JP2009530412A - Methods and methods for treating damaged heart tissue - Google Patents

Abstract

  Disclosed are methods and methods for treating heart tissue by delivering a platelet composition that promotes neovascularization in heart tissue followed by a cell formulation that promotes regeneration in the revascularized tissue. The platelet composition can additionally contain structural materials and / or bioactive agents.

Description

  This application is a related patent of U.S. Patent Applications No. 11 / 426,211 and No. 11 / 426,219 filed on June 23, 2006, which are hereby incorporated by reference in their entirety. In accordance with United States Code, Volume 35, §119 (e), replaces US Provisional Patent Application No. 60 / 693,749 filed June 23, 2005, and No. 60 / 743,686 filed March 23, 2006. It is what I insist.

  The present invention relates generally to schemes and schemes for the treatment of damaged heart tissue. In particular, the present invention discloses compositions, methods, and methods for regeneration in damaged tissue.

  The human heart wall, called the endocardium, overlaps the myocardium or myocardium with a constant thickness and is composed of the inner layer of a single-layer squamous epithelium, enclosed in a multi-layered tissue structure called the pericardium. ing. The innermost layer of the pericardium, called the visceral pericardium or epicardium, covers the myocardium. The epicardium is the origin of an aortic arch that forms an outer tissue layer called the wall-side pericardium that is spaced from a sac extending around the ventricle and the visceral pericardium of the atria. Flip outside at the beginning. The outermost layer of the pericardium, called the fibrous pericardium, attaches the parietal pericardium to the sternum, large vessels, and diaphragm so that the heart is trapped within the mediastinum. Originally, the visceral pericardium and the wall pericardium are in close proximity to each other and separated by a thin layer of serous pericardial fluid that allows the heart to move without friction within the sac. The gap between the visceral and parietal pericardium is called the pericardial space. In common terminology, the visceral pericardium is usually referred to as the epicardium, hereinafter the epicardium is used. Similarly, the sidewall pericardium is usually referred to as the pericardium, and the pericardium is used to refer to the wall-side pericardium.

  Heart disease, including myocardial infarction (MI), is a leading cause of death and disability, especially in Western countries, especially in men. Various heart diseases progress to heart failure by a common mechanism called remodeling. Remodeling often leads to clinical heart failure and concomitant symptoms, which gradually worsens heart function. Heart disease can then harm other physiological systems. Every year, more than 1.1 million Americans have myocardial infarction (MI). Myocardial infarction results in an acute decline in ventricular function and expansion of infarcted tissue under stress. This causes a gradual chain of myocyte events known as remodeling. In many cases, this progressive myocardial infarction expansion and remodeling leads to worsening ventricular function and heart failure. Such ischemic cardiomyopathy is a leading cause of heart failure in the United States. It is an object of the present invention to improve vascular supply to patients at high risk for developing heart disease (cardiac ischemia). Increased blood supply will be helpful to acutely or chronically diseased heart tissue. Studies have shown that even in adults, normal repair mechanisms (eg, recruitment of endogenous neoplastic cells) can be derived following heart failure. Inadequate blood supply limits the survival of such cells and prevents healing. Blood supply provides the necessary oxygen and nutrients for the injured area, blood components (cells, inflammatory cell migration factors, etc.), and it is necessary to clear out metabolites. Treatments that improve blood supply to such areas are likely to help patients by promoting greater recovery.

  Heart tissue can be acute or chronic ischemic. Severe ischemia resulting from cardiac cell death is called infarction. Increased vascular supply to or around the lesion area can improve acute or chronic repair.

  Stenotic or occluded coronary arteries are one example of heart disease. Coronary arteries that are completely or fairly occluded can cause short-, medium-, and long-term adverse effects. In the short term, when the coronary artery is occluded, a myocardial infarction occurs, and the resulting myocardial cell death cannot supply blood to the myocardial tissue. When a myocardial infarction occurs, myocardial tissue that is no longer receiving adequate blood flow will die and eventually scar tissue will be replaced.

  Myocardial cells that are underperfused within a few seconds of myocardial infarction no longer undergo contractions that cause abnormal wall motion, stress in the infarct and surrounding high walls, and reduced ventricular function. High stress at the junction between infarcted tissue and normal tissue will, over time, cause an expansion to the infarcted area of the heart and remodeling. These high stresses harm the still viable cardiomyocytes and ultimately reduce their function. This results in enlargement of the tissue including the original myocardial infarct region and causing further damage and dysfunction.

  According to the American Heart Association, approximately 1,100,000 new myocardial infarctions occurred in the United States in 2000. 650,000 patients had the first myocardial infarction, while the other 450,000 patients had recurrent events. Before reaching the hospital, 220,000 people with MI have died. Within one year of myocardial infarction, 25% of men and 38% of women die. Within 6 years, 22% of men and 46% of women develop heart failure, 67% of whom are disabled. Despite the latest medical therapy, the above results are shown.

  The consequences of myocardial infarction are often severe and interfere with daily life. When a myocardial infarction occurs, myocardial tissue that is no longer receiving adequate blood flow dies and scar tissue is replaced. Since this infarcted tissue cannot contract during the systole, it actually lengthens the systole and causes a rapid decline in ventricular function. This abnormal movement of the infarcted tissue can lead to delayed electrical activity or abnormal conduction in the peri-infarct tissue (the tissue at the junction between the normal tissue and the infarcted tissue) that is still alive. Even give extra structural stress.

  The area receiving reduced blood flow is known as the ischemic area. In addition, elevated matrix metalloproteinases, decreased matrix metalloproteinases in tissue inhibitors (collagenase inhibition), and the resulting degradation of collagen can play additional roles in ischemic cardiomyopathy. In order to improve cardiac function in patients with ischemic cardiomyopathy, it is necessary to re-establish blood flow in the ischemic region and / or to the increased contractility of the damaged tissue.

  In addition to immediate hemodynamic effects, it performs cardiac infarct tissue and three main processes: infarct enlargement, infarct extension, and chamber remodeling. Individually and in combination, these factors contribute to the eventual dysfunction found in heart tissue away from the infarct site.

  Infarct enlargement is a permanent, unsuitable area unthinning and tissue expansion fixed within the infarct area. After myocardial infarction, infarct enlargement occurs early. The mechanism is the tissue page slippage.

  Infarct enlargement is additional myocardial necrosis following myocardial infarction. Infarct expansion results in an increase in the total mass of the infarct tissue, and additional infarct tissue can also undergo infarct expansion. Infarct expansion occurs days after myocardial infarction. The mechanism of infarct expansion appears to be an imbalance in blood supply to the peri-infarct tissue against the increased oxygen demand of the tissue.

  Remodeling is a progressive hypertrophy of the ventricle usually associated with a decrease in ventricular function. Myocyte function in heart tissue away from the original myocardial infarction is reduced. Remodeling occurs from weeks to years after myocardial infarction. Such remodeling usually occurs on the left side of the heart. If remodeling occurs on the right side of the heart, it can generally be related to remodeling on the left side of the heart (or some other bad event). Less frequently than the left side, but remodeling can occur alone in the right heart. There are many potential mechanisms for remodeling, and it is generally believed that high stress on peri-infarct tissue plays an important role. Due to a variety of factors such as different shapes and sizes, the stress on the wall in the heart tissue surrounding infarction is much higher than normal.

  The changing processes associated with infarct expansion and remodeling are believed to be the result of high stress exerted on the junction between the infarct tissue and normal heart tissue (ie, around the infarct region). In the absence of interventional treatment, these high stresses eventually kill or severely reduce the function of neighboring cells. As a result, the area around the infarct region will develop over time from the initial infarct site over time. As a result, tissues that have become dysfunctional from the original myocardial infarction region can continue to exacerbate the nature of the disease and often progress to a much advanced stage of heart failure.

  There are various treatments for myocardial infarction used now and in the past. Immediately following myocardial infarction, preventing and treating ventricular fibrillation and stabilizing hemodynamics are well-established therapies.

  Ischemic heart disease can occur acutely or chronically. Mild disease will result in inadequate blood supply during increased demand (eg, during intense activity). Serious illness results in an inadequate blood supply even at rest. Both symptoms will benefit from an increased blood supply, as it is expected to have an obvious clinical clinical sequelae. This may include any or all of increased effort performance, reduced symptoms, increased organ blood perfusion, improved cardiac output per minute, and / or improved cardiac contractility.

  Newer approaches include more aggressive attempts to reopen occluded blood vessels. This has been achieved through thrombolytic therapy or angioplasty and stents. Resuming an occluded artery (ie, vascular regeneration) within hours of the original occlusion reduces tissue death, thereby reducing the total size of infarct expansion and extension, and thereby a stimulant for remodeling Can be limited.

  The interest in re-establishing blood flow in an ischemic area that reduces symptoms and / or improves cardiac function remains unchanged. Blood flow re-establishment can be accomplished through angiogenic stimulation that the body provides or expands blood supply to a specific area. Conventional methods for re-establishing blood flow and returning the heart to the original state often required invasive surgery such as bypass surgery or angioplasty. Other methods used a laser to drill through the infarct and ischemic areas to promote blood flow. These surgeries are complex and dangerous. Therefore, there is a need for safer and less invasive methods for reestablishing blood flow.

  Delivery of agents directly or selectively to heart tissue is desirable over global delivery for a number of reasons. One reason is the small amount of drugs that are substantial and available, eg, agents used in gene therapy. Another reason is a substantially greater concentration of the agent that can be delivered directly to the heart tissue compared to the diluted concentration that would be obtained through systemic delivery. Yet another reason is related to the systemic toxicity of systemic administration at the doses required to obtain the desired drug concentration in the heart tissue.

  One way to deliver drugs to the heart tissue is direct epicardial injection into the heart tissue during an open chest surgery. Another approach taken for drug delivery to heart tissue is intravascular access. A catheter may be advanced through the vasculature and into the heart to inject material from the heart into the heart tissue. Another approach is the endocardial approach, which delivers material from the heart chamber to the heart wall. In addition, additional therapies that have been developed for the treatment of damaged heart tissue include injection of cells and / or other biological agents into ischemic heart tissue, or cells and / or into ischemic tissue. Alternatively, it includes the placement of an agonist. One therapy for treating infarcted heart tissue involves the delivery of cells that can mature and actively contract cardiomyocytes, or regenerate the heart tissue. Examples of such cells include myocytes, myoblasts, mesenchymal stem cells and pluripotent cells. Such cell delivery into heart tissue is beneficial and is believed to prevent or treat heart failure in particular. However, cell therapy of myocardial tissue to date has not reached its full potential. At least one reason is that the transplanted cells survive with poor vascularity and are insufficient to regenerate damaged tissue in the area. Such a therapeutic strategy would greatly assist in the re-establishment of the vascularized tissue layer in which the transplanted cells are placed in or near.

After acute or chronic heart damage, attempts have been made to try to restore some or all of the functions of the endogenous neoplastic cells to the damaged tissue. Reduced blood flow and vascular supply to the injured area appears to inhibit this recovery mechanism. Proper perfusion delivery may promote earlier, faster, and / or more complete repair.
Establishment of an improved or normal blood supply in the damaged tissue can prevent further deterioration and promote regeneration of the damaged tissue. Such therapy is advantageous over previously used treatments. For these reasons, it would be desirable to have an agent that can be delivered directly to the heart tissue to promote the survival and neogenesis of the damaged tissue and to cause neovascularization.

  The present invention provides methods and compositions for promoting neovascularization and treating cardiac tissue by administration of a platelet composition and cell therapy. Induction of neovascularization in damaged heart tissue prior to cell preparation in vivo transplantation increases the survival, coalescence, and maintenance of cells transplanted into the damaged tissue.

  In one embodiment of the present invention, the composition comprises a platelet composition to a treatment site in heart tissue that promotes neovascularization of heart tissue, comprising injecting a cell preparation into regenerated heart tissue Provide treatment. In another embodiment, the heart tissue is damaged tissue or healthy tissue in a damaged heart. In another embodiment, the method results in the regeneration of the aforementioned heart tissue. In another example, the target heart tissue is healthy tissue in an undamaged heart.

  In another embodiment of the invention, the platelet composition is selected from the group consisting of platelet gel, platelet rich plasma and platelet poor plasma. In another example, the platelet composition is autologous. In another embodiment, the platelet gel is formed from platelet poor plasma or platelet rich plasma and an activator. In another example, the activator is thrombin. In another embodiment, the thrombin is selected from the group consisting of recombinant thrombin, human thrombin, animal thrombin, modified thrombin and autothrombin. In another embodiment, the platelet gel comprises platelet rich plasma or platelet poor plasma and thrombin in a ratio between about 5: 1 and about 25: 1. In another example, the ratio of platelet rich plasma or platelet poor plasma to thrombin is about 10: 1.

  In another embodiment of the invention, the platelet composition comprises platelet rich plasma.

In another embodiment of the invention, the platelet composition is delivered to the treatment site and forms a solid or gel in the aforementioned heart tissue at the treatment site.
In another embodiment of the invention, the bloodplate composition further comprises a collagenous, biocompatible polymer, alginate, synthetic / natural compound, fibrinogen, silk elastin polymer, hydrogel and dental complex material. A structural material selected from the group consisting of: In another embodiment, the structural material becomes solid or chemically as a result of activation selected from the group consisting of physical or chemical cross-linking or activation of the aforementioned composition from enzyme, chemical, thermal, or photoactivation. Form a gel.

  In another embodiment of the invention, the platelet composition further comprises a bioactive agent. In another embodiment, the bioactive agent is a pharmaceutically effective compound, corticosteroid, growth factor, enzyme, DNA, ribonucleic acid, siRNA, virus, protein, lipid, polymer, hyaluronic acid, antibody, antibiotic, Selected from the group consisting of anti-inflammatory agents, antisense nucleotides and transforming nucleic acids, and combinations thereof.

  In another embodiment of the invention, the platelet composition further comprises a contrast agent.

  In another embodiment of the invention, the cell preparation comprises cells of one or more cell types selected from the group consisting of somatic, germ cells, fetuses, embryonic, postnatal cells and mature cells. In another embodiment, the cell preparation comprises cells isolated from one or more tissue types selected from the group consisting of fat, brain, muscle, endothelium, blood, bone marrow, heart, testis and ovary. In another example, the cell is autologous.

  In another embodiment of the invention, the cells are modified prior to transplantation. In another example, the modification includes cellular genetic manipulation to secrete one or more biologically active molecules. In another example, the biologically active molecule is a growth factor. In another embodiment, the cell preparation further comprises a bioactive agent. In another example, the bioactive agent is a growth factor. In another embodiment, the cell preparation further comprises a platelet composition.

  In another embodiment of the invention, the platelet composition is supplied to the damaged heart tissue between about 1 hour and 1 year after the injury has occurred in the heart tissue. In another example, either the platelet composition or the cell preparation is provided by injection at approximately 1 to 20 sites. In another embodiment, the injections are performed sequentially. In another embodiment, the injections are made almost simultaneously. In another embodiment, the infusion includes a total infusion volume of up to about 15 ml. In another embodiment of the invention, the injection includes a volume of up to 1100 microliters per injection.

  In another embodiment of the invention, the platelet composition or the cell preparation described above is injected into the heart tissue at a right angle or at an angle to the tissue surface.

  In another embodiment of the invention, the cell preparation is delivered to the damaged heart tissue between about 1 hour and 1 year after the injury has occurred in the heart tissue. In another example, the cell preparation is provided to the heart tissue between about 1 hour and 1 year after administration of the platelet composition. In another example, the cell preparation is provided to the heart tissue after neovascularization is initiated in the damaged heart tissue. In another example, the platelet composition and cell preparation are provided to the heart tissue almost simultaneously.

  In another embodiment of the invention, the treatment site for cardiac tissue is selected from the group consisting of subendocardial, epicardial and intramyocardial sites. In another embodiment of the invention, the platelet composition or cell preparation is injected into the heart tissue at a depth in the middle of the heart muscle thickness.

  In another embodiment of the invention, the method further comprises a delivery device adapted to deliver the platelet composition or cell preparation to the injured heart tissue. In another embodiment, the delivery device is an infusion catheter selected from the group consisting of an endocardial infusion catheter, a transvascular infusion catheter, and an epicardial infusion catheter.

  In another embodiment of the present invention, the treatment site is selected from the group consisting of an injured area, an injured area, and healthy tissue surrounding the injured area. In another example, the platelet composition and cell preparation are injected into the same treatment site. In another example, the platelet composition and cell preparation are injected at different treatment sites. In another example, the cell preparation is injected adjacent to the injection site of the platelet gel composition.

  In one embodiment of the invention, the method provides a platelet composition within a heart tissue treatment site, and progressively promotes angiogenic cells from which the angiogenic cells revascularize from the tissue or blood to the treatment site. Providing for the treatment of heart tissue. In another example, the platelet composition further comprises a molecule that attracts angiogenic cells to the treatment site. In another embodiment, the molecule is selected from the group consisting of growth factors, growth factor receptors and chemoattractants.

  In one embodiment of the present invention, a method for cardiac tissue regeneration includes at least one delivery device for introducing a platelet composition into the heart tissue such that cell tissue and regeneration of the heart tissue by the cell formulation are promoted. including.

  In one embodiment of the present invention, a method for treating heart tissue includes providing a platelet composition within a heart tissue treatment site and injecting a cell preparation into the heart tissue. In another example, the platelet composition and cell preparation are provided almost simultaneously.

(Definition of terms)
In general, all terminology and phrases appearing herein are used by those skilled in the art to understand in their ordinary sense. However, for the convenience of the reader, the selected terms are more specifically defined as follows:

  Angiogenesis: As used in this agreement, “angiogenesis” means a process that involves the development of new blood vessel physiology from existing blood vessels.

  Bioactive agent: As used herein, “bioactive agent” includes therapeutic agents and drugs, and also includes pharmaceutically active compounds, corticosteroids, growth factors, enzymes, DNA, ribonucleic acid, siRNA, Virus, protein, lipid, polymer, hyaluronic acid, antibody, antibiotic, anti-inflammatory agent, antisense nucleotide and transforming nucleic acid, remodeling (eg, angiotensin II inhibitor, angiotensin converting enzyme, atrial natriuretic peptide, aldosterone , Renin, norepinephrine, epinephrine, endothelin, etc.) and inhibitors of compounds deemed to be related, and combinations thereof.

  Chamber Remodeling: As used herein, “chamber remodeling” means atrial or ventricular remodeling. “Remodeling” means a sequence of events (including changes in gene expression, molecules, cells and stroma) that cause changes in the size, shape and function of heart tissue after stress or injury. Remodeling after myocardial infarction (MI), pressure overload (eg aortic stenosis, hypertension), volume overload (eg valve insufficiency), inflammatory heart disease (eg myocarditis) or idiopathic May occur in sex cases (eg, idiopathic dilated cardiomyopathy). Remodeling results in progressively worsening cardiac function and ultimately heart failure and is often pathological. In the present disclosure, as described above, pathological remodeling is referred to as remodeling.

  Cardiac tissue injury: As used herein, “cardiac tissue injury” means any area of abnormal tissue in the heart caused by a disease, disorder, or injury, such as epicardium, endocardium and / or Includes damage to myocardium. Non-limiting examples of causes of cardiac tissue damage include acute or chronic stress (systemic hypertension, pulmonary hypertension, valve dysfunction, etc.), coronary artery disease, ischemia or infarction, inflammatory diseases and cardiomyopathy Including. Cardiac tissue damage is most often accompanied by damage to the myocardium, and therefore for the present disclosure, myocardial damage is equivalent to heart tissue damage. In addition, the damage may be acute, such as acute myocardial infarction, where the damage is referred to as a disability event. Injured heart tissue includes ischemic, infarcted, or otherwise focally or diffusely diseased tissue.

  Composition: As used herein, “composition” means a combination of substances that can be delivered into an injection solution, substance, or tissue and used interchangeably herein. Exemplary compositions include, but are not limited to, platelet gelation, autologous platelet gel, platelet rich plasma and platelet poor plasma with or without the addition of bioactive agents, structural agents, and the like.

  Delivery: As used herein, “delivery” means providing the composition to the treatment site in the damaged tissue through any method suitable for delivering the functional composition to the treatment site. Non-limiting examples of delivery methods are well known to those skilled in the art: direct injection at the treatment site, direct topical application at the treatment site, transdermal delivery for injection, transdermal delivery for topical application, and the like. Including other delivery methods.

  Damaged area: As used herein, “damaged area” means damaged tissue. “Injured peripheral area” means tissue directly adjacent to damaged tissue. That is, tissue at the junction between damaged tissue and normal tissue.

  Damaged tissue: As used herein, “damaged tissue” means tissue that is damaged by damaged, ischemic tissue, infarcted tissue, or damaged tissue that causes totally normal blood flow obstruction to the tissue. To do. With respect to the heart, “damaged tissue” includes tissue that has undergone any changes described in “Heart Tissue Damage”.

  Isolated: As used herein, the term “isolated” means a cell that has been separated from at least some components of its natural environment. The term includes the overall physical sorting of cells from their natural environment, such as separation from the donor, and further changes in the association of surrounding cells with direct contact. In this context, the term “isolated” includes cell dissociation. The term “isolated”, as disclosed herein, can also encompass a cell population resulting from the culture and / or proliferation of isolated cells.

  Neovascularization: As used herein, “neovascularization” refers to the formation of a functional vascular network that can be perfused by blood or blood components. Neovascularization includes angiogenesis, budding angiogenesis, intussusceptive angiogenesis, sprouting angiogenesis, therapeutic angiogenesis and angiogenesis.

  Percutaneous: As used herein, the term “percutaneous” refers to the patient's form, whether in the form of a small incision, incision, perforation, tubular access to a cannula, sleeve or port, or the like. Means any penetration through the skin. Percutaneous penetration may be performed at other sites such as the interstitial space between the patient's ribs or the patient's groin area.

  Structural support: As used herein, the term “structural support” means a mechanical reinforcement that provides resistance to stress and maladaptive processes of remodeling.

  Angiogenesis: As used herein, the term “angiogenesis” is a novel process of endothelial cells that is a process of change that occurs during development and also during adulthood (eg, after injury or after myocardial injury). Means angiogenesis by production.

  The present invention provides methods and compositions for promoting neovascularization and treating heart tissue by subsequent administration of a platelet composition after a period of cell therapy. Induction of neovascularization in damaged heart tissue prior to cell preparation in vivo transplantation increases the survival, coalescence, and maintenance of cells transplanted into the damaged tissue.

  After injury, such as ischemic injury, the blood supply to the tissue is often insufficient to maintain the remaining healthy tissue. This changing process, in which persistent ischemic tissue dies and adjacent tissue is at increased risk of ischemia, leads to an increase in the ischemic area over time. Increasing blood supply to damaged or adjacent tissue can prevent further ischemia, as does chamber remodeling.

  In an embodiment of the invention, the method promotes angiogenesis in the heart tissue by directly injecting the platelet composition into the damaged or surrounding heart tissue, and then cell therapy to promote regeneration of the damaged tissue. Providing.

  Neovascularization refers to the development of new blood vessels from endothelial progenitor cells by any means, such as by angiogenesis, angiogenesis, or the formation of new blood vessels from endothelial progenitor cells that link to existing blood vessels. Angiogenesis is the process by which new blood vessels grow from the endothelium of existing blood vessels in a growing animal. Endothelial progenitor cells circulate in the blood and selectively migrate or “home” to sites of active neovascularization (see US Pat. No. 5,980,887, Isne et al., The contents of which are hereby incorporated by reference in their entirety. It is considered as described in the description).

  In one embodiment of the invention, the method provides the platelet composition to a treatment site in heart tissue where the composition promotes neovascularization of heart tissue and injects the cell preparation into the neovascularized heart tissue. And where the cell preparation includes providing for the regeneration of the heart tissue described above.

  The present invention will be described in detail below with reference to the figures where like numerals indicate like structures. With reference to FIGS. 1 and 4, a depiction of a normal heart 10 can be seen. The cross-sectional view of FIG. 4 shows a right heart 44 and a left ventricle 42 of a normal heart that has not undergone chamber remodeling.

  FIG. 2 represents that the heart 20 has an infarct region periphery 26 surrounded by an ischemic or infarct region 24 and a healthy non-ischemic myocardium 28. When an infarct occurs, heart tissue that is no longer receiving adequate blood flow dies and scar tissue is replaced. Insufficient blood flow in the damaged tissue that triggers the injury, the chain of events (remodeling) thins and widens the wall, and eventually stops functioning would otherwise cause regrowth of the damaged tissue and Prevents cells and mechanisms originating within the body part that may lead to recovery.

  FIG. 3 is an enlarged view of a portion cut off by a broken line in FIG. 2 and 3 represent the extent of myocardium 24 that has undergone some ischemic injury, such as myocardial infarction or other injury. When necrosis occurs, that part of the myocardium that has suffered from necrosis is completely infarcted. The area directly surrounding the ischemic / infarcted area 26 is known as the peri-infarct area and is surrounded by a healthy myocardium 28. As discussed above, the peri-infarct region 26 can suffer from some degree of ischemic activity, but the blood supply has not yet been interrupted to the same extent as the ischemic / infarct area 24.

  Remodeling is usually a progressive hypertrophy of the ventricle with a worsening of ventricular function and can occur for weeks to years after myocardial infarction. There are many potential mechanisms for remodeling, and it is generally believed that high stress on peri-infarct tissue plays an important role. Due to the altered shape and size, the wall stress is much higher than the normal myocardial tissue surrounding the infarct formation. FIG. 5 is a cross-sectional view of the heart shown in FIG. FIG. 5 shows a left ventricle 52 having a right ventricle 54 and a remodeled left ventricular region 50. As can be seen in the figure, the heart wall is thinner than the expanded region 50.

  A limited amount of remodeling can be advantageous for the patient and occurs mainly in two contexts. The first is termed “physiological remodeling” that occurs in athletes with some high performance as an adaptive response to above-average demands on the heart. Compensatory changes in heart shape and size and function in a physiologically remodeled heart allow the heart to work better in high performance environments. The second situation is during the earliest stages of post-damage remodeling. Sometimes this early stage of remodeling can actually be adaptive and protective. If to a limited extent, some cellular rearrangements in the heart wall and increased chamber volume can maintain or even increase cardiac output. These changes can be beneficial. However, the wall composition changes further, the shape and size of the ventricle gradually becomes dysfunctional, and this “pathological remodeling”, which sometimes leads to heart failure, in most cases proceeds beyond what is adaptive. To do. “Pathological remodeling” as described above is referred to as “remodeling” in this disclosure.

  Details are under study, but the remodeling mechanism appears to involve a chain of events. When muscle cells are stretched under mechanical stress, the local activity of some molecules increases (eg noreprenamine, angiotensin, endothelin and others). These molecules promote the expression of specific proteins, leading to hypertrophy of existing muscle cells. This leads to further deterioration in cardiac function (eg applied mechanical stress) and increased neurohormonal activation. Some termination factors further promote local collagen synthesis leading to fibrosis and scarring of the affected area. These changes are often beyond compensatory and lead to progressive heart failure.

  In the absence of adequate blood flow in the affected area, the intrinsic repair mechanism cannot restore heart tissue or function. Although cells from body parts have been shown to "home" to damaged cells even in the adult heart, blood flow limitation can prevent cells from body parts from living and promoting healing.

  Progressive deterioration in cardiac function can occur early without symptoms. Over time, however, symptoms of clinical heart failure such as shortness of breath, swelling, dyspnea in the supine position, arrhythmia, organ failure, etc. develop. It is important to pay attention even to patients with asymptomatic cardiac dysfunction, and mild heart failure is at high risk of sudden cardiac death. Therefore, there is a great incentive to treat this disease process early and effectively.

  Means for assessing cardiac remodeling include heart mass, heart shape, heart mass, systolic ejection fraction, end-diastolic and end-systolic volumes, and peak systolic forces. Left ventricular volume (particularly left ventricular end systolic volume) is the best predictor of human mortality after myocardial infarction.

  Drug therapy providing enzyme inhibition to convert angiotensin (eg, captopril, enalapril) and beta sympathetic blockade (eg, carvedilol, metoprolol, propranolol, timolol) has been shown to slow down certain parameters of cardiac remodeling ing. The intention of these therapies is to reduce the body's remodeling response to harmful or mechanically stressful stimuli and to reduce mortality and morbidity in patients with myocardial infarction and heart failure in clinical trials It is shown. Other therapies, such as antihypertensive agents, have been used to reduce the chronic burden placed on the heart that can cause or exacerbate pathological remodeling. Despite the use of the aforementioned drugs, remodeling remains at best a partially treatable process. Furthermore, none of these agents cause cardiovascularization in damaged tissue as a means to prevent further heart damage or to restore heart tissue or function.

  As further described below, embodiments of the present invention handle cardiac injury and remodeling by injecting the composition into the heart wall to produce cardiovascularization, and thus remodeling in advance. prevent. The injected composition can occupy some of the interstitial space between cells in the heart wall region, providing neovascularization as well as providing structural reinforcement for the tissue. The present invention contemplates providing neovascularization at every heart wall site and includes both the atria and ventricles.

  The infused platelet composition may be a substance that can provide some level of structural support in the tissue as well as desirable neovascularization. Substances that can provide both tissue reinforcement and stimulated neovascularization can be included in the platelet compositions disclosed herein. For the purposes of this document, the term “platelet gel” refers to a platelet composition that is administered with an activator and may comprise both tissue reinforcement and biological therapy such as neovascularization. Furthermore, a platelet composition can mean rich blood or platelet poor plasma administered without an activator. Platelet compositions such as hyperemic or platelet poor plasma can be activated by tissue thrombin in situ in addition to provide both structural support and neovascularization. Illustratively, non-limiting platelet compositions include platelet gel, autologous platelet gel, platelet rich plasma and platelet poor plasma. Residence in the target tissue poses a significant challenge to the currently developing somatic cell-derived technology. When delivered almost simultaneously with the cells, the platelet composition comprising the structural support will further act to increase the tissue retention of the delivered cells.

  Platelet compositions that cause neovascularization of the present invention include, but are not limited to, collagen, cyanoacrylate, adhesive that heals by injection into tissue, solidifies or gels after injection into tissue Liquids, suture materials, agar, gelatin, light activated dental composites, other dental composites, silk elastin polymers, Matrigel® (BD Biosciences), hydrogels and other suitable organisms It can be administered with other compositions that can provide a structural support comprising the polymer. Such compositions can include single or multi-component compounds. These compositions can include an agent that is delivered as a liquid and gels or hardens into a solid after delivery. Curing / gelling can be caused by temperature, pH, protein, or other environmental factors that are specific to or created in the target tissue. These platelet compositions can be infused separately or with each other and / or in combination with the platelet composition. In addition, the composition or combination can include other additives. Further, some of these compositions and / or additives will be described below.

  The platelet composition of the present invention can be reinforced with a biocompatible fluid that solidifies and / or cross-links in situ to provide a structural support structure upon delivery to the heart wall. Other embodiments of the platelet composition of the present invention may include, for example, synthetic or naturally occurring substances and / or non-degradable or biodegradable substances to provide strength. In one embodiment, the structural material comprises cyanoacrylate or silk elastin protein polymer.

  The platelet compositions of various embodiments of the present invention can include additives such as fibrinogen to increase the structural strength of the heart wall. Fibrinogen can be autologous, allogeneic, recombinant, human, modified, purified from animal sources. At least one embodiment includes elastin to increase the elasticity of the treated heart wall. Compositions as liquids (without cross-linking or coagulation components) so that important soluble factors are trapped in the target tissue (either physically or by binding to the site) without having an intrinsic structural element It can be delivered. Alternatively, the composition can be delivered with one or more structural materials to provide additional structural support to the tissue.

  The present invention may be practiced to use materials that contain synthetic biodegradable materials that provide strength during a specific time interval after delivery and are resorbed. Such substances include genetically modified or modified compounds such as collagen or fibrin. Provides strength during a specific time interval after delivery, and then resorbs cartilage, bone or bone components, gelatin, collagen, glycosaminoglycan, starch, polysaccharides, any other substance, but this Without being limited to, naturally occurring materials can be used as well.

  Other embodiments of the invention may include any combination of various compounds that can create the desired local effects of tissue expansion. Components that provide local edema, tissue hypertrophy, structural reinforcement of the tissue, or any other effect that obviates remodeling are included in this invention. Such compounds include suture materials that have been ground to cause edema and hydrogels for tissue structural reinforcement. These substances can be added to PRP (platelet rich plasma), or PRP (platelet rich plasma) + thrombin.

If the desired effect is tissue structural reinforcement, 50-100 μm (at the maximum width of the particle) that is biodegradable that is small enough for injection with a needle but too large to fit into capillaries and venules Medium sized microparticles can be added to the platelet composition. In one embodiment of the present invention where microparticles can be infiltrated with a drug that elutes as the particulate matter degrades, only the microparticles are delivered to the heart tissue by infusion into the coronary sinus. Based on size characteristics, they are expected to remain in the organization and provide structural reinforcement of the organization. The microparticles used will have a glass transition temperature (Tg) > 37 ° C and will gel over several days after insertion. The injected microparticles provide a “mass” and volume for immediate tissue structure reinforcement, but will soften into a gel to become a single member over time.

  Embodiments of the platelet composition of the present invention can be covalently linked directly to one or more proteins located on the surface of one or more cell types so as to maintain the polymer at the site of injection. Polymers can be included. In one example, a polymer that covalently binds to the primary amine group (—NH 3) of the protein may be used.

  The cells used in the cell preparation of the present invention include cells that grow and transplant into the heart muscle of the patient. The cells may be obtained from a single individual or a plurality of individuals, and may be the same or different from the recipient individual. In one embodiment, the cell is autologous.

  Suitable cell sources for use in the cell preparations of the present invention include embryonic, embryonic, postnatal or adult stem or progenitor cells, cardiomyocytes, skeletal myocytes, skeletal myoblasts, mesenchymal stem cells, endothelial progenitor cells, Includes hematological cells, immune cells, and combinations thereof. Stem cell sources include bone marrow, blood, adipose tissue, gonadal, skeletal or heart muscle, or any tissue containing stem cells. These cells can be obtained by any suitable method known by those skilled in the art.

  Suitable physiological carrier solutions include, for example, solvents or dispersion media including water, ethanol, polyols (including but not limited to glycerol, polyethylene glycol and propylene glycol), and combinations thereof.

  The number of cells administered to a patient will vary depending on the patient being treated. In one example, 1 × 10 4 to 1 × 10 9 cells are administered. However, accurate determination of cell volume is based on individual factors for each patient, including weight, age, size of damaged tissue, and amount of time since injury. Based on the current disclosure and general knowledge in the industry, those skilled in the art will similarly determine the dose of cells, the amount of platelet composition, the amount of carrier solution, and other parameters related to the administration of platelet compositions and cell preparations. Can be easily determined.

  In another embodiment of the invention, after injection at the treatment site, the platelet composition attracts angiogenic cells to the treatment site, where the angiogenic cells promote neovascularization of heart tissue. In another embodiment, the platelet composition further comprises molecules that attract angiogenic cells to the treatment site. Non-limiting examples of such molecules include cell growth factors, growth factor receptors and chemoattractants.

  In addition, each or both of the platelet composition and the cell preparation can include one or more bioactive agents that promote the healing or regeneration of damaged heart tissue. Suitable bioactive agents include pharmaceutically active compounds, corticosteroids, cell growth factors, enzymes, DNA, ribonucleic acid, siRNA, viruses, proteins, lipids, polymers, hyaluronic acid, pro-inflammatory molecules, antibodies, Including but not limited to antibiotics, anti-inflammatory agents, antisense nucleotides and transforming nucleic acids or combinations thereof.

  These cells can spontaneously secrete one or more biologically active molecules, or can be the product of genetic engineering that secretes a therapeutically effective amount of one or more biologically active proteins. Suitable biologically active proteins include, but are not limited to, growth factors and cytokines. The secretion can be controlled by the presence of an inducible promoter, or the secretion can be structural.

  Growth factors and cytokines useful in the methods of the present invention include, but are not limited to, stem cell factor (SCF), granulocyte colony stimulating factor (G-CSF)), granulocyte macrophage colony stimulating factor (GM-CSF), stroma Cell-derived factor-1, hematopoietic stem cell factor (SteelFactor), vascular endothelial growth factor, macrophage colony, granule cell macrophage stimulating factor, hepatocyte growth factor (HGF), insulin-like growth factor (IGF-1), interleukin (IL ) -3, IL-1- and IL-1-IL-6, IL-7, IL-8, IL-11 and IL-13, colony stimulating factor, thrombopoietin, erythropoietin, fit3-ligand, mass necrosis factor α, leukemia inhibitory factor, tumor growth factor-β1 and -β3; macrophage inflammatory protein 1α, angiogenic factor (fibroblast growth factor 1 and 2, vascular endothelial growth factor), platelet-derived growth factor A, epithelial cell growth Long factor, tumor growth factor-β1 and β2, oncostatin M, insulin-like growth factor-1, nerve growth factor, HIF-1, endothelial PAS domain protein 1 (EPAS1), monocyte chemotactic protein-1 (MCP-1), keratinocyte growth factor, platelet-derived growth factor, heparin-binding epidermal growth factor, or any cytokine or growth factor that can stimulate, maintain, and / or mobilize cells.

  The site and extent of the injection site is identified before any composition is injected into the heart with the damaged tissue region to cause neovascularization or to provide structural reinforcement of the tissue. A number of techniques and techniques are available for clinicians to identify and assess heart tissue that is normal, injured and not viable, and injured but can survive. These include visual inspection during open chest surgery, regional blood flow measurements, local electrical and structural activity, nuclear cardiology, echocardiography, echocardiographic stress test, coronary angiography, nuclear magnetic resonance imaging (MRI), computed tomography (CT) scanning and ventricular imaging, including but not limited to.

  In one embodiment of the invention, the platelet composition is prepared using a Medtronic Magellan® platelet separator. Anticoagulated whole blood is prepared by combining an anticoagulant with fresh whole blood voted from a subject. The MagellanR instrument is then used to extract platelet rich plasma (PRP) and platelet poor plasma (PPP) from anticoagulated whole blood specimens. Platelet gels are prepared by binding the resulting PRP or PPP with an activator. In one embodiment, the activator is bovine thrombin returned to 1000 units / ml in a 10% calcium chloride solution. In another embodiment, PRP is combined with bovine thrombin in a ratio of about 10: 1.

  In one embodiment of the present invention, the platelet gel composition is a platelet gel made using PRP and thrombin to a ratio of about 10: 1. In another example, PRP is used in a ratio of about 11: 1 to thrombin. Other embodiments of the invention have a PRP to thrombin ratio of about 5: 1 to about 25: 1. In another embodiment, the ratio of PRP to thrombin is from about 7: 1 to about 20: 1. In another embodiment, the ratio of PRP to thrombin is from about 9: 1 to about 15: 1. In another embodiment, the ratio of PRP is from about 10: 1 to about 12: 1 with respect to thrombin. In at least one embodiment, thrombin is not included and PRP alone is injected into the heart tissue. Other embodiments of the invention include a number of components of the composition in the proportions required to achieve or optimize the desired effect.

  When PRP and thrombin are injected in a mixed manner to form a platelet gel in heart tissue (see description of delivery device below), they become a gel in the tissue. Some embodiments of the invention have an accelerated gel time. Apply local heat to the injection site through a delivery catheter or other device, increase thrombin concentration, or combine PRP and thrombin in the mixing chamber, and inject the mixture into the heart tissue after the mixture begins to gel Thus, the gelation time at the in vivo position can be accelerated. The description also applies to other component compositions in which the components gel, crosslink, and / or polymerize after being mixed together.

  PRP contains high concentrations of platelets that can aggregate during gelation, as well as free cytokines, cell growth factors or enzymes after activation. Some of the many factors released by platelets and leukocytes that make up PRP are free platelet-derived growth factor (PDGF), platelet-derived epidermal growth factor (PDEGF), fibroblast growth factor (FGF), transforming growth factor β ( Mutant growth factor-β) and platelet derived angiogenic growth factor (PDAF). These factors have been considered to be involved in wound healing by increasing the rate of collagen secretion, blood vessel ingrowth and fibroblast proliferation.

  Once the site, size and shape of the injury area has been determined, the clinician can access and begin injecting the platelet composition into the heart wall. In one example, the platelet composition comprises PRP and thrombin. In another example, the platelet composition comprises only PRP. In another example, the platelet composition comprises PPP and thrombin. In another example, the platelet composition comprises only PPP. The components of the platelet composition may be derived from humans and / or animals and / or recombinant sources. The component may be artificially created as well. The components for platelet compositions that can be classified as autologous or non-self-derived and non-self-derived components are as described above (ie, animal, recombinant, (Modified type, human self-type, etc.) Autologous platelet gel (APG) refers to a composition made from autologous PRP or autologous platelet poor plasma and self or non-self activator. One advantage of using autologous and / or recombinant components in injecting compositions is that it reduces the risk of inflammatory reactions or exposure of recipients to infectious and foreign agents. It is.

  Further, each or both of the platelet composition and the cell preparation of the present invention may contain a contrast agent for detection by X-ray, nuclear magnetic resonance imaging (MRI) or ultrasound. Suitable contrast agents are known to those skilled in the art and include, but are not limited to, radiopaque materials, echogenic materials and paramagnetic materials. Contrast agents can be used in some example compositions for visual confirmation of successful injection. Examples of such contrast agents include, but are not limited to, such examples, X-ray contrast (eg, IsoVue or other contrast agents with high X-ray attenuation factors), magnetic resonance imaging (eg, MRI signal) Or including, but not limited to, cadolinium or other contrast agents detectable as signal voids (no signal) and ultrasound contrast (echogenic or echopaque compounds).

  As explained in more detail in the Examples, the platelet composition of the present invention was injected into the damaged heart wall of test subjects (sheep and pig). Experiments show that when performed on infarcted or non-infarcted tissues, infusions of PRP and thrombin are safe and well tolerated and can be performed safely even as early as 1 hour after MI. It was. Controlled infusion was possible with or without a cardiac stabilizer and infusion was possible without external cardiac pacing. Injections were made at right angles and at an angle to the heart surface at intervals of 0.5 to 2.5 cm. Multiple injections can be performed per heart without safety issues. The total volume that can be injected can be up to 15 mL, and the volume of individual injections can be up to 1100 ul per injection site.

  In addition, administration of autologous platelet gel following myocardial injury can partially or completely restore the infarct's deleterious acute effects on systolic ejection fraction (EF), or to bring EF to pre-infarct levels or It can be expanded further. In some surprising results, administration of autologous platelet gel to ischemic tissue after myocardial injury promotes neovascularization in the damaged tissue (FIGS. 16-17). This angiogenesis was not observed in infarct animals that did not receive platelet gel therapy. All or some components of platelet gel (PRP or PPP components with or without thrombin) can be used to produce such an effect.

  To practice the present invention and deliver platelet compositions and cell preparations to target sites within the heart wall, clinicians may use one of a variety of access techniques. These include surgical (sternotomy, thoracotomy, mini-thoracotomy, xiphoid) and percutaneous (transvascular and endocardial) approaches. Once achieved, the composition can be delivered by epicardium, endocardium, or transvascular delivery. Platelet compositions can be delivered to heart wall tissue at one or more locations. This includes intramyocardial, subendocardial and / or subepicardial administration. The following delivery methods are suitable for delivery of platelet compositions and / or cell preparations in accordance with the teachings of the present invention.

  One way to deliver the platelet composition as expected into such moving target tissue is to synchronize the infusion timing for delivery, particularly during the selected portion of the cardiac cycle. is there. In one embodiment of the invention, one or more electrodes may be used as stimulation electrodes, for example, to keep the pace of the heart during composition delivery. In this way, the cardiac cycle is made predictable and the timing of the injection can be timed and synchronized. In fact, the heartbeat interval time can be artificially increased so as not to interfere with complete delivery during a particular (and relatively) stationary phase of the cardiac cycle. In one example, the delivery device includes one or more stimulation and / or detection electrodes. In one embodiment of the invention, a sensor may be used to sense heart contraction, thereby allowing the delivery of platelet composition to be timed with the heart contraction. For example, it may be desirable to deliver one or more components of a platelet composition between heart contractions.

  Regardless of the method used to access or stabilize the heart with a damaged heart tissue region, the delivery device used can inject multiple components separately into the heart wall. There is a need to get. One embodiment of the invention allows repeated infusions with a single instrument. This can be achieved with a proximal one-handed trigger that allows for predictable delivery of a determinable amount (eg dial-in) of a single or multiple component composition or cell preparation in a determinable ratio. Different embodiments of the present invention utilize a delivery device having two lumen needles / delivery catheters, and at least one other embodiment has three or more lumen needles / A delivery device having a delivery catheter is used. The lumen of the needle / delivery catheter can be coaxial or biaxial.

  At least one embodiment of the present invention includes two or more side-by-side injectors for single-hand injection of multiple composition components. In one embodiment, the device of FIG. 9 is used to inject multiple component platelet compositions into an injured heart 100. In the embodiment of FIG. 9, the two components of the composition of the present invention are housed separately in syringes 102 and 104. Injectors 102 and 104 are placed in cradle 112 within handle assembly 106 to allow for one-hand injection of the composition. An adapter 108 connects the injectors 102 and 104 to the biaxial needle 110. A biaxial needle 110 allows delivery of PRP and thrombin in a non-limiting example, which is two components of the platelet composition at the treatment site in the heart 100.

  FIG. 10 represents an enlarged view of two component composition injections according to the present invention using a biaxial injection needle containing a delivery device 300. Component 310 is kept in the reservoir or in the injector 306, and component 308 is kept in the reservoir or in the injector 304. Components 310 and 308 will pass through a biaxial injection needle 318 that includes an injection needle lumen 314 of component 310 and an injection needle lumen 312 of component 308. Components 310 and 308 are simultaneously injected into treatment site 302 and the two components are combined to form composition 316. Immediately after injection, components 310 and 308 and some of the composition 316 diffuse through the tissue at the therapy site 302. Components and compositions were observed to diffuse up to 2 centimeters in heart tissue (see also FIG. 11).

  The delivery system may deliver the components of the platelet composition at a defined ratio. This ratio may be preset (and fixed) or dialed (and changed). One embodiment of the present invention allows the delivery of multiple compounds at different ratios without creating a pressure gradient between the injectors (with a configurable tooth ratio or lever ratio) with a separate gear or lever. For use. Other component delivery devices according to the present invention include different caliber lumens that allow a predetermined ratio of each component. Some multi-component delivery devices of the present invention include different lengths of lumens so that one component is released more distally than the other. Nevertheless, other equipment incorporates one or more mixing chambers in the equipment. At least one embodiment of the delivery device of the present invention includes a single lumen needle / catheter that is used for the delivery of multiple component series (next to next).

  Some embodiments of the delivery device are placed in a blood vessel adjacent to a target treatment site and penetrate the heart wall by penetrating the vessel wall and successfully progressing to the desired site with a needle tip or a miniature catheter contained in the injection needle. It can be used to deliver a platelet composition. The catheter or needle may include a local diagnostic imaging system to identify the target area and place the delivery device in place. The instrument may include one or more needles having a closed distal end and one or more armpits for directing material from the distal end sufficiently laterally into the heart wall. Desirably, the injection needle has a sufficiently small gauge diameter so that the needle insertion pathway in the heart is sufficiently self-sealing to prevent leakage of the composition upon withdrawal. Recent data (obtained in the context of epicardial delivery) demonstrates hemostatic effects when platelet gel is injected even through a large 18 gauge needle. This result may be attributed to the rapid blood clotting achieved by the hemostatic effect inherent in the injected components and platelet gel. In another embodiment, the needle gauge is less than 18 gauge. In one embodiment, the needle gauge is 26 gauge.

  As an alternative, the delivery assembly may include one or more needles having multiple lumens spanning between multiple line collection tubes on the proximal end adjacent to the exit port. Multiple lumen needle assemblies can allow a component of a substance to be injected alone, thereby delivering the component within a selected tissue region, as described herein. Allows to react with each other later.

  In one embodiment, as described herein, a plurality of lumen needle assemblies can be obtained that allows two components of the composition to be injected simultaneously and independently, and then selected from each other. Can react once in a different tissue area. In another embodiment having multiple lumen needle assemblies, the lumens are injected into a mixing chamber located near the distal end of the needle and before being injected into a selected tissue region. The components of the injected substance are immediately mixed with each other.

    The platelet composition and cell preparation of the present invention can be delivered to the heart wall by a catheter system. Suitable catheter delivery schemes for the present invention include those having multiple biaxial or coaxial lumens with twisted or flat tips. The catheter system of the present invention can include a needle or other injection device located at the distal end, and an injector at the proximal end of the catheter. The catheters and other delivery devices of the present invention may have lumens that are sized differently to ensure that multiple component compositions can be delivered to the heart tissue in the desired ratio. Another example of a possible catheter system may be used to create a composition reservoir in the heart wall itself for sustained release. The catheter may be vascularized into the blood vessel until the distal portion is adjacent to the desired therapy site. The needle assembly can be placed in the correct orientation to puncture the vessel wall and enter the heart tissue. The composition can then be injected into the heart tissue, thereby forming a reservoir. When using catheterization, clinicians access the heart through blood vessels, or epicardial (FIG. 12), endocardial (Figure 13A-B), or transvascular (Figure 14A- B) One of several routes that successfully travel to the heart chamber for delivery of the composition can be used to successfully navigate the patient's heart. 12 and 13 show a cross section of the entire heart. In FIG. 14, the right ventricle and atrium are shown in cross-section, while the left ventricle and atrium are shown closed so that the epicardial surface and its coronary vessels can be seen.

  The epicardial delivery of the platelet composition can be achieved by accessing the treatment site 520 in the left ventricle 516 of the heart 200 from the epicardium, the outer surface of the heart, as shown in FIG. 520, injecting the composition.

  Endocardial delivery of platelet compositions (FIGS. 13A and B) can be achieved, for example, via the superior vena cava (delivery device 540) or inferior vena cava (delivery device 540) into the right ventricle 504 (FIG. 13A). Transcutaneously, including accessing the treatment site 520 in the left ventricle of the heart 200 with the delivery device 540, 540 ′. The delivery device 540 passes through the atrial septum to enter the left atrium 508 and then into the left ventricle 516 to reach the therapy site 520 where the composition is injected by the delivery device 540. An alternative endocardial delivery method depicted in FIG. 13B is a treatment site that enters the left atrium 508 through the aorta 512, for example, percutaneously retrogradely with the delivery device 560, and then injects the composition with the delivery device 560. Entering the left ventricle 516 to reach 520 and accessing the treatment site 520 in the left ventricle of the heart 200.

  Transvascular delivery of platelet compositions (FIGS. 14A and B) can be achieved, for example, via the superior vena cava 500 (delivery device 580) or inferior vena cava 502 (delivery device 580 ′) into the right ventricle 504 (FIG. 13A). ) Percutaneously through the delivery device 580, 580 ′ to access the treatment site 520 in the left ventricle of the heart 200. The delivery device 580 passes through these veins through the coronary sinus 503 to the cardiac venous system and, if necessary, leaves by tracing these veins through the heart tissue and the composition is injected with the delivery device 580. To reach the treatment site 520. An alternative transvascular delivery method depicted in FIG. 14B is, for example, to reach the treatment site 520 where the composition is injected through the coronary artery 595 through the coronary artery 595 percutaneously with the delivery device 590. Access to the treatment site 520 in the left ventricle of the heart 200.

  Devices for injecting the platelet compositions and cell preparations of the present invention can include a refrigerated portion to keep the various composition components cool. Various embodiments of a delivery device for practicing the present invention include a refrigerated / cooled chamber for thrombin refill, a refrigerated / cooled chamber for thrombin, and / or a stirring mechanism for PRP refill, or sediment. An injection chamber can be included to prevent this from occurring. The delivery device can include a heating or cooling device used to warm or cool the heart tissue or composition to speed up or slow down the gelation / curing rate after delivery. Some devices of the present invention include a single lumen or multiple lumen catheter that is cooled to keep the components of the infusion composition cool while traveling through the device lumen. , Or other delivery devices. As noted above, some devices may include a mixing chamber for mixing the components of the composition to be injected before the substance is delivered into the tissue. In one embodiment of the invention, the PRP is stored in an agitation / vibration chamber that provides sufficient agitation to keep the PRP homogeneous. In another example, the clinician provides sufficient agitation to the delivery device by tilting or otherwise manipulating the device to keep the PRP homogeneous.

  A clinician practicing the present invention may need to make multiple infusions using a single delivery assembly. As such, at least one embodiment of the delivery device of the present invention includes a device having at least one reusable needle. Some embodiments of the present invention may include an automatic dispensing system, eg, a delivery device that has a mode for advancing infusion with an injector. The automatic dosing system can determine each amount in advance and allow dialing (which can be variable or fixed), eg, a screw-type setting scheme. One embodiment of the invention may include a base handle in which a predetermined dosage is delivered at a predetermined or manually controllable rate each time the base handle is pushed.

  In a further alternative embodiment, the delivery system may include a plurality of needle assemblies (similar to the individual needle assemblies described above) to be implemented in a pre-determined arrangement along the outer surface of the catheter. In one embodiment, the needle assembly may be arranged in one or more rows. It is particularly desirable to access a wide range of distant tissue regions within the myocardium that extend considerably parallel to, for example, blood vessels. With a multi-needle transvascular catheter system, a single device may be delivered into the blood vessel and placed in the correct position. The plurality of arranged needles may be placed sequentially or simultaneously to inject the composition into a wide range of tissue regions, thereby providing a selected trajectory pattern. A catheter-based device as described above is disclosed in US Pat. No. 6,283,951, the disclosure of which is hereby incorporated by reference.

  When clinicians practice this invention using minimally surgical or percutaneous techniques, some kind of real-time visualization or guidance may be required to ensure site-specific injection. is there. As such, at least one embodiment of the present invention uses pre-operative MRI or MRI guidance techniques that overlay CT images on fluoroscopic images of the delivery catheter to track the catheter to the target site in real time. In one embodiment, the clinician uses a guidance technique that tracks the contrast agent and / or needle tip during injection in a virtual three-dimensional environment. This technique records previous injections to ensure adequate space for future injections.

  The needle assembly (or other instrument component) may include a feedback element or a sensor for measuring physiological condition reflexes to direct delivery of the composition to the desired site. For example, EKG guidance may be included at the distal end, otherwise to detect and guide injections towards electrically quiet regions of heart tissue, or within the heart monitored during delivery of the composition It may be sent into a selected tissue region to account for electrical events. During treatment, for example, the composition may be delivered into the tissue region until it meets the desired conditions. Similarly, a local EKG monitor can target an electrically quiet area of heart tissue and guide the injection towards it.

  Regardless of the equipment used to deliver the platelet compositions, or how the clinician accesses the heart wall, the clinician practicing the present invention will have accurate local placement and accuracy for each infusion. Deep control is needed. In one embodiment of the invention, the platelet composition and / or cell preparation is delivered / injected to a depth approximately midway between the outer and inner walls of the heart wall. In other embodiments, the compositions are delivered to a depth closer to the inner or outer wall. The compositions can be delivered endocardial, quasi-endocardial, or quasi-epicardial. In another embodiment of the invention, the depth of injection varies with the thickness of the target tissue, and the depth is shallower at the apex than at other parts of the heart.

  In order to obtain depth control, at least one delivery device of the present invention can be applied to the needle body to prevent it from penetrating tissue beyond a specified depth at a desired distance from the distal end of the needle. Includes a fixed stopper (or fixed for adjustment). In some embodiments, at least one embodiment of the present invention uses a method of inserting one or more needles into the tissue tangentially to the tissue to control the depth of injection. It can be placed at the injection position at any angle between vertical (90 degrees), tangent to tissue (0 degrees), and so on. Suction can facilitate injector positioning and intrusion control.

  Referring to FIG. 6, one can see an example of an injection in which the needle 65 of a delivery device (not shown) according to one embodiment of the present invention is approaching the remodeling portion of the heart wall 60 at a normal vertical angle. it can. A needle penetrates the heart wall directly at the point 61 above the desired delivery site 62 in the heart tissue. The device may include one or more means to ensure that the needle achieves the desired penetration depth in the myocardium, for example as described above.

  FIG. 7 shows an example of an injection according to one embodiment of the present invention when the needle 75 of the delivery device (not shown) approaches the desired injection point 71 in the heart wall at an almost tangential angle. The needle penetrates the heart wall at a point 71 located at a desired distance tangentially from the desired delivery site 72 in the heart wall. Although not shown, an aspiration-type stabilization device may be used on the surface of the heart near the location or near the injection site to stabilize the target area or heartbeat adjacent area. The device secures the dome-shaped portion of the myocardium 70 so that the composition can be delivered. In order to ensure that the needle achieves but does not exceed the penetration into the myocardium, the device may include one or more means as described above.

  At least one embodiment of the present invention includes a “Smart-Needle” that detects the distance from the needle tip to the ventricular blood septum or endocardial surface so that the needle tip is maintained within the heart wall. use. Such a needle can be relied upon for diagnostic imaging around or beyond the tip of the needle by an image diagnostic method such as ultrasound.

  Sometimes it may be desirable to distribute the platelet composition and / or cell preparation as widely as possible around the injection site. It may be desirable to have a uniform distribution of the platelet composition around the injection site as well. One way to increase the distribution of platelet composition around the injection site is to use a needle with a side hole as opposed to the use of a needle with a hole at the end. A large number of side holes can provide a wide distribution of the composition around the injection site. The side holes also provide access to the tissue from multiple locations rather than only from the tip of the needle, thereby requiring less movement due to the wide distribution of the composition. The potential benefit of the side holes of these needles is that even if the needle tip happens to penetrate the heart wall and enter the heart chamber, it is still in this case, as opposed to being injected into the intracardiac blood flow. Note that the composition will be injected into the heart tissue. Another way to increase the distribution of the composition around the injection site is to increase the number of needles used at the injection site. If desired, the multi-needle delivery device of the present invention allows multiple needles to be placed close together to provide a uniform distribution over a large area compared to the use of a single needle device. The combination of side holes in the needles of a multi-needle device can provide a wide distribution of the composition around the injection site.

  In one embodiment of the invention, suction may be used to improve the distribution of the composition around the injection site. Negative pressure can be created in the interstitial space through the use of suction. Since the composition is moved by a negative pressure gradient, this negative pressure in the interstitial space can help the composition move more freely and far away. The combination of suction and side holes on the needles of the multi-needle device can provide a more thorough and wide distribution of the composition around the injection site.

  In one embodiment of the invention, the platelet composition from the delivery device to the tissue can be enhanced via the application of electrical current, for example via electrophoresis. In general, the delivery of ionized material into the tissue can be enhanced via a small current flowing between the two electrodes. Positive ions can be introduced into the tissue from the cathode to the anode or from negative ions. The use of iontophoresis can greatly facilitate the transport of certain ionized substances through tissues.

  In one example, one or more needles of the delivery device can serve as positive and / or negative electrodes. For example, a ground electrode may be used in combination with a needle electrode via a monopolar array to ionically migrate an ionized composition to a target tissue. In one example, the composition can first be dispersed from the needle to the tissue. Following delivery, the composition can also be driven deeper into the tissue iontophoretically through the application of an electric current. In one example, a delivery device having multiple needles may include positive and negative bipolar with a bipolar array. Further, in one embodiment, multiple needle electrodes can be used to inject material and / or to provide current simultaneously or sequentially.

  One goal in practicing the present invention is to avoid inadvertent delivery of substances or cells into one or more chambers of the heart, coronary arteries, or venous system. Delivery to one or more areas can have negative consequences such as, for example, pulmonary or systemic embolization, stroke, cardiac congestion, and / or remotely located thromboembolism. The present invention addresses the obstruction of these negative consequences with a variety of methods. In at least one embodiment of the invention, in order to minimize the movement of one or more of the components, the composition of the composition is gelled or polymerized in situ almost instantaneously. Select component ratios. In one example, a balloon catheter is placed in the coronary sinus and inflated during delivery until gelation is complete. This obviates the migration of liquid components from the tissue to the coronary vein branch, and instead promotes residence and gelation in the target tissue. At least one embodiment includes a pressure control system in the delivery device that ensures that the infusate pressure never exceeds the ventricular chamber pressure. This facilitates storage in the tissue and obviates pressure-driven transfer of the composition through the Thebesian venous system into the chamber of the heart. One embodiment of the present invention uses a “smart needle” as described above to obviate negative outcomes.

  At least one embodiment of the present invention provides a local negative pressure to obviate the outward flow of components from the tip of the needle between injections that cover or require multiple injections. Including a proximal manual end sleeve. In at least one embodiment, the column of catheter components is kept below a certain minimum pressure to prevent spillage between injections. In at least one embodiment, a check valve may be placed in each line to prevent one component from flowing into the line containing the other. This is important when the gelling reaction is rapid and the different components must be kept separate at the time of injection and to the injection site. This prevents clogging of the delivery device, thereby allowing repeated infusions using one device.

  At least one embodiment of the present invention may be in place for several seconds during or after withdrawal of the needle so as to utilize the infusion as a “plug” to prevent retrograde blood that has bleed prior to withdrawal following the infusion. Keep the needle (eg, 5-30 seconds beyond the blood clotting time) to prevent retrograde bleeding blood from the needle insertion path. In at least one embodiment of the present invention, the needle is left in place and withdrawn from during the expected gelation time of the material to be injected. In one embodiment of the invention, the gel time of the composition to be injected is 5 seconds.

  Some embodiments of the invention are sensors that guide the delivery device to the desired site, ensuring that the injection occurs at the desired depth, ensuring that the delivery device is at the treatment site, and providing the desired amount of It can be ensured that the composition is delivered and can include any type of sensor or other function that may require imaging means to be used. For example, real-time recording of electrical activity (eg, EKG), pH, oxygenation, lactate metabolism, CO2, or other local indicators of cardiac tissue viability or activity can lead to infusion at the desired site To help. In some embodiments of the present invention, the delivery device may include one or more sensors. For example, the sensor may be one or more electrical sensors, optical fiber sensors, chemical sensors, diagnostic imaging sensors, structural sensors and / or proximity sensors that measure conductance. In one example, the sensor may be a tissue depth sensor for determining the depth of tissue surrounding the delivery device. In one example, sensors that detect pH, oxygenation, blood metabolites, tissue metabolites, etc. are used at the end of the delivery device to alert the user when and when the tip enters the chamber blood. May be. This will cause the operator to reposition the delivery device prior to delivering the composition. One or more depth sensors may be used to control the depth of the needle that penetrates into the tissue. In this way, the needle penetration depth can be controlled according to the thickness of the tissue, for example, the tissue of the heart chamber wall, for example. In some embodiments, the sensor may be installed or placed on one or more needles of the delivery device. In some embodiments, the sensor may be placed or placed on the tissue contacting surface of the delivery device. In other embodiments of the invention, the delivery device may include one or more indicators. For example, various visual or acoustic indicators may be used to indicate to the physician that the desired tissue depth has been achieved.

  In addition, the delivery device may include a sensor that allows the surgeon or clinician to ensure that the delivery device is in the heart wall rather than the ventricle at the time of infusion. Non-limiting examples of sensors that allow determination of the position of the injector include pressure sensors, pH sensors, and sensors for dissolved gases such as oxygen. An additional sensor associated with a delivery device that is suitable for use with the present invention provides the surgeon or clinician that the backflow port or needle portion of the delivery device is in a region having blood flow rather than in tissue. A sensor that indicates blood flow, such as a back flow lumen to inform, may also be included.

  Although the amount of platelet composition injected will vary depending on the size of the heart and the area to be treated, at least one embodiment of the present invention will inject up to about 1100 μL of platelet composition into the heart wall at each injection site. To do. In another example, about 200 μL to 1000 μL of the platelet composition is delivered per injection site. In at least one other embodiment, about 100 μL to 10,000 μL of the platelet composition is delivered to the injection site. In another example, about 50 μL of the platelet composition is delivered per injection site. In one example, the clinician adjusts the injection volume, number of injection sites and injection site spacing, and total amount of platelet composition per heart while optimizing clinical benefit while minimizing clinical risk To do.

  The total infusion volume per heart may be dose-dependent based on the size of the heart, the size of the heart wall injury area, the desired range of tissue structure reinforcement, and / or the size of angiogenesis needed. good. In at least one embodiment, the total amount of platelet composition injected into the heart wall is an amount that can be accepted by the tissue at a reasonable number of injection sites. In another embodiment, the total volume of composition to be injected is 15000 μL (15 mL) or less.

  The number of injection sites per heart can be based on the size and shape of the injury area, the location where injection is desired, and the distance separating the injection sites. In at least one embodiment, the number of injection sites can range from 5 to 25 sites. The distance separating the injection sites will vary based on the desired amount of platelet composition to be injected at each injection site, the desired total amount to be injected, and the condition of the injured tissue. In at least one embodiment, the distance between injection sites is about 2 cm, and in at least one other embodiment, the distance between injection sites is 1 cm. In yet another example, the separation distance at the injection site can range between about 50 mm and about 2 cm. In another example, the distance between injection sites can range from 0.5 cm to 2.5 cm. In another example, the distance between the injection sites is greater than 2.5 cm. Instead of as a separate single injection, the injection can be continuous or intermittent infusion along the needle insertion path.

  In one embodiment of the invention, the platelet composition is injected into heart tissue in a pattern that promotes blood vessel formation. One exemplary pattern is a linear pattern that connects two target regions of tissue so that angiogenesis is stimulated along the linear pattern. In another embodiment, the pattern is branched. In particular, angiogenesis involves the formation of thick conduits.

  FIG. 11 shows a region of injured heart tissue after multiple injection of the platelet composition of the present invention. The composition is injected into the injured myocardium midway between the epicardial surface 404 and the endocardial surface 406 along the plane 402 of the ventricle or chamber. The composition is injected into multiple injection sites 410, 420, 430, and 450, which will result in infusion solution diffusion a few centimeters from the injection site. The composition to be injected diffuses so that the multiple injections are about 2 cm apart, the composition forms an overlapping field of structural support material. For example, composition 412 is injected into injection site 410 and diffuses as represented in FIG. Further, the composition 422 is injected into the injection site 420, diffuses, and mixes with the composition 412. This is repeated at the injection sites 430, 440 and 450 so that the compositions 412, 422, 432, 442 and 452 form successive overlapping fields of structural support material. In this example, compositions 412, 422, 432, 442 and 452 are the same composition, in a non-limiting example, autologous platelet gelation. In another example, one or more compositions can be injected at the treatment site.

  The location of delivery can vary based on the size and shape of the heart tissue injury area and the desired range of structural reinforcement of the tissue. In at least one embodiment of the present invention, platelet compositions and / or cell preparations are individually delivered only to damaged heart tissue, while in other embodiments the peri-injured zone around the damaged area is treated and at least other In one embodiment, the compositions and / or cells are delivered only to healthy tissue adjacent to the damaged area. In other examples, the composition and / or cells may be delivered to any combination of damaged heart tissue, surrounding injury zone tissue, and healthy tissue.

  The timing of platelet composition and cell product delivery in relation to the disorder event will be based on the severity of the injury, the extent of the injury, the patient's condition, and any tissue remodeling progress. In at least one embodiment, the platelet composition is delivered within 1 to 8 hours after a disorder event such as MI, for example within 1 to 8 hours after ischemia-reperfusion (immediately after reperfusion in a catheterization laboratory setting) To do. In another example, the platelet composition is delivered to the heart wall within 1 hour of the disorder event. In another example, the platelet composition is infused three to four days after injury (after the patient's clinical stability making it safe for the patient to receive a separate treatment). In at least one embodiment, the platelet composition is delivered more than a week after injury, including up to months or years after injury. Other dates and times for injecting the platelet composition into the heart tissue also include prior to any disorder event, and immediately upon discovery of the area of the disordered heart tissue (preventing additional remodeling in older disorders in advance) To be predicted). In another embodiment of the invention, the platelet composition can be injected into the heart tissue many years after the disorder event. In another example, the platelet composition is infused into the heart tissue about 1 hour to about 2 years after the disorder event. In another example, the platelet composition is infused into the heart tissue about 6 hours to about 1 year after the failure event. In another example, the platelet composition is infused into the heart tissue about 12 hours to about 9 months after the failure event. In another example, the platelet composition is infused into the heart tissue about 24 hours to about 6 months after the failure event. In another example, the platelet composition is infused into the heart tissue about 48 hours to about 3 months after the disorder event. In another example, the platelet composition is infused into the heart tissue by up to 10 years after the failure event.

  The timing of cell product delivery relative to platelet composition infusions and damaging events will be based on the patient and clinical scenario. Important factors include the presence of injury, severity and extent, patient condition, progression of any tissue remodeling and progression of neovascularization. Examples of the invention include promoting neovascularization in damaged tissue prior to administration of cell therapy. The time required for neovascularization of the damaged tissue will vary from patient to patient and the determination will be based on factors including, but not limited to, the size of the damaged tissue and the time since injury. In one example, the cell preparation is delivered to the treatment site up to 10 years after the disorder event. In another embodiment of the invention, the cell preparation is delivered to the treatment site between about 1 hour and 1 year after administration of the platelet composition. In another example, the cell preparation is administered between about 24 hours and about 9 months after the platelet composition. In another example, the cell preparation is administered between about 1 week and about 6 months after the platelet composition. Other dates and times for injecting cell preparations into heart tissue are intended to be the same as before any damaging event, immediately to find the area of damaged heart tissue, or almost at the same time as the platelet composition Is done. In another embodiment of the invention, the cell preparation can be injected into the heart tissue many years after the disorder event.

  The invention, its compositions, methods and methods, in which other damaged tissues in addition to the aforementioned right to use for platelets will benefit from delivery of treatments that promote neovascularization and regeneration to damaged heart tissue in addition The scheme is obvious to those skilled in the art. Examples of such tissues include wounds, gastrointestinal tissues, kidneys, liver, skin, and neural tissues such as brain, spinal cord and nerves, but are not limited to ischemic tissues in organs or sites. Including.

  Experiments were conducted in laboratory conditions to test the method and instrument of the present invention disclosed herein. These include in vitro experiments (described in Examples 1 and 2), in vivo studies conducted in healthy pig tissues (Examples 3 and 4), and in vivo studies conducted in damaged sheep tissues (Example 5). Including.

Example No. 1
Various combinations of components for autologous platelet gel (APG) were tested in vitro using human blood, pig blood and sheep blood. One composition consists of 60 mL whole blood (52.5 ml whole blood + 7.5 mL anticoagulant [ACD-A with citrate, sodium citrate and dextrose, anticoagulant dextrose citrate solution A]) Extraction of platelet plasma was required. This PRP was mixed with bovine thrombin (1000 U / ml in 10% CaCl2) at approximately 10: 1 (vol: vol) so that mixing occurred only in the target tissue. This is a composition tested in vivo as described below.

Example No. 2
The ability of fibrinogen to affect gelation and / or the physical properties of autologous platelet gel (APG) were tested directly in vitro. PRP and platelet poor plasma (PPP) were pre-made from fresh sheep blood using a Medtronic Magellan® platelet separator. Autologous fibrinogen was further extracted from the resulting PPP using the ethanol precipitation method. Alternative methods such as cryoprecipitation can be used for fibrinogen separation. Precipitated fibrinogen was resuspended in PRP to produce enhanced autologous fibrinogen PRP (AFFPRP). Made from the same animal (1) conventional APG made from 10: 1 ratio cattle thrombin PRP + 1000U / ml and (2) made from cattle thrombin AFFPRP + 1000U / ml in 10: 1 ratio Two preparations of enhanced fibrinogen APG were compared. Enhanced fibrinogen APG purified from the same animal was noticeably harder / stronger than conventional APG. This demonstrates the usefulness of serfibrinogen to augment the mechanical properties of APG without reducing the gelation rate.

Example No. 3
It has been successfully demonstrated that intramural transport of autologous platelet gel (APG) as two separate compositions (self-PRP and bovine thrombin) that contact and coagulate in tissue can be safely achieved in vivo. It was.

  Model and approach: A healthy pig model was used to test the safety and effectiveness of delivery. 180 ml of non-heparinized blood was obtained and used to make PRP18cc using a Medtronic Magellan® autologous platelet separator on the day of treatment. The animals were then heparinized for an active clotting time (ACT) ranging from 250-300. A median sternotomy provided an access path to the epicardial surface of the heart.

  Injection: Three injection schemes were tested. Method 1 is a 27 gauge syringe that delivers PRP alone. Method 2 is an 18-gauge stainless needle and two independent proximal syringes (12 mL and 1 mL in size) containing a 2 lumen tilt catheter with a luer lock into the needle (each ID [0.0085 inch ID)]. The injector was operated using a single-handed collection tube that ensured simultaneous injection of the two compositions at the desired ratio (about 11: 1 in this example). This was used to inject autologous PRP and bovine thrombin. Method 3 is a suction injector combined with a suction head (located on the epicardial surface of the heart) equipped with two needle injectors. The suction member is driven by a suction pump that achieves local stabilization of the heart beat. In addition, the heart wall is pulled into the suction cup so that the needle (entering tissue parallel to the plane of the chamber) can be delivered to a controllable depth. As shown in FIG. 10, the needle is driven by two separate injectors and is also secured to the one-handed injection collection tube. 12 mL and 1 mL syringes were used to ensure delivery of the desired ratio of autologous PRP and bovine thrombin (11: 1 in this example).

  Small multiple injections (200-400 μl / each) are performed by epicardial surgical approach. To inject using methods 1 and 2 above, the infusion was made perpendicular to the target myocardium and a “depth anchoring device” was used to ensure that the infusion was at the desired depth. The target depth was 5 mm in the left ventricle and 3 mm in the right ventricle. The depth fixing device is composed of a C-shaped member having a central hole through which the injection needle passes. Side screws (the depth fixator lumen size narrowed as it was screwed) were used to stop the depth fixator along its outside at the desired location along the length of the needle. As the needle is gently advanced into the target myocardium by applying force, the needle reaches the level of the depth fixation device and does not advance beyond it. In this way, this method ensures a fixed depth of needle penetration into the tissue, and intrawall injection occurs if the wall thickness is known or can be estimated. It is certain that it is.

  For all infusions in this study, MedtronicStarfish® (available from Minneapolis, Minnesota, Medtronic, Inc.) was used as a stabilization procedure for heart beats. Target tissues: Injections were made into the left ventricle (LV, bottom, middle position, apex) and right ventricle (RV, bottom, middle position, apex). Injection into LV was targeted to a depth of 5mm. Injection into the RV was targeted at a depth of 3 mm.

Composition: Different injections were tested.
1) Autologous PRP only-to determine whether clot formation occurs in the absence of exogenous thrombin
2) Self-PRP + cattle thrombin
3) Each of the above injections was performed with or without the addition of toluidine blue pigment to the self-PRP. This was to test the usefulness and effectiveness of the tracking dye for experimental purposes.
4) Saline control

  Results: The hemostatic effect after APG injection was outstanding. In particular, it was possible to inject a large amount of APG up to 1000 μl / each into the left ventricle in healthy pig myocardium, which was clinically safe. No adverse events were observed in up to 3 days follow-up. In particular, it was possible to inject a maximum of 200 μl / each of APG into the right ventricle in healthy pig myocardium, which was clinically safe. No adverse events were observed at follow-up times greater than 2 hours.

  The 23 infusions were well tolerated during the infusion or for 1 hour after the last infusion without arrhythmia, hypoxemia, or any clinically dangerous signs. After death, no thrombus or thromboembolic sequela was found. All 23 injections were successful and the injection site was examined during autopsy.

  Furthermore, we demonstrated that APG injection into the myocardium has a protective effect against arrhythmias. In this pig model, a 5600 μl infusion in a left ventricular split infusion is relatively resistant to lethal arrhythmias caused by an intravascular dose of potassium chloride (KCl). Instead of causing the expected fibrillation in 10-15 seconds with the standard dose of KCl, no arrhythmia was observed over 1.5 minutes. A second dose of KCl was required before any arrhythmia developed.

  Platelet gel can be formed from PRP alone without the addition of exogenous thrombin. The platelet-rich plasma injected into the myocardium surprisingly gels in situ (without thrombin) by itself. The inventor has formulated an unconstrained hypothesis that tissue thrombin may be present in an amount sufficient to trigger this gelation reaction. Therefore, when injected alone into the myocardium in vivo, PRP may be used to create APG in the tissue.

Example No. 4
Platelet rich plasma can be detected in tissues by adding toluidine blue pigment to PRP. This dye does not change the characteristics of PRP gelation (gelation rate, extent of gelation, hardness of the resulting gel) clearly in combination with thrombin.

The pattern of APG distribution by injection into the myocardium was evaluated in vivo. In 3 pigs, injections of APG labeled with toluidine blue demonstrated that each injection resulted in a distribution of APG in all directions within the tissue. The most important diffusion is along the plane of the ventricle. APG travels up to 1.5 cm radially in the plane of the ventricle. Depending on the injection, APG was detected more than 1.5 cm away from the injection site. It is likely that APG will migrate during the gelation process until sufficient gelation has occurred that prevents further diffusion of the material in the tissue.
Example No. 5

  The acute effect of APG injection into ischemic myocardium was investigated in a sheep anterior wall infarction model. In this model, myocardial infarction results in deleterious structural and functional changes that occur within minutes of the injury. Early features of remodeling include ventricular relaxation, wall thinning, ataxia and often movement disorders. Over time, these changes will progress as remodeling continues. It was determined that early post-infarction interventions could interfere with the remodeling process by supplying APG to the damaged myocardium. In addition, such APG treatment resulted in a significant increase in neovascularization of heart tissue embolizing over control embolic animals that did not receive APG.

  When performed on infarcted or non-infarcted tissues, the injection was safe and the tolerability was good. The experiment also showed that it can be safely implemented as early as 1 hour after MI. Controlled infusion was possible with or without a cardiac stabilizer and infusion was possible without external cardiac pacing. Injections were made at right angles and obliquely to the myocardial surface at intervals of 0.5 to 2.5 cm. The total injection volume was tested to be safe, even as high as 15.0 ml per heart and individual injections as high as 1100 μl per injection site.

  In 13 sheep that received APG 1 hour after infarction and followed up for 2 weeks, APG received infarct plus APG from 25-30% of existing control animals that received infarct arrhythmia-related post-infarction mortality Of 8%.

  Remodeling was rapidly prevented after infarction and APG injection. In this 13 sheep study, heart morphology and function were evaluated at different time points before and after APG injection. APG injection 1 hour after MI resulted in a markedly thickened heart wall and correction of post-MI movement disorders after injection. If post-MI remodeling was prevented partially or wholly for existing animals that had undergone infarction without APG injection, this effect was being triggered at 2 weeks of follow-up.

In this anterior wall infarct model, the ventricle expanded to 152.4% of the pre-infarction diastolic volume within minutes of infarct formation. The systolic ejection fraction (EF) also dropped dramatically to 62.1% of baseline after infarct formation. APG was injected into the damaged myocardium 1 hour after infarction in 5 animals. Treated hearts received between 10 and 13.6 cc of APG each by dispensing delivered into the myocardium. This treatment reduced the expected post-infarct diastolic volume increase from 152.4% to 108.6% of the pre-infarction volume. This demonstrates the substantial effect of APG that prevents the expected post-MI chamber expansion, which is one of the key metrics for remodeling. In this study, APG delivery resulted in an EF that was 111.1% of the pre-MI level, in one animal that also had post-MI EF efficacy, as the previous MI level recovered from 62.1% to 70.3%. It was. That is, in this animal, EF was 45% at baseline, 35% immediately after infarction, and 50% after administration of APG. This demonstrates that APG medication after cardiac injury can restore the partial or complete adverse acute effects of infarct formation on EF and, in some conditions, can extend EF to pre-infarction levels. To do.

  APG was surprisingly associated with neovascularization in target ischemic tissues in 12 sheep that received APG 1 hour after infarction and 8 weeks later. This effect was not expected because the target tissue is clearly ischemic and does not grow to generate a functional structure, but has a poor environment for cell survival. In 12 of the 12 animals, many small blood vessels were observed in the APG-treated infarct area at 8 weeks (FIGS. 16 and 17). Red blood cells observed in blood vessels are rich in perfusion suggestions that can be considered functional blood vessels that can provide a fresh blood supply to the surrounding tissue. Such blood vessels are usually not observed in animals suffering from infarct formation without APG injection.

  Experiments have shown that there are variations in the mechanical properties of APG between animals (and possibly between patients) and in the coagulation rate of APG. An improved method of anterior pituitary gonadotropin clotting rate / intensity inclusion using high-dose bovine thrombin at 1000 U / ml makes anterior pituitary gonadotropins, and uses to do anterior pituitary gonadotropins (~ 0 ° C) Thrombin was cooled. In addition, clotting rate / intensity can be improved by enhancing self-PRP with high concentrations of fibrinogen (eg, self-fibrinogen pre-formed with ethanol extraction or refrigeration regulator). Similarly, post-injection clotting rate / strength can be improved by very careful handling of pre-injection PRP to ensure minimal deactivation.

  Several methods have been clarified to enhance the retention of infusate in the target tissue and address possible leak / bleeded blood retrograde problems. These methods include using a high dose of bovine thrombin 1000 U / ml to make APG. A stirrer mechanism can be used in the replenishment chamber to prevent PRP delivery and / or PRP precipitation or degradation, thereby ensuring homogeneous PRP delivery to the target tissue and improved coagulation Will be promoted. Allow the needle to pause 5-10 seconds at the injection site after the infusion is delivered, use bevel to lengthen the injection path in the tissue, and local stability of the injection site with respect to needle insertion including. Each of these methods was tested in the previous examples.

  Making APG using chilled (˜0 ° C.) thrombin also increases the retention of the infusate in the target tissue. For embodiments using chilled thrombin, a refrigerated / cooled chamber in the thrombin delivery and / or refill chamber can be used.

  Appropriate needle placement and intraoperative ECHO (ultrasound cardiography) injected composition can be visualized that can be used to confirm retention. ECHO can be used as a separate device or can be included in a delivery system (eg, similar to intravascular ultrasound [IVUS]).

  Unintentional perforation of the heart chamber and / or delivery to room blood (or blood vessels) can be avoided by using a diagnostic imaging guidance device such as that provided by ECHO or IVUS during infusion. In addition, it has been found that direct pericardial membrane injection into the heart apex should be avoided to avoid chamber puncture. Instead, bevel injection should be used to access the apical tissue. Similarly, the device can be used to notify the operator when the delivery portion of the delivery device is in an undesired location, such as a ventricle or coronary vessel. Such devices include, but are not limited to, at least one sensor for the delivery system to represent backflow blood under pressure that emits a unique signal when its target tissue is in a location such as a blood cavity, Includes pressure sensors, color detectors, oxygen sensors, carbon dioxide sensors, or lumens. If the user is warned, the device can be repositioned prior to delivering the composition.

  These experiments showed that the method disclosed herein can be used to restore infarct left ventricular wall thickness to pre-MI (or higher) levels immediately after injection. This favorable effect lasts up to a week (in a reproducible manner). Similarly, this method allows the left ventricular systolic ejection fraction (EF) to be restored to the pre-myocardial infarction level immediately after injection. In addition, the individually disclosed therapies can improve post-MI cardiac mechanics and function by providing a motor impairment portion of left ventricular tissue akinetic properties.

  The present invention injects a substance that promotes neovascularization with or without a composition that structurally strengthens the tissue, and injects a cell preparation into the newly revascularized tissue to promote regeneration. Discloses a method of treating damaged heart tissue. As is apparent from FIG. 8, the method typically identifies and / or images the ischemic region of the heart tissue where treatment is desired 101, the desired effect (neovascularization with or without structural enhancement of the tissue). To determine the appropriate material (biological therapy with or without structural support) to be injected into the myocardium to achieve this, and select the appropriate device to inject the material into the heart tissue 102, Access 103 includes delivering 104 the substance and delivery device to the desired treatment location, injecting substance 105 into the heart tissue, and retrieving the device 106. Methods and equipment for injecting the composition (substance / infusion solution), composition, and delivery processes have been discussed herein.

  Further, as seen in FIG. 15, the system of the present invention identifies the injury region and treatment site of the heart tissue, provides access to the treatment site by the delivery device, and accesses one or more locations of the treatment site in the heart tissue. Including injection of the composition and recovery of the delivery device from the patient.

  Unless otherwise stated, all numbers representing the amounts of ingredients used in the specification and claims, properties such as molecular weight, reaction conditions, etc. are amended with the term “about” in all examples It should be. Accordingly, unless stated to the contrary, the numerical parameters set forth in the specification and claims are approximations that vary based on the desired properties obtained by the present invention. Each numerical parameter should be interpreted at least by taking into account the number of significant figures published and applying the usual rounding method, not at least as an attempt to limit the application of the doctrine to the scope of the claims. Despite the approximate numerical values and parameters described in the broad scope of the invention, the numerical values set forth in the specific examples are reported as accurately as possible. Every numerical value, however, necessarily contains certain errors resulting from the deviations inherent in each test measurement.

  Unless otherwise specified herein or otherwise clearly contradicted by context, the terms used in the context of describing the present invention, “a”, “its” and similar referents, include both the singular and the plural. Should be interpreted as covering. The description of a range of values herein is merely intended to serve as a convenient way to individually reference each separate value that falls within the range. Unless otherwise stated, each single value is stated in the specification as if it were individually listed. All methods described herein can be performed in any suitable order unless otherwise indicated herein or otherwise clearly contradicted by context. The use of any and all examples listed individually, or exemplary language (eg, “etc.”) merely makes the invention clearer or the scope of the claimed invention. It is not limited. The language herein should not be construed as indicating any non-patentable element essential to the practice of the invention.

  The groupings of alternative elements or the embodiments of the invention disclosed herein should not be construed as limiting. Each group of components may refer to and claim patents individually or in any combination with other components of the group or other components found herein. It is anticipated that one or more members of the group may be included or excluded from the group for convenience and / or for patent qualification reasons. In the event of any such inclusion or deletion, this document will satisfy all the Markush group written specifications used in the appended claims and include groups so modified It is considered.

  Certain specific embodiments of the present invention are described herein, including the best mode known to the inventors for carrying out the invention. Of course, variations on these described embodiments will become apparent to those of ordinary skill in the art upon reading the foregoing description. The inventor expects those skilled in the art to employ such variations as appropriate, or is intended to practice the invention in ways other than those specifically described herein. Accordingly, this invention includes all modifications and equivalents of the subject matter recited in the claims appended hereto as permitted by applicable law. Moreover, any combination of the above-described elements in all possible variations thereof is encompassed by the invention unless otherwise indicated herein or otherwise clearly contradicted by context.

  In addition, numerous references to patents and publications are cited throughout this specification. Each of the references cited above, and publications, are deemed to be incorporated herein by reference in their entirety.

  In closing, it should be understood that the embodiments of the present invention disclosed herein are illustrative of the principles of the present invention. Other modifications that can be used are within the scope of the invention. Thus, by way of example but not limitation, alternative configurations of the present invention may be utilized in accordance with what is taught herein. Accordingly, the present invention is not limited to that precisely as shown and described.

1 is a diagram of a normal, healthy heart. It is a figure of the heart with an injured myocardial region. FIG. 3 is an enlarged view of the damaged myocardium shown in FIG. 2 represents a cross-sectional view of the heart shown in FIG. Fig. 4 represents a cross-section of a heart showing a region of cardiac tissue that has been damaged and remodeled by a left ventricular wall. Eligible damaged heart tissue has different wall thicknesses and geometries. An example is shown with a slightly thinner and relaxed (aneurysm) wall. 1 represents a needle used to deliver a composition to the heart wall according to an embodiment of the present invention. 1 represents a needle used to deliver a composition to the heart wall according to an embodiment of the present invention. FIG. 5 is a block diagram illustrating steps for treating heart tissue in accordance with the teachings of the present invention. In accordance with one embodiment of the present invention, the delivery of the composition into the heart is schematically represented. FIG. 4 schematically represents a detailed view of composition delivery into heart tissue, in accordance with another embodiment of the present invention. Figure 7 schematically represents the transfer of a composition within myocardial tissue after delivery, according to an embodiment of the present invention. Fig. 4 schematically represents an epicardial delivery method for delivering a composition to heart tissue in accordance with the teachings of the present invention. FIGS. 13A-B schematically represent an epicardial delivery method for delivering a composition to heart tissue in accordance with the teachings of the present invention. FIG. 13A represents antegrade endocardial access through the venous system. FIGS. 13A-B schematically represent an epicardial delivery method for delivering a composition to heart tissue in accordance with the teachings of the present invention. FIG. 13B represents retrograde endocardial access through the arterial system. 14A-B schematically represent a transvascular delivery method for delivering a composition to heart tissue in accordance with the teachings of the present invention. FIG. 14A represents the vein access method. 14A-B schematically represent a transvascular delivery method for delivering a composition to heart tissue in accordance with the teachings of the present invention. FIG. 14B represents arterial access through the coronary artery. 2 shows a flowchart of the method of the present invention. In accordance with the teachings of the present invention, represents a photomicrograph of infarcted myocardium 8 weeks after infusion of autologous platelet gel (ratio of platelet rich plasma and bovine thrombin 10: 1) 1 hour after infarction. Many blood vessels (arrow A) are observed in the infarcted tissue region (arrow C). These blood vessels carry red blood cells (arrow B). In accordance with the teachings of the present invention, a representation of an intensified micrograph of infarcted myocardium 8 weeks after infusion of autologous platelet gel (platelet rich plasma and bovine thrombin in a 10: 1 ratio) 1 hour after infarct formation. Many blood vessels (arrow A) are observed in the infarcted tissue region (arrow C). These blood vessels carry red blood cells (arrow B).

Claims (42)

A method of treating heart tissue,
Providing a platelet composition within a treatment site in the heart tissue, the platelet composition promoting neovascularization of the heart tissue;
Injecting a cell preparation into the revascularized heart tissue, and treating the heart tissue.   2. The method of claim 1, wherein the heart tissue is damaged tissue or healthy tissue.   2. The method of claim 1, wherein the method causes regeneration of the heart tissue.   2. The method of claim 1, wherein the platelet composition is selected from the group consisting of platelet gel, platelet rich plasma and platelet poor plasma.   2. The method of claim 1, wherein the platelet composition is autologous.   5. The method of claim 4, wherein the platelet gel is formed from platelet poor plasma or platelet rich plasma and an activator.   7. The method of claim 6, wherein the activator is thrombin.   8. The method of claim 7, wherein the platelet gel comprises platelet rich plasma or platelet poor plasma and thrombin in a ratio between about 5: 1 and about 25: 1.   5. The method of claim 4, wherein the platelet composition comprises platelet rich plasma without an exogenous source of thrombin.   The platelet composition further comprises a structural material selected from the group consisting of collagen, biocompatible polymer, alginate, synthetic / natural compound, fibrinogen, silk elastin polymer, hydrogel and dental complex material. The method of claim 1.   The platelet composition is a pharmaceutically active compound, corticosteroid, growth factor, enzyme, DNA, ribonucleic acid, siRNA, virus, protein, lipid, polymer, hyaluronic acid, antibody, antibiotic, anti-inflammatory action 2. The method of claim 1, further comprising a bioactive agent selected from the group consisting of drugs, antisense nucleotides and transforming nucleic acids, and combinations thereof.   2. The method of claim 1, wherein the platelet composition further comprises a contrast agent.   2. The method of claim 1, wherein the cell preparation comprises cells of one or more cell types selected from the group consisting of somatic, germ cells, fetuses, embryonic, postnatal cells and mature cells.   14. The method of claim 13, wherein the cell preparation comprises cells isolated from one or more tissue types selected from the group consisting of fat, brain, muscle, endothelium, blood, bone marrow, heart, testis and ovary. .   14. The method according to claim 13, wherein the cells are autologous.   14. The method of claim 13, wherein the cell has been modified prior to transplantation.   27. The method of claim 26, wherein said modification comprises genetic manipulation of said cell to secrete one or more biologically active molecules.   28. The method of claim 27, wherein the biologically active molecule is a growth factor.   2. The method of claim 1, wherein the cell preparation further comprises a bioactive agent.   2. The method of claim 1, wherein the cell preparation further comprises a platelet composition.   2. The method of claim 1, wherein the platelet composition is provided to the damaged heart tissue between about 1 hour and about 1 year after the heart tissue is damaged.   2. The method of claim 1, wherein the platelet composition and any of the cell preparations described above are injected into about 1 to 20 sites.   24. The method of claim 22, wherein the infusion is provided over time.   24. The method of claim 22, wherein the infusion comprises a total infusion volume of up to about 15 mL.   24. The method of claim 22, wherein the injection comprises an injection volume of up to about 1100 microliters per injection.   2. The method of claim 1, wherein the cellular preparation is provided to the damaged heart tissue between about 1 hour and about 1 year after the heart tissue is damaged.   2. The method of claim 1, wherein said cell preparation is provided to said damaged heart tissue between about 1 hour and about 1 year after administration of said platelet composition.   28. The method of claim 27, wherein the cell preparation is provided to the heart tissue after neovascularization is initiated in the damaged heart tissue.   2. The method of claim 1, wherein the platelet composition and the cell preparation are provided to the heart tissue almost simultaneously.   2. The method of claim 1, wherein in the cardiac tissue, the treatment site is selected from the group consisting of subendocardial, subepicardial and intramyocardial sites.   2. The method of claim 1, further comprising a delivery device for delivering the platelet composition or cell preparation to the damaged heart tissue.   32. The method of claim 31, wherein the delivery device is an infusion catheter selected from the group consisting of an endocardial infusion catheter, a transvascular infusion catheter, and an epicardial infusion catheter.   The method of claim 1, wherein the treatment site is selected from the group consisting of a damaged area, a damaged peripheral area, and healthy tissue surrounding the damaged area.   2. The method of claim 1, wherein the platelet composition and the cell preparation are injected into the same treatment site.   2. The method of claim 1, wherein the platelet composition and the cell preparation are injected at different treatment sites.   36. The method of claim 35, wherein the cell preparation is injected adjacent to the injection site of the platelet gel composition. A method of treating heart tissue,
Providing a platelet composition to a treatment site in the heart tissue;
Replenishing cells forming blood vessels from the tissue or blood to the treatment site, wherein the heart tissue is revascularized by the cells forming the blood vessels. how to.   38. The method of claim 37, wherein the platelet composition further comprises molecules that attract angiogenic cells to a treatment site.   38. The method of claim 37, wherein the molecules are selected from the group consisting of growth factors, growth factor receptors and chemoattractants. A system for regenerating heart tissue,
Platelet composition,
A cell preparation, and at least one delivery device for introducing the platelet composition into the heart tissue,
A system for regenerating heart tissue, wherein the platelet composition causes neovascularization of the heart tissue, and regeneration of the heart tissue by the cell preparation is promoted. A method of treating heart tissue,
Providing a platelet composition within a treatment site in the heart tissue, and injecting a cell preparation into the heart tissue.   42. The method of claim 41, wherein said platelet composition and said cell preparation are provided almost simultaneously.

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