WO2019135237A1 - Échafaudage pour le soudage de tissus - Google Patents

Échafaudage pour le soudage de tissus Download PDF

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Publication number
WO2019135237A1
WO2019135237A1 PCT/IL2019/050023 IL2019050023W WO2019135237A1 WO 2019135237 A1 WO2019135237 A1 WO 2019135237A1 IL 2019050023 W IL2019050023 W IL 2019050023W WO 2019135237 A1 WO2019135237 A1 WO 2019135237A1
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WO
WIPO (PCT)
Prior art keywords
composition
tissue
scaffold
particles
radiation
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PCT/IL2019/050023
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English (en)
Inventor
Tal Dvir
Maayan MALKI
Sharon FLEISCHER
Original Assignee
Ramot At Tel-Aviv University Ltd.
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Publication date
Application filed by Ramot At Tel-Aviv University Ltd. filed Critical Ramot At Tel-Aviv University Ltd.
Publication of WO2019135237A1 publication Critical patent/WO2019135237A1/fr

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Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
    • A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
    • A61L27/56—Porous materials, e.g. foams or sponges
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
    • A61L27/36—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix
    • A61L27/38—Materials for grafts or prostheses or for coating grafts or prostheses containing ingredients of undetermined constitution or reaction products thereof, e.g. transplant tissue, natural bone, extracellular matrix containing added animal cells
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L27/00—Materials for grafts or prostheses or for coating grafts or prostheses
    • A61L27/50—Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61F—FILTERS IMPLANTABLE INTO BLOOD VESSELS; PROSTHESES; DEVICES PROVIDING PATENCY TO, OR PREVENTING COLLAPSING OF, TUBULAR STRUCTURES OF THE BODY, e.g. STENTS; ORTHOPAEDIC, NURSING OR CONTRACEPTIVE DEVICES; FOMENTATION; TREATMENT OR PROTECTION OF EYES OR EARS; BANDAGES, DRESSINGS OR ABSORBENT PADS; FIRST-AID KITS
    • A61F2/00—Filters implantable into blood vessels; Prostheses, i.e. artificial substitutes or replacements for parts of the body; Appliances for connecting them with the body; Devices providing patency to, or preventing collapsing of, tubular structures of the body, e.g. stents
    • A61F2/0063—Implantable repair or support meshes, e.g. hernia meshes
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61F—FILTERS IMPLANTABLE INTO BLOOD VESSELS; PROSTHESES; DEVICES PROVIDING PATENCY TO, OR PREVENTING COLLAPSING OF, TUBULAR STRUCTURES OF THE BODY, e.g. STENTS; ORTHOPAEDIC, NURSING OR CONTRACEPTIVE DEVICES; FOMENTATION; TREATMENT OR PROTECTION OF EYES OR EARS; BANDAGES, DRESSINGS OR ABSORBENT PADS; FIRST-AID KITS
    • A61F2/00—Filters implantable into blood vessels; Prostheses, i.e. artificial substitutes or replacements for parts of the body; Appliances for connecting them with the body; Devices providing patency to, or preventing collapsing of, tubular structures of the body, e.g. stents
    • A61F2/02—Prostheses implantable into the body
    • A61F2/24—Heart valves ; Vascular valves, e.g. venous valves; Heart implants, e.g. passive devices for improving the function of the native valve or the heart muscle; Transmyocardial revascularisation [TMR] devices; Valves implantable in the body
    • A61F2/2478—Passive devices for improving the function of the heart muscle, i.e. devices for reshaping the external surface of the heart, e.g. bags, strips or bands
    • A61F2/2481—Devices outside the heart wall, e.g. bags, strips or bands
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61F—FILTERS IMPLANTABLE INTO BLOOD VESSELS; PROSTHESES; DEVICES PROVIDING PATENCY TO, OR PREVENTING COLLAPSING OF, TUBULAR STRUCTURES OF THE BODY, e.g. STENTS; ORTHOPAEDIC, NURSING OR CONTRACEPTIVE DEVICES; FOMENTATION; TREATMENT OR PROTECTION OF EYES OR EARS; BANDAGES, DRESSINGS OR ABSORBENT PADS; FIRST-AID KITS
    • A61F2250/00—Special features of prostheses classified in groups A61F2/00 - A61F2/26 or A61F2/82 or A61F9/00 or A61F11/00 or subgroups thereof
    • A61F2250/0058—Additional features; Implant or prostheses properties not otherwise provided for
    • A61F2250/0091—Additional features; Implant or prostheses properties not otherwise provided for transparent or translucent

Definitions

  • the present invention in some embodiments thereof, relates to tissue engineering and, more particularly, but not exclusively, to an artificial tissues designed for tissue welding.
  • Engineered tissues are considered a promising approach for regenerating infarcted organs.
  • cardiac patches are prepared by seeding cardiac cells within a 3D biomaterial scaffold, which provide a physical, structural and biochemical supporting microenvironment.
  • the scaffolds encourage cell-cell and cell-matrix interactions, which lead to the formation of a functioning tissue.
  • Cardiac patches are usually attached to the scar tissue of the heart by a surgical operation, involving synthetic sutures or staples. Although these procedures can attach a patch to a desired site, this surgical approach suffers from several limitations, which include the blockage of blood supply to the patch, bleeding, injury to a healthy tissue and risk of infection. Such trauma may cause an additional deterioration of the ventricle’s function, expanding the damage.
  • nanoparticle solution was used to glue two hydrogels by connecting to their polymer chains and reorganizing their structure.
  • Other approaches include neat glues that do not contain any solvents and their mechanical properties can be tuned.
  • these materials exhibit excellent adhesive properties, several properties limit their application for engineering thick tissues. For example, they are not porous, do not supply a supporting fibrous biomimetic microenvironment for tissue assembly, and in some cases their water-free environment does not allow cell penetration into their core.
  • Gold nanostructures such as spheres or particles with higher aspect ratio can be incorporated into 3D scaffolds to increase the transfer of the electrical signal between electrogenic cells.
  • Gold nanostructures can also be used for tissue welding and wound sealing.
  • tissue welding and wound sealing For example, there were some proposals to use gold nanoshells or particles in solutions that can absorb light, heat up, and locally melt tissues and fuse them. However, such techniques of fusion of tissues by heating might be destructive for viability of cells.
  • using of particles in a solution in combination with a three dimensional assembly of a tissue is limited.
  • aspects and embodiments of the present invention relate to an artificial scaffold which can be safely attached to an organ or extracellular matrix in a subject by using low-energy radiation welding.
  • composition-of-matter that includes a porous scaffold that includes a polymer; and a plurality of particles attached to the polymer, wherein the scaffold is having at least one region that is translucent to radiation, the particles are capable of generating heat upon interacting with the radiation, and the heat renders the polymer reactive to bonding with a tissue in a subject
  • the region is located at a perimeter, an edge, a frame, and/or an end of the scaffold.
  • the region is located where the scaffold is making a close intimate contact with the tissue.
  • the amount of the particles ranges from 0.001 mg to 10 mg per 1 gram of the scaffold.
  • the radiation is near infrared radiation having a wavelength that ranges from 700 nm to 950 nm.
  • the region admits at least 20 % of the radiation.
  • the particles comprise a substance and/or have a size and/or have a shape that is conducive to the generation of heat upon exposure to NIR.
  • the shape of the particles is an anisotropic shape.
  • the size of the particles ranges from 0.5 nm to 2 pm.
  • the substance of the particles is having a physical property selected from the group consisting of electric conductance, anisotropy, ferromagnetism, ferrimagnetism, diamagnetism, superparamagnetism or paramagnetism.
  • the particles comprise a metal and/or a metal oxide.
  • the metal is selected from the group consisting of gold, silver, aluminum, iron, nickel, copper, platinum, titanium, and any blends and alloys thereof.
  • the particles comprise an organic material.
  • the particles comprise a substance selected from the group consisting of a carbon allotrope, a composite material, polyaniline, polypyrrole, and blends thereof.
  • the particles have a core-coat structure.
  • the substance of the core and/or the coat is selected from the group consisting of a conductive substance, a dielectric substance, a metal, an organic substance, an inorganic substance, and any mixture thereof.
  • the substance of the core and/or the coat is selected from the group consisting of gold, silver, an organic polymer, a ceramic and a metal oxide.
  • the metal is gold
  • the particles are having an aspect ratio that ranges from 2 to 10.
  • the radiation is near IR characterized by a wavelength that ranges from 750 nm to 850 nm.
  • the translucent region in the scaffold is having a thickness that ranges from 50 pm to 100 pm.
  • the scaffold is having a form selected from the group consisting of a patch, a sheet, a mesh, a rod, a tube, a ring, and a three- dimensional object.
  • the polymer is having a shape selected from the group consisting of fibers, ribbons, flakes, spheres, coils, helicoils, beads-on-a-string, and any combination thereof.
  • the ribbons are having a mean width that ranges from 100 nm to 10 pm, and mean thickness that ranges from 100 nm to 10 pm.
  • the fibers are having a thickness that ranges from 1 pm to 0.1 pm.
  • the polymer is in a form of electrospun fibers or ribbons.
  • the polymer exhibits a plurality of functional groups capable of binding the particles.
  • the polymer exhibits a plurality of functional groups capable of the bonding upon exposure to the heat.
  • the polymer comprises a protein, a polysaccharide, alginate, chitosan, a polycaprolactone, polyglycolic acid, poly(L-lactide), a poly(ester urethane), polydioxanone, polyethylene glycol, and any copolymer thereof.
  • the protein is selected from the group consisting of albumin, collagen, fibrin, elastin, fibronectin, glycoprotein, laminin, silk, and any combination thereof.
  • the polymer includes albumin.
  • the polymer includes electrospun albumin fibers.
  • the porous scaffold is having an average pore size that ranges from 50 nm to 2 mm.
  • composition-of-matter presented herein further includes a plurality of cells distributed within and/or on the scaffold.
  • the cells are viable cells selected from the group consisting of seed cells, stem cells, mature cells, and a tissue.
  • the cells correspond to the tissue in the subject.
  • the polymer includes electrospun albumin fibers
  • the plurality of particles includes gold particles having an anisotropic shape characterized by an aspect ratio of at least 2, and further includes a plurality of viable cells distributed within and/or on the scaffold.
  • the tissue to which the composition- of-matter presented herein is attached is selected from the group consisting of a connective tissue, a muscle tissue, a nervous tissue and an epithelial tissue.
  • the muscle tissue is selected from the group consisting of a smooth muscle, a visceral muscle, a skeletal muscle, and cardiac muscle.
  • the connective tissue is selected from the group consisting of an extracellular matrix, a fibrous connective tissue, a skeletal connective tissue, a fascia, a bone, a tendon, a ligament, an adipose tissue and an areolar tissue.
  • a method of attaching/fusing/welding the composition-of-matter presented herein to a tissue in a subject in need thereof includes:
  • the radiation is emitted on the composition by a radiation source capable of emitting a flux of at least 1 W/cm 2 .
  • the irradiating is effected for at least 60 seconds.
  • the composition-of-matter presented herein is used as a patch and the tissue in the subject is selected from the group consisting of a cardiac muscle, a visceral muscle, a skeletal muscle, a fascia, and an abdominal wall.
  • composition-of-matter presented herein includes:
  • the process further includes, prior or subsequent to the contacting, introducing viable cells into the porous scaffold under conditions suitable for maintaining the viable cells.
  • a method of treating a tissue defect in a subject in need thereof is effected by:
  • composition-of-matter presented herein with the tissue in the subject such that at least the radiation-translucent region(s) overlap with healthy parts of the tissue surrounding the tissue defect, and the composition is covering the tissue defect;
  • the radiation is emitted on the composition by a radiation source capable of emitting a flux of at least 1 W/cm2.
  • the irradiating is effected for at least 60 seconds.
  • the tissue defect is selected from the group consisting of a damaged cardiac muscle, an inguinal hernia, an incisional hernia, a femoral hernia, a skeletal muscle tear, and a damaged skin.
  • FIGs. 1A-D present schematic illustrations of the process of preparing a composition-of- matter, according to some embodiments of the present invention, designed as a cardiac patch, and using the same in a tissue welding process, showing the adsorption of gold nanorods (FIG.
  • FIG. 1C cardiac patch ready for use
  • FIG. 1D NIR laser source
  • FIG. 2A shows HRTEM micrographs of AuNRs (scale bar: 50 nm)
  • FIG. 2B shows the UV-visible spectra wavescan of AuNRs
  • FIG. 2C shows SEM micrographs of electrospun albumin fiber scaffolds (scale bar: 100 pm)
  • FIG. 2D shows SEM micrographs of
  • FIG. 2F shows ESEM micrographs of AuNRs adsorbed to electrospun albumin fiber scaffold (scale bar: 2 pm)
  • FIG. 2G shows ESEM micrographs AuNRs adsorbed to electrospun albumin fiber scaffold (scale bar: 500 nm)
  • FIG. 2H shows XRD spectroscopy confirming the existence of Au on the albumin fibers;
  • FIGs. 3A-F present mechanical properties and viability of the engineered tissues, according to some embodiments of the present invention, welded to a porcine heart, wherein FIG. 3A shows a stress versus laser power flux plot, FIG. 3B shows a stress versus time of NIR irradiation plot, FIG. 3C shows schematics of the geometry of the albumin fiber scaffold used for tissue welding, FIG. 3D shows a stress versus thickness of albumin scaffold plot, FIG. 3E shows viability of cardiomyocyte on electrospun albumin fiber scaffolds before and after irradiation, and FIG. 3F shows immunofluorescence image of cardiac a-sarcomeric actinin (pink), connexin- 43 (green) and cell nuclei (blue) (scale bar: 20 pm); and
  • FIGs. 4A-E present illustrations and characterization of the cardiac patch welded to a heart
  • FIG. 4A shows a schematic representation of the welding/integration process
  • FIG. 4B shows the cardiac patch welded to rat heart
  • FIG. 4C is a SEM micrograph of cross section of the welded heart showing the interaction between the patch and the heart (scale bar: 200 pm)
  • FIG. 4D is a larger magnification SEM micrograph of a cross section of the welded heart showing the interaction between the patch and the heart (scale bar: 50 pm)
  • FIG. 4E shows the H&E staining of interaction between the patch and the heart (scale bar: 500 pm).
  • the present invention in some embodiments thereof, relates to tissue engineering and, more particularly, but not exclusively, to an artificial tissues designed for tissue welding.
  • a composite scaffold (a composition-of-matter), comprising albumin electrospun fibers and gold nanorods (AuNRs) is provided herein.
  • AuNRs gold nanorods
  • the engineered tissue was then positioned on the myocardium and irradiated with near IR laser (808 nm). Due to the definitive and purposeful design of the patch, the AuNRs were able to absorb the low-energy NIR radiation sufficiently to convert it into thermal energy, which locally changed the molecular structure of the fibrous scaffold, allowing it to attach itself to the wall of the heart tissue strongly but safely.
  • Such hybrid biomaterials can be used to integrate any engineered tissue with any defected organs, while minimizing the risk of additional injury for the subject, caused by the conventional stitching methods.
  • aspects of the present invention are drawn to a scaffold which can be safely attached to a living tissue, an organ or an extracellular matrix in a live subject by using near infrared (NIR) radiation.
  • NIR near infrared
  • a composition-of-matter which is based on a porous scaffold made of a polymer; and a plurality of particles attached to the polymer, wherein the scaffold is having at least one region that is translucent to radiation, the particles are capable of generating heat upon interacting with the radiation, and the heat renders the polymer in the scaffold reactive to bonding with a tissue in a subject.
  • bonding is also referred to as “fusing”, welding” and“merging” between the scaffold and the tissue. Bonding is formed upon irradiation of the particles-containing scaffold and the tissue, whereas the radiation is converted into heat by the particles, and the heat renders the polymer in the scaffold and the parts of the tissue that are in close intimate contact with the scaffold reactive towards one-another, thereby effecting the bonding.
  • the term“bonding” refers to any form of chemical, physical or mechanical attachment of the at least part of the polymer in the scaffold to the tissue in the subject. Bonding, in the sense of embodiments of the present invention, includes forming hydrogen bonds, covalent bonds, salt-bridge bonds, disulfide bonds, aromatic interactions and the likes, between functional groups in the scaffold and corresponding functional groups in the tissue.
  • the term“bonding” also encompasses entanglement of filament, strands and chains in the scaffold and the tissue.
  • the radiation-translucent region(s) is/are located at a perimeter, an edge, a frame, and/or an end of the scaffold, or in the alternative, the translucent region is located where the scaffold is intended for attachment with the tissue, or intended for making a close intimate contact with the tissue.
  • the amount of the particles per 1 gram of the scaffold ranges from 0.001 mg to 10 mg, or at least 0.001 mg, 0.01 mg, 0.1 mg, 1 mg, or at least 10 mg per 1 gram of the scaffold.
  • the radiation is a low-energy radiation which is substantially non-deleterious to living cells, such as near infrared radiation having a wavelength that ranges from 700 nm to 950 nm.
  • the radiation-translucent region(s) in the scaffold admit at least 5, 10, 15, 20, 30, 40 or 50 percent of the radiation.
  • the particles are selected, manufactured and designed to interact with the low-energy radiation and generate heat upon exposure to the radiation; hence, the particles comprise a certain substance and/or have a certain size and/or have a certain shape that is conducive to the generation of heat upon exposure to low- energy radiation such as NIR.
  • the shape of the particles is an anisotropic shape (non-spherical, elongated, elliptical, or oddly/irregularly shaped.
  • the size of the particles ranges from 0.5 nm to 2 pm, or less than 100 pm, less than 50 pm, less than 10 pm, less than 10 pm, less than 1 pm, less than 0.1 pm, less than 50 nm, less than 10 nm, less than 5 nm, or less than 1 nm.
  • the substance of the particles is having a physical property selected from the group consisting of electric conductance, anisotropy, ferromagnetism, ferrimagnetism, diamagnetism, superparamagnetism or paramagnetism.
  • Such physical property may also be conducive to interaction with electric, magnetic or electromagnetic radiation that is harmless to cells and can further be employed by the generated tissue.
  • electrically conductive particles may assist in the process of forming nerve cell connections.
  • the particles comprise a metal and/or a metal oxide, wherein the metal may be, without limitation, gold, silver, aluminum, iron, nickel, copper, platinum, titanium, and any blends and alloys thereof.
  • the particles comprise an organic material.
  • the particles comprise a substance selected from the group consisting of a carbon allotrope, a composite material, polyaniline, polypyrrole, and blends thereof.
  • the particles have a core-coat structure, as these radiation-responsive particles are known in the art.
  • the substance of the core and/or the coat is selected from the group consisting of a conductive substance, a dielectric substance, a metal, an organic substance, an inorganic substance, and any mixture thereof.
  • the substance of the core and/or the coat is selected from the group consisting of gold, silver, an organic polymer, a ceramic and a metal oxide.
  • the metal is gold
  • the particles are having an aspect ratio that ranges from 2 to 10.
  • the radiation is near IR characterized by a wavelength that ranges from 750 nm to 850 nm.
  • the translucent region in the scaffold is having a thickness that is configured to admit sufficient flux of radiation to allow the particles adsorbed thereon/therein to generate sufficient heat to case welding of the scaffold to the tissue.
  • the optimal thickness depends on the type of polymer used in the scaffold, the amount and type of particles adsorbed therein, and other practical considerations. Hence, the thickness of the region is about 40 pm, 50 pm, 60 pm, 70 pm, 80 pm, 90 pm, 100 pm, 110 pm, 120 pm, 150 pm, 200 pm, or ranges from 50 to 100 pm.
  • the scaffold is having a form selected from the group consisting of a patch, a sheet, a mesh, a rod, a tube, a ring, and a three- dimensional object.
  • the polymer is having a shape selected from the group consisting of fibers, ribbons, flakes, spheres, coils, helicoils, beads-on-a-string, and any combination thereof.
  • the ribbons are having a mean width that ranges from 100 nm to 10 pm, and mean thickness that ranges from 100 nm to 10 pm.
  • the fibers are having a thickness that ranges from 1 pm to 0.1 pm.
  • the polymer is in a form of electrospun fibers or ribbons.
  • the polymer exhibits a plurality of functional groups capable of binding the particles.
  • a protein-based scaffold may exhibit a plurality of thiol functional groups, to which gold nanoparticles can bind spontaneously.
  • the polymer exhibits a plurality of functional groups capable of the bonding to a tissue upon exposure to the heat generated locally by the interaction of the radiation with the particles.
  • hydrogen-bond donor/acceptor functional groups may be rendered free for inter-molecular bonding between the polymer of the scaffold and the tissue.
  • the polymer comprises a protein, a polysaccharide, alginate, chitosan, a polycaprolactone, polyglycolic acid, poly(L-lactide), a poly(ester urethane), polydioxanone, polyethylene glycol, and any copolymer thereof.
  • the protein is selected from the group consisting of albumin, collagen, fibrin, elastin, fibronectin, glycoprotein, laminin, silk, and any combination thereof.
  • the polymer includes albumin.
  • the polymer includes electrospun albumin fibers.
  • the porous scaffold is having an average pore size that ranges from 50 nm to 2 mm.
  • composition-of-matter presented herein further includes a plurality of cells distributed within and/or on the scaffold.
  • the cells are viable cells selected from the group consisting of seed cells, stem cells, mature cells, and a tissue.
  • the cells correspond to the tissue in the subject, namely the type of cells that is being introduced into the scaffold corresponds to the type of cells of the tissue, to which the composition-of-matter presented herein is intended to be fused.
  • the cells are harvested from the subject, and in some embodiments the cells are harvested from a healthy section of the tissue that is being treated.
  • the polymer includes electrospun albumin fibers
  • the plurality of particles includes gold particles having an anisotropic shape characterized by an aspect ratio of at least 2, and further includes a plurality of viable cells distributed within and/or on the scaffold.
  • the tissue to which the composition- of-matter presented herein can be attached/fused/welded includes without limitation, a connective tissue, a muscle tissue, a nervous tissue and an epithelial tissue.
  • the muscle tissue includes without limitation, a smooth muscle, a visceral muscle, a skeletal muscle, and cardiac muscle.
  • the connective tissue includes without limitation, an extracellular matrix, a fibrous connective tissue, a skeletal connective tissue, a fascia, a bone, a tendon, a ligament, an adipose tissue and an areolar tissue.
  • a method of attaching/fusing/welding the composition-of-matter presented herein to a tissue in a subject in need thereof includes:
  • composition placed on the tissue; and irradiating at least the translucent region(s) in the composition with radiation suitable for absorption by the particles.
  • the radiation is emitted on the composition by a radiation source capable of emitting a flux of at least 0.5, 1, 2, 3, 4, 5, 6, 7, 8,
  • the irradiating is effected for at least 5,
  • composition-of-matter presented herein includes:
  • the process further includes, prior or subsequent to the contacting, introducing viable cells into the porous scaffold under conditions suitable for maintaining the viable cells.
  • a method of treating a tissue defect in a subject in need thereof is effected by:
  • composition-of-matter presented herein with the tissue in the subject such that at least the radiation-translucent region(s) overlap with healthy parts of the tissue surrounding the tissue defect, and the composition is covering the tissue defect;
  • the tissue defect is a necrotic or gangrenous tissue
  • the composition-of-matter presented herein is fabricated as patch designed to reconstruct the tissue after the damaged part of the tissue had been surgically removed or cleared.
  • the patch is an engineered tissue containing viable cells that correspond to the type of cells in the treated tissue in the subject.
  • the tissue defect is selected from the group consisting of a damaged cardiac muscle, an inguinal hernia, an incisional hernia, a femoral hernia, a skeletal muscle tear, and a damaged skin.
  • the composition-of-matter presented herein is useful in treating cardiac muscle defects, hernia and muscle tear or rupture.
  • all technical and/or scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the invention pertains.
  • methods and materials similar or equivalent to those described herein can be used in the practice or testing of embodiments of the invention, exemplary methods and/or materials are described below. In case of conflict, the patent specification, including definitions, will control.
  • the materials, methods, and examples are illustrative only and are not intended to be necessarily limiting
  • composition-of-matter comprising a porous scaffold that comprises a polymer; and a plurality of particles attached to said polymer, wherein the scaffold is having at least one region that is translucent to radiation, the particles are capable of generating heat from said radiation, and the heat renders the polymer reactive to bonding with an extracellular matrix in a subject, is intended to include all such new technologies a priori.
  • compositions, method or structure may include additional ingredients, steps and/or parts, but only if the additional ingredients, steps and/or parts do not materially alter the basic and novel characteristics of the claimed composition, method or structure.
  • the phrases “substantially devoid of” and/or “essentially devoid of” in the context of a certain substance refer to a composition that is totally devoid of this substance or includes less than about 5, 1, 0.5 or 0.1 percent of the substance by total weight or volume of the composition.
  • the phrases "substantially devoid of” and/or “essentially devoid of” in the context of a process, a method, a property or a characteristic refer to a process, a composition, a structure or an article that is totally devoid of a certain process/method step, or a certain property or a certain characteristic, or a process/method wherein the certain process/method step is effected at less than about 5, 1, 0.5 or 0.1 percent compared to a given standard process/method, or property or a characteristic characterized by less than about 5, 1, 0.5 or 0.1 percent of the property or characteristic, compared to a given standard.
  • the term“exemplary” is used herein to mean“serving as an example, instance or illustration”. Any embodiment described as“exemplary” is not necessarily to be construed as preferred or advantageous over other embodiments and/or to exclude the incorporation of features from other embodiments.
  • a compound or “at least one compound” may include a plurality of compounds, including mixtures thereof.
  • range format is merely for convenience and brevity and should not be construed as an inflexible limitation on the scope of the invention. Accordingly, the description of a range should be considered to have specifically disclosed all the possible subranges as well as individual numerical values within that range. For example, description of a range such as from 1 to 6 should be considered to have specifically disclosed subranges such as from 1 to 3, from 1 to 4, from 1 to 5, from 2 to 4, from 2 to 6, from 3 to 6 etc., as well as individual numbers within that range, for example, 1, 2, 3, 4, 5, and 6. This applies regardless of the breadth of the range.
  • process and “method” refer to manners, means, techniques and procedures for accomplishing a given task including, but not limited to, those manners, means, techniques and procedures either known to, or readily developed from known manners, means, techniques and procedures by practitioners of the chemical, material, mechanical, computational and digital arts.
  • the term“treating” includes abrogating, substantially inhibiting, slowing or reversing the progression of a condition, substantially ameliorating clinical or aesthetical symptoms of a condition or substantially preventing the appearance of clinical or aesthetical symptoms of a condition. It is appreciated that certain features of the invention, which are, for clarity, described in the context of separate embodiments, may also be provided in combination in a single embodiment. Conversely, various features of the invention, which are, for brevity, described in the context of a single embodiment, may also be provided separately or in any suitable subcombination or as suitable in any other described embodiment of the invention. Certain features described in the context of various embodiments are not to be considered essential features of those embodiments, unless the embodiment is inoperative without those elements.
  • a cetyl trimethylammonium bromide solution (CTAB, 15.3 mL, 0.20 M) was mixed with 1.7 mL of 1.016 mM HAuCL. The solution was separated into two solutions, seed and growth (2 ml and 16 ml, respectively). Approximately 0.12 mL of ice-cold 0.01 M NaBH 4 was added to the seed solution, resulting in the formation of a brownish-yellow solution. The seed solution was vigorously stirred for 3 minutes. Following, it was kept at 25 °C for 1 hour to ensure total decomposition of the borohydride. Ascorbic acid (0.15 ml, 0.115 M) and of AgN0 3 (0.15 ml, 20 mM) were added to the growth solution.
  • Ascorbic acid (0.15 ml, 0.115 M
  • AgN0 3 (0.15 ml, 20 mM
  • the seed solution was diluted in distilled deionized water at a ratio of 1 to 10. Finally, 0.18 mL of the seed solution was added to the growth solution. The final solution was kept for at least 4 hours at 32 °C, reaching a concentration of 20 mg/ml.
  • the AUNRs exhibited strong absorbance at approximately 808 nm.
  • Bovine serum albumin [BSA; Fraction V, MP Biomedicals, Aurora, OH; 14 % (w/v)] was dissolved in tetrafluoroethylene (TFE) and distilled water (9:1, respectively), following, excess of b-mercaptoethanol (Merck, Darmstadt, Germany) was added for overnight reaction.
  • TFE tetrafluoroethylene
  • b-mercaptoethanol Merck, Darmstadt, Germany
  • AuNRs with concentration of 50 mg/ml were deposited on carbon coated copper grids (SPI) for transmission electron microscopy (TEM) analysis. Images were captured using FEITM F20 Philip s-Tecnai equipped with FEG (Schottky type).
  • Samples were mounted onto aluminum stubs with conductive paint and sputter-coated with an ultrathin (150 A) layer of gold in a Polaron E 5100 coating apparatus (Quorum technologies, Laughton, UK). The samples were viewed under JCM-6000PLUS NeoScope Benchtop (JEOL USA Inc., Peabody, MA).
  • the constructs were fixed with 2.5 % glutaraldehyde (24 h at 4 °C), followed by graded series of ethanol-water solutions for dehydration (25-100 %). All samples were critical point dried, sputter-coated with gold in a Polaron E 5100 coating apparatus (Quorum technologies, Lewis, UK) and observed under JSM-840A SEM (JEOL, Tokyo, Japan).
  • AuNRs solution were suspended on albumin scaffold and imaged without additional coating using a Quanta 200 FEG Environmental Scanning Electron Microscope (ESEM) with a field-emission gun (FEG) electron source. Imaging was carried out under low vacuum with a high tension of 20 kV and a working distance of 10 mm. Elemental composition was performed using X-Max 20, Oxford EDX and acquired using INCA Software.
  • ESEM FEG Environmental Scanning Electron Microscope
  • FEG field-emission gun
  • Cardiac cells were isolated according to Tel Aviv University ethical use protocols. Briefly, left ventricles of 0-3-day-old neonatal Sprague-Dawley rats (Envigo Laboratories, Israel) were harvested, and cells were isolated using six cycles (30 min each at 37 °C) of enzyme digestion with collagenase type II (95 U/mL; Worthington, Lakewood, NJ) and pancreatin (0.6 mg/mL; Sigma- Aldrich) in Dulbecco’s modified Eagle Medium (DMEM, CaCl 2 -2H 2 0 (1.8 mM), KC1 (5.36 mM), MgS0 4 -7H 2 0 (0.81 mM), NaCl (0.1 M), NaHCOs (0.44 mM), NaH 2 P0 4 (0.9 mM)).
  • DMEM Dulbecco’s modified Eagle Medium
  • KC1 5.36 mM
  • Cardiac cells (8-10 5 ) were seeded onto the scaffolds by adding 10 pL of the suspended cells followed by a 45 minutes incubation period (Humidified incubator, 37 °C, 5 % C0 2 ). Following, cell constructs were supplemented with culture medium (5 % FBS) and further incubated.
  • Cardiac tissue integration to heart tissues by laser irradiation Cardiac tissue integration to heart tissues by laser irradiation:
  • AuNRs solution (3.5 pl, 20 mg/ml) was suspended on the edges of albumin scaffolds. Following, the scaffolds were placed on the rat heart tissue and irradiated with ranging laser power fluxes (1, 1.2, and 1.5 W/cm 2 ) and ranging time (60, 90, 120 sec).
  • An 808 nm diode laser was used as a radiation source (MDL-III-808, 0-2.5W continuous wave output; Optoengine, Midvale, UT) and a 400 pm fiber optic cable was used.
  • the fiber optic was attached to a silica- lens collimator with a 22.2-mm aperture (CeramOptec Industries, Changchun, China). The adhesion was tested using a Lloyd tensile testing instrument (model LS1) with a 20 N load cell at a rate of 5 mm per minute.
  • compositions-of-matter in the form of cardiac tissues, prepared according to some embodiments of the present invention, were integrated to porcine heart as previously described. Following, 6 replicates of the samples were cultured immediately after irradiation in Presto Blue viability assay (Life Technologies, NY). As a control group 6 replicates of cardiac tissues with AuNRs, but no irradiation was cultured in Presto Blue as well.
  • Cell constructs were fixed and permeabilized in 100 % cold methanol for 10 minutes, washed three times in DMEM-based buffer and then blocked for 1 hour at room temperature in DMEM -based buffer containing 2 % FBS, after which the samples were washed three times.
  • Cardiac tissues were incubated with primary mouse anti a-sarcomeric actinin antibody (1:750, Sigma- Aldrich) and connexin 43 (1:250, Invitrogen, Carlsbad, CA), washed three times and incubated for 1 hour with Alexa Fluor 647 conjugated goat anti-mouse antibody (1:500; Jackson, West Grove, PA) and Alexa Fluor 488 conjugated goat anti-rabbit antibody (1:500; Jackson).
  • H&E Haematoxylin and eosin stain
  • Cardiac tissues were soldered to rat tissue as previously described. Following constructs were fixed in 4% formalin (24 h at 40° C) and embedded in OCT. Sections (6 pm thick) were prepared using a cryotomeTM FSE (Thermo scientific) and affixed to X-tra® adhesive glass slides (Leica Biosystems, Wetzler, Germany), dehydrated in graduated ethanol steps (70-100%), and stained with hematoxylin and eosin. Samples were visualized using an inverted fluorescence microscope (Nikon Eclipse TI).
  • FIGs. 1A-D present schematic illustrations of the process of preparing a composition-of- matter, according to some embodiments of the present invention, designed as a cardiac patch, and using the same in a tissue welding process, showing the adsorption of gold nanorods (FIG.
  • FIG. 1C cardiac patch ready for use
  • FIG. 1D NIR laser source
  • a proof of concept of some embodiments of the present invention was carried out by preparing engineered tissue comprising electrospun albumin scaffolds and gold nanorods, of which their attachment to tissues is made by NIR radiation.
  • FIGs. 2A-D illustrate an example of a 3D porous scaffold of albumin electrospun fiber, according to an embodiment of the present invention, where mono-dispersed AuNRs with a mean size of about 60 nm x 20 nm were incorporated.
  • the AuNRs which are shown in FIG. 2A, were synthesized so as to allow rapid and efficient conversion of a laser radiation with a wavelength of about 800 nm to thermal energy.
  • Such AuNRs can be characterized by TEM and UV-vis-NIR. The UV-vis spectrum of those AuNRs showed a sharp and strong absorption centered at about 810 nm as shown in FIG. 2B.
  • the preparing of the structure of the scaffolds included dissolving albumin in trifluoroethanol and distilled water, and adding b-mercaptoethanol after 24 hours.
  • the solution was electrospun at room temperature at a flow rate of 2 mL/hour under an electrical field of 12.5 kV.
  • the resulting scaffolds were composed of ribbon-like fibers with a mean width of 2.4 pm pm and thickness of 0.5 pm (FIG. 2C and FIG. 2D).
  • the AuNRs were integrated into the albumin scaffolds by soaking the scaffolds in an AuNRs solution during 60 min to allow quick adsorption.
  • the spontaneous attachment of the AuNRs onto the structures of the scaffolds turned the color of the scaffolds into brown, while leaving the solution transparent (FIG. 2E).
  • Surface analysis by scanning electron microscope images and EDX revealed a homogenous distribution of the AuNRs on the surface of the scaffolds fibers (FIGs. 2F-H).
  • FIG. 3C Shaping thin edges at the perimeters of thick scaffolds facilitated the penetration of sufficient radiation through the scaffolds.
  • FIG. 3D The functionality of different thicknesses, irradiated with 1.5 W/m 2 during 120 seconds, is shown in FIG. 3D. As shown, a thickness of 60- 80 pm facilitates significantly stronger attachment to a tissue, as compared to 100-120 pm probably due to better penetration of the light.
  • Non-destructiveness for cells of the thermal energy generated by the AuNRs after irradiated with near IR for 120 seconds, at 1.5 W/cm 2 was demonstrated by isolating cardiac cells from the left ventricles of neonatal rat hearts and seeding them within the hybrid scaffolds. After 7 days of incubation, which is considered sufficient period for cells’ organization and tissue assembly, the cardiac patches prepared as presented herein, were irradiated. As shown in FIG. 4E, exposure to near IR did not affect cells’ viability. Following tissue irradiation, the patches were fixed and immunostained for cardiac sarcomeric actinin and connexin 43 (Cx43), proteins associated with cell contraction and electrical coupling, respectively.
  • Cx43 connexin 43
  • the cardiac cells exhibited massive actinin staining, indicating on the contraction ability of the assembled tissue. Furthermore, pronounced Cx43 expression was observed between the cardiomyocytes, indicating on the ability of the cells to function synchronously. Such hallmarks further indicated that the exposure to near IR and the local heating by the AuNRs did not harm the engineered tissue.
  • Demonstration for attachment to a heart in wet and dynamic conditions was carried by opening the chest of Sprague Dawley rats, exposing the heart and placing a patch prepared as presented herein on the left ventricle. Then the patch was irradiated with near IR for 120 seconds as illustrated in FIG. 1A.
  • FIG. 1B Scanning electron microscope images of the sliced heart reveal a tight interaction between the heart and the patch (FIG. 1C). Furthermore, the thin edges were able to strongly secure the thick tissue layer on the heart, ensuring a strong fixation (FIG. 4C-E). As shown, no changes in the scaffolds’ structure could be observed. This further indicates on the safety of the procedure.
  • the physical properties of the AuNRs were utilized to provide a suture- free engraftment of a cardiac patch to the heart.
  • Such technology embodying aspects of the present invention may be implemented to integrate various engineered tissues or pure biomaterials with other defected organ, minimizing the risk of additional injury for the patient.

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Abstract

La présente invention concerne une composition de matière, comprenant un échafaudage poreux qui comprend un polymère ; et une pluralité de particules fixées audit polymère, l'échafaudage comprenant au moins une région qui est translucide à un rayonnement de faible énergie, les particules pouvant générer de la chaleur dès l'interaction avec le rayonnement, et la chaleur générée rendant le polymère réactif pour se lier à des tissus chez un sujet.
PCT/IL2019/050023 2018-01-04 2019-01-03 Échafaudage pour le soudage de tissus WO2019135237A1 (fr)

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Citations (4)

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Publication number Priority date Publication date Assignee Title
US5292362A (en) * 1990-07-27 1994-03-08 The Trustees Of Columbia University In The City Of New York Tissue bonding and sealing composition and method of using the same
US20030093092A1 (en) * 2001-09-26 2003-05-15 West Jennifer L. Optically-absorbing nanoparticles for enhanced tissue repair
US6773699B1 (en) * 2001-10-09 2004-08-10 Tissue Adhesive Technologies, Inc. Light energized tissue adhesive conformal patch
US20120277773A1 (en) * 2011-04-27 2012-11-01 Tyco Healthcare Group Lp Attachment of a Biomaterial to Tissue

Patent Citations (4)

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Publication number Priority date Publication date Assignee Title
US5292362A (en) * 1990-07-27 1994-03-08 The Trustees Of Columbia University In The City Of New York Tissue bonding and sealing composition and method of using the same
US20030093092A1 (en) * 2001-09-26 2003-05-15 West Jennifer L. Optically-absorbing nanoparticles for enhanced tissue repair
US6773699B1 (en) * 2001-10-09 2004-08-10 Tissue Adhesive Technologies, Inc. Light energized tissue adhesive conformal patch
US20120277773A1 (en) * 2011-04-27 2012-11-01 Tyco Healthcare Group Lp Attachment of a Biomaterial to Tissue

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Title
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