WO2011031301A2 - Intrapericardial delivery of periostin - Google Patents

Abstract

Compositions and methods for delivery of periostin or fragments or variants thereof to or near the pericardial space are provided. In some embodiments, a sustained release vehicle is used to deliver periostin or fragments or variants thereof in a controlled manner to treat the heart of a subject. The sustained release vehicle may be, for example, a hydrogel particle. In certain embodiments, compositions and methods are provided for inhibiting formation of tissue adhesions within the pericardial space.

Description

INTRAPERICARDIAL DELIVERY OF PERIOSTIN

RELATED APPLICATIONS

This application claims the benefit of U.S. Provisional Patent Application Serial No. 61/237,971 , filed August 28, 2009, entitled "Intrapericardial Injections," by Ladage et al.

FIELD OF INVENTION

The present invention relates to compositions and methods for delivery of periostin to the pericardial space.

BACKGROUND

One of the major challenges in the pharmacological treatment of heart diseases is to achieve delivery of suitable concentrations of therapeutic agents to the specific target site. Various approaches for local delivery to the heart include intramyocardial injections, epicardial deposition, and intracoronary or transvascular application. The efficacy of intramyocardial injection is limited by retention and survival rates of 2% or less. Epicardial deposition of can be more effective, but is highly invasive. In the setting of vascular obstruction, such as myocardial infarction, reduced local blood supply can significantly impair targeted agent delivery via the vasculature.

It has been discovered that periostin, a component of the extracellular matrix, and fragments thereof promote cardiomyocyte proliferation and myocardial regeneration. After myocardial infarction,recombinant periostin induces cardiomyocyte cell cycle reentry, improves cardiac remodeling and function, reduces fibrosis and infarct size, and increases angiogenesis. While periostin has been found to be an effective treatment for myocardial infarction and other vascular and cardiac conditions, improved methods of delivering periostin to the heart in an effective and safe manner would be advantageous.

SUMMARY OF THE INVENTION

This invention relates generally to compositions and methods for delivery of periostin, or fragments or variants thereof to the pericardial space of a patient. The subject matter of this invention involves, in some cases, interrelated products, alternative solutions to a particular problem, and/or a plurality of different uses of one or more systems and/or articles.

In one aspect, a method for treating the heart of a human or animal subject is provided. The method comprises injecting a polymer containing and active agent comprising periostin into the pericardial space of the subject, wherein the polymer prevents adhesion from forming between a first tissue and a second tissue in the pericardial space.

In another aspect, a method for treating the heart of a human or animal subject is provided. The method comprises injecting a plurality of particles comprising gelatin into the pericardial space of the subject, wherein the particles further comprise periostin or fragments or variants thereof and are configured to act as a sustained release vehicle for the periostin or fragments or variants thereof.

In yet another aspect, a method for treating the heart of a human or animal subject is provided. The method comprises injecting a plurality of particles having an average particle sizegreater than 500 microns into the pericardial space of the subject, wherein the particles further comprise periostin or fragments or variants thereof and are configured to act as a sustained release vehicle for the periostin.

The subject matter of this application may involve, in some cases, interrelated products, alternative solutions to a particular problem, and/or a plurality of different uses of a single system or article.

The present invention also relates to pharmaceutical compositions comprising any of the compositions and/or particles described above and herein, as well as one or more pharmaceutically acceptable carriers, additives, and/or diluents. The present invention also relates to compositions for treating a subject having a heart disease or cardiovascular condition, wherein the composition comprises any of the compositions and/or particles described above and herein. The present invention also relates to the use of any of the compositions and/or particles described above and herein in the preparation of a medicament for treating a subject having a heart disease or cardiovascular condition.

Other aspects, embodiments and features of the invention will become apparent from the following detailed description when considered in conjunction with the accompanying drawings. All patent applications and patents incorporated herein by reference are incorporated by reference in their entirety. In case of conflict, this specification, including definitions, will control.

BRIEF DESCRIPTION OF THE DRAWINGS

The accompanying drawings are schematic and are not intended to be drawn to scale. In the figures, each identical, or substantially similar component that is illustrated in various figures is typically represented by a single numeral or notation. For purposes of clarity, not every component is labeled in every figure, nor is every component of each embodiment of the invention shown where illustration is not necessary to allow those of ordinary skill in the art to understand the invention. In the drawings:

FIG. 1 shows a schematic of a heart and pericardium;

FIGs. 2A-2E are photocopies of showing various images (FIGs. 2A and 2C-2E) and a plot (FIG. 2B) demonstrating the reabsorption time of particles, according to one set of embodiments;

FIGs. 3 A-3E are plots demonstrating the study design (FIG. 3A), myocardial function determined by magnetic resonance imaging (FIG. 3B), catheterization (FIGs. 3C and 3D), and quantification of scar area visualized by delayed enhancement (FIG. 3E), according to one set of embodiments;

FIGs. 3F-3H are photocopies showing various images (FIGs. 3F and 3G) and a plot (FIG. 3H) demonstrating cardiac properties and effects of treatment, according to one set of embodiments;

FIG. 4 shows a schematic of an injection technique, according to an embodiment; FIG. 5A shows fluoroscopic images of catheter insertion, according to an embodiment;

FIG. 5B shows fluoroscopic images of liquid dye, gelfoam mixed with dye, and liquid dye after closure of the puncture site to assess possible leakage, according to an embodiment;

FIG. 5C shows the position of the injected gelfoam as well as the IVUS probe in relation to the infarct zone, according to an embodiment; and

FIGs. 6A-6C show various images and a plot demonstrating assessment of the size of the space between the heart and the pericardial membrane, according to an embodiment. DETAILED DESCRIPTION OF THE INVENTION

The present invention relates to compositions and methods for delivery of periostin or fragments or variants thereof to the pericardial space. In some embodiments, the invention utilizes a sustained release vehicle to deliver periostin or fragments or variants thereof in a controlled manner to treat the heart of a subject. The sustained release vehicle may be, for example, a hydrogel particle. In certain embodiments, compositions and methods are provided for also inhibiting formation of tissue adhesions near or within the pericardial space.

Additional description relating to the structure, properties and production of periostin or fragments or variants thereof is descrbed in International Patent Application Serial No. PCT/US2008/051659, filed January 22, 2008, entitled "Periostin Induces Proliferation of Cardiomyocytes and Promotes Cardiac Regeneration," by Kuhn, which is incorporated herein by reference in its entirety for all purposes.

As defined in the above referenced International Patent Application as is also applicable herein, the term "portion" or "fragment" as used herein refers to an amino acid sequence of the periostin genes that has fewer amino acids than the entire sequence of the periostin genes. For example, a periostin fragment can comprise one, two, three or four of the fasciclin 1 (fasl) domains. In some embodiments, the periostin fragment comprises the four fasciclin 1 (fasl) domains. In some embodiments, the periostin fragment that comprise a fasl domain can include additional amino acids to facilitate the binding of the protein fragment. For example, a periostin fragment comprising fasl can include 10%, 20%, 30%, 40% , or 50%, etc. of the amino acids comprising fas2.

"Variant" as the term is used herein, is a protein that differs from a reference protein (i.e. a periostin protein or fragment thereof consistent with embodiments of the present invention), but retains essential properties (i.e., biological activity). A typical variant of a polynucleotide differs in nucleotide sequence from another, reference polynucleotide. Changes in the nucleotide sequence of the variant may or may not alter the amino acid sequence of a polypeptide encoded by the reference polynucleotide.

Nucleotide changes may result in amino acid substitutions, additions, deletions, fusions and truncations in the polypeptide encoded by the reference sequence, as discussed below. Generally, differences are limited so that the sequences of the reference polypeptide and the variant are closely similar overall and, in many regions, identical. A variant and reference protein may differ in amino acid sequence by one or more substitutions, additions, and deletions in any combination. A substituted or inserted amino acid residue may or may not be one encoded by the genetic code. A variant of a protein may be naturally occurring such as an allelic variant, or it may be a variant that is not known to occur naturally. Non-naturally occurring variants of polynucleotides and polypeptides may be made by mutagenesis techniques or by direct synthesis. For instance, a conservative amino acid substitution may be made with respect to the amino acid sequence encoding the polypeptide.

Variant proteins encompassed by the present application are biologically active, that is they continue to possess the desired biological activity of the native protein, as described herein. The term "variant" includes any polypeptide having an amino acid residue sequence substantially identical to a sequence specifically shown herein in which one or more residues have been conservatively substituted with a functionally similar residue, and which displays the ability to mimic the biological activity of periostin, such as for example, activating integrins, phosphorylating ERK 1/2 and Akt, and/or increasing proliferation of cardiomyocytes. "Biological activity," as used herein refers to the ability of the protein to increase DNA synthesis in cardiomyocytes, as can be tested by methods known to one skilled in the art, such as, but not limited to, BrdU uptake assay. Variants may result from, for example, genetic polymorphism or from human manipulation.

Biologically active variants of a periostin protein of the invention will have at least about 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or more sequence identity to the amino acid sequence for the human periostin protein as determined by sequence alignment programs and parameters described elsewhere herein. A biologically active variant of a protein consistent with an embodiment of the invention may differ from that protein by as few as 1-15 amino acid residues, as few as 1-10, such as 6-10, as few as 5, as few as 4, 3, 2, or even 1 amino acid residue.

Upon injury, adult human hearts respond by producing fibrotic tissue. For example, myocardial infarction typically results in loss of cardiomyocytes and replacement of these cells with scar tissue. Scar tissue that connects tissue surfaces that are normally separated are known as adhesions and can cause significant complications for a patient. Agents that can cause regeneration of cardiomyocytes are therefore in great need as are compositions and methods for delivering such agents effectively, in a sustained fashion, and without causing further damage to the heart.

The pericardial space offers a convenient location for sustained delivery of an agent to the heart because of its proximity to the myocardium. The substantially closed volume of the pericardial sac also offers the benefit of localized containment of an agent and accessibility by minimally invasive techniques (i.e., injection).

Certain existing techniques for delivering agents to the heart in the pericardial space are deficient in that the delivery materials can cause adhesions to form, which can cause complications in the patient. Furthermore, certain controlled delivery devices implanted by open surgical methods may require more recovery time for the patient and can also lead to adhesion formation. Thus, an aim of certain embodiments described herein is to provide a sustained release vehicle that can be implanted using a minimally invasive technique. Another aim is to provide a composition and/or technique that minimizes or prevents adhesions.

The compositions and methods of certain embodiments of the invention may be used to prevent the onset, slow progression, and/or reduce symptoms of cardiac disease caused by, for example, myocardial ischemia, hypoxia, stroke, myocardial infarction, etc. Other examples of cardiac diseases or conditions that can be treated using the inventive compositions and methods are provided below.

FIG. 1 shows a schematic of a heart 100, which includes an aorta 110, a superior vena cava 112, a pulmonary artery 114, a myocardium 120, an epicardium (also known as the visceral pericardium) 130, and a pericardium (also known as the parietal pericardium) 140. Heart 100 also includes a pericardial space 150, a region between the epicardium 130 and the pericardium 140. The pericardial space may contain a pericardial fluid. The pericardium envelopes the heart and a portion of the great vessels (i.e., the aorta, superior vena cava, and pulmonary artery).

In some embodiments, a method of treating the heart of a human or animal subject involves introducing a polymer into or near pericardial space 150 of the subject. The polymer may be introduced into or near the pericardial space of the subject by any suitable method such as by injection or by implantation of a device, as described in more detail below. The delivery technique used may be minimally-invasive. In some cases, the polymer is delivered locally to a site of injury (e.g., an infarction), the location of which may be determined by any suitable method (e.g., by echocardiography or by cardiac MRI).

The polymer may be in any suitable form while or after being introduced into the subject. For example, in one embodiment the polymer is in the form of a liquid or a gel that can be injected into the subject. After injection, the liquid or gel may remain in a liquid or gel form, respectively, or in other embodiments may solidify after being introduced into the subject. In another embodiment, the polymer may be in solid form while being introduced into the subject. The polymer may remain as a solid after being introduced into the subject, or may become a liquid or a gel, e.g., by chemical reaction or physical interaction with one or more components delivered along with the polymer, or by interaction with one or more components already present at the place of injection. As described in more detail below, the polymer may be in the form of a plurality of particles in some embodiments.

In some instances, the polymer (e.g., in particulate or other form) becomes substantially immobilized in or near the pericardial space after injection. For example, the polymer may be held in a localized region between pericardium 140 and myocardium 120 (e.g., in pericardial space 150, at or near the parietal pericardium 140 and/or epicardium 130, between the parietal pericardium and the myocardium, or between the pericardium and a heart vessel (e.g., aorta 110, superior vena cava 112, or pulmonary artery 114)). In certain cases, the polymer forms a gel or a solid mass that localizes at a particular region within the pericardial space, such as those noted above. For instance, the polymer may form a membrane or a film on a surface of the pericardium (e.g., the parietal pericardium or the visceral pericardium). In some cases, the membrane or film forms at a site of injury to be treated by the composition and methods described herein (e.g., at a site of infarction). In other embodiments, the polymer may at least partially distribute within the pericardial space after injection. For example, the polymer may be dispersed or suspended in the pericardial fluid. Advantageously, as described in more detail below, the polymer includes an active agent comprising periostin or fragments or variants thereof that can be delivered to one or more locations within the heart.

Polymers described herein (e.g., in particulate or other form) may have desirable properties, such as the ability to substantially inhibit or reduce tissue adhesion formation. For example, the polymer may be injected into the pericardial space and may

substantially prevent adhesion between a first tissue and a second tissue near or within the pericardial space. The first tissue and the second tissue may be the same (i.e., the first tissue and the second tissue may be different regions of the pericardium) or different (i.e., the first tissue may be the pericardium and the second tissue may be the

myocardium). In certain embodiments, tissue adhesion formation is substantially inhibited or reduced between the parietal pericardium and the visceral pericardium, between the parietal pericardium and the myocardium, or between the pericardium and a heart vessel. It should be understood that tissue adhesion formation can be substantially inhibited or reduced between other tissues or layers within or near the pericardial space. Sometimes, the polymer can substantially prevent or reduce tissue adhesions between a tissue of the heart and a tissue of another organ (e.g., lung tissue). Furthermore, in some cases the polymer can substantially prevent or reduce the amount of scar tissue formed at a tissue site.

It has been discovered within the context of the invention that certain forms of a polymer can reduce or substantially prevent tissue adhesion and/or scar formation compared to other forms of the same polymer, all other factors being equal. For example, it has beenfound that polymers in certain particulate forms reduce tissue adhesion and/or scar formation to a greater extent than the same polymer delivered in a patch form. Formation of tissue adhesion and/or scar formation may be reduced by, for example, at least 10%, at least 20%, at least 40%, at least 60%, at least 80%, or at least 90%, when delivered in a particulate form compared to a non-particulate form (e.g., in the form of a patch), all other factors being equal.

In other embodiments, formation of tissue adhesions and/or scar tissue is reduced by at least 10%, at least 20%, at least 40%, at least 60%, at least 80%, or at least 90%, when the polymer is delivered by injection, compared to when the polymer is delivered by a non-injection method (e.g., by surgical insertion), all other factors being equal. In other embodiments, formation of tissue adhesions and/or scar formation is reduced by at least 10%, at least 20%, at least 40%, at least 60%, at least 80%, or at least 90%, by injecting a polymer into or near the pericardial space of the subject compared to not delivering any polymer into or near the pericardial space.

The amount of tissue adhesions and/or scar tissue formation can be determined by one of ordinary skill in the art by gross inspection and/or by methods such as tissue staining, echocardiography, and cardiac MRI. Accordingly, in one set of embodiments, a method for treating the heart of a human or animal subject includes injecting a polymer into the pericardial space of the subject, and preventing or reducing the formation of adhesion between a first tissue and a second tissue in or near the pericardial space. The polymer, which may be in particulate or other form, may reduce the amount of adhesion between the first and second tissues in or near the pericardial space by at least 10%, at least 20%, at least 40%, at least 60%, at least 80%, or at least 90% compared to either the absence of such delivery, or compared to a different method of delivery (e.g., a non-injection method).

As described herein, in some embodiments, a method of treating the heart of a human or animal subject comprises injecting a plurality of particles. The particles are configured for controlled release of an active agent comprising periostin and may have other desirable properties such as the ability to inhibit tissue adhesion. The particles may be any suitable size. For example, in some cases the particles may have an average particle size greater than 50 nm, greater than 200 nm, greater than 500 nm, greater than 10 microns, greater than 100 microns, greater than 500 microns, greater than 1 mm, greater than 2 mm, etc. In some cases, the particles have an average particle sizebetween 500 microns and 2mm (e.g., between 500 microns and 1 mm, or between 1 mm and 2 mm) or in other cases between about 1mm and 4mm. The particle size may be chosen to elicit certain properties (i.e., release rate of an agent, degradation rate, agent loading capacity, etc.) or accommodate certain methods of administration (i.e., injection), as discussed in more detail below. As used herein, "particle size" refers to the largest characteristic dimension (i.e. of a line passing through the geometric center of the particle e.g., diameter) that can be measured along any orientation of a particle (e.g., a polymer particle). In the case of hydrogel particles, particle size refers to the size of the swelled particle (e.g., in a solution). Particle size as used herein may be measured or estimated, for example, using a sieve analysis, wherein particles are passed through openings of a standard size in a screen. The particle-size distribution may be reported as the weight percentage of particles retained on each of a series of standard sieves of decreasing size, and the percentage of particles passed of the finest size. That is, the average particle size may correspond to the 50% point in the weight distribution of particles.

Furthermore, the particles may have any suitable shape. For example, they may be substantially spherical, pyramidal, cubical, rod-like, or irregularly shaped. After a plurality of particles are introduced into the pericardial space of a subject, the particles may remain in particulate form. For example, the particles may be suspended or dispersed within the pericardial fluid. In some embodiments, the particles aggregate with one another to form a solid or gel-like mass. In other embodiments, the particles dissolve or degrade after being delivered to the subject. In yet other

embodiments, the particles form a film or membrane of material on a tissue surface at or near the site of delivery, or at or near a site of injury. Such a film may, in some cases, extend between two different surfaces at the site of delivery. The film may have any suitable thickness, e.g., between 0.1-5 microns thick, between 5-10 microns thick, between 10-50 microns thick, between 50-100 microns thick, between 100-200 microns thick, between 200-500 microns thick, between 0.5-1 mm thick, or between 1-2 mm thick. The film may be elastic or inelastic, e.g., depending on the polymer used.

A particle or other delivery agent may be contracted of any suitable material. In some embodiments, a particle or delivery agent comprises a polymer. For example, the polymer may be a biodegradable polymer such as a polyester (i.e., polylactic acid, polyglycolic acid, polycaprolactone, etc.), polyanhydride, polycarbonate, copolymers thereof, etc. In some cases, the polymer may form a hydrogel. Examples of polymers capable of forming hydrogels include gelatin (i.e., Gelfoam®, commercially available from Pfizer, Inc.), hyaluronic acid, chitosan, alginate, agarose, polyethylene glycol- polypropylene glycol copolymers, etc. A polymer may be crosslinked, for example through covalent bonds, ionic bonds, hydrophobic bonds, metal binding, etc. A polymer may be obtained from natural sources or be created synthetically. In other

embodiments, the particle is non-biodegradable, or is degradable only after application of energy from an external source (e.g., light or heat).

Those of ordinary skill in the art can chose appropriate materials to control the rates of degradation of the material after it has been delivered to the subject. For instance, the polymer in a particulate or other form may substantially or completely degrade within the subject after or within at least one day, at least three days, at least one week, at least two weeks, at least one month, at least six months, or at least one year. The rate of degradation will depend on the condition to be treated among other factors.

In some embodiments, a polymer may be modified to improve one or more properties. For example, a polymer may be crosslinked or at least partially degraded, or an existing crosslinking density may be increased or descreased. Such changes may be advantageous, for instance, for changing the degradation time of the polymer or the rate of release of an agent from the polymer.

The polymer may be a homopolymer or a copolymer. In certain embodiments, the polymer is a diblock copolymer, a triblock copolymer, etc., e.g., where one block is a hydrophobic polymer and another block is a hydrophilic polymer, or where both blocks are hydrophilic or both block are hydrophobic. For example, the polymer may be a copolymer of an a-hydroxy acid (e.g., lactic acid) and polyethylene glycol. In some cases, a particle includes a hydrophobic polymer, such as polymers that may include certain acrylics, amides and imides, carbonates, dienes, esters, ethers, fluorocarbons, olefins, sytrenes, vinyl acetals, vinyl and vinylidene chlorides, vinyl esters, vinyl ethers and ketones, and vinylpyridine and vinylpyrrolidones polymers. In other cases, a particle includes a hydrophilic polymer, such as polymers including certain acrylics, amines, ethers, styrenes, vinyl acids, and vinyl alcohols. The polymer may be charged or uncharged. As noted herein, the particular components of the particle can be chosen so as to impart certain functionality to the structures.

In some embodiments, the particles may swell upon absorption of fluid. This effect may be used, for example, to load the particles with the active agent comprising periostin or fragments or variants thereof, as discussed in more detail below. As discussed above, in certain embodiments, the particles may be hydrogels. A short description of the properties and behavior of certain hydrogels is provided below. It should be noted that the list is not exhaustive, and those of ordinary skill in the art may readily select or form other suitable absorbent materials using available information regarding the absorbency and swelling properties of various materials and no more than routine experimentation and screening tests.

Polymer gels are typically characterized by long chain polymer molecules that are crosslinked to form a network. This network can trap and hold fluid, which can give gels properties somewhere between those of solids and liquids. Depending on the level of crosslinking, various properties of a particular gel can be tailored. For example, a highly crosslinked gel generally is structurally strong and tends to resist releasing fluid under pressure, but may exhibit slow transition times. A lightly crosslinked gel may be weaker structurally, but may react more quickly during its phase transition. In the design of gels for a particular application, the degree of crosslinking may be adjusted to achieve the desired compromise between speed of absorption and level of structural integrity. Those of ordinary skill in the art would be able to identify methods for modulating the degree of crosslinking in such gels.

Particles may be made by any suitable method. In some embodiments, particles may be made by rasping a larger piece of polymeric material. For example, a Gelfoam® patch may be rasped into particles. In some cases, particles may be made by oitin-water emulsion techniques, crosslinking of polymers, etc. Other methods for fabricating particles will be known to those of ordinary skill in the art.

The compositions and methods disclosed herein may be used to treat a variety of diseases and/or conditions, for example: cardiac arrhythmia, congenital heart diseases, dilated cardiomyopathy, hypertrophic cardiomyopathy, aortic regurgitation, aortic stenosis, mitral regurgitation, mitral stenosis, Ellis-van Cleveld syndrome, familial hypertrophic cardiomyopathy, Holt-Orams syndrome, Marfan syndrome, Ward-Romano syndrome, pericarditis, myocarditis, tumors of the heart (e.g., myxoma, metastasis, etc.), atherosclerosis, hypertension, etc.

In some cases, the compositions and methods described herein can reduce the amount of pericardial effusions (i.e., abnormal amounts of accumulated fluid in the pericardial cavity). For example, a delivery method may include removal of an amount of pericardial fluid prior to delivery of a composition described herein, and then delivery of the same or similar amount of volume of the composition. After delivery, the amount of fluid in the pericardial may be maintained at normal amounts (e.g., about 15 - about 50 mL).

Furthermore, compositions and methods described herein may facilitate healing in a subject, and therefore may be employed during or after after surgery, tissue grafting, organ or tissue transplant, or treatment of heart disease or a cardiovascular condition. The compositions and methods may modify or reduce scar tissue, promote generation of new tissue, preserve the viability of impaired tissues (e.g., ischemic tissue), or prevent or reduce adhesions.

As described herein, a polymer or composition may be configured to release an active agent comprising periostin or fragments or variants thereof. In some

embodiments, the polymer may form a particle with a core-shell configuration, where the shell comprises a polymer and the core may contain, for example, an active agent. In other embodiments, a particle may be substantially uniform throughout.

The polymer or particles injected into the pericardium according to the invention include a therapeutic amount of an active agent comprising periostin or fragments or variants thereof. Periostin, a component of the extracellular matrix, or fragments or variants thereof may be used to promote cardiomyocyte proliferation and myocardial regeneration. Periostin can induce cell cycle re-entry of differentiated mammalian cardiomyocytes. Periostin stimulates mononuclear cardiomyocytes, present in the adult mammalian heart, to undergo the full mitotic cell cycle. After myocardial infarction, recombinant periostin induces cardiomyocyte cell cycle re-entry, improves cardiac remodeling and function, reduces fibrosis and infarct size, and increases angiogenesis. Additional advantages of periostin are descrbed in International Patent Application Serial No. PCT US2008/051659, filed January 22, 2008, entitled "Periostin Induces

Proliferation of Cardiomyocytes and Promotes Cardiac Regeneration," by Kuhn, which is incorporated herein by reference in its entirety for all purposes.

In some embodiments, a polymer may be loaded with the active agent by soaking the polymer in a solution containing the agent. Generally, the loading of agent can be increased by increasing the concentration of the agent in the soaking solution and/or increasing the contact time between the polymer and the soaking solution. In some cases, the polymer is in the form of a particle, and the agent may diffuse into the particle. An agent may also adsorb onto the surface of the particle. The association of an agent with a polymer may result from non-covalent interactions. Alternatively, the agent may be reacted with a polymer to form a covalent bond. As known to those in the art, an agent- polymer covalent bond may be chosen such that under certain conditions (i.e.,

physiological conditions), the bond may break thereby releasing the agent. Depending on the ratio of the active agent to polymer, the nature of the particular polymer employed, the type of association between the active agent and the particle, and the size of the particle, the rate of release of the active agent can be controlled.

The polymers and particles described herein may be used in "pharmaceutical compositions" or "pharmaceutically acceptable" compositions, which comprise a therapeutically effective amount of the active agent associated with one or more of the polymers or particles described herein, formulated together with one or more pharmaceutically acceptable carriers, additives, and/or diluents. The pharmaceutical compositions described herein may be useful for diagnosing, preventing, treating or managing a disease or bodily condition including cardiac and certain vascular conditions.

The pharmaceutical compositions may be specially formulated for administration in gel or liquid form, including those adapted for the following: a sterile solution or suspension, a sustained-release formulation, or as a cream or foam. In some cases, a composition includes a plurality of particles encapsulated in a hydrogel or hydrogel precursor and injected into the pericardial space. The hydrogel or hydrogel precursor may be able to inhibit the formation of tissue adhesions.

The phrase "pharmaceutically acceptable" is employed herein to refer to those structures, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.

The phrase "pharmaceutically-acceptable carrier" as used herein means a pharmaceutically-acceptable material, composition or vehicle, such as a liquid, gel or solid filler, diluent, excipient, or solvent encapsulating material, involved in carrying or transporting the subject compound, e.g., from a device or from one organ, or portion of the body, to another organ, or portion of the body. Each carrier must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not injurious to the patient. Some examples of materials which can serve as

pharmaceutically-acceptable carriers include: sugars, such as lactose, glucose and sucrose; starches, such as corn starch and potato starch; cellulose, and its derivatives, such as sodium carboxymethyl cellulose, ethyl cellulose and cellulose acetate; powdered tragacanth; malt; gelatin; talc; excipients, such as cocoa butter and suppository waxes; oils, such as peanut oil, cottonseed oil, safflower oil, sesame oil, olive oil, corn oil and soybean oil; glycols, such as propylene glycol; polyols, suchas glycerin, sorbitol, mannitol and polyethylene glycol; esters, such as ethyl oleate and ethyl laurate; agar; buffering agents, such as magnesium hydroxide and aluminum hydroxide; alginic acid; pyrogen-free water; isotonic saline; Ringer's solution; ethyl alcohol; pH buffered solutions; polyesters, polycarbonates and/or polyanhydrides; and other non-toxic compatible substances employed in pharmaceutical formulations. Examples of pharmaceutically-acceptable antioxidants include: water soluble antioxidants, such as ascorbic acid, cysteine hydrochloride, sodium bisulfate, sodium metabisulfite, sodium sulfite and the like; oil-soluble antioxidants, such as ascorbyl palmitate, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate, alpha-tocopherol, and the like; and metal chelating agents, such as citric acid, ethylenediamine tetraacetic acid (EDTA), sorbitol, tartaric acid, phosphoric acid, and the like.

The amount of active agent which can be combined with a particle or other carrier material to produce a single dosage form will vary depending upon the host being treated, and the particular mode of administration. The amount of active agent that can be combined with a particle or other carrier material to produce a single dosage form will generally be that amount of the compound which produces a therapeutic effect.

Generally, this amount will range from about 1% to about 99% of active ingredient, from about 5% to about 70%, or from about 10% to about 30%.

Polymers and particles described herein suitable for injection may be

administered in the form of a solution, dispersion, or a suspension in an aqueous or nonaqueous liquid, as an emulsion or microemulsion (e.g., an oil-in-water or water-in-oil liquid emulsion), or as an elixir or syrup, or as pastilles (using an inert base, such as gelatin and glycerin, or sucrose and acacia), each containing a predetermined amount of a particle described herein, and optionally including an active ingredient.

Examples of suitable aqueous and nonaqueous carriers, which may be employed in the pharmaceutical compositions described herein include water, ethanol, polyols (such as glycerol, propylene glycol, polyethylene glycol, and the like), and suitable mixtures thereof, vegetable oils, such as olive oil, and injectable organic esters, such as ethyl oleate. Proper fluidity can be maintained, for example, by the use of coating materials, such as lecithin, by the maintenance of the required particle size in the case of dispersions, and by the use of surfactants.

In addition to the polymers and/or particles, a liquid dosage form may contain inert diluents commonly used in the art, such as, for example, water or other solvents, solubilizing agents and emulsifiers, such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1 ,3-butylene glycol, oils (in particular, cottonseed, groundnut, corn, germ, olive, castor and sesame oils), glycerol, tetrahydrofuryl alcohol, polyethylene glycds and fatty acid esters of sorbitan, and mixtures thereof.

Suspensions, in addition to the polymers and/or particles, may contain suspending agents as, for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bento ite, agar-agar and tragacanth, and mixtures thereof.

These compositions and particles described herein may also contain adjuvants such as preservatives, wetting agents, emulsifying agents, lubricating agents and dispersing agents. Prevention of the action of microorganisms upon the particles may be facilitated by the inclusion of various antibacterial and antifungal agents, for example, paraben, chlorobutanol, phenol sorbic acid, and the like. It may also be desirable to include isotonic agents, such as sugars, sodium chloride, and the like into the

compositions. In addition, prolonged absorption of the injectable pharmaceutical form may be brought about by the inclusion of agents which delay absorption such as aluminum monostearate and gelatin.

Delivery systems suitable for use with polymers, particles and compositions described herein include time-release, delayed release, sustained release, or controlled release delivery systems. Such systems may avoid repeated administrations of the particles and/or active agents in many cases, increasing convenience to the subject and the physician. Many types of release delivery systems are available and known to those of ordinary skill in the art. Specific examples include, but are not limited to, erosional systems in which the composition is contained in a form within a matrix, or diffusional systems in which an active component controls the release rate. The compositions may be as, for example, particles (e.g., microparticles, microspheres), hydrogels, polymeric reservoirs, or combinations thereof. In some embodiments, the system may allow sustained or controlled release of an active agent to occur, for example, through control of the diffusion or erosion/degradation rate of the formulation or particle. The polymers, particles and compositions described herein can also be combined (e.g., contained) with delivery devices such as syringes, catheters, tubes, and implantable devices.

In some embodiments, a pericardial injection (e.g., using a catheter and injection needle) may be used to deliver the polymers, particles and compositions described. A non-limiting schematic of such an approach is shown in FIG. 4. In some cases, it may be desirable to use a guiding instrument to guide insertion of a catheter and/or needle. For example, in some embodiments, a fluoroscope may be used to guide insertion of the catheter and/or needle. In another embodiment, ultrasound may be used. For example, in some cases, intravascular ultrasound (IVUS) may be used. In one embodiment, an IVUS probe may be advanced after the subxiphoid access and positioned next to the pericardial sac at the proposed puncture site. The ultrasound probe may be used to produce a real-time picture so as to inspect the size of the space between the heart and the pericardial membrane in the diastole and systole. This may be advantageous, in some cases, since the distance between the heart and the pericardial membrane can become altered from post-infarction effusion or adhesion. In some cases, use of a guiding instrument may allow more precise positioning of the catheter over the anterior wall of the LV before injection of the gelfoam.

In some instances, the puncture site for the pericardial injection may be closed, for example, to prevent leakage of the injected material. For instance, an injection procedure may be performed and a Starclose SE vascular closure device (Abbott, Abbott Park, IL) may be used to seal the pericardium. In some embodiments, closing the puncture site may result in less than 20% leakage of injected material, less than 10% leakage of injected material, less than 5% leakage of injected material, less than 1% leakage of injected material, or essentially no leakage of injected material, Use of a long- term release implant may be particularly suitable in some cases. "Long-term release," as used herein, means that the implant is constructed and arranged to deliver therapeutic levels of the composition for at least about 30 or about 45 days, for at least about 60 or about 90 days, or even longer in some cases. Long-term release implants are well known to those of ordinary skill in the art. In some embodiments, a long-term release implant can be formed by delivering a plurality of particles to a subject, after which the particles remain within the subject for an extended period.

When the particles described herein are administered as pharmaceuticals, to humans and animals, they can be given per se or as a pharmaceutical composition containing, for example, about 0.1% to about 99.5%, about 0.5% to about 90%, or the like, of particles in combination with a pharmaceutically acceptable carrier.

The particles and compositions described herein may be given in dosages, e.g., at the maximum amount while avoiding or minimizing any potentially detrimental side effects. The particles and compositions can be administered in effective amounts, alone or in a combinations with other compounds.

The phrase "therapeutically effective amount" as used herein means that amount of a material or composition comprising an inventive structure which is effective for producing some desired therapeutic effect in a subject at a reasonable benefit/risk ratio applicable to any medical treatment. Accordingly, a therapeutically effective amount may, for example, prevent, minimize, or reverse disease progression associated with a disease or bodily condition. Disease progression can be monitored by clinical observations, laboratory and imaging investigations apparent to a person skilled in the art. A therapeutically effective amount can be an amount that is effective in a single dose or an amount that is effective as part of a multi-dose therapy, for example an amount that is administered in two or more doses or an amount that is administered chronically.

The effective amount of any one or more particles or the active agent therein described herein may be from about 10 ng/kg of body weight to about 1000 mg/kg of body weight, and the frequency of administration may range from once a day to a once a month basis, to an as-needed basis. However, other dosage amounts and frequencies also may be used as the invention is not limited in this respect. A subject may be

administered one or more particles described herein in an amount effective to treat one or more diseases or bodily conditions described herein.

The effective amounts will depend on factors such as the severity of the condition being treated; individual patient parameters including age, physical condition, size and weight; concurrent treatments; the frequency of treatment; or the mode of administration. These factors are well known to those of ordinary skill in the art and can be addressed with no more than routine experimentation. In some cases, a maximum dose be used, that is, the highest safe dose according to sound medical judgment.

The selected dosage level can also depend upon a variety of factors including the activity of the active agent employed, the route of administration, the time of

administration, the rate of excretion or metabolism of the particular particles or active agents being employed, the duration of the treatment, other drugs, compounds and/or materials used in combination with the particular particle employed, the age, sex, weight, condition, general health and prior medical history of the patient being treated, and like factors well known in the medical arts. A physician or veterinarian having ordinary skill in the art can readily determine and prescribe the effective amount of the pharmaceutical composition required. For example, the physician or veterinarian could start doses of periostin or fragments or variants thereof at levels lower than that required to achieve the desired therapeutic effect and then gradually increasing the dosage until the desired effect is achieved.

In some embodiments, a polymer, particle or pharmaceutical composition described herein is provided to a subject chronically. Chronic treatments include any form of repeated administration for an extended period of time, such as repeated administrations for one or more months, between a month and a year, one or more years, or longer. In many embodiments, a chronic treatment involves administering a particle or pharmaceutical composition repeatedly over the life of the subject. For example, chronic treatments may involve regular administrations, for example one or more times a week, or one or more times a month.

While it is possible for a polymer, particle or pharmaceutical composition described herein described herein to be administered alone, it may be administered as a pharmaceutical composition as described above. The present invention also provides any of the above-mentioned compositions useful for preventing, treating, or managing a disease or bodily condition packaged in kits, optionally including instructions for use of the composition. That is, the kit can include a description of use of the composition for participation in a particular disease or bodily condition, The kits can further include a description of use of the compositions as discussed herein. Instructions also may be provided for administering the composition to the pericardial space by any of the suitable techniques described herein.

The kits described herein may also contain one or more containers, which can contain components such as the polymer, particle or pharmaceutical composition described herein, the active agent as described herein. The kits also may contain instructions for mixing, diluting, and/or administrating the polymer, particle or pharmaceutical composition described herein. The kits also can include other containers with one or more solvents, surfactants, preservatives, and/or diluents (e.g., normal saline (0.9% NaCl), or 5% dextrose) as well as containers for mixing, diluting or administering the polymer, particle or pharmaceutical composition described herein to the patient in need of such treatment. The compositions of the kit may be provided as any suitable form, for example, as liquid solutions or as dried powders. When the composition provided is a dry powder, the powder may be reconstituted by the addition of a suitable solvent, which may also be provided. In embodiments where liquid forms of the composition are used, the liquid form may be concentrated or ready to use. The solvent will depend on the particular particle and the mode of use or administration. Suitable solvents for compositions are well known and are available in the literature.

The kit, in one set of embodiments, may comprise one or more containers such as vials, tubes, syringes, and the like, each of the containers comprising one or more of the elements to be used in the method. For example, one of the containers may contain a solution or suspension of polymer, particle or pharmaceutical composition described herein. Additionally, the kit may include containers for other components, for example, buffers or diluents to be mixed with the polymer, particle or pharmaceutical composition described herein prior to delivery.

As used herein, a "subject" or a "patient" refers to any mammal (e.g., a human), for example, a mammal that may be susceptible to a disease or bodily condition.

Examples of subjects or patients include a human, a non-human primate, a cow, a horse, a pig, a sheep, a goat, a dog, a cat or a rodent such as a mouse, a rat, a hamster, or a guinea pig. Generally, the invention is directed toward use with humans. A subject may be a subject diagnosed with a certain disease or bodily condition or otherwise known to have a disease or bodily condition. In some embodiments, a subject may be diagnosed as, or known to be, at risk of developing a disease or bodily condition.

These above descriptions of applications for the inventive compositions and methods devices are not intended to be exhaustive, and merely illustrate some of the possible embodiments and uses of this invention.

The function and advantage of these and other embodiments of the present invention may be more fully understood from the examples below. The following examples, while illustrative of certain embodiments of the invention, do not exemplify the full scope of the invention.

EXAMPLES

Example 1 This example demonstrates development of a composition for pericardial injection. To achieve meaningful infarct regression by inducing cardiomyocyte proliferation, the constant presence of a mitogenic stimulus was required. A controlled delivery system was designed to apply recombinant periostin, an extracellular matrix molecule, to the cardiac extracellular matrix. It was discovered that periostin binds non- covalently to Gelfoam^®, a preparation of collagen, and is gradually released, indicating that Gelfoam^®can provide a suitable delivery vehicle. The solid Gelfoam was homogenized by rasping into small particles to make it injectable (FIG. 2A).

It was determined how fast homogenized Gelfoam would be degraded in pig pericardial fluid and found that Gelfoam^®particles persisted for 9 days at 38 °C in vitro versus greater than 60 days at 38 °C in saline (FIG. 2B). The Gelfoam^® particles in pericardial fluid or saline were slowly agitated in an Eppendorf tube on a shaker, and degradation of the Gelfoam^®particles was scored daily by visual inspection.

The homogenized Gelfoam^®once homogenized by rasping into small particles could now be injected through a 5F introducer. A minimally invasive approach was then developed to insert the introducer through the pericardium into the pericardial space (FIG. 2C). The so injected Gelfoam^® material covered the left ventricular free wall containing the myocardial infarct scar and after closure of the puncture site was completely retained within the pericardium (FIG. 2D). Echocardiography was performed immediately after, one week after, and four weeks after injection and did not find pericardial effusions, indicating that the periostin delivery system is biocompatible. Importantly, at one week (FIG. 2E), the Gelfoam^® had formed an elastic membrane overlying the area of the infarction. No Gelfoam^® was visible on the inferior wall. One month after application, the Gelfoam^® was completely degraded, indicating good biodegradation. The pericardial sheet could be easily peeled off the epicardial surface 3 months after administration, thus demonstrating the absence of major pericardial adhesions.

Example 2

This example demonstrates the effect of periostin delivered using the composition of Example 1. To determine whether periostin peptide has a beneficial effect after myocardial injury in a preclinical large animal model, a pig animal model was used. Permanent occlusion of the left anterior descending coronary artery (LAD) was chosen as an injury model. Periostin peptide was applied two days (2 d) after infarct generation (MI) (see Example 4), and the effect on cardiac function and structure one and three months later was determined (FIG. 3 A). Cardiac MRI showed both groups to have similar ejection fractions at the day of application of the periostin (FIG. 3B). However after 3 months, the ejection fraction in periostin treated animals was significantly increased compared to control. Peak LV pressure rate of rise (dP/dt)max and the cardiac output normalized to weight as cardiac index (CI) were comparable in both groups 48 hours after creation of the infarct, but differed significantly at the 1 month follow-up with periostin treated animals recovering in their cardiac function and untreated animals further declining (FIGs. 3C-3D). Two months later at the time of sacrifice, the gain in cardiac function with the application of periostin was sustained, again being significantly higher than in untreated control animals. Blood pressures as well as the diastolic function of the heart expressed with the parameter Tau did not differ between groups over the course of the study. The improvements in systolic function in the treatment group were supported by a significantly increased peak ejection rate (PER), while the peak filling rate (PFR), a marker for diastolic function, did not differ between groups.

Example 3

This example demonstrates the effect of pericardial periostin administration on scar size in animals treated as described in Example 2. Cardiac MRI imaging, using delayed enhancement of scar areas after injection of gadolinium, was performed to analyze scar size in all animals at time of periostin application and sacrifice (FIG. 3E). A correlation of r=.875 (PO.001) was observed between the in-vivo imaging method and the scar size analysis performed with tissue staining after sacrifice. The quantitative analysis confirmed that animals of both treatment arms had a similar extent of infracted area upon administration of periostin. While untreated animals had similar or larger scar areas 3 months later, periostin-treated animals showed a significant decrease in the area marked by delayed enhancement as scar. The extent of necrotic tissue was confirmed after sacrifice by staining 5 slices of the left ventricle with TTC, as shown in a representative panel (FIG. 3F) (Scale bars, 5mm). Quantification (n = 13) revealed that the percentage of viable myocardium in the periostin group was significantly higher than in untreated swine (FIG. 3H). Moreover, in a macroscopic comparison of the treatment groups, it was found that periostin-treated animals had islands or strips of viable myocardial tissue incorporated in the immediate scar area, while none of the untreated animals showed this phenomenon (FIG. 3G). In summary, minimally invasive application of cardiac regeneration factors is biologically effective and safe. Periostin peptide induced sustained improvement of cardiac function after myocardial infarction.

Example 4

This example provides additional description of the materials and methods employed for Examples 1-3 above.

Animal Studies. All procedures were approved by the Institutional Animal Care Committee. Non-surgical procedures were performed under anesthesia with propofol (8- 15 mg/h) and surgery under isoflurane anesthesia (0.8-1.2 % in 100% oxygen). In the experiments, 13 female Yorkshire pigs (weight approximately 20 kg) underwent percutaneous transluminal coil embolization of the left anterior descending coronary artery (LAD). The animals were randomized to receive either control Gelfoam^®

(Surgifoam, Johnson & Johnson, USA) with the buffer used to dissolve periostin or Gelfoam^® with periostin. Structural and functional assessment at baseline, 2 days after MI generation, at 1 month and after 3 months was performed.

Experimental myocardial infarction. An 8F sheet was introduced into the femoral artery and cannulated the LAD with an 8F hockey stick guiding catheter (Cordis Infiniti, Johnson & Johnson, USA), 100 micrograms nitroglycerin were injected, and baseline coronary angiography was performed. A platinum embolic coil (0.035 in, 40 mm length, 5><3-mm diameter, Cook Medical Inc, Bloomington, IN) was placed with a 4F AR catheter (Cordis Infiniti, Johnson & Johnson, USA) into the LAD after the takeoff of the first diagonal branch. This completely occluded 2/3 of the LAD tributary, determined by coronary angiography, resulting in infarction of approximately 18% of the left ventricle, determined by TTC staining. The 48 hour rate of survival after infract creation was 75%.

Controlled release system and delivery strategy. Gelfoam^® sheets were homogenized with rasping to prepare an injectable Gelfoam^® slurry. 0.1 mg of dissolved periostin (treatment group) were added per mL of Gelfoam or the same volume of buffer (control group). A 4 cm lateral thoracotomy was made to puncture the pericardial sac with a 5F introducer. Approximately 7 mL of pericardial fluid was aspirated, and 7 mL of Gelfoam^® slurry containing Periostin (0.7 mg) or control buffer was injected. The sheet was retracted and the puncture hole was closed with a purse string suture.

Assessment of myocardial function and structure. To determine myocardial repair, myocardial function and structure was assessed at baseline (i.e., before MI generation), 48 hours after myocardial infarction (i.e., before application of the delivery system), 1 month, and 3 months after application of the delivery system.

Echocardiography. Images were acquired with an iE33 ultrasound machine (Philips Medical Systems) equipped with an X3-1 and S8-3 transducer during end- expiratory breath-hold in an R-wave-trigged mode.

Cardiac MRI. Contiguous short-axis cine images covering the LV from base to apex were acquired with a 1.5 T magnet (Magneton Sonata, Siemens Medical Solutions) using a phased-array cardiac coil with ECG gating during end-expiratory breath-hold. Imaging of delayed enhancement (DE) was performed 15 minutes after the

administration of 0.2 mmol/kg gadopentate dimeglumine (Magnevist, Bayer Medical Solutions, Germany) in an inversion-recovery fast gradient-echo sequence. All images were acquired and analyzed by an investigator blinded to the study arm. LV function analysis was performed with Argus software (Argus, Siemens Medical Solutions). DE was quantified with prototype analysis software (QMass v7, medis, Leiden, Netherlands).

Catheterization. The femoral artery and vein were accessed with 7F sheets. A 6F Millar Micro-Tip catheter system (Millar Instruments Inc.) was placed into the aorta, the left ventricle, and the right ventricle. The following parameters were collected and analyzed: systolic pressure, enddiastolic pressure, peak LV pressure rate of rise

(dP/dt)max and Tau value (time constant of isovolumic relaxation). (dP/dt)max/P was calculated as (dP/dt)max/(systolic - end-diastolic pressure). The mean of at least 3 consecutive cardiac cycles was calculated for each measurement. Coronary angiography and left ventriculography (LVG) were performed at day 2, after one, and three months using a Integris H5000 single-plane fluoroscopy system (Philips Medical Systems) to determine the LV end-diastolic volume (EDV), end-systolic volume (ESV), and ejection fraction (EF).

Assessment of periostin effects at the tissue level. Using a pathology knife (Tissue Tek, Sakura Finetek, USA), the left ventricle was cut into 6 slabs of the same thickness. Viable myocardium was visualized by staining 5 of these slabs with 2,3,5- triphenyltetrazolium chloride and digitally quantified the scar area.

Statistical analyses. Four Investigators quantified observations independently from one another and in a blinded manner. Numeric data are presented as mean ± s.e.m. Statistical significance was tested using SPSS (ver 16) software with one and two-sided ANOVA, where appropriate. Sigmoidal nonlinear or linear regression was used to fit data (GraphPad). The avalue was set at 0.05.

Example 5

This example demonstrates a pericardial injection technique. For large animal in vivo studies, a flouroscopic-guided approach to the pericardial sac was developed (FIG. 4). As depicted in FIG. 4, a needle 410 was advanced under the sternum 420 towards the pericardium 430. The procedure allowed precise positioning of the catheter over the anterior wall of the LV before injection of the Gelfoam^® (FIG. 5A).

The presence of the Gelfoam^®in the pericardium was confirmed by mixing the gelfoam with 50% saline and 50% contrast dye before injection. Fluoroscopic pictures were acquired every 10 minutes for up to 90 minutes to assess the amount of leakage after removal of the catheter (FIG. 5B). Leakage occurred to a large extent when only liquid contrast dye was injected. This effect is presumably enhanced by gravity in combination with the higher density of the liquid dye. In contrast, almost no

Gelfoam^®was visible in the chest cavity, even when the puncture site was not closed. In order to improve this approach further, these procedures were performed using a

Starclose SE vascular closure device (Abbott, Abbott Park, IL) to seal the pericardium. This strategy resulted in elimination of any visible leakage, even after injection of pure liquid contrast_.dye.

The distribution of the Gelfoam^®in relation to the infarct zone is depicted in FIG. 5C. To increase safety of the percutaneous puncture, an IVUS-guided approach was further established. The IVUS probe was advanced after the subxiphoid access and positioned next to the pericardial sac at the proposed puncture site (FIG. 5C). The ultrasound probe at the tip of the catheter produces a real-time picture to inspect the size of the space between the heart and the pericardial membrane in diastole and systole (FIG. 6A). The distance between the heart and the pericardial membrane can become altered from post-infarction effusion (upper panel) or adhesion (lower panel). With this instrument, a decrease in pericardial space due to severe pericardial effusion was visualized (FIG. 6B) with removal of two times 20 mL of fluid. Statistical analysis showed significant differences in the setting of severe adhesion and pericardial effusion versus the pericardial space of a naive animal in systole (baseline: 2.1±0.2 mm vs.

adhesion: 0.9±0.08 mm vs. effusion: 3.98±1.1 mm; P < 0.05) (FIG. 6C).

Example 6

This example provides additional description of the materials and methods employed for Example 5 above.

Pericardial injection of Gelfoam^®particles. In order to make this approach suitable for preclinical testing, a percutaneous method of accessing the pericardial space and injecting the liquified Gelfoam^®was developed. Injections were performed 48 hours after myocardial infarction. Under fluoroscopic guidance a 18G puncture needle (Cook Medical) was advanced under the sternum towards the pericardium. After confirming successful puncture with a small bolus injection of contrast dye a wire was placed in the pericardial space. After puncture clear pericardial fluid could be aspirated. Only in a few cases the fluid contained small amounts of blood, which had no implications for the safety of the procedure. A 8F vascular sheath was then placed over the wire and thereafter a 5 French Amplatz Right catheter was inserted and positioned over the anterior wall of the LV.

Intravascular ultrasound (IVUS). Measurement of the distance between the pericardial sac and the heart was performed with a Galaxy Intravascular Ultrasound catheter (Boston Scientific, Ample Groove, MN, USA). After subxyphoid puncture, the ultrasound probe was advanced over the 8F sheath towards the pericardial sac. The procedure was performed under fluoroscopic guidance and during breath-hold.

While several embodiments of the invention have been described and illustrated herein, those of ordinary skill in the art will readily envision a variety of other means and structures for performing the functions and/or obtaining the results or advantages described herein, and each of such variations, modifications and improvements is deemed to be within the scope of the present invention. More generally, those skilled in the art would readily appreciate that all parameters, dimensions, materials, and configurations described herein are meant to be exemplary and that actual parameters, dimensions, materials, and configurations will depend upon specific applications for which the teachings of the present invention are used. Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments of the invention described herein. It is, therefore, to be understood that the foregoing embodiments are presented by way of example only and that, within the scope of the appended claims and equivalents thereto, the invention may be practiced otherwise than as specifically described. The present invention is directed to each individual feature, system, material and/or method described herein. In addition, any combination of two or more such features, systems, materials and/or methods, provided that such features, systems, materials and/or methods are not mutually inconsistent, is included within the scope of the present invention.

In the claims (as well as in the specification above), all transitional phrases or phrases of inclusion, such as "comprising," "including," "carrying," "having,"

"containing," "composed of," "made of," "formed of," "involving" and the like shall be interpreted to be open-ended, i.e., to mean "including but not limited to" and, therefore, encompassing the items listed thereafter and equivalents thereof as well as additional items. Only the transitional phrases or phrases of inclusion "consisting of and

"consisting essentially of are to be interpreted as closed or semi-closed phrases, respectively. The indefinite articles "a" and "an," as used herein in the specification and in the claims, unless clearly indicated to the contrary, should be understood to mean "at least one."

All references cited herein, including patents and published applications, are incorporated herein by reference. In cases where the present specification and a document incorporated by reference and/or referred to herein include conflicting disclosure, and/or inconsistent use of terminology, and/or the incorporated/referenced documents use or define terms differently than they are used or defined in the present specification, the present specification shall control. What is claimed is:

Claims

1. A method for treating the heart of a human or animal subject, comprising:

injecting a polymer containing an active agent comprising periostinor fragments or variants thereof into the pericardial space of the subject, wherein the polymer prevents adhesion from forming between a first tissue and a second tissue in the pericardial space.

2. The method of claim 1 , wherein the polymer containing an active agent comprises periostin.

3. The method of claim 1 or 2, wherein the polymer is configured to act as a sustained release vehicle for the periostin.

4. The method of claim 1 or 2, wherein the polymer is biodegradable.

5. The method of claim 1 or 2, wherein the polymer forms a hydrogel.

6. The method of claim 1 or 2, wherein the polymer comprises collagen.

7. The method of claim 1 or 2, wherein the polymer comprises gelatin.

8. The method of claim 1 or 2, wherein the polymer becomes substantially immobilized in the pericardial space after injection.

9. The method of claim 1 or 2, wherein the first tissue and the second tissue are substantially the same type of tissue.

10. The method of claim 1 or 2, wherein the first tissue and the second tissue are different types of tissue.

11. The method of claim 1 or 2, wherein the polymer is injected in particulate form.

12. A method for treating the heart of a human or animal subject, comprising: injecting a plurality of particles comprising gelatin into the pericardial space of the subject, wherein the particles further comprise periostin or fragments or variants thereof and are configured to act as a sustained release vehicle for the periostin or fragments or variants thereof.

13. The method of claim 12, wherein the particles further comprise periostin.

14. The method of claim 12 or 13, wherein the particles are capable of preventing adhesion from forming between a first tissue and a second tissue in the pericardial space.

15. The method of claim 14, wherein the first tissue and the second tissue are substantially the same type of tissue.

16. The method of claim 14, wherein the first tissue and the second tissue are different types of tissue.

17. The method of claim 12 or 13, wherein the particles become substantially immobilized in the pericardial space after injection.

18. The method of claim 12 or 13, wherein the particles have an average particle size greater than 500 microns.

19. The method of claim 18, wherein the particles have an average particle sizegreater than 1 millimeter.

20. The method of claim 18, wherein the particles have an average particle size between 500 microns and 2 millimeters.

21. The method of claim 20, wherein the particles have an average particle size between 500 microns and 1 millimeter.

22. The method of claim 20, wherein the particles have an average particle size between 1 millimeter and 2 millimeters.

23. A method for treating the heart of a human or animal subject, comprising:

injecting a plurality of particles having an average particle size greater than 500 microns into the pericardial space of the subject, wherein the particles further comprise periostin or fragments or variants thereof and are configured to act as a sustained release vehicle for the periostin or fragments or variants thereof.

24. The method of claim 23, wherein the particles further comprise periostin.

25. The method of claim 23 or 24, wherein the particles have an average particle size greater than 1 millimeter.

26. The method of claim 25, wherein the particles have an average particle size between 500 microns and 2 millimeters.

27. The method of claim 26, wherein the particles have an average particle size between 500 microns and 1 millimeter.

28. The method of claim 26, wherein the particles have an average particle size between 1 millimeter and 2 millimeters.

29. The method of claim 23 or 24, wherein the particles are capable of preventing adhesions from forming between a first tissue and a second tissue in the pericardial space.

30. The method of claim 29, wherein the first tissue and the second tissue are substantially the same type of tissue.

31. The method of claim 29, wherein the first tissue and the second tissue are different types of tissue.

32. The method of claim 23 or 24, wherein the particles become substantially immobilized in the pericardial space after injection.

33. The method of claim 23 or 24, wherein the particles comprise gelatin.