HK1111346A - Use of ppar agonists for the treatment of congestive heart failure - Google Patents

Description

Use of PPAR agonists for the treatment of congestive heart failure

Congestive heart failure is a devastating disease in which the heart fails to pump enough blood to cause the accumulation of blood flow in the lungs. Typical symptoms include shortness of breath, difficulty breathing when lying down and swelling of the legs and ankles. The progressive impairment of physical performance can eventually lead to death. There are many causes of heart failure, but the most common are myocardial infarction (about 60% of all cases), chronic hypertension (about 25%), genetic predisposition (10%) and cardiomyopathy, or a combination of these factors.

In patients, the severity of CHF is classified according to clinical symptoms based on a classification method established by the New York Heart Association (NYHA). The patient's physical performance can be determined as NYHA class I (no symptoms), NYHA class II (symptoms at moderate exertion), NYHA class III (symptoms at mild exertion), or NYHA class IV (symptoms at rest).

Current CHF treatments that slow down the progression of CHF greatly prolong patient survival. However, overall mortality remains high for any given NYHA stage, with a recent average annual mortality of 15% in large-scale trials conducted in predominantly NYHHA class II and III patients. A common drawback of all currently approved CHF drugs (e.g., diuretics, ACE inhibitors, beta-blockers) that have proven beneficial for mortality is their hypotensive effect. Combination therapy is often not possible because many patients have too much reduced blood pressure. Therefore, alternative treatment strategies directed to new mechanisms of action are urgently needed to further improve the drug treatment of CHF.

Peroxisome proliferator-activated receptors (PPARs) represent a nuclear hormone receptor, two of which (PPAR α and PPAR γ) are expressed in many tissues, including the myocardium and the blood vessels. Activation of the PPARs results in the expression of a large number of genes and the consequent production of proteins. PPAR γ activators (e.g., rosiglitazone) have been approved for the treatment of type II diabetes based on their effectiveness in increasing sensitivity to insulin and delaying the progression of insulin resistance to overt diabetes (Malinowski and Bolesta, Clin. therapeutics, (2000), 22, 1151-. In addition to some PPAR α activators, fibrates are used clinically because they are capable of lowering blood cholesterol levels (Sacks-FM, am.J. Cardiol. (2001), 88(12A), 14N-18N). Novel PPAR α activators, structurally different from fibrates and more potent, are clinically developed for lipid disorders and diabetes (Inoue and Katayama, current drug targets: Cardiovasular & Haematologica disorders (2004), 4, 35-52).

In failing myocardium, metabolic disturbances are accompanied by a shift from fatty acid oxidation to glucose oxidation. This effect results in a reduction in the efficiency of myocardial productivity, which in turn results in a loss of contractile function in CHF. In the aging rat model, CHF was improved by physical training, accompanied by normalization of PPAR α expression in myocardium.

In addition to their metabolic effects, little is known about the direct effects of PPAR activators on the heart. In vitro in neonatal cardiomyocytes, the PPAR α agonist fenofibrate and the WY14,643 and the PPAR γ activator rosiglitazone are able to prevent endothelin-1 from inducing myocardial hypertrophy. Similarly, PPAR γ activators reduce cardiac hypertrophy induced by mechanical stress in isolated cardiomyocytes. In the arterial hypertension model, both PPAR α and PPAR γ activation can reduce cardiac fibrosis. In mouse models of acute myocardial ischemia and reperfusion, PPAR α and PPAR γ activation has been shown to reduce myocardial infarction size. PPAR γ activators have been shown to improve myocardial remodeling and heart failure symptoms in the chronic phase following myocardial infarction (Liang et al, Endocrinology 2003, 144: 4187-. On the other hand, there is evidence that PPAR γ agonists may exacerbate heart failure in type II diabetic patients.

Thus, the benefits of PPAR γ activation in CHF are controversial, and there is no data on the role of selective activation of PPAR α in CHF.

One embodiment of the present invention is the use of a compound of formula (I) or a pharmaceutically acceptable salt or physiologically functional derivative thereof for the manufacture of a medicament for the treatment of Congestive Heart Failure (CHF).

Another embodiment is the use of a compound of formula (II) or a pharmaceutically acceptable salt or physiologically functional derivative thereof for the manufacture of a medicament for the treatment of Congestive Heart Failure (CHF).

Preferred compounds of formula (II) are compounds of formula (III).

The compound of formula (I) is prepared according to example 5 of International patent application WO 2004/085377 and the compounds (II) and (III) are prepared according to examples I and II of International patent application WO 03/020269.

Pharmaceutically acceptable salts are particularly useful for medical applications because of their higher solubility in water than the original or base compound. These salts must have a pharmaceutically acceptable anion or cation. Suitable pharmaceutically acceptable acid addition salts of the compounds of the invention are salts of inorganic acids such as hydrochloric acid, hydrobromic acid, phosphoric acid, metaphosphoric acid, nitric acid and sulfuric acid, and salts of organic acids such as, for example, acetic acid, benzenesulfonic acid, benzoic acid, citric acid, ethanesulfonic acid, fumaric acid, gluconic acid, glycolic acid, isethionic acid, lactic acid, lactobionic acid, maleic acid, malic acid, methanesulfonic acid, succinic acid, p-toluenesulfonic acid and tartaric acid. Suitable pharmaceutically acceptable base salts are ammonium salts, alkali metal salts (e.g. sodium and potassium salts), alkaline earth metal salts (e.g. magnesium and calcium salts).

The term "physiologically functional derivative" as used herein refers to any physiologically tolerable derivative of a compound of formula (I) of the invention, e.g. an ester, which on administration to a mammal such as, for example, a human, is capable of forming (directly or indirectly) a compound of formula (I), (II) or (III) or an active metabolite thereof. Physiologically functional derivatives also include prodrugs of the compounds of the invention, such as, for example, those described in H.Okada et al, chem.pharm.Bull.1994, 42, 57-61. Such prodrugs can be metabolized in vivo to the compounds of the invention. These prodrugs may themselves be active or inactive.

The compounds of the present invention may also exist in various polymorphic forms, such as amorphous and crystalline polymorphic forms. All polymorphic forms of the compounds of the invention are within the scope of the invention and are a further aspect of the invention.

The amount of a compound of formula (I), (II) or (III) necessary to achieve a desired biological effect depends on many factors, such as the particular compound selected, the desired application, the mode of administration and the clinical condition of the patient. The daily dosage is generally from 0.3mg to 100mg (generally from 3mg to 50mg) per kg body weight per day, for example from 3 to 10 mg/kg/day. The intravenous dose may be, for example, from 0.3mg to 1.0mg/kg, which may suitably be administered as an infusion of from 10ng to 100ng per kg per minute. Suitable infusion solutions for these purposes may be present in an amount of, for example, about 0.1ng to 10mg per ml, typically about 1ng to 10 mg. Single doses may contain, for example, from about 1mg to 10g of active ingredient. Thus, ampoules for injection may contain, for example, from 1mg to 100mg, and single-dose preparations which can be administered orally, such as capsules or tablets, may contain from 1.0 to 1000mg, typically from 10 to 600 mg. For the treatment of the above-mentioned conditions, the compounds of formula (I), (II) or (III) may be used as such, but are preferably in the form of a pharmaceutical composition with a pharmaceutically acceptable carrier. Of course, the carrier must be acceptable in the sense that it is compatible with the other ingredients of the composition and not deleterious to the health of the patient. The carrier may be a solid and/or liquid and is preferably formulated with the compound as a single dose, for example as a tablet, which may contain from 0.05% to 95% by weight of the active ingredient. Other pharmaceutically active substances, including other compounds of formula I, may also be present. The pharmaceutical compositions of the present invention may be prepared by one of the known pharmaceutical methods which essentially consist of mixing the ingredients together with pharmacologically acceptable carriers and/or excipients.

The pharmaceutical compositions of the invention include those suitable for oral, rectal, topical, peroral (e.g. sublingual) and parenteral (e.g. subcutaneous, intramuscular, intradermal or intravenous) administration, although the most suitable mode of administration will depend on the nature and severity of the condition being treated and on the nature of the compound of formula (I), (II) or (III) used in each case. Coated formulations and coated sustained release formulations are also within the scope of the invention. Acid and gastric juice resistant formulations are preferred. Suitable coatings resistant to gastric juices include cellulose acetate phthalate, polyvinyl acetate phthalate, hydroxypropyl methylcellulose phthalate and anionic polymers of methacrylic acid and methyl methacrylate.

Suitable pharmaceutical compounds for oral administration may be in the form of individual units such as, for example, capsules, wafers or tablets, each containing a defined amount of a compound of formula (I), (II) or (III); may be in the form of a powder or granules, in the form of a solution or suspension in an aqueous or non-aqueous liquid; or may be in the form of an oil-in-water or water-in-oil emulsion. As already mentioned, these compositions may be prepared by any suitable pharmaceutical method which includes a step in which the active compound is contacted with a carrier, which may consist of one or more additional ingredients. The compositions are generally prepared by uniformly mixing the active compound with a liquid and/or finely divided solid carrier, and thereafter, if desired, shaping the product. Thus, for example, tablets may be prepared by compressing or molding a powder or granules of the compound (and where appropriate one or more additional ingredients). Compressed tablets may be prepared by compressing in a suitable machine the compound in a free-flowing form such as, for example, a powder or granules, which may be mixed with binders, glidants, inert diluents and/or surfactant (s)/dispersing agents where appropriate. Molded tablets may be prepared by molding the compound in powder form and moistened with an inert liquid diluent in a suitable machine.

Pharmaceutical compositions suitable for oral (sublingual) administration include tablets containing a compound of formula (I), (II) or (III) and a flavouring agent, typically sucrose, and acacia or tragacanth, and pastilles comprising the compound in an inert base such as gelatin and glycerin, or sucrose and acacia.

Suitable pharmaceutical compositions for parenteral administration may include sterile aqueous preparations of a compound of formula (I), (II) or (III) which are isotonic with the blood of the intended recipient. These formulations may be administered intravenously, but may also be administered by subcutaneous, intramuscular or intradermal injection. These formulations can be prepared by mixing the compound with water and rendering the resulting solution sterile and isotonic with blood. The injectable compositions of the invention generally comprise from 0.1 to 5% by weight of active compound.

Pharmaceutical compositions suitable for rectal administration may be in the form of single dose suppositories. These formulations may be prepared by mixing a compound of formula (I) with one or more conventional solid carriers, for example cocoa butter, and shaping the resulting mixture.

Pharmaceutical compositions suitable for topical application to the skin may be in the form of ointments, creams, lotions, pastes, sprays, aerosols or oils. Suitable carriers are, for example, petrolatum, lanolin, polyethylene glycols, alcohols and combinations of two or more of these substances. The active compound is typically present at a concentration of about 0.1 to 15% by weight of the composition, for example about 0.5 to 2%.

Transdermal administration may also be performed. Pharmaceutical compositions suitable for transdermal application may be in the form of a single plaster suitable for prolonged contact with the epidermis of a patient. Such plasters suitably comprise the active ingredient dissolved and/or dispersed in an adhesive or dispersed in a polymer in a suitably buffered aqueous solution. Suitable active ingredient concentrations are about 1% to 35% by weight, or about 3% to 15%. The active compound may be, for example, as in Pharmaceutical Research, 2 (6): 318(1986) by electrotransport or iontophoresis.

To test the effect of specific activation of PPAR γ (rosiglitazone) or PPAR α (compounds of formulae (I), (II) and (III)) on the improvement of CHF, a rat model of chronic coronary artery ligation was used, since myocardial infarction is the most common cause of CHF in industrialized countries.

Activation of PPAR α is shown herein to be beneficial in congestive heart failure. With respect to myocardial function, systolic and diastolic LV function-and hence cardiac output affected-is improved upon treatment with a compound of formula (I), (II) or (III) other than a PPAR γ activator.

Activation of PPAR α also improves pulmonary congestion as evidenced by normalization of right ventricular and lung weights (lower lung weight may be indicative of better cardiac function, higher lung weight indicates pulmonary congestion, which is often caused by decreased cardiac function (i.e., congestion).

In previous studies it has been reported that activation of PPAR γ and PPAR α reduces myocardial hypertrophy, reduces fibrosis in mineralocorticoid dependent hypertension, and limits myocardial infarct size in an acute ischemia reperfusion model in vitro (see above). Activation of these two PPAR subtypes is therefore not expected to play opposite roles in post-myocardial infarction heart failure. One reason for this differential finding may be the tissue-specific differential expression of the two PPAR subtypes.

Abbreviations and acronyms:

CHF ═ congestive heart failure

LV left ventricle/ventricle

MI ═ myocardial infarction

PPAR (peroxisome proliferator-activated receptor)

SEM-mean standard error

Example 1: proof of concept study for the treatment of congestive heart failure Using Compounds of formula (I)

Male Wistar rats were housed 3 per cage, each under standardized conditions of temperature, humidity and light. Rats had free access to standardized feed (sodium content 0.2%, Altromin, Lage, germany) and drinking water. Induction of chronic heart failure is achieved by permanent occlusion of the left coronary artery approximately 2mm distal to the origin of the aorta, resulting in a large infarct of the free left ventricular wall. Chronic treatment begins on the day after myocardial infarction is produced and continues for 8 weeks.

At the end of the treatment period, cardiac function was determined using ex vivo working heart specimens (modified Langendorff instrument, ref Linz et al, j.ren.angiotensin Aldosterone syst, 2003): heart perfusion was performed by Langendorff with oxygenated (95% O)₂，5％CO₂) Non-circulating Krebs-Henseleit solution (mmol/L) of the following composition: NaCl, 118; KCl, 4.7; CaCl₂，2.5；MgSO₄，1.6；NaHCO₃，24.9；KH₂PO₄1.2; glucose, 5.5; sodium pyruvate, 2.0. The left atrium is cannulated by the incision in the left atrial appendage. After an equilibrium period of 15min at a fixed perfusion pressure of 60mmHg, the heart was switched to the operating mode at a fixed filling pressure of 11 mmHg. Thereafter, the afterload pressure was gradually increased from 40mm Hg to 140mmHg at intervals of two minutes. The data are representative and are given a constant afterload pressure of 80mm Hg. The flow and pressure signals were sampled at 500Hz, on average every 2 seconds. Cardiac output measures the total volume of blood pumped by the heart to the body. LVdP/dtmax is an indicator of myocardial contractile force, i.e., the ability of the heart to produce force; LV dP/dtmin is an indicator of cardiac relaxation ability. In addition, lung weight is an indicator of measured pulmonary congestion, which is an indirect indicator of CHF.

Treatment with the compound of formula (I) was started on the day after myocardial infarction (pressed in feed at a dose of 20 mg/kg/d). Rosiglitazone, a PPAR γ agonist alone, was used as a control in another group (3 mg/kg/day, added to feed).

Chronic treatment with PPAR α activators of formula (I) improves different aspects of heart failure, whereas activation of PPAR γ (with rosiglitazone) does not. All of these data provide a strong rationale for the beneficial effects of the compounds of formula (I) in the treatment of congestive heart failure.

Table 1:

test conditions	Lung weight gram	Cardiac outputAmount of mL/min	LVdP/dtmaxmmHg/s	LVdP/dtminmmHg/s
Sham (No MI, untreated)						1.88±0.10*	37.3±3.5*	5780±191*	3527±217*
	MI placebo	2.96±0.40	18.9±2.9	3748±176	2119±75
MI rosiglitazone 3mg/kg/d						3.67±0.38*	10.2±3.8*	3491±147	2081±65
	MI Compound (I)20mg/kg/d	1.62±0.06*	25.8±4.3*	4614±253*	2502±77*

Data are presented as mean ± mean standard error. Each group N is 6-12.^*P < 0.05 compared to placebo. Cardiac output, LV dP/dtmax and LV dP/dtmin were measured at a afterload of 80mmHg (working heart).

Example 2: dose response in chronic myocardial infarction

Male Sprague Dawley rats were pretreated with chronic ligation of the left coronary artery in order to cause Myocardial Ischemia (MI) and subsequent development of heart failure. Treatment was initiated on the day following myocardial ischemia with the compound of formula (I) (pressed in the feed, resulting in different daily doses). After 8 weeks of treatment, the animals were sacrificed, the lungs weighed, and the ex vivo heart function was analyzed in the same manner as described in example 1 (see above) in the mode of working heart. The method can evaluate different aspects of myocardial function. For comparison with the role of the established therapeutic principles in the current trial series, another group used known ACE/NEP dual inhibitors or vasopeptidase inhibitors (7- (2-acetylthio-3-methyl-butyrylamino) -6-oxo-1, 2, 3, 4,6, 7, 8, 12 b-octahydro-benzo [ c ] pyrido [1, 2-a ] azepine  -4-carboxylic acid, international patent application No. WO 02/083671) which are active in the treatment of CHF.

Chronic treatment with compounds of formula (I) ameliorates different aspects of heart failure. Together, these data demonstrate the beneficial effect of the compounds of formula (I) in congestive heart failure.

Table 2:

test conditions	Lung weight g/100g BW	Cardiac output mL/min	LV dP/dtmaxMm Hg/s	LV dP/dtminmm Hg/s
					Sham (No MI, untreated)	0.39±0.01*	36.0±2.7*	5807±192*	2985±109*
MI placebo	0.71±0.07	7.3±1.8	3170±247	2056±138
					MI Compound (I)1mg/kg/d	0.80±0.08	12.5±3.3	3280±250	1886±137
MI Compound (I)3mg/kg/d	0.54±0.08*	21.5±5.4*	3665±166*	2353±77*
					MI Compound (I)10mg/kg/d	0.54±0.09*	21.0±4.0*	4043±256*	2339±111*
MI VPI30mg/kg/d	0.56±0.07*	27.2±3.5*	3868±172*	2379±120*

Data are presented as mean ± mean standard error. Each group N is 6-12.^*P < 0.05 compared to placebo. Cardiac output, LV dP/dtmax and LV dP/dtmin were measured at a afterload of 80mmHg (working heart).

Example 3: agonist effectiveness of the compound of formula (I)

The agonist activity of the compounds of formula (I) is determined according to WO 03/020269 as follows: to analyze the effectiveness of substances that bind to human PPAR α and activate it in an agonist manner, a stably transfected HEK cell line known as a "PPAR α reporter cell line" (HEK ═ human embryonic kidney) was used.

PPAR α agonist activity was determined in a 3 day assay as follows:

day 1: PPAR α reporter cell lines were cultured to 80% confluency in DMEM medium (Life Technology) mixed with the following supplements: 10% cs-FCS (fetal calf serum, Hyclone), antibiotics (0.5mg/ml zeocin [ Invitrogen)]0.5mg/ml G418(Life Technology), 1% penicillin-streptomycin solution [ Life Technology]) And 2mM L-glutamine (Life Technology). Said culturing is carried out in a standard cell culture flask (Becton Dickinson) in a cell culture incubator at 37 ℃ in the presence of 5% CO₂Is carried out in the case of (1). The 80% confluent cells were washed once with 30ml PBS (Life Technology), treated with 2ml trypsin solution (Life Technology) at 37 ℃ for 2 minutes, absorbed in 5ml of the above medium and counted in a cell counter. After dilution to 500000 cells/ml, 100000 cells were seeded in each case into the individual wells of a 96-well microtiter plate (Corning Costar) with a transparent plastic bottom. These plates were placed in a cell culture incubator at 37 ℃ and 5% CO₂Cultured under the conditions of (1) for 24 hours.

Day 2: the PPAR α agonist to be tested was dissolved in DMSO at a concentration of 10 mM. This stock was diluted in phenol red free DMEM medium (Life Technology) mixed with 5% cs-fcs (hyclone), 2mM L-glutamine (Life Technology) and the antibiotics previously described (zeocin, G418, penicillin and streptomycin). The test substances were tested at 11 different concentrations (10. mu.M; 3.3. mu.M; 1. mu.M; 0.33. mu.M; 0, 1. mu.M; 0.033. mu.M; 0.01. mu.M; 0.0033. mu.M; 0.001. mu.M; 0.00033. mu.M and 0.0001. mu.M). More potent compounds were tested in a concentration range of 1. mu.M to 10pM or 100nM to 1 pM. The culture medium in the PPAR α reporter cell line inoculated on day 1 was completely removed by aspiration and the test substance diluted with the culture medium was immediately added to these cells. Both dilution and addition of the test substances were carried out with an automatic apparatus (robot) (Beckman Biomek 2000). The final volume of test substance diluted in culture medium was 100. mu.l per well of a 96-well microtiter plate. In said assay, the concentration of DMSO is below 0.1% v/v to avoid the cytotoxic effect of the solvent. Adding a standard P to each platePAR alpha agonists (which were also diluted to 11 different concentrations) were used to demonstrate that the assay was effective in each plate. These test panels were placed in an incubator at 37 ℃ and 5% CO₂Cultured under the conditions of (1) for 24 hours.

Day 3: the PPAR α receptor cells treated with the test substance were removed from the incubator and frozen at-20 ℃ for 1 hour to promote cell lysis. After thawing the plates (at room temperature for at least 30min), 50. mu.l of buffer 1(Luc-Screen kit # LS1000, PE Biosystems Tropix) were pipetted into each well and the plates were transferred to a fluorescence measuring device equipped with a pipetting unit (Luminoscan Ascent, LabSystems). Transferring 50. mu.l of buffer 2(Luc-Screen kit # LS1000, PEbiosystems Tropix) to each well of a 96-well plate initiated the luciferase reaction performed in the measuring instrument. The addition of buffer to each well was performed at defined, equal intervals according to the manufacturer's instructions (LabSystems). All samples were measured exactly 16min after addition of buffer 2. The measurement time for each sample was 10 seconds.

The PPAR α agonist activity of compounds (II) and (III) and the PPAR γ agonist activity of compounds (I), (II) and (III) were determined in a similar manner.

Claims (3)

1. Use of a compound of formula (I) or a pharmaceutically acceptable salt or physiologically functional derivative thereof for the manufacture of a medicament for the treatment of Congestive Heart Failure (CHF)

2. Use of a compound of formula (II) or a pharmaceutically acceptable salt or physiologically functional derivative thereof for the manufacture of a medicament for the treatment of Congestive Heart Failure (CHF)

3. Use according to claim 2, characterized in that the compound (II) is a compound of formula (III)