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Inhibition of histone deacetylase as a treatment for cardiac hypertrophyRelated Patent Categories: Drug, Bio-affecting And Body Treating Compositions, Designated Organic Active Ingredient Containing (doai), Peptide Containing (e.g., Protein, Peptones, Fibrinogen, Etc.) Doai, CyclopeptidesInhibition of histone deacetylase as a treatment for cardiac hypertrophy description/claimsThe Patent Description & Claims data below is from USPTO Patent Application 20060069014, Inhibition of histone deacetylase as a treatment for cardiac hypertrophy. Brief Patent Description - Full Patent Description - Patent Application Claims
BACKGROUND OF THE INVENTION [0002] 1. Field of the Invention [0003] The present invention relates generally to the fields of developmental biology and molecular biology. More particularly, it concerns gene regulation and cellular physiology in cardiomyocytes. Specifically, the invention relates to the use of HDAC inhibitors to treat cardiac hypertrophy and heart failure. [0004] 2. Description of Related Art [0005] Cardiac hypertrophy in response to an increased workload imposed on the heart is a fundamental adaptive mechanism. It is a specialized process reflecting a quantitative increase in cell size and mass (rather than cell number) as the result of any or a combination of neural, endocrine or mechanical stimuli. Hypertension, another factor involved in cardiac hypertrophy, is a frequent precursor of congestive heart failure. When heart failure occurs, the left ventricle usually is hypertrophied and dilated and indices of systolic function, such as ejection fraction, are reduced. Clearly, the cardiac hypertrophic response is a complex syndrome and the elucidation of the pathways leading to cardiac hypertrophy will be beneficial in the treatment of heart disease resulting from a various stimuli. [0006] A family of transcription factors, the myocyte enhancer factor-2 family (MEF2), are involved in cardiac hypertrophy. For example, a variety of stimuli can elevate intracellular calcium, resulting in a cascade of intracellular signaling systems or pathways, including calcineurin, CAM kinases PKC and MAP kinases. All of these signals activate MEF2 and result in cardiac hypertrophy. However, it is still not completely understood how the various signal systems exert their effects on MEF2 and modulate its hypertrophic signaling. It is known that certain histone deacetylase proteins, HDAC 4, HDAC 5, HDAC 7, HDAC 9, and HDAC 10, are involved in modulating MEF2 activity. [0007] Eleven different HDACs have been cloned from vertebrate organisms. All share homology in the catalytic region. Histone acetylases and deacetylases play a major role in the control of gene expression. The balance between activities of histone acetylases, usually called acetyl transferases (HATs), and deacetylases (HDACs) determines the level of histone acetylation. Consequently, acetylated histones cause relaxation of chromatin and activation of gene transcription, whereas deacetylated chromatin is generally transcriptionally inactive. In a previous report, the inventors' laboratory demonstrated that HDAC 4 and 5 dimerize with MEF2 and repress the transcriptional activity of MEF2 and, further, that this interaction requires the presence of the N-terminus of the HDAC 4 and 5 proteins. McKinsey et al. (2000a,b). [0008] In a distinct context, recent research has also highlighted the important role of HDACs in cancer biology. In fact, various inhibitors of HDACs are being tested for their ability to induce cellular differentiation and/or apoptosis in cancer cells. Marks et al. (2000). Such inhibitors include suberoylanilide hydroxamic acid (SAHA) (Butler et al., 2000; Marks et al., 2001); m-carboxycinnamic acid bis-hydroxamide (Coffey et al., 2001); and pyroxamide (Butler et al., 2001). These studies have been summarized as indicating "that the hydroxamic acid-based HPCs, in particular SAHA and pyroxamide--are potent inhibitors of HDAC in vitro and in vivo and induce growth arrest, differentiation, or apoptotic cell death of transformed cells . . . [and thus] are lead compounds among the family of hydroxamic acid-based HPCs and are currently in phase I clinical trials." Marks et al. (2000). To date, no reports on the effects of HDAC inhibitors on muscle cell hypertrophy and response to stress have been reported. SUMMARY OF THE INVENTION [0009] Thus, in accordance with the present invention, there is provided a method of treating pathologic cardiac hypertrophy and heart failure comprising (a) identifying a patient having cardiac hypertrophy; and (b) administering to the patient a histone deacetylase inhibitor. Administering may comprise intravenous, oral, transdermal, sustained release, suppository, or sublingual administration. The method may further comprise administering a second therapeutic regimen, such as a beta blocker, an iontrope, diuretic, ACE-I, AII antagonist or Ca.sup.++-blocker. The second therapeutic regimen may be administered at the same time as the histone deacetylase inhibitor, or either before or after the histone deacetylase inhibitor. The treatment may improve one or more symptoms of cardiac failure such as providing increased exercise capacity, increased blood ejection volume, left ventricular end diastolic pressure, pulmonary capillary wedge pressure, cardiac output, cardiac index, pulmonary artery pressures, left ventricular end systolic and diastolic dimensions, left and right ventricular wall stress, wall tension and wall thickness, quality of life, disease-related morbidity and mortality. [0010] In yet another embodiment, there is provided a method of preventing pathologic cardiac hypertrophy and heart failure comprising (a) identifying a patient at risk of developing cardiac hypertrophy; and (b) administering to the patient a histone deacetylase inhibitor. Administration may comprise intravenous, oral, transdermal, sustained release, suppository, or sublingual administration. The patient at risk may exhibit one or more of long standing uncontrolled hypertension, uncorrected valvular disease, chronic angina and/or recent myocardial infarction. [0011] In accordance with the preceding embodiments, the histone deacetylase inhibitor may be any molecule that effects a reduction in the activity of a histone deacetylase. This includes proteins, peptides, DNA molecules (including antisense), RNA molecules (including RNAi and antisense) and small molecules. The small molecules include, but are not limited to, trichostatin A, trapoxin B, MS 275-27, m-carboxycinnamic acid bis-hydroxamide, depudecin, oxamflatin, apicidin, suberoylanilide hydroxamic acid, Scriptaid, pyroxamide, 2-amino-8-oxo-9,10-epoxy-decanoyl, 3-(4-aroyl-1H-pyrrol-2-yl)-N-hydroxy-2-propenamide and FR901228. Additionally, the following references describe histone deacetylase inhibitors which may be selected for use in the current invention: AU 9,013,101; AU 9,013,201; AU 9,013,401; AU 6,794,700; EP 1,233,958; EP 1,208,086; EP 1,174,438; EP 1,173,562; EP 1,170,008; EP 1,123,111; JP 2001/348340; U.S. 2002/103192; U.S. 2002/65282; U.S. 2002/61860; WO 02/51842; WO 02/50285; WO 02/46144; WO 02/46129; WO 02/30879; WO 02/26703; WO 02/26696; WO 01/70675; WO 01/42437; WO 01/38322; WO 01/18045; WO 01/14581; Furumai et al. (2002); Hinnebusch et al. (2002); Mai et al. (2002); Vigushin et al. (2002); Gottlicher et al. (2001); Jung (2001); Komatsu et al. (2001); Su et al. (2000). [0012] In still another embodiment, there is provided a method of identifying inhibitors of cardiac hypertrophy comprising (a) providing a histone deacetylase inhibitor; (b) treating a myocyte with the histone deacetylase inhibitor; and (c) measuring the expression of one or more cardiac hypertrophy parameters, wherein a change in the one or more cardiac hypertrophy parameters, as compared to one or more cardiac hypertrophy parameters in a myocyte not treated with the histone deacetylase inhibitor, identifies the histone deacetylase inhibitor as an inhibitor of cardiac hypertrophy. The myocyte may be subjected to a stimulus that triggers a hypertrophic response in the one or more cardiac hypertrophy parameters, such as expression of a transgene or treatment with a drug. [0013] The one or more cardiac hypertrophy parameters may comprise the expression level of one or more target genes in the myocyte, wherein the expression level of the one or more target genes is indicative of cardiac hypertrophy. The one or more target genes may be selected from the group consisting of ANF, .alpha.-MyHC, .beta.-MyHC, .alpha.-skeletal actin, SERCA, cytochrome oxidase subunit VIII, mouse T-complex protein, insulin growth factor binding protein, Tau-microtubule-associated protein, ubiquitin carboxyl-terminal hydrolase, Thy-1 cell-surface glycoprotein, or MyHC class I antigen. The expression level may be measured using a reporter protein coding region operably linked to a target gene promoter, such as luciferase, .beta.-gal or green fluorescent protein. The expression level may be measured using hybridization of a nucleic acid probe to a target mRNA or amplified nucleic acid product. [0014] The one or more cardiac hypertrophy parameters also may comprise one or more aspects of cellular morphology, such as sarcomere assembly, cell size, or cell contractility. The myocyte may be an isolated myocyte, or comprised in isolated intact tissue. The myocyte also may be a cardiomyocyte, and, may be located in vivo in a functioning intact heart muscle, such as functioning intact heart muscle that is subjected to a stimulus that triggers a hypertrophic response in one or more cardiac hypertrophy parameters. The stimulus may be aortic banding, rapid cardiac pacing, induced myocardial infarction, or transgene expression. The one or more cardiac hypertrophy parameters comprises right ventricle ejection fraction, left ventricle ejection fraction, ventricular wall thickness, heart weight/body weight ratio, or cardiac weight normalization measurement. The one or more cardiac hypertrophy parameters also may comprise total protein synthesis. [0015] In still yet another embodiment, there is provided a method of identifying inhibitors of cardiac hypertrophy comprising (a) providing at least one class I and one class II histone deacetylase; (b) contacting the histone deacetylases with a candidate inhibitor substance; and (c) measuring the activity of the histone deacetylases, wherein a greater decrease in class I histone deacetylase activity than class II histone deacetylase activity identifies the candidate inhibitor substance as an inhibitor of cardiac hypertrophy. The histone deacetylases may be purified away from whole cells or located in an intact cell. The cell may be a myocyte, such as a cardiomyocyte. Measuring HDAC activity may comprise measuring release of a labeled acetyl group from a histone. The label may be a radiolabel, a fluorescent label or a chromophore. [0016] The class I histone deacetylase may be HDAC1, HDAC2, HDAC3, or HDAC 8. The class II histone deacetylase may be HDAC4, HDAC5, HDAC6, HDAC7, HDAC9, or HDAC 10. The activity of more than one class I histone deacetylase may be measured. The activity of more than one class II histone deacetylase may be measured. The activity of more than one class I histone deacetylase and more than one class II histone deacetylase may be measured. The candidate inhibitor substance may have inhibitory activity against at least one class I histone deacetylase and have no activity against at least one class II histone deacetylase. The candidate inhibitor substance may have inhibitory activity against multiple class I histone deacetylases and have no activity against multiple class II histone deacetylases. The candidate inhibitor substance may have inhibitory activity against at least one class I histone deacetylase that is at least two-fold greater than its inhibitory activity against at least one class II histone deacetylase. The candidate inhibitor substance may have inhibitory activity against at least one class I histone deacetylase that is at least five-fold greater than its inhibitory activity against at least one class II histone deacetylase. BRIEF DESCRIPTION OF THE DRAWINGS [0017] The following drawings form part of the present specification and are included to further demonstrate certain aspects of the present invention. The invention may be better understood by reference to one or more of these drawings in combination with the detailed description of specific embodiments presented herein. [0018] FIGS. 1A-C. TSA alters agonist-induced gene repression. Cultured cardiac myocytes were treated with PE (20 .mu.mol/L) or IL-1 (1 ng/mL) for 48 hrs. TSA (30 nmol/L) was added 30 min. prior to treatment with PE or IL-1. Myocyte-specific mRNA expression (SERCA2a (FIG. 1A), .alpha.MyHC (FIG. 1B), .beta.MyHC (FIG. 1C) was assayed in 5 .mu.g total RNA by RNase protection assay. Mean data are from 4 cultures, and are presented as % of control after normalization to GAPDH signal. [0019] FIGS. 2A-D. Over-expression of HDACs repress muscle-specific promoters. Cultured myocytes were transfected for 72 hrs. with expression vectors (2 .mu.g per .about.3.times.105 cells) for Flag-tagged HDAC1, 4, 5 (or its backbone vector) and CAT reporter (5 .mu.g) constructs for SERCA (FIG. 2A), .alpha.MyHC (FIG. 2B), .beta.MyHC (FIGS. 2C and 2D) genes, plus SV40-driven secreted alkaline phosphatase (SEAP 1 .mu.g, Clontech). TSA was used at 30 nmol/L, and added just after the transfection. PE 22 (20 .mu.mol/L) was added 24 hrs. later and CAT assays performed after an additional 48 hours. Mean data are from n=3 different cultures, and are presented as % of pCMV after normalization to SEAP activity in the media. Over-expression of HDAC was confirmed by Western blot for Flag. [0020] FIGS. 3A-D. HDAC inhibitors block the activation of ANF reporter by phenylphrine without cytotoxicity. Neonatal rat cardiomyocytes were co-transfected with a total of 1 .mu.g of the mouse 3 kb ANF promoter fragment and CMV-Lac Z plasmids. (FIG. 3A) The ANF promoter is minimally active in unstimulated cardiomyocytes. The addition of HDAC inhibitors does not induce ANF promoter activity. Addition of phenylphrine activated the ANF promoter, but co-treatment of cardiomyoyctes with phenylephrine and a HDAC inhibitor (TSA (85 nM), NaBut (5 mM), or HC-toxin (5 ng/ml)) prevented the activation of the ANF promoter by phenylphrine (100 .mu.M). (FIG. 3B) Lac Z expression by the constitutive promoter CMV. Treatment with HDAC inhibitors augmented CMV activity with and without phenylephrine co-treatment. (FIGS. 3C and 3D) The graphs show the measurements of adenylate kinase activity remains constant in the medium after X hours of culturing cardiomyocytes in the absence or presence of hypertrophic stimulants FBS, PE or ET-1. [0021] FIGS. 4A-C. HDAC inhibitors prevent endogenous ANF expression normally induced by hypertrophic agonists. (FIG. 4A) Graph shows the summation of several dot blot experiments (n=4). As in the transfection experiments, phenylephrine (100 .quadrature.M) induces ANF expression over three-fold. Treatment with HDAC inhibitors (TSA 85 nM; sodium butyrate, 5 mM; HC-toxin 5 ng.ml) blocks the accumulation of ANF message. (FIGS. 4B and 4C) The graphs show a reduction of the chemiluminent detection of ANF in the culture medium with increasing concentrations of the HDAC inhibitors TSA (FIG. 4B) and sodium butyrate (FIG. 4C) when co-cultured with the hypertrophic stimulants FBS, PE or ET-1. [0022] FIGS. 5A-B. Treatment of cardiomyocytes with HDAC inhibitors blocks the fetal gene program associated with cardiomyocyte hypertrophy. (FIG. 5A) Graph shows fold changes in .alpha.SK-actin expression by phenylephrine and the lack of gene activation in TSA-treated samples. (FIG. 5B) Graph shows the fold changes of .alpha.MyHC and .beta.MyHC RNA expression. Phenylephrine treatment induces the activation of .alpha.MyHC expression (fetal gene) in cardiomyocytes; whereas, phenylephrine alone does not active the .alpha.MyHC gene, the adult isoform. Treatment with TSA prevented the activation of .beta.MyHC but stimulated the expression of .alpha.MyHC. The graphs represent three or more independent experiments Continue reading about Inhibition of histone deacetylase as a treatment for cardiac hypertrophy... Full patent description for Inhibition of histone deacetylase as a treatment for cardiac hypertrophy Brief Patent Description - Full Patent Description - Patent Application Claims Click on the above for other options relating to this Inhibition of histone deacetylase as a treatment for cardiac hypertrophy patent application. ###  How KEYWORD MONITOR works... a FREE service from FreshPatents1. Sign up (takes 30 seconds). 2. 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