound to cyclodextrins that are not non- covalently bound to the second polymer.

16. The dynamic supramolecular hydrogel composition of claim 15 , wherein the hydrophobic molecule is a hydrophobic drug.

17. The composition of claim 1 , wherein the ratio of second polymer subunits comprising covalently-bound hydrophobic moieties to the total second polymer subunits is about 0.01 to about 0.8, or about 0.34.

18. The dynamic supramolecular hydrogel composition of claim 1 , wherein the covalently -bound hydrophobic moieties are adamantane moieties.

19. The dynamic supramolecular hydrogel composition of claim 2, wherein the second polymer is a polysaccharide.

20. The dynamic supramolecular hydrogel composition of claim 19 wherein the polysaccharide is hyaluronic acid.

21. The dynamic supramolecular hydrogel composition of claim 20 wherein the hyaluronic acid has a molecular weight of about 10 kDa to about 400 kDa.

22. The dynamic supramolecular hydrogel composition of claim 21 wherein the hyaluronic acid has a molecular weight of about 60 kDa to about 400 kDa.

23. The dynamic supramolecular hydrogel composition of claim 22 whereon the hyaluronic acid has a molecular weight of between about 60 kDa and about 80 kDa.

24. The dynamic supramolecular hydrogel composition of claim 18 wherein the polymer comprising covalently-bound adamantane moieties comprises a polymer of Formula I:

wherein y/(x + y) is between about 0. 01 and about 0.8.

25. The dynamic supramolecular hydrogel composition of claim 1 wherein the second polymer further comprises one or more bioactive peptide, antibody or bioactive protein.

26. The dynamic supramolecular hydrogel composition of claim 25, wherein the bioactive peptide is a neuron growth and differentiation-inductive peptide.

27. The dynamic supramolecular hydrogel composition of claim 25, wherein the bioactive peptide, an antibody, or a bioactive protein is selected from or comprises nerve growth factor, RGD or a peptide comprising RGD, an integrin binding peptide, a fragment of any of the foregoing, or a mimetic of any of the foregoing.

28. The dynamic supramolecular hydrogel composition of claim 27, wherein the bioactive peptide comprises RGD, such as GCGYGRGDSPG (SEQ ID NO:1).

29. The dynamic supramolecular hydrogel composition of claim 25, wherein the one or more bioactive peptide, antibody or bioactive protein is covalently bound to the second polymer.

30. The dynamic supramolecular hydrogel composition of any one of claims 1 -29, wherein the composition remodels in response to a traction force during cell spreading.

31. The dynamic supramolecular hydrogel composition of any one of claims 1 -29, wherein the ratio of polymer subunits comprising covalently-bound hydrophobic moieties to the total polymer subunits provides for the dynamic property.

32. The dynamic supramolecular hydrogel composition of any one of claims 1 -29, wherein the dynamic property is stiffness, strength, toughness, self-healing, shearthinning, fast stress relaxation, and easy matrix remodeling.

33. The dynamic supramolecular hydrogel composition of any one of claims 1-29, wherein the composition mimics the dynamic properties of extracellular matrix.

34. The dynamic supramolecular hydrogel composition of any one of claims 1 -29, wherein the hydrogel is biodegradable.

35. The dynamic supramolecular hydrogel composition of any one of claims 1 -29, further comprising cells.

36. The dynamic supramolecular hydrogel composition of claim 35, wherein the cells are stem cells.

37. The dynamic supramolecular hydrogel composition of claim 35, wherein the cells are immune cells.

38. The dynamic supramolecular hydrogel composition of claim 37, wherein the immune cells are T cells, macrophages or microglia.

39. The dynamic supramolecular hydrogel composition of any one of claims 1-38, further comprising a non-covalently bound bioactive peptide, antibody, bioactive protein or drug.

40. A wound dressing composition comprising the dynamic supramolecular hydrogel composition of any one of claims 1-39.

41. A medical device for in vivo application or implantation comprising the dynamic supramolecular hydrogel composition of any one of claims 1-39.

42. A method for enhancing tissue remodeling, tissue engineering or wound healing, comprising applying to a tissue repair site or wound site the dynamic supramolecular hydrogel composition of any one of claims 1-39.

43. The method of any one of claims 40-42, wherein the dynamic supramolecular hydrogel composition further comprises cells.

44. The method of claim 43, wherein the cells are stem cells, immune cells, or the combination thereof.

45. The method of claim 44, wherein the immune cells are T cells, macrophages, or microglia.

46. The method of claim 43, wherein the cells are obtained from a patient, a donor or a cell line.

47. The method of claim 42 or 43, wherein the composition is applied to the site together with the cells.

48. The method of claim 47, wherein the cells are stem cells, immune cells, or the combination thereof

49. The method of claim 48, wherein the immune cells are T cells, macrophages or microglia.

50. A method for delivering one or more drugs to a bodily site comprising formulating the one or more drugs with the dynamic supramolecular hydrogel composition of any one of claims 1-39, and administering the one or more drugcontaining composition to a bodily site in a subject.

51. The method of claim 50, wherein the each of the one or more drugs is independently covalently bound to the composition, is non-covalently bound to the composition, or the combination thereof.

52. The method of claim 51 , wherein the covalently bound is a labile covalent bond.

53. The method of claim 50, wherein the administering is topical, parenteral, or to a specific bodily site.

54. A method of evaluating in vitro the activities of cells that interact in vivo with extracellular matrix comprising evaluating the cells in vitro in contact with the dynamic supramolecular hydrogel composition of any one of claims 1-39.

55. The method of claim 54, wherein the activities include tissue regeneration, cell migration, cell proliferation, cell differentiation and cell morphogenesis.

56. The method of claim 54, wherein the cells are stem cells.

57. A method for making a dynamic supramolecular hydrogel composition of claim 1 comprising the steps of: a. preparing a hyaluronic acid covalently bound to adamantane moieties; b. preparing a P-cyclodextrin having a sulfhydryl group; c. preparing gelatin substituted with maleimidyl groups ; d. combining the hyaluronic acid covalendy bound to adamantane moieties with the P-cyclodextrin having a sulfhydryl group; e. combining the combination of step d with the gelatin substituted with maleimidyl groups; whereby the maleimidyl groups react with the sulfhydryl groups, thereby forming the composition.

58. The method of claim 57 wherein a portion of the maleimidyl groups on the gelatin are bound to a RGD containing peptide.

59. The method of claim 57 wherein step (e) further comprises including cells.

60. The method of claim 57 wherein the degree of substitution of the gelatin substituted with maleimidyl groups is about 0.01 to about 0.8.

61. The method of claim 60, wherein the degree of substitution of the gelatin substituted with maleimidyl groups is about 0.17 to about 0.36.

62. The method of claim 57, wherein the degree of substitution of the hyaluronic acid covalently conjugated to adamantane moieties is about 0.05 to about 0.5.

63. The method of claim 57, wherein the degree of substitution of the hyaluronic acid covalently conjugated to adamantane moieties is about 0.34.

64. The method of claim 57, wherein the hyaluronic acid has a molecular weight of about 10 kDa to about 400 kDa.

65. The method of claim 64, wherein the hyaluronic acid has a molecular weight of about 60 kDa to about 400 kDa.

66. The method of claim 57, wherein the hyaluronic acid has a molecular weight of about 60 kDa to about 80 kDa.