Glucose-triggered gelation of supramolecular peptide nanocoils with glucose-binding motifs

Abstract

Described herein are self-assembled peptides and hydrogels comprising self-assembled peptides and glucose. Exemplary hydrogels comprising self-assembled peptides and glucose may be used to encapsulate glucagon and / or a glucagon analogue. Hydrogels with glucagon and / or a glucagon analogue encapsulated therein may be used to treat hypoglycemia and related disorders in subjects in need thereof.

Description

[0001] GLUCOSE-TRIGGERED GELATION OF SUPRAMOLECULAR PEPTIDE NANOCOILS WITH GLUCOSE-BINDING MOTIFS CROSS-REFERENCE TO RELATED APPLICATION(S) This application claims priority to U.S. Provisional Patent Application No.63 / 603,502, filed on November 28, 2023, which is incorporated by reference herein in its entirety. REFERENCE TO SEQUENCE LISTING This application was filed with a Sequence Listing XML in ST.26 XML format accordance with 37 C.F.R. § 1.831 and PCT Rule 13ter. The Sequence Listing XML file submitted in the USPTO Patent Center, “092012-0034-WO01_sequence_listing_xml_22-NOV-2024.xml,” was created on November 22, 2024, contains 3 sequences, has a file size of 8.0 kilobytes (8,192 bytes), and is incorporated by reference in its entirety into the specification. BACKGROUND Molecular constituents designed for self-assembly offer a powerful and thermodynamically driven route to achieve nanoscale order and recreate elegant and organized structures found in the living world. Proteins are key building blocks underlying self-assembly of many natural materials. The presence of certain analytes in the physiologic milieu can regulate the state and / or stability of biological matter arising from non-covalent filamentous protein assemblies, as seen in cytoskeletal actin and microtubules. These natural materials have inspired the design of synthetic analogues of small molecule, protein, or colloidal assemblies that form explicitly from the binding and / or chemical reaction of small analyte molecules (e.g., ATP). Accordingly, dynamic, reversible, and / or non-equilibrium assemblies arising from the binding or consumption of small molecule triggers have been demonstrated. Peptides, in particular, are a useful platform from which to design self-assembling nanomaterials, with some designs integrating functionality to further modulate the formation or properties of these assemblies in response to a specific trigger. In the context of analytes found in the human body, glucose is of particular interest; though ubiquitous, poorly controlled glucose levels are a symptomatic hallmark of diabetes. Numerous glucose-responsive material platforms have been designed by leveraging synthetic or protein- derived glucose-binding motifs in order to better treat diabetes. Other related works have used enzymatic conversion of glucose by glucose oxidase (GOx) to actuate pH-dependent material outcomes. The self-assembly of a peptide gelator was designed to be governed by continuous conversion of glucose using GOx, with the material dissipating under conditions of low glucose to release glucagon, a therapeutic remedy for low blood glucose levels. The gelation of this material, driven by the presence of glucose, offered opposing function to most glucose-responsive materials reported thus far, which typically de-gel, swell, or dissipate under conditions of high glucose to release insulin. However, the use of a xenogeneic GOx enzyme to achieve glucose analyte-dependent self-assembly has attendant complications related to its immunogenicity along with toxic hydrogen peroxide byproducts that result from its conversion of glucose to gluconic acid. Phenylboronic acids (PBAs) are Lewis acids that are capable of dynamic-covalent bonding to cis-1,2 and cis-1,3 diols at pH at or above their pKa, stabilizing the negatively charged tetrahedral boronate. As glucose is a cis-1,2 diol, its binding therefore increases the overall charge state of the PBA motif, a feature often used to introduce electrostatic repulsion or swelling in a material for glucose-responsive release of insulin. However, PBA motifs are not specific for glucose binding and are able to bind to many other cis-diol molecules found in the normal physiological environment, and in many cases bind to these better than to glucose. SUMMARY One embodiment described herein is self-assembling peptide comprising: a boronic acid moiety attached to the N-terminus of a sequence of 5–12 amino acid residues, wherein: the terminal amino acid residues of the sequence are positively charged amino acid residues; the non-terminal amino acid residues of the sequence are hydrophobic amino acid residues; the C-terminus of the sequence ; and the boronic acid moiety is a anion thereof, wherein: n RX, at each occurrence, is independently halo, –C1-4haloalkyl, –CN, –NO2, –C(O)H, –CO2H, –CO2C1-4alkyl, or –SO2C1-4alkyl; and X−is an anion having a net charge of −1. In another aspect, the self-assembling peptide is a peptide of formula (I): (I), wherein: B1is the boronic acid moiety; and is the sequence of amino acid residues, wherein: AA1and AA8are the terminal amino acid residues; AA2and AA6are aromatic amino acid residues; AA3, AA4, AA5and AA7are non-aromatic amino acid residues; and the C- . In another aspect, AA1is an an ornithine residue, or a histidine residue. In another aspect, AA2is a phenylalanine residue, a tyrosine residue, or a tryptophan residue. In another aspect, AA3, AA4, and AA5are each independently a glycine residue, an alanine residue, a valine residue, an isoleucine residue, or a leucine residue. In another aspect, AA6is a tryptophan residue, a phenylalanine residue, or a tyrosine residue. In another aspect, AA7is a glycine residue, an alanine residue, a valine residue, an isoleucine residue, or a leucine residue. In another aspect, AA8is an arginine residue, a lysine residue, an ornithine residue, or a histidine residue. In another aspect, the peptide of formula (I) is a peptide of formula (I-a): a), ; R3, R4, and R5are each independently hydrogen, ; . In In another . In another . In another In another aspect, R5is methyl. . In another aspect, R7is . . or In another aspect, RXis –F or –C1-4fluoroalkyl. In another aspect, the self-assembling peptide is a peptide of formula: . peptide and glucose. In another aspect, glucagon and / or a glucagon analogue is encapsulated within the hydrogel. In another aspect, the glucagon analogue comprises dasiglucagon and / or a depsi- glucagon analogue. In another aspect, the glucagon and / or glucagon analogue is co-formulated with a cationic polymer within the hydrogel. In another aspect, the cationic polymer is protamine. In another aspect, the molar ratio of the self-assembling peptide to glucose is from about 1:100 to about 1:1000. Another embodiment described herein is a pharmaceutical composition comprising glucagon and / or a glucagon analogue encapsulated within a hydrogel, and a pharmaceutically acceptable excipient. Another embodiment described herein is a method of modulating glucose levels in a subject, the method comprising: administering a therapeutically effective amount of a hydrogel or a pharmaceutical composition. In another aspect, the subject in need thereof is experiencing a hypoglycemic event or at risk of experiencing a hypoglycemic event. In another aspect, the subject in need thereof has diabetes. Another embodiment described herein is a kit comprising a hydrogel or a pharmaceutical composition; delivery or administration apparata or devices; and optionally, packaging, a label, or instructions for use. DESCRIPTION OF THE DRAWINGS FIG. 1 depicts the chemical structure of multidomain peptide PBA–MDP and control molecules (COOH–MDP and OH–MDP) used in this study (top) which demonstrate a preference for antiparallel dimer formation. PBA–MDP contains phenylboronic acid (PBA) which is Lewis acid capable of forming a charged tetrahedral boronate through dynamic-covalent interactions with glucose. Short nanocoils arise from PBA–MDP self-assembly, and upon addition of glucose these nanocoils elongate and entangle (bottom). FIG.2A–J show analyses of PBA–MDP. FIG.2A shows cryogenic transmission electron microscopy (cryo-TEM) performed on PBA–MDP without glucose. FIG. 2B shows cryogenic transmission electron microscopy (cryo-TEM) performed on PBA–MDP with 100 mg / dL glucose. FIG.2C shows liquid-phase atomic force microscopy (AFM) on PBA–MDP without glucose. FIG. 2D shows small angle X-ray scattering (SAXS) performed on PBA–MDP with or without addition of 100 mg / dL glucose. FIG.2E shows 2D class average (inset) and power spectrum of vertically aligned nanocoil segments. FIG. 2F shows the cryo-EM density map of the nanocoil (left) and the atomic model of nanocoil fitted into the density map (right). FIG.2G graphically shows circular dichroism (CD) spectroscopy of PBA–MDP with increasing concentration of glucose, along with controls of COOH–MDP and OH–MDP. FIG. 2H graphically shows Fourier transform infrared (FTIR) spectroscopy of PBA–MDP with increasing concentration of glucose, along with controls of COOH–MDP and OH–MDP. The dashed vertical lines denote peaks of interest in the Amide I region of the spectra. FIG. 2I graphically shows thioflavin-T (ThT) fluorescence of PBA–MDP upon immediate addition of glucose or when glucose is added after 4 h of continuous monitoring. FIG. 2J graphically shows zeta potential measurements for PBA–MDP with increasing concentration of glucose, along with controls of COOH–MDP and OH–MDP (n = 3 samples / group, mean ± SD shown). FIG.3A–E show hydrogelation of PBA–MDP. FIG.3A shows hydrogelation of PBA–MDP as observed by vial inversion under different glucose conditions. FIG.3B graphically shows the evolution of the storage modulus (G′, solid, left axis) and complex viscosity (η, dashed, right axis) over time following addition of glucose and application onto the rheometer. FIG.3C graphically shows glucose-dependent changes in the plateau storage modulus (G′, solid, left axis) and complex viscosity (η, open, right axis), from n = 2 samples / group (mean ± SD shown). FIG.3D is a bar graph showing shear viscosity ramp for a PBA–MDP hydrogel prepared in 100 mg / dL glucose. FIG.3E graphically shows step-strain cycling from 1% to 100% strain for a PBA–MDP hydrogel prepared in 100 mg / dL glucose. FIG. 4A–D show PBA–MDP hydrogel release and in vivo studies. FIG. 4A graphically shows the release of encapsulated dasiglucagon from PBA–MDP hydrogels prepared at 100 mg / dL glucose and incubated in a buffer of varying glucose levels (n = 3 samples / group, mean ± SD shown). FIG.4B schematically depicts the mouse model for prophylactic glucagon delivery prior to severe hypoglycemia caused by insulin overdose. FIG. 4C graphically shows blood glucose monitored over time, focusing specifically on the region following insulin overdose (zoomed shaded region). FIG.4D shows a bar graph illustrating the lowest (nadir) blood glucose level observed at 4 h after the insulin overdose. FIG.4E shows a bar graph illustrating the final blood glucose level at 4 h after the insulin overdose. For FIG.4C–4E, n = 9–10 mice / group, mean ± SEM shown. For FIG. C, multiple unpaired t-tests were performed at each timepoint, using the Holm-Šídák method to correct for multiple comparisons (*-P < 0.05 for PBA–MDP vs. all other groups from 180 min onward). For FIG.4D-4E, a one-way ANOVA was performed with Bonferroni multiple comparisons post-hoc testing (*-P < 0.05, **-P < 0.01 for PBA–MDP vs. all other groups). FIG.5 shows the chemical structures of PBA–MDP, COOH–MDP, and OH–MDP. FIG.6 shows transmission electron microscopy (TEM) images of PBA–MDP over a range of glucose concentrations (0, 25, 50, 100, and 200 mg / dL. FIG.7 shows TEM images of COOH–MDP. FIG.8 shows TEM images of OH–MDP. FIG.9 shows graphs depicting the rheology of COOH–MDP. FIG. 10 graphically shows the release profile of methoxycoumarin (MCA)-dasiglucagon when not co-formulated is protamine. DETAILED DESCRIPTION Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. For example, any nomenclatures used in connection with, and techniques of biochemistry, molecular biology, immunology, microbiology, genetics, cell and tissue culture, and protein and nucleic acid chemistry described herein are well known and commonly used in the art. In case of conflict, the present disclosure, including definitions, will control. Exemplary methods and materials are described below, although methods and materials similar or equivalent to those described herein can be used in practice or testing of the embodiments and aspects described herein. As used herein, the terms “amino acid,” “nucleotide,” “polynucleotide,” “vector,” “polypeptide,” and “protein” have their common meanings as would be understood by a biochemist of ordinary skill in the art. Standard single letter nucleotides (A, C, G, T, U) and standard single letter amino acids (A, C, D, E, F, G, H, I, K, L, M, N, P, Q, R, S, T, V, W, or Y) are used herein. As used herein, terms such as “include,” “including,” “contain,” “containing,” “having,” and the like mean “comprising.” The present disclosure also contemplates other embodiments “comprising,” “consisting essentially of,” and “consisting of” the embodiments or elements presented herein, whether explicitly set forth or not. As used herein, “comprising,” is an “open- ended” term that does not exclude additional, unrecited elements or method steps. As used herein, “consisting essentially of” limits the scope of a claim to the specified materials or steps and those that do not materially affect the basic and novel characteristics of the claimed invention. As used herein, “consisting of” excludes any element, step, or ingredient not specified in the claim. As used herein, the term “a,” “an,” “the” and similar terms used in the context of the disclosure (especially in the context of the claims) are to be construed to cover both the singular and plural unless otherwise indicated herein or clearly contradicted by the context. In addition, “a,” “an,” or “the” means “one or more” unless otherwise specified. As used herein, the term “or” can be conjunctive or disjunctive. As used herein, the term “and / or” refers to both the conjunctive and disjunctive. As used herein, the term “substantially” means to a great or significant extent, but not completely. As used herein, the term “about” or “approximately” as applied to one or more values of interest, refers to a value that is similar to a stated reference value, or within an acceptable error range for the particular value as determined by one of ordinary skill in the art, which will depend in part on how the value is measured or determined, such as the limitations of the measurement system. In one aspect, the term “about” refers to any values, including both integers and fractional components that are within a variation of up to ± 10% of the value modified by the term “about.” Alternatively, “about” can mean within 3 or more standard deviations, per the practice in the art. Alternatively, such as with respect to biological systems or processes, the term “about” can mean within an order of magnitude, in some embodiments within 5-fold, and in some embodiments within 2-fold, of a value. As used herein, the symbol “~” means “about” or “approximately.” All ranges disclosed herein include both end points as discrete values as well as all integers and fractions specified within the range. For example, a range of 0.1–2.0 includes 0.1, 0.2, 0.3, 0.4 . . . 2.0. If the end points are modified by the term “about,” the range specified is expanded by a variation of up to ±10% of any value within the range or within 3 or more standard deviations, including the end points, or as described above in the definition of “about.” As used herein, the terms “room temperature,” “RT,” or “ambient temperature” refer to the typical temperature in an indoor laboratory setting. In one aspect, the laboratory setting is climate controlled to maintain the temperature at a substantially uniform temperature or with a specific range of temperatures. In one aspect, “room temperature” refers a temperature of about 15–30 °C, including all integers and endpoints within the specified range. In another aspect, “room temperature” refers a temperature of about 15–30 °C; about 20–30 °C; about 22–30 °C; about 25–30 °C; about 27–30 °C; about 15–22 °C; about 15–25 °C; about 15–27 °C; about 20–22 °C; about 20–25 °C; about 20–27 °C; about 22–25 °C; about 22–27 °C; about 25–27 °C; about 15 °C ± 10%; about 20 °C ± 10%; about 22 °C ± 10%; about 25 °C ± 10%; about 27 °C ± 10%; ~20 °C, ~22 °C, ~25 °C, or ~27 °C, at standard atmospheric pressure. As used herein, the terms “active ingredient” or “active pharmaceutical ingredient” refer to a pharmaceutical agent, active ingredient, compound, or substance, compositions, or mixtures thereof, that provide a pharmacological, often beneficial, effect. As used herein, the terms “control,” or “reference” are interchangeable. A “reference” or “control” level may be a predetermined value or range, which is employed as a baseline or benchmark against which to assess a measured result. “Control” also refers to control experiments or control cells. As used herein, the term “dose” denotes any form of an active ingredient formulation or composition, including cells, that contains an amount sufficient to initiate or produce a therapeutic effect with at least one or more administrations. “Formulation” and “composition” are used interchangeably herein. As used herein, the term “prophylaxis” refers to preventing or reducing the progression of a disorder, either to a statistically significant degree or to a degree detectable by a person of ordinary skill in the art. As used herein, the terms “effective amount” or “therapeutically effective amount,” refers to a substantially non-toxic, but sufficient amount of an action, agent, composition, or cell(s) being administered to a subject that will prevent, treat, or ameliorate to some extent one or more of the symptoms of the disease or condition being experienced or that the subject is susceptible to contracting. The result can be the reduction or alleviation of the signs, symptoms, or causes of a disease, or any other desired alteration of a biological system. An effective amount may be based on factors individual to each subject, including, but not limited to, the subject’s age, size, type or extent of disease, stage of the disease, route of administration, the type or extent of supplemental therapy used, ongoing disease process, and type of treatment desired. As used herein, the term “subject” refers to an animal. Typically, the subject is a mammal. A subject also refers to primates (e.g., humans, male or female; infant, adolescent, or adult), non- human primates, rats, mice, rabbits, pigs, cows, sheep, goats, horses, dogs, cats, fish, birds, and the like. In one embodiment, the subject is a primate. In one embodiment, the subject is a human. As used herein, a subject is “in need of treatment” if such subject would benefit biologically, medically, or in quality of life from such treatment. A subject in need of treatment does not necessarily present symptoms, particular in the case of preventative or prophylaxis treatments. As used herein, the terms “inhibit,” “inhibition,” or “inhibiting” refer to the reduction or suppression of a given biological process, condition, symptom, disorder, or disease, or a significant decrease in the baseline activity of a biological activity or process. As used herein, “treatment” or “treating” refers to prophylaxis of, preventing, suppressing, repressing, reversing, alleviating, ameliorating, or inhibiting the progress of biological process including a disorder or disease, or completely eliminating a disease. A treatment may be either performed in an acute or chronic way. The term “treatment” also refers to reducing the severity of a disease or symptoms associated with such disease prior to affliction with the disease. “Repressing” or “ameliorating” a disease, disorder, or the symptoms thereof involves administering a cell, composition, or compound described herein to a subject after clinical appearance of such disease, disorder, or its symptoms. “Prophylaxis of” or “preventing” a disease, disorder, or the symptoms thereof involves administering a cell, composition, or compound described herein to a subject prior to onset of the disease, disorder, or the symptoms thereof. “Suppressing” a disease or disorder involves administering a cell, composition, or compound described herein to a subject after induction of the disease or disorder thereof but before its clinical appearance or symptoms thereof have manifest. Definitions of specific functional groups and chemical terms are described in more detail herein. The chemical elements are identified in accordance with the Periodic Table of the Elements, CAS version, Handbook of Chemistry and Physics, 75thed., inside cover, and specific functional groups are generally defined as described therein. Additionally, general principles of organic chemistry, as well as specific functional moieties and reactivity, are described in Thomas Sorrell, Organic Chemistry, University Science Books, Sausalito, 1999; Smith and March, March’s Advanced Organic Chemistry, 5thed, John Wiley & Sons, Inc., New York, 2001; Larock, Comprehensive Organic Transformations, VCH Publishers, Inc., New York, 1989; and Carruthers, Some Modern Methods of Organic Synthesis, 3rded, Cambridge University Press, Cambridge, 1987. The term “alkyl,” as used herein, means a straight or branched, saturated hydrocarbon chain. The term “lower alkyl” or “C1-6alkyl” means a straight or branched chain hydrocarbon containing from 1 to 6 carbon atoms. The term “C1-4alkyl” means a straight or branched chain hydrocarbon containing from 1 to 4 carbon atoms. Representative examples of alkyl include, but are not limited to, methyl, ethyl, n-propyl, iso-propyl, n-butyl, sec-butyl, iso-butyl, tert-butyl, n- pentyl, isopentyl, neopentyl, n-hexyl, 3-methylhexyl, 2,2-dimethylpentyl, 2,3-dimethylpentyl, n- heptyl, n-octyl, n-nonyl, and n-decyl. The term “halogen” or “halo,” as used herein, means Cl, Br, I, or F. The term “haloalkyl,” as used herein, means an alkyl group, as defined herein, in which one, two, three, four, five, six, seven or eight hydrogen atoms are replaced by a halogen. The term “haloalkoxy,” as used herein, means at least one haloalkyl group, as defined herein, is appended to the parent molecular moiety through an oxygen atom. The term “substituted” refers to a group that may be further substituted with one or more non-hydrogen substituent groups. Substituent groups include, but are not limited to, halogen, =O (oxo), =S (thioxo), cyano, nitro, fluoroalkyl, alkoxyfluoroalkyl, fluoroalkoxy, alkyl, alkenyl, alkynyl, haloalkyl, haloalkoxy, heteroalkyl, cycloalkyl, cycloalkenyl, aryl, heteroaryl, heterocycle, cycloalkylalkyl, heteroarylalkyl, arylalkyl, hydroxy, hydroxyalkyl, alkoxy, alkoxyalkyl, alkylene, aryloxy, phenoxy, benzyloxy, amino, alkylamino, acylamino, aminoalkyl, arylamino, sulfonylamino, sulfinylamino, sulfonyl, alkylsulfonyl, arylsulfonyl, aminosulfonyl, sulfinyl, -COOH, ketone, amide, carbamate, and acyl. For compounds described herein, groups and substituents thereof may be selected in accordance with permitted valence of the atoms and the substituents, such that the selections and substitutions result in a stable compound, e.g., which does not spontaneously undergo transformation such as by rearrangement, cyclization, elimination, etc. The term “peptide,” as used herein, means a compound comprising two or more amino acids. As used herein, the term “peptide” encompasses peptides terminated with a carboxyl group and peptides terminated with a free amide group (–C(O)NH2). The term “amino acid,” as used herein, means an organic compound that contains both amino and carboxyl functional group. As used herein, the term “amino acid” may refer to a native amino acid residue (i.e., a proteogenic amino acid residue) or a non-native amino acid residue (i.e., a non-proteogenic amino acid residue). The term “amino acid residue,” as used herein, means the part of an amino acid that remains after two or more amino acids combine to form a peptide. As used herein, the term “amino acid residue” may refer to a native amino acid residue (i.e., a proteogenic amino acid residue) or a non-native amino acid residue (i.e., a non-proteogenic amino acid residue). The term “N-terminus,” as used herein, means the nitrogen terminus of an amino acid, peptide, or protein. The term “C-terminus,” as used herein, means the carbon terminus of an amino acid, peptide, or protein. Herein, a self-assembling multidomain peptide (MDP) bearing a terminal PBA motif (PBA– MDP) is reported to assemble through antiparallel stacking into an uncommon nanocoil morphology (FIG.1). Upon addition of glucose, preexisting short nanocoils elongate and entangle to form self-supporting hydrogels. The hydrogels can encapsulate glucagon and analogues thereof, with their controlled release inversely related to the bulk glucose concentration. When explored in an animal model of acute hypoglycemia, prophylactic injection of glucagon-loaded PBA–MDP hydrogels offers protection against severe outcomes and more rapid blood glucose recovery. Accordingly, the glucose-triggered entanglement and gelation of PBA–MDP nanocoils provides a synthetic and enzyme-free route to link material form to glucose level. Self-Assembling Peptides In one aspect, the present disclosure provides peptides that are capable of forming ordered structures through self-assembly (i.e., “self-assembling peptides”). The self-assembling peptides disclosed herein comprise: a boronic acid moiety attached to the N-terminus of a sequence of 5–12 amino acid residues, wherein: the terminal amino acid residues of the sequence are positively charged amino acid residues; the non-terminal amino acid residues of the sequence are hydrophobic amino acid residues; OH the C-terminus of the sequencethe boronic acid moiety is a anion thereof, wherein: n –CN, –NO2, –C(O)H, –CO2H, –CO2C1-4alkyl, or –SO2C1-4alkyl; and X−is an anion having a net charge of −1. In various instances, the self-assembling peptide may be a peptide of formula (I): (I), wherein: B1is the boronic acid moiety; and is the sequence of amino acid residues, wherein: AA1and AA8are the terminal amino acid residues; AA2and AA6are aromatic amino acid residues; AA3, AA4, AA5and AA7are non-aromatic amino acid residues; and OHthe C-. In arginine residue, a lysine residue, an ornithine residue, or a histidine residue. In various instances, AA2may be a phenylalanine residue, a tyrosine residue, or a tryptophan residue. In various instances, AA3, AA4, and AA5each independently may be a glycine residue, an alanine residue, a valine residue, an isoleucine residue, or a leucine residue. In various instances, AA6may be a tryptophan residue, a phenylalanine residue, or a tyrosine residue. In various instances, AA7may be a glycine residue, an alanine residue, a valine residue, an isoleucine residue, or a leucine residue. In various instances, AA8may be an arginine residue, a lysine residue, an ornithine residue, or a histidine residue. In various instances, the peptide of formula (I) may be a peptide of formula (I-a): a), ; 12In various instances, R ma . In various instances, R2may . 3 In various instances, R may . In various instances, R4may In various instances, R5may In various instances, R6. In various instances, R7may . In various instances, R8. In various instances, B1. In various instances, B1may be . In various instances, n may be In various instances, RXmay be halo or –C1-4haloalkyl. In various instances, RXmay be –F or –C1-4fluoroalkyl. In various instances, the self-assembling peptide may be a peptide of formula:

[0002] . enters are present. The stereoisomer is “R” or “S” depending on the configuration of substituents around the chiral carbon atom. The terms “R” and “S” used herein are configurations as defined in IUPAC 1974 Recommendations for Section E, Fundamental Stereochemistry, in Pure Appl. Chem., 1976, 45: 13-30. The disclosure contemplates various stereoisomers and mixtures thereof and these are specifically included within the scope of this invention. Stereoisomers include enantiomers and diastereomers, and mixtures of enantiomers or diastereomers. Individual stereoisomers of the compounds may be prepared synthetically from commercially available starting materials, which contain asymmetric or chiral centers or by preparation of racemic mixtures followed by methods of resolution well-known to those of ordinary skill in the art. These methods of resolution are exemplified by (1) attachment of a mixture of enantiomers to a chiral auxiliary, separation of the resulting mixture of diastereomers by recrystallization or chromatography and optional liberation of the optically pure product from the auxiliary as described in Furniss, Hannaford, Smith, and Tatchell, “Vogel’s Textbook of Practical Organic Chemistry”, 5th edition (1989), Longman Scientific & Technical, Essex CM202JE, England, or (2) direct separation of the mixture of optical enantiomers on chiral chromatographic columns or (3) fractional recrystallization methods. It should be understood that the peptide may possess tautomeric forms, as well as geometric isomers, and that these also constitute an aspect of the invention. In the peptides, e.g., peptides of formula (I) and any subformulas, any “hydrogen” or “H,” whether explicitly recited or implicit in the structure, encompasses hydrogen isotopes1H (protium) and2H (deuterium). The present disclosure also includes isotopically-labeled peptides (e.g., deuterium labeled), where an atom in the isotopically-labeled peptides is specified as a particular isotope of the atom. Examples of isotopes suitable for inclusion in the peptides of the invention are hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, and chlorine, such as, but not limited to2H,3H,13C,14C,15N,18O,17O,31P,32P,35S,18F, and36Cl, respectively. Isotopically-enriched forms of peptides of formula (I), or any subformulas, may generally be prepared by conventional techniques known to those skilled in the art or by processes analogous to those described in the accompanying Examples using an appropriate isotopically- enriched reagent in place of a non-isotopically-enriched reagent. The extent of isotopic enrichment can be characterized as a percent incorporation of a particular isotope at an isotopically-labeled atom (e.g., % deuterium incorporation at a deuterium label). Pharmaceutical Salts The disclosed peptides may exist as pharmaceutically acceptable salts. The term “pharmaceutically acceptable salt” refers to salts or zwitterions of the compounds which are water or oil-soluble or dispersible, suitable for treatment of disorders without undue toxicity, irritation, and allergic response, commensurate with a reasonable benefit / risk ratio and effective for their intended use. The salts may be prepared during the final isolation and purification of the compounds or separately by reacting an amino group of the compounds with a suitable acid. For example, a compound may be dissolved in a suitable solvent, such as but not limited to methanol and water and treated with at least one equivalent of an acid, like hydrochloric acid. The resulting salt may precipitate out and be isolated by filtration and dried under reduced pressure. Alternatively, the solvent and excess acid may be removed under reduced pressure to provide a salt. Representative salts include acetate, adipate, alginate, citrate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, camphorate, camphorsulfonate, digluconate, glycerophosphate, hemisulfate, heptanoate, hexanoate, formate, isethionate, fumarate, lactate, maleate, methanesulfonate, naphthylenesulfonate, nicotinate, oxalate, pamoate, pectinate, persulfate, 3-phenylpropionate, picrate, oxalate, maleate, pivalate, propionate, succinate, tartrate, thrichloroacetate, trifluoroacetate, glutamate, para-toluenesulfonate, undecanoate, hydrochloric, hydrobromic, sulfuric, phosphoric and the like. The amino groups of the compounds may also be quaternized with alkyl chlorides, bromides, and iodides such as methyl, ethyl, propyl, isopropyl, butyl, lauryl, myristyl, stearyl and the like. Basic addition salts may be prepared during the final isolation and purification of the disclosed compounds by reaction of a carboxyl group with a suitable base such as the hydroxide, carbonate, or bicarbonate of a metal cation such as lithium, sodium, potassium, calcium, magnesium, or aluminum, or an organic primary, secondary, or tertiary amine. Quaternary amine salts can be prepared, such as those derived from methylamine, dimethylamine, trimethylamine, triethylamine, diethylamine, ethylamine, tributylamine, pyridine, N,N-dimethylaniline, N- methylpiperidine, N-methylmorpholine, dicyclohexylamine, procaine, dibenzylamine, N,N- dibenzylphenethylamine, 1-ephenamine and N,N′-dibenzylethylenediamine, ethylenediamine, ethanolamine, diethanolamine, piperidine, piperazine, and the like. General Synthesis The self-assembling peptides disclosed herein may be prepared according to peptide synthesis methods known in the art. In some instances, the self-assembling peptides may be prepared using solid-phase synthesis. Optimum reaction conditions and reaction times for each individual step can vary depending on the particular reactants employed and substituents present in the reactants used. Specific procedures are provided in the Examples section. Reactions can be worked up in the conventional manner, e.g., by eliminating the solvent from the residue and further purified according to methodologies generally known in the art such as, but not limited to, crystallization, distillation, extraction, trituration, and chromatography. Unless otherwise described, the starting materials and reagents are either commercially available or can be prepared by one skilled in the art from commercially available materials using methods described in the chemical literature. Starting materials, if not commercially available, can be prepared by procedures selected from standard organic chemical techniques, techniques that are analogous to the synthesis of known, structurally similar compounds, or techniques that are analogous to the above-described schemes or the procedures described in the synthetic examples section. Routine experimentations, including appropriate manipulation of the reaction conditions, reagents and sequence of the synthetic route, protection of any chemical functionality that cannot be compatible with the reaction conditions, and deprotection at a suitable point in the reaction sequence of the method are included in the scope of the invention. Suitable protecting groups and the methods for protecting and deprotecting different substituents using such suitable protecting groups are well known to those skilled in the art; examples of which can be found in PGM Wuts and TW Greene, in Greene’s book titled Protective Groups in Organic Synthesis (4thed.), John Wiley & Sons, NY (2006), which is incorporated herein by reference in its entirety. Synthesis of the compounds of the invention can be accomplished by methods analogous to those described in the synthetic schemes described hereinabove and in specific examples. When an optically active form of a disclosed compound is required, it can be obtained by carrying out one of the procedures described herein using an optically active starting material (prepared, for example, by asymmetric induction of a suitable reaction step), or by resolution of a mixture of the stereoisomers of the compound or intermediates using a standard procedure (such as chromatographic separation, recrystallization, or enzymatic resolution). Similarly, when a pure geometric isomer of a compound is required, it can be obtained by carrying out one of the above procedures using a pure geometric isomer as a starting material, or by resolution of a mixture of the geometric isomers of the compound or intermediates using a standard procedure such as chromatographic separation. It can be appreciated that the synthetic schemes and specific examples as described are illustrative and are not to be read as limiting the scope of the invention as it is defined in the appended claims. All alternatives, modifications, and equivalents of the synthetic methods and specific examples are included within the scope of the claims. Hydrogels In another aspect, the present disclosure provides hydrogels comprising a self-assembling peptide and glucose. As used here, these hydrogels arise from physical entanglements of the self-assembled peptide nanostructures that from upon glucose binding to PBA motifs on the peptide. In various instances, the molar ratio of the self-assembling peptide to glucose is from about 1:100 to about 1:1000. In some instances, the molar ratio of the self-assembling peptide to glucose is from about 1:100 to about 1:900; about 1:150 to about 1:850; about 1:200 to about 1:800; about 1:250 to about 1:750; about 1:300 to about 1:700; about 1:350 to about 1:650; about 1:400 to about 1:600; or from about 1:450 to about 1:550. In some instances, the molar ratio of the self-assembling peptide to glucose is no greater than about 1:1000; no greater than about 1:900; no greater than about 1:800; no greater than about 1:700; no greater than about 1:600; no greater than about 1:500; no greater than about 1:400; no greater than about 1:300; or no greater than about 1:200. In some instances, the molar ratio of the self-assembling peptide to glucose is no less than 1:100; no less than 1:200; no less than 1:300; no less than 1:400; no less than 1:500; no less than 1:600; no less than 1:700; no less than 1:800; or no less than 1:900. In various instances, glucagon and / or a glucagon analogue may be encapsulated within the hydrogel. The glucagon analogue may comprise dasiglucagon and / or a depsi-glucagon analogue. In various instances, the glucagon and / or glucagon analogue may be co-formulated with a cationic polymer within the hydrogel. The cationic polymer may be protamine. Glucagon and Glucagon Analogues Glucagon administered at low doses may prevent insulin-induced hypoglycemia or improve the ability to recover from hypoglycemia. However, glucagon is of limited use in pharmaceuticals due to fast clearance from circulation with a half-life of approximately 5 min. Compared to glucagon, glucagon-like analogues (i.e., “glucagon analogues”) may demonstrate improved physical stability toward gel and fibril formation, improved chemical stability and increased half-life, while also showing improved aqueous solubility at neutral pH or slightly basic pH. Glucagon analogues mimic the endogenous hormone glucagon-like peptide 1 (GLP-1), a gastrointestinal hormone that is released into the circulation in response to ingested nutrients. Various exemplary glucagon-based analogues are described in U.S. Patent No.9,486,506 B2. In various instances, the glucagon analogue comprises one or more of: dasiglucagon and / or a depsi- glucagon analogue. Various exemplary depsi-glucagon analogues are described in international patent publication WO 2017 / 210168 A1. Pharmaceutical Compositions Pharmaceutical compositions of the present invention may comprise glucagon and / or a glucagon analogue encapsulated within the hydrogels disclosed herein (i.e., “hydrogel- encapsulated glucagon and / or glucagon analogue”). The hydrogel-encapsulated glucagon / glucagon analogue may be incorporated into pharmaceutical compositions suitable for administration to a subject (such as a patient, which may be a human or non-human). The pharmaceutical compositions may include a “therapeutically effective amount” or a “prophylactically effective amount” of the active agent (glucagon and / or glucagon analogue). A “therapeutically effective amount” refers to an amount effective, at dosages and for periods of time necessary, to achieve the desired therapeutic result. A therapeutically effective amount of the composition may be determined by a person skilled in the art and may vary according to factors such as the disease state, age, sex, and weight of the individual, and the ability of the composition to elicit a desired response in the individual. A “therapeutically effective amount” is also one in which any toxic or detrimental effects are outweighed by the therapeutically beneficial effects. A “prophylactically effective amount” refers to an amount effective, at dosages and for periods of time necessary, to achieve the desired prophylactic result. Typically, since a prophylactic dose is used in subjects prior to or at an earlier stage of disease, the prophylactically effective amount will be less than the therapeutically effective amount. The pharmaceutical compositions may include pharmaceutically acceptable carriers. The term “pharmaceutically acceptable carrier,” as used herein, means a non-toxic, inert solid, semi- solid or liquid filler, diluent, encapsulating material, or formulation auxiliary of any type. Some examples of materials which can serve as pharmaceutically acceptable carriers are sugars such as, but not limited to, lactose, glucose and sucrose; starches such as, but not limited to, corn starch and potato starch; cellulose and its derivatives such as, but not limited to, sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; powdered tragacanth; malt; gelatin; talc; excipients such as, but not limited to, cocoa butter and suppository waxes; oils such as, but not limited to, peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; glycols; such as propylene glycol; esters such as, but not limited to, ethyl oleate and ethyl laurate; agar; buffering agents such as, but not limited to, magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer’s solution; ethyl alcohol, and phosphate buffer solutions, as well as other non-toxic compatible lubricants such as, but not limited to, sodium lauryl sulfate and magnesium stearate, as well as coloring agents, releasing agents, coating agents, sweetening, flavoring and perfuming agents, preservatives and antioxidants can also be present in the composition, according to the judgment of the formulator. Thus, the hydrogels and their physiologically acceptable salts and solvates may be formulated for administration by, for example, solid dosing, eyedrop, in a topical oil-based formulation, injection, inhalation (either through the mouth or the nose), implants, or oral, buccal, parenteral, or rectal administration. Techniques and formulations may generally be found in “Remington’s Pharmaceutical Sciences,” (Meade Publishing Co., Easton, Pa.). Therapeutic compositions must typically be sterile and stable under the conditions of manufacture and storage. The route by which the hydrogel-encapsulated glucagon / glucagon analogue is administered, and the form of the composition will dictate the type of carrier to be used. The composition may be in a variety of forms, suitable, for example, for systemic administration (e.g., oral, rectal, nasal, sublingual, buccal, implants, or parenteral) or topical administration (e.g., dermal, pulmonary, nasal, aural, ocular, liposome delivery systems, or iontophoresis). Carriers for systemic administration typically include at least one of diluents, lubricants, binders, disintegrants, colorants, flavors, sweeteners, antioxidants, preservatives, glidants, solvents, suspending agents, wetting agents, surfactants, combinations thereof, and others. All carriers are optional in the compositions. Suitable diluents include sugars such as glucose, lactose, dextrose, and sucrose; diols such as propylene glycol; calcium carbonate; sodium carbonate; sugar alcohols, such as glycerin; mannitol; and sorbitol. The amount of diluent(s) in a systemic or topical composition is typically about 50 to about 90%. Suitable lubricants include silica, talc, stearic acid and its magnesium salts and calcium salts, calcium sulfate; and liquid lubricants such as polyethylene glycol and vegetable oils such as peanut oil, cottonseed oil, sesame oil, olive oil, corn oil and oil of theobroma. The amount of lubricant(s) in a systemic or topical composition is typically about 5 to about 10%. Suitable binders include polyvinyl pyrrolidone; magnesium aluminum silicate; starches such as corn starch and potato starch; gelatin; tragacanth; and cellulose and its derivatives, such as sodium carboxymethylcellulose, ethyl cellulose, methylcellulose, microcrystalline cellulose, and sodium carboxymethylcellulose. The amount of binder(s) in a systemic composition is typically about 5 to about 50%. Suitable disintegrants include agar, alginic acid and the sodium salt thereof, effervescent mixtures, croscarmellose, crospovidone, sodium carboxymethyl starch, sodium starch glycolate, clays, and ion exchange resins. The amount of disintegrant(s) in a systemic or topical composition is typically about 0.1 to about 10%. Suitable colorants include a colorant such as an FD&C dye. When used, the amount of colorant in a systemic or topical composition is typically about 0.005 to about 0.1%. Suitable flavors include menthol, peppermint, and fruit flavors. The amount of flavor(s), when used, in a systemic or topical composition is typically about 0.1 to about 1.0%. Suitable sweeteners include aspartame and saccharin. The amount of sweetener(s) in a systemic or topical composition is typically about 0.001 to about 1%. Suitable antioxidants include butylated hydroxyanisole (“BHA”), butylated hydroxytoluene (“BHT”), and vitamin E. The amount of antioxidant(s) in a systemic or topical composition is typically about 0.1 to about 5%. Suitable preservatives include benzalkonium chloride, methyl paraben and sodium benzoate. The amount of preservative(s) in a systemic or topical composition is typically about 0.01 to about 5%. Suitable glidants include silicon dioxide. The amount of glidant(s) in a systemic or topical composition is typically about 1 to about 5%. Suitable solvents include water, isotonic saline, ethyl oleate, glycerine, hydroxylated castor oils, alcohols such as ethanol, and phosphate buffer solutions. The amount of solvent(s) in a systemic or topical composition is typically from about 0 to about 100%. Suitable suspending agents include AVICEL RC-591 (from FMC Corporation of Philadelphia, PA) and sodium alginate. The amount of suspending agent(s) in a systemic or topical composition is typically about 1 to about 8%. Suitable surfactants include lecithin, Polysorbate 80, and sodium lauryl sulfate, and the TWEENS from Atlas Powder Company of Wilmington, Delaware. Suitable surfactants include those disclosed in the C.T.F.A. Cosmetic Ingredient Handbook, 1992, pp.587-592; Remington’s Pharmaceutical Sciences, 15th Ed.1975, pp.335–337; and McCutcheon’s Volume 1, Emulsifiers & Detergents, 1994, North American Edition, pp. 236–239. The amount of surfactant(s) in the systemic or topical composition is typically about 0.1% to about 5%. Although the amounts of components in the systemic compositions may vary depending on the type of systemic composition prepared, in general, systemic compositions include 0.01% to 50% of actives and 50% to 99.99% of one or more carriers. Compositions for parenteral administration typically include 0.1% to 10% of actives and 90% to 99.9% of a carrier including a diluent and a solvent. Compositions for oral administration can have various dosage forms. For example, solid forms include tablets, capsules, granules, and bulk powders. These oral dosage forms include a safe and effective amount, usually at least about 5%, and more particularly from about 25% to about 50% of actives. The oral dosage compositions include about 50% to about 95% of carriers, and more particularly, from about 50% to about 75%. Tablets can be compressed, tablet triturates, enteric-coated, sugar-coated, film-coated, or multiple-compressed. Tablets typically include an active component, and a carrier comprising ingredients selected from diluents, lubricants, binders, disintegrants, colorants, flavors, sweeteners, glidants, and combinations thereof. Specific diluents include calcium carbonate, sodium carbonate, mannitol, lactose, and cellulose. Specific binders include starch, gelatin, and sucrose. Specific disintegrants include alginic acid and croscarmellose. Specific lubricants include magnesium stearate, stearic acid, and talc. Specific colorants are the FD&C dyes, which can be added for appearance. Chewable tablets preferably contain sweeteners such as aspartame and saccharin, or flavors such as menthol, peppermint, fruit flavors, or a combination thereof. Capsules (including implants, time release and sustained release formulations) typically include an active and a carrier including one or more diluents disclosed above in a capsule comprising gelatin. Granules typically comprise an active, and preferably glidants such as silicon dioxide to improve flow characteristics. Implants can be of the biodegradable or the non- biodegradable type. The selection of ingredients in the carrier for oral compositions depends on secondary considerations like taste, cost, and shelf stability, which are not critical for the purposes of this invention. Solid compositions may be coated by conventional methods, typically with pH or time- dependent coatings, such that the hydrogel-encapsulated glucagon / glucagon analogue is released in the gastrointestinal tract in the vicinity of the desired application, or at various points and times to extend the desired action. The coatings typically include one or more components selected from the group consisting of cellulose acetate phthalate, polyvinyl acetate phthalate, hydroxypropyl methyl cellulose phthalate, ethyl cellulose, EUDRAGIT coatings (available from Rohm & Haas G.M.B.H. of Darmstadt, Germany), waxes and shellac. Compositions for oral administration can have liquid forms. For example, suitable liquid forms include aqueous solutions, emulsions, suspensions, solutions reconstituted from non- effervescent granules, suspensions reconstituted from non-effervescent granules, effervescent preparations reconstituted from effervescent granules, elixirs, tinctures, syrups, and the like. Liquid orally administered compositions typically include the hydrogel-encapsulated glucagon / glucagon analogue and a carrier, namely, a carrier selected from diluents, colorants, flavors, sweeteners, preservatives, solvents, suspending agents, and surfactants. Peroral liquid compositions preferably include one or more ingredients selected from colorants, flavors, and sweeteners. Other compositions useful for attaining systemic delivery of the subject compounds include sublingual, buccal and nasal dosage forms. Such compositions typically include one or more of soluble filler substances such as diluents including sucrose, sorbitol, and mannitol; and binders such as acacia, microcrystalline cellulose, carboxymethyl cellulose, and hydroxypropyl methylcellulose. Such compositions may further include lubricants, colorants, flavors, sweeteners, antioxidants, and glidants. The disclosed compositions can be topically administered. Topical compositions that can be applied locally to the skin may be in any form including solids, solutions, oils, creams, ointments, gels, lotions, shampoos, leave-on and rinse-out hair conditioners, milks, cleansers, moisturizers, sprays, skin patches, and the like. Topical compositions include: a disclosed hydrogel and a carrier. The carrier of the topical composition preferably aids penetration of the hydrogels into the skin. The carrier may further include one or more optional components. The amount of the carrier employed in conjunction with the hydrogel-encapsulated glucagon / glucagon analogue is sufficient to provide a practical quantity of composition for administration per unit dose of the medicament. Techniques and compositions for making dosage forms useful in the methods of this invention are described in the following references: Modern Pharmaceutics, Chapters 9 and 10, Banker & Rhodes, eds. (1979); Lieberman et al., Pharmaceutical Dosage Forms: Tablets (1981); and Ansel, Introduction to Pharmaceutical Dosage Forms, 2nded., (1976). A carrier may include a single ingredient or a combination of two or more ingredients. In the topical compositions, the carrier includes a topical carrier. Suitable topical carriers include one or more ingredients selected from phosphate buffered saline, isotonic water, deionized water, monofunctional alcohols, symmetrical alcohols, aloe vera gel, allantoin, glycerin, vitamin A and E oils, mineral oil, propylene glycol, PPG-2 myristyl propionate, dimethyl isosorbide, castor oil, combinations thereof, and the like. More particularly, carriers for skin applications include propylene glycol, dimethyl isosorbide, and water, and even more particularly, phosphate buffered saline, isotonic water, deionized water, monofunctional alcohols, and symmetrical alcohols. The carrier of a topical composition may further include one or more ingredients selected from emollients, propellants, solvents, humectants, thickeners, powders, fragrances, pigments, and preservatives, all of which are optional. Suitable emollients include stearyl alcohol, glyceryl monoricinoleate, glyceryl monostearate, propane-1,2-diol, butane-1,3-diol, mink oil, cetyl alcohol, isopropyl isostearate, stearic acid, isobutyl palmitate, isocetyl stearate, oleyl alcohol, isopropyl laurate, hexyl laurate, decyl oleate, octadecan-2-ol, isocetyl alcohol, cetyl palmitate, di-n-butyl sebacate, isopropyl myristate, isopropyl palmitate, isopropyl stearate, butyl stearate, polyethylene glycol, triethylene glycol, lanolin, sesame oil, coconut oil, arachis oil, castor oil, acetylated lanolin alcohols, petroleum, mineral oil, butyl myristate, isostearic acid, palmitic acid, isopropyl linoleate, lauryl lactate, myristyl lactate, decyl oleate, myristyl myristate, and combinations thereof. Specific emollients for skin include stearyl alcohol and polydimethylsiloxane. The amount of emollient(s) in a skin-based topical composition is typically about 5% to about 95%. Suitable propellants include propane, butane, isobutane, dimethyl ether, carbon dioxide, nitrous oxide, and combinations thereof. The amount of propellant(s) in a topical composition is typically about 0% to about 95%. Suitable solvents include water, ethyl alcohol, methylene chloride, isopropanol, castor oil, ethylene glycol monoethyl ether, diethylene glycol monobutyl ether, diethylene glycol monoethyl ether, dimethylsulfoxide, dimethyl formamide, tetrahydrofuran, and combinations thereof. Specific solvents include ethyl alcohol and homotopic alcohols. The amount of solvent(s) in a topical composition is typically about 0% to about 95%. Suitable humectants include glycerin, sorbitol, sodium 2-pyrrolidone-5-carboxylate, soluble collagen, dibutyl phthalate, gelatin, and combinations thereof. Specific humectants include glycerin. The amount of humectant(s) in a topical composition is typically 0% to 95%. The amount of thickener(s) in a topical composition is typically about 0% to about 95%. Suitable powders include beta-cyclodextrins, hydroxypropyl cyclodextrins, chalk, talc, fullers earth, kaolin, starch, gums, colloidal silicon dioxide, sodium polyacrylate, tetra alkyl ammonium smectites, trialkyl aryl ammonium smectites, chemically-modified magnesium aluminum silicate, organically- modified Montmorillonite clay, hydrated aluminum silicate, fumed silica, carboxyvinyl polymer, sodium carboxymethyl cellulose, ethylene glycol monostearate, and combinations thereof. The amount of powder(s) in a topical composition is typically 0% to 95%. The amount of fragrance in a topical composition is typically about 0% to about 0.5%, particularly, about 0.001% to about 0.1%. Suitable pH adjusting additives include HCl or NaOH in amounts sufficient to adjust the pH of a topical pharmaceutical composition. Methods of Treatment The disclosed compositions may be used to modulate glucose levels in a subject. The method may comprise administering a therapeutically effective amount of a pharmaceutical composition comprising glucagon and / or a glucagon analogue encapsulated within a hydrogel, as described herein, to a subject in need thereof. In various instances, the subject in need thereof may have an insulin disorder, such as diabetes. In various instances, the subject in need thereof may be at risk of experiencing a hypoglycemic event. In various instances, the subject in need thereof may be experiencing a hypoglycemic event. In some instances, when the subject is hypoglycemic, the glucagon and / or glucagon analogue may be released from the hydrogel, however, conversely, when the subject is normoglycemic or hyperglycemic, the glucagon and / or glucagon analogue will not be released from the hydrogel. In various instances, the therapeutically effective amount comprises 0.1–10 mg. In various instances, the therapeutically effective amount comprises 0.2–9.8 mg; 0.5–9.5 mg; 1.0– 9.0 mg; 2.0–8.0 mg; 3.0–7.0 mg; or 4.0–6.0 mg. In various instances, the therapeutically amount comprises no greater than 10 mg; no greater than 9.0 mg; no greater than 8.0 mg; no greater than 7.0 mg; no greater than 6.0 mg; no greater than 5.0 mg; no greater than 4.0 mg; no greater than 3.0 mg; no greater than 2.0 mg; no greater than 1.0 mg; no greater than 0.5 mg; or no greater than 0.2 mg; or no greater than 0.1 mg. In various instances, the therapeutically amount comprises no less than 0.1 mg; no less than 0.2 mg; no less than 0.5 mg; no less than 1.0 mg; no less than 2.0 mg; no less than 3.0 mg; no less than 4.0 mg; no less than 5.0 mg; no less than 6.0 mg; no less than 7.0 mg; no less than 8.0 mg; no less than 9.0 mg; or no less than 10 mg. In various instances, following administration of the pharmaceutical composition, the subject has blood glucose levels of about 20–600 mg / dL. In various instances, following administration of the pharmaceutical composition, the subject has blood glucose levels of about 30–500 mg / dL; about 40–400 mg / dL; about 50–300 mg / dL; about 60–200 mg / dL; or about 70– 200 mg / dL. In various instances, following administration of the pharmaceutical composition, the subject has blood glucose levels of no greater than about 600 mg / dL; no greater than about 500 mg / dL; no greater than about 400 mg / dL; no greater than about 300 mg / dL; no greater than about 200 mg / dL; no greater than about 100 mg / dL; no greater than about 90 mg / dL; no greater than about 80 mg / dL; no greater than about 70 mg / dL; no greater than about 60 mg / dL; no greater than about 50 mg / dL; or no greater than about 40 mg / dL. It will be apparent to one of ordinary skill in the relevant art that suitable modifications and adaptations to the compositions, formulations, methods, processes, and applications described herein can be made without departing from the scope of any embodiments or aspects thereof. The compositions and methods provided are exemplary and are not intended to limit the scope of any of the specified embodiments. All of the various embodiments, aspects, and options disclosed herein can be combined in any variations or iterations. The scope of the compositions, formulations, methods, and processes described herein include all actual or potential combinations of embodiments, aspects, options, examples, and preferences herein described. The exemplary compositions and formulations described herein may omit any component, substitute any component disclosed herein, or include any component disclosed elsewhere herein. The ratios of the mass of any component of any of the compositions or formulations disclosed herein to the mass of any other component in the formulation or to the total mass of the other components in the formulation are hereby disclosed as if they were expressly disclosed. Should the meaning of any terms in any of the patents or publications incorporated by reference conflict with the meaning of the terms used in this disclosure, the meanings of the terms or phrases in this disclosure are controlling. Furthermore, the foregoing discussion discloses and describes merely exemplary embodiments. All patents and publications cited herein are incorporated by reference herein for the specific teachings thereof. Various embodiments and aspects of the inventions described herein are summarized by the following clauses: Clause 1. A self-assembling peptide comprising: a boronic acid moiety attached to the N-terminus of a sequence of 5–12 amino acid residues, wherein: the terminal amino acid residues of the sequence are positively charged amino acid residues; the non-terminal amino acid residues of the sequence are hydrophobic amino acid residues; OH the C-terminus of the sequence; and the boronic acid moiety is a moiety of formula: an anion thereof, wherein: n i RX, a eac occurrence, s n epen en y ao, –1-4aoalkyl, –CN, –NO2, –C(O)H, –CO2H, –CO2C1-4alkyl, or –SO2C1-4alkyl; and X−is an anion having a net charge of −1. Clause 2. The self-assembling peptide of clause 1, wherein the self-assembling peptide is a peptide of formula (I): (I), wherein: B1is the boronic acid moiety; and is the sequence of amino acid residues, wherein: AA1and AA8are the terminal amino acid residues; AA2and AA6are aromatic amino acid residues; AA3, AA4, AA5and AA7are non-aromatic amino acid residues; and OH the C-. Clause 3. The self-1 AA is an arginine residue, a lysine residue, an ornithine residue, or a histidine residue. Clause 4. The self-assembling peptide of clause 2 or 3, wherein AA2is a phenylalanine residue, a tyrosine residue, or a tryptophan residue. Clause 5. The self-assembling peptide of any one of clauses 2–4, wherein AA3, AA4, and AA5are each independently a glycine residue, an alanine residue, a valine residue, an isoleucine residue, or a leucine residue. Clause 6. The self-assembling peptide of any one of clauses 2–5, wherein AA6is a tryptophan residue, a phenylalanine residue, or a tyrosine residue. Clause 7. The self-assembling peptide of any one of clauses 2–6, wherein AA7is a glycine residue, an alanine residue, a valine residue, an isoleucine residue, or a leucine residue. Clause 8. The self-assembling peptide of any one of clauses 2–7, wherein AA8is an arginine residue, a lysine residue, an ornithine residue, or a histidine residue. Clause 9. The self-assembling peptide of any one of clauses 2–8, wherein the peptide of formula (I) is a peptide of formula (I-a): a), R3, R4, and R5are each independently hydrogen, ; . H2Clause 10. The self-assembling peptide of clause 9, wherei . Clause 11. The self-assembling peptide of clause 9 or 10, wherein R2. Clause 12. The self-assembling peptide of any one of clauses 9–11, is . Clause 13. The self-assembling peptide of any one of clauses 9–12, wherein Clause 14. The self-assembling peptide of any one of clauses 9–13, wherein R5Clause 15. The self-assembling peptide of any one of clauses 9–14, . Clause 16. The self-assembling peptide of any one of clauses 9–15, wherein R7. Clause 17. The self-assembling peptide of any one of clauses 9–16, . Clause 18. The self-assembling peptide of any one of clauses 2–17, wherein B1is . Clause 19. The self-assembling peptide of clause 18, wherein B1. Clause 20. The self-assembling peptide of any one of clauses 1. Clause 21. The self-assembling peptide of any one of clauses 1–20, wherein RXis halo or –C1-4haloalkyl. Clause 22. The self-assembling peptide of clause 21, wherein RXis –F or –C1-4fluoroalkyl. Clause 23. The self-assembling peptide of any one of clauses 1–22, wherein the self- assembling peptide is a peptide of formula: . the self-assembling peptide of any one of clauses 1–23; and glucose. Clause 25. The hydrogel of clause 24, wherein glucagon and / or a glucagon analogue is encapsulated within the hydrogel. Clause 26. The hydrogel of clause 25, wherein the glucagon analogue comprises dasiglucagon and / or a depsi-glucagon analogue. Clause 27. The hydrogel of clause 25 or 26, wherein the glucagon and / or glucagon analogue is co-formulated with a cationic polymer within the hydrogel. Clause 28. The hydrogel of clause 27, wherein the cationic polymer is protamine. Clause 29. The hydrogel of any one of clauses 24–28, wherein the molar ratio of the self- assembling peptide of clause 1 to glucose is from about 1:100 to about 1:1000. Clause 30. A pharmaceutical composition comprising glucagon and / or a glucagon analogue encapsulated within the hydrogel of any one of clauses 24–29, and a pharmaceutically acceptable excipient. Clause 31. A method of modulating glucose levels in a subject, the method comprising: administering a therapeutically effective amount of the hydrogel of any one of clauses 24– 29 or the pharmaceutical composition of clause 30. Clause 32. The method of clause 31, wherein the subject in need thereof is experiencing a hypoglycemic event or at risk of experiencing a hypoglycemic event. Clause 33. The method of clause 31 or 32, wherein the subject in need thereof has diabetes. Clause 34. A kit comprising the hydrogel of any one of clauses 24–29 or the pharmaceutical composition of clause 30; delivery or administration apparata or devices; and optionally, packaging, a label, or instructions for use. Clause 35. Use of the hydrogel of any one of clauses 24–29 or the pharmaceutical composition of clause 30 to deliver glucagon and / or a glucagon analogue to a subject in need thereof. EXAMPLES Peptide Synthesis and Purification All multidomain proteins (MDPs) were synthesized by solid-phase methods using a CEM Liberty Blue synthesizer. Fmoc-protected amino acids and Rink amide resin (0.94 emq / g, 100- 200 mesh) were purchased from ChemImpex. Coupling reactions were performed using diisopropylcarbodiimide (DIC) and Oxyma in DMF with microwave heating. At the N-terminal positions, peptides were modified on resin using 4-carboxyphenylboronic acid pinacol ester (PBA–RFVAAWVK–NH2, “PBA–MDP”), 4-(tert-butoxycarbonyl)benzoic acid (COOH– RFVAAWVK–NH2, “COOH–MDP”), or 4-(tert- butoxy)benzoic acid (OH–RFVAAWVK–NH2, “OH– MDP”) by hand using standard coupling methods. Completed peptides were cleaved from resin by treatment with trifluoroacetic acid (TFA) / triisopropylsilane / H2O (95:2.5:2.5, v / v / v) for 2 h at room temperature. The cleavage liquid was then concentrated under a vacuum to remove most TFA and precipitated in cold diethyl ether. Crude material was collected by centrifugation, with excess ether being decanted and the remaining solid air-dried overnight. The crude peptide was then purified using by dissolving in hexafluoro-2-propanol (HFIP) at a concentration of 100–150 mg / mL and injecting onto a Biotage®Isolera flash chromatography system using a reversed-phase bio- C18cartridge (25 g) at a flow rate of 40 mL / min with a linear gradient from 0 to 100% (v / v) acetonitrile (+0.1% TFA) in water. Wavelengths of 220 and 280 nm were monitored for fraction collections, and collected fractions were then verified by electrospray ionization mass spectrometry (ESI-MS, Advion) and high-performance liquid chromatography (HPLC) using a C18Gemini (Phenomenex) column. Fractions with verified purity were combined and lyophilized, yielding a slightly yellowish powder product. The synthesis of dasiglucagon and a fluorescent variant (methoxycoumarin (MCA)-dasiglucagon) was performed according to previous reports. Table 1. Peptide Structures 4-carboxyphenylboronic acid pinacol ester–RFVAAWVK–NH2, “PBA–MDP” SEQ ID NO: 1 PBA–MDP was first dissolved in deionized (DI) water at a concentration of 3% (w / v) and adjusted to pH 7.4 using 0.1 M NaOH. Then an equal volume of 2× HEPES buffer (20 mM HEPES, 280 mM NaCl) containing different glucose concentrations was added for final glucose concentrations of 0, 25, 50, 100, 200 mg / dL and final peptide concentration of 1.5% (w / v). COOH– MDP and OH–MDP were prepared identically at a final glucose concentration of 0 mg / dL. Fourier Transform Infrared Spectroscopy (FTIR) Samples were prepared as described in Section 2.2 and lyophilized overnight. The dried powder was then analyzed using a Jasco FT / IR-6300 spectrometer. A background of air was subtracted for all spectra. Circular Dichroism (CD) Spectroscopy CD was collected using a Jasco J-1700 instrument. PBA–MDP were first prepared as reported in Section 2.2 at 1.5% w / v and diluted immediately before measurement by 10 times with 5 mM HEPES buffer at varying glucose concentration for a final peptide concentration of 0.15% w / v and glucose concentrations of 0, 25, 50, 100, and 200 mg / dL. COOH–MDP and OH–MDP samples were prepared identically, with a final glucose concentration of 0 mg / dL. To collect spectra, 50 μL of the diluted peptide solution was transferred to a quartz plate cuvette with a path length of 0.1 mm. Three spectra were collected (range of 250−190 nm, 50 nm / min scanning speed) and averaged for each sample. A background of 5 mM HEPES buffer was subtracted from all samples. Small-angle X-ray Scattering PBA–MDP samples prepared as noted in Section 2.2 at 1.5% w / v in HEPES buffer with a final glucose concentration of 0 or 100 mg / dL were subjected to small angle X-ray scattering with a SAXSLAB GANESHA 300-XL. CuKα radiation was generated by a Genix 3D Cu-source with an integrated monochromator, 3-pinhole collimation, and a two-dimensional Pilatus 300 K detector. The scattering intensity, q, was recorded at intervals of 0.012 < q < 0.3 Å−1(corresponding to lengths of 10–800 Å). Measurements were performed under vacuum at ambient temperature. The scattering curves were corrected for counting time and sample absorption. The PBA–MDP sample was sealed in thin-walled quartz capillaries about 1.5 mm in diameter and 0.01 mm wall thickness. The scattering spectra of the solvent were subtracted from the corresponding solution data using the Irena package for analysis of small-angle scattering data. Quantitative data analysis of KsEEKs was based on fitting its scattering curve to a model of spherical units packed into a helical structure, with aid from SASfit software. For complete details of this model, please refer to a prior report on its use to ascertain structure of related peptide nanocoil assemblies. Transmission Electron Microscopy (TEM) Peptide samples were prepared at 1.5% w / v as described in Section 2.2 and diluted 5 times in HEPES buffer immediately before TEM sample preparation. TEM samples were prepared by depositing a volume of 3 μL peptide solutions onto plasma-treated copper grids with a carbon film (Ted Pella, 200 mesh) for 45 s. After wicking away excess fluid and allowing the grid to dry in air, negative staining was applied using 2% uranyl acetate. TEM visualization was performed at an accelerating voltage of 120 kV (JOEL 2011). Cryogenic Transmission Electron Microscopy TEM at cryogenic temperature (Cryo-TEM) was used for direct imaging of solutions and dispersions. Vitrified specimens were prepared on a copper grid coated with a perforated lacey carbon 300 mesh (Ted Pella Inc.). A 3 μL drop from the solution was applied to the grid and blotted with filter paper to form a thin liquid film of solution. The blotted samples were immediately plunged into liquid ethane at its freezing point (−183 °C). The procedure was performed automatically in the Plunger (Lieca EM GP). The vitrified specimens were then transferred into liquid nitrogen for storage. The samples were studied using a FEI Talos F200C TEM, at 200 kV maintained at −180 °C; and images were recorded by FEI Ceta 16 M camera (4 k × 4 k CMOS sensor) at low dose conditions, to minimize electron beam radiation damage. Structural Cryo-EM The nanocoil solution was applied on a Quantifoil®holey carbon support grid. Excess sample was blotted away and the grid with thin film of solution was plunge-frozen in liquid ethane (−180 °C) using an EM GP Plunge Freezer (Leica). Cryo-EM data was collected on a 300 keV Titan Krios electron microscope having a K3 camera housed at University of Virginia. Each recorded movie had an accumulated exposure time of approximately 50 electrons per square angstrom (~50 e / Å2), while the pixel size of the images was ~1.08 Å. The data processing was performed in cryoSPARC (v4.2.1). Micrographs were preprocessed by “patch motion correction” and “patch CTF estimation” jobs for motion correction and contrast transfer function (CTF) estimation, respectively. The nanocoil segments were picked manually by “manual picker” from several micrographs to generate the initial template for further automated picking. The “helical tracer” was used for automated picking from whole dataset. Bad picks were removed by “inspect pick” and “2D classification” jobs. The averaged power spectrum of the aligned nanocoil segments was generated by the job “Average Power Spectra” while the 3D reconstruction was done by the “Helix Refine” function. The structure of PBA–MDP was predicted by AlphaFold, manually fit in the density using COOT, and used for the model-to-map fit representation. Atomic Force Microscopy (AFM) AFM samples were prepared by depositing a drop of peptide solution (0.2% w / v in pH 7.4 HEPES buffer) on a mica surface. AFM was performed in liquid non-contact mode using a Park XE7 instrument and a cantilever with a spring constant of 2.8 N / m (FMR 10 M, Park Systems). Zeta Potential Zeta potential was measured using a Malvern Zetasizer. PBA–MDP solutions at a concentration of 0.2% (w / v) were prepared in pH 7.4 HEPES buffer at glucose concentrations of 0, 25, 50, 100, and 200 mg / dL. COOH-MPD and OH–MDP were prepared the same as PBA– MDP at a concentration of 0.2% (w / v) and a glucose concentration of 0 mg / dL. For all samples, an 800 μL sample was added into a clear folded cuvette and evaluated by using the Huckel model at 25 °C. Each sample was prepared and measured in triplicate. Thioflavin T (ThT) assay PBA–MDP was dissolved in DI water at a concentration of 0.4% (w / v) and adjusted to pH 7.4 by 0.1 M NaOH. An aliquot of 97 μL of the peptide solution was mixed with 97 μL of 2× HEPES buffer (pH 7.4, 20 mM HEPES and 280mM NaCl), 2 μL of a 10 mM ThT stock solution, and 2 μL of HEPES buffer containing 0 or 100,000 mg / dL glucose, for a final glucose concentration of 0 or 100 mg / dL, respectively. Fluorescence intensity was serially measured on a Tecan M200 microplate reader at excitation of 485 nm and emission of 528 nm with 5 min intervals. After incubating for 4 h, 2 μL of 101,000 mg / dL glucose in 1× HEPES buffer was added into the 0 mg / dL glucose sample to rapidly raise its final glucose concentration to 100 mg / dL. For the sample initially prepared at 100 mg / dL glucose sample, 2 μL of 1× HEPES buffer was added. Fluorescence intensity was recorded every 5 min for another 36 h. A background of ThT in pH 7.4 HEPES buffer was subtracted from all spectra. Rheological Characterization To evaluate the mechanical properties of PBA–MDP hydrogels, dynamic oscillatory rheology was performed on a TA Instruments Discovery HR-2 rheometer fitted with a Peltier stage using a parallel plate geometry with a diameter of 25 mm. Samples at 1.5% (w / v) were prepared as described above at glucose concentrations of 0, 25, 50, 100, and 200 mg / dL and incubated for 45 min before the measurement. An amplitude sweep was performed to determine the linear viscoelastic range. Then a frequency sweep was performed at a constant strain (1% strain). To evaluate the time course of gelation, a time sweep was performed immediately after sample preparation (1% strain, 10 rad / s). A step-strain cycling study was also performed on 1.5% (w / v) PBA–MDP hydrogels prepared at 100 mg / dL glucose, cycling between 1% strain for 2 min and 100% strain for 30 s at an angular frequency of 10 rad / s. To assess shear-thinning, PBA–MDPs prepared at glucose concentrations of 0 and 100 mg / dL were studied under steady shear flow over a shear rate range of 0.001 to 10 s−1. In vitro Glucagon Release PBA–MDP hydrogels (1.5% w / v) were first prepared at a glucose concentration of 100 mg / dL, as described above. Mixed into each 50 μL was added a pre-formed complex of 0.1 mg MCA-dasiglucagon and 0.05 mg protamine. Hydrogels were incubated for 45 min after preparation in 6-well plates, after which 2 mL of pH 7.4 HEPES buffer that contained 0, 25, 50, 100, and 200 mg / dL glucose was added. As a control, COOH–MDP was prepared, loaded with the MCA-dasiglucagon and protamine complex, and tested for release in a glucose-free buffer. At each time point, a 20 μL solution was taken and diluted 10 times for fluorescence analysis (Ex: 322 nm, Em: 398 nm). The MCA-dasiglucagon concentration of each sample was determined using a standard curve. After the measurement, 20 μL of fresh buffer containing the corresponding glucose concentration was added to maintain constant bulk buffer volume. In vivo hypoglycemia protection. Using a previously established hypoglycemia mouse model, the prophylactic (preventative) use of this material was explored for its ability to reduce the severity of a sudden hypoglycemic onset. Briefly, male C57BL6 / J mice, aged 8 weeks, were induced diabetic by streptozotocin (STZ) dosed at 150 mg / kg i.p. Once non-fasted blood glucose (BG) levels were 600+ mg / dL, 7–8 days after STZ treatment, the study was initiated. After fasting for 8 h, mice with BG > 450 mg / dL were dosed with 0.5 IU / kg basal insulin detemir (Levemir, Novo Nordisk) via subcutaneous (s.c.) injection in a total volume of ~100 μL. After 4 h BG levels were corrected to within a normal range (~180 mg / dL) and mice were randomly divided into 4 groups (n = 9–10 per group) and treated with buffer, dasiglucagon (10 μg), PBA–MDP hydrogel loaded with 10 μg dasiglucagon, or COOH–MDP hydrogel loaded with 10 μg dasiglucagon in 100 μL total volume injected s.c.. BG levels were monitored after the treatment (t = 0 min). To trigger hypoglycemia, AOF recombinant human insulin (Gibco) was administered 2 h following treatment i.p. at a dose of ~3 IU / kg in saline. BG levels were monitored for another 4 h. Throughout the experiment, mice exhibiting “high” readings were noted with a BG value of 600 mg / dL, while “low” readings were noted as a BG value of 20 mg / dL and indicated as dead. These studies were detailed in a protocol approved by the University of Notre Dame Animal Care and Use Committee and adhered to all relevant Institutional, State, and Federal guidelines. Statistical testing between treatment and control groups was performed using one-way ANOVA with Tukey multiple comparison post hoc testing (GraphPad Prism v9.0). Multidomain Peptide Design The general design of the PBA–MDP molecule (FIG.1, FIG.5) consisted of eight amino acid residues functionalized with a PBA motif at the N-terminus (PBA–RFVAAWVK). This design was inspired by prior reports of MDPs that note a criteria of ABA oligopeptide patterning, wherein two charged / hydrophilic “A” blocks flank a hydrophobic “B” block. However, in addition to the inclusion of the prosthetic PBA motif, the design here differed from typical MDP designs by having only a single cationic amino acid for the “A” blocks, and also broke from the typical approach in its asymmetric sequence. The design rationale for each component was as follows: i) cationic residues at the N- and C-terminal ends consisting of an arginine (R) and lysine (K), respectively, serve as charge-bearing units and enhance solubility; ii) a hydrophobic “B” segment that included aromatic phenylalanine (F) and tryptophan (W) residues shielding an internal β-sheet forming region of valine (V) and alanine (A) residues; and iii) a glucose-binding PBA motif intended to shift the net charge on the N-terminal domain of the PBA–MDP as a function of glucose level. The particular amide-linked PBA motif chosen would be expected to have a pKaof ~8.4 on its own. While this would not typically be effective for glucose-binding function under conditions of physiological pH, its positioning next to the cationic arginine residue was envisioned to significantly reduce its effective pKa, a phenomenon also observed in PBA-modified cationic polymers. In addition to PBA–MDP, two control molecules were prepared to capture the general structure and range of charge states of the prosthetic PBA motif. The first (COOH–MDP) was designed with a benzoic acid group in place of the boronate (FIG. 1, FIG. 5). The pKaof this benzoic acid group is expected to be ~4, meaning it would be fully charged at neutral pH conditions used in these experiments. The second (OH–MDP) was designed with a phenol group in place of the boronate (FIG.1, FIG. 5). As the pKaof this phenol is expected to be ~10, the motif should be uncharged at neutral pH conditions used in these experiments. It is possible that proximity to arginine could likewise shift pKaof the phenol, as postulated for the PBA motif. It is also important to note that the expected pKaof amino acids and prosthetic motifs used to construct all of the MDPs here could shift as a result of their position on the peptide and / or due to aggregation-induced pKashifts. Efforts to quantify pKavalues for the various ionizable groups in these peptides via titration did not yield precise measurements and instead suggested a range of charged states, likely due to a broadened charge distribution resulting from peptide assembly. Nanocoil Assembly Given the design rationale for PBA–MDP, it was of interest to first study its propensity for self-assembly into nanostructures, as is commonly observed for other classes of MDPs and related oligopeptide motifs. When samples at 0.2% w / v were examined using cryogenic transmission electron microscopy (cryoTEM), the PBA–MDP was observed to self-assemble into relatively short and small nanocoils (FIG.2A). This nanocoil structure, resembling a telephone cord, is uncommon in MDPs and related oligopeptide assemblies formed from β-sheet segments, which instead more commonly form high aspect-ratio cylindrical micelles or ribbon-like filaments. To date, the formation of nanocoils of this type has only been observed from the self-assembly of tetrapeptides functionalized at both termini with S-aroylthiooximes for use as H2S donors. Upon addition of glucose at a concentration of 100 mg / dL, recreating normal glucose levels in the human body, the nanocoil structures were retained though appeared longer with a higher extent of nanostructure bundling (FIG.2B). The lengths of nanocoils in these two states were quantified by measuring every particle visible in four separate cryoTEM images collected from each glucose condition, revealing lengths in the case of PBA–MDP prepared at 100 mg / dL glucose (122 ± 63 nm, mean ± SD) that were significantly longer (P < 0.0001, Student’s t-test) than in 0 mg / dL glucose (77 ± 46 nm). Apart from differences in length, the nanocoil morphology was consistent over a range of different glucose levels (FIG.6). Solution-phase AFM was also performed (FIG. 2C), revealing nanocoil structures with features matching those seen by cryoTEM. TEM analysis of the control molecules showed nanocoils for the COOH–MDP molecule (length of 127 ± 45 nm from cryoTEM images) that were similar to those observed for PBA–MDP (FIG. 7), while OH– MDP formed anomalous small aggregates (FIG. 8). Accordingly, the formation of nanocoils seemingly depends on the availability of a negative charge at the position where the PBA has been inserted, thereby rendering the N-terminal end of the molecule effectively neutral. This data from the two control MDPs also suggests that the boronates on PBA–MDP are at least partially in their negatively charged conformation even in the absence of glucose, confirming expectations that the adjacent arginine residue reduces the effective pKaof the boronate. These structures were further characterized using small angle x-ray scattering (SAXS) (FIG.2D), with data fit to a model of spherical objects packed into a helical structure. This is the same model used to analyze SAXS data from the only other reported peptide-based nanocoil assembly. In the absence of glucose, PBA–MDP nanocoils had a coil diameter of 47 Å with a helical pitch of 126 Å and length of 625 Å. Upon addition of glucose at a concentration of 100 mg / dL, the fitted coil diameter (49 Å) and helical pitch (124 Å) did not change considerably, but the nanocoil length increased by ~4-fold (2790 Å). It is noted that PBAs can form a bisdentate complex with glucose – two PBAs bound to different sites on the same glucose molecule – when in its furanose form, though the second binding event is less favorable in part due to a low fraction of glucose in this anomeric form. As no structural changes to the nanocoils were observed upon addition of glucose, apart from their length, it is not anticipated that bisdentate interactions are readily occurring in PBA–MDP assemblies. The observation of this uncommon nanocoil morphology inspired additional studies using high-resolution structural analysis with cryoEM to uncover the mode of molecular packing. The average power spectrum of aligned nanocoil segments revealed the typical pattern for a helical assembly (FIG.2E). Four layer lines were visible in the power spectrum, which corresponded to four orders of a first layer line at ~1 / (123 Å) arising from a nanocoil pitch of ~123 Å. This matches well with observations from SAXS. The ring at ~1 / (4.8 Å) in the power spectrum appears to arise from the unpolymerized peptides in the background. Since the spacing of β-strands in a β-sheet is ~4.8 Å, PBA–MDP likely exists in the background as dimers of two PBA–MDP monomers in either a parallel or antiparallel state. An unambiguous determination of the helical rise of the subunits in the nanocoil was not possible due to the lack of high-resolution information in the average power spectrum. Amongst many tested possible symmetries, a twist / rise of 15.7° / 5.1 Å was found to give the best density map, showing separation of β-strands with reasonable density for bulky side chains of the residues (FIG.2F). In this configuration, twenty-four PBA–MDP dimers are aligned roughly parallel to the helical axis with an axial rise of 5.1 Å to generate a nanocoil with ~123 Å pitch. Impact of Glucose Binding Results from TEM and SAXS showed limited impact from glucose binding on nanocoil morphology, though the addition of glucose led to an increase in the length and apparent bundling of these nanocoils. As part of the original design rationale, the central region was intended to drive β-sheet hydrogen bonding; though envisioned as a fibrilizing domain, the close head-to-tail stacking of the monomers revealed through structural elucidation of the nanocoils by cryoEM could still allow for H-bonding of this domain. Indeed, a β-sheet dimer is likely the building block for the eventual nanocoil assemblies. Circular dichroism (CD) spectroscopy was first performed on PBA–MDP over a range of glucose concentrations from 0–200 mg / dL (FIG.2G). Though these spectra did not conform to the typical canonical secondary structures of a β-sheet, the emergence of a small negative peak at 223 nm and a relatively larger negative peak at 201 nm were generally consistent with predictions for a right-twisted antiparallel β-sheet; red-shifting of these minima as glucose concentration increased suggests glucose acts to increase twisting of these antiparallel interactions. This interpretation of CD data is further supported by the characteristic β-sheet spacing of ~4.8 Å revealed in structural cryoEM studies. Though TEM and SAXS demonstrate that the nanocoil morphology does not change upon addition of glucose, it is postulated that these glucose-dependent changes in secondary structure present in CD arise from the antiparallel β- sheet dimers observed in the background of cryoEM studies being recruited to participate in nanocoil assemblies when glucose is present. Both control molecules COOH–MDP and OH– MDP generally revealed similar secondary structure signatures by CD, with signal intensity increased for the COOH–MDP variant. It is likewise noted that the inclusion of aromatic amino acids on self-assembling peptides can confound interpretation of CD as a result of π-π* and n-π* transitions present specifically in the spectral region of interest here. Given the uncharacteristic nature of these peaks relative to typical self-assembling oligopeptides, Fourier Transform Infrared Spectroscopy (FTIR) was next performed to further characterize stretches in the Amide I region as a means of assessing peptide secondary structure (FIG.2H). A major peak was observed at 1627 cm−1with a minor peak at 1693 cm−1; these peaks in the Amide I region correspond to those reported for a model peptide that is known to form antiparallel β-sheets. These signatures were enhanced by the addition of glucose, supporting an increased propensity to form antiparallel β-sheets for PBA–MDP in its glucose-bound state. Control molecules demonstrated the impact of prosthetic charge on this β-sheet formation as well, with the uncharged prosthetic of the OH–MDP leading to limited antiparallel β-sheet signal in this region while the charged prosthetic of the COOH–MDP corresponded to a very strong signal. Accordingly, the PBA motif is expected to become increasingly charged as glucose level increases, a known outcome of PBA motifs wherein the equilibrium shifts to the charged tetrahedral boronate in water upon glucose binding. The presence of a negative prosthetic charge at this position, and thus a more neutralized N-terminal domain, appears to promote enhanced antiparallel β-sheet formation. However, given microscopy results showing nanocoils from PBA– MDP in both the presence and absence of glucose, the existence of well-ordered antiparallel β- sheet interactions is seemingly not essential to nanocoil formation though may have implications on the overall increase in nanocoil length through internal stabilization. Moreover, the CD signal of an antiparallel β-sheet in all samples irrespective of glucose levels could also arise from H- bonding of PBA–MDP dimers existing in equilibrium with the nanocoil structures that are accentuated in the presence of glucose. Indeed, the likely presence of a background of peptide dimers spaced by ~4.8 Å revealed in structural cryoEM support this hypothesis. To further verify the glucose-dependent emergence of β-sheet structures, thioflavin T (ThT) was used (FIG.2I). The fluorescence of this dye increases when embedded in β-sheet- rich domains, offering a means to sense and monitor amyloid structures. When PBA–MDP was prepared in the presence of 100 mg / dL glucose, the ThT fluorescence showed a steady increase over a period of ~20 h. When PBA–MDP was prepared in the absence of glucose, limited ThT fluorescence was observed. However, when this sample was transferred to 100 mg / dL glucose at 4 h after preparation, the ThT fluorescence began to increase immediately upon introduction of glucose. Taken together with results from CD and FTIR, these data support a role for glucose binding in triggering increased antiparallel β-sheet ordering in PBA–MDP assemblies. It is worth reiterating that TEM and SAXS data demonstrate the existence of these β-sheet structures is not essential to the formation of the nanocoil assemblies, though a greater antiparallel β-sheet propensity along with recruitment of antiparallel β-sheet dimers to extending nanocoils could underlie the increase in nanocoil length observed by TEM and SAXS in the presence of glucose. The original design rationale was that glucose binding to the prosthetic PBA motif would stabilize a negative charge on the boronate and make the N-terminal end of the molecule net- neutral. Accordingly, Zeta potential was also measured for PBA–MDP as a function of glucose (FIG. 2J). At pH 7.4, it is expected that both the arginine and lysine residues would be fully charged, while the boronate would likely exist in both its negatively charged tetrahedral configuration and uncharged trigonal form; the extent to which the neighboring arginine residue lowers the pKaof this boronate from its nominal value of ~8.4 is not clear. Thus, the molecule would be expected to be net-positive overall, with the magnitude of its charge state depending on the extent to which the boronate was charged. In the absence of glucose, PBA–MDP had a substantial positive zeta potential of ~29.3 mV. The magnitude of this positive zeta potential decreased corresponding to an increase in glucose up to 200 mg / dL, wherein the zeta potential was reduced to ~12.6 mV. As a control, the uncharged prosthetic of OH–MDP, with a nominal net charge of +2 for the overall molecule, had the most positive zeta potential of ~31.8 mV. Meanwhile, the negatively charged prosthetic of the COOH–MDP, with a nominal net charge of +1, had the least positive zeta potential of ~5.5 mV. These data support a gradient in the charge state of the PBA motif on the PBA–MDP, becoming increasingly negative as glucose binding shifts the equilibrium to its tetrahedral configuration. These data likely also support a role for charge screening in the increased nanocoil length and extent of bundling observed in cryoTEM for PBA– MDP as glucose level is increased. Glucose-Driven Hydrogelation When PBA–MDP was prepared at concentrations of 1.5% w / v in the presence of 100 mg / dL glucose, a self-supporting hydrogel was observed (FIG.3A). This observation was further evaluated over a range of glucose concentration from 0 to 200 mg / dL. Over a period of minutes following preparation, samples with 100 mg / dL and 200 mg / dL glucose formed translucent self- supporting hydrogels. Meanwhile, samples prepared at lower glucose levels (0, 25, 50 mg / dL) remained translucent but were not able to be inverted to demonstrate self-supporting character. Accordingly, by gross macroscopic inspection, the addition of glucose promotes hydrogelation of solutions of PBA–MDP. To further support these observations for steady gelation upon addition of glucose, a rheological time-course study was next performed (FIG.3B). Samples of PBA–MDP were freshly prepared immediately before measurements at 1.5% w / v in pH 7.4 HEPES buffer and placed onto the rheometer stage. The initial storage modulus (G′) and complex viscosity (η*) were similar in both conditions (G′ = ~23 Pa and η* = ~5 Pa·s). In the sample prepared with 100 mg / dL glucose, G′ and η* increased rapidly over the first 10 min to ~240 Pa and ~48 Pa·s, respectively. Further measurements over the next 35 min demonstrated a plateau in G′ = 507 Pa and η* = 102 Pa·s. Comparatively, the PBA–MDP sol prepared with no glucose, G′ and η* stayed relatively stable with only a small increase in G′ = ~41.3 Pa and η* = ~9 Pa·s. These data support a role for glucose in promoting hydrogelation of PBA–MDP, possibly resulting from the overall increase in nanocoil length and extent of bundling that was observed in TEM studies performed in the presence of glucose. The effect of glucose concentration on gelation was further studied in samples brought to steady-state over 45 min in various glucose concentrations (0, 25, 50, 100, and 200 mg / dL) prior to rheological measurements. G′ and η* both increased by almost 2 orders of magnitude with an increase in glucose concentration, from G′ = 16.7 Pa and η* = 3.4 Pa·s at 0 mg / dL to G’ = 1009.8 Pa and η* = 203.3 Pa·s at 100 mg / dL (FIG.3C). With the further increase in glucose to 200 mg / dL glucose, negligible additional increase was observed for G′ or η*. A large increase in G′ occurred for an increase in glucose level from 50 mg / dL (G′ = 165.2 Pa) to 100 mg / dL (G′ = 1009.8 Pa); importantly this range in glucose concentration corresponds to a low to normal blood glucose level, suggesting the possibility for function in physiologically relevant conditions. The trends in gelation and viscosity as a function of glucose align with observations from cryoTEM and SAXS pointing to a greater extent of nanocoil bundling and entanglement and longer nanocoil structures upon addition of glucose. These measurements performed on bulk gels aged in vials result in higher measured G′ values than when gels were aged on the rheometer, likely resulting from being aged in the absence of continuous deformation. COOH–MDP also formed hydrogels at 1.5% w / v; rheology on samples prepared and equilibrated overnight revealed these to be stiffer with a higher complex viscosity (FIG.9) than PBA–MDP hydrogels, further supporting increased intermolecular interactions and reduced electrostatic repulsion for COOH–MDP that was observed by CD, FTIR, and zeta potential measurements. Toward injection-relevant applications, the PBA–MDP hydrogel should be shear-thinning and self-healing, allowing it to pass through a needle under shear and immediately reform its gel structure once in situ. A hydrogel prepared in 100 mg / dL glucose exhibited reduced viscosity of over three orders of magnitude from a “zero-shear” level of ~1400 Pa·s to <1 Pa·s at a shear rate of 10 s−1 (FIG.3D). Another crucial property of an injectable hydrogels is its ability to self-heal following exposure to high strain. To probe this feature, the PBA–MDP hydrogel prepared with 100 mg / dL glucose was thus exposed to cyclic strain between 1% and 100% at a constant frequency of 10 rad / s (FIG.3E). Mechanical properties of the hydrogel recovered immediately after cessation of high strain, a process repeated for multiple cycles. Glucose-Responsive Drug Delivery Evidence for glucose binding-induced formation and stabilization of PBA–MDP nanocoil hydrogels supported their further exploration in the context of glucagon therapy. As an antagonist to insulin, glucagon functions in normal physiology to reverse low blood glucose (hypoglycemia). Commonly only used in the context of emergency rescue, the integration of glucose-responsive glucagon delivery may also enable enhanced precision in insulin-centered blood glucose control by mitigating the risks of hypoglycemia. An ideal glucagon delivery approach should thus exhibit release and / or activity inversely related to blood glucose level. As an active glucagon payload, dasiglucagon was selected for its improved solubility and stabilized secondary structure relative to native glucagon. To first monitor release as a function of ambient glucose level, a fluorescent variant (MCA-dasiglucagon) was used. MCA-dasiglucagon alone had rapid release that was effectively independent of glucose (FIG.10), likely due to the relatively small payload compared to the mesh size of the nanocoil network. Thus, MCA-dasiglucagon was instead co-formulated with protamine, an arginine-rich polypeptide, to increase the hydrodynamic size of the payload and limit its leakage from the hydrogels. Protamine is also used in complex with insulin in the long-used clinical NPH Insulin product. Release profiles of dasiglucagon following its complexation with protamine revealed an inverse glucose dependence in the release profiles (FIG.4A). Fitting to a standard first-order model demonstrated decreased rates of release and plateau values when glucose was increased over a range from 0 to 200 mg / dL. It is noted that the control hydrogel of COOH–MDP exhibited a similar release behavior as the PBA–MDP hydrogel at a glucose concentration of 200 mg / dL and, when taken together with material characterization data, supports a role for glucose binding-induced neutralization of the peptide N- terminal domain in nanocoil stabilization, entanglement, and gelation, thereby slowing dasiglucagon release. Finally, glucose-induced stability and increased retention of dasiglucagon was explored in a previously established model of prophylactic delivery to prevent a subsequent hypoglycemic emergency (FIG. 4B). Streptozotocin (STZ) was used to induce diabetes in mice; following a stable hyperglycemic phenotype, mice were fasted overnight and then administered the clinically used Insulin Detemir to induce a state of normoglycemia. Four hours after administering Insulin Detemir (t = 0), control mice were treated with either buffer or 0.01 mg dasiglucagon while experimental groups were administered 0.01 mg dasiglucagon encapsulated in 50 μL hydrogels of either PBA–MDP or COOH–MDP. After two additional hours, mice were overdosed by injection of 3 IU / kg recombinant human insulin. Blood glucose levels were monitored throughout the procedure (FIG. 4C). A threshold for hypoglycemia, set nominally at 60 mg / dL, was used to monitor the severity of hyperglycemic response following insulin overdose. Among the four groups, the lowest average blood glucose level observed (nadir value, FIG.4D) was above this hypoglycemic threshold of 60 mg / dL for only the mice treated with the PBA–MDP hydrogel containing dasiglucagon (74 ± 8 mg / dL). The buffer-treated (39 ± 7 mg / dL) and dasiglucagon- treated (40 ± 5 mg / dL) groups were both significantly less (P < 0.01) than the group treated with the PBA–MDP hydrogel. The lack of function for dasiglucagon administered 2 h prior to insulin overdose is not surprising given the relatively short half-life of the hormone, and also is supported by previous studies in this same model. The glucose-responsive function of the hydrogel was further supported by the performance of the COOH–MDP (49 ± 10 mg / dL), which resulting in a nadir value significantly lower (P < 0.05) than PBA–MDP and demonstrates function from glucose- independent controlled release. In addition to a prevention of low blood glucose levels, the PBA– MDP hydrogel treatment offered protection against onset of severe hypoglycemia; there were no overdose-related deaths in the PBA–MDP group (0 / 9) compared to groups treated with the COOH–MDP hydrogel (1 / 10), buffer (4 / 9), and dasiglucagon (2 / 9). The recovery from hypoglycemia was also evaluated at the endpoint of the study (FIG.4E). Mice treated with PBA– MDP hydrogel demonstrated significantly better recovery (148 ± 22 mg / dL) than mice treated with COOH–MDP hydrogel (83 ± 11 mg / dL, P < 0.05), buffer (63 ± 14 mg / dL, P < 0.01), and dasiglucagon alone (62 ± 10 mg / dL, P < 0.01). Accordingly, the glucose-directed retention and release of dasiglucagon from within the PBA–MDP hydrogels offered advantages as a prophylactic (protective) measure against a subsequent sudden hypoglycemic episode brought on by insulin overdose. The performance of this approach to deliver dasiglucagon is likewise comparable in this same model to an orthogonal approach that used enzymatic actuation by the glucose oxidase enzyme to endow self-assembled peptide hydrogels with glucose-driven stability and afford dasiglucagon release under states of low glucose. The xenogeneic origin of glucose oxidase, along with its toxic hydrogen peroxide byproduct, may limit the therapeutic utility of this prior work in the therapeutic context of a routine glucagon prophylactic. By comparison, the approach presented here using a single-component system of glucose-binding nanocoils may offer distinct advantages. Namely, the current system does not require a xenogeneic enzyme with immunogenic risk, nor does it entail its toxic H2O2, instead achieving its function through a discrete small molecule assembly process including a synthetic PBA glucose binder. Certain drawbacks of this prior system, including undesirable glucagon leakage after injection and lower responsibility than would be desirable in emergency- use contexts, remain challenges in the PBA–MDP platform as well.

Claims

CLAIMS What is claimed:

1. A self-assembling peptide comprising: a boronic acid moiety attached to the N-terminus of a sequence of 5–12 amino acid residues, wherein: the terminal amino acid residues of the sequence are positively charged amino acid residues; the non-terminal amino acid residues of the sequence are hydrophobic amino acid residues; OH the C-terminus of the sequence is; and the boronic acid moiety is a moiety of formula:nRX, at each occurrence, is independently halo, –C1-4haloalkyl, –CN, –NO2, –C(O)H, –CO2H, –CO2C1-4alkyl, or –SO2C1-4alkyl; and X−is an anion having a net charge of −1.

2. The self-assembling peptide of claim 1, wherein the self-assembling peptide is a peptide of formula (I): (I), wherein: B1is the boronic acid moiety; and is the sequence of amino acid residues, wherein: AA1and AA8are the terminal amino acid residues; AA2and AA6are aromatic amino acid residues; AA3, AA4, AA5and AA7are non-aromatic amino acid residues; andOH the C-.

3. The self-assemblingis an arginine residue, a lysine residue, an ornithine residue, or a histidine residue.

4. The self-assembling peptide of claim 2, wherein AA2is a phenylalanine residue, a tyrosine residue, or a tryptophan residue.

5. The self-assembling peptide of claim 2, wherein AA3, AA4, and AA5are each independently a glycine residue, an alanine residue, a valine residue, an isoleucine residue, or a leucine residue.

6. The self-assembling peptide of claim 2, wherein AA6is a tryptophan residue, a phenylalanine residue, or a tyrosine residue.

7. The self-assembling peptide of claim 2, wherein AA7is a glycine residue, an alanine residue, a valine residue, an isoleucine residue, or a leucine residue.

8. The self-assembling peptide of claim 2, wherein AA8is an arginine residue, a lysine residue, an ornithine residue, or a histidine residue.

9. The self-assembling peptide of claim 2, wherein the peptide of formula (I) is a peptide of formula (I-a): a),; R3, R4, and R5are each independently hydrogen, ;.

10. The self-assembling peptide of claim 9, .

11. The self-assembling peptide of claim 9, wherein R2.

12. The self-assembling peptide of claim 9, wherein R3is .

13. The self-assembling peptide of claim 9, wherein R414. The self-assembling peptide of claim 9, wherein R5is methyl.

15. The self-assembling peptide of claim 9, .

16. The self-assembling peptide of claim 9, wherein R7is .

17. The self-assembling peptide of claim 9, .

18. The self-assembling peptide of claim 2, .

19. The self-assembling peptide of claim 18, .

20. The self-assembling peptide of claim 2, wherein n is 0 or 1.

21. The self-assembling peptide of claim 2, wherein RXis halo or –C1-4haloalkyl.

22. The self-assembling peptide of claim 21, wherein RXis –F or –C1-4fluoroalkyl.

23. The self-assembling peptide of claim 1, wherein the self-assembling peptide is a peptide of formula: .

24. A hydrogel comprising: the self-assembling peptide of claim 1; and glucose.

25. The hydrogel of claim 24, wherein glucagon and / or a glucagon analogue is encapsulated within the hydrogel.

26. The hydrogel of claim 25, wherein the glucagon analogue comprises dasiglucagon and / or a depsi-glucagon analogue.

27. The hydrogel of claim 25, wherein the glucagon and / or glucagon analogue is co- formulated with a cationic polymer within the hydrogel.

28. The hydrogel of claim 27, wherein the cationic polymer is protamine.

29. The hydrogel of claim 24, wherein the molar ratio of the self-assembling peptide of claim 1 to glucose is from about 1:100 to about 1:1000.

30. A pharmaceutical composition comprising glucagon and / or a glucagon analogue encapsulated within the hydrogel of claim 24, and a pharmaceutically acceptable excipient.

31. A method of modulating glucose levels in a subject, the method comprising: administering a therapeutically effective amount of the hydrogel of claim 24 or the pharmaceutical composition of claim 30.

32. The method of claim 31, wherein the subject in need thereof is experiencing a hypoglycemic event or at risk of experiencing a hypoglycemic event.

33. The method of claim 31, wherein the subject in need thereof has diabetes.

34. A kit comprising the hydrogel of claim 24 or the pharmaceutical composition of claim 30; delivery or administration apparata or devices; and optionally, packaging, a label, or instructions for use.

35. Use of the hydrogel of claim 24 or the pharmaceutical composition of claim 30 to deliver glucagon and / or a glucagon analogue to a subject in need thereof.

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