# Patent

## Title
Threads of Hyaluronic acid and/or derivatives thereof, methods of making thereof and uses thereof

## Abstract

The present invention provides threads of hyaluronic acid, and/or derivatives thereof, methods of making thereof and uses thereof, for example, in aesthetic applications (e.g., dermal fillers), surgery (sutures), drug delivery, etc.

## Background
CROSS REFERENCE TO RELATED APPLICATIONS
This application is a National Phase application under 35 U.S.C. §371 of PCT Application PCT/US2009/055704, filed Sep. 2, 2009, which claims benefit under 35 U.S.C. §119(e) from U.S. Provisional Application Ser. No. 61/190,866, filed Sep. 2, 2008, which are hereby incorporated by reference in their entirety.


FIELD
The present invention relates generally to threads of hyaluronic acid, and/or derivatives thereof, methods of making thereof and uses thereof, for example, in aesthetic applications (e.g., dermal fillers), surgery (e.g., sutures), drug delivery, negative pressure wound therapy, moist wound dressing, etc.
BACKGROUND
Hyaluronic acid is a linear polysaccharide (i.e., non-sulfated glycosaminoglycan) consisting of a repeated disaccharide unit of alternately bonded β-D-N-acetylglucoamine and β-D-glucuronic acid (i.e., (-4GlcUAβ1-3GlcNAcβ1-)n) which is a chief component of the extracellular matrix and is found, for example, in connective, epithelial and neural tissue. Natural hylauronic acid is highly biocompatible because of its lack of species and organ specificity and thus is often used as a biomaterial in tissue engineering and as a common ingredient in various dermal fillers.
Various chemically modified forms of hyaluronic acid (e.g., cross linked forms, ionically modified forms, esterified forms, etc.) have been synthesized to address a significant problem associated with natural hyaluronic acid which has poor in vivo stability due to rapid enzymatic degradation and hydrolysis. Currently, hyaluronic acid or cross linked versions thereof are used in various gel forms, for example as dermal fillers, adhesion barriers, etc.
However, substantial issues exist with the use of gels of hyaluronic acid or cross linked versions thereof. First, the force required to dispense gels of hyaluronic acid or cross linked versions thereof is non-linear which causes the initial “glob” that many physicians report when injecting hyaluronic acid or cross linked versions thereof. Second, precisely dispensing hyaluronic gels to specific locations is very difficult because such gels have little mechanical strength. Further, the gel will occupy the space of least resistance which makes its use in many applications (e.g., treatment of fine wrinkles) problematic.
Accordingly, what is needed are new physical forms of hyaluronic acid or cross linked versions thereof which can be dispensed uniformly to specific locations regardless of tissue resistance. Such new forms may have particular uses, for example, in aesthetic and surgical applications, drug delivery, wound therapy and wound dressing.
SUMMARY
The present invention satisfies these and other needs by providing, in one aspect, a thread of hyaluronic acid or salts, hydrates or solvates thereof and, in a second aspect, a thread of cross linked hyaluronic acid or salts, hydrates or solvates thereof. In some embodiments, the thread is a combination of a thread of hyaluronic acid or salts, hydrates or solvates thereof and a thread of cross linked hyaluronic acid or salts, hydrates or solvates thereof.
In a third aspect, a method of making a thread of hyaluronic acid or salts, hydrates or solvates thereof is provided. Hyaluronic acid or salts, hydrates or solvates thereof are mixed with water or a buffer to form a gel. The gel is extruded to form a thread. The thread is then dried to provide a thread of hyaluronic acid.
In a fourth aspect, a method of making a thread of cross linked hyaluronic acid or salts, hydrates or solvates thereof is provided. Hyaluronic acid or salts, hydrates or solvates thereof are mixed with water or a buffer and a cross linking agent to form a gel. The gel is extruded to form a thread. The thread is then dried to provide a thread of cross linked hyaluronic acid.
In a fifth aspect a method of treating a wrinkle in a subject in need thereof is provided. A thread of hyaluronic acid or salts, hydrates or solvates thereof or a thread of cross linked hyaluronic acid or salts, hydrates or solvates thereof or a combination thereof is attached to the proximal aspect of a needle. The distal end of the needle is inserted through the skin surface of the subject into the dermis adjacent to or within the wrinkle. The dermis of the subject in the base of the wrinkle is traversed with the needle. The needle then exits the skin surface of the subject and is pulled distally until it is removed from the skin of the subject such that the thread is pulled into the location previously occupied by the needle. The excess thread is cut from the needle at the skin surface of the subject.
In still other aspects, methods of using threads of hyaluronic acid or salts, hydrates or solvates thereof or threads of cross linked hyaluronic acid or salts, hydrates or solvates thereof or combinations thereof, for example, as dermal fillers, adhesion barriers, wound dressings including negative pressure wound dressings, sutures, etc. is provided. Further provided are methods of using threads of hyaluronic acid or salts, hydrates or solvates thereof or threads of cross linked hyaluronic acid or salts, hydrates or solvates thereof or combinations thereof, for example, in surgery, opthamology, wound closure, drug delivery, etc.



BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1 illustrates a thread attached to the proximal end of a needle, in its entirety;
FIG. 2A illustrates a close-up view of a thread inserted into the inner-diameter of a needle;
FIG. 2B illustrates a close-up view of the proximal end of a solid needle with the thread overlapping the needle;
FIG. 3A illustrates a fine, facial wrinkle in the peri-orbital region of a human;
FIG. 3B illustrates a needle and thread being inserted into the dermis of the wrinkle at the medial margin;
FIG. 3C illustrates the needle being adjusted to traverse beneath the wrinkle;
FIG. 3D illustrates the needle exiting at the lateral margin of the wrinkle;
FIG. 3E illustrates the needle having pulled the thread into the location it previously occupied beneath the wrinkle;
FIG. 3F illustrates the thread implanted beneath the wrinkle, with excess thread having been cut off;
FIG. 4A illustrates a top-down view of a male with typical male-pattern baldness;
FIG. 4B illustrates where hair re-growth is desired, taking hair-lines into consideration;
FIG. 4C illustrates a curved needle with attached thread being inserted into one imaginary line where hair re-growth is desired;
FIG. 4D illustrates the needle traversing the imaginary line, and exiting the skin;
FIG. 4E illustrates the needle pulled through distally, pulling along the thread into the desired location;
FIG. 4F illustrates scissors being used to cut excess thread;
FIG. 5A illustrates a cross-sectional view of a fold or a wrinkle;
FIG. 5B illustrates a thread implanted beneath a wrinkle that is not yet hydrated;
FIG. 5C illustrates a thread implanted beneath a wrinkle that is fully hydrated and has flattened the surface appearance of the wrinkle;
FIG. 6A illustrates a human pancreas with a tumor;
FIG. 6B illustrates a curved needle with a thread attached thereto;
FIG. 6C illustrates a curved needle traversing the tumor within the pancreas;
FIG. 6D illustrates the end-result of repeated implantations of thread;
FIG. 7A illustrates multiple layers of concentric coils of thread, shaped to represent a human nipple;
FIG. 7B illustrates the implant of FIG. 7A in cross-section;
FIG. 7C illustrates how an implant of coiled thread would be used for nipple reconstruction; and
FIG. 8 illustrates how a needle and thread could be used to place a thread in a specific, linear location to promote nerve or vessel regrowth in a specific line.



DETAILED DESCRIPTION
Definitions
“Buffer” includes, but is not limited to, 2-amino-2-methyl-1,3-propanediol, 2-amino-2-methyl-1-propanol, L-(+)-tartaric acid, D-(−)-tartaric acid, ACES, ADA, acetic acid, ammonium acetate, ammonium bicarbonate, ammonium citrate, ammonium formate, ammonium oxalate, ammonium phosphate, ammonium sodium phosphate, ammonium sulfate, ammonium tartrate, BES, BICINE, BIS-TRIS, bicarbonate, boric acid, CAPS, CHES, calcium acetate, calcium carbonate, calcium citrate, citrate, citric acid, diethanolamine, EPP, ethylenediaminetetraacetic acid disodium salt, formic acid solution, Gly-Gly-Gly, Gly-Gly, glycine, HEPES, imidazole, lithium acetate, lithium citrate, MES, MOPS, magnesium acetate, magnesium citrate, magnesium formate, magnesium phosphate, oxalic acid, PIPES, phosphate buffered saline, phosphate buffered saline, piperazine potassium D-tartrate, potassium acetate, potassium bicarbonate, potassium carbonate, potassium chloride, potassium citrate, potassium formate, potassium oxalate, potassium phosphate, potassium phthalate, potassium sodium tartrate, potassium tetraborate, potassium tetraoxalate dehydrate, propionic acid solution, STE buffer solution, sodium 5,5-diethylbarbiturate, sodium acetate, sodium bicarbonate, sodium bitartrate monohydrate, sodium carbonate, sodium citrate, sodium formate, sodium oxalate, sodium phosphate, sodium pyrophosphate, sodium tartrate, sodium tetraborate, TAPS, TES, TNT, TRIS-glycine, TRIS-acetate, TRIS buffered saline, TRIS-HCl, TRIS phosphate-EDTA, tricine, triethanolamine, triethylamine, triethylammonium acetate, triethylammonium phosphate, trimethylammonium acetate, trimethylammonium phosphate, Trizma® acetate, Trizma® base, Trizma® carbonate, Trizma® hydrochloride or Trizma® maleate.
“Salt” refers to a salt of hyaluronic acid, which possesses the desired activity of the parent compound. Such salts include, but are not limited to: (1) acid addition salts, formed with inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, phosphoric acid and the like; or formed with organic acids such as acetic acid, propionic acid, hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid, lactic acid, malonic acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid, citric acid, benzoic acid, 3-(4-hydroxybenzoyl) benzoic acid, cinnamic acid, mandelic acid, methanesulfonic acid, ethanesulfonic acid, 1,2-ethane-disulfonic acid, 2-hydroxyethanesulfonic acid, benzenesulfonic acid, 4-chlorobenzenesulfonic acid, 2-naphthalenesulfonic acid, 4-toluenesulfonic acid, camphorsulfonic acid, 4-methylbicyclo[2.2.2]-oct-2-ene-1-carboxylic acid, glucoheptonic acid, 3-phenylpropionic acid, trimethylacetic acid, t-butylacetic acid, lauryl sulfuric acid, gluconic acid, glutamic acid, hydroxynaphthoic acid, salicylic acid, stearic acid, muconic acid and the like; or (2) salts formed when an acidic proton present in the parent compound is replaced by an ammonium ion, a metal ion, e.g., an alkali metal ion (e.g., sodium or potassium), an alkaline earth ion (e.g., calcium or magnesium), or an aluminum ion; or coordinates with an organic base such as ethanolamine, diethanolamine, triethanolamine, N-methylglucamine, morpholine, piperidine, dimethylamine, diethylamine and the like. Also included are salts of amino acids such as arginates and the like, and salts of organic acids like glucurmic or galactunoric acids and the like.
Threads of Hyaluronic Acid and Derivatives Thereof
The present invention generally provides threads of hyaluronic acid or salts, hydrates or solvates thereof, threads of cross linked hyaluronic acid or salts, hydrates or solvates thereof and combinations thereof. In some embodiments, the hyaluronic acid is isolated from an animal source. In other embodiments, the hyaluronic acid is isolated from bacterial fermentation
In some embodiments, the lifetime of the threads of hyaluronic acid or salts, hydrates or solvates thereof, in vivo is between about 1 minute and about 1 month. In other embodiments, the lifetime of the thread of hyaluronic acid or salts, hydrates or solvates thereof, in vivo is between about 10 minutes and about 1 week. In still other embodiments, the lifetime of the thread of hyaluronic acid or salts, hydrates or solvates thereof, in vivo is between about 1 hour and about 3 days.
In some embodiments, the lifetime of the thread of cross linked hyaluronic acid or salts, hydrates or solvates thereof, in vivo is between about 1 week and about 24 months. In other embodiments, the lifetime of the thread of cross linked hyaluronic acid or salts, hydrates or solvates thereof, in vivo is between about 1 month and about 12 months. In still other embodiments, the lifetime of the thread of hyaluronic acid or salts, hydrates or solvates thereof, in vivo is between about 3 months and about 9 months.
In some embodiments, hyaluronic acid or salts, hydrates or solvates thereof have been cross linked with butanediol diglycidyl ether (BDDE), divinyl sulfone (DVS) or 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC). Those of skill in the art will appreciate that many other cross linking agents may be used to crosslink hyaluronic acid or salts, hydrates or solvates thereof. Accordingly, the above list of cross linking agents is illustrative rather than comprehensive.
In some of the above embodiments, the degree of cross linking between hyaluronic acid or salts, hydrates or solvates thereof and the cross linking agent is between about 0.01% and about 20%. In other of the above embodiments, the degree of cross linking between hyaluronic acid or salts, hydrates or solvates thereof and the cross linking agent is between about 0.1% and about 10%. In still other of the above embodiments, the degree of cross linking between hyaluronic acid or salts, hydrates or solvates thereof and the cross linking agent is between about 1% and about 8%.
In some of the above embodiments, the thread includes one or more therapeutic or diagnostic agents. In other of the above embodiments, the diagnostic agent is soluble TB (tuberculosis) protein. In still other of the above embodiments, the therapeutic agent is an anesthetic, including but not limited to, lidocaine, xylocaine, novocaine, benzocaine, prilocaine, ripivacaine, propofol or combinations thereof. In still other of the above embodiments, the therapeutic agent is epinephrine, adrenaline, ephedrine, aminophylline, theophylline or combinations thereof. In still other of the above embodiments, the therapeutic agent is botulism toxin. In still other of the above embodiments, the therapeutic agent is laminin-511. In still other of the above embodiments, the therapeutic agent is glucosamine, which can be used, for example, in the treatment of regenerative joint disease. In still other of the above embodiments, the therapeutic agent is an antioxidant, including but not limited to, vitamin E or all-trans retinoic acid such as retinol. In still other of the above embodiments, the therapeutic agent includes stem cells. In still other of the above embodiments, the therapeutic agent is insulin, a growth factor such as, for example, NGF (nerve growth factor), BDNF (brain-derived neurotrophic factor), PDGF (platelet-derived growth factor) or Purmorphamine Deferoxamine NGF (nerve growth factor), dexamethasone, ascorbic acid, 5-azacytidine, 4,6-disubstituted pyrrolopyrimidine, cardiogenols, cDNA, DNA, RNAi, BMP-4 (bone morphogenetic protein-4), BMP-2 (bone morphogenetic protein-2), an antibiotic agent such as, for example, β lactams, quinolones including fluoroquinolones, aminoglycosides or macrolides, an anti-fibrotic agent, including but not limited to, hepatocyte growth factor or Pirfenidone, an anti-scarring agent, such as, for example, anti-TGF-b2 monoclonal antibody (rhAnti-TGF-b2 mAb), a peptide such as, for example, GHK copper binding peptide, a tissue regeneration agent, a steroid, fibronectin, a cytokine, an analgesic such as, for example, Tapentadol HCl, opiates, (e.g., morphine, codone, oxycodone, etc.) an antiseptic, alpha- beta or gamma-interferon, EPO, glucagons, calcitonin, heparin, interleukin-1, interleukin-2, filgrastim, a protein, HGH, luteinizing hormone, atrial natriuretic factor, Factor VIII, Factor IX, or a follicle-stimulating hormone. In still other of the above embodiments, the thread contains a combination of more than one therapeutic agent or diagnostic agent. In some of these embodiments, different threads comprise different therapeutic agents or diagnostic agents.
In some of the above embodiments, the thread has an ultimate tensile strength of between about 0 kpsi and about 250 kpsi. In other of the above embodiments, the thread has an ultimate tensile strength of between about 1 kpsi and about 125 kpsi. In still other of the above embodiments, the thread has an ultimate tensile strength of between about 5 kpsi and about 100 kpsi.
In some of the above embodiments, the thread has an axial tensile strength of between about 0.01 lbs and about 10 lbs. In other of the above embodiments, the thread has an axial tensile strength of between about 0.1 lbs and about 5 lbs. In still other of the above embodiments, the thread has an axial tensile strength of between about 0.5 lbs and about 2 lbs.
In some of the above embodiments, the thread has a cross-section area of between about 1*106 in2 and about 1,000*106 in2. In other of the above embodiments, the thread has a cross-section area of between about 10*106 in2 and about 500*106 in2. In still other of the above embodiments, the thread has a cross-section area of between about 50*106 in2 and about 250*106 in2.
In some of the above embodiments, the thread has a diameter of between about 0.0001 in and about 0.100 in. In other of the above embodiments, the thread has a diameter of between about 0.001 in and about 0.020 in. In still other of the above embodiments, the thread has a diameter of between about 0.003 and about 0.010 in.
In some of the above embodiments, the thread has an elasticity of between about 1% and 200%. In other of the above embodiments, the thread has an elasticity of between about 5% and about 100%. In still other of the above embodiments, the thread has an elasticity of between about 10% and 50%. Herein, elasticity is the % elongation of the thread while retaining ability to return to the initial length of the thread.
In some of the above embodiments, the thread has a molecular weight of between about 0.1 MD and about 8 MD (MD is a million Daltons). In other of the above embodiments, the thread has a molecular weight of between about 0.5 MD to about 4 MD. In still other of the above embodiments, the thread has a molecular weight of between about 1 MD to about 2 MD.
In some of the above embodiments, the thread has a persistent chain length of between about 10 nm and about 250 nm. In other of the above embodiments, the thread has a persistent chain length of between about 10 nm and about 125 nm. In still other of the above embodiments, the thread has a persistent chain length of between about 10 nm and about 75 nm.
In some of the above embodiments, the cross-sectional area of the thread when fully hydrated swells to between about 0% to about 10,000%. In other of the above embodiments, the cross-sectional area of the thread when fully hydrated swells to between about 0% to about 2,500%. In still other of the above embodiments, the cross-sectional area of the thread when fully hydrated swells to between about 0% to about 900%.
In some of the above embodiments, the thread elongates when fully hydrated to between about 0% to about 1,000%. In other of the above embodiments, the thread elongates when fully hydrated to between about 0% to about 100%. In still other of the above embodiments, the thread elongates when fully hydrated to between about 0% to about 30%.
In some of the above embodiments, the thread is fully hydrated after submersion in an aqueous environment in between about 1 second and about 24 hours. In other of the above embodiments, the thread is fully hydrated after submersion in an aqueous environment in between about 1 second and about 1 hour. In still other of the above embodiments, the thread is fully hydrated after submersion in an aqueous environment in between about 1 second to about 5 minutes.
In some embodiments, the thread is cross linked and has an ultimate tensile strength of between about 50 kpsi and about 75 kpsi, a diameter of between 0.005 in and about 0.015 in, the thickness or diameter of the thread when fully hydrated swells between about 50% to about 100% and the lifetime of the thread in vivo is about 6 months.
In some embodiments, braids may be formed from the threads described above. In other embodiments, cords may be formed from the threads described above. In still other embodiments, a woven mesh may be formed from the threads described above. In still other embodiments, a woven mesh may be formed from the braids or cords described above.
In some embodiments, a three-dimensional structure may be constructed by weaving or wrapping or coiling or layering the threads described above. In other embodiments, a three-dimensional structure may be constructed by weaving or wrapping or coiling or layering the braids described above. In still other embodiments, a three-dimensional structure may be constructed by weaving or wrapping or coiling or layering the cords described above. In still other embodiments, a three-dimensional structure may be constructed by weaving or wrapping or coiling or layering the meshes described above.
In some embodiments, a three-dimensional, cylindrical implant is made of any of the threads is provided. An exemplary use for such an implant is for nipple reconstruction. In some embodiments, the threads used to make the cylindrical implant are cross linked and include chondrocyte adhesion compounds. In other embodiments, the cylindrical shape is provided by multiple, concentric coils of threads.
Threads of hyaluronic acid and/or derivatives thereof may contain one or more chiral centers and therefore, may exist as stereoisomers, such as enantiomers or diastereomers. In general, all stereoisomers (i.e., all possible enantiomers and stereoisomers of the illustrated compounds including the stereoisomerically pure form (e.g., enantiomerically pure or diastereomerically pure) and enantiomeric and stereoisomeric mixtures are within the scope of the present invention.
Threads of hyaluronic acid and/or derivatives thereof may exist in several tautomeric forms and mixtures thereof all of which are within the scope of the present invention. Threads of hyaluronic acid and/or derivatives thereof may exist in unsolvated forms as well as solvated forms, including hydrated forms. In general, hydrated and solvated forms are within the scope of the present invention. Accordingly, all physical forms of threads of hyaluronic acid and/or derivatives thereof are equivalent for the uses contemplated by the present invention and are intended to be within the scope of the present invention.
Methods of Making Threads of Hyaluronic Acid and Derivatives Thereof
The present invention also provides methods for making threads of hyaluronic acid and derivatives thereof as described above. In some embodiments, a method of making threads of hyaluronic acid or salts, hydrates or solvates thereof, is provided Hyaluronic acid or salts, hydrates or solvates thereof are mixed with water or a buffer to form a gel. The gel is then extruded to form a thread of gel. The gel can be extruded, for example, by placing the gel in a syringe with a nozzle, pressurizing the syringe, and linearly translating the syringe as gel is extruded from the nozzle. Nozzle characteristics such as taper, length and diameter, the syringe pressure, and the speed of linear translation may be adjusted to make threads of different sizes and mechanical characteristics. Another method of making a thread of gel is by rolling the gel, i.e., like dough, or by placing it into a mold. Still another method of making a thread of gel is to allow the gel to stretch into a thread under the influence of gravity or using centrifugal force. Still another method of making a thread of gel is by shearing the gel in between charged parallel glass plates. Yet another method of making a thread of gel is by confining the gel into a groove patterned on an elastomer and then stretching the elastomer. Yet another method of making a thread of gel is by confining the gel into a permeable tubular structure that allows dehydration of the thread, and if necessary controlling the nature of the dehydration by adjusting environmental parameters such as temperature, pressure and gaseous composition. The thread of hyaluronic acid or salts, hydrates or solvates thereof is then dried after preparation.
In other embodiments, a method of making threads of cross linked hyaluronic acid or salts, hydrates or solvates thereof, is provided. Hyaluronic acid or salts, hydrates or solvates thereof are mixed with water or a buffer and a cross linking agent to form a gel. The gel is then extruded to form a thread as described above or the thread can be made by any of the methods described above. Generally, the gel should be extruded or otherwise manipulated soon after addition of the cross linking agent so that cross linking occurs as the thread dries. The thread of cross linked hyaluronic acid or salts, hydrates or solvates thereof is then dried after preparation.
In some embodiments, the ratio of cross linking agent to hyaluronic acid is between about 0.01% and about 10%. In other embodiments, the ratio of cross linking agent to hyaluronic acid is between about 0.02% and about 5%. In still other embodiments, the ratio of cross linking agent to hyaluronic acid is between about 0.1% and about 3%.
In some of the above embodiments, one or more therapeutic or diagnostic agents are included in the gel forming step.
In some of the above embodiments, the gel has a concentration by weight of hyaluronic acid of between about 0.1% and about 10%. In other of the above embodiments, the gel has a concentration by weight of hyaluronic acid of between about 1% and about 7%. In still other of the above embodiments, the gel has a concentration by weight of hyaluronic acid of between about 4% and about 6%.
In some of the above embodiments, the polymer chains are further oriented along the axis of the thread by being stretched axially. In other of the above embodiments, the polymer chains are oriented along the axis of the thread by gravimetric force or centrifugal force. In still other of the above embodiments, gravimetric force is applied by hanging the thread vertically. In still other of the above embodiments, the polymer chains are oriented along the axis of the thread by application of an electric potential along the length of the thread. In still other of the above embodiments, the polymer chains are oriented along the axis of the thread by a combination of the above methods.
In some of the above embodiments, the threads are hydrated with water and then dried again. In other of the above embodiments, the hydration and drying steps are repeated multiple times. In still other of the above embodiments, the polymer chains are oriented along the axis of the thread by being stretched axially, by application of gravimetric force or centrifugal force, by application of an electric potential along the length of the thread or by combinations thereof. In still other of the above embodiments, a therapeutic agent or a diagnostic agent or a cross linking agent is applied to the thread during the hydration step.
In some of the above embodiments, the gel is extruded over a previously made thread to provide a layered thread.
In another of the above embodiments, after the drying step, the thread is submerged or rinsed with an agent. In some of the above embodiments, the agent is a cross linking agent, therapeutic or diagnostic agent.
In another of the above embodiments, while the thread is hydrated, for example after a rinsing step, the thread is submerged or rinsed with an agent. In some of the above embodiments, the agent is a cross linking agent, therapeutic or diagnostic agent.
In still other of the above embodiments, the thread is frozen and then thawed. In still other of the above embodiments, the thread is frozen and then thawed at least more than once.
In still other of the above embodiments, a dried thread is irradiated to promote cross linking. In some of the above embodiments, a hydrated thread is irradiated to promote cross linking.
In still other of the above embodiments, a dried or hydrated thread is coated to alter the properties of the thread, with a bioabsorbable biopolymer, such as for example, collagen, PEG or PLGA. Alternatively, woven constructs, whether single layer or 3D, can be coated in their entirety to create weaves or meshes with altered physical properties from that of a free-woven mesh.
Methods of Using Threads of Hyaluronic Acid and Derivatives Thereof
The threads, braids, cords, woven meshes or three-dimensional structures described herein can be used, for example, to fill aneurysms, occlude blood flow to tumors, (i.e., tumor occlusion), in eye-lid surgery, in penile augmentation (e.g., for enlargement or for sensitivity reduction, i.e., pre-mature ejaculation treatment), inter-nasal (blood-brain barrier) delivery devices for diagnostic and/or therapeutic agents, corneal implants for drug delivery, nose augmentation or reconstruction, lip augmentation or reconstruction, facial augmentation or reconstruction, ear lobe augmentation or reconstruction, spinal implants (e.g., to support a bulging disc), root canal filler (medicated with therapeutic agent), glottal insufficiency, laser photo-refractive therapy (e.g., hyaluronic acid thread/weave used as a cushion), scaffolding for organ regrowth, spinal cord treatment (BDNF and NGF), in Parkinson's disease (stereotactic delivery), precise delivery of therapeutic or diagnostic molecules, in pulp implantation, replacement pulp root canal treatment, shaped root canal system, negative pressure wound therapy, adhesion barriers and wound dressings.
In some embodiments, the threads, braids, cords, woven meshes or three-dimensional structures described herein are used as dermal fillers in various aesthetic applications. In other embodiments, the threads, braids, cords, woven meshes or three-dimensional structures described herein are used as sutures in various surgical applications. In still other embodiments, the threads, braids, cords, woven meshes or three-dimensional structures described herein are used in ophthalmologic surgery, drug delivery and intra-articular injection.
In some embodiments, the threads, braids, cords, woven meshes or three-dimensional structures described herein are used in wound dressings including negative pressure wound dressings.
In some embodiments, wound dressing remains in contact with the wound for at least 72 hours. In other embodiments, the negative pressure wound dressing remains in contact with the wound for at least 1 week. In still other embodiments, the wound dressing remains in contact with the wound for at least 2 weeks. In still other embodiments, the wound dressing remains in contact with the wound for at least 3 weeks. In still other embodiments, the wound dressing remains in contact with the wound for at least 4 weeks. In the above embodiments, it should be understood that granulation tissue is not retaining the threads, braids, cords, woven meshes or three-dimensional structures described herein as these components are fully absorbable. In some of these embodiments, the wound dressing is between about 1 cm and about 5 cm thick. Accordingly, in some of these embodiments, wound bed closure may be achieved without changing the dressing.
In some embodiments, the woven meshes described herein are used in wound dressings including negative pressure wound dressings. In other embodiments, the dressing include between 2 and about 10 layers of woven meshes.
In still other embodiments, the woven meshes comprise identical threads. In still other embodiments, the woven meshes comprise different threads.
In some embodiments, the woven meshes are between about 1 mm and about 2 mm thick when dry. In other embodiments, the woven meshes are between about 2 mm and about 4 mm thick when dry.
In some embodiments, the pore size of the woven mesh is between about 1 mm and about 10 mm in width. In other embodiments, the pore size of the woven mesh is between about 0.3 mm and about 0.6 mm in width. In still other embodiments, the pores of the woven mesh are aligned. In still other embodiments, the pores of the woven mesh are staggered. In still other embodiments, the woven meshes are collimated to create pores of desired size.
In some embodiments, the woven mesh is mechanically stable at a vacuum up to about 75 mm Hg. In other embodiments, the woven mesh is mechanically stable at a vacuum up to about 150 mm Hg.
In some embodiments, the woven mesh includes collagen. In other embodiments, the dressing is attached to a polyurethane foam. In still other embodiments, the polyurethane foam is open celled. In still other embodiments, the dressing is attached to a thin film. In still other embodiments, the thin film is silicone or polyurethane. In still other embodiments, the dressing is attached to the thin film with a water soluble adhesive.
In some embodiments, the thread used in the dressing includes a therapeutic agent or a diagnostic agent.
In some embodiments, a negative pressure wound dressing (Johnson et al., U.S. Pat. No. 7,070,584, Kemp et al., U.S. Pat. No. 5,256,418, Chatelier et al., U.S. Pat. No. 5,449,383, Bennet et al., U.S. Pat. No. 5,578,662, Yasukawa et al., U.S. Pat. Nos. 5,629,186 and 5,780,281 and Ser. No. 08/951,832) is provided for use in vacuum induced healing of wounds, particularly open surface wounds (Zamierski U.S. Pat. Nos. 4,969,880, 5,100,396, 5,261,893, 5,527,293 and 6,071,267 and Argenta et al., U.S. Pat. Nos. 5,636,643 and 5,645,081). The dressing includes a pad which conforms to the wound location, an air-tight seal which is removably adhered to the pad, a negative pressure source in fluid communication with the pad and the threads, braids, cords, woven meshes or three-dimensional structures described herein attached to the wound contacting surface of the pad. The pad, seal and vacuum source are implemented as described in the prior art.
In other embodiments, the threads, braids, cords, woven meshes or three-dimensional structures described herein are mechanically stable at a vacuum up to about 75 mm Hg. In still other embodiments, the threads, braids, cords, woven meshes or three-dimensional structures described herein are mechanically stable at a vacuum up to about 150 mm Hg. In still other embodiments, the dressing includes at least one layer of woven mesh. In still other embodiments, the dressing include between 2 and about 10 layers of woven mesh. In still other embodiments, the pad is a foam. In still other embodiments, the pad is an open cell polyurethane foam.
In some embodiments a tube connects the pad to the negative pressure source. In still other embodiments, a removable canister is inserted between the pad and the negative pressure source and is in fluid communication with both the pad and the negative pressure source.
In some embodiments, the threads, braids, cords, woven meshes or three-dimensional structures described herein are not hydrated. Accordingly, in these embodiments, the dressing could absorb wound exudates when placed in contact with the wound. In other embodiments, the threads, braids, cords, woven meshes or three-dimensional structures described herein are hydrated. Accordingly, in these embodiments, the dressing could keep the wound moist when placed in contact with the wound.
In some embodiments, an input port attached to a fluid is connected with the pad. Accordingly, in these embodiments, fluid could be dispensed in the wound. In some embodiments, the fluid is saline. In other embodiments, the fluid contains diagnostic or therapeutic agents.
In some embodiments, the threads, braids, cords, woven meshes or three-dimensional structures described herein are used as adhesion barriers. In some embodiments, the woven meshes described herein are used in adhesion barriers.
In some embodiments, a method of treating a wrinkle in a subject is provided. For example, the wrinkle may be in the peri-orbital region as illustrated in FIG. 3A. The thread may be attached to a needle as illustrated, for example, in FIGS. 1, 2A and 2B. The distal end of the needle may be inserted through the skin surface of the subject into the dermis adjacent to or within the wrinkle as illustrated, for example, in FIG. 3B. In some embodiments, the thread is inserted into the subcutaneous space instead of the dermis. The needle then may traverse the dermis of the subject beneath the wrinkle as illustrated, for example, in FIG. 3C. The needle then may exit the skin of the subject at the opposite margin of the wrinkle, as illustrated, for example, in FIG. 3D. The needle may then be pulled distally until it is removed from the subject such that the thread is pulled into the location previously occupied by the needle beneath the wrinkle, as illustrated, for example, in FIG. 3E. Finally, excess thread is cut from the needle at the skin surface of the subject which leaves the thread implanted as illustrated, for example, in FIG. 3F.
While not wishing to be bound by theory, the method above may successfully treat wrinkles as shown in FIGS. 5A, 5B and 5C. A typical wrinkle is illustrated in FIG. 5A. FIG. 5B illustrates a thread implanted beneath a wrinkle that is not yet hydrated. As the thread implanted beneath the wrinkle becomes fully hydrated the surface appearance of the wrinkle is concurrently flattened as illustrated in FIG. 5C.
In some embodiments, the above method may be used to rejuvenate the skin of a subject in need thereof. In many of these embodiments, the thread includes substantial amounts of non-cross linked hyaluronic acid. In some of these embodiments, the thread includes antioxidants, vitamin E or retinol or combinations thereof.
In some embodiments, a method of treating hair loss in a subject is provided. A subject such as, for example, a male with typical male-pattern baldness is illustrated in FIG. 4A and the area where hair growth (with imaginary hairlines) is desired is shown in FIG. 4B. The thread may be attached to a needle as illustrated, for example, in FIGS. 1, 2A, 2B and 4C. The distal end of the needle may be inserted into one of the hair lines as illustrated, for example, in FIG. 4C. The needle then may traverse the area beneath the hairline of the subject and then may exit the skin of the subject as illustrated, for example, in FIG. 4D. The needle may then be pulled distally until it is removed from the subject such that the thread is pulled into the location previously occupied by the needle as illustrated, for example, in FIG. 4E. Finally, excess thread is cut from the needle at the skin surface of the subject which leaves the thread implanted as illustrated, for example, in FIG. 4D.
In some embodiments, a method for treating tumors in a subject in need thereof is provided. The thread may be attached to a needle as illustrated, for example, in FIGS. 1, 2A and 2B. The distal end of the needle may be inserted into the tumor of the subject. The needle then may traverse the tumor and then may exit the tumor. The needle may then be pulled distally until it is removed from the tumor of the subject such that the thread is pulled into the location previously occupied by the needle. Finally, excess thread is cut from the needle which leaves the thread implanted in the tumor of the subject. In some of the above embodiments, the thread includes an anti-cancer agent. In some embodiments, the thread is cross linked and includes Bcl-2 inhibitors.
In an exemplary embodiment, methods of the current invention may be used to treat pancreatic tumors. FIG. 6A illustrates a human pancreas with a tumor while FIG. 6B illustrates a needle with a thread attached thereto. The pancreas may be accessed by surgery or minimally invasively methods such as by laparoscopy. The distal end of the needle may be inserted into the pancreatic tumor. The needle then may traverse the pancreatic tumor as illustrated in FIG. 6C and then may exit the tumor. The needle may then be pulled distally until it is removed from the pancreatic tumor such that the thread is pulled into the location previously occupied by the needle. Finally, excess thread is cut from the needle which leaves the thread implanted in the pancreatic tumor. The process may be repeated any number of times to provide, as illustrated in FIG. 6D, a pancreatic tumor which has been implanted with a number of threads. In some embodiments, the thread includes an anti-cancer agent.
In some embodiments, a method for treating a varicose vein in subject in need thereof is provided. The thread may be attached to a needle as illustrated, for example, in FIGS. 1, 2A and 2B. The distal end of the needle may be inserted into the varicose vein of the subject. The needle then may traverse the varicose vein and then may exit the vein. The needle may then be pulled distally until it is removed from the varicose vein of the subject such that the thread is pulled into the location previously occupied by the needle. Finally, excess thread is cut from the needle which leaves the thread implanted in the varicose vein of the subject. In some embodiments, the needle is a flexible. In other embodiments, the thread coils when hydrated, more readily occluding the vessel.
In some embodiments, a method for nipple reconstruction is provided where a three-dimensional, cylindrical implant comprised of cross linked threads is implanted underneath the skin. The implant may include therapeutic agents, for example chrondrocyte adhesion compounds. FIG. 7A illustrates an implant of multiple layers of concentric coils of threads shaped to represent a nipple while FIG. 7B shows a cross-section of the implant of FIG. 7A. FIG. 7C illustrates how the implant of FIG. 7A could be used for nipple reconstruction.
In some embodiments, methods for nerve or vessel regrowth are provided. As illustrated in FIG. 8, a needle can be used to place a thread in a specific line which could promote nerve or vessel regeneration.
EXAMPLES
The present invention is further defined by reference to the following examples. It will be apparent to those skilled in the art that many modifications, both to materials and methods, may be practiced without departing from the scope of the current invention.
Example 1
Synthesis of a Cross Linked Thread
A cross linked thread of a diameter between 0.004 in and 0.006 in was made by forming a gel with a concentration of 5% hyaluronic acid and 0.4% BDDE, by weight with the remainder comprised of water. A tapered tip nozzle with an inner diameter of 0.02 in, a syringe pressure of 20 psi and a linear translation speed commensurate with the speed of gel ejection from the syringe was used to extrude the gel into a thread form. However, numerous combinations of extrusion parameters that can make a thread of the desired thickness exist. The thread was dried and then rinsed with water which hydrated the thread, which was then stretched during drying. Over the course of multiple rinsing and drying cycles the overall length of the thread was increased by between about 25% and about 100%. The thread made as described above will fail at a tensile force of about between about 0.25 kg and about 1.50 kg and will swell in diameter by about 25% and about 100% when hydrated. It may persist as a thread in vivo between 1 and 9 months.
Example 2
Treatment of Wrinkles of a Cadaver with Hyaluronic Acid Threads
Hypodermic needles (22 to 25 Ga) were affixed with single or double strands of hyaluronic acid threads, ranging from thicknesses of 0.004 in to 0.008 in. Both non-crosslinked threads and threads crosslinked using BDDE were used. The needles were able to traverse wrinkles in a cadaveric head of a 50 y/o woman such as the naso-labial fold, peri-orals, peri-orbitals, frontalis (forehead), and glabellar. The needle was able to pull the thread through the skin such that the thread was located where the needle was previously inserted.
Example 3
Placement of Hyaluronic Acid Threads in Dogs
Acute and chronic canine studies were performed. Hypodermic needles (22 to 25 Ga) were affixed with single or double strands of hyaluronic acid threads, ranging from thicknesses of 0.004 in to 0.008 in. Both non-crosslinked threads and threads cross linked using BDDE were used. In all cases, the needle was able to pull the attached thread or threads into the dermis. Within minutes most threads produced a visible impact on the skin surface of the animals in the form of a linear bump.
Example 4
Comparison of Tensile Strength of Different Hyaluronic Acid Threads
The tensile strength of an autocrosslinked thread of hyaluronic acid was compared to a thread cross linked using the method of Example 1. A thread of non-crosslinked hyaluronic acid was repeatedly frozen and thawed, replicating a method of autocrosslinking hyaluronic acid (Ref. U.S. Pat. No. 6,387,413). All such samples had less tensile force at failure than a thread made using the same extrusion parameters and cross-linked using BDDE as described above.
Finally, it should be noted that there are alternative ways of implementing the present invention. Accordingly, the present embodiments are to be considered as illustrative and not restrictive, and the invention is not to be limited to the details given herein, but may be modified within the scope and equivalents of the appended claims. All references and publications cited herein are incorporated by reference in their entirety.

## Claims

1. A dried thread comprised of (i) cross linked hyaluronic acid or a salt(s) thereof, wherein the cross linked hyaluronic acid has been cross linked with butanediol diglycidyl ether (BDDE), and (ii) non-cross linked hyaluronic acid or a salt(s) thereof.

2. The thread of claim 1, further comprising a needle, and wherein the dried thread includes an end attached to the needle.

3. The thread of claim 1, wherein the dried thread has a diameter of between about 0.0001 and about 0.020 in.

4. The thread of claim 1, wherein the degree of cross linking between the hyaluronic acid or salt thereof and the cross linking agent is between about 0.01% and about 20%.

5. The thread of claim 1, wherein the dried thread has sufficient mechanical strength for dermal implantation.

6. The thread of claim 1, wherein the dried thread will fail at a tensile force of between about 0.1 lbs and about 5 lbs.

7. The thread of claim 1, wherein the dried thread has an ultimate tensile strength of between about 1 kpsi and about 125 kpsi.

8. The thread of claim 1 wherein the thread is swellable when hydrated.

9. A dermal filler comprising the thread of claim 1.

10. A method of treating a wrinkle comprising implanting in a subject in need thereof the thread of claim 1 beneath the wrinkle.

11. The method of claim 10, wherein the method comprises attaching the thread to a needle; inserting the distal end of the needle through the skin surface of the subject into the dermis of the subject adjacent to the wrinkle; traversing the dermis of the subject under the wrinkle with the needle; forcing the needle through the skin surface of the subject; and cutting the thread from the needle at the skin surface of the subject.

12. A dried thread comprised of crosslinked hyaluronic acid or a salt(s) thereof, wherein the cross linked hyaluronic acid has been cross linked with butanediol diglycidyl ether (BDDE).

13. A dried thread comprised of crosslinked hyaluronic acid or a salt(s) thereof, wherein the crosslinked hyaluronic acid consists essentially of hyaluronic acid that has been crosslinked with butanediol diglycidyl ether (BDDE).

14. A dried thread comprised of cross linked hyaluronic acid or a salt(s) thereof, wherein the cross linked hyaluronic acid has been cross linked with butanediol diglycidyl ether (BDDE);
wherein the thread is a single strand thread; and
wherein the thread is capable of being hydrated, and when hydrated will swell in diameter by about 25% and about 100%.

15. The thread of claim 14 wherein the degree of crosslinking is between about 0.1% and about 10%.

16. The thread of claim 15 wherein the degree of crosslinking is between about 1% and about 8%.

17. The thread of claim 14 wherein the thread persists in vivo between 1 and 9 months after implantation in the dermis.