The application is a divisional application of a patent application with the application number of 201280073745.X, the application date of 2012, 12 and 5, and the invention name of "medical organogel method and composition".
This patent application claims priority to U.S. serial No. 61/566,768, filed on 5.12.2011, which is hereby incorporated by reference herein.
Detailed Description
Embodiments of the invention are xerogels comprising a protein powder or other water-soluble biologic powder dispersed in a matrix of the xerogel. The xerogel may be hydrated at the point of use and placed in tissue where it controllably releases the protein over time. The powder contains fine particles of protein. The xerogel matrix, when hydrated, is a hydrogel made from a cross-linked matrix. The protein is in the solid phase and is substantially insoluble until the matrix begins to erode, thereby allowing the protein to go into solution. The matrix protects the protein from cells, enzymatic denaturation, and unwanted local reactions. The protein is in a substantially solid phase until released by gradual solvation (solubilization), and is thus protected from denaturation, autohydrolysis, proteolysis, and local chemical reactions, which may result in loss of potency or antigenicity.
Fig. 1A depicts an embodiment of the method, which starts with a protein particle 100, the protein particle 100 being prepared by conventional means to preserve the secondary structure and, if present, the tertiary or quaternary structure of the protein. These are combined with the precursors 102, 104 into an organic solvent 106. The mixture is processed to achieve the desired shape of the biomaterial, for example by casting 108, as rods 110, as granules and/or spheres 112, and molding 114. The solvent is removed (strip) from the shape and the material will form a hydrogel when exposed to water. The entire process up to the moment of actual use of the xerogel for the patient can be carried out in the absence of water and/or in the absence of hydrophobic materials. Fig. 1B depicts the microstructure of the biomaterial 120 produced by this method. The structure represents a material spanning its manufacturing and use processes: organogels, xerogels, and then hydrogels. The cross-linked matrix is made of precursors 124 that have covalently reacted with each other. Particles 124 of a water-soluble biological agent are dispersed within the matrix. The matrix is a continuous phase and the particles spread inside and are a discontinuous phase, also called dispersed phase.
An alternative embodiment involves the use of block copolymer precursors that are physically crosslinked through the formation of hydrophobic domains, as depicted in fig. 1C. The biomaterial 130 has bioreagent particles 132 dispersed in a matrix. The precursor has a hydrophilic block 134 and a hydrophobic block 136. The hydrophobic blocks 136 self-assemble to form hydrophobic domains 138, which create physical cross-linking between the precursors. The term physical crosslinking means crosslinking that is not covalently bonded. Hydrophobic domains are one such example, as well as hard and soft segments of polyurethanes or other multi-block copolymers. Ionic crosslinking is another example. The term cross-linking is well understood by the skilled person, who will immediately be able to distinguish covalent cross-linking from physical cross-linking, and the sub-types of physical cross-linking, such as ionic, hydrophobic and crystalline domains.
Other drug delivery approaches have used, for example, liposomes or micelles to encapsulate proteins, or to make nanoparticles, using polymers or other agents in the production of the particles. Protein delivery in hydrogels generally involves sequestering the protein from the hydrogel: for example, by placing the hydrogel in liposomes, micelles, or in a mixture with a binder such as a polymer. Other approaches involve direct adsorption of materials to proteins to inhibit their dissolution. Another approach is to precipitate the protein during delivery, as disclosed in U.S. publication No. 2008/0187568. Other approaches use hydrogels in which soluble proteins are dispersed throughout the hydrogel, where the hydrogel is eroded for controlled release.
Despite all these efforts, the effectiveness and success of sustained release therapies using biological agents (including proteins) is still limited because the stability of the biological agents in vivo tends to be poor. And loss of conformation can not only lead to loss of potency, but it can be detrimental by causing unwanted effects or eliciting an immune response. Despite much effort, there are no universally applicable solutions that are effective enough to have real-world clinical value, as reviewed in Wu and Jin, AAPS PhamSech 9(4): 1218-.
Surprisingly, however, the embodiments provided herein show that the solubility and release of proteins or other biological agents from a matrix can be controlled by: disposing the bioreagent as solid phase particles in a suitable matrix eliminates the need for these other approaches involving polymers, encapsulants, binders, and the like. In addition, biological agents are resistant to denaturation even in aqueous in vivo environments. The particles in the matrix are water soluble, but, despite not having any coating (coating) or the like, dissolve slowly and their dissolution in physiological solution, which is usually measured in minutes or hours, can be extended to days, weeks or months. Moreover, another unexpected and surprising result has been observed: that is, the biological agents do not tend to aggregate even though they must be present in the matrix at very high concentrations. It appears that the bioreagent leaves the particles very slowly. The first principle of operation (to which the invention is not limited) is to use a highly mobile polymer-for example the following: molecular chains of e.g. polyethylene glycol (PEG) or polyethyleneimine-made matrices form exclusion volumes around themselves, which limit the solubility of any other macromolecules in the immediate vicinity. This structural property not only limits the proteins to the solid phase by physical entrapment (entrampment) within the matrix, but also limits the dissolution of macromolecules so that protein particles cannot move into solution; as the particles and proteins begin to swell by solvation with water, they are constrained by the matrix until the matrix at least partially dissolves. Therefore, as the crosslinking density decreases and the molecular chains move further apart, the gradual dissolution of the entrapped macromolecular particles is promoted. These processes thus provide unexpected and surprising results: the biological agents remain in the solid phase until they become adjacent to the moment of their release from the matrix: thus, the protein or other biological agent is stable because it does not suffer from deleterious effects in solution over a long period of time. Release is also limited by diffusion of the macromolecule out of the matrix and is affected by the molecular weight of the macromolecule and the characteristics of the polymer forming the matrix. The second operating principle is complementary to the first operating principle and as such is not the mechanism to which the invention is limited: the molecular chains of the matrix associate with water molecules in the vicinity of the protein, so that the protein cannot be dissolved. This second principle can be applied to polymers with highly mobile, hydrophilic linear chains such as PEG. In addition to PEG, other water-soluble polymers or copolymers may be selected that exhibit a repulsive volume effect with the selected protein. For example, polymers such as polyacrylic acid, polyvinyl alcohol, polyvinylpyrrolidone (PVP), and Polyhydroxyethylmethacrylate (PHEMA) will typically have such an effect. Some polysaccharides also have these effects. PEG and/or these other polymers may also be incorporated into the organogel as a solid. They will dissolve in the presence of water, i.e. in the hydrogel (solubilisation). Also, non-crosslinked PEG and/or PEG copolymers such as PLURONIC are additives that may be entrapped in the hydrogel along with the protein to promote a repulsive volume effect, thereby keeping the protein in a solid state.
Aspects of the systems disclosed herein relate to a large improvement in the control of release over time by placing protein particles in hydratable xerogels. Examples 1-2 detail methods for forming xerogels comprising particles of water-soluble biological agents. Protein albumin and immunoglobulin (IgG) were used as models of water-soluble therapeutic agent proteins. Powders of these proteins were prepared. The powder particles are combined with a hydrogel precursor in an organic solvent to form an organogel. Tables 1-5 of example 1 illustrate examples of organogels including dispersed protein powder. The organogel is broken up and sieved into aggregates of particles, which are evacuated of organic solvent to form a xerogel. Working example 2 records the release of protein from xerogels.
As illustrated in fig. 2-5, the protein is fully (completely) released; unexpectedly, there is no detectable reaction of the organogel precursors with the proteins, which prevents them from being dissolved when the matrix degrades. Indeed, these proteins, and in general proteins, contain amine and thiol functional groups, which are potentially very reactive towards strong electrophiles such as the electrophilic precursors used. Although reactions with these electrophilic functional groups are expected, the lack of reaction suggests that these reactions are prevented by: prior to gelation, the protein is in an insoluble or substantially solid phase, while the gel-forming precursors are in a liquid phase. The release profile shows good control of the release rate and ranges from a rapid release of hours to a release of months.
Moreover, the rate and kinetics of release can be further controlled by combining groups of particles with each other, as illustrated in fig. 6 and 7. These demonstrate a substantially zero order release, which is the ability to deliver the drug at a time independent rate and the concentration of the drug within the drug dosage form is desirable. The zero order release mechanism ensures that a steady amount of drug is released over time, minimizing potential peak/trough fluctuations and side effects, while maximizing the amount of time (efficacy) that the drug concentration remains within the therapeutic window.
Methods and materials for preparing organogel-hydrogels, dual solvent delivery systems for water-soluble biological agents
A first embodiment involves forming a covalently cross-linked matrix. A fine powder of a water-soluble biological agent is prepared and suspended in an organic solvent that does not solvate the water-soluble biological agent, e.g., a protein. The term powder is used broadly herein to refer to an aggregate of dried particles. The term particle is broad and includes spheres, teardrop shapes, rods, and other irregular shapes. Typically, the powder is processed to provide a controlled particle composition having its known size, shape and distribution (as a difference from the average or mean). Protein powders typically contain stabilizing sugars such as sucrose or trehalose. These sugars are generally water soluble and not organic soluble. It was found that throughout the process, these will remain with the protein until such time as it hydrates to form a hydrogel. A matrix precursor is prepared that has the ability to form a cross-linked organogel by reacting with each other in an organic solvent. The precursor is selected to be soluble in the organic solvent. The precursor and the water-soluble biologic powder are mixed in an organic solvent such that the water-soluble biologic particles are dispersed throughout the matrix formed when covalent bonds are formed between the precursors. The matrix formed in the organic solvent is called organogel. The solvent is removed to form a xerogel. Upon hydration in water, the matrix forms an internal covalently crosslinked hydrogel. The process is a sequential (serial) two-solvent process, in that the organic solvent must be effective, removable (i.e., removable without leaving a pharmaceutically unacceptable residue) for the biological agent and precursor, but the precursor must be effective in an in vivo aqueous environment. The protein is never exposed to both the organic and aqueous phases. It is believed that exposure of proteins in aqueous solution to interfaces such as with organic liquids or solids or air bubbles contributes to protein adsorption and denaturation. The sequential organogel to xerogel to hydrogel approach eliminates the possibility of interface exposure, i.e., embodiments include the approach as described herein performed without exposure of the water-soluble biologic agent to an interface between any combination of the following: air, gas, water, organic solvent.
Another embodiment is the formation of covalently cross-linked gels (also referred to herein as pseudo-organogels) by using a liquid reactive polymer as a matrix precursor. A matrix precursor is prepared that has the ability to form a cross-linked organogel by reacting with each other in the absence of an organic solvent, such as when in a molten state. The precursor and water-soluble bioreagent powder are mixed at a temperature high enough to liquefy the precursor polymer, but low enough to maintain protein stability. Examples of such temperatures are from about 10 ℃ to about 75 ℃, or up to about 60 ℃ or up to about 75 ℃; the skilled person will immediately understand that all values and ranges between the explicitly stated values are designed and incorporated herein as if written in detail. Mixing conditions are employed such that the water-soluble bioreagent particles are dispersed throughout the matrix formed upon formation of covalent bonds between the precursors. The reaction is thus carried out in the melt of the polymer, the term melt meaning that no solvent is present. However, other materials may be present in the melt, such as biological agents, sugars, proteins, buffers. Embodiments include materials and methods of making medical materials comprising forming a gel around a powder of a water-soluble biologic, wherein the powder is dispersed in the gel, wherein forming the gel comprises preparing a melt of one or more precursors and covalently crosslinking the precursors. A majority of the volume of the gel, e.g., about 30% to about 95% v/v, may be occupied by the biological agent or other solid (e.g., sugar, buffer salt); the skilled artisan will immediately appreciate that all values and ranges between the explicitly stated values are contemplated and incorporated herein, as if written in detail, for example, at least 30% volume/volume or from about 40% to about 75%.
Another embodiment of the sequential two-solvent process involves forming a crosslinked material having physical crosslinks. One such embodiment uses a block copolymer as a precursor. The precursor has a lyophilic (solvent-philic) block and a lyophobic (solvent-phobic) block. These precursors are added to an organic solvent and form a physically crosslinked matrix. The block (also referred to as segment) that precipitates in a particular organic solvent to form an organogel may or may not be the same segment that precipitates in water to form a hydrogel. After removal of the solvent, the resulting xerogel forms a hydrogel in aqueous solution because one or more of the block or segment portions are hydrophobic and one or more of the block or segment portions are hydrophilic. Related embodiments use two organic solvents: the block copolymeric precursor is dissolved in a first organic solvent. The copolymer solution is then mixed with a second organic solvent that is miscible with the first solvent but is non-solvent for at least one of the segments of the copolymer. The lyophobic domains form organogels. Another embodiment uses the first and second organic solvents and also the second organic solvent to precipitate the bioreagent such that an organogel and particles of bioreagent are formed in the same step.
Another embodiment of the sequential two-solvent process involves thermal gelation. The precursor that is converted from solution to an organogel in an organic solvent at a temperature in the range of about-20 ℃ to about 70 ℃ is placed in the organic solvent along with the bioreagent at a temperature where the precursor is in a dissolved state. The solution is then cooled to a second temperature below the gelation point, and the precursor forms an organogel. Thus, the method for making the organogel is to heat the solvent to dissolve the copolymer and then cool the solution to precipitate at least one of the segments of the copolymer. The solvent is then removed to produce a xerogel. The precursors are selected such that the xerogel is a hydrogel at physiological temperatures.
Block copolymers that can be suitable for use in these methods include a number of PEG copolymers. PEG is hydrophilic and lyophilic with respect to many organic solvents. Other hydrophilic polymers and polymer type chain segments are polyvinyl alcohol, polyacrylic acid, polymaleic anhydride, PVP, PHEMA, polysaccharide, polyethyleneimine, polyvinylamine, polyacrylamide and the like. The other blocks are chosen to be hydrophobic and lyophobic to organic solvents. Examples of such other blocks are: polybutylene terephthalate (PBT), polylactic acid, polyglycolic acid, polytrimethylene carbonate, polydioxanone, and polyalkyl ethers such as polypropylene oxide (PLURONICS, POLOXAMERS). The copolymer may have one or more of each block type.
These methods may be performed such that the water-soluble bioreagent is never contacted with water from the time it is initially prepared until placed in the body. The water-soluble bioreagent may be further processed such that, once obtained in purified form at the source or manufacturing location, it is later insoluble in water and/or never exposed to water during the gel manufacturing process. Exposure to water can cause a variety of problems. One problem is that the protein will undergo hydrolysis over time, so that it degrades slowly. Another problem is that proteins, once in solution, can rearrange or form metastable aggregates such as dimers or trimers.
Embodiments of the invention include these methods performed in the absence of hydrophobic polymers and/or hydrophobic solvents. Embodiments requiring hydrophobic block polymers cannot be performed in a non-hydrophobic manner, but the skilled person can easily discern which methods are applicable. One embodiment provides for covalent crosslinking of the hydrophilic precursor in an organic solvent in the presence of the bioreagent particles and in the absence of the hydrophobic material at both the organogel step and the subsequent step. In some embodiments, a hydrophobic solvent may be present without damage, depending on the solvent, and thus embodiments include the absence of hydrophobic materials other than solvents; and/or the absence of hydrophobic polymers; and/or the absence of hydrophobic polymer segments.
Common sense teaches that organic solvents typically denature proteins. Some life science processes can tolerate some degree of denaturation, for example, in diagnostic or analytical devices. However, in the medical field, even a small degree of denaturation is undesirable. Denatured proteins may exhibit a wide range of properties, from loss of solubility to co-aggregation (communalaggregation). Co-aggregation involves aggregation of hydrophobic proteins closer to each other to reduce the total area exposed to water. The reduction in distance may result in a permanent or quasi-stable association. When a protein is denatured, its secondary and tertiary structures change, but the peptide bonds of the primary structure between amino acids generally remain intact.
Surprisingly, however, it has been found that proteins that remain in the solid phase can be exposed to some organic solvent without substantial denaturation. Completely anhydrous organic solvents which are treated under anhydrous conditions are preferred. Denaturation by exposure to organic solvents can occur when the protein is already in aqueous solution and/or if the organic solvent, or organic/aqueous mixed solvent (e.g. ethanol/water), has a tendency to dissolve or even swell the protein particles in a limited manner. Protein-solvent compatibility can be established experimentally by: exposure followed by characterization tests to determine if the protein has denatured and/or has undergone replacement or alteration by one or more chemical groups. Organic solvent compatibility can be tested simply by: the test protein is immersed in the test solvent for a suitable period of time, the protein is removed, for example by filtration and vacuum drying, and recovery of the protein is then tested by HPLC or other suitable analytical method. The solvent most likely to leave the protein intact is anhydrous and hydrophobic, but must also be a good solvent for the gel-forming precursor molecules. In the case of polyethylene glycol (PEG) precursors, solvents such as methylene chloride and dimethyl carbonate are employed. Other solvents such as acetone (or acetone/water), ethyl acetate, tetrahydrofuran may also be useful. Supercritical fluid solvents such as carbon dioxide may also be useful for forming organogels.
Precursors are described in detail elsewhere herein. Many useful precursors are available as a variety of precursors. A first precursor is added to the solvent-protein mixture followed by the addition of a second precursor, which is reactive with the first precursor to form crosslinks. The first precursor may be selected to have only those functional groups as follows: which is non-reactive to form covalent bonds with proteins in the absence of further chemical components. Proteins have amines and thiols available for reaction with some electrophilic functional groups to form covalent bonds, and carboxyl and hydroxyl groups available for other chemical reactions. The precursor may thus be selected to be non-reactive to these functional groups. For example, the precursor may have amine and/or thiol and/or hydroxyl and/or carboxyl groups and be non-reactive with proteins. Thus, embodiments of the present invention involve adding a protein-non-reactive first precursor to a protein-organic solvent mixture followed by adding a second precursor that is reactive to the first precursor.
The water-soluble biologic agent particles can be free of one or more of the following: binders, fatty acids, hydrophobic materials, surfactants, fats, phospholipids, oils, waxes, micelles, liposomes, and nanocapsules (capsules). The organogel or xerogel comprising particles of the water-soluble biologic may also be free of one or more of the above. The protein or other water-soluble biologic agent in the xerogel may be entirely in the solid phase, may be entirely crystalline, partially crystalline, or substantially free of crystals (meaning more than 90% free of crystals, weight/weight; the skilled artisan will immediately appreciate that all ranges and values within the explicitly stated ranges are contemplated).
The xerogel-water soluble biologic material can be formed in the desired shape. One method is to react the precursor in a mold having the desired shape. The shape is removed from the mold before or after the solvent is removed. The material may also be fragmented into particles, as described in more detail elsewhere herein.
After forming the matrix in the organic solvent, the solvent may be removed to form a xerogel. Possible methods include, for example, precipitation using a non-solvent, nitrogen purge drying, vacuum drying, freeze drying, a combination of heat and vacuum, and lyophilization.
If the molten precursor is used in the absence of a third (tertiary) solvent, no solvent removal process is employed. Upon cooling, the material forms a rubbery solid (if above Tm), a semi-rigid semi-crystalline material (if below Tm), or a rigid glassy solid (if below Tg). These materials are denser than xerogels formed from organic solvents. When filled with particles of other materials, such as therapeutic agents, buffer salts, visualization agents, they can be highly porous, as solid particles create and fill the pores.
All of these methods can be performed in the absence of water-soluble biological agents. Materials (including particles) have utility for many applications without biological agents. Uses include, for example, tissue augmentation (augmentation), bulking agents, and tissue separation in radiotherapy.
Moreover, all of these methods can be performed with additional reagents in place of or in addition to the biological reagents. Such additional agents include visualization agents visible to the naked eye and radiopaque agents or materials.
Preparation of granules
An organogel may be formed and then comminuted into particles that are subsequently treated to remove the organic solvent to form a xerogel. For injectable forms, the organogel may be macerated (macerate), homogenized, extruded, screened, chopped, diced, or otherwise comminuted into particulate form. Alternatively, the organogel may be formed as a molded article or drop (droplet) containing suspended protein particles.
One method for making organogel particles involves creating a matrix, which is broken up to make organogel particles. Thus, a matrix is fabricated from a precursor as described herein, which is then broken up. One technique involves preparing an organogel with protein particles and grinding it, for example, in a ball mill or using a mortar and pestle. The substrate may be chopped or diced with a knife or wire. Alternatively, the matrix may be chopped in a blender or homogenizer. Another method involves forcing the organogel through a mesh, collecting the debris, and passing them through the same mesh or another mesh until the desired size is achieved.
The water-soluble biological agent, such as a protein, is prepared as particles prior to dispersion into the organogel. Various protein granulation (granulation) techniques such as spray drying or precipitation exist and may be employed provided that the protein of interest is compatible with such processing. Embodiments of particle preparation involve receiving a biological agent without substantial (significant) denaturation, e.g., from a supplier or animal or recombinant (recombiant) source. For proteins, the solid phase is in a stable form. The protein is lyophilized or concentrated or used as received. The protein is then prepared as a fine powder without denaturation by processing the protein in the solid state and avoiding high temperatures, moisture and optionally in an oxygen-free environment. The powder can be prepared, for example, by: milling, ball milling, cryogenic milling (cryomilling), microfluidization, or mortar and pestle milling, followed by sieving of the solid protein. Proteins may also be processed while maintaining the protein in solid form in a compatible anhydrous organic solvent in which the protein in question is insoluble. The reduction of the particle size to the desired range can be achieved, for example, by: attrition milling, ball milling, jet milling of solid protein suspensions in compatible organic solvents. High shear rate processing, high pressure and sudden temperature changes should be minimized as they lead to protein instability. Thus, care must be taken to treat the protein or other water-soluble biological agent in a manner that avoids damage, and without appropriate redesign and testing of the results, the use of conventional methods for making particles should not be considered appropriate, and would not be expected to be useful.
The term powder of proteins refers to a powder made from one or more proteins. Similarly, a powder of a water-soluble biological agent is a powder having particles made of one or more water-soluble biological agents. The proteins in the protein particles or the bioreagent in the bioreagent particles associate with each other to provide mechanical integrity and structure to the dried particles, even in the absence of a binder or encapsulant. These powders are distinguished from protein or biological agent delivery using encapsulation or other techniques such as liposomal, micellar, or nanocapsule approaches, essentially encapsulating proteins or biological agents. The powders and/or xerogels or hydrogels comprising them may be free of encapsulating material and free of one or more of liposomes, micelles, or nanocapsules. In addition, proteins can be made that do not contain one or more of the followingPlasmid particles or water-soluble biological agent particles: a binder, a non-peptidic polymer, a surfactant, an oil, a fat, a wax, a hydrophobic polymer, comprising more than 4 CH2Polymers with long alkyl chains, phospholipids, micelle-forming polymers, micelle-forming compositions, amphiphilic molecules, polysaccharides of three or more sugars, fatty acids, and lipids. Lyophilized, spray-dried, or otherwise processed proteins are often formulated with sugars such as trehalose to stabilize the protein by lyophilization or other methods used to prepare the protein. These sugars can be allowed to persist in the particles throughout the organogel/xerogel process. The particles can be made to include from about 20% to about 100% (dry, weight/weight) protein; the skilled artisan will immediately appreciate that all ranges and values within the explicitly stated ranges are contemplated, for example, from about 50% to about 80% or at least 90% or at least about 99%.
The particles of the biological agent or of the organogel or of the xerogel can be separated into aggregates having the desired size range and size distribution by a variety of methods. Very fine control of the classification (sizing) is available, wherein the size ranges from 1 micron to several mm, and wherein the mean and range of particle sizes are controllable for narrow distributions. The skilled artisan will immediately appreciate that all ranges and values within the explicitly stated ranges are contemplated, for example, from about 1 to about 10 μm or from about 1 to about 30 μm. About 1 to about 500 microns is another such useful range where the size falls within the entire range and has an average size at one value within the range, and a standard deviation centered on the average, e.g., from about 1% to about 100%. A simple method for classifying particles involves the use of a custom or standardized mesh size. In addition to standard U.S. and Tyler mesh sizes, sieves in commercial grades (Market Grade), grinding grades (Mill Grade) and stretch Bolting Cloth (tensle Bolting Cloth) are also commonly used. The material forced through the mesh may exhibit deformation such that the particle size does not exactly match the mesh size; however, the mesh size may be selected to achieve the desired particle size range. Particle size analyzers, in which protein particles are dispersed in an organic or oil phase, are commonly used. Particle size is also commonly determined using microscopy. Spherical particles refer to particles in which the longest central axis (a straight line passing through the geometric center of the particle) does not exceed about twice the length of the other central axes, wherein the particles are literally spherical or have an irregular shape. Rod-shaped particles refer to particles in which the longitudinal central axis exceeds about twice the length of the shortest central axis. The implementation mode comprises the following steps: producing a plurality of collections of particles, wherein the collections have different rates of in vivo degradation, and mixing the collections to produce a biomaterial having a desired degradation property.
Delivery of water-soluble biological agents without denaturation
These processes can be performed using proteins or other water-soluble biological agents. These include peptides and proteins. The term protein as used herein refers to a peptide of at least about 5000 daltons. The term peptide as used herein refers to a peptide of any size. The term oligopeptide refers to peptides having a mass of up to about 5000 daltons. Peptides include therapeutic proteins and peptides, antibodies, antibody fragments, short chain variable regions (fragments) (scFv), growth factors, angiogenic factors, and insulin. Other water-soluble biological agents are carbohydrates, polysaccharides, nucleic acids, antisense nucleic acids, RNA, DNA, small interfering RNA (sirna), and aptamers. The description herein is often in terms of proteins, but the methods are generally applicable to other water-soluble biological agents.
Proteins are easily denatured. However, as described herein, proteins can be delivered substantially without denaturation, including where no binders, lipophilic materials, surfactants, or other prophylactic components are used. The term substantially no denaturation is a protein that is processed into particles without a change in the chemical structure of the protein (no addition of chemical groups or change in existing chemical groups) and without a change in the conformation of the protein, i.e. the secondary and/or tertiary and/or quaternary structure. In this context, the term essentially means that no significant difference between the processed protein and the control protein was observed (p value <0.05) for the averaged test group tested under conventional conditions by enzyme linked immunosorbent assay (ELISA) for epitope (epitope) denaturation and by isoelectric focusing (IEF) for shifts exceeding 0.2 in the isoelectric point (pI), see us serial No. 13/234,428, which is hereby incorporated by reference herein for testing or protein stability and all purposes; in case of conflict, the present specification will control. Primary protein structure refers to the amino acid sequence. To be able to perform their biological functions, proteins fold into one or more specific spatial conformations, which are driven by a number of non-covalent interactions such as hydrogen bonding, ionic interactions, van der waals forces, and hydrophobic packing (packing). The term secondary structure refers to local protein structure, e.g. local folding. Tertiary structure refers to a specific three-dimensional conformation, including folding. Proteins with secondary and/or tertiary structure thus present local and global structural organization. In contrast, linear peptides that do not have a specific conformation do not have secondary and/or tertiary structure. The term native means as found naturally (in nature) in vivo, such that the protein can be processed into particles and released in the native conformation.
Denaturation of proteins can be tested by a variety of techniques, including enzyme-linked immunosorbent assay (ELISA), isoelectric focusing (IEF), Size Exclusion Chromatography (SEC), High Pressure Liquid Chromatography (HPLC), Circular Dichroism (CD), and fourier transform infrared spectroscopy (FTIR). These tests report, for example, the following parameters: changes in molecular weight, changes in end groups, changes in bonds, changes in hydrophobicity or size exclusion, and exposure/concealment of antigenic sites (antigenic groups). Generally, tests by IEF and ELISA can be designed that are sufficient to show native conformation after processing, although other tests and combinations of tests can alternatively be used.
Experiments have shown that many factors contributing to processing and delivering proteins without denaturation can be controlled. The protein may be prepared as a powder, wherein the powder particle size is selected according to the size of the final organogel. All organic solvents for the protein can be selected such that the protein is not solvated by the organic solvent and is compatible with the protein. Another factor is oxygen, and the elimination of oxygen is useful in the process to avoid denaturation. Another factor is chemical reaction. These can be avoided by: the protein is held in a solid phase and does not contain a solvent to solubilize the protein until such time as the protein is implanted.
One embodiment of particle preparation involves receiving the protein without substantial (significant) denaturation, e.g., from a supplier or animal or recombinant source. The protein is lyophilized, spray dried or concentrated or used as received. The protein is then prepared as a fine powder without denaturation by processing it in the solid state and avoiding high temperatures, moisture and optionally in an oxygen-free environment. Powders can be prepared by, for example, grinding, ball milling, or mortar and pestle grinding of solid proteins.
The manufacture of a proteinaceous agent or other water-soluble biological agent as a particle can be a useful first step for delivery of the agent from a solid phase. However, it is not a sufficient step for achieving a well-controlled release, or an effective release, from the matrix over an extended period of time. However, upon implantation, the particles will tend to dissolve rapidly as water contacts the particles and solvates the agent. In the case of particles in a hydrogel, for example, water permeates the hydrogel and contacts the particles. However, unexpectedly, it is possible to prevent the dissolution of water-soluble biological agents in the particles in the hydrogel. Some mechanisms for doing so are set forth herein, but will not be used to limit the invention to a particular theory of action. One mechanism apparently involves the use of a matrix that prevents the agent from dislodging from the particles. Moreover, even if the molecules of the agent dissolve, they are held at the local (local) location and will saturate the local location to prevent further solvation of other agent molecules. Another mechanism involves the solvation of a matrix that competes with potentially soluble agents for water, where the matrix has a size exclusion effect to interfere with agent solvation.
These mechanisms involve the achievement of a space between molecular chains of a dense matrix. The crosslink density of the organogel matrix (and thus the xerogel and hydrogel matrices) is controlled by the total molecular weight of the precursor and other precursors used as crosslinking agents and the number of functional groups available per precursor molecule. A lower molecular weight, e.g. 500, between crosslinks will provide a much higher crosslink density than a higher molecular weight, e.g. 10,000, between crosslinks. The crosslink density can also be controlled by the total solids percentage of the crosslinker and the functional polymer solution. Yet another method for controlling crosslink density is by adjusting the stoichiometry of nucleophilic to electrophilic functional groups. The one-to-one ratio results in the highest crosslink density. Precursors with longer distances between crosslinkable sites form gels that are generally softer, more complex and more elastic. The increased length of the water soluble segment, such as polyethylene glycol, tends to increase the elasticity to produce desirable physical properties. Some embodiments thus relate to precursors having water-soluble segments with molecular weights in the range of 2,000-100,000; the skilled artisan will immediately appreciate that all ranges and values within the explicitly stated ranges are contemplated, such as 10,000 and 35,000. The solids content of the hydrogel can affect its mechanical properties and biocompatibility and reflect a balance between competing requirements. Relatively low solids content is useful, for example, from about 2.5% to about 20%, including all ranges and values therebetween, for example, from about 2.5% to about 10%, from about 5% to about 15%, or less than about 15%. The skilled person will appreciate that the same material may be used to fabricate substrates having a wide range of structures, which will have very different mechanical properties and performance, such that the achievement of a particular property should not be based solely on the general type of precursor involved.
Delivery of water-soluble biological and other therapeutic agents
Various water-soluble biological agents and/or other therapeutic agents can be delivered using the systems described herein. Xerogel particles containing protein powders can be used to deliver water-soluble biological and/or other therapeutic agents. The particles may be applied within a xerogel. The xerogel may be of preformed structure, e.g. having at least 2cm3Of (the skilled person will immediately understand, within the scope explicitly statedAre designed such that all ranges and values of (A) are, for example, from about 2 to about 20cm3) Or as an aggregate of particles. Alternatively, the xerogel particles may be administered directly, or in a pharmaceutically acceptable binder or carrier. Other materials may include xerogel particles. Water soluble agents are one class of agents that can be delivered as a powder within a xerogel. Other drugs such as hydrophobic agents or small molecule drugs (water soluble or hydrophobic) can also be mixed into or with the xerogels.
Proteins are a class of water-soluble agents. The xerogel particles can be processed such that the proteins are introduced and released without substantial denaturation and/or in their native conformation. Some anti-vascular endothelial growth factor (anti-VEGF) agents are therapeutic proteins. anti-VEGF therapy is important in the treatment of some cancers and in age-related macular degeneration. They may relate to monoclonal antibodies such as bevacizumab (AVASTIN), antibody derivatives such as ranibizumab (LUCENTIS), or small molecules that inhibit tyrosine kinases stimulated (stimulated) by VEGF: lapatinib (TYKERB), Sunitinib (SUTENT), sorafenib (neraVAR), axitinib (axitinib), and pazopanib (pazopanib). (some of these treatments target the VEGF receptor as opposed to VEGF).
Some conventional ophthalmic drug delivery systems deliver drugs in topical (topical) eye drops. For example, antibiotics are administered dropwise every few hours for several days after cataract and vitreoretinal surgery. In addition, it may be desirable to frequently provide other drugs such as nonsteroidal anti-inflammatory drugs (NSAIDS). Some of these eye drops, such as restasi (allergan), also have stinging and burning sensations associated with their administration. Restasi is indicated for dry eye and must be used several times a day by a patient. Similarly, the administration of steroids or NSAID drugs is also required for the treatment of other eye diseases such as cystoid macular edema, Diabetic Macular Edema (DME), and diabetic retinopathy. Several vascular proliferative diseases such as macular degeneration are treated with intravitreal injections of VEGF inhibitors. These include drugs such as LUCENTIS and AVASTIN (Genentech) and MACUGEN (OSI). Such drugs may be delivered using the hydrogel-and-particle systems described herein, wherein repeated dosing steps are avoided; for example, no new drug application is performed daily, weekly, or monthly, or no topical eye drops are used to administer the drug.
A variety of drug delivery systems are known. These various other systems typically include intravitreal implant reservoir (reservoir) type systems, biodegradable reservoir (depot) systems, or (non-erodible) implants that require removal. The level of skill in this regard has been described in textbooks such as "Intraocular Drug Delivery" (Jaffe et al, Taylor & Francis published, 2006). However, most of these implants need to be removed at an expiration date, can detach from their target site, can cause visual impairment in the posterior part of the eye, or can themselves be inflammatory due to the release of significant amounts of acidic degradation products. These implants are therefore made very small, with very high drug concentrations. Even if they are small, they still require deployment with needles over 25G (25 gauge) in size, or surgical access delivery systems to be implanted or removed when needed. Typically, these are topical injections of drug solutions into the vitreous or intravitreal implants using biodegradable routes or removable depot routes. For example, topical injections for delivery into the vitreous include the anti-VEGF agent LUCENTIS or AVASTIN. Posurdex (allergan) is a biodegradable implant, where the indication used is Diabetic Macular Edema (DME) or retinal vein occlusion, where a 22-gauge syringe delivery system is used for delivery into the vitreous cavity; these are powerful drugs in a short drug delivery duration setting. The therapeutic agent is dexamethasone (dexamethasone) with a polylactic acid/polyglycolic acid polymer matrix. Tests using posuredex for diabetic retinopathy are underway. Also, for example, MEDIDURE implants (psidiida) are used for DME indications. The therapeutic agent of the implant is fluocinolone acetonide and has a nominal delivery lifetime of 18 months or 36 months (of both forms). Intravitreal removable implants containing triamcinolone acetonide are being tested. Its nominal delivery life is about two years and requires surgical implantation. The indication is for DME.
In contrast to these conventional systems, these or other therapeutic agents can be delivered using a collection of xerogel particles or a system comprising the particles. Xerogel particles comprise the agent. The xerogels, when exposed to physiological fluids, absorb the fluids to form hydrogels that are biocompatible to the eye, an environment that is distinctly different from other environments. The use of minimally inflammatory materials avoids angiogenesis, which is harmful in many cases in the eye. Biocompatible ocular materials thus avoid unintended angiogenesis; in some aspects, avoiding acidic degradation products achieves this goal. Furthermore, by using hydrogels and hydrophilic materials (components having a solubility of at least one gram per liter in water, e.g., polyethylene glycol/oxide), the influx of inflammatory cells is also minimized; this procedure is in contrast to conventional use of non-hydrogel or rigid reservoir-based ocular implants. Moreover, some proteins can be avoided to improve biocompatibility; collagen or fibrin glue, for example, tends to promote inflammation or unwanted cellular reactions, as these release signals that promote biological activity as they degrade. Instead, synthetic materials or peptide sequences not normally found in nature are used. In addition, biodegradable materials can be used to avoid chronic foreign body reactions, for example, as with a thermally formed gel that does not degrade. In addition, soft materials or materials manufactured in situ to conform to the shape of the surrounding tissue can minimize ocular distortion, and low-swelling materials can be used to eliminate visual distortion caused by swelling. High or low pH materials may be avoided during the formation, introduction or degradation stages.
Xerogels can be prepared with and used to deliver a wide variety of drugs (as well as drugs to other parts of the body for topical and systemic delivery), including steroids, non-steroidal anti-inflammatory drugs (NSAIDS), anti-cancer drugs, antibiotics, and the like. The xerogels are useful for the delivery of drugs and therapeutics, such as anti-inflammatory drugs (e.g., Diclofenac), analgesic drugs (e.g., Bupivacaine), calcium channel blockers (e.g., Nifedipine), antibiotics (e.g., Ciprofloxacin), cell cycle inhibitors (e.g., Simvastatin), proteins (e.g., insulin). The particles can be used to deliver a wide variety of drugs including, for example, steroids, NSAIDS, antibiotics, analgesics, Vascular Endothelial Growth Factor (VEGF) inhibitors, chemotherapy, antiviral drugs. Examples of NSAIDS are Ibuprofen (Ibuprofen), meclofenamate sodium, mefenamic acid (mefanamic acid), salsalate, sulindac (sulindac), tolmetin sodium (tolmetin sodium), ketoprofen (ketoprofen), diflunisal (diflunisal), piroxicam (piroxicam), naproxen (naproxen), etodolac (etodolac), flurbiprofen (flurbiprofen), fenoprofen calcium (fenoprofen calcium), Indomethacin (Indomethacin), celecoxib, ketorolac (ketorolac) and nepafenac (nepafenac). The drug itself can be a small molecule, protein, RNA fragment, protein, glycosaminoglycan (glycosaminoglycan), carbohydrate, nucleic acid, inorganic and organic bioactive compound, wherein specific bioactive agents include, but are not limited to: enzymes, antibiotics, antineoplastic agents, local anesthetics, hormones, angiogenic agents, anti-angiogenic agents, growth factors, antibodies, neurotransmitters, psychotropic drugs, anti-cancer drugs, chemotherapeutic drugs, drugs affecting the genitalia, genes, and oligonucleotides, or other configurations.
These xerogel particles or other xerogel structures can be used to deliver a variety of drugs or other therapeutic agents. A list of examples of agents or series of drugs and indications for the agents is provided. The agents may also be used as part of a method of treating an indicated disorder or making a medicament for treating an indicated condition. For example, AZOPT (brinzolamide) ophthalmic suspension) may be used for the treatment of elevated intraocular pressure in patients with ocular hypertension or open angle glaucoma. BETADINE in povidone-iodine ophthalmic solution is useful for preparation of the periocular region and irrigation of the ocular surface. Betotic (betaxolol HCl) can be used to reduce intraocular pressure, or for chronic open-angle glaucoma and/or ocular hypertension. CILOXAN (Ciprofloxacin HCl) ophthalmic solution) can be used to treat infections caused by susceptible strains of microorganisms (susceptable strains). NATACYN (Natamycin ophthalmic suspension) can be used for the treatment of fungal blepharitis, conjunctivitis and keratitis. NEVANAC (nepafenac) ophthalmic suspension is useful in the treatment of inflammation and pain associated with cataract surgery. TRAVATAN (Travoprost ophthalmic solution) is useful for lowering elevated intraocular pressure-open angle glaucoma or ocular hypertension. FML FORTE (Fluorometholone ophthalmic suspension) is useful in the treatment of corticosteroid-reactive inflammation of the palpebral and bulbar conjunctiva, cornea and anterior segment of the eyeball. LUMIGAN (Bimatoprost) ophthalmic solution) is used for lowering elevated intraocular pressure-open angle glaucoma or ocular hypertension. PRED FORTE (Prednisolone acetate) is useful for the treatment of steroid-reactive inflammation of the palpebral and bulbar conjunctiva, cornea and anterior segment of the eyeball. PROPINE (dipivefrin hydrochloride) is used for intraocular pressure control in chronic open-angle glaucoma. RESTASIS (Cyclosporine) ophthalmic emulsions) can be used to increase tear production in patients such as those with ocular inflammation associated with keratoconjunctivitis sicca. ALREX (Loteprednol etabonate) ophthalmic suspension) is useful for temporary relief of seasonal allergic conjunctivitis. LOTEMAX (loteprednol etabonate ophthalmic suspension) is useful for the treatment of steroid-reactive inflammation of the palpebral and bulbar conjunctiva, cornea and anterior segment of the eyeball. MACUGEN (injectable Pegaptanib sodium) can be used for the treatment of neovascular (wet) age-related macular degeneration. OPTIVAR (Azelastine hydrochloride) is used for the treatment of ocular itching associated with allergic conjunctivitis. XALATAN (Latanoprost ophthalmic solution) is useful for reducing elevated intraocular pressure in a patient (e.g., with open angle glaucoma or ocular hypertension). BETIMOL (Timolol) ophthalmic solution) is useful for the treatment of elevated intraocular pressure in patients with ocular hypertension or open angle glaucoma. Latanoprost is a free acid form prodrug that is a prostanoid-selective FP receptor agonist. Latanoprost lowers intraocular pressure in glaucoma patients with few side effects. Latanoprost has relatively low solubility in aqueous solutions, but is readily soluble in organic solvents typically used for microsphere manufacture using solvent evaporation.
Further embodiments of agents for delivery include those that specifically bind a target peptide in vivo to prevent interaction of the target peptide with its natural receptor or other ligand. AVASTIN is, for example, an antibody that binds VEGF. Also aflibecrcept is a fusion protein comprising a portion of a VEGF receptor for capturing VEGF. IL-1 capture agents (IL-1trap) which utilize the extracellular domain of the IL-1 receptor are also known; the capture agent blocks IL-1 binding and activation of receptors on the cell surface. Embodiments of agents for delivery include nucleic acids, such as aptamers. Pegaptanib (MACUGEN) is, for example, a pegylated (pegylated) anti-VEGF aptamer. An advantage of the particle-and-hydrogel delivery method is that aptamers are protected from the in vivo environment until they are released. Further embodiments of agents for delivery include macromolecular drugs, which is a term referring to drugs that are significantly larger than classical small molecule drugs (i.e., drugs such as oligonucleotides (aptamers, antisense, RNAi), ribozymes, gene therapy nucleic acids, recombinant peptides, and antibodies).
One embodiment comprises extended release of a drug for allergic conjunctivitis. For example, ketotifen (a antihistamine and mast cell stabilizer) can be provided in particles and released to the eye in an effective amount for treating allergic conjunctivitis as described herein. Seasonal Allergic Conjunctivitis (SAC) and Perennial Allergic Conjunctivitis (PAC) are allergic conjunctival disorders (conditions). Symptoms include itching and pink to reddish eyes. Both of these ocular conditions are mediated by mast cells. Non-specific measures to ameliorate symptoms routinely include: cold compress, eye wash with tear substitute, and avoidance of allergens. Treatment conventionally consists of an antihistamine mast cell stabilizer, a dual mechanism (dual mechanism) anti-allergen agent, or a topical antihistamine. Corticosteroids may be effective, but due to side effects they are retained for more severe forms of allergic conjunctivitis such as Vernal Keratoconjunctivitis (VKC) and Atopic Keratoconjunctivitis (AKC).
Moxifloxacin (moxifloxacin) is an active ingredient in vigmox, a fluoroquinolone approved for the treatment or prevention of bacterial infections of the eye. The dose is typically one drop of a 0.5% solution, which is applied 3 times daily for a week or more.
VKC and AKC are slow allergic diseases in which eosinophils (eosinophil), conjunctival fibroblasts, epithelial cells, mast cells, and/or TH2 lymphocytes worsen the biochemistry and histology of the conjunctiva. VKC and AKC can be treated by drugs used to combat allergic conjunctivitis.
Osmotic agents are agents and may also be included in gels, hydrogels, organogels, xerogels, and biomaterials as described herein. These are agents that aid in the penetration of the drug into the intended tissue. The osmotic agent may be selected as desired for the tissue, for example, for the skin, for the tympanic membrane, for the eye.
Xerogel particle blending and aggregation
The collection of particles (powder particles and/or xerogel/hydrogel particles of the agent) may comprise a population (set) of particles. The term xerogel/hydrogel refers to xerogels and/or xerogels that are hydrated into hydrogels. For example, an assembly can include some xerogel particles that contain a radiopaque agent, where those particles form a population in the assembly. Other populations relate to particle size, where the populations have different shapes or size distributions. As discussed, the particles can be manufactured in well controlled sizes and can therefore be manufactured and divided into multiple populations to be combined into an aggregate.
Some populations are made of particles with specific degradability (xerogel/hydrogel). One embodiment relates to a plurality of clusters each having a different degradability profile. Different degradation rates provide different release profiles. Combinations of different particle populations can be made to achieve the desired curve, as demonstrated in fig. 6 and 7, reference example 2. The degradation time comprises 3-1000 days; the skilled person will immediately understand that all ranges and values within the explicitly stated ranges are contemplated. For example, a first population may have a median degradation time of about 5 to about 8 days, a second population may have a median time of about 30 to about 90 days, and a third population may have a median time of about 180 to about 360 days.
The xerogel/hydrogel particles can be blended to achieve the desired protein release profile. Gels (e.g., hydrogels) having different degradation rates can be combined to provide a constant or nearly constant release that compensates for the inherent non-linear release profile of a single gel.
The assembly of xerogel/hydrogel particles may comprise a population of agents. For example, some particles can be made to contain a first therapeutic agent, where those particles form a cluster in the aggregate. And other populations may have another reagent. Examples of agents are water-soluble biological agents, proteins, peptides, nucleic acids, small molecule drugs, and hydrophobic agents. Other populations may relate to particle size, where the populations have different shapes or size distributions. As discussed, the particles can be manufactured in well controlled sizes and divided into multiple populations to be combined into aggregates. These multiple clusters can be freely mixed and matched in combinations and sub-combinations such as: size, degradability, therapeutic agent, and visualization agent.
The xerogel/hydrogel may further comprise an agent which is not in powder form. The agent may be disposed in a xerogel/hydrogel or mixed with a solution of other excipients used with the xerogel/hydrogel. For example, the assembly of xerogel particles may be hydrated to form a hydrogel at the point of use by the addition of saline or water which further comprises a drug solution. Such drugs or agents may be the same as the agents in powder form in the xerogel/hydrogel to provide an initial burst release, or may be used for secondary therapy or visualization.
Lubricity of
The aggregate can be made by the size and lubricity of a small gauge needle for manual injection. The hydrophilic hydrogel, comminuted into spherical particles about 40 to about 100 microns in diameter, is small enough to be manually injected through a 30 gauge needle. It was observed that hydrophilic hydrogel particles were difficult to pass through a small gauge needle/catheter, as reported in U.S. publication No.2011/0142936, which was hereby incorporated herein for all purposes; in case of conflict, the present specification will control. The particle size contributes to the resistance and viscosity of the solution. The particles tend to clog the needle. The resistance is proportional to the viscosity of the fluid, with a more viscous fluid requiring more force to push through a small opening.
As reported in U.S. publication No.2011/0142936, it was unexpectedly found that increasing the viscosity of the solvent used for the particles can reduce the resistance through the catheter and/or needle. This reduction can be attributed to the use of solvents with high osmolarity. Without being bound by a particular theory, the addition of these agents to improve injectability results from: the particles shrink, increased free water between particles that reduces the contribution of the particles to the viscosity, and increased viscosity of the free water, which helps pull the particles into and out of the syringe, preventing strain and clogging. The use of linear polymers may further contribute to thixotropic properties, which are useful to prevent settling of the particles and to promote movement of the particles with the solvent, but exhibit shear thinning when forced out of the small openings. It was also observed that this approach solves another problem, namely the difficulty in moving particles from solution through the needle/catheter, as the particles tend to settle and otherwise escape pick-up (pick-up). Expulsion of the solution of particles in aqueous (aqueous) solvent through small pore openings is observed; the solvent tends to be preferentially removed from the applicator, leaving an excess of particles that cannot be removed from the applicator, or that clog the applicator, or in some cases can be removed (but only by using an undesirably high force that is not appropriate for the average user handling the hand-held syringe). However, the addition of the penetrant aids in the viscosity and/or thixotropic behavior, which aids in emptying the particles from the applicator.
Embodiments of the invention include the addition of an osmotic agent to the plurality of xerogel/hydrogel particles. Examples of such agents include salts and polymers. Embodiments include polymers, linear polymers and hydrophilic polymers, or combinations thereof. Embodiments include polymers having a molecular weight of about 500 to about 100,000; the skilled artisan will immediately appreciate that all ranges and values within the explicitly stated ranges are contemplated, for example, from about 5000 to about 50,000 molecular weights. Embodiments include an osmotic agent concentration of, for example, about 1% to about 50% w/w; the skilled person will immediately understand that all ranges and values within the explicitly stated ranges are contemplated, e.g. 10% -30%. The agents and hydrogels may be introduced into a patient and may be part of a kit for the same.
Precursor body
The matrix may be prepared and used for particles comprising water-soluble biological agents. Thus, embodiments are provided herein for making implantable matrices. Such substrates include substrates having a porosity greater than about 20% v/v; the skilled person will immediately understand that all ranges and values within the explicitly stated ranges are contemplated. The precursor may be dissolved in an organic solvent to produce an organogel. Organogels are amorphous, non-glassy solid materials composed of a liquid organic phase entrapped in a three-dimensional crosslinked network. The liquid may be, for example, an organic solvent, mineral oil, or vegetable oil. The solubility and size (dimension) of the solvent are important characteristics for the elastic properties and robustness of the organogel. Alternatively, the precursor molecules themselves may be capable of forming their own organic matrix, eliminating the need for a third organic solvent. The term precursor refers to the component that becomes part of the crosslinked matrix. The polymer that becomes cross-linked into the matrix is a precursor, while the salt or protein that is only present in the matrix is not a precursor.
Removal of the solvent (if used) from the organogel provides a xerogel, i.e., a dried gel. Xerogels formed by, for example, freeze-drying can have a high porosity (at least about 20%), a large surface area, and a small pore size. Xerogels made from hydrophilic materials form hydrogels when exposed to aqueous solutions. Xerogels of high porosity hydrate faster than more dense xerogels. Hydrogels are materials that are insoluble in water and retain a significant fraction (over 20%) of the water within their structure. In practice, water contents of more than 90% are often known. Hydrogels can be formed by cross-linking water-soluble molecules to form a network of essentially infinite molecular weight. Hydrogels with high water content are typically soft, pliable materials. Hydrogels and drug delivery systems as described in U.S. publication nos. 2009/0017097, 2011/0142936, and 2012/0071865 may be suitable for use with the materials and methods herein by following the guidance provided herein; these references are hereby incorporated by reference herein for all purposes and, in case of conflict, the present specification will control.
Organogels and hydrogels can be formed from natural, synthetic, or biosynthetic polymers. Natural polymers may include glycosaminoglycans, polysaccharides, and proteins. Some examples of glycosaminoglycans include dermatan sulfate, hyaluronic acid, chondroitin sulfate, chitin, heparin, keratan sulfate, cutin sulfate, and derivatives thereof. Typically, glycosaminoglycans are extracted from natural sources and purified and derivatized. However, they may also be synthetically produced or synthesized by modifying microorganisms, such as bacteria. These materials can be synthetically altered from a naturally soluble state to a partially soluble or water swellable or hydrogel state. This modification can be achieved by a variety of well-known techniques, for example by conjugation or substitution of ionizable or hydrogen-bondable functional groups, such as carboxyl and/or hydroxyl or amine groups, with other more hydrophobic groups.
For example, carboxyl groups on hyaluronic acid may be esterified by an alcohol to reduce the solubility of hyaluronic acid. Such methods are used by various manufacturers of hyaluronic acid products (e.g., Genzyme core, Cambridge, MA) to produce hyaluronic acid-based sheets (sheets), fibers, and fabrics, which form hydrogels. Other natural polysaccharides such as carboxymethylcellulose or oxidized regenerated cellulose, natural gums, agar, agarose, sodium alginate, carrageenan, fucoidan, furcellaran, laminarin, hypnea, eucheuma, acacia, gum ghatti, karaya, tragacanth, locust bean, arabinogalactan, pectin, pullulan, gelatin, hydrocolloids such as carboxymethylcellulose or alginate cross-linked with polyols such as propylene glycol, and the like, also form hydrogels when in contact with aqueous environments.
The synthetic organogel or hydrogel may be biostable or biodegradable. Examples of biostable hydrophilic polymeric materials are poly (hydroxyalkyl methacrylates), poly (electrolyte complexes), poly (vinyl acetate) crosslinked with hydrolyzable or otherwise degradable linkages, and water swellable N-vinyl lactams. Other hydrogels include those referred to asHydrophilic hydrogels of (i) acid carboxyl polymers (Carbomer resin is a high molecular weight, allyl pentaerythritol crosslinked, acrylic acid based polymer modified with C10-C30 alkyl acrylate), polyacrylamides, polyacrylic acid, starch graft copolymers, acrylate polymers, ester crosslinked polydextrose (polyglucan). Such hydrogels are described, for example, in U.S. patent No.3,640,741 to ets, U.S. patent No.3,865,108 to Hartop, U.S. patent No.3,992,562 to Denzinger et al, U.S. patent No.4,002,173 to Manning et al, U.S. patent No.4,014,335 to Arnold, and U.S. patent No.4,207,893 to Michaels, all of which are incorporated herein by reference, where in the event of conflict, the present specification governs.
Hydrogels and organogels can be made from precursors. The precursors are not hydrogels/organogels, but rather are crosslinked to each other to form a hydrogel/organogel. The crosslinks may be formed by covalent or physical bonds. Examples of physical bonds are ionic bonds, hydrophobic associations of segments of precursor molecules, and crystallization of segments of precursor molecules. The precursors can be triggered to react to form a crosslinked hydrogel. The precursor may be polymerizable and include a crosslinking agent, which is often, but not always, a polymerizable precursor. Polymerizable precursors are thus precursors having functional groups that react with each other to form a polymer and/or matrix made of repeating units. The precursor may be a polymer.
Some precursors therefore react by chain-growth polymerization (also known as polyaddition) and involve crosslinking together of monomers incorporating double or triple chemical bonds. These unsaturated monomers have additional internal bonds that can be cleaved and linked to other monomers to form repeating chains. Monomers are polymerizable molecules having at least one group that reacts with other groups to form polymers. A macromer (or macromer) is a polymer or oligomer having at least one reactive group, often terminal, which enables it to function as a monomer; each macromonomer molecule is attached to the polymer through the reaction of a reactive group. Thus, macromonomers having two or more monomers or other functional groups tend to form covalent crosslinks. In the manufacture of, for example, polypropylene or polyvinyl chloride, addition polymerization is involved. One type of addition polymerization is living polymerization.
Some precursors therefore pass through a condensation polymerization reaction that occurs when the monomers are bonded together by a condensation reaction. Typically, these reactions are achieved by reacting molecules that incorporate alcohol, amine, or carboxylic acid (or other carboxylic derivative) functionality. When an amine reacts with a carboxylic acid, amide or peptide bonds are formed and water is released. Some condensation reactions are substituted according to nucleophilic acyl groups, for example, as in U.S. patent No.6,958,212, which is hereby incorporated by reference in its entirety to the extent that it does not contradict the explicit disclosure herein.
Some precursors react by a chain growth mechanism. A chain-growth polymer is defined as a polymer formed by the reaction of a monomer or macromer with a reactive center. A reactive center is a specific location within a compound that is an initiator of a reaction chemically involved therein. In chain-growth polymer chemistry, this is also the point for propagation of the growing chain. The reactive center is typically free-radical, anionic, or cationic in nature, but may take other forms as well. Chain extension systems include free radical polymerization, which involves processes of initiation, propagation, and termination. Initiation is the generation of free radicals necessary for propagation, as produced by free radical initiators such as organic peroxide molecules. Termination occurs when the free radicals react in a manner that prevents further propagation. The most common termination method is by coupling (coupling), in which two radical species react with each other to form a single molecule.
Some precursors react by a step-growth mechanism and are polymers formed by a step-reaction between functional groups of monomers. Most step-growth polymers are also classified as condensation polymers, but not all step-growth polymers release condensates.
The monomer may be a polymer or a small molecule. A polymer is a high molecular weight molecule formed by combining many small molecules (monomers) in a regular pattern. Oligomers are polymers having less than about 20 monomer repeat units. Small molecules generally refer to molecules less than about 2000 daltons.
The precursor may thus be a small molecule such as acrylic acid or vinyl caprolactam, a larger molecule containing polymerizable groups such as acrylate-terminated polyethylene glycol (PEG-diacrylate), or other polymers containing ethylenically unsaturated groups such as those described below: U.S. Pat. No.4,938,763 to Dunn et al, U.S. Pat. Nos. 5,100,992 and 4,826,945 to Cohn et al, or U.S. Pat. Nos. 4,741,872 and 5,160,745 to Deluca et al, each of which is hereby incorporated by reference in its entirety to the extent that it does not contradict the express disclosure herein.
To form a covalently crosslinked hydrogel, the precursors must be covalently crosslinked together. Typically, a polymeric precursor is a polymer that will bind to other polymeric precursors at two or more points, where each point is a link to the same or different polymer (link). Precursors having at least two reactive centers (e.g., in free radical polymerization) can be used as crosslinkers because each reactive group can participate in the formation of a different growing polymer chain. In the case of functional groups without reactive centers, where crosslinking requires three or more such functional groups on at least one of the precursor types. For example, many electrophilic-nucleophilic reactions consume both electrophilic and nucleophilic functional groups, such that a third functional group is required for the precursor to form the crosslink. Such precursors may thus have three or more functional groups and may be crosslinked by precursors having two or more functional groups. The cross-linked molecules may be cross-linked via ionic or covalent bonds, physical forces, or other attractive forces. However, covalent crosslinking will typically provide stability and predictability in the structure of the reaction product.
In some embodiments, each precursor is multifunctional, meaning that it includes two or more electrophilic or nucleophilic functional groups, such that a nucleophilic functional group on one precursor can react with an electrophilic functional group on another precursor to form a covalent bond. At least one of the precursors includes more than two functional groups such that the precursors combine to form a crosslinked polymeric product as a result of an electrophilic-nucleophilic reaction.
The precursor may have a biologically inert and hydrophilic moiety, e.g., a core. In the case of branched polymers, the core refers to the adjacent portion of the molecule that is bound to arms extending from the core, where the arms have functional groups, often at the ends of the branches. The hydrophilic precursor or precursor portion has a solubility in aqueous solution of at least 1g/100 mL. The hydrophilic moiety can be, for example, a polyether, such as a polyalkylene oxide, e.g., polyethylene glycol (PEG), polyethylene oxide (PEO), polyethylene oxide-co-polypropylene oxide (PPO), co-ethylene oxide block or random copolymers, and polyvinyl alcohol (PVA), poly (vinyl pyrrolidone) (PVP), polyamino acids, dextran, or proteins. The precursor may have polyalkylene glycol moieties and may be polyethylene glycol-based, wherein at least about 80% or 90% by weight of the polymer comprises polyethylene oxide repeats. The polyethers and more particularly poly (alkylene oxide) or poly (ethylene glycol) or polyethylene glycol are generally hydrophilic. As is customary in these areas, the term PEG is used to refer to PEO with or without hydroxyl end groups.
The precursor may also be a macromolecule (or macromer), which is a molecule having a molecular weight in the range of one thousand to many millions. However, in some embodiments, at least one of the precursors is a small molecule of about 1000Da or less. A large molecule, preferably at least five to fifty times as large in molecular weight as a small molecule of about 1000Da or less and preferably less than about 60,000Da when reacted in combination with the small molecule; the skilled person will immediately understand that all ranges and values within the explicitly stated ranges are contemplated. More preferred ranges are macromolecules having a molecular weight of about seven to about thirty times as large as the crosslinking agent, and most preferred ranges are about ten to twenty times the difference in molecular weight. In addition, a molecular weight of 5,000-50,000 is useful, such as a molecular weight of 7,000-40,000 or a molecular weight of 10,000-20,000.
Some of the macromer precursors are crosslinkable, biodegradable, water-soluble macromers described in U.S. Pat. No.5,410,016 to Hubbell et al, which is hereby incorporated by reference in its entirety to the extent that it does not contradict the explicit disclosure. These macromers are characterized by having at least two polymerizable groups separated by at least one degradable region.
Synthetic precursors may be used. Synthetic refers to molecules not found in nature or not normally found in humans. Some synthetic precursors do not contain amino acids or amino acid sequences that occur in nature. Some synthetic precursors are polypeptides that are not found in nature or are not normally found in the human body, e.g., dimeric, trimeric, or tetrameric-lysine. Some synthetic molecules have amino acid residues, but only have adjacent one, two, or three, where the amino acids or clusters thereof are separated by non-natural polymers or groups. Polysaccharides or their derivatives are therefore not synthetic.
Alternatively, natural proteins or polysaccharides may be suitable for use in these methods, such as collagen, fibrin (ogen), albumin, alginate, hyaluronic acid, and heparin. These natural molecules may further include chemical derivatization, e.g., synthetic polymer decoration (decotion). The natural molecules may be crosslinked via their natural nucleophiles or after they are derivatized with functional groups, for example, as in U.S. patent nos. 5,304,595, 5,324,775, 6,371,975, and 7,129,210, each of which is hereby incorporated by reference to the extent that it does not contradict the explicit disclosure herein. Natural refers to molecules found in nature. Natural polymers, such as proteins or glycosaminoglycans such as collagen, fibrinogen, albumin, and fibrin, can be crosslinked using reactive precursor species having electrophilic functional groups. Natural polymers commonly found in the body are proteolytically degraded by proteases present in the body. Such polymers may be reacted or derivatized with activatable functional groups via functional groups on their amino acids, such as amine, thiol, or carboxyl groups. Although natural polymers can be used in hydrogels, their gelation time and final mechanical properties must be controlled by appropriate introduction of additional functional groups and selection of appropriate reaction conditions such as pH.
The precursor can be made to have hydrophilic portions, provided that the resulting hydrogel retains the desired amount of water, e.g., at least about 20%. In some cases, the precursor is still soluble in water because it also has a hydrophilic moiety. In other cases, the precursor is dispersed in water (suspension), but is still reactive to form a crosslinked material. Some hydrophobic moieties may include multiple alkyl groups, polypropylene, alkyl chains, or other groups. Some precursors having hydrophobic moieties are sold under the tradenames PLURONICF68, JEFFAMINE or TECTRONIC. The hydrophobic portion of the hydrophobic molecule or copolymer, etc., is a hydrophobic molecule or hydrophobic portion: which is sufficiently hydrophobic to cause aggregation of molecules (e.g., polymers or copolymers) to form micelles or microphases comprising hydrophobic domains in an aqueous continuous phase; or which when tested alone is sufficiently hydrophobic to precipitate, or otherwise change phase, from an aqueous solution of water at a pH of about 7 to about 7.5 at a temperature of about 30 to about 50 degrees celsius while within the aqueous solution.
The precursors may have, for example, 2-100 arms, with each arm having a terminal end, bearing in mind that some precursors may be dendrimers or other highly branched materials. An arm on a hydrogel precursor refers to a straight chain of chemical groups that attach a crosslinkable functional group to the polymer core. Some embodiments are precursors having 3-300 arms; the skilled person will immediately understand that all ranges and values within the explicitly stated ranges are contemplated, e.g. 4-16, 8-100, or at least 6 arms.
Thus, a hydrogel can be made, for example, from a multi-armed precursor having a first set of functional groups and a low molecular weight precursor having a second set of functional groups. For example, a six-or eight-arm precursor can have hydrophilic arms, e.g., polyethylene glycol terminated with a primary amine, wherein the arms have a molecular weight of about 1,000 to about 40,000; the skilled person will immediately understand that all ranges and values within the explicitly stated ranges are contemplated. Such precursors can be mixed with relatively small precursors, such as molecules having a molecular weight of about 100 to about 5000, or not more than about 800, 1000, 2000, or 5000 having at least about three functional groups, or about 3 to about 16 functional groups; those of ordinary skill will understand that all ranges and values between these explicitly stated values are contemplated. Such small molecules may be polymeric or non-polymeric and may be natural or synthetic.
Precursors other than dendrimers may be used. Dendrimers are highly branched, radiation-symmetric polymers in which atoms are arranged in a number of arms and sub-arms radiating out from a central core. Dendrimers are characterized by their structural perfection (as evaluated based on both symmetry and polydispersity) and require specific chemical methods to synthesize. Thus, the skilled person can easily distinguish between branched and non-branched macromolecular precursors. Dendrimers have a shape that is typically dependent on the solubility of its constituent polymers in a given environment, and can vary significantly depending on the changes in the surrounding solvents and solutes, e.g., temperature, pH, or ionic content.
The precursor may be a dendrimer, for example, as in U.S. publication nos. 2004/0086479 and 2004/0131582 and PCT publication nos. WO07005249, WO07001926, and WO06031358, or their U.S. counterparts; dendrimers may also be useful as multifunctional precursors, for example, as in U.S. publication nos. 2004/0131582 and 2004/0086479 and PCT publication nos. WO06031388 and WO 06031388; each of the U.S. and PCT applications are hereby incorporated by reference in their entirety to the extent that they do not contradict the explicit disclosure herein. Dendrimers are highly ordered, have a high surface area to volume ratio, and exhibit many terminal groups for potential functionalization. Embodiments include multifunctional precursors that are not dendrimeric macromolecules.
Some embodiments include precursors consisting essentially of oligopeptide sequences of no more than five residues, e.g., amino acids including at least one amine, thiol, carboxyl, or hydroxyl side chain. Residues are amino acids, as occur in nature or derived therefrom. The backbone of such oligopeptides may be natural or synthetic. In some embodiments, a peptide of two or more amino acids is combined with a synthetic backbone to make a precursor; some embodiments of such precursors have a molecular weight in the range of from about 100 to about 10,000 or from about 300 to about 500. The skilled person will immediately understand that all ranges and values between these explicitly stated ranges are contemplated.
The precursor can be prepared free of amino acid sequences cleavable by enzymes present at the site of introduction, including free of sequences susceptible to attachment by metalloproteases and/or collagenases. In addition, precursors can be made to contain no amino acids, or to contain no more than about 50, 30, 20, 10, 9, 8,7, 6, 5,4, 3, 2, or 1 amino acid sequence. Precursors may be non-proteins, meaning that they are not naturally occurring proteins and cannot be made by cleaving naturally occurring proteins and cannot be made by adding synthetic materials to proteins. The precursors may be non-collagens, non-fibrous proteins, non-fibrinogen and non-albumin, meaning that they are not one of these proteins and are not chemical derivatives of one of these proteins. The use of non-protein precursors and limited use of amino acid sequences may be useful to avoid immune responses, avoid unwanted cellular recognition, and avoid the risks associated with the use of proteins derived from natural sources. The precursor can also be non-saccharide (free of saccharide) or substantially non-saccharide (free of saccharide in excess of about 5% weight/weight of the molecular weight of the precursor.
Peptides may be used as precursors. Generally, peptides having less than about 10 residues are preferred, although larger sequences (e.g., proteins) can be used. The skilled artisan will immediately appreciate that each range and value included within these explicit ranges, for example, 1-10, 2-9, 3-10, 1, 2, 3, 4,5, 6, or 7, is included. Some amino acids have nucleophilic groups (e.g., primary amines or thiols) or groups that can be derivatized to introduce nucleophilic or electrophilic groups (e.g., carboxyl or hydroxyl groups) if desired. Synthetically produced polyamino acid polymers are generally considered synthetic if they are not found in nature and are engineered to be different from naturally occurring biomolecules.
Some organogels and hydrogels are made with precursors containing polyethylene glycol. Polyethylene glycol (PEG, also known as polyethylene oxide when present at high molecular weight) refers to a polymer having repeating groups (CH) where n is at least 32CH2O)nThe polymer of (1). Polymeric precursors having polyethylene glycol thus have at least three of these repeating groups connected to each other in a linear series. The polyethylene glycol content of a polymer or arm is calculated by summing all polyethylene glycol groups on the polymer or arm, even if they are interrupted by other groups. Thus, an arm with at least 1000MW of polyethylene glycol has sufficient CH2CH2O groups to a total of at least 1000 MW. As is customary in these arts, polyethylene glycol polymers do not necessarily refer to molecules that end-cap in a hydroxyl group. The molecular weight is abbreviated in thousand using the symbol K, for example 15K means 15,000 molecular weight, i.e. 15,000 daltons. SG refers to succinimidyl glutarate. SS refers to succinimidyl succinate. SAP refers to succinimidyl adipate. SAZ refers to succinimidyl azelate. SS, SG, SAP and SAZ are succinimidyl esters having ester groups that degrade by hydrolysis in water. Hydrolytically degradable thus refers to a material that spontaneously degrades in excess water in vitro without the presence of any enzymes or cells to mediate degradation. The degradation time refers to the effective disappearance of the material as judged by the naked eye. Tripolylysine (also abbreviated as LLL)) Is a synthetic tripeptide. PEG and/or hydrogels, and compositions comprising the same, may be provided in a pharmaceutically acceptable form, meaning that they are highly purified and free of contaminants such as pyrogens.
Functional group
The precursors used for covalent cross-linking have functional groups that react with each other to form a material, either externally to the patient, or in situ. The functional groups typically have polymerizable groups for polymerization or are reactive with each other in an electrophile-nucleophile reaction or are configured to participate in other polymerization reactions. Various aspects of the polymerization reaction are discussed in the precursors section herein.
Thus, in some embodiments, the precursor has a polymerizable group that is activated by, for example, a photoinitiating or redox system as used in the polymerization art, or an electrophilic functional group that is a carbodiimide (carbodiimidazole), sulfonyl chloride, chlorocarbonate, n-hydroxysuccinimide, succinimidyl, or sulfonamido (sulfonamido) ester, or as in U.S. Pat. No.5,410,016 or 6,149,931, each of which is hereby incorporated herein by reference in its entirety to the extent that they do not contradict the explicit disclosure herein. Nucleophilic functional groups can be, for example, amines, hydroxyl groups, carboxyl groups, and thiols. Another type of electrophile is an acyl group, for example, as in U.S. patent No.6,958,212, which describes, among other things, the Michael (Michael) addition scheme for reacting polymers.
Some functional groups, such as alcohols or carboxylic acids, are generally unreactive with other functional groups, such as amines, under physiological conditions (e.g., pH 7.2-11.0, 37 ℃). However, such functional groups can be made more reactive by using activating groups such as N-hydroxysuccinimide. Some activating groups include carbonyldiimidazole, sulfonyl chlorides, aryl halides, sulfosuccinimidyl esters, N-hydroxysuccinimidyl esters, succinimidyl esters, epoxides, aldehydes, maleimides, imidoesters, and the like. N-hydroxysuccinimide ester or N-hydroxysulfosuccinimide (NHS) groups are useful groups for the crosslinking of proteins or amine-containing polymers such as amino-terminated polyethylene glycol. The NHS-amine reaction has the advantage that the reaction kinetics are favourable, but the gelation rate can be adjusted by pH or concentration. The NHS-amine crosslinking reaction results in the formation of N-hydroxysuccinimide as a by-product. The sulfonated or ethoxylated forms of N-hydroxysuccinimide have relatively increased solubility in water and therefore their rapid clearance from the body. The NHS-amine crosslinking reaction can be carried out in aqueous solution and in the presence of a buffer such as a phosphate buffer (pH 5.0-7.5), triethanolamine buffer (pH 7.5-9.0), or borate buffer (pH 9.0-12), or sodium bicarbonate buffer (pH 9.0-10.0). Due to the reaction of NHS groups with water, it is preferred to make an aqueous solution of NHS-based crosslinker and functional polymer just prior to the crosslinking reaction. The reaction rate of these groups can be retarded by keeping the solutions at a lower pH (pH 4-7). Buffers may also be included in the hydrogel introduced into the body.
In some embodiments, each precursor includes only nucleophilic functional groups or only electrophilic functional groups, so long as both nucleophilic and electrophilic precursors are used in the crosslinking reaction. Thus, for example, if the crosslinking agent has nucleophilic functional groups such as amines, the functional polymer may have electrophilic functional groups such as N-hydroxysuccinimide. On the other hand, if the crosslinker has electrophilic functional groups such as sulfosuccinimide, the functional polymer may have nucleophilic functional groups such as amines or thiols. Thus, functional polymers such as proteins, poly (allylamine), or amine-terminated di-or polyfunctional poly (ethylene glycol) may be used.
One embodiment has reactive precursor species with 3-16 nucleophilic functional groups each and reactive precursor species with 2-12 electrophilic functional groups each; the skilled person will immediately understand that all ranges and values within the explicitly stated ranges are contemplated.
The functional group can be, for example, an electrophile that can react with a nucleophile, a group that can react with a particular nucleophile, such as a primary amine, a group that forms an amide bond with a material in a biological fluid, a group that forms an amide bond with a carboxyl group, an activated acid functional group, or a combination thereof. The functional group can be, for example, a strong electrophilic functional group, meaning an electrophilic functional group that effectively forms a covalent bond with a primary amine in aqueous solution at pH9.0 at room temperature and pressure and/or an electrophilic group that reacts by a michael-type reaction. The strong electrophile may be of a type that does not participate in a michael-type reaction or of a type that participates in a michael-type reaction.
The Michael type reaction refers to the 1,4 addition reaction of a nucleophile to a conjugated unsaturated system. The addition mechanism can be purely polar or by an intermediate state like a radical; lewis acids or appropriately designed hydrogen bonding species may act as catalysts. The term conjugated may refer to either carbon-carbon, carbon-heteroatom, heteroatom-heteroatom multiple bonds alternating with single bonds, or functional groups attached to macromolecules such as synthetic polymers or proteins. The michael type reaction is discussed in detail in U.S. patent No.6,958,212, which is hereby incorporated by reference in its entirety for all purposes to the extent that it does not contradict the explicit disclosure herein.
Examples of strong electrophiles that do not participate in the michael-type reaction are: succinimide, succinimidyl ester, or NHS-ester. Examples of michael-type electrophiles are acrylates, methacrylates, methyl methacrylate, and other unsaturated polymerizable groups.
Initiating system
Some precursors are reacted using an initiator. An initiator group is a chemical group capable of initiating a free radical polymerization reaction. For example, it may be present as a separate component, or as a pendant group on a precursor. Initiator groups include thermal initiators, photoactivatable initiators, and oxidation-reduction (redox) systems. Long wave UV and visible light photoactivated initiators include, for example, ethyl eosin groups, 2-dimethoxy-2-phenylacetophenone groups, other acetophenone derivatives, thioxanthone groups, benzophenone groups, and camphorquinone groups. Examples of thermally reactive initiators include 4, 4' azobis (4-cyanovaleric acid) groups, and the like of benzoyl peroxide groups. Several commercially available low temperature free radical initiators may be used, such as V-044 available from Wako Chemicals USA, inc., Richmond, Va., to initiate a free radical crosslinking reaction at body temperature to form a hydrogel coating with the aforementioned monomers.
The metal ions can be used as an oxidizing or reducing agent in redox initiation systems. For example, ferrous ions may be used in combination with peroxides or hydroperoxides to initiate polymerization, or as part of a polymerization system. In this case, ferrous ions will be used as the reducing agent. Alternatively, metal ions may be used as the oxidizing agent. For example, the eerie ion (the 4+ valence state of cerium) interacts with various organic groups (including carboxylic acids and carbamates) to move electrons to the metal ion and leave an initiating radical on the organic group. In such systems, the metal ion acts as an oxidizing agent. Potentially suitable metal ions for either role are any transition metal ions, lanthanides and actinides having at least two readily accessible oxidation states. Particularly useful metal ions have at least two states separated by only one difference in charge. Among these, ferric/ferrous is most commonly used; divalent copper/cuprous copper; high cerium/trivalent cerium; trivalent cobalt/divalent cobalt; vanadate V vs IV; a permanganate salt; and trivalent/divalent manganese. Peroxy (peroxygen) -containing compounds may be used, such as peroxides and hydroperoxides, including hydrogen peroxide, t-butyl hydroperoxide, t-butyl peroxide, benzoyl peroxide, cumyl peroxide.
An example of an initiating system is a combination of a peroxy compound in one solution and a reactive ion such as a transition metal in another solution. In this case, when the moieties containing two complementary reactive functional groups interact at the application site, no external polymerization initiator is required and the polymerization proceeds spontaneously and without the application of external energy or the use of an external energy source.
Visualization reagent
The visualisation agent may be used as a powder in a xerogel/hydrogel; which reflects or emits light at a wavelength detectable by the human eye such that a user applying the hydrogel can view the subject when the subject contains an effective amount of the agent. Agents that require mechanical assistance for imaging are referred to herein as imaging agents, and examples include: radiopaque contrast agents and ultrasound contrast agents.
Some biocompatible visualization agents are FD & C BLUE #1, FD & C BLUE #2, and methylene BLUE. These agents are preferably present in the final electrophilic-nucleophilic reactive precursor substance mixture at a concentration in excess of 0.05mg/ml and preferably in the range of at least 0.1 to about 12mg/ml, and more preferably in the range of 0.1 to 4.0mg/ml, although greater concentrations up to the limit of solubility of the visualization agent can potentially be used. The visualization agent may be covalently attached to the molecular network of the xerogel/hydrogel, thus remaining visualized after application to the patient until the hydrogel hydrolyzes for dissolution.
The visualization agent may be selected from any of a variety of non-toxic coloring substances suitable for use in medical implantable medical devices, such as FD & C BLUE dyes 3 and 6, eosin, methylene BLUE, indocyanine green, or coloring dyes commonly found in synthetic surgical sutures. Reactive visualization agents such as NHS-fluorescein can be used to introduce the visualization agent into the molecular network of the xerogel/hydrogel. The visualization agent may be present with a reactive precursor species such as a cross-linking agent or a solution of a functional polymer. Preferred coloring substances may or may not become chemically bound to the hydrogel. The visualization agent may be used in small amounts, e.g., concentrations of 1% weight/volume, more preferably less than 0.01% weight/volume, and most preferably less than 0.001% weight/volume; the skilled person will immediately understand that all ranges and values within the explicitly stated ranges are contemplated. The agent tends to label the location of the particles and provide an indication of their presence and dissolution rate.
Biodegradation
Xerogels may be formed from organogels such that upon hydration in a physiological solution, a hydrogel is formed which is water degradable, such as measurable by: the hydrogels lose their mechanical strength in excess water in vitro and eventually dissipate through hydrolytic degradation of water-degradable groups. This test predicts hydrolysis-driven dissolution in vivo, which is the opposite process to cell or protease-driven degradation. Notably, however, polyanhydrides or other conventionally used degradable materials that degrade acidic components tend to cause inflammation in tissue. However, hydrogels may exclude such materials, and may be free of polyanhydrides, anhydride linkages, or precursors that degrade to acids or diacids. The term degradation by solvation in water, also called dissolution in water, refers to the process of gradual dissolution of the matrix, which is a process that cannot occur for covalently cross-linked materials and materials that are insoluble in water.
For example, electrophilic groups such as SG (N-hydroxysuccinimidyl glutarate), SS (N-hydroxysuccinimidyl succinate), SC (N-hydroxysuccinimidyl carbonate), SAP (N-hydroxysuccinimidyl adipate) or SAZ (N-hydroxysuccinimidyl azelate) may be used and have hydrolytically unstable ester linkages. More linear hydrophobic linkages such as pimelate, suberate, azelate or sebacate linkages may also be used, where these linkages are less hydrolyzable than succinate, glutarate or adipate linkages. Branched, cyclic, or other hydrophobic linkages may also be used. Polyethylene glycol and other precursors can be prepared using these groups. When using water-degradable materials, the crosslinked hydrogel degradation can be carried out by water-driven hydrolysis of the biodegradable segments. Polymers comprising ester linkages may also be included to provide a desired degradation rate, with groups added or subtracted near the ester to increase or decrease the degradation rate. Thus, hydrogels having desired degradation profiles from days to many months can be constructed using degradable segments. For example, if polyglycolic acid ester is used as the biodegradable segment, the crosslinked polymer may be degraded in about 1 to about 30 days depending on the crosslink density of the network. Similarly, the polycaprolactone-based crosslinked network can be degraded in about 1 to about 8 months. The degradation time generally varies according to the type of degradable segment used in the following order: polyglycolide < polylactate < polytrimethylene carbonate < polycaprolactone. Thus, hydrogels having desired degradation profiles from days to many months can be constructed using degradable segments.
The biodegradable linkages in the organogel and/or xerogel and/or hydrogel and/or precursor may be water-degradable or enzymatically degradable. Illustrative water-degradable biodegradable linkages include polymers, copolymers and oligomers of glycolide, dl-lactide, 1-lactide, dioxanone, esters, carbonates and trimethylene carbonate. Illustrative enzymatically biodegradable linkages include peptide linkages cleavable by metalloproteases and collagenases. Examples of biodegradable linkages include the following polymers and copolymers: poly (hydroxy acids), poly (orthocarbonates), poly (anhydrides), poly (lactones), poly (amino acids), poly (carbonates), and poly (phosphonates).
If it is desired that the biocompatible cross-linked matrix be biodegradable or absorbable, one or more precursors having biodegradable linkages present between functional groups may be used. The biodegradable linkages may also optionally be used as a water soluble core for one or more of the precursors used to make the matrix. For each pathway, the biodegradable linkage can be selected such that the resulting biodegradable, biocompatible crosslinked polymer will degrade or be absorbed within a desired period of time.
The matrix material may be selected such that the degradation products are absorbed into the circulatory system and substantially cleared from the body via renal filtration. The matrix material may be a hydrogel in physiological solution. One approach is to select precursors that do not decompose in the body, where the links between the precursors degrade to return to the precursor or precursors with small changes resulting from the covalent crosslinking process. This approach is in contrast to selecting biological matrix materials that are destroyed by enzymatic processes and/or materials that are eliminated by macrophages, or materials that result in byproducts that are not actually water soluble. Material cleared from the body by renal filtration can be labeled using techniques known to the skilled artisan and detected in the urine. Although there may be at least some theoretical loss of these materials to other body systems, the usual outcome of the materials is the renal clearance process. The term substantially cleared thus refers to material that is normally cleared by the kidneys.
Administration of
The application of the xerogel can be carried out directly into the site of interest. For example, a xerogel microlens (lenticule) may be applied to the cornea, or a membrane may be applied to the dermis or epidermis. The xerogel particles may be administered by inhalation. Also, powder delivery systems can be used to inject xerogel powder directly into tissue.
The application of a xerogel may also involve hydration at the point of, or just about, use. The xerogel is exposed to an aqueous solution, such as physiological saline, and allowed to absorb water to form a hydrogel. The hydrogel is implanted directly, surgically, or via injection through a syringe or catheter.
Embodiments of the invention include administration at or near the eye. The structure of the mammalian eye can be divided into three main layers or membranes: fibrous membranes, vascular membranes, and neural membranes. The fibrous membrane, also known as the ocular fibrous membrane, is the outer layer of the eyeball consisting of the cornea and the sclera. The sclera is the supporting wall of the eye and imparts to the eye most of its white color. Which extends from the cornea (the clear anterior portion of the eye) to the optic nerve at the back of the eye. The sclera is a fibrous, elastic and protective tissue, composed of closely packed collagen fibrils, containing about 70% water.
The fiber membrane is covered with a conjunctiva. The conjunctiva is a membrane that covers the sclera (the white part of the eye) and lines the interior of the eyelids. It helps lubricate the eye by producing mucus and tears, although a smaller volume of tears is produced compared to the lacrimal gland. The conjunctiva is typically divided into three parts: (a) the palpebral conjunctiva or the tarsal conjunctiva, which is the conjunctiva lining the eyelids; the palpebral conjunctiva folds back at the upper and lower fornices to become the bulbar conjunctiva, (b) the fornix conjunctiva: conjunctiva where the inner part of the eyelid and the eyeball meet, (c) bulbar conjunctiva: the conjunctiva above the sclera covering the eyeball. This region of the conjunctiva is tightly (firmly) bound and moves with the movement of the eyeball. The conjunctiva effectively surrounds, covers, and adheres to the sclera. It has cells and connective tissue, is somewhat elastic, and may be removed, excised (tease), or otherwise removed to expose a surface region of the sclera.
The vascular membrane, also known as the ocular vascular membrane, is the intermediate vascularized layer that includes the iris, ciliary body, and choroid. The choroid contains blood vessels that supply oxygen to retinal cells and remove respiratory waste products. The neural membrane, also known as the ocular neural membrane, is the internal sense that includes the retina. The retina contains the sensitive rods and cones and the associated neurons. The retina is a relatively smooth (but curved) layer. It has two points: at the two points, it is different; fovea centralis (fovea) and optic disc. The fovea is a depression in the retina directly opposite the lens, which is densely packed with cone cells. The fovea is part of the macula (macula). The fovea is largely responsible for the human color vision and enables a high acuity, which is necessary in reading. The optic disc is a point on the retina where the optic nerve penetrates the retina to connect to nerve cells within it.
The mammalian eye can also be divided into two main segments: a front section and a rear section. The anterior segment consists of the anterior chamber and the posterior chamber. The anterior chamber is located in front of the iris and behind the corneal endothelium and includes the pupil, iris, ciliary body, and aqueous humor. The posterior chamber is located behind the iris and in front of the vitreous face, with the lens and zonule fibers located between the anterior and posterior capsules in a water (aqueous humor) environment.
The cornea and lens help to focus the light rays onto the retina. The lens behind the iris is a convex, elastic disc that focuses light through a second body fluid onto the retina. It is attached to the ciliary body via a ring of zonules called the Zonule of zinn. The ciliary muscle is relaxed to focus on a distant object, which stretches the fibers connecting it with the lens, thereby flattening the lens. Light enters the eye, passes through the cornea, and enters the first of the two body fluids (aqueous humor). About two thirds of the total refractive power of the eye comes from the cornea, which has a fixed curvature. Aqueous humor is a transparent substance that connects the cornea of the eye with the lens, helps maintain the convex shape of the cornea (necessary for the convergence of light at the lens), and provides nutrients to the corneal endothelium.
The posterior segment is located behind the lens and in front of the retina. It accounts for about two thirds of the eye, which includes the anterior vitreous membrane and all structures behind it: vitreous humor, retina, c and optic nerve. On the other side of the lens is a second body fluid (vitreous fluid) which is confined on all sides as follows: lens, ciliary body, zonules, and retina. Which passes light without refraction, helps to maintain the shape of the eye and suspend the delicate (lens).
Fig. 8 shows some delivery points at or near the eye 200. Eye 200 includes sclera 212, iris 214, cornea 222, vitreous 232, zonular space 242, fovea 236, retina 238, and optic nerve 225. One area for delivery is locally at 260, where area 260 is indicated by a point on the surface of eye 200. The other region is intravitreal, as indicated by the numeral 262, or transscleral, as indicated by the numeral 264. In use, the xerogel (or gel or hydrogel or precursor thereof) is delivered into the eye, optionally through needle 268, intravitreally as at 262, or periocularly as at 272, using, for example, syringe 266, a catheter (not shown), or other device. Another area is under the conjunctiva (not shown), below the conjunctiva 211 and above the sclera 212. The drug or other therapeutic agent is released into the intraocular space. In the case of posterior ocular diseases, drugs may be targeted to the general target region 274 via periocular or intravitreal routes, where they interact with biological features to effect treatment. An embodiment is to place a xerogel in contact with retina 238 or near retina 238 without contacting it. For example, xerogels, hydrogels, and/or particles (or rods, microspheres, unitary materials, beads, or other shapes set forth herein) can be delivered to a location adjacent to retina 238 or on retina 238. The hydrogel is advantageously anchored (anchor) in the vitreous gel and does not allow diffusion of the particles. In contrast, other systems using rods or sliding microspheres do not provide anchoring and diffusion or migration in response to movement or rubbing of the eye. Placing the library at or near the retina (or other location) allows for high concentrations to be achieved at the intended site, where small particles can be used to deliver drugs for effective treatment. Conversely, a sphere, rod, or other shape that is too large to diffuse or migrate has a volume to surface ratio that is disadvantageous for effective controlled release. Another area for placement of xerogels, hydrogels, and/or particles, or other materials including the particles, is in the punctum (not shown), for example, by placing the particles in punctal plugs (silicones, polysaccharides, hydrogels, or other materials) that are inserted into the punctum of the eye.
Sites where drug delivery reservoirs may be formed in or near the eye include, inter alia, the anterior chamber, the vitreous (placed intravitreally), episcleral, in the posterior sub-tenon space (inferior fornix), subconjunctivally, on the surface of the cornea or conjunctiva. Periocular drug delivery using subconjunctival, retrobulbar, or sub-tenon's placed ocular hydrogel implants has the potential to provide a safer and improved drug delivery system to the retina compared to topical and systemic approaches.
An example of in situ placement is illustrated in fig. 9A for intravitreal implantation. In fig. 9A, the xerogel implant is injected intravitreally approximately 2.5mm behind the limbus (limbus) through the pars plana incision 390 using a subretinal cannula 392, which can be done after cutting away (disnect away) or otherwise clearing the conjunctiva when needed, as shown by the depiction of the magnifying glass 394 holding to visualize the incision 390 on the eye 310. A subretinal cannula 392 (or other suitable cannula) is then inserted through incision 390 and positioned intraocularly at a desired target site, e.g., at least one of sites 396, 398, 300 (fig. 9B), where a xerogel is introduced and then a hydrogel is formed in situ. The xerogels are formed into absorbable gels 302, 304 and/or 306, which attach to the desired target site. Particles comprising a therapeutic agent may be included in one or more gels. Notably, a nine gauge needle may be used to place the precursor. Embodiments include placement with a 25 gauge needle. Further embodiments include the use of needles having a diameter smaller than 25 gauge, such as 26, 27, 30, 31, 32 gauge.
Intravitreal in situ implant embodiments can improve the efficacy and pharmacokinetics of effective therapeutic agents in the treatment of ocular diseases and minimize patient side effects in several ways. First, the implant can be placed at a specific disease site in the vitreous cavity, bypassing local or systemic routes and thereby increasing drug bioavailability. Second, the implant maintains a local therapeutic concentration at a particular target tissue site for an extended period of time. Again, during the 12 month treatment regimen, the number of intravitreal injections will be significantly reduced, thereby reducing the patient's risk of: infection, retinal detachment, and transient visual impairment (white spots floating in the vitreous), which can occur until the drug in the vitreous migrates down the lower wall of the eye and away from the central vitreous or part of the macula.
The xerogels or xerogels hydrated into hydrogels (xerogel/hydrogel) may be placed on scleral tissue with or without the presence of conjunctiva. The xerogel/hydrogel may be attached to the sclera or other tissue near the sclera to facilitate diffusion of the drug through the desired tissue or to provide a stable reservoir to direct the therapeutic agent as desired. Hydrogel adhesives may be used, for exampleThe sealant acts as an adhesion aid. In some embodiments, the conjunctiva of the eye may be removed, macerated, dissected away, or excised so that the tissue may be lifted away from the sclera to access the specific area of the sclera for implantation or injection of the xerogel/hydrogel. The xerogel/hydrogel is placed to create a layer on and adhere to the surface. The conjunctiva may be allowed to contact the tissue if it is still present or remains sufficiently mechanically intactUnity in order to do so. In some embodiments, the xerogel/hydrogel is composed of at least 50%, 75%, 80%, 90%, or 99% weight/weight of water-soluble precursors (calculated by measuring the weight of the hydrophilic precursors and dividing by the weight of all precursors so that the weight of the water or solvent or non-hydrogel components is ignored) to promote non-adhesiveness of the hydrogel. In some embodiments, such hydrophilic precursors substantially (substantially) comprise PEO. In some embodiments, agents for reducing tissue adhesion mediated by biological mechanisms including cell mitosis, cell migration, or macrophage migration or activation are included, such as anti-inflammatory agents, antimitotic agents, antibiotics, PACLITAXEL (PACLITAXEL), MITOMYCIN (MITOMYCIN), or taxol.
In other embodiments, the sclera is substantially free of conjunctiva. The conjunctiva is a mass of important tissue that covers many or all of the sclera. The conjunctiva may be punctured or penetrated with a needle or catheter or trocar and the precursors introduced into the space between the sclera and conjunctiva. This placement of the implant is referred to as subconjunctival placement. In some cases, the conjunctiva may be punctured to access the natural potential gaps between the tissues filled with the precursor. In other cases, potential or actual gaps are created mechanically with trocars, stents, etc. that break the adhesion between the sclera and conjunctiva so that precursors can be introduced. The conjunctiva is sufficiently elastic to permit the introduction of useful amounts of xerogel or to be forced into such natural or created spaces. Thus, in some cases, the xerogel/hydrogel volume is from about 0.25 to about 5 ml; the skilled person will immediately understand that all ranges and values within the explicitly stated ranges are contemplated, e.g. about 1ml or from 0.5ml to about 1.5 ml.
Moreover, removal of the hydrogel-formed xerogel, whether present in or around the eye, is also readily achieved using a vitrectomy cutter (if the implant is located in the vitreous cavity) or a manual I/a syringe and cannula (if the implant is located on the scleral surface) or an irrigation/aspiration handpiece. This is in contrast to the primary surgical procedure required to remove some conventional non-absorbable implants.
In further embodiments, the xerogel/hydrogel material may be placed into a patient, for example, in a tissue or organ, including subcutaneous, intramuscular, intraperitoneal, in a potential space of the body, or in a natural cavity or opening. The material provides a depot for the release of the agent over time. Embodiments thus include a volume for placement of about 0.5 to about 500ml (referred to as the total volume in the case of a collection of delivered particles); the skilled person will immediately understand that all ranges and values within the explicitly stated ranges are contemplated, e.g. 1-10ml or 5-50 ml. Intraperitoneal or intramuscular injection, for example, is a useful area for extended controlled release of an agent over a period of hours, days, or weeks.
The materials described herein can be used to deliver drugs or other therapeutic agents (e.g., imaging agents or markers). One mode of application is to apply a mixture of xerogel/hydrogel particles and other materials (e.g., therapeutic agents, buffers, accelerators, initiators) to the site through a needle, cannula, catheter or hollow wire. The mixture may be delivered, for example, using a manually controlled syringe or a mechanically controlled syringe, such as a syringe pump. Alternatively, xerogel/hydrogel particles may be mixed with hydration fluid and/or other reagents at or near the site using a double or multi-tube syringe or multi-lumen (multi-lumen) system.
The xerogels may be provided to the site in a flowable form, for example, as flowable particles. The xerogel may be suspended in a liquid and applied to the site. The xerogel particles can be made to have the largest diameter for manual discharge from a syringe through a 3 to 5French catheter or a 10-30 gauge needle. The skilled person will immediately understand that all ranges and values within the explicitly stated ranges are contemplated, e.g. numbers 25-30. The use of small needles is particularly advantageous in the eye as sensitive organs. Application to other organs is also advantageous, for example, to control bleeding or other damage. The particles may be formed by: a hydrogel is produced which is then broken into smaller pieces. The material can be ground, for example, in a ball mill or with a mortar and pestle, or chopped or diced with a knife or wire. Alternatively, the material may be chopped in a blender. Another method involves forcing the organogel or the material from the gel step through a mesh, collecting the debris, and passing them through the same mesh or another mesh until the desired size is achieved, followed by the manufacture of a xerogel. The xerogel/hydrogel may contain particles loaded with a therapeutic agent. Some or all of the hydrogel particles may contain particles loaded with a therapeutic agent. In some embodiments, a first population of therapeutic agent-loaded particles loaded with a first therapeutic agent is included within a first population of xerogel particles, and a second population of therapeutic agent-loaded particles loaded with a second therapeutic agent is included within a second population of xerogel particles. In this manner, multiple agents may be released from a single implant. Embodiments of particles include those having a particular shape, such as a sphere, rod, or disk.
Embodiments include the placement of a plurality of xerogel/hydrogel particles. The xerogel/hydrogel particles may include a therapeutic agent, for example a protein such as anti-VEGF. The particles may be sized for manual passage through a 27 gauge or smaller diameter needle. Pressure for forcing the particles through the needle may be provided manually.
An alternative for the delivery of particles is to pre-form the gel into shaped particles and then introduce the material into the body. For example, the xerogel/hydrogel may be formed into a sphere, rod, cylinder, or other shape. Embodiments include solid rods of xerogels/hydrogels for subcutaneous implantation and delivery of one or more agents.
Xerogels/hydrogels as set forth herein may be used for tissue expansion. The use of collagen for skin augmentation is well known. Xerogels/hydrogels, such as particles, can be used for dermal fillers or for tissue augmentation. Embodiments include injecting or otherwise placing a plurality of particles in tissue, or forming a hydrogel in situ. The material may be injected or otherwise placed at the desired site.
A xerogel/hydrogel as set forth herein may be used to separate tissues to reduce the dose of radioactivity received by one of the tissues. The spacer material may be placed in the patient as set forth in U.S. patent No.7,744,913, which is hereby incorporated by reference for all purposes, where the present specification dictates, in case of conflict. Some embodiments are methods comprising: the spacer is introduced to a location between the first tissue location and the second tissue location to increase a distance between the first tissue location and the second tissue location. Further, there may be the step of administering a dose of radiation energy to at least the first tissue site or the second tissue site. For example, a method is to deliver a therapeutic dose of radiation to a patient, comprising introducing a biocompatible, biodegradable particulate xerogel, e.g., an aggregate of particles optionally with radiopaque content, between a first tissue site and a second tissue site to increase the distance between the first tissue site and the second tissue site, and treating the second tissue site with the therapeutic dose of radiation such that the presence of the filler device results in the first tissue site receiving a lesser dose of radiation than the first tissue site would receive without the presence of the spacer. The spacer may be introduced as a xerogel that forms a hydrogel in the patient that is removed by biodegradation of the spacer-hydrogel in the patient. An example is where the first tissue location is associated with the rectum and the second tissue location is associated with the prostate. The amount of reduction of the radiation may vary. Embodiments include at least about 10% to about 90%; the skilled artisan will immediately appreciate that all ranges and values within the explicitly stated ranges are contemplated, for example at least about 50%. The radiation may alternatively be directed to a third tissue such that the first tissue or the second tissue receives a lower amount of radiation as a result of its separation from other tissue. The first tissue and the second tissue may be adjacent to each other in the body, or may be separated from each other by other tissue. The spacer volume for separating the tissues depends on the configuration of the tissue to be treated and the tissues to be separated from each other. In many cases, a volume of about 20 cubic centimeters (cc or ml) is suitable. In other embodiments, as little as about 1cc may be required. Other volumes are in the range of about 5-1000 cc; the skilled person will immediately understand that all ranges and values within the explicitly stated ranges are contemplated, e.g. 10-30 cc. In some embodiments, the spacer is administered in two doses at different times to allow the tissue to stretch and accommodate the spacer and thereby receive a larger spacer volume than would otherwise be readily possible. The tissue to be separated by the spacer includes, for example, at least one of rectum, prostate and chest, or a portion thereof. For example, a first portion of the chest may be separated from a second portion.
Reagent kit
A kit or system for making a hydrogel from a xerogel can be prepared such that the xerogel is stored in the kit and the hydrogel is made when needed for use on a patient. Furthermore, a kit can be made for applying the xerogel in the form of a xerogel. The applicator may be used in combination with a xerogel and/or a hydrogel. Kits are made using medically acceptable conditions and contain components having sterility, purity, and pharmaceutically acceptable preparations. The kit may contain an applicator (when appropriate) and instructions. Xerogel particles comprising a therapeutic agent may be useful for mixing with a solution in a kit or provided separately. The xerogel component may be provided as follows: one or more containers of xerogels which form hydrogels, wherein the xerogels are in the form of a plurality of particles that are placed into the patient, or as a monolithic implant. The solvent/solution may be provided in the kit or separately, or the components may be pre-mixed with the solvent. The kit may include a syringe and/or needle for mixing and/or delivery. The kit or system may include the components set forth herein.
Some embodiments provide a single applicator, e.g., one syringe, comprising xerogel particles for delivery, wherein an aqueous solution is added to the applicator for hydration, followed by placement of the material in the patient using the syringe. The xerogel particle solvent may be substantially water, meaning that about 99% of the solvent is water volume/volume, with salts or buffers being present when desired. Other solvents that are safe and biocompatible may be used, such as dimethylsulfoxide. The xerogel particles may further comprise powders of proteins and/or other agents.
The packaging of the precursors and/or the entire kit is preferably carried out under dry conditions in the absence of oxygen. The precursor and/or kit components may be placed in a hermetically sealed container, such as a glass or metal (foil) container, that is impermeable to moisture or oxygen.
Xerogels containing protein powders or other solid phase, water-soluble biological agents can be gamma sterilized at the end of the implantable material manufacturing process. Alternatively or further, there may be a sterilization process before and/or after assembly and sealing of the kit. In this technique, low moisture conditions are often useful. It has been observed that solid phase dispersed powders resist aggregate formation and cross-linking under gamma irradiation. This result is unexpected and surprising because gamma radiation sterilization is generally considered to damage protein or peptide biological agents. Without being bound by a particular theory of operation, it is believed that the small particle size and absence of moisture is detrimental to these unwanted reactions.
Further description of the invention
(1) A first embodiment of the present invention is directed to a method of making a medical material comprising forming an organogel around a powder of a water-soluble biologic, wherein the powder is dispersed in the organogel. (2) A second embodiment of the invention is directed to a method of making a medical material comprising forming a gel around a powder of a water-soluble biologic, wherein the powder is dispersed in the gel, wherein forming the gel comprises preparing a melt of one or more precursors and covalently crosslinking the precursors. (3) A third embodiment of the invention is directed to a method of making a medical material comprising forming an organogel around particles of a powder of a biologic agent, wherein the particles are dispersed in the organogel, and removing solvent from the organogel, thereby forming a xerogel, the method being performed in the absence of water. (4) A fourth embodiment of the invention is directed to a method of making a medical material comprising forming an organogel or gel from a melt, making a xerogel from the (organic) gel, and providing the xerogel as an assembly of particles, wherein the xerogel is a hydrogel upon exposure to an aqueous solution. (5) A fifth embodiment of the invention is directed to a pharmaceutically acceptable material as in any one of embodiments I-IV. (6) A sixth embodiment of the invention relates to a medical material comprising a pharmaceutically acceptable biodegradable xerogel comprising dispersed protein particles, the protein being a therapeutic agent and having a secondary and/or tertiary structure. In addition, the protein may be released from the particle in aqueous solution in a conformation that is substantially non-denatured. (7) A seventh embodiment of the invention is directed to a (pharmaceutically acceptable) biomaterial for the controlled release of a therapeutic water-soluble biologic, comprising a pharmaceutically acceptable xerogel comprising solid particles of the biologic dispersed therein, (optionally wherein the xerogel is free of hydrophobic material) and wherein the xerogel is a hydrogel when exposed to water. (8) An eighth embodiment is a method of making any of the materials of embodiments VI or VII.
The further implementation mode is as follows: (9) the method of any one of claims 1-8, wherein the (water-soluble) biological agent is a protein. (10) The method of any one of claims 1-9, wherein the protein has a molecular weight of at least about 10,000 daltons and the saccharide is associated with the protein. (11) The method of any one of claims 1-10, wherein the powder is used and is a first powder, wherein the method further comprises a second powder comprising a second water-soluble biologic, wherein the first powder and the second powder are dispersed throughout the organogel. (12) The method of any one of claims 1-11, wherein the powder is used and has an average particle size of about 1 μm to about 10 μm. (13) The method of any one of claims 1-12, wherein the organogel is formed in the absence of an aqueous solution. (14) As in any of claims 1-13, comprising removing solvent from the organogel to thereby form a xerogel, if desired. (15) As in any one of claims 1-14, comprising removing the solvent by a method selected from the group consisting of vacuum removal, lyophilization, and freezing followed by application of vacuum. (16) As in any one of claims 1-15, comprising the xerogel, wherein the xerogel is a hydrogel upon exposure to an aqueous solution. (17) As in any of claims 1-15, comprising the powder, wherein the (water-soluble) biological agent remains in the powder substantially in a solid phase when the hydrogel is formed and slowly dissolves over a period of time when the hydrogel is exposed to a physiological solution in the mammalian body. (18) The method of claim 17, wherein said dissolving during a period of time is in the range of about 1 week to about 52 weeks. (19) Such as in any of 1-18, wherein the biological agent in the gel is a protein having a secondary and/or tertiary structure, wherein the protein is released in a conformation that is substantially free of denaturation, as can be measured by, for example, enzyme-linked immunosorbent assay and isoelectric focusing. (20) Such as in any of claims 1-19, wherein the gel or organogel or xerogel comprises a covalently crosslinked hydrophilic polymer. (21) As in any of claims 1-20, wherein the gel organogel or xerogel organogel comprises a covalently crosslinked hydrophilic polymer selected from the group consisting of: polyethylene oxide, polyvinylpyrrolidone, hyaluronic acid, polyhydroxyethylmethacrylate, and block copolymers thereof. (22) The method of any one of claims 1 to 21, wherein, when the hydrogel is present, the hydrogel is biodegradable by spontaneous hydrolysis through a hydrolytically degradable linkage selected from the group consisting of an ester, a carbonate, an anhydride, and an orthocarbonate. (23) The method of any one of claims 1-22, wherein the organogel comprises block copolymers that form the organogel when present, and forms a hydrogel upon exposure to an aqueous solution after removal of the solvent to form a xerogel. (24) As in any of claims 1-23, wherein the organogel, when present, comprises a polymer wherein the organogel (and the hydrogel) comprises ionic crosslinks. (25) The method of any one of claims 1-24, wherein, when present, the organogel comprises a member selected from the group consisting of alginate, gellan, collagen, and polysaccharide. (25) As in any of claims 1-24, comprising forming a plurality of particles from: (a) said gel, (b) said organogel, (c) a xerogel made from said gel or said organogel, or (d) a hydrogel made from said gel or organogel. (26) The method of any one of claims 1-25, wherein the organogel is formed from a precursor in an organic solvent when the organogel is present, wherein the precursor chemically reacts to form covalent bonds to thereby form the organogel, wherein the organogel is covalently crosslinked. (27) The method of any one of claims 1-26, wherein the precursor is reacted by free radical polymerization to form the organogel. (28) The method of any one of claims 1-27, wherein the precursor is a first precursor comprising a first functional group and further comprising a second precursor comprising a second functional group, wherein the first functional group and the second functional group are reactive in the organic solvent to form a covalent bond. (29) The method of claim 28, wherein the first functional group and the second functional group are each selected from an electrophile and a nucleophile, and the reaction between the first functional group and the second functional group is an electrophilic-nucleophilic reaction that forms a covalent bond. (30) As in 28 or 29, wherein the electrophilic group comprises a succinimide, a succinimide ester, an n-hydroxysuccinimide, a maleimide, a succinate, a nitrophenylcarbonate, an aldehyde, a vinyl sulfone, an azide, a hydrazide, an isocyanate, a diisocyanate, a tosyl, a trifluoroethanesulfonyl (tresyl), or a carbonyldiimidazole. (31) Such as any one of 28-30, wherein the nucleophile group comprises a primary amine or a primary thiol. (32) The method of any one of claims 28-31, wherein the first precursor and the second precursor are water soluble. (33) As in any of 28-32, wherein at least one of the first precursor and the second precursor comprises a synthetic polymer. (34) The method of any of claims 28-33, wherein the first precursor comprises a polymer selected from the group consisting of: polyethylene glycol, polyacrylic acid, polyvinylpyrrolidone, and block copolymers thereof. (35) The method as in any one of claims 1-34, comprising preparing the organogel into a structure selected from the group consisting of: a rod, a sheet, a particle, a sphere, and an aggregate of at least one thereof. (36) As in any one of claims 1-35, comprising, or further comprising, a therapeutic agent, wherein the therapeutic agent comprises a fluoroquinolone, moxifloxacin, travoprost, dexamethasone, an antibiotic, or vestibiotoxin. (37) The method of claim 36, wherein said organogel further comprises a penetration enhancer. (38) The method of any one of claims 1-8, wherein the organogel is physically crosslinked by domain formation, the method further comprising forming the organogel from a precursor in an organic solvent, wherein the precursor is a block copolymer comprising a first block and a second block. (39) As in 38, comprising heating a mixture of the precursor and the organic solvent and allowing the solution to cool, thereby precipitating at least a first block of a copolymeric precursor, wherein the domains comprise at least the first block. (40) As in 38 or 39, comprising mixing precursors in a first organic solvent that dissolves a copolymeric precursor, wherein all blocks of said copolymeric precursor are soluble in said first organic solvent, and adding a second organic solvent that is miscible with said first organic solvent, wherein said first block of said copolymeric precursor is insoluble in said second organic solvent, wherein said second solvent is effective to form said domains, wherein said domains comprise said first block of said copolymer. (41) Such as in any of 38-40, wherein the copolymeric precursor comprises a block selected from the group consisting of polyethylene glycol. (42) The method of any one of claims 38-41, wherein the copolymeric precursor further comprises a second block selected from the group consisting of: polylactic acid, polyglycolic acid, polytrimethylene carbonate, polydioxanone, polyalkyl, polybutylene terephthalate, and polylysine. (43) The organogel of any of claims 1-37, wherein the organogel is free of hydrophobic materials; or no hydrophobic polymer, or no hydrophobic material except the solvent (which may be slightly hydrophobic). (44) As in any of claims 1-43, comprising preparing a powder of the biological agent according to a method that avoids denaturation of the biological agent, and preventing exposure of the powder to water once the powder has been prepared. (45) The method of any one of claims 1-44, wherein the biological agent is a therapeutic protein having a secondary and/or tertiary structure. (46) As in any of claims 1-45, comprising a xerogel, wherein the xerogel is a hydrogel upon exposure to water. (47) The method of any one of claims 1 to 46, wherein said hydrogel, or a hydrogel made from said gel/organogel/xerogel, is biodegradable. (48) The xerogel as in any one of claims 1-47, wherein the cumulative release of the agent reaches 90% weight/weight of the agent at a time of about 1 month to about 6 months after placing the hydrogel and particles in a saline solution. (49) The biomaterial as in any one of claims 1-48. (50) A biomaterial as in any one of claims 1-49 wherein the xerogel comprises a covalently crosslinked hydrophilic polymer. (51) The biomaterial of any one of claims 1-50, wherein the water-soluble biological agent is a protein having a secondary and/or tertiary structure. (52) The biomaterial of any one of claims 1-51, wherein the water-soluble biologic remains substantially in a solid phase when the hydrogel is formed and slowly dissolves over a period of time when the hydrogel is exposed to a physiological solution in the mammalian body. (53) The biomaterial of any one of claims 1-52, comprising the organogel, wherein the organogel comprises a covalently crosslinked hydrophilic polymer. (54)53, wherein the polymer comprises a member selected from the group consisting of: polyethylene oxide, polyvinylpyrrolidone, hyaluronic acid, polyhydroxyethylmethacrylate, and block copolymers thereof. (54) The method of any one of claims 1-53, wherein the material is a structure selected from the group consisting of: rods, sheets, particles, spheres, and aggregates thereof. (55)1-54, comprising said xerogel, or providing said xerogel as an assembly of particles, for example by a method selected from the group consisting of: (a) producing the organogel and breaking it to form particles for the aggregates, (b) producing the xerogel and breaking the xerogel to form particles for the aggregates, and (c) producing the organogel as a plurality of particles for the aggregates, the particles being depleted of organic solvent to produce the xerogel. (56) The method of 55, comprising producing a plurality of collections of particles, wherein the collections have different rates of degradation in vivo, and mixing the collections to produce a biomaterial having a desired degradation profile.
These embodiments 1-56 can be further prepared as a kit having a polymer, a biological agent or protein, and an applicator, wherein the kit is in a sterile container. These embodiments 1-56 may further be practiced by placing the material, or a material made by one of the methods, in contact with the tissue of a patient. Examples of tissues are the intraperitoneal space, muscles, dermis, epidermis, natural lumen or void, abdominal cavity, prostate, rectum, location between prostate and rectum, thorax, tissue between the irradiation target and healthy tissue, and vasculature.
The invention also relates to the following embodiments:
1. a method of making a medical material comprising forming an organogel around a powder of a water-soluble biologic, wherein the powder is dispersed in the organogel.
2. The method of clause 1, wherein the water-soluble biological agent is a protein having a molecular weight of at least about 10,000 daltons and a sugar is associated with the protein.
3. The method of item 1 or 2, wherein the powder is a first powder, wherein the method further comprises a second powder comprising a second water-soluble biologic, wherein the first powder and the second powder are dispersed throughout the organogel.
4. The process of any of items 1 to 3, wherein the organogel is formed in the absence of an aqueous solution.
5. The method of any of clauses 1-4, further comprising removing solvent from the organogel to thereby form a xerogel.
6. The method of item 5, wherein the solvent is removed by a method selected from the group consisting of: vacuum removal, lyophilization, and freezing followed by application of vacuum.
7. The method of item 5, wherein the xerogel is a hydrogel upon exposure to an aqueous solution.
8. The method of clause 7, wherein the water-soluble biologic remains in the powder substantially in a solid phase when the hydrogel is formed and slowly dissolves over a period of time when the hydrogel is exposed to a physiological solution in the mammalian body.
9. The method of item 7, wherein the biological agent is a protein having a secondary and/or tertiary structure, wherein the protein is released in a conformation that is substantially free of denaturation, which can be measured by, for example, enzyme-linked immunosorbent assay and isoelectric focusing.
10. The method of any of items 1-9, wherein the organogel comprises covalently crosslinked hydrophilic polymers.
11. The method of any of items 1-9, wherein the organogel comprises a block copolymer that forms the organogel and forms a hydrogel upon exposure to an aqueous solution after removal of the solvent to form a xerogel.
12. The method of any one of items 1 to 11, wherein the organogel is desolventized to produce a xerogel that forms a hydrogel upon exposure to an aqueous solution, wherein the hydrogel is biodegradable by spontaneous hydrolysis of a hydrolytically degradable linkage selected from the group consisting of: esters, carbonates, anhydrides and orthocarbonates.
13. The method of any of items 1-9 or 12, wherein the organogel comprises an ionically crosslinked polymer.
14. The method of item 1, wherein the organogel comprises a member selected from the group consisting of alginate, gellan, collagen, and polysaccharide.
15. The method of any of items 1-13, further comprising forming a plurality of particles from: the organogel, a xerogel made from the organogel, or a hydrogel made from the organogel.
16. The method of any of items 1-10 or 14-15, comprising forming the organogel from a precursor in an organic solvent, wherein the precursor chemically reacts to form a covalent bond to thereby form the organogel, wherein the organogel is covalently crosslinked.
17. The method of clause 16, wherein the precursor reacts by free radical polymerization to form the organogel.
18. The method of clause 16, wherein the precursor is a first precursor comprising a first functional group and, further comprising a second precursor comprising a second functional group, wherein the first functional group and the second functional group are reactive in the organic solvent to form a covalent bond.
19. The method of item 18, wherein the first functional group and the second functional group are each selected from an electrophile and a nucleophile, and the reaction between the first functional group and the second functional group is an electrophilic-nucleophilic reaction that forms a covalent bond.
20. The method of item 18 or 19, wherein the electrophilic group comprises a succinimide, a succinimide ester, an n-hydroxysuccinimide, a maleimide, a succinate, a nitrophenylcarbonate, an aldehyde, a vinyl sulfone, an azide, a hydrazide, an isocyanate, a diisocyanate, a tosyl, a trifluoroethanesulfonyl, or a carbonyldiimidazole.
21. The method of any one of items 18 to 20, wherein the nucleophile group comprises a primary amine or a primary thiol.
22. The method of any of items 18-21, wherein the first precursor and the second precursor are water soluble.
23. The method of any of items 18-22, wherein either or both of the first precursor and the second precursor comprise a synthetic polymer.
24. The method of any of items 18-23, wherein the first precursor comprises a polymer selected from the group consisting of: polyethylene glycol, polyacrylic acid, polyvinylpyrrolidone, and block copolymers thereof.
25. The method of any one of items 1-24, wherein the water-soluble biological agent comprises a therapeutic agent selected from the group consisting of: fluoroquinolone, moxifloxacin, travoprost, dexamethasone, antibiotics, or vestibiotoxin.
26. The method of any of clauses 1-9, 11-12, 14-15, or 24-25, wherein the organogel is physically crosslinked by formation of domains, the method further comprising forming the organogel from a precursor in an organic solvent, wherein the precursor is a block copolymer comprising a first block and a second block.
27. The method of item 26, comprising heating a mixture of the precursor and the organic solvent and allowing the solution to cool, thereby precipitating at least the first block of the copolymeric precursor, wherein the domains comprise at least the first block.
28. The method of item 26, comprising:
mixing the precursor in a first organic solvent that dissolves a copolymeric precursor, wherein all blocks of the copolymeric precursor are soluble in the first organic solvent, and
adding a second organic solvent that is miscible with the first organic solvent, wherein the first block of the copolymeric precursor is insoluble in the second organic solvent, wherein the second solvent is effective to form the domain, wherein the domain comprises the first block of the copolymer.
29. The method of item 26, wherein the copolymeric precursor comprises a block selected from the group consisting of polyethylene glycol.
30. The method of any of clauses 1-10 or 12-25, wherein the organogel is free of hydrophobic materials.
31. The method of any one of items 1-30, comprising preparing a powder of the biological agent according to a method that avoids denaturation of the biological agent, and once the powder is prepared, preventing exposure of the powder to water until the medical material is used on a patient.
32. A biomaterial made by the method of any one of claims 1-31.
33. A biomaterial for the controlled release of a therapeutic water-soluble biologic comprising a pharmaceutically acceptable xerogel comprising solid particles of the biologic dispersed therein, wherein the xerogel is a hydrogel when exposed to water.
34. The biomaterial of item 33 wherein the xerogel comprises a covalently crosslinked hydrophilic polymer and/or wherein the water-soluble biologic is a protein having a secondary and/or tertiary structure.
35. The biomaterial of item 33 or 34, wherein the water-soluble biologic remains substantially in a solid phase when the hydrogel is formed and slowly dissolves over a period of time when the hydrogel is exposed to a physiological solution in the mammalian body.
36. The biomaterial of any one of claims 33-35, wherein the organogel comprises covalently crosslinked hydrophilic polymers.
37. The biomaterial of any one of claims 33-36 wherein the xerogel is biodegradable after hydration in water by spontaneous hydrolysis of a hydrolytically degradable linkage selected from the group consisting of: esters, carbonates, anhydrides and orthocarbonates.
38. The biomaterial of any of claims 33-37 wherein the xerogel is a chemical reaction product of a first precursor comprising a first functional group and a second precursor comprising a second functional group, wherein the first functional group and the second functional group react to form a covalent bond.
39. A method of controlled delivery of a drug, comprising placing the biomaterial of any of items 33-38 in contact with a tissue of a patient.
40. The method of item 39, wherein the tissue comprises an intraperitoneally space, muscle, dermis, epidermis, natural lumen or void, abdominal cavity, prostate, rectum, a location between prostate and rectum, thorax, tissue between a radiation target and healthy tissue, and vasculature.
41. A kit comprising the biomaterial of any one of claims 1-40.
42. The kit of item 41, wherein the biomaterial is an assembly of xerogel particles that form hydrogel particles upon exposure to water.
43. A method of making a medical material comprising
The formation of an organogel is carried out,
a xerogel is made from the organogel,
and providing the xerogel as an assembly of particles, wherein the xerogel is a hydrogel upon exposure to an aqueous solution.
44. The method of item 43, comprising: producing a plurality of said assemblies of particles, wherein said assemblies have different in vivo degradation rates, and mixing the assemblies to produce a biomaterial having a desired degradation property.