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Posts Tagged ‘Thymosin’


Heart Vasculature – Regeneration and Protection of Coronary Artery Endothelium and Smooth Muscle: A Concept-based Pharmacological Therapy of a Combination Three Drug Regimen including THYMOSIN 

Author & Curator: Aviva Lev-Ari, PhD, RN

 

ABSTRACT

A concept-based original pharmacological therapy was developed for the research results presented in Cell by Wu, Fujiwara, Cibulsky et al. (2006), Moretti, Caron, Nakano, et al. (2006) and for the research results in Nature by Smart, Risebro, Melville, et al. (2007). We propose the following concept-based original pharmacological therapy design for Preoperative and Postoperative management of cardiac injury to heart tissue, smooth muscle, to aorta and coronary artery disease. This is a treatment for Coronary Vasculogenesis, Anti-hypertention (short-acting), Vascular Anti-inflammation (vasculitis), Neovascularization of ischemic tissue and release of adult epicardium from a quiescent state while restoring its pluripotency.

VIEW VIDEO

What are Induced Pluripotent Stem Cells? (iPS Cells)

 http://www.youtube.com/watch?v=i-QSurQWZo0

Lasker Lecture: Dr. Shinya Yamanaka, 2 of 3:

Induced Pluripotent Stem Cells? (iPS Cells)

http://www.youtube.com/watch?v=DQNoyDwCPzM

Multipotent Embryonic Isl1^+ Progenitor Cells Lead to Cardiac, Smooth Muscle, and Endothelial Cell Diversification

Alessandra A MorettiLeslie L CaronAtsushi A NakanoJason T JT LamAlexandra A Bernshausen,Yinhong Y ChenYibing Y QyangLei L BuMika M SasakiSilvia S Martin-PuigYunfu Y SunSylvia M SM EvansKarl-Ludwig KL LaugwitzKenneth R KR Chien
Cell 127(6):15 (2006), PMID 17123592

Cardiogenesis requires the generation of endothelial, cardiac, and smooth muscle cells, thought to arise from distinct embryonic precursors. We use genetic fate-mapping studies to document that isl1^+ precursors from the second heart field can generate each of these diverse cardiovascular cell types in vivo. Utilizing embryonic stem (ES) cells, we clonally amplified a cellular hierarchy of isl1^+ cardiovascular progenitors, which resemble the developmental precursors in the embryonic heart. The transcriptional signature of isl1^+/Nkx2.5^+/flk1^+ defines a multipotent cardiovascular progenitor, which can give rise to cells of all three lineages. These studies document a developmental paradigm for cardiogenesis, where muscle and endothelial lineage diversification arises from a single cell-level decision of a multipotent isl1^+ cardiovascular progenitor cell (MICP). The discovery of ES cell-derived MICPs suggests a strategy for cardiovascular tissue regeneration via their isolation, renewal, and directed differentiation into specific mature cardiac, pacemaker, smooth muscle, and endothelial cell types.

http://pubget.com/paper/17123592/Multipotent_Embryonic_Isl1___Progenitor_Cells_Lead_to_Cardiac__Smooth_Muscle__and_Endothelial_Cell_Diversification

 

Thymosin beta 4 (Tβ4)

is a highly conserved, 43-amino acid acidic peptide (pI 4.6) that was first isolated from bovine thymus tissue over 25 years ago. It is present in most tissues and cell lines and is found in high concentrations in blood platelets, neutrophils, macrophages, and other lymphoid tissues. Tβ4 has numerous physiological functions, the most prominent of which being the regulation of actin polymerization in mammalian nucleated cells and with subsequent effects on actin cytoskeletal organization, necessary for cell motility, organogenesis, and other important cellular events.

Recently,

  • Tβ4 was shown to be expressed in the developing heart and found to stimulate migration of cardiomyocytes and endothelial cells, promote survival of cardiomyocytes (Nature, 2004), and most recently
  • to play an essential role in all key stages of cardiac vessel development: vasculogenesis, angiogenesis, and arteriogenesis (Nature 2006).
  • These results suggest that Tβ4 may have significant therapeutic potential in humans to protect myocardium and promote cardiomyocyte survival in the acute stages of ischemic heart disease.

RegeneRx Biopharmaceuticals, Inc. is developing Tβ4 for the treatment of patients with acute myocardial infarction (AMI). Such efforts presented will include the formulation, development, and manufacture of a suitable drug product for use in the clinic, the performance of nonclinical pharmacology and toxicology studies, and the implementation of a phase 1 clinical protocol to assess the safety, tolerability, and the pharmacokinetics of Tβ4 in healthy volunteers.

http://onlinelibrary.wiley.com/doi/10.1196/annals.1415.051/abstract;jsessionid=BB7CC897572B7DDB60370EA64A81FC3F.d01t03?deniedAccessCustomisedMessage=&userIsAuthenticated=false

EXPLORATIONS with THYMOSIN beta4 for INDUCING ADULT EPICARDIAL PROGENETOR MOBILIZATION AND NEOVASCULARIZATION is presented in

Resident-cell-based Therapy in Human Ischaemic Heart Disease: Evolution in the PROMISE of Thymosin beta4 for Cardiac Repair

https://pharmaceuticalintelligence.com/2012/04/30/93/

EXPLORATIONS with THYMOSIN beta4 for INDUCTION of ARTERIOGENESIS, Prevention and repair of damaged cardiac tissue post MI and other CVD related research projects are presented in

Arteriogenesis and Cardiac Repair: Two Biomaterials – Injectable Thymosin beta4 and Myocardial Matrix Hydrogel

https://pharmaceuticalintelligence.com/2013/02/27/arteriogenesis-and-cardiac-repair-two-biomaterials-injectable-thymosin-beta4-and-myocardial-matrix-hydrogel/

Recent research results with THYMOSIN beta4 in use for Cardiovascular Disease

appeared in 2010:

Annals of the New York Academy of Sciences, May 2010 Volume 1194 Pages ix–xi, 1–230

http://onlinelibrary.wiley.com/doi/10.1111/nyas.2010.1194.issue-1/issuetoc

appeared in 2012:

  • Thymosins in Health and Disease II: 3rd International Symposium on The Emerging Clinical Applications of Tymosin beta 4 in Cardiovascular Disease

Annals of the New York Academy of Sciences, October 2012 Volume 1270 Pages vii-ix, 1–121.

http://onlinelibrary.wiley.com/doi/10.1111/nyas.2012.1270.issue-1/issuetoc

Allan L. Goldstein, Enrico Garaci, Editors, Thymosins in Cardiovascular Disease, November 2012, Wiley-Blackwell (paperback)

http://www.wiley.com/WileyCDA/WileyTitle/productCd-1573319104.html?cid=RSS_WILEY2_LIFEMED

Selected for this article are the abstracts of the following research projects, all were presented at the 2nd International Symposium, May 2010:

Thymosin β4: structure, function, and biological properties supporting current and future clinical applications

Published studies have described a number of physiological properties and cellular functions of thymosin β4 (Tβ4), the major G-actin-sequestering molecule in mammalian cells. Those activities include the promotion of cell migration, blood vessel formation, cell survival, stem cell differentiation, the modulation of cytokines, chemokines, and specific proteases, the upregulation of matrix molecules and gene expression, and the downregulation of a major nuclear transcription factor. Such properties have provided the scientific rationale for a number of ongoing and planned dermal, corneal, cardiac clinical trials evaluating the tissue protective, regenerative and repair potential of Tβ4, and direction for future clinical applications in the treatment of diseases of the central nervous system, lung inflammatory disease, and sepsis. A special emphasis is placed on the development of Tβ4 in the treatment of patients with ST elevation myocardial infarction in combination with percutaneous coronary intervention, pp.179-189, May 2010.

  

Thymosin β4 and cardiac repair

Hypoxic heart disease is a predominant cause of disability and death worldwide. As adult mammals are incapable of cardiac repair after infarction, the discovery of effective methods to achieve myocardial and vascular regeneration is crucial. Efforts to use stem cells to repopulate damaged tissue are currently limited by technical considerations and restricted cell potential. We discovered that the small, secreted peptide thymosin β4 (Tβ4) could be sufficiently used to inhibit myocardial cell death, stimulate vessel growth, and activate endogenous cardiac progenitors by reminding the adult heart on its embryonic program in vivo. The initiation of epicardial thickening accompanied by increase of myocardial and epicardial progenitors with or without infarction indicate that the reactivation process is independent of injury. Our results demonstrate Tβ4 to be the first known molecule able to initiate simultaneous myocardial and vascular regeneration after systemic administration in vivo. Given our findings, the utility of Tβ4 to heal cardiac injury may hold promise and warrant further investigation, pp. 87-96, May 2010.

 

Thymosin β4 facilitates epicardial neovascularization of the injured adult heart

Ischemic heart disease complicated by coronary artery occlusion causes myocardial infarction (MI), which is the major cause of morbidity and mortality in humans

http://www.who.int/cardiovascular_diseases/resources/atlas/en/index.html

After MI the human heart has an impaired capacity to regenerate and, despite the high prevalence of cardiovascular disease worldwide, there is currently only limited insight into how to stimulate repair of the injured adult heart from its component parts. Efficient cardiac regeneration requires the replacement of lost cardiomyocytes, formation of new coronary blood vessels, and appropriate modulation of inflammation to prevent maladaptive remodeling, fibrosis/scarring, and consequent cardiac dysfunction. Here we show that thymosin β4 (Tβ4) promotes new vasculature in both the intact and injured mammalian heart. We demonstrate that limited EPDC-derived endothelial-restricted neovascularization constitutes suboptimal “endogenous repair,” following injury, which is significantly augmented by Tβ4 to increase and stabilize the vascular plexus via collateral vessel growth. As such, we identify Tβ4 as a facilitator of cardiac neovascularization and highlight adult EPDCs as resident progenitors which, when instructed by Tβ4, have the capacity to sustain the myocardium after ischemic damage, pp. 97-104, May 2010.

 

Thymosin β4: a key factor for protective effects of eEPCs in acute and chronic ischemia

Acute myocardial infarction is still one of the leading causes of death in the industrial nations. Even after successful revascularization, myocardial ischemia results in a loss of cardiomyocytes and scar formation. Embryonic EPCs (eEPCs), retroinfused into the ischemic region of the pig heart, provided rapid paracrine benefit to acute and chronic ischemia in a PI-3K/Akt-dependent manner. In a model of acute myocardial ischemia, infarct size and loss of regional myocardial function decreased after eEPC application, unless cell pre-treatment with thymosin β4 shRNA was performed. Thymosin ß4 peptide retroinfusion mimicked the eEPC-derived improvement of infarct size and myocardial function. In chronic ischemia (rabbit model), eEPCs retroinfused into the ischemic hindlimb enhanced capillary density, collateral growth, and perfusion. Therapeutic neovascularization was absent when thymosin ß4 shRNA was introduced into eEPCs before application. In conclusion, eEPCs are capable of acute and chronic ischemia protection in a thymosin ß4 dependent manner, pp. 105-111, May 2010.

Clinical Study Data of Thymosin beta 4 Presented

Published on October 3, 2009 at 5:10 AM

REGENERX BIOPHARMACEUTICALS, INC. (NYSE Amex:RGN) today reported on several clinical studies with Thymosin beta 4 (Tβ4) presented the Second International Symposium on Thymosins in Health and Disease, in Catania, Italy. The following are synopses of the presentations:

Myocardial Development of RGN-352 (Injectable Tβ4 Peptide)

David Crockford, RegeneRx’s vice president for clinical and regulatory affairs presented an overview of the biological properties that support Tβ4’s near term and long term clinical applications. Mr. Crockford noted that special emphasis is being placed on the development of RGN-352 for the systemic (injectable) treatment of patients with ST-elevation myocardial infarction (STEMI) in combination with percutaneous coronary intervention, the current standard of care in most western countries for this common type of heart attack. The goal with RGN-352 is to prevent or repair continued damage to cardiac tissue post-heart attack, when such tissue around the damaged site remains at risk.

Dr. Dennis Ruff, vice president and medical director of ICON, and principal investigator, presented the most current results on the Phase I safety study with RGN-352 entitled, “A Randomized, Double-blind, Placebo-controlled, Dose-response Phase I Study of the Safety and Tolerability of the Intravenous Administration of Thymosin Beta 4 and its Pharmacokinetics After Single and Multiple Doses in Healthy Volunteers.” Dr. Ruff discussed key aspects of the study and concluded with, “There were no dose limiting or serious adverse events throughout the dosing period. Synthetic Tβ4 administered intravenously up to 1260 mg, and for up to 14 days, appears to be well tolerated with low incidence of adverse events and no evidence of serious adverse events.”

http://www.news-medical.net/news/20091003/Clinical-study-data-of-Thymosin-beta-4-presented.aspx

RegeneRx Receives Notice of Allowance from Chinese Patent Office for Treatment and Prevention of Heart Disease

RegeneRx Receives Notice of Allowance from Chinese Patent Office for Treatment and Prevention of Heart Disease

February 7, 2013 — Rockville, MD

RegeneRx Biopharmaceuticals, Inc. (OTC Bulletin Board: RGRX) (“the Company” or “RegeneRx”) today announced that it has received a Notice of Allowance of a Chinese patent application for uses of Thymosin beta 4 (TB4) for treating, preventing, inhibiting or reducing heart tissue deterioration, injury or damage in a subject with heart failure disease. Claims also include uses for restoring heart tissue in those subjects. The patent will expire July 26, 2026 http://www.regenerx.com/wt/page/pr_1360265259

Theoretical treatment protocol differential between the Preoperative which may be between 3 to 6 month, and the Postoperative which may prolong to one year.

Proposal for Preoperative Treatment – Three drug combination involves

  • Drug # 1: Thymosin fraction 5 (a sublingual composition)
  • Drug # 2: Indomethacin (Nonsteroidal anti-inflammatory drugs (NSAID))
  • Drug # 3: Clevidipine (blood pressure lowering drug, (no effect on heart rate))

 

Proposal for Postoperative Treatment – Three drug combination consists of

  • Drug # 1: Thymosin fraction 5 (a sublingual composition)
  • Drug # 4: ACEI (Captopril (50mg))
  • Drug # 5: Beta Blocker and Diuretic (Metoprolol and hydrochlorothiazide (50 mg/25 mg)) Lopressor HCT

Unprecedented novel paradigm development in the scientific understanding of the origin of

  • (a) myocardial cells
  • (b) smooth muscle cells
  • (c) endothelial cells
  • (d) pace maker cells and
  • (e) heart vasculature: aorta, pulmonary artery and coronary arteries, occurred in 2006.

In a seminal article in Cell, “Developmental Origin of a Bipotential Myocardial and Smooth Muscle Cell Precursor in the Mammalian Heart” Wu, et al., (2006), described their discovery as follows:

“Despite recent advances in delineating the mechanisms involved in cardiogenesis, cellular lineage specification remains incompletely understood.” To explore the relationship between developmental fate and potential.” They “isolated a cardiac-specific Nkx2.5+ cell population from the developing mouse embryo. The majority of these cells differentiated into cardiomyocytes and conduction system cells. Some, surprisingly, adopted a smooth muscle fate. To address the clonal origin of these lineages, we isolated Nkx2.5+ cells from in vitro differentiated murine embryonic stem cells and found ~28% of these cells expressed c-kit. These c-kit+ cells possessed the capacity for long-term in vitro expansion and differentiation into both cardiomyocytes and smooth muscle cells from a single cell.” They “confirmed these findings by isolating c-kit+Nkx2.5+ cells from mouse embryos and demonstrated their capacity for bipotential differentiation in vivo. Taken together, these results support the existence of a common precursor for cardiovascular lineages in the mammalian heart.”

Another breakthrough article in Cell, “Multipotent Embryonic Isl1+ Progenitor Cells Lead to Cardiac, Smooth Muscle, and Endothelial Cell Diversification” Moretti, et al., (2006) described their discovery as follows:

“Cardiogenesis requires the generation of endothelial, cardiac, and smooth muscle cells, thought to arise from distinct embryonic precursors.” They “use genetic fate-mapping studies to document that isl1+ precursors from the second heart field can generate each of these diverse cardiovascular cell types in vivo. Utilizing embryonic stem (ES) cells”, they “clonally amplified a cellular hierarchy of isl1+ cardiovascular progenitors, which resemble the developmental precursors in the embryonic heart. The transcriptional signature of isl1+/Nkx2.5+/flk1+ defines a multipotent cardiovascular progenitor, which can give rise to cells of all three lineages. These studies document a developmental paradigm for cardiogenesis, where muscle and endothelial lineage diversification arises from a single cell-level decision of a multipotent isl1+ cardiovascular progenitor cell (MICP). The discovery of ES cell-derived MICPs suggests a strategy for cardiovascular tissue regeneration via their isolation, renewal, and directed differentiation into specific mature cardiac, pacemaker, smooth muscle, and endothelial cell types.” (Moretti, et al., 2006).

Third scientific breakthrough was reported in Nature on the roles that Thymosin beta4 play in

  • (a) coronary vessel development
  • (b) induction of adult epicardial cell migration
  • (c) cardiomyocyte survival by vascularization which is dependent on Thymosin beta4 and
  • (d) identification of the pro-angiogenic tetrapeptide AcSDKP which is produced by endoproteinase activity of Thymosin beta4 (Smart, et al., 2007).

That new level of understanding has the potential to generate new pharmaco therapies to upregulate biological processes that underlie the function of the various compartments of the cardiovascular system, as new scientific explanations became available in 2006.

We have developed a methodology for discovery of concept-based original pharmacological therapy designs for combination of several drug regimens. We carry out two types of research strategy. Methodology Strategy Type One: we develop an original pharmacological therapy design specialized in addressing medical problems identified in targeted follow up studies on mortality and morbidity of cardiovascular patients. Methodology Strategy Type One is implemented in Lev-Ari & Abourjaily (2006a, 2006b, 2006c). We designed a specialized pharmaco therapy for the research results presented in NEJM, on “Circulating Endothelial Progenitor Cells and Cardiovascular Outcomes” (Werner, Kosiol, Schiegl, et al., 2005a) and the editorial interpretation of these research results by Rosenzweig  (2005). We proposed the following concept-based original pharmacological therapy design for Endogenous Augmentation of circulating Endothelial Progenitor Cells for Reduction of Risk for Macrovascular Cardiac Events.

 

Proposal of Treatment – Three drug combination

  • Inhibition of ET-1, ETA and ETA-ETB (Bosentan)
  • Induction of NO production and stimulation of eNOS (Nebivolol)
  • Stimulation of PPAR-gamma (substitute to Rosiglitazone)

Our Methodology Strategy Type Two involves discovery of concept-based original pharmacological therapy design for combination of several drug regimens for underlying biological processes discovered in the pursuit of basic researchers conducted in wet lab experiments by vascular biologists and molecular cardiologists. Here, we developed a concept-based original pharmacological therapy for the research results presented in Cell by Wu, Fujiwara, Cibulsky et al. (2006), Moretti, Caron, Nakano, et al. (2006) and for the research results in Nature by Smart, Risebro, Melville, et al. (2007). We propose the following concept-based original pharmacological therapy design for Preoperative and Postoperative management of cardiac injury to heart tissue, smooth muscle and to aorta and coronary artery disease. This is a treatment for Coronary Vasculogenesis, Anti-hypertention (short-acting), Vascular Anti-inflammation (vasculitis), Neovascularization of ischemic tissue and release of adult epicardium from a quiescent state and restoring its pluripotency.

 

Proposal for Preoperative Treatment – Three drug combination

  • Drug # 1:

Thymosin fraction 5 (a sublingual composition)

  • Drug # 2:

Indomethacin (Nonsteroidal anti-inflammatory drugs (NSAID))

  • Drug # 3:

Clevidipine (blood pressure lowering drug, no effect on heart rate)

Proposal for Postoperative Treatment – Three drugs combination

  • Drug # 1:

Thymosin fraction 5 (a sublingual composition)

  • Drug # 4:

ACEI (Captopril (50mg))

  • Drug # 5:

HCTBeta Blocker and Diuretic (Metoprolol and hydrochlorothiazide (50 mg/25 mg)) Lopressor

 

Thymosin beta4 Induces Adult Epicardial Progenitor Mobilization and Neovascularization

 

Smart et al. (2007) implicate Thymosine beta4 (Tb4) with the following functions: (a) Tb4 in regulating all three key stages of cardiac vessel development: coronary vasculogenesis, angiogenesis and arteriogenesis – collateral growth; (b) identify the adult epicardium as a potential source of vascular progenitors which, when stimulated by Tb4, migrate and differentiate into smooth muscle and endothelial cells; (c) the ability of Tb4 to promote coronary vascularization both during development and in the adult, enhances cardiomyocyte survival and contributes significantly towards Tb4-induced cardioprotection.

The reaction in the scientific community to these investigative results was most favorable.

“These results are very exciting because most humans suffering from ischemic cardiac events, either acutely or chronically, do not develop the collateral vessel growth necessary to preserve and restore heart tissue. If, in humans, we see the same effects as seen in mice, TB4 would be the first drug to prevent loss of (heart) muscle cells and restore blood flow in this manner and provide a new and much needed treatment modality for these patients,”

commented Deepak Srivastava, M.D., Professor and Director, Gladstone Institute of Cardiovascular Disease, University of California San Francisco, CA. Dr. Srivastava and his colleagues published the first paper on TB4’s effects on myocardial infarction in Nature in November 2004.

http://phx.corporate-ir.net/phoenix.zhtml?c=144396&p=irol-newsArticle&ID=932573&highlight=

VIEW VIDEO

http://www.youtube.com/watch?v=Vjj7LSuSMAo

 

Review of the Chemistry and the Mechanism of action supporting the process by which, N-acetyl-seryl-aspartyl-lysyl- proline (Ac-SDKP) stimulates endothelial cell differentiation from adult epicardium, is presented in

Resident-cell-based Therapy in Human Ischaemic Heart Disease: Evolution in the PROMISE of Thymosin beta4 for Cardiac Repair

https://pharmaceuticalintelligence.com/2012/04/30/93/

A Concept-based Pharmacologic Therapy of a Combined Three Drug Regimen for Regeneration and Protection of Coronary Artery Endothelium and Smooth Muscle.

This is a treatment for Coronary Vasculogenesis, Anti-hypertention (short-acting), Vascular Anti-inflammation (vasculitis), Neovascularization of ischemic tissue and release of adult epicardium from a quiescent state and restoring its pluripotency.

 

Preoperative Treatment – Three drugs

  • Drug # 1:
  • Thymosin fraction 5 (a sublingual composition)
  • Drug # 2:
  • Indomethacin (Nonsteroidal anti-inflammatory drugs (NSAID)) (25 mg PO bid)
  • Drug # 3:
  • Clevidipine (Blood pressure lowering drug, no effect on heart rate)

 

Postoperative Treatment – Three drugs

  • Drug # 1:
  • Thymosin fraction 5 (a sublingual composition)
  • Drug # 4:
  • ACEI (Captopril (50mg))
  • Drug # 5:
  • Beta Blocker and diuretic (Metoprolol and hydrochlorothiazide (50 mg/25 mg)) Lopressor HCT

Original Drug Therapy Combination Proposed

Drug # 1: Thymosin fraction 5

Drug # 2: Indomethacin

Drug # 3: Clevidipine

Drug # 1:

Sublingual compositions comprising Thymosin fraction 5

United States Patent:  6,733,791

http://www.pharmcast.com/Patents100/Yr2004/May2004/051104/6733791_Sublingual051104.htm

http://www.google.com/patents/US6733791

The compositions comprise a room temperature stable peptide or complex of peptides that may be administered in a dosage of between 0.0001 mg/ml or gm and 600 mg/ml or gm.

Thymosin beta4 is released from human blood platelets and attached by factor XIIIa (transglutaminase) to fibrin and collagen (Huff et al. 2002). They suggest that Thymosin beta4 cross-linking is mediated by factor XIIIa, a transglutaminase that is co-released from stimulated platelets. This provides a mechanism to increase the local concentration of Thymosin beta4 near sites of clots and tissue damage, where it may contribute to wound healing, angiogenesis and inflammatory responses (Al-Nedawi, et al., 2004). The beta-Thymosins constitute a family of highly conserved and extremely water-soluble 5 kDa polypeptides. Thymosin beta4 is the most abundant member; it is expressed in most cell types and is regarded as the main intracellular G-actin sequestering peptide. There is increasing evidence for extracellular functions of Thymosin beta4. For example, Thymosin beta4 increases the rate of attachment and spreading of endothelial cells on matrix components and stimulates the migration of human umbilical vein endothelial cells. They show that Thymosin beta4 can be cross-linked to proteins such as fibrin and collagen by tissue transglutaminase. Thymosin beta4 is not cross-linked to many other proteins and its cross-linking to fibrin is competed by another family member, Thymosin beta10 (Huff et al. 2002).

Rationale for selection of Sublingual compositions comprising Thymosin fraction 5

The actin binding motif of Thymosin beta4 is an essential site for its angiogenic activity (Philip, et al. (2003). Thymosin beta4 is presented in Smart, et al. (2007) in the Nature article as a single factor that can potentially couple myocardial and coronary vascular regeneration in failing mouse hearts. They have shown that cells in the heart’s outer layer can migrate deeper into a failing organ to carry out essential repairs. The migration of progenitor cells is controlled by the protein Thymosin beta 4, already known to help reduce muscle cell loss after a heart attack.

http://news.bbc.co.uk/2/hi/health/6143286.stm

The discovery opens up the possibility of using the protein to develop more effective treatments for heart disease. Previously it was thought that cells within the adult heart are in a state of permanent rest and that any progenitor cells that can contribute to heart tissue repair travel into the heart from the bone marrow. See 150 references on that perspective on cEPCs origin and roles, which was the scientific frontier on this topic, prior to the publication of Smart et al., (2007), in Lev-Ari & Abourjaily (2006a, 2006b, 2006c).

However, researchers at University College London have demonstrated that beneficial cells actually reside in the heart itself (Smart et al. (2007). This approach would bypass the risk of immune system rejection, a major problem with the use of stem cell transplants from another source. Allogenic rejection was the main reason for the selection of an endogenous augmentation method for cEPCs using drug therapy by Lev-Ari & Abourjaily  (2006a, 2006b, 2006c). Closer examination revealed that without the Thymosin beta 4 protein, the progenitor cells failed to move deeper into the heart and change the cells needed to build healthy blood vessels and sustain muscle tissue.

http://www.irishhealth.com/clin/cholesterol/newsstory.php?id=10581

Drug # 2:

Indomethacin

Nonsteroidal anti-inflammatory drugs (NSAIDs) are widely used for their anti-inflammatory effects and have been shown to have chemopreventive effects as well. NSAIDs inhibit cyclooxygenase (COX) activity to exert their anti-inflammatory effects, but it is not clear whether their antitumorigenic ability is through COX inhibition. Using subtractive hybridization, Jain et al. (2004) identified a novel member of the transforming growth factor- superfamily that has antitumorigenic activity from Indomethacin-treated HCT-116 human colorectal cancer cells. On further investigation of this library, they now report the identification of a new cDNA corresponding to the Thymosin beta-4 gene. Thymosin beta-4 is a small peptide that is known for its actin-sequestering function, and it is associated with the induction of angiogenesis, accelerated wound healing, and metastatic potential of tumor cells. However, only selective NSAIDs induce Thymosin beta-4 expression in a time- and concentration-dependent manner. For example,

Indomethacin and SC-560 [5-(4-chlorophenyl)-1-(4-methoxyphenyl)-3-(trifluoromethyl)-1H-pyrazole] induce Thymosin beta-4 expression whereas sulindac sulfide does not.

They show that selective NSAIDs induce actin cytoskeletal reorganization, a precursory step to many dynamic processes regulating growth and motility including tumorigenesis. This is the first report to link Thymosin beta-4 induction with NSAIDs. These data suggest that NSAIDs alter the expression of a diverse number of genes and provide new insights into the chemopreventive and biological activity of these drugs (Jain et al. 2004).

Rationale for Indomethacin selection

 

Inhibitor of prostaglandin synthesis. Inhibits cyclooxygenase (COX) 1 selective.

Suggested dosage: 25 mg PO bid.

Jain et al. (2004) report a link between Thymosin beta-4 induction with NSAIDs. We selected both drugs (drug classes) and anticipate strong synergistic therapeutic effects.

Drug # 3:

Clevidipine

Clevidipine is the first third-generation calcium channel blocker, Dr. Papadakos said. It has what he called an “ultrashort” clinically relevant half-life of about one minute and then is rapidly metabolized. The effect on blood pressure is seen within one to two minutes.

http://www.medpagetoday.com/MeetingCoverage/SCCM/tb/5091

Clevidipine is an investigational agent undergoing late-stage clinical development to evaluate its potential as an innovative, targeted, fast acting intravenous product under investigation for lowering blood pressure before, during and after surgery.

http://www.themedicinescompany.com/products_Clevidipine.shtml

The Medicines Company entered into agreements with AstraZeneca PLC in March of 2002 for the development, licensing and commercialization of Clevidipine. If approved, the product could be an excellent fit with The Medicines Company’s emerging acute cardiovascular care franchise, which is led by Angiomax® (bivalirudin), an anticoagulant approved in the U.S. and other countries for use during coronary angioplasty procedures. If Clevidipine passes further clinical hurdles — phase III trials are under way — the drug may form a useful addition to the medications available to physicians in the perioperative setting

Mechanism of Action

Clevidipine belongs to a well-known class of drugs called dihydropyridine calcium channel antagonists. In vitro studies demonstrated that Clevidipine acts by selectively relaxing the smooth muscle cells that line small arteries, resulting in widening of the artery opening and reducing blood pressure within the artery (Levy, Huraux, Nordlander, 1997, 345-358).

Phase III Clinical Trials

The Medicines Company is currently sponsoring a Phase III clinical program of five studies to evaluate safety and efficacy of Clevidipine:

Early Development

The Medicines Company’s development program for Clevidipine follows upon the data sets generated by AstraZeneca, which completed clinical pharmacology, dose-finding and efficacy studies in almost 300 patients or volunteers. In clinical studies, Clevidipine has shown to provide the desired blood pressure lowering effect without causing an increase in heart rate (Kotrly, et al. 1984). Further studies demonstrate that reductions in blood pressure are dose-dependent, are not associated with an increase in heart rate and cease rapidly after stopping Clevidipine infusions (Ericsson, et al., 2000), (Schwieler, et al., 1999). In clinical studies Clevidipine was rapidly metabolized independent of the liver and the kidneys, allowing rapid clearance of the drug from the bloodstream (Ericsson, et al., 1999a), (Ericsson, et al., 1999b). Therefore, the effects of Clevidipine are short-lived, which translates into a rapid cessation of its effect on reducing blood pressure.

The two efficacy studies are known as ESCAPE-1 and ESCAPE-2. The primary objective of these studies is to determine the efficacy of Clevidipine injection versus placebo in treating pre-operative (ESCAPE-1) and post-operative (ESCAPE-2) high blood pressure. Three safety studies are collectively known as ECLIPSE. The primary objective is to establish the safety of Clevidipine in the treatment of perioperative high blood pressure, as measured by a comparison of the incidences of death, stroke, myocardial infarction and renal dysfunction between the Clevidipine and comparative treatment groups. The comparative treatments are nitroglycerin, sodium nitroprusside and nicardipine.  The ECLIPSE trial randomized 589 patients at 40 centers in the U.S. to get either sodium nitroprusside or Clevidipine. Sodium nitroprusside was administered according to institutional practice; Clevidipine was begun at 2 mg/kg and doubled every 90 seconds until blood pressure was lowered. The primary endpoint was the difference in major clinical events — death, myocardial infarction, stroke, and renal dysfunction 30 days after surgery. The secondary endpoint was blood pressure control during the first 24 hours after surgery.

The study showed no significant differences in the elements of the primary endpoint, except for mortality, Dr. Papadakos said, where 1.7% of Clevidipine patients died, compared with 4.7 of those getting sodium nitroprusside.  The difference was statistically significant at P<0.05, but Dr. Papadakos characterized the improvement as “slight.” On the other hand, the drug did show an important difference in blood pressure control over the first 24 hours, he said:

  • Patients on Clevidipine spent an average of 4.37 minutes per hour outside the desired blood pressure range.
  • Sodium nitroprusside patients spent, on average, 10.5 minutes per hour outside the desired range.
  • The difference was statistically significant at P<0.003.

Dr. Papadakos concluded that Clevidipine is a new drug that is effective, safe, and easy to use. On 2/20/2007, Dr. Deutschman, who moderated the late-breaking session at which Dr. Papadakos spoke, said that a better comparison, would be intravenous nicardipine (Cardene IV), a second-generation calcium channel blocker that is also in wide use and is considered the standard of care. “We don’t know yet if this drug is going to be better than nicardipine,” he said.

http://www.medpagetoday.com/MeetingCoverage/SCCM/tb/5091

Rationale for Clevidipine selection

Clevidipine is an acute care product. Blood pressure management is a major component of care during the 13.4 million inpatient surgeries conducted in the U.S. each year. Blood pressure control, which is managed by an anesthesiologist, is often important in patients with both normal and high blood pressure undergoing surgery or other interventional procedures. Some of these patients require rapid, precise control of blood pressure to avoid compromising key organ function such as the heart, brain and kidney.

CONCLUSION 

This is the first study to design a novel combination drug treatment for Coronary Vasculogenesis, Anti-hypertention (short-acting), Vascular Anti-inflammation (vasculitis), Neovascularization of ischemic tissue and release of adult epicardium from a quiescent state and restoring its pluripotency. This treatment is based on the new three paradigms that were presented in Cell (2006) and Nature (2007). This combination drug therapy of three drugs, one in current use (Indomethacin), and two in clinical trials (Thymosin beta4 & Clevidipine), has not been proposed before. It represents an original concept drug combination design by Lev-Ari & Abourjaily (2007). This combination represents the cutting edge conceptualization of the field of treatment of cardiac injury based on a protein produced in the heart cells, Thymosin beta4, which function as a tissue and artery healer. Its upregulation by drug therapy will revolutionize cardiology and treatment for cardiovascular disease. The combination drug therapy consists of the following drugs:

  • Drug # 1:

Thymosin fraction 5 (a sublingual composition)

  • Drug # 2:

Indomethacin (Nonsteroidal anti-inflammatory drugs (NSAID)) (25 mg PO bid)

  • Drug # 3:

Clevidipine (Blood pressure lowering drug, (no effect on heart rate))

 

REFERENCES

Al-Nedawi, K.N.I., Malgorzata, C., Bednarek, R., Szemraj, J., Swiatkowska, M., Cierniewska-Cieslak, A., Wyczolkowska, J., Cierniewski, C.S. (2004, February) “Thymosin 4 induces the synthesis of plasminogen activator inhibitor 1 in cultured endothelial cells and increases its extracellular expression.” Blood, 103, (4), 1319-1324.

Ericsson, H., Fakt. C., Hoglund. L., et al. (1999a). “Pharmacokinetics and pharmacodynamics of Clevidipine in healthy volunteers after intravenous infusion.” Eur J Clin Pharm., 55 (1), 61-67.

Ericsson, H., Fakt, C., Jolin-Mellgard A., et al. (1999b). “Clinical and pharmacokinetic results with a new ultrashort-acting calcium antagonist, Clevidipine, following gradually increasing intravenous doses to healthy volunteers.” Br J Clin Pharm., 47 (5), 531-538.

Ericsson, H., Bredberg, U., Eriksson, U., et al. (2000). “Pharmacokinetics and arteriovenous differences in Clevidipine concentration following a short and a long-term intravenous infusion in healthy volunteers.” Anesthesiology, 92 (4), 993-1001.

Fleming, I. (2006). “Signaling by the Angiotensin-Converting Enzyme” Circulation Research, 98, 887.

Heart can carry out own repairs, 16.11.2006

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http://www.medicalprogress.org/benefits/heartdis/news.cfm?news_id=478

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http://news.bbc.co.uk/2/hi/health/6143286.stm

Heart may be able to repair itself

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http://www.irishhealth.com/clin/cholesterol/newsstory.php?id=10581

Huff, T., Otto, A., Muller, C.S.G., Meier, M., Hannappel, E. (2002). “Thymosin ß4 is released from human blood platelets and attached by factor XIIIa (transglutaminase) to fibrin and collagen.”The FASEB Journal, 16, 691-696.

Jain, A.K., Moore, S.M., Yamaguchi, K., Eling, T.E., Baek, S.J. (2004, August). “Selective Nonsteroidal Anti-Inflammatory Drugs Induce Thymosin beta-4 and Alter Actin Cytoskeletal Organization in Human Colorectal Cancer Cells.” Journal of Pharmacology and Experimental Therapeutics, 311 (3) 885-891.

Kotrly, K. J., Ebert, T. J., Vucins, E. et al. (1984). “Baroreceptor reflex control of heart rate during isoflurane anesthesia in humans.” Anesthesiology,  60, 173-179.

Lev-Ari, A. & Abourjaily, P. (2006a) “An Investigation of the Potential of circulating Endothelial Progenitor Cells (cEPC) as a Therapeutic Target for Pharmacologic Therapy Design for Cardiovascular Risk Reduction.” Part I: Macrovascular Disease – Therapeutic Potential of cEPCs – Reduction methods for CV risk. Unpublished manuscript.

Lev-Ari, A. & Abourjaily, P. (2006b) “An Investigation of the Potential of circulating Endothelial Progenitor Cells (cEPC) as a Therapeutic Target for Pharmacologic Therapy Design for Cardiovascular Risk Reduction.” Part II: Therapeutic Strategy for cEPCs Endogenous Augmentation: A Concept-based Treatment Protocol for a Combined Three Drug Regimen. Unpublished manuscript.

Lev-Ari, A. & Abourjaily, P. (2006c) “An Investigation of the Potential of circulating Endothelial Progenitor Cells (cEPC) as a Therapeutic Target for Pharmacological Therapy Design for Cardiovascular Risk Reduction.” Part III: Biomarker for Therapeutic Targets of Cardiovascular Risk Reduction by cEPCs Endogenous Augmentation using New Combination Drug Therapy of Three Drug Classes and Several Drug Indications. A Theoretical Design for Quantification of the Endogenous EPCs Augmentation for Differential Level of CV Risk Reduction and Diagnostic Device Design for Drug Delivery. Unpublished manuscript.

Lev-Ari, A. & Abourjaily, P. (2007). Heart Vasculature – Regeneration and Protection of Coronary Artery Endothelium and Smooth Muscle: A Concept-based Pharmacological Therapy of a Combined Three Drug Regimen. Unpublished manuscript.

Levy, J. H., Huraux, C., Nordlander, M. (1997). “Treatment of perioperative hypertension.” In: Epstein M, Ed. Chapter in Calcium Antagonists in Clinical Medicine. Philadelphiea: Hanely & Belfus, pp. 345-358.

Liu, J-M, Lawrence, F., Kovacevic, M., Bignon, J., Papadimitriou, E., Lallemand, J-Y., Katsoris, P., Potier, P., Fromes, Y., Wdzieczak-Bakala, J. (2003, April) “The tetrapeptide AcSDKP, an inhibitor of primitive hematopoietic cell proliferation, induces angiogenesis in vitro and in vivo.” Blood, 101 (8), 3014-3020

Moretti, A., Caron, L., Nakano, A., Lam, J.T., Bernshausen, A., Chen, Y., Qyang, Y., Bu, L., Sasaki, M., Martin-Puig, S., Sun, Y., Evans, S.M., Laugwitz, K-L, Chien, K.R. (2006, December) “Multipotent Embryonic Isl1+ Progenitor Cells Lead to Cardiac, Smooth Muscle, and Endothelial Cell Diversification.” Cell, 127, 1151-1165.

Protein Discovered That Can Tell Human Heart to Heal Itself

Retrieved 3/1/2007

http://www.cathlabdigest.com/displaynews.cfm?newsid=1122065

Philp D, Huff T, Gho YS, Hannappel E, Kleinman HK. (2003). “The actin binding site on Thymosin beta4 promotes angiogenesis.” FASEB Journal, published on line 9/18/2003.

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http://www.fasebj.org/cgi/reprint/03-0121fjev1.pdf

Putting the art in heart research, 15 February 2007

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http://www.ich.ucl.ac.uk/pressoffice/pressrelease_00498

Rosenzweig A., (2005). Circulating Endothelial Progenitors – Cells as Biomarkers. NEJM, 353 (10), 1055-1057.

Schwieler, J.H., Ericsson, H., Lofdahl, P., et al. (1999). “Circulatory effects and pharmacology of Clevidipine, a novel ultra short acting and vascular selective calcium antagonist, in hypertensive humans.” J Cardiovasc Pharmacology, 34 (2), 268-274.

Smart, N., Risebro, C.A., Melville, A.D., Moses, K., Schwartz, R.J., Chien, K.R., Riley, P.R. (2007, January) “Thymosin Beta4 induces adult epicardial progenitor mobilization and neovascularization.” Nature, 445, 177-182.

 

Sublingual compositions comprising Thymosin fraction 5 and methods for administration

Retrieved 3/1/2007

http://www.pharmcast.com/Patents100/Yr2004/May2004/051104/6733791_Sublingual051104.htm

 

TMSB4X  Thymosin, beta 4, X-linked

Retrieved on 3/1/2007

http://www.ihop-net.org/UniPub/iHOP/gs/92756.html

Waeckel, L., Jérôme Bignon, J., Jian-Miao Liu, J-M., Markovits, D., Ebrahimian, T.G., Vilar, J., Mees, B., Blanc-Brude, O., Barateau, V., Sophie Le ricousse-Roussanne. S., Duriez, M. Tobelem, G.,  Wdzieczak-Bakala, J., Bernard I Lévy, B.I., Silvestre, J-S. (2006) “Tetrapeptide AcSDKP Induces Postischemic Neovascularization Through Monocyte Chemoattractant Protein-1 Signaling.” Arteriosclerosis, Thrombosis, and Vascular Biology, 26, 773

Wang, D., Oscar A. Carretero, O.A.,Yang, X-Y., Rhaleb, N-E., Liu, Y-H., Liao, T-D., Yang, X-P. (2004). “N-acetyl-seryl-aspartyl-lysyl-proline stimulates angiogenesis in vitro and in vivo.” Am J Physiol Heart Circ Physiol., 287, H2099-H2105.

Werner N, Junk S, Laufs L, Link A, Walenta K, Bohm M, Nickenig G., (2003).  Intravenous transfusion of endothelial progenitor cells reduces neointima formation after vascular injury. Circ Res., 93, e17– e24.

Werner N, Kosiol S, Schiegl T, Ahlers P, Walenta K, Link A, Böhm M, Nickenig G. (2005a). Circulating Endothelial Progenitor Cells and Cardiovascular Outcomes, NEJM, 353, 999-1007

Werner, N. & Nickenig, G. (2005b). Authors Reply to Correspondence to the Editor on Circulating Endothelial Progenitor Cells. NEJM, 353 (24), 2613-2616

Wu, S.M., Fujiwara, Y., Cibulsky, S.M., Clapham, D.E., Lien, C., Schultheiss, T.M., Orkin, S.H. (2006, December). “Developmental Origin of a Bipotential Myocardial and Smooth Muscle Cell Precursor in the Mammalian Heart.” Cell, 127, 1137-1150.

Other related articles on this Open Access Online Scientific Journal, include the following:

Saha, S. (2012b) Innovations in Bio instrumentation for Measurement of Circulating Progenetor Endothelial Cells in Human Blood.
https://pharmaceuticalintelligence.com/2012/07/08/innovations-in-bio-instrumentation-for-measurement-of-circulating-progenitor-endothelial-cells-in-human-blood/

 

Saha, S. (2012c) Endothelial Differentiation and Morphogenesis of Cardiac Precursor
https://pharmaceuticalintelligence.com/2012/07/17/endothelial-differentiation-and-morphogenesis-of-cardiac-precursors/

Saha, S. (2012e). Human Embryonic-Derived Cardiac Progenitor Cells for Myocardial Repair

https://pharmaceuticalintelligence.com/2012/08/01/human-embryonic-derived-cardiac-progenitor-cells-for-myocardial-repair/

Lev-Ari, A. 12/29/2012. Coronary artery disease in symptomatic patients referred for coronary angiography: Predicted by Serum Protein Profiles

https://pharmaceuticalintelligence.com/2012/12/29/coronary-artery-disease-in-symptomatic-patients-referred-for-coronary-angiography-predicted-by-serum-protein-profiles/

 

Bernstein, HL and Lev-Ari, A. 11/28/2012. Special Considerations in Blood Lipoproteins, Viscosity, Assessment and Treatment

https://pharmaceuticalintelligence.com/2012/11/28/special-considerations-in-blood-lipoproteins-viscosity-assessment-and-treatment/

 

Lev-Ari, A. 11/13/2012 Peroxisome proliferator-activated receptor (PPAR-gamma) Receptors Activation: PPARγ transrepression for Angiogenesis in Cardiovascular Disease and PPARγ transactivation for Treatment of Diabetes

https://pharmaceuticalintelligence.com/2012/11/13/peroxisome-proliferator-activated-receptor-ppar-gamma-receptors-activation-pparγ-transrepression-for-angiogenesis-in-cardiovascular-disease-and-pparγ-transactivation-for-treatment-of-dia/

 

Lev-Ari, A. 10/19/2012 Clinical Trials Results for Endothelin System: Pathophysiological role in Chronic Heart Failure, Acute Coronary Syndromes and MI – Marker of Disease Severity or Genetic Determination?

https://pharmaceuticalintelligence.com/2012/10/19/clinical-trials-results-for-endothelin-system-pathophysiological-role-in-chronic-heart-failure-acute-coronary-syndromes-and-mi-marker-of-disease-severity-or-genetic-determination/

 

Lev-Ari, A. 10/4/2012 Endothelin Receptors in Cardiovascular Diseases: The Role of eNOS Stimulation

https://pharmaceuticalintelligence.com/2012/10/04/endothelin-receptors-in-cardiovascular-diseases-the-role-of-enos-stimulation/

 

Lev-Ari, A. 10/4/2012 Inhibition of ET-1, ETA and ETA-ETB, Induction of NO production, stimulation of eNOS and Treatment Regime with PPAR-gamma agonists (TZD): cEPCs Endogenous Augmentation for Cardiovascular Risk Reduction – A Bibliography

https://pharmaceuticalintelligence.com/2012/10/04/inhibition-of-et-1-eta-and-eta-etb-induction-of-no-production-and-stimulation-of-enos-and-treatment-regime-with-ppar-gamma-agonists-tzd-cepcs-endogenous-augmentation-for-cardiovascular-risk-reduc/

 

Lev-Ari, A. 8/28/2012 Cardiovascular Outcomes: Function of circulating Endothelial Progenitor Cells (cEPCs): Exploring Pharmaco-therapy targeted at Endogenous Augmentation of cEPCs

https://pharmaceuticalintelligence.com/2012/08/28/cardiovascular-outcomes-function-of-circulating-endothelial-progenitor-cells-cepcs-exploring-pharmaco-therapy-targeted-at-endogenous-augmentation-of-cepcs/

 

Lev-Ari, A. 8/27/2012 Endothelial Dysfunction, Diminished Availability of cEPCs, Increasing CVD Risk for Macrovascular Disease – Therapeutic Potential of cEPCs

https://pharmaceuticalintelligence.com/2012/08/27/endothelial-dysfunction-diminished-availability-of-cepcs-increasing-cvd-risk-for-macrovascular-disease-therapeutic-potential-of-cepcs/

 

Lev-Ari, A. 8/24/2012 Vascular Medicine and Biology: CLASSIFICATION OF FAST ACTING THERAPY FOR PATIENTS AT HIGH RISK FOR MACROVASCULAR EVENTS Macrovascular Disease – Therapeutic Potential of cEPCs

https://pharmaceuticalintelligence.com/2012/08/24/vascular-medicine-and-biology-classification-of-fast-acting-therapy-for-patients-at-high-risk-for-macrovascular-events-macrovascular-disease-therapeutic-potential-of-cepcs/

 

Lev-Ari, A. 7/30/2012 Biosimilars: Intellectual Property Creation and Protection by Pioneer and by Biosimilar Manufacturers

https://pharmaceuticalintelligence.com/2012/07/30/biosimilars-intellectual-property-creation-and-protection-by-pioneer-and-by-biosimilar-manufacturers/

 

Lev-Ari, A. 7/29/2012 Biosimilars: Financials 2012 vs. 2008

https://pharmaceuticalintelligence.com/2012/07/30/biosimilars-financials-2012-vs-2008/

 

Lev-Ari, A. 7/29/2012 Biosimilars: CMC Issues and Regulatory Requirements

https://pharmaceuticalintelligence.com/2012/07/29/biosimilars-cmc-issues-and-regulatory-requirements/

 

Lev-Ari, A. 7/19/2012 Cardiovascular Disease (CVD) and the Role of agent alternatives in endothelial Nitric Oxide Synthase (eNOS) Activation and Nitric Oxide Production

https://pharmaceuticalintelligence.com/2012/07/19/cardiovascular-disease-cvd-and-the-role-of-agent-alternatives-in-endothelial-nitric-oxide-synthase-enos-activation-and-nitric-oxide-production/

 

Lev-Ari, A. 4/30/2012 Resident-cell-based Therapy in Human Ischaemic Heart Disease: Evolution in the PROMISE of Thymosin beta4 for Cardiac Repair

https://pharmaceuticalintelligence.com/2012/04/30/93/

Lev-Ari, A. 5/29/2012 Triple Antihypertensive Combination Therapy Significantly Lowers Blood Pressure in Hard-to-Treat Patients with Hypertension and Diabetes

https://pharmaceuticalintelligence.com/2012/05/29/445/

 

Lev-Ari, A. 7/2/2012 Macrovascular Disease – Therapeutic Potential of cEPCs: Reduction Methods for CV Risk

https://pharmaceuticalintelligence.com/2012/07/02/macrovascular-disease-therapeutic-potential-of-cepcs-reduction-methods-for-cv-risk/

 

Lev-Ari, A. 7/9/2012 Mitochondria Dysfunction and Cardiovascular Disease – Mitochondria: More than just the “powerhouse of the cell”

https://pharmaceuticalintelligence.com/2012/07/09/mitochondria-more-than-just-the-powerhouse-of-the-cell/

 

Lev-Ari, A. 7/16/2012 Bystolic’s generic Nebivolol – positive effect on circulating Endothelial Proginetor Cells endogenous augmentation

https://pharmaceuticalintelligence.com/2012/07/16/bystolics-generic-nebivolol-positive-effect-on-circulating-endothilial-progrnetor-cells-endogenous-augmentation/

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Arteriogenesis and Cardiac Repair: Two Biomaterials – Injectable Thymosin beta4 and Myocardial Matrix Hydrogel

Curator: Aviva Lev-Ari, PhD, RN

 

Thymosin beta 4 (Tβ4)

is a highly conserved, 43-amino acid acidic peptide (pI 4.6) that was first isolated from bovine thymus tissue over 25 years ago. It is present in most tissues and cell lines and is found in high concentrations in blood platelets, neutrophils, macrophages, and other lymphoid tissues. Tβ4 has numerous physiological functions, the most prominent of which being the regulation of actin polymerization in mammalian nucleated cells and with subsequent effects on actin cytoskeletal organization, necessary for cell motility, organogenesis, and other important cellular events.

Recently,

  • Tβ4 was shown to be expressed in the developing heart and found to stimulate migration of cardiomyocytes and endothelial cells, promote survival of cardiomyocytes (Nature, 2004), and most recently
  • to play an essential role in all key stages of cardiac vessel development: vasculogenesis, angiogenesis, and arteriogenesis (Nature 2006).

These results suggest that Tβ4 may have significant therapeutic potential in humans to protect myocardium and promote cardiomyocyte survival in the acute stages of ischemic heart disease.

RegeneRx Biopharmaceuticals, Inc. is developing Tβ4 for the treatment of patients with acute myocardial infarction (AMI). Such efforts presented will include the formulation, development, and manufacture of a suitable drug product for use in the clinic, the performance of nonclinical pharmacology and toxicology studies, and the implementation of a phase 1 clinical protocol to assess the safety, tolerability, and the pharmacokinetics of Tβ4 in healthy volunteers.

 

SOURCE:
EXPLORATIONS with THYMOSIN beta4 FOR INDUCING ADULT EPICARDIAL PROGENETOR MOBILIZATION AND NEOVASCULARIZATION is presented in
Resident-cell-based Therapy in Human Ischaemic Heart Disease: Evolution in the PROMISE of Thymosin beta4 for Cardiac Repair

https://pharmaceuticalintelligence.com/2012/04/30/93/

Clinical Study Data of Thymosin beta 4 Presented

Published on October 3, 2009 at 5:10 AM

REGENERX BIOPHARMACEUTICALS, INC. (NYSE Amex:RGN) today reported on several clinical studies with Thymosin beta 4 (Tβ4) presented the Second International Symposium on Thymosins in Health and Disease, in Catania, Italy. The following are synopses of the presentations:

Myocardial Development of RGN-352 (Injectable Tβ4 Peptide)

David Crockford, RegeneRx’s vice president for clinical and regulatory affairs presented an overview of the biological properties that support Tβ4’s near term and long term clinical applications. Mr. Crockford noted that special emphasis is being placed on the development of RGN-352 for the systemic (injectable) treatment of patients with ST-elevation myocardial infarction (STEMI) in combination with percutaneous coronary intervention, the current standard of care in most western countries for this common type of heart attack. The goal with RGN-352 is to prevent or repair continued damage to cardiac tissue post-heart attack, when such tissue around the damaged site remains at risk.

Dr. Dennis Ruff, vice president and medical director of ICON, and principal investigator, presented the most current results on the Phase I safety study with RGN-352 entitled, “A Randomized, Double-blind, Placebo-controlled, Dose-response Phase I Study of the Safety and Tolerability of the Intravenous Administration of Thymosin Beta 4 and its Pharmacokinetics After Single and Multiple Doses in Healthy Volunteers.” Dr. Ruff discussed key aspects of the study and concluded with, “There were no dose limiting or serious adverse events throughout the dosing period. Synthetic Tβ4 administered intravenously up to 1260 mg, and for up to 14 days, appears to be well tolerated with low incidence of adverse events and no evidence of serious adverse events.”

http://www.news-medical.net/news/20091003/Clinical-study-data-of-Thymosin-beta-4-presented.aspx

RegeneRx Receives Notice of Allowance from Chinese Patent Office for Treatment and Prevention of Heart Disease

RegeneRx Receives Notice of Allowance from Chinese Patent Office for Treatment and Prevention of Heart Disease

February 7, 2013 — Rockville, Md.

RegeneRx Biopharmaceuticals, Inc. (OTC Bulletin Board: RGRX) (“the Company” or “RegeneRx”) today announced that it has received a Notice of Allowance of a Chinese patent application for uses of Thymosin beta 4 (TB4) for treating, preventing, inhibiting or reducing heart tissue deterioration, injury or damage in a subject with heart failure disease. Claims also include uses for restoring heart tissue in those subjects. The patent will expire July 26, 2026.

http://www.regenerx.com/wt/page/pr_1360265259

Active Research on Thymosins in Cardiovascular Disease Reported in 2010 and 2012 Annual Conference on Thymosins, Proceedings by NY Academy of Sciences

Use of the cardioprotectants thymosin β4 and dexrazoxane during congenital heart surgery: proposal for a randomized, double-blind, clinical trial

Neonates and infants undergoing heart surgery with cardioplegic arrest experience both inflammation and myocardial ischemia-reperfusion (IR) injury. These processes provoke myocardial apoptosis and oxygen-free radical formation that result in cardiac injury and dysfunction. Thymosin β4 (Tβ4) is a naturally occurring peptide that has cardioprotective and antiapoptotic effects. Similarly, dexrazoxane provides cardioprotection by reduction of toxic reactive oxygen species (ROS) and suppression of apoptosis. We propose a pilot pharmacokinetic/safety trial of Tβ4 and dexrazoxane in children less than one year of age, followed by a randomized, double-blind, clinical trial of Tβ4 or dexrazoxane versus placebo during congenital heart surgery. We will evaluate postoperative time to resolution of organ failure, development of low cardiac output syndrome, length of cardiac ICU and hospital stays, and echocardiographic indices of cardiac dysfunction. Results could establish the clinical utility of Tβ4 and/or dexrazoxane in ameliorating ischemia-reperfusion injury during congenital heart surgery.[1]

Cardiac repair with thymosin β4 and cardiac reprogramming factors

Heart disease is a leading cause of death in newborns and in adults. We previously reported that the G-actin–sequestering peptide thymosin β4 promotes myocardial survival in hypoxia and promotes neoangiogenesis, resulting in cardiac repair after injury. More recently, we showed that reprogramming of cardiac fibroblasts to cardiomyocyte-like cells in vivo after coronary artery ligation using three cardiac transcription factors (Gata4/Mef2c/Tbx5) offers an alternative approach to regenerate heart muscle. We have combined the delivery of thymosin β4 and the cardiac reprogramming factors to further enhance the degree of cardiac repair and improvement in cardiac function after myocardial infarction. These findings suggest that thymosin β4 and cardiac reprogramming technology may synergistically limit damage to the heart and promote cardiac regeneration through the stimulation of endogenous cells within the heart.[2]

NMR structural studies of thymosin α1 and β-thymosins

Thymosin proteins, originally isolated from fractionation of thymus tissue, represent a class of compounds that we now know are present in numerous other tissues, are unrelated to each other in a genetic sense, and appear to have different functions within the cell. Thymosin α1 (generic drug name thymalfasin; trade name Zadaxin) is derived from a precursor molecule, prothymosin, by proteolytic cleavage, and stimulates the immune system. Although the peptide is natively unstructured in aqueous solution, the helical structure has been observed in the presence of trifluoroethanol or unilamellar vesicles, and these studies are consistent with the presence of a dynamic helical structure whose sides are not completely hydrophilic or hydrophobic. This helical structure may occur in circulation when the peptide comes into contact with membranes. In this report, we discuss the current knowledge of the thymosin α1 structure and similar properties of thymosin β4 and thymosin β9, in different environments.[3]

Thymosin β4 sustained release from poly (lactide-co-glycolide) microspheres: synthesis and implications for treatment of myocardial ischemia

 A sustained release formulation for the therapeutic peptide thymosin β4 (Tβ4) that can be localized to the heart and reduce the concentration and frequency of dose is being explored as a means to improve its delivery in humans. This review contains concepts involved in the delivery of peptides to the heart and the synthesis of polymer microspheres for the sustained release of peptides, including Tβ4. Initial results of poly(lactic-co-glycolic acid) microspheres synthesized with specific tolerances for intramyocardial injection that demonstrate the encapsulation and release of Tβ4 from double-emulsion microspheres are also presented.[4]
Thymosin β4 and cardiac repair
Hypoxic heart disease is a predominant cause of disability and death worldwide. As adult mammals are incapable of cardiac repair after infarction, the discovery of effective methods to achieve myocardial and vascular regeneration is crucial. Efforts to use stem cells to repopulate damaged tissue are currently limited by technical considerations and restricted cell potential. We discovered that the small, secreted peptide thymosin β4 (Tβ4) could be sufficiently used to inhibit myocardial cell death, stimulate vessel growth, and activate endogenous cardiac progenitors by reminding the adult heart on its embryonic program in vivo. The initiation of epicardial thickening accompanied by increase of myocardial and epicardial progenitors with or without infarction indicate that the reactivation process is independent of injury. Our results demonstrate Tβ4 to be the first known molecule able to initiate simultaneous myocardial and vascular regeneration after systemic administration in vivo. Given our findings, the utility of Tβ4 to heal cardiac injury may hold promise and warrant further investigation.[7]
Thymosin β4 facilitates epicardial neovascularization of the injured adult heart
Ischemic heart disease complicated by coronary artery occlusion causes myocardial infarction (MI), which is the major cause of morbidity and mortality in humans (http://www.who.int/cardiovascular_diseases/resources/atlas/en/index.html). After MI the human heart has an impaired capacity to regenerate and, despite the high prevalence of cardiovascular disease worldwide, there is currently only limited insight into how to stimulate repair of the injured adult heart from its component parts. Efficient cardiac regeneration requires the replacement of lost cardiomyocytes, formation of new coronary blood vessels, and appropriate modulation of inflammation to prevent maladaptive remodeling, fibrosis/scarring, and consequent cardiac dysfunction. Here we show that thymosin β4 (Tβ4) promotes new vasculature in both the intact and injured mammalian heart. We demonstrate that limited EPDC-derived endothelial-restricted neovascularization constitutes suboptimal “endogenous repair,” following injury, which is significantly augmented by Tβ4 to increase and stabilize the vascular plexus via collateral vessel growth. As such, we identify Tβ4 as a facilitator of cardiac neovascularization and highlight adult EPDCs as resident progenitors which, when instructed by Tβ4, have the capacity to sustain the myocardium after ischemic damage.[8]
Thymosin β4 enhances repair by organizing connective tissue and preventing the appearance of myofibroblasts
Incisional wounds in rats treated locally with thymosin β4 (Tβ4) healed with minimal scaring and without loss in wound breaking strength. Treated wounds were significantly narrower in width. Polarized light microscopy treated wounds had superior organized collagen fibers, displaying a red birefringence, which is consistent with mature connective tissue. Control incisions had randomly organized collagen fibers, displaying green birefringence that is consistent with immature connective tissue. Immunohistology treated wounds had few myofibroblasts and fibroblasts with α smooth muscle actin (SMA) stained stress fibers. Polyvinyl alcohol sponge implants placed in subcutaneous pockets received either carrier or 100 μg of Tβ4 on days 2, 3, and 4. On day 14, treated implants revealed longer, thicker collagen fiber bundles with intense yellow-red birefringence by polarized light microscopy. In controls, fine, thin collagen fiber bundles were arranged in random arrays with predominantly green birefringence. Controls contained mostly myofibroblasts, while few myofibroblasts appeared in Tβ4 treated implants. Electron microscopy confirmed both cell types and the degree of collagen fiber bundle organization. Our results demonstrate that Tβ4 treated wounds appear to mature earlier and heal with minimal scaring.[9]
Thymosin β4: a key factor for protective effects of eEPCs in acute and chronic ischemia
Acute myocardial infarction is still one of the leading causes of death in the industrial nations. Even after successful revascularization, myocardial ischemia results in a loss of cardiomyocytes and scar formation. Embryonic EPCs (eEPCs), retroinfused into the ischemic region of the pig heart, provided rapid paracrine benefit to acute and chronic ischemia in a PI-3K/Akt-dependent manner. In a model of acute myocardial ischemia, infarct size and loss of regional myocardial function decreased after eEPC application, unless cell pre-treatment with thymosin β4 shRNA was performed. Thymosin ß4 peptide retroinfusion mimicked the eEPC-derived improvement of infarct size and myocardial function. In chronic ischemia (rabbit model), eEPCs retroinfused into the ischemic hindlimb enhanced capillary density, collateral growth, and perfusion. Therapeutic neovascularization was absent when thymosin ß4 shRNA was introduced into eEPCs before application. In conclusion, eEPCs are capable of acute and chronic ischemia protection in a thymosin ß4 dependent manner. [10]
Thymosin β4: a candidate for treatment of stroke?
Neurorestorative therapy is the next frontier in the treatment of stroke. An expanding body of evidence supports the theory that after stroke, certain cellular changes occur that resemble early stages of development. Increased expression of developmental proteins in the area bordering the infarct suggest an active repair or reconditioning response to ischemic injury. Neurorestorative therapy targets parenchymal cells (neurons, oligodendrocytes, astrocyes, and endothelial cells) to enhance endogenous neurogenesis, angiogenesis, axonal sprouting, and synaptogenesis to promote functional recovery. Pharmacological treatments include statins, phosphodiesterase 5 inhibitors, erythropoietin, and nitric oxide donors that have all improved funtional outcome after stroke in the preclinial arena. Thymosin β4 (Tβ4) is expressed in both the developing and adult brain and it has been shown to stimulate vasculogenesis, angiogenesis, and arteriogenesis in the postnatal and adult murine cardiac myocardium. In this manuscript, we describe our rationale and techniques to test our hypothesis that Tβ4 may be a candidate neurorestorative agent. [11]
Prothymosin α as robustness molecule against ischemic stress to brain and retina

Following stroke or traumatic damage, neuronal death via both necrosis and apoptosis causes loss of functions, including memory, sensory perception, and motor skills. As necrosis has the nature to expand, while apoptosis stops the cell death cascade in the brain, necrosis is considered to be a promising target for rapid treatment for stroke. We identified the nuclear protein, prothymosin alpha (ProTα) from the conditioned medium of serum-free culture of cortical neurons as a key protein-inhibiting necrosis. In the culture of cortical neurons in the serum-free condition without any supplements, ProTα inhibited the necrosis, but caused apoptosis. In the ischemic brain or retina, ProTα showed a potent inhibition of both necrosis and apoptosis. By use of anti-brain-derived neurotrophic factor or anti-erythropoietin IgG, we found that ProTα inhibits necrosis, but causes apoptosis, which is in turn inhibited by ProTα-induced neurotrophins under the condition of ischemia. From the experiment using anti-ProTα IgG or antisense oligonucleotide for ProTα, it was revealed that ProTα has a pathophysiological role in protecting neurons in stroke.[12]

 
Thymosin β4 and cardiac repair
Hypoxic heart disease is a predominant cause of disability and death worldwide. As adult mammals are incapable of cardiac repair after infarction, the discovery of effective methods to achieve myocardial and vascular regeneration is crucial. Efforts to use stem cells to repopulate damaged tissue are currently limited by technical considerations and restricted cell potential. We discovered that the small, secreted peptide thymosin β4 (Tβ4) could be sufficiently used to inhibit myocardial cell death, stimulate vessel growth, and activate endogenous cardiac progenitors by reminding the adult heart on its embryonic program in vivo. The initiation of epicardial thickening accompanied by increase of myocardial and epicardial progenitors with or without infarction indicate that the reactivation process is independent of injury. Our results demonstrate Tβ4 to be the first known molecule able to initiate simultaneous myocardial and vascular regeneration after systemic administration in vivo. Given our findings, the utility of Tβ4 to heal cardiac injury may hold promise and warrant further investigation.[13]
Thymosin β4 facilitates epicardial neovascularization of the injured adult heart
schemic heart disease complicated by coronary artery occlusion causes myocardial infarction (MI), which is the major cause of morbidity and mortality in humans (http://www.who.int/cardiovascular_diseases/resources/atlas/en/index.html). After MI the human heart has an impaired capacity to regenerate and, despite the high prevalence of cardiovascular disease worldwide, there is currently only limited insight into how to stimulate repair of the injured adult heart from its component parts. Efficient cardiac regeneration requires the replacement of lost cardiomyocytes, formation of new coronary blood vessels, and appropriate modulation of inflammation to prevent maladaptive remodeling, fibrosis/scarring, and consequent cardiac dysfunction. Here we show that thymosin β4 (Tβ4) promotes new vasculature in both the intact and injured mammalian heart. We demonstrate that limited EPDC-derived endothelial-restricted neovascularization constitutes suboptimal “endogenous repair,” following injury, which is significantly augmented by Tβ4 to increase and stabilize the vascular plexus via collateral vessel growth. As such, we identify Tβ4 as a facilitator of cardiac neovascularization and highlight adult EPDCs as resident progenitors which, when instructed by Tβ4, have the capacity to sustain the myocardium after ischemic damage. [14]
Thymosin β4: a key factor for protective effects of eEPCs in acute and chronic ischemia

Acute myocardial infarction is still one of the leading causes of death in the industrial nations. Even after successful revascularization, myocardial ischemia results in a loss of cardiomyocytes and scar formation. Embryonic EPCs (eEPCs), retroinfused into the ischemic region of the pig heart, provided rapid paracrine benefit to acute and chronic ischemia in a PI-3K/Akt-dependent manner. In a model of acute myocardial ischemia, infarct size and loss of regional myocardial function decreased after eEPC application, unless cell pre-treatment with thymosin β4 shRNA was performed. Thymosin ß4 peptide retroinfusion mimicked the eEPC-derived improvement of infarct size and myocardial function. In chronic ischemia (rabbit model), eEPCs retroinfused into the ischemic hindlimb enhanced capillary density, collateral growth, and perfusion. Therapeutic neovascularization was absent when thymosin ß4 shRNA was introduced into eEPCs before application. In conclusion, eEPCs are capable of acute and chronic ischemia protection in a thymosin ß4 dependent manner.[15]

 
Thymosin β4: a candidate for treatment of stroke?
Neurorestorative therapy is the next frontier in the treatment of stroke. An expanding body of evidence supports the theory that after stroke, certain cellular changes occur that resemble early stages of development. Increased expression of developmental proteins in the area bordering the infarct suggest an active repair or reconditioning response to ischemic injury. Neurorestorative therapy targets parenchymal cells (neurons, oligodendrocytes, astrocyes, and endothelial cells) to enhance endogenous neurogenesis, angiogenesis, axonal sprouting, and synaptogenesis to promote functional recovery. Pharmacological treatments include statins, phosphodiesterase 5 inhibitors, erythropoietin, and nitric oxide donors that have all improved funtional outcome after stroke in the preclinial arena. Thymosin β4 (Tβ4) is expressed in both the developing and adult brain and it has been shown to stimulate vasculogenesis, angiogenesis, and arteriogenesis in the postnatal and adult murine cardiac myocardium. In this manuscript, we describe our rationale and techniques to test our hypothesis that Tβ4 may be a candidate neurorestorative agent.[16]
Thymosin β4: structure, function, and biological properties supporting current and future clinical applications

Published studies have described a number of physiological properties and cellular functions of thymosin β4 (Tβ4), the major G-actin-sequestering molecule in mammalian cells. Those activities include the promotion of cell migration, blood vessel formation, cell survival, stem cell differentiation, the modulation of cytokines, chemokines, and specific proteases, the upregulation of matrix molecules and gene expression, and the downregulation of a major nuclear transcription factor. Such properties have provided the scientific rationale for a number of ongoing and planned dermal, corneal, cardiac clinical trials evaluating the tissue protective, regenerative and repair potential of Tβ4, and direction for future clinical applications in the treatment of diseases of the central nervous system, lung inflammatory disease, and sepsis. A special emphasis is placed on the development of Tβ4 in the treatment of patients with ST elevation myocardial infarction in combination with percutaneous coronary intervention.[17]

The effect of thymosin treatment of venous ulcers

Venous ulcers are responsible for about 70% of the chronic ulcers of the lower limbs. Standard of care includes compression, dressings, debridement of devitalized tissue, and infection control. Thymosin beta 4 (Tβ4), a synthetic copy of the naturally occurring 43 amino-acid peptide, has been found to have wound healing and anti-inflammatory properties, and is thought to exert its therapeutic effect through promotion of keratinocyte and endothelial cell migration, increased collagen deposition, and stimulation of angiogenesis. To assess the safety, tolerability, and efficacy of topically administered Tβ4 in patients with venous stasis ulcers, a double-blind, placebo-controlled, dose-escalation study was conducted in eight European sites (five in Italy and three in Poland) that enrolled and randomized 73 patients. The safety profile of all doses of administered Tβ4 was deemed acceptable and comparable to placebo. Efficacy findings from this Phase 2 study suggest that a Tβ4 dose of 0.03% may have the potential to accelerate wound healing and that complete wound healing can be achieved within 3 months in about 25% of the patients, especially among those whose wounds are small to moderate in size or mild to moderate in severity.[18]

A randomized, placebo-controlled, single and multiple dose study of intravenous thymosin β4 in healthy volunteers

Synthetic thymosin beta 4 (Tβ4) may have a potential use in promoting myocardial cell survival during acute myocardial infarction. Four cohorts, with 10 healthy subjects each, were given a single intravenous dose of placebo or synthetic Tβ4. Cohorts received ascending doses of either 42, 140, 420, or 1260 mg. Following safety review, subjects were given the same dose regimen daily for 14 days. Safety evaluations, incidence of Treatment-Emergent Adverse Events, and pharmacokinetic parameters were evaluated. Adverse events were infrequent, and mild or moderate in intensity. There were no dose limiting toxicities or serious adverse events. Pharmacokinetic profile for single dose showed a dose proportional response, and an increasing half-life with increasing dose. Synthetic Tβ4 given intravenously as a single dose or in multiple daily doses for 14 days over a dose range of 42–1260 mg was well tolerated with no evidence of dose limiting toxicity. Further development for use in cardiac ischemia should be considered.[19]

Safety and Efficacy of an Injectable Extracellular Matrix Hydrogel for Treating Myocardial Infarction

  1. Sonya B. Seif-Naraghi1,*,
  2. Jennifer M. Singelyn1,*,
  3. Michael A. Salvatore2,
  4. Kent G. Osborn1,
  5. Jean J. Wang1,
  6. Unatti Sampat1,
  7. Oi Ling Kwan1,
  8. G. Monet Strachan1,
  9. Jonathan Wong3,
  10. Pamela J. Schup-Magoffin1,
  11. Rebecca L. Braden1,
  12. Kendra Bartels1,
  13. Jessica A. DeQuach2,
  14. Mark Preul4,
  15. Adam M. Kinsey2,
  16. Anthony N. DeMaria1,
  17. Nabil Dib1 and
  18. Karen L. Christman1,

+Author Affiliations

  1. 1University of California, San Diego, La Jolla, CA 92093, USA.
  2. 2Ventrix, Inc., San Diego, CA 92109, USA.
  3. 3Biologics Delivery Systems, Irwindale, CA 91706, USA.
  4. 4Barrow Neurological Institute, Phoenix, AZ 85013, USA.

+Author Notes

  • * These authors contributed equally to this work.
  1. †To whom correspondence should be addressed. E-mail: christman@eng.ucsd.edu

ABSTRACT

New therapies are needed to prevent heart failure after myocardial infarction (MI). As experimental treatment strategies for MI approach translation, safety and efficacy must be established in relevant animal models that mimic the clinical situation. We have developed an injectable hydrogel derived from porcine myocardial extracellular matrix as a scaffold for cardiac repair after MI. We establish the safety and efficacy of this injectable biomaterial in large- and small-animal studies that simulate the clinical setting. Infarcted pigs were treated with percutaneous transendocardial injections of the myocardial matrix hydrogel 2 weeks after MI and evaluated after 3 months. Echocardiography indicated improvement in cardiac function, ventricular volumes, and global wall motion scores. Furthermore, a significantly larger zone of cardiac muscle was found at the endocardium in matrix-injected pigs compared to controls. In rats, we establish the safety of this biomaterial and explore the host response via direct injection into the left ventricular lumen and in an inflammation study, both of which support the biocompatibility of this material. Hemocompatibility studies with human blood indicate that exposure to the material at relevant concentrations does not affect clotting times or platelet activation. This work therefore provides a strong platform to move forward in clinical studies with this cardiac-specific biomaterial that can be delivered by catheter.

  • Copyright © 2013, American Association for the Advancement of Science
Citation: S. B. Seif-Naraghi, J. M. Singelyn, M. A. Salvatore, K. G. Osborn, J. J. Wang, U. Sampat, O. L. Kwan, G. M. Strachan, J. Wong, P. J. Schup-Magoffin, R. L. Braden, K. Bartels, J. A. DeQuach, M. Preul, A. M. Kinsey, A. N. DeMaria, N. Dib, K. L. Christman, Safety and Efficacy of an Injectable Extracellular Matrix Hydrogel for Treating Myocardial Infarction.

RELATED RESOURCES ON SCIENCE SITES

In Science Translational Medicine

REFERENCES OF THYMOSIN IN CARDIOVASCULAR DISEASE

Thymosins in Health and Disease II: 3rd International Symposium on The Emerging Clinical Applications of Tymosin beta 4 in Cardiovascular Disease

Annals of the New York Academy of Sciences, October 2012 Volume 1270 Pages vii-ix, 1–121.

Allan L. Goldstein, Enrico Garaci, Editors, Thymosins in Cardiovascular Disease, November 2012, Wiley-Blackwell

http://onlinelibrary.wiley.com/doi/10.1111/nyas.2012.1270.issue-1/issuetoc

http://www.wiley.com/WileyCDA/WileyTitle/productCd-1573319104.html?cid=RSS_WILEY2_LIFEMED

1


Use of the cardioprotectants thymosin β4 and dexrazoxane during congenital heart surgery: proposal for a randomized, double-blind, clinical trial (pages 59–65) Daniel Stromberg, Tia Raymond, David Samuel, David Crockford, William Stigall, Steven Leonard, Eric Mendeloff and Andrew Gormley
Article first published online: 10 OCT 2012 | DOI: 10.1111/j.1749-6632.2012.06710.x

2


Cardiac repair with thymosin β4 and cardiac reprogramming factors (pages 66–72) Deepak Srivastava, Masaki Ieda, Jidong Fu and Li Qian
Article first published online: 10 OCT 2012 | DOI: 10.1111/j.1749-6632.2012.06696.x

3 NMR structural studies of thymosin α1 and β-thymosins (pages 73–78) David E. Volk, Cynthia W. Tuthill, Miguel-Angel Elizondo-Riojas and David G. Gorenstein
Article first published online: 10 OCT 2012 | DOI: 10.1111/j.1749-6632.2012.06656.x

4

Thymosin β4 sustained release from poly(lactide-co-glycolide) microspheres: synthesis and implications for treatment of myocardial ischemia (pages 112–119) Jeffrey E. Thatcher, Tré Welch, Robert C. Eberhart, Zoltan A. Schelly and J. Michael DiMaio
Article first published online: 10 OCT 2012 | DOI: 10.1111/j.1749-6632.2012.06681.x

5 Corrigendum for Ann. N.Y. Acad. Sci. 2012. 1254: 57–65 (page 121) Article first published online: 10 OCT 2012 | DOI: 10.1111/j.1749-6632.2012.06793.x
This article corrects:
A bird’s-eye view of cell therapy and tissue engineering for cardiac regeneration
Vol. 1254, Issue 1, 57–65, Article first published online: 30 APR 2012

Thymosins in Health and Disease: 2nd International Symposium,
Annals of the New York Academy of Sciences, May 2010 Volume 1194 Pages ix–xi, 1–230 

http://onlinelibrary.wiley.com/doi/10.1111/nyas.2010.1194.issue-1/issuetoc

6. Preface to Thymosins in Health and Disease (pages ix–xi) Enrico Garaci and Allan L. Goldstein
Article first published online: 3 MAY 2010 | DOI: 10.1111/j.1749-6632.2010.05493.x

7.
Thymosin β4 and cardiac repair (pages 87–96) Santwana Shrivastava, Deepak Srivastava, Eric N. Olson, J. Michael DiMaio and Ildiko Bock-Marquette
Article first published online: 3 MAY 2010 | DOI: 10.1111/j.1749-6632.2010.05468.x

8.
Thymosin β4 facilitates epicardial neovascularization of the injured adult heart (pages 97–104) Nicola Smart, Catherine A. Risebro, James E. Clark, Elisabeth Ehler, Lucile Miquerol, Alex Rossdeutsch, Michael S. Marber and Paul R. Riley
Article first published online: 3 MAY 2010 | DOI: 10.1111/j.1749-6632.2010.05478.x

9.
Thymosin β4 enhances repair by organizing connective tissue and preventing the appearance of myofibroblasts (pages 118–124) H. Paul Ehrlich and Sprague W. Hazard III
Article first published online: 3 MAY 2010 | DOI: 10.1111/j.1749-6632.2010.05483.x

10. Thymosin β4: a key factor for protective effects of eEPCs in acute and chronic ischemia (pages 105–111) Rabea Hinkel, Ildiko Bock-Marquette, Antonis K. Hazopoulos and Christian Kupatt
Article first published online: 3 MAY 2010 | DOI: 10.1111/j.1749-6632.2010.05489.x
Corrected by:
Corrigendum for Ann. N. Y. Acad. Sci. 1194: 105–111
Vol. 1205, Issue 1, 284, Article first published online: 14 SEP 2010

11.

Thymosin β4: a candidate for treatment of stroke? (pages 112–117) Daniel C. Morris, Michael Chopp, Li Zhang and Zheng G. Zhang
Article first published online: 3 MAY 2010 | DOI: 10.1111/j.1749-6632.2010.05469.x

12. Prothymosin α as robustness molecule against ischemic stress to brain and retina (pages 20–26) Hiroshi Ueda, Hayato Matsunaga, Hitoshi Uchida and Mutsumi Ueda
Article first published online: 3 MAY 2010 | DOI: 10.1111/j.1749-6632.2010.05466.x

13.
Thymosin β4 and cardiac repair (pages 87–96) Santwana Shrivastava, Deepak Srivastava, Eric N. Olson, J. Michael DiMaio and Ildiko Bock-Marquette
Article first published online: 3 MAY 2010 | DOI: 10.1111/j.1749-6632.2010.05468.x

14.

Thymosin β4 facilitates epicardial neovascularization of the injured adult heart (pages 97–104) Nicola Smart, Catherine A. Risebro, James E. Clark, Elisabeth Ehler, Lucile Miquerol, Alex Rossdeutsch, Michael S. Marber and Paul R. Riley
Article first published online: 3 MAY 2010 | DOI: 10.1111/j.1749-6632.2010.05478.x

15.

Thymosin β4: a key factor for protective effects of eEPCs in acute and chronic ischemia (pages 105–111) Rabea Hinkel, Ildiko Bock-Marquette, Antonis K. Hazopoulos and Christian Kupatt
Article first published online: 3 MAY 2010 | DOI: 10.1111/j.1749-6632.2010.05489.x
Corrected by:
Corrigendum for Ann. N. Y. Acad. Sci. 1194: 105–111
Vol. 1205, Issue 1, 284, Article first published online: 14 SEP 2010

16.

Thymosin β4: a candidate for treatment of stroke? (pages 112–117) Daniel C. Morris, Michael Chopp, Li Zhang and Zheng G. Zhang
Article first published online: 3 MAY 2010 | DOI: 10.1111/j.1749-6632.2010.05469.x

17.Thymosin β4: structure, function, and biological properties supporting current and future clinical applications (pages 179–189) David Crockford, Nabila Turjman, Christian Allan and Janet Angel
Article first published online: 3 MAY 2010 | DOI: 10.1111/j.1749-6632.2010.05492.x

18.

The effect of thymosin treatment of venous ulcers (pages 207–212) G. Guarnera, A. DeRosa and R. Camerini, on behalf of 8 European sites
Article first published online: 3 MAY 2010 | DOI: 10.1111/j.1749-6632.2010.05490.x

19.
A randomized, placebo-controlled, single and multiple dose study of intravenous thymosin β4 in healthy volunteers (pages 223–229) Dennis Ruff, David Crockford, Gino Girardi and Yuxin Zhang
Article first published online: 3 MAY 2010 | DOI: 10.1111/j.1749-6632.2010.05474.x

Other related articles on this Open Access Online Scientific Journal include the following:

Gene Therapy Into Healthy Heart Muscle: Reprogramming Scar Tissue In Damaged Hearts

https://pharmaceuticalintelligence.com/2013/01/09/gene-therapy-into-healthy-heart-muscle-reprogramming-scar-tissue-in-damaged-hearts/

Human Embryonic-Derived Cardiac Progenitor Cells for Myocardial Repair

https://pharmaceuticalintelligence.com/2012/08/01/human-embryonic-derived-cardiac-progenitor-cells-for-myocardial-repair/

Human embryonic pluripotent stem cells and healing post-myocardial infarction

https://pharmaceuticalintelligence.com/2012/08/07/human-embryonic-pluripotent-stem-cells-and-healing-post-myocardial-infarction/

Resident-cell-based Therapy in Human Ischaemic Heart Disease: Evolution in the PROMISE of Thymosin beta4 for Cardiac Repair

https://pharmaceuticalintelligence.com/2012/04/30/93/

Heart Renewal by pre-existing Cardiomyocytes: Source of New Heart Cell Growth Discovered

https://pharmaceuticalintelligence.com/2012/12/23/heart-renewal-by-pre-existing-cardiomyocytes-source-of-new-heart-cell-growth-discovered/

Absorb™ Bioresorbable Vascular Scaffold: An International Launch by Abbott Laboratories

https://pharmaceuticalintelligence.com/2012/09/29/absorb-bioresorbable-vascular-scaffold-an-international-launch-by-abbott-laboratories/

Heart patients’ skin cells turned into healthy heart muscle cells

https://pharmaceuticalintelligence.com/2012/06/04/heart-patients-skin-cells-turned-into-healthy-heart-muscle-cells/

Telling NO to Cardiac Risk

https://pharmaceuticalintelligence.com/2012/12/10/telling-no-to-cardiac-risk/

Read Full Post »


Author, Editor: Tilda Barliya, PhD

Although melanoma accounts for only 4 percent of all dermatologic cancers, it is responsible for 80 percent of deaths from skin cancer; only 14 percent of patients with metastatic melanoma survive for five years (1). The incidence of melanoma is increasing worldwide, with a growing fraction of patients with advanced disease for which prognosis remains poor despite advances in the field (2). Treatment options are limited despite advances in immunotherapy and targeted therapy. For patients with surgically resected, thick (≥2 mm) primary melanoma with or without regional lymph node metastases, the only effective adjuvant therapy is interferon-α (IFN-α). However, because of the limited benefit upon disease-free survival and the smaller potential improvement of overall survival, the indication for IFN-α treatment remains controversial (2). A better understanding of melanoma immunosurveillance is therefore essential to enable the design of better, targeted melanoma therapies (4).

Risk factors (2):

  • Family history of melanoma, multiple benign or atypical nevi, and a previous melanoma
  • Immunosuppression
  • Sun sensitivity
  • Exposure to ultraviolet radiation

Each of these risk factors corresponds to a genetic predisposition or an environmental stressor that contributes to the genesis of melanoma and each factor is understood to various degrees at a molecular level. The Clark model of the progression of melanoma emphasizes the stepwise transformation of melanocytes to melanoma. The model depicts the proliferation of melanocytes in the process of forming nevi and the subsequent development of dysplasia, hyperplasia, invasion, and metastasis.

 

This Clark’s multi-step model, and predict that the acquisition of a BRAF mutation can be a founder event in melanocytic neoplasia. While mutations of the BRAF gene are frequent in melanomas on non-chronic sun damaged skin which are prevalent in Caucasians, acral and mucosal melanomas harbor mutations of the KIT gene as well as the amplifications of cyclin D1 or cyclin-dependent kinase 4 gene.

The choice of target antigens is key to the success of tumour vaccination or tumour immunotherapy. Melanoma candidate antigens include: (A) mutated or aberrantly expressed molecules (e.g. CDK4, MUM-1, beta-catenin) (B) cancer/testis antigens (e.g. MAGE, BAGE and GAGE) and (C) melanoma- associated antigens (MAA).

MAAs are self-antigens normally expressed during the differentiation of melanocytes and play a role in different enzymatic steps of melanogenesis. However, in transformed melanocytes (melanoma cells), MAAs are often overexpressed (4).

The main MAAs are tyrosinase, an enzyme that catalyses the production of melanin from tyrosine by oxidation, the tyrosinase-related proteins (TRP-1) and 2 (TRP-2), the glycoprotein (gp)100 (silver-gene) and MelanA/MART. It is thought that the specialized cell biology of melanin synthesis may favour the loading of MAA peptides into the antigen presentation pathway. 50% of melanoma patients have tumour-infiltrating lymphocytes (TILs) recognising tyrosinase and Melan A, indicating that these antigens are important in the natural melanoma immunosurveillance. Moreover, MAAs are well characterized in mice and humans, allowing the development of tetramers to detect antigen-specific immune responses.

Tα1 Mechanism of action

Tα1, a 28 amino acid peptide of ∼3.1 kDa, is endogenously produced by the thymus gland by the cleavage of its precursor pro-Ta1.  Although the fine immunologic mechanism(s) of action of T1 have not fully been elucidated, experimental evidence points to its strong immunomodulatory properties. In particular, it was reported that Ta1 enhances T cell–mediated immune responses by several mechanisms, including increased T cell production (i.e., CD4+, CD8+, and CD3+ cells), stimulation of T cell differentiation and/or maturation, reduction of T cell apoptosis, and restoration of T cell–mediated antibody production (5).

Furthermore, it was demonstrated that T1 acts on the immune system by modulating the release of proinflammatory cytokines (i.e., interleukin-2 (IL-2), interferon-gamma (IFN-)),12–14 and through the activation of natural killer and dendritic cells.12 In addition, T1 was also demonstrated to have direct effects on cancer cells by increasing the levels of expression of different tumor antigens and of components of the major histocompatibility complex class I, as well as by reducing cancer cell growth.

Together, these experimental findings bear relevance for cancer immunotherapy and suggest that T1 can activate innate and adaptive immune responses and modulate the immunophenotype of cancer cells, improving their immunogenicity and their recognition by the immune system.

Danielli R and colleagues have very nicely outlined the use of the Thymosin a1 in the clinical setting for treating melanoma (5) titled :”Thymosin a1 in melanoma: from the clinical trial setting to the daily practice and beyond”.  A large body of available preclinical in vitro and in vivo evidence points to thymosin alpha 1 (Ta1) as a useful immunomodulatory peptide,with significant therapeutic potential in metastatic melanoma in the absence of clinically meaningful toxicity.  The results emerging frominitial trials provide support of the ability of T1 to improve the clinical outcome of advanced melanoma patients through the activation of the immune system.

Ta1 and Clinical Trials in Melanoma

A large scale, randomized, phase II study was conducted at 64 European centers between 2002 and 2006 to investigate the efficacy of Ta1 administered in combination with DTIC (Dacarbazine) or with DTIC + IFNa, versus only DTIC + IFNa, in 488 previously untreated patients with cutaneous metastatic melanoma. The study was designed to evaluate the ability of Ta1 to potentiate the therapeutic efficacy of DTIC.

Patients were randomly assigned to five treatment groups: DTIC + IFNa and 1.6 mg of Ta1; DTIC + IFNa and 3.2 mg of T1; DTIC + IFN-a and 6.4 mg of Ta1; DTIC + 3.2 mg of Ta1; and DTIC + IFNa

Results:

The clinical rate (CBR), defined as the proportion of patients with a complete response, partial response, or stable disease, was significantly higher in patients who received Ta1 + DTIC than in those who received control therapy. Results in patients who received T1 (all groups combined) compared with those who received the control treatment

  • Improved progression-free survival (hazard ratio (HR): 0.80;
  • 95%confidence interval (CI): 0.63–1.01; P = 0.06) and
  • OS (median: 9.4 vs. 6.6 months)

These outcomes suggested to addition of Thymosin a1 to the treatment resulted in the reduction in the risk of mortality and disease progression in patients with metastatic malignant melanoma, and pointed to a poor effect of IFN- in the combination. More so, the poor results of the IFN group is not surprising due to the limited therapeutic activity of IFN observed in phase III clinical trials.

This study however have some limitations as standard assessment criteria, such as RECIST and WHO indications,  conventionally applied to cytotoxic agents, do not adequately capture some patterns of response observed in the course of immunotherapy; stemming from these considerations, immune-related response criteria (irRC) were developed to measure primary and secondary endpoints in immunotherapy clinical trials.

Therefore the above study might underestimate the therapeutic efficacy of Thymosin a1 since irRC criteria were not used.

In Summary:

A large scale phase III clinical trial should be designed to further explore the therapeutic activity of Thymosine a1 in melanoma patients with defined endpoints and irRC criteria. Moreover, combination studies should explore the activity of T1 in association with other approved agents, such as ipilimumab and vemurafenib or as maintenance therapy in melanoma patients who experience clinical benefit after treatment with these agents.

Also, because of the pleiotropic immunemechanism(s) of action of T1, including the upregulation of T cell–driven immune responses against specific tumor antigens, priming of immune responses and potentiation of antitumor T cell–mediated immune responses through the activation of Toll-like receptor 9 on dendritic cells, coupling Ta1 to cancer vaccines should be an additional useful therapeutic strategy to pursue. T1 could, in fact, prove helpful in overcoming the limited immunogenicity and the short-lived persistency of adequate immunologic antitumor responses frequently reported as potential causes of failure of therapeutic vaccines.

Ref:

1. Arlo J. Miller, M.D.,., and Martin C. Mihm, Jr. Mechanisms of disease: Melanoma. N Engl J Med 2006 (6); 355:51-65.

http://www.nejm.org.rproxy.tau.ac.il/doi/pdf/10.1056/NEJMra052166

http://www.nejm.org/doi/full/10.1056/NEJMra052166

2. Garbe C., Eigentler TK., Keilholz U., Hauschild A and Kirkwood JM. Systematic review of medical treatment in melanoma: current status and future prospects. Oncologist 2011;16(1):5-24.

http://theoncologist.alphamedpress.org/content/16/1/5.long

3. http://flipper.diff.org/app/items/info/1983

4.  Träger U, Sierro S, Djordjevic G, Bouzo B, Khandwala S, et al. (2012) The Immune Response to Melanoma Is Limited by Thymic Selection of Self-Antigens. PLoS ONE 7(4): e35005. doi:10.1371/journal.pone.0035005.

http://www.plosone.org/article/info:doi/10.1371/journal.pone.0035005

5. Riccardo Danielli, Ester Fonsatti, Luana Calabr` o, Anna Maria Di Giacomo, and Michele Maio. Thymosin 1 in melanoma: from the clinical trial setting to the daily practice and beyond. Ann. N.Y. Acad. Sci. 1270 (2012) 8–12.

http://www.ncbi.nlm.nih.gov/pubmed/16822996

http://onlinelibrary.wiley.com/doi/10.1111/j.1749-6632.2012.06757.x/abstract

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Resident-cell-based Therapy in Human Ischaemic Heart Disease: Evolution in the PROMISE of Thymosin beta4 for Cardiac Repair

Curator: Aviva Lev-Ari, PhD, RN

2012 – STATE OF PHARMACEUTICAL RESEARCH

Scientists Report that Process of Converting Non-Beating Heart Cells into Functional, Beating Heart Cells is Enhanced Using Thymosin Beta 4 in Conjunction with Gene Therapy

ROCKVILLE, Md.–(BUSINESS WIRE)–Apr. 18, 2012– Regenerx Biopharmaceuticals, Inc. (OTC Bulletin Board: RGRX) (“the Company” or “RegeneRx”) announced today that scientists at the Gladstone Institute of Cardiovascular Disease, University of California, San Francisco, have published new animal data in the current issue of Nature showing that the process of converting non-beating heart cells (which normally form scar tissue after a heart attack), into functional, beating heart muscle cells can be enhanced using Thymosin beta 4 (Tβ4). Delivery of Tβ4, in conjunction with GMT (an acronym for three genes that normally guide embryonic heart development), into the damaged region resulted in reduction of scar area and improvement in cardiac function compared to GMT or Tβ4 alone. Within a month, non-beating cells that normally form scar tissue transformed into beating heart-muscle cells. Within three months, the hearts were beating even stronger and pumping more blood.

“Our experiments in mice are a proof of concept that we can reprogram non-beating cells directly into fully functional, beating heart cells – offering an innovative and less invasive way to restore heart function after a heart attack,” stated Dr. Deepak Srivastava, who directs cardiovascular and stem cell research at Gladstone and a member of RegeneRx’s scientific advisory board.

“These findings could have a significant impact on heart-failure patients whose damaged hearts make it difficult for them to engage in normal activities like walking up a flight of stairs,” said Dr. Li Qian, PhD, who is a postdoctoral scholar at Gladstone and a member of Dr. Srivastava’s research team. “This research may result in a much needed alternative to heart transplants for which donors are extremely limited. And because we are reprogramming cells directly in the heart, we eliminate the need to surgically implant cells that were created in a petri dish,” he further commented.

According to the Institute news release, “The results have broad human health implications” and are a “medical breakthrough [that] holds promise for millions with heart failure.”

The results are described in the latest issue of Nature, available online today.

About RegeneRx Biopharmaceuticals, Inc. (www.regenerx.com)

RegeneRx is focused on the development of a novel therapeutic peptide, Thymosin beta 4, or Tβ4, for tissue and organ protection, repair and regeneration. RegeneRx currently has three drug candidates in Phase 2 clinical development and has an extensive worldwide patent portfolio covering its products.

2012 –  STATE OF SCIENCE: MOLECULAR CELL CARDIOLOGY
Recent research by Zhou BHonor LBMa QOh JHLin RZMelero-Martin JMvon Gise AZhou PHu THe LWu KHZhang HZhang YPu WT of the Key Laboratory of Nutrition and Metabolism, Institute for Nutritional Sciences, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China concluded that Thymosin beta 4 treatment after myocardial infarction does not reprogram epicardial cells into cardiomyocytes J Mol Cell Cardiol. 2012 Jan;52(1):43-7. Epub 2011 Aug 26.
Kispert A. commented:“No muscle for a damaged heart: thymosin beta 4 treatment after myocardial infarction does not induce myocardial differentiation of epicardial cells.” http://www.ncbi.nlm.nih.gov/pubmed?term=Kispert%20A.
2012 – 2003 MILESTONES IN THE EVOLUTION OF THE PROMISE OF THYMOSIN beta4 FOR CARDIAC REPAIR

A significant bottleneck in cardiovascular regenerative medicine is the identification of a viable source of stem/progenitor cells that could contribute new muscle after ischaemic heart disease and acute myocardial infarction. A therapeutic ideal–relative to cell transplantation–would be to stimulate a resident source, thus avoiding the caveats of limited graft survival, restricted homing to the site of injury and host immune rejection. Thymosin β4, a peptide previously shown to restore vascular potential to adult epicardium-derived progenitor cells with injury, indicating that  an epicardial origin of the progenitor population, and embryonic reprogramming results in the mobilization of this population and concomitant differentiation to give rise to de novo cardiomyocytes. Derived cardiomyocytes are shown here to structurally and functionally integrate with resident muscle; as such, stimulation of this adult progenitor pool represents a significant step towards resident-cell-based therapy in human ischaemic heart disease.

Shrivastava SSrivastava DOlson ENDiMaio JMBock-Marquette I. of Department of Cardiovascular and Thoracic Surgery, University of Texas, Southwestern Medical Center, Dallas, Texas, USA in Ann N Y Acad Sci. 2010 Apr;1194:87-96. asserted that Tbeta4 to be the first known molecule able to initiate simultaneous myocardial and vascular regeneration.

Another a study by Smart NRisebro CAClark JEEhler EMiquerol LRossdeutsch AMarber MSRiley PR, of the Molecular Medicine Unit, UCL Institute of Child Health, London, UK. concluded that Thymosin beta4 facilitates epicardial neovascularization of the injured adult heart, Ann N Y Acad Sci. 2010 Apr;1194:97-104

Additional research on De novo cardiomyocytes from within the activated adult heart after injury by Smart NBollini SDubé KNVieira JMZhou BDavidson SYellon DRiegler JPrice ANLythgoe MFPu WTRiley PR. Molecular Medicine Unit, UCL Institute of Child Health, London,Nature. 2011 Jun 8;474(7353):640-4 . They demonstrate in mice that the adult heart contains a resident stem or progenitor cell population, which has the potential to contribute bona fide terminally differentiated cardiomyocytes after myocardial infarction. They reveal a novel genetic label of the activated adult progenitors via re-expression of a key embryonic epicardial gene, Wilm’s tumour 1 (Wt1), through priming by thymosin β4, a peptide previously shown to restore vascular potential to adult epicardium-derived progenitor cells with injury.

 In search for new strategies to repair and/or regenerate the myocardium after ischemia and infarction in order to prevent maladaptive remodeling and cardiac dysfunction, Cavasin MA, of the Hypertension and Vascular Research Division, Henry Ford Health System, Detroit, Michigan, in Therapeutic potential of thymosin-beta4 and its derivative N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) in cardiac healing after infarction, Am J Cardiovasc Drugs. 2006;6(5):305-11 compiles and analyzes the available experimental data regarding the potential therapeutic effects of thymosin-beta4 and its derivative N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) in cardiac healing after myocardial infarction (MI) as well as discussing the possible mechanisms involved. More recently it has been shown that thymosin-beta4 facilitates cardiac repair after infarction by promoting cell migration and myocyte survival. Additionally, the tetrapeptide Ac-SDKP was reported to reduce left ventricular fibrosis in hypertensive rats, reverse fibrosis and inflammation in rats with MI, and stimulate both in vitro and in vivo angiogenesis. Ac-SDKP also reduced cardiac rupture rate in mice post-MI. Some of the effects of Ac-SDKP, such as the enhancement of angiogenesis and the decrease in inflammation and collagenase activity, are similar to those described for thymosin-beta4. Thus, it is possible that Ac-SDKP could be mediating some of the beneficial effects of its precursor. Although the experimental evidence is very promising, there are no data available from a clinical trial supporting the use of thymosin-beta(4) or Ac-SDKP as means of healing the myocardium after MI in patients.
EXPLORATIONS with THYMOSIN beta4 FOR INDUCING ADULT EPICARDIAL PROGENETOR MOBILIZATION AND NEOVASCULARIZATION
 
Smart et al. (2007) implicate Thymosine beta4 (Tb4) with the following functions: (a) Tb4 in regulating all three key stages of cardiac vessel development: coronary vasculogenesis, angiogenesis and arteriogenesis – collateral growth; (b) identify the adult epicardium as a potential source of vascular progenitors which, when stimulated by Tb4, migrate and differentiate into smooth muscle and endothelial cells; (c) the ability of Tb4 to promote coronary vascularization both during development and in the adult, enhances cardiomyocyte survival and contributes significantly towards Tb4-induced cardioprotection.

Nature, 2007, 445, 177-182.

The reaction in the scientific community to these investigative results was most favorable.

“These results are very exciting because most humans suffering from ischemic cardiac events, either acutely or chronically, do not develop the collateral vessel growth necessary to preserve and restore heart tissue. If, in humans, we see the same effects as seen in mice, TB4 would be the first drug to prevent loss of (heart) muscle cells and restore blood flow in this manner and provide a new and much needed treatment modality for these patients,”

commented Deepak Srivastava, M.D., Professor and Director, Gladstone Institute of Cardiovascular Disease, University of California San Francisco, CA. Dr. Srivastava and his colleagues published the first paper on TB4’s effects on myocardial infarction in Nature in November 2004.

http://phx.corporate-ir.net/phoenix.zhtml?c=144396&p=irol-newsArticle&ID=932573&highlight=

 Dr. Paul Riley, Institute of Child Health, University College London, Research Lead on the Smart et al. (2007) article in Nature, said: “In 2006, through (British Heart Foundation) BHF-funded work, we discovered that a protein called Thymosin beta4 could mobilise dormant cells from the epicardium to form new blood vessels in the heart. This is a major step towards finding a DIY repair mechanism to repair injury following heart attack.”

http://www.ich.ucl.ac.uk/pressoffice/pressrelease_00498

 “To investigate whether Thymosin beta 4 could have a therapeutic effect on damaged adult hearts, my research team took cells from the outermost layer of adult mice and grew them in the lab. We found that, when treated with the protein, these adult cells have as much potential as embryonic cells to create healthy heart tissue. This suggests that the protein could have a therapeutic use,” explained lead researcher, Dr. Paul Riley. Furthermore, “current treatments for a damaged heart are limited by the ability of the adult tissue to respond. By using this protein to guide progenitor cells from the outer layer of the heart, to form new blood vessels and nourish tissue, it could be possible to better repair damaged adult hearts.”

“Our research has shown that blood vessel regeneration is still possible in the adult heart. In the future, if we can figure out how to direct the progenitor cells using Thymosin beta 4, there could be potential for therapy based on the patients’ own heart cells”, Dr. Riley explained. He said that this process has the added benefit in that the cells are already located in the right place – within the heart itself.

“All these cells need is the appropriate instructions to guide them towards new blood vessel formation that will help in the repair of muscle damage following a heart attack”, Dr. Riley added.

http://www.irishhealth.com/clin/cholesterol/newsstory.php?id=10581

Professor Jeremy Pearson, BHF associate medical director, said: “These results are important and exciting.” By identifying for the first time a molecule that can cause cells in the adult heart to form new blood vessels, Dr. Riley’s group have taken a large step towards practical therapy to encourage damaged hearts to repair themselves, a goal that researchers are urgently aiming for.” Here, we target pharmaco-therapy for their discovery.

Professor Colin Blakemore, MRC chief executive, said: ”Finding out how this protein helps to heal the heart offers enormous potential in fighting heart disease, which kills more than 105,000 people in the UK every year.”

http://news.bbc.co.uk/2/hi/health/6143286.stm

Philip et al., (2003) reported that Thymosin beta4 is angiogenic and can promote endothelial cell migration and adhesion, tubule formation, aortic ring sprouting, and angiogenesis. It also accelerates wound healing and reduces inflammation when applied in dermal wound-healing assays. Using naturally occurring Thymosin beta4, proteolytic fragments, and synthetic peptides, they found that a seven amino acid actin binding motif of Thymosin beta4 was essential for its angiogenic activity. Migration assays with human umbilical vein endothelial cells and vessel sprouting assays using chick aortic arches showed that Thymosin beta4 and the actin-binding motif of the peptide display near-identical activity at ~50 nM, whereas peptides lacking any portion of the actin motif were inactive. Furthermore, adhesion to Thymosin beta4 was blocked by this seven amino acid peptide demonstrating it as the major Thymosin beta4 cell binding site on the molecule. The adhesion and sprouting activity of Thymosin beta4 was inhibited with the addition of 5-50 nM soluble actin. These results demonstrate that the actin binding motif of Thymosin beta4 is an essential site for its angiogenic activity. FASEB Journal,2003, published on line 9/18/2003. Retrieved 3/1/2007, FASEB Journal,2007.

Smart et al. (2007) describe the mechanism by which Thymosin beta4 stimulates coronary vessel development which in this regard involves Thymosin beta4 directly promoting Epicardium-Derived Cells (EPDC) migration from the epicardium via its previously known function of actin binding, filament assembly and lamellipodia formation. Thymosin beta4 is presented in their Nature article as a single factor that can potentially couple myocardial and coronary vascular regeneration in failing mouse hearts. A major shortcoming of current angiogenic therapy in response to myocardial ischaemia in humans is that the outcome may be limited to capillary growth without concomitant collateral support of arterioles. Smart et al. (2007) findings that, in mice, Thymosin beta4 can promote vessel formation and collateral growth not only during development but also critically from adult epicardium, suggest Thymosin beta4 has considerable therapeutic potential in humans. They revealed the mechanism by which Thymosin beta4 may act to promote cardiomyocyte survival following acute myocardial damage in mice and identify the biopeptide AcSDKP as a small molecule that potentially offers further protection following cardiac injury. Nature, 2007, 445, 177-182.

N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) stimulates endothelial cell differentiation from adult epicardium

The first report demonstrating the ability of AcSDKP to interact directly with endothelial cells and to elicit an angiogenic response in vitro and in vivo was reported by Liu, et al., (2003). A novel biologic function of AcSDKP, is its function as a mediator of angiogenesis, as measured by its ability to modulate endothelial cell function in vitro and angiogenesis in vivo. The tetrapeptide acetyl-Ser-Asp-Lys-Pro (AcSDKP), purified from bone marrow and constitutively synthesized in vivo, belongs to the family of negative regulators of hematopoiesis. It protects the stem cell compartment from the toxicity of anticancer drugs and irradiation and consequently contributes to a reduction in marrow failure. AcSDKP at nanomolar concentrations stimulates in vitro endothelial cell migration and differentiation into capillary-like structures on Matrigel as well as enhances the secretion of an active form of matrix metalloproteinase-1 (MMP-1). In vivo, AcSDKP promotes a significant angiogenic response in the chicken embryo chorioallantoic membrane (CAM) and in the abdominal muscle of the rat. Moreover, it induces the formation of blood vessels in Matrigel plugs implanted subcutaneously in the rat (Liu, et al., 2003). Blood,2003, 101 (8), 3014-3020

Wang et al., (2004) reported three findings, that N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) (a) stimulated endothelial cell proliferation and migration and tube formation in a dose-dependent manner, (b) enhanced corneal neovascularization, and (c) increased myocardial capillary density. Endothelial cell proliferation and angiogenesis stimulated by Ac-SDKP could be beneficial in cardiovascular diseases such as hypertension and MI. Furthermore, they wrote, because Ac-SDKP is mainly cleaved by ACE, it may partially mediate the cardioprotective effect of ACE inhibitors. .” Am J Physiol Heart Circ Physiol., 2004, 287, H2099-H2105.

Fleming (2006) reports that recent evidence suggests that some of the beneficial effects of ACE inhibitors on cardiovascular function and homeostasis can be attributed to novel mechanisms. These include the accumulation of the ACE substrate N-acetyl-seryl-aspartyl-lysyl-proline, which blocks collagen deposition in the injured heart, as well as the activation of an ACE signaling cascade that involves the activation of the kinase CK2 and the c-Jun N-terminal kinase in endothelial cells and leads to changes in gene expression. Circulation Research,2006, 98, 887.

Waeckel et al. (2006) report the putative proangiogenic activity and molecular pathway(s) of the tetrapeptide acetyl-N-Ser-Asp-Lees-Pro (AcSDKP) in a model of surgically induced hind limb ischemia. AcSDKP stimulated MCP-1 mRNA and protein levels in cultured endothelial cells and ischemic tissue. AcSDKP stimulates postischemic neovascularization through activation of a proinflammatory MCP-1-related pathway. Arteriosclerosis, Thrombosis, and Vascular Biology,2006, 26, 773

Smart et al. (2007) in Nature, identified the biopeptide AcSDKP as a small molecule that potentially offers further protection following cardiac injury. Their research on AcSDKP, continue the work of Fleming (2006), Liu et al. (2003), Waeckel at al. (2006), Wang et al. (2004) and references 19 to 25 in Smart (2007). In their paper they report that scope exists for a non actin-mediated vasculo-, angio- and arteriogenic function for Thymosin beta4 by virtue of its endoproteinase activity to produce the pro-angiogenic tetrapeptide N-acetyl-seryl-aspartyl-lysyl-proline (AcSDKP). They therefore quantified AcSDKP levels in their mutant knockdown hearts by competitive enzyme immunoassay on extracted myocardium, and found they were decreased to 62% and 60%, respectively, of that of controls.

They concluded that (a) this is robust evidence for a peptide and precursor peptide relationship between Thymosin beta4 and AcSDKP in a physiological setting. They investigated whether AcSDKP could rescue any of the vasculogenesis defects observed in the Thymosin beta4 mutant hearts, and (b) stated that their results support their interpretation of the primary phenotype in Thymosin beta4 mutants. AcSDKP lacks actin binding function, rendering it unable to stimulate filamentous actin assembly and lamellipodia-based cell migration and consequently unable to rescue the Epicardium-Derived Cells (EPDC) defect.

However, in the adult, consistent with reported cardioprotective effects of AcSDKP (refs 23–25 in Smart et al. (2007)), they observed a significant upregulation in levels of both endogenous Thymosin beta4 and AcSDKP in response to ischaemia after 1 day and 1 week. They reported that addition of AcSDKP to adult epicardial explants resulted in a striking increase in differentiated (Flk1-positive) endothelial cells. Although unable to promote epicardial outgrowth beyond control levels, AcSDKP brought about rapid differentiation of any emerging EPDCs. The differentiated cells were almost exclusively endothelial, with only very few smooth muscle cells observed in AcSDKP-treated cultures.  They stated  that it suggests that cleavage of AcSDKP from Thymosin beta4 exclusively promotes EPDC endothelial cell differentiation, and may underlie a compound vasculogenic effect of Thymosin beta4 aside from simply promoting EPDC migration into overlying myocardium as an instructive cue for differentiation.

Lastly, they concluded that crucial to the further understanding of this two-step function will be the identification of the respective receptors for Thymosin beta4 and AcSDKP. Receptors for Thymosin beta4 and AcSDKP will promote research activity for drug discovery.

Leading drug developer of synthetic Thymosin beta4 is RegeneRx Biopharmaceuticals, Inc. They are developing Thymosin beta4, a 43 amino acid peptide as a potential therapeutic target in part, under an exclusive world-wide license from the National Institutes of Health. RegeneRx holds nearly 60 world-wide patents and patent applications related to dermal, ocular, and internal wounds and tissue repair, cardiac and neurological injuries, septic shock and several consumer product areas. RegeneRx is currently sponsoring three Phase 2 chronic dermal wound healing clinical trials and has additionally targeted ophthalmic and cardiac trials in 2007 as part of its ongoing clinical development program. J.J. Finkelstein is RegeneRx’s president and chief executive officer.

http://phx.corporate-ir.net/phoenix.zhtml?c=144396&p=irol-newsArticle&ID=953115&highlight=

Scientists Find Heart Stem Cells

by Constance Holden on 2 July 2009, 12:00 AM in http://news.sciencemag.org/sciencenow/2009/07/02-01.html

Scientists have identified a cardiac stem cell that gives rise to all of the major cell types in the human heart. The find opens the way to using patients’ own cells to heal their damaged hearts.

The cells in question express a protein, called Islet 1, which is present in the early stages of fetal heart formation. In recent years, scientists have identified the cells in embryonic mouse hearts. And now, a team in the laboratory of Kenneth Chien, director of the Cardiovascular Research Center at Massachusetts General Hospital in Boston, has found the same cell type in human fetal hearts.

Once the group pinpointed the cells, it took the next important step: generating new cardiac stem cells from human embryonic stem cells. Using fluorescent tags to identify the ones containing Islet 1, the researchers obtained a purified population. They then proved that the Islet 1 cells are what Chien calls “master stem cells” by showing that single cells could be made to grow into any of the heart’s major cell types: heart muscle (cardiomyocytes), smooth muscle, and blood vessel lining (endothelium). The team reports its work today in Nature.

Chien cautions that these primordial stem cells, which are found only in fetuses, could not be used for therapy because they could develop into undesired cell types. Instead, he says, researchers need to isolate “intermediate” cells that are already heading for a particular fate. In the meantime, the primordial cells could be used for disease modeling and drug screening. They may also help shed light on congenital heart malformations. In the fetal heart, Islet 1 cells are clustered in areas that are “hot spots” for congenital heart defects, says Chien: “Congenital heart disease may be a stem cell disease.”

Ultimately, researchers may be able to use the cells to grow human “heart parts” such as strips of heart muscle or a valve on scaffolds that could be inserted into patients, Chien says.

“The findings are very important if they can be reproduced,” says cardiologist Richard Schatz of Scripps Clinic in San Diego, California. But Eduardo Marbán, director of the Cedars-Sinai Heart Institute in Los Angeles, says he’s doubtful that the identification of Islet 1 cells will hasten new therapies. Marbán is currently heading a trial that involves removing a tiny chunk of heart tissue from a patient, cultivating cells from it, and reinjecting them into the patient’s heart. He says Islet 1 cells “do appear to be important in development” but that “normal heart tissue can and does form in the complete absence” of the protein.

Chien has a different view, saying that there’s little or no evidence that scientists can obtain stem cells by “grinding up hearts and culturing cells from them.” It’s important to identify authentic progenitor cells, he says, in order to identify cells that will help repair damaged hearts.

http://news.softpedia.com/newsImage/Stem-Cells-for-the-Hear-Found-2.jpg/

http://news.sciencemag.org/sciencenow/2009/07/02-01.html

http://www.readcube.com/articles/10.1038/nature08191

Pluripotent stem cell-based heart regeneration: from the developmental and immunological perspectives.

Kathy O KO LuiLei L BuRonald A RA LiCamie W CW Chan
Birth Defects Res A Clin Mol Teratol 96(1):98-108 (2012), PMID 22457181

Heart diseases such as myocardial infarction cause massive loss of cardiomyocytes, but the human heart lacks the innate ability to regenerate. In the adult mammalian heart, a resident progenitor cell population, termed epicardial progenitors, has been identified and reported to stay quiescent under uninjured conditions; however, myocardial infarction induces their proliferation and de novo differentiation into cardiac cells. It is conceivable to develop novel therapeutic approaches for myocardial repair by targeting such expandable sources of cardiac progenitors, thereby giving rise to new muscle and vasculatures. Human pluripotent stem cells such as embryonic stem cells and induced pluripotent stem cells can self-renew and differentiate into the three major cell types of the heart, namely cardiomyocytes, smooth muscle, and endothelial cells. In this review, we describe our current knowledge of the therapeutic potential and challenges associated with the use of pluripotent stem cell and progenitor biology in cell therapy. An emphasis is placed on the contribution of paracrine factors in the growth of myocardium and neovascularization as well as the role of immunogenicity in cell survival and engraftment.

Multipotent Embryonic Isl1^+ Progenitor Cells Lead to Cardiac, Smooth Muscle, and Endothelial Cell Diversification

Alessandra A MorettiLeslie L CaronAtsushi A NakanoJason T JT LamAlexandra A Bernshausen,Yinhong Y ChenYibing Y QyangLei L BuMika M SasakiSilvia S Martin-PuigYunfu Y SunSylvia M SM EvansKarl-Ludwig KL LaugwitzKenneth R KR Chien
Cell 127(6):15 (2006), PMID 17123592

Cardiogenesis requires the generation of endothelial, cardiac, and smooth muscle cells, thought to arise from distinct embryonic precursors. We use genetic fate-mapping studies to document that isl1^+ precursors from the second heart field can generate each of these diverse cardiovascular cell types in vivo. Utilizing embryonic stem (ES) cells, we clonally amplified a cellular hierarchy of isl1^+ cardiovascular progenitors, which resemble the developmental precursors in the embryonic heart. The transcriptional signature of isl1^+/Nkx2.5^+/flk1^+ defines a multipotent cardiovascular progenitor, which can give rise to cells of all three lineages. These studies document a developmental paradigm for cardiogenesis, where muscle and endothelial lineage diversification arises from a single cell-level decision of a multipotent isl1^+ cardiovascular progenitor cell (MICP). The discovery of ES cell-derived MICPs suggests a strategy for cardiovascular tissue regeneration via their isolation, renewal, and directed differentiation into specific mature cardiac, pacemaker, smooth muscle, and endothelial cell types.

http://pubget.com/paper/17123592/Multipotent_Embryonic_Isl1___Progenitor_Cells_Lead_to_Cardiac__Smooth_Muscle__and_Endothelial_Cell_Diversification

 

 

 

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