Claims
- 1. A pyridone of the following formula (1): ##STR8## wherein: R.sup.1 and R.sup.2 represent, independently, hydrogen, C.sub.1-3 alkyloxy, cyano, halogen trifluoromethyl, C.sub.1-6 alkyl, c.sub.1-6 alkylsulfonyl, C.sub.1-6 alkyloxy-C.sub.1-6 -alkyl, C.sub.3-7 cycyloalkyl-C.sub.1-6 -alkyloxy-C.sub.1-6 -alkyl, nitro, hdyroxy, C.sub.2-3 alkenyoxy, amno or amino substituted by one or two C.sub.1-6 alkyl groups;
- L represents a linking moiety of the following formula (III) ##STR9## in which; R.sup.10 -R.sup.11 represents, independently, hydrogen or methyl;
- p represents the integer 2, 3, 4, 5 or 6, or a pharmaceutically acceptable acid addition salt thereof.
- 2. The pyridone of claim 1, wherein L is a moiety of the formula (III).
- 3. The pyridone of claim 1, wherein R.sup.10 and R.sup.11 are hydrogen.
- 4. The pyridone of claim 1, wherein R.sup.1 is hydrogen.
- 5. The pyridone of claim 1, wherein R.sup.2 is cyano, chlorine or methyl.
- 6. The pyridone of claim 1, wherein R.sup.1 is hydrogen and R.sup.2 is other than hydrogen and is substituted at the 2-position of the phenyl ring.
- 7. The pyridone of claim 1, wherein aid pyridone has the S configuration at the 2-hdyroxy position shown in formula (I).
- 8. A pharmaceutical composition comprising an effective amount of a pyridone of claim 1 and a pharmaceutically acceptable diluent or carrier.
- 9. A method for the treatment of congestive heart failure which comprises administering to a patient in need of such an effective amount of the pharmaceutical composition of claim 8.
- 10. 5-{4-phenyl}-6-methyl -2-oxo-1,2-dihydro-3-pyridinecarbonitrile.
- 11. 5-{4-phenyl}-6-methyl-2-oxo-1,2-dihydro-3-pyridinecarbonitrile.
BACKGROUND OF THE INVENTION
This is a division of U.S. Ser. No. 07/565,297 filed Aug. 9, 1990, and issued as Pat. No. 5,051,431 on Sep. 24, 1991, which is a continuation-in-part of U.S. Ser. No. 07/411,065 filed Sep. 22, 1989 now abandoned.
Congestive heart failure (CHF) is the disease state wherein a weakened heart results in the inability to adequately pump blood throughout the body. CHF is a common cause of death in the hospital and is an expensive and time consuming condition to treat. Positive inotropic pharmaceuticals such as amrinone act by increasing the force of contraction of the heart without increasing heart rate and have been proposed to treat CHF. Presumably these agents produce their cardiotonic effects at least partly through inhibition of type III phosphodiesterase.
Beta-blockers such as atenolol and propranolol may be given to persons who have suffered a heart attack in order to lessen oxygen consumption by the heart and prevent sudden death. However, if there is significant damage to the heart, there may be a lack of ability to pump forcefully and the negative inotropic effects of a beta-blocker may exacerbate an already dangerous situation.
Propanolamines having a heterocyclic moiety which may be pyridine are set forth in U.S. Pat. No. 4,608,383. Hydroxyalkylaminoalkyl substituted salicylamides having beta blocking or beta-stimulating activities are taught in European Patent 39,892 published Nov. 18, 1981. N-Heterocyclyl amines as beta agonists are taught in European Patent 236,624 published Sep. 16, 1987.
European Patent 178,189 published Apr. 16, 1986 teaches pyridazinones having a phenyl group at the 6-position. Pyridazinones having an alkylaminophenyl group at the 6-position are taught in European Patent 259,835 published Mar. 16, 1988.
This invention relates to novel pyridones of the following formula (I) or salts thereof as well as pharmaceutical compositions, methods for their use in treating cardiovascular conditions, processes used in syntheses and intermediates used in such processes.
This invention relates to novel chemical compounds and pharmaceutical compositions thereof. The subject chemical compounds are 5-phenyl-2-pyridones of the formula (I): ##STR2## wherein: R.sup.1 and R.sup.2 represents, independently, hydrogen, alkyloxy, morpholino, cyano, halogen, trifluoromethyl, alkyl, alkylsulfonyl, alkyloxyalkyl, cycloalkylalkyloxyalkyl, nitrok hdyroxy, alkenyloxy, amino or amino substituted by one or two alkyl groups;
As used herein, "lower alkyl" per se or as part of another group such as lower alkoxy may be 1 to 3 carbons, straight or branched chain; "alkyl" may be of about 1 to 6 carbons, straight or branched chain "cycloalkyl" may be 3 to 7 carbons; "independently" indicates that members, where two or more are present, need not be identical as in the definitions of R.sup.1 and R.sup.2 or the various possibilities for R.sup.3 when n is 2 or 3; "halogen" is fluoro, chloro, bromo, or iodo; the L group is attached as shown in the definition i.e. the carbon carrying R.sup.3 and R.sup.4 in formula (II) is attached to the left most oxygen of --OLNH--of formula (I) rather than the nitrogen; the wavy lines in formulae (II) and (III) indicated the bond of attachment of L; and morpholino may be attached via the nitrogen or any ring carbon. Placement of the --O--L--moiety on the phenyl ring in formula (I) which is attached to the 5-position of the pyridone may be at any of the 2-, 3- or 4-positions.
Particular compounds of this invention are those of formula (I) with one or more of the following definitions: L is the linking group of formula (II); n is 1 or 3; R.sup.3, R.sup.4, R.sup.5, R.sup.6, R.sup.7, and R.sup.11 are hydrogen; R.sup.8 and R.sup.9 are methyl groups; R.sup.1 is hydrogen, and R.sup.2 is a cyano, chlorine, or methyl substituted at position 2 of the phenyl ring. The --O--L--moiety is particularly at the 4-position of phenyl ring which is, in turn, attached to the 5-position of the pyridone ring.
The compounds of formula (I) contain a basic nitrogen atom and hence can form pharmaceutically acceptable acid addition salts. A wide variety of acids may be employed to form such salts and representative examples of such acids include inorganic acids, e.g. hydrochloric acid, hydrobromic acid, hydroiodic acid, phosphoric acid, nitric acid, and sulfuric acid; and organic acids, e.g. maleic acid, fumaric acid, acetic acid, benozoic acid, p-toluenesulfonic acid, tartaric acid, citric acid, succinic acid, lactic acid, and propionic acid. These acid addition salts are prepared by conventions methods. Compounds of the formula (I) may also exist as a solvate, e.g. a hydrate or hemihydrate, and such are within the scope of the invention.
The compounds of formula (I) have one or more asymmetric carbon atoms in their structure and consequently they may exist in different optical isomeric forms or mixtures, e.g. racemates or mixtures of diastereomers. Enantiomeric forms and mixtures of such forms may be obtained separate by application of methods of resolution known to those skilled in the art such as, for example, salt formation with an optically active acid followed by selective crystallization or chiral derivatization followed by selective crystallization or silica gel chromatography. All stereoisomeric forms of the compounds of formula (I) including mixtures of diastereomers, pure diastereomers enantiomers, and mixtures thereof are understood to be within the scope of this invention.
The compounds of formula (I) in which the linking moiety L is of the formula (II) may be prepared as shown in Scheme I. ##STR4## In Scheme I, the various R groups, except R.sup.12, R.sup.13 and R.sup.14, and n are as defined for formula (I).
The compounds of formula (IV) employed as starting materials in Scheme I may be prepared by the methods described by G.Y. Lesher et al, U.S. Pat. No. 4,465,686. In the compounds of formula (V), Y represents a leaving group that is reactive toward displacement by nucleophiles. Suitable Y groups include halogen or the p-toluenesulfonate ester, p-nitrobenzenesulfonate ester, methansulfonate ester and trifluoromethanesulfonate ester. R.sup.12 in formula (V) represents a lower alkyl group. The compounds of formula (V) are known compounds or may be prepared from known compounds by conventional methods.
Step 1 of Scheme I is effected by reacting the phenol of formula (IV) with a compound of formula (V) in the presence of a suitable base and appropriate solvent to give compounds of formula (VI). Bases which may be used include sodium hydride, sodium t-butoxide, and similar non-nucleophilic basic reagents. The preferred base is sodium hydride.
A wide variety of solvents may be used in Step 1 of Scheme I with the only restriction being that the solvent be inert toward starting materials (IV) and (V) as well as to the basic reagent and the product (VI). Suitable solvents include DMF, dimethylsulfoxide, aromatic hydrocarbons such as benzene or toluene, and ethers such as tetrahydrofuran. Step may be conducted over a wide temperature range, with the preferred temperature being about 0.degree. C. to 80.degree. C. The preferred conditions for effecting Step 1 of Scheme I is to use sodium hydride as the base, dimethylformamide as the solvent, and a temperature of 0.degree. C. with gradual warming to 80.degree. C. Under these conditions, Step 1 is completed in 2-4 hr.
Step 2 of Scheme I is effected by heating the ester of formula (VI) in an aqueous solution containing a suitable base. Suitable bases which may be used include sodium hydroxide, potassium hydroxide, potassium carbonate, and the like. Suitable co-solvents with water for use in Step 2 include alcohols, e.g. methanol, ethanol, and propanol ethers, e.g. tetrahydrofuran and dioxane, and dimethylsulfoxide. Step 2 may be conducted over a wide temperature range, with the preferred conditions for effecting Step 2 of Scheme I being to use potassium hydroxide as the base, 1:1 water-ethanol as the solvent, and a temperature of 80.degree. C. Under these conditions, Step 2 is completed in 2-4 hr.
In the compound of formula (VII), X is a leaving group such as hdyroxy whereby (VII) is a carboxylic acid. Alternatively, the compound of formula (VII) where X is hdyroxy may be converted to a suitable reactive derivative which is then reacted with an amine of formula (V). Suitable reactive derivatives of the carboxylic acid (VII) include: acid halides, such as the acid chloride; mixed anhydrides of the carboxylic acid with another organic acid, such as acetic acid, propionic acid, or pivalic acid whereby X is --OCOR where R is an organic moiety such as alkyl; acyl imidazoles; and active esters of carboxylic acid, such as the 4-nitrophenyl ester. With the exception of the acyl imidazole, these reactive derivatives can be prepared by treating the carboxylic acid with a suitable halogen compound, such as thionyl or oxalyl chloride, acetyl chloride, pivaloyl chloride, or isobutoxycarbonyl chloride in the presence of a proton acceptor and an inert solvent. Suitable proton acceptors include both organic bases such as triethylamine or 4-dimethylaminopyridine and inorganic bases such as anhydrous potassium carbonate. Suitable solvents for forming reactive derivatives of (VII) where X.dbd.OH include diethyl ether, tetrahydrofuran, aromatic hydrocarbon solvents such as benzene or toluene, methylene chloride, and acetonitrile. Acyl imidazoles can be prepared from (VII) where X.dbd.OH by reaction of (VII) with 1,1'-carbonyldiimidazole. Thus, X in formula (VII) may specifically be hdyroxy, chloro, acetoxy, propionoxy, pivaloxy, isobutoxycarbonyloxy or an imidazole group.
In the compounds of formula (VIII) and (IX) in Scheme I, R.sup.13 represents hydrogen and R.sup.14 represents any of several monovalent amine protecting groups including, but not limited to carbamates, e.g. --CO.sub.2 C'(CH.sub.3).sub.3 or --CO.sub.2 CH.sub.2 CH.sub.3, or N-benzyl derivatives, e.g. benzyl, or R.sup.13 and R.sup.14 together represent a divalent amine protecting group such as phthalimide, e.g. by reaction of the free amine with phthallic anhydride. Detailed examples of the use and removal of these amine protecting groups are described by T.W. Greene in Protective Groups in Organic Synthesis, John Wiley & Sons, 1981, pp. 218-323.
Depending upon the definition of R.sup.6, R.sup.7, R.sup.8 and R.sup.9, an amine protecting group R.sup.14 or R.sup.13 and R.sup.14 may not be necessary. In those cases, Step 4 of Scheme I, removal of the amine protecting group is unnecessary and the intermediate (X) is prepared directly from (VII) via Step 3' with a diamine of the formula HR.sup.5 NC(R.sup.6 R.sup.7)C(R.sup.8 R.sup.9)NH.sub.2.
The amines of formula (VIII) are commercially available or may be prepared by conventional methods, for example, see the Journal of Medicinal Chemistry, 31, 898-901 (1988).
Steps 3 and 3'in Scheme I are coupling reactions and may be executed by treating a mixture of compounds (VII) and (VIII) in the presence of inert solvent with suitable dehydrating agents such as diethylcyanophosphonate or dicyclohexylcarbodiimide. The reaction may be carried out over a wide range of temperatures, with the preferred temperature being 0.degree. C. to 75.degree. C. Suitable solvents for the coupling reaction are tetrahydrofuran, acetonitrile, benzene, toluene, methylene chloride, chloroform, and DMF. The preferred conditions for effecting steps 3 or 3'is to use diethylcyanophosphonate as the coupling agent, dimethyformamide as the solvent, and a temperature of 0.degree. C. to 25.degree. C. Under these conditions the time required for reaction is about 1-12 hr.
Step 4 of Scheme I, removal of the amine protective group R.sup.14 or and R.sup.14, is executed under conditions appropriate to the particular amine protective group. A preferred amine protective group is the tert-butoxycarbonyl group. When R.sup.14 in Scheme I represents the tert-butoxycarbonyl group, Step 4 is effected by treating the compound of formula (IX) with an acid solution which may be either a mineral acid such as hydrochloric acid, hydrobromic acid, or sulfuric acid or an organic acid such as p-toluenesulfonic acid or trifluoroacetic acid. A wide range of solvents may be used for removal of the tert-butoxycarbonyl group as long as the solvent is stable to acids and does not react with the amine product, (X). Suitable solvents include the halogenated hydrocarbons, such as methylene chloride and chloroform, and aromatic solvents such as benzene and toluene. The reaction may be run over a wide range of temperatures, in particular in the temperature range of 0.degree. C. to 25.degree. C. The time required for the reaction is about 15 min to 2 hr and depends upon the solvent and temperature of the reaction. Removal of other amine protecting groups may be as set forth in the description below of Step 2 of Scheme IV.
Step 5 in Scheme I is effected by reacting an amine of formula (X) with an epoxide of formula (XI). Epoxides of the formula (XI) are either known compounds or can be prepared by conventional procedures well known to those skilled in the art of organic synthesis. Particular compounds of formula (I) are the enantiomers wherein the carbon bearing the hydroxyl group has the S-configuration. It may be thus of particular utility to utilize in Step 5 epoxides of the formula (XI) having the S-configuration at the asymmetric carbon, since these epoxides will yield structures having the S-configuration at the carbon bearing the hydroxyl group. Many of the examples of compounds of formula (I) prepared and tested have been mixtures of the 2S- and 2R-hdyroxy configurations and thus the invention covers all such stereoisomers. The desired S-epoxides are prepared as shown in Scheme II using the procedures described by K.B. Sharpless, J.M. Klunder and T. Onami in the Journal of Organic Chemistry, 1989, 54, 1295-1304. ##STR5##
In Scheme II, R.sup.1 and R.sup.2 are as defined above for formula (I). In Scheme II, a phenolic salt of the formula (XII) is reacted with a sulfonate of formula (XIII) at 0.degree. C. to 75.degree. C. to yield the epoxide (XI). The (2S)-(.+-.)-glycidyl 3-nitrobenzenesulfonate form of the compound of formula (XIII) shown in Scheme II is commercially available from the Aldrich Chemical Company or may be prepared by the methods cited in the Sharpless et al reference given above. Suitable salts for the reaction shown in Scheme II include the sodium and potassium salts, i.e. M+=Na+or K+The phenolic salts (XII) are prepared from the corresponding phenols which are known compounds that are commercially available or can be prepared by conventional methods.
A wide variety of solvents may be employed in Step 5 of Scheme I, with the only restriction being that the solvent must be inert with respect to the amines (X), epoxides (XI), and products (I). Suitable solvents for Step 5 include alcohols such as methanol, ethanol, or isopropanol; aromatic hydrocarbon solvents such as benzene or toluene, and ethers such as tetrahydrofuran or dioxane. The reaction in Step 5 may be run over a wide range of temperatures, in particular in the range of 25.degree. C. to 100.degree. C. The time required for the reaction of Step 5 is dependent upon temperature and the nature of the substituents R.sup.8 and R.sup.9 ; however, a time span of 3-24 hr is usually sufficient for completion of the reaction.
An alternative method that has been employed in certain instances to prepare compounds of formula (I) in which the L group is (II) is shown in Scheme III: ##STR6##
In Scheme III all R groups are as previously defined. Step 1 in Scheme III is executed as previously described for Step 5 of Scheme I and Step 2 of Scheme III is effected as described for Step 3 of Scheme I. Thus, the epoxide (XI) is reacted with the diamine (XIV), or a protected derivative as explained below, to yield the intermediate (XV) which is then condensed with the pyridinecarbonitrile (VII) to yield the product of the invention (I) where L is moiety (II).
The method shown in Scheme III for the preparation of compounds of formula (I) with L=(II) is most advantageously employed when R.sup.5 -R.sup.9 are all hydrogen or when R.sup.6 .dbd.R.sup.7 .dbd.CH.sub.3 and R.sup.8 .dbd.R.sup.9 .dbd.H. A protected form of the ethylenediamine compound of formula (XIV), e.g. 2-(tert-butoxycarbamoyl)ethylamine, may be used to avoid formation of the bis derivative although simply using an excess of the formula (XIV) amine will usually insure that only the 1:1 adduct is formed. Thus, the amine-protected form of the amine of formula (XIV) having an amine protecting group in the place of the hydrogen on the nitrogen bearing R.sup.5 is reacted in Step 1 of Scheme III and the product, bearing the protecting group on the nitrogen bearing R.sup.5, is deprotected by conventional means to yield the product of formula (XV).
The compounds of formula (I) in which L is a group of the formula (III) may be prepared as shown in Scheme IV: ##STR7##
The compounds of formula (XVI) represent protected alkylamines with a leaving group, Y, at one end of the chain that is reactive toward display element by nucleophiles. Suitable Y groups include halogen and p-toluenesulfonate and p-nitrobenzenesulfonate esters. In Scheme IV, R.sup.15 and R.sup.16 are as defined in Scheme I for and R.sup.14 Suitable amine protecting group R.sup.15 and R.sup.16 include, together with the nitrogen to which they are attached, a phthalimide group, carbamates, and N-benzylated amines. The compounds of formula (XV) are either known or may be prepared from the corresponding halo- or hydroxyalkylamines by conventional methods well known to those skilled in the art.
Step 1 in Scheme IV is effected by reacting the phenol of formula (IV) with a compound of the formula (XVI) in the presence of a suitable base and appropriate solvent to give compounds of formula (XVII). Bases which may be used in Step 1 include sodium hydride, sodium t-butoxide, and similar non-nucleophilic basic reagents. The preferred base is sodium hydride.
A wide variety of solvents may be used in Step 1 of Scheme IV with the only restriction being that the solvent be inert toward starting materials (IV) and (XVI) as well as to the basic reagent and the product (XVII). Suitable solvents include DMF, dimethylsulfoxide, aromatic hydrocarbons such as benzene or toluene, and ethers such as tetrahydrofuran. Step 1 may be conducted over a wide temperature range, with the preferred temperature being 25.degree. C. to 100.degree. C. The preferred conditions for effecting Step 1 of Scheme IV is to use sodium hydride (2.0 equivalents) as the base, dimethylformamide as the solvent, and a temperature of 60.degree. C. to 80.degree. C. Under these conditions, Step 1 is completed in 2-6 hr.
The nature of the reaction conditions for Step 2 of Scheme IV are dependent upon the amine protecting group R.sup.16 or R.sup.15 and R.sup.16 that has been employed. If the amine protective group is a carbamate moiety such as the tert-butoxycarbamoyl group, it may be removed under acid hydrolysis conditions. Reaction conditions and suitable acids are the same as those described earlier for Step 4 of Scheme I. If the protecting group is a phthalimide, it is conveniently removed by treatment of a compound of the formula (XVII) with hydrazine in a suitable solvent. Solvents that may be used for this reaction include alcohols, e.g. ethanol or isopropanol, ethers such as tetrahydrofuran, acetonitrile, or aromatic hydrocarbon solvents such as benzene or toluene. The reaction may be executed over a wide temperature range, with the preferred temperature range being about 25.degree. C. to 100.degree. C. If the protecting group in formula (XVII) is an N-benzylated amine, then the removal of the protection group (Step 2) in Scheme IV is conveniently accomplished by catalytic reduction. Suitable catalysts for this reaction include platinum or palladium supported on activated charcoal. The reaction is carried out under a pressure of 1-3 atmospheres of hydrogen in the temperature range of about 25.degree. C. to 70.degree. C.
Step 3 in Scheme IV is carried out by reacting the amine (XVIII) with an epoxide of the formula (XI) as described for Step 5 of Scheme I.
Specific examples of the compounds of the present invention are those of the formula (I) set forth in the following Table I where the "--O--L--Position" refers to position of substitution on the phenyl ring.
Also part of the present invention are intermediates used in the various processes of the invention including those of formulae (VI), VII), (X), (XVII) and (XVIII).
The efficacy of compounds of the present invention as both inotropic and beta-andrenergic blocking agents can be evaluated and measured using pharmacological methods known in the art or as described in detail below based on similarly established methodologies.
It has been shown by R. F. Kauffman et.al. in J. of Pharmacol. Exp. Ther. 242:864-872 (1987) that inotropic agents such as milrinone and enoximone produce marked relaxation of rate aorta. Such vasorelaxation appears to be related to inhibitaion of the phosphodiesterase (PDE) isozyme related to the cardiac sarcoplasmic reticulum. Thus, relaxation of rat aorta can be used as a screen to eliminate compounds which are not PDE inhibitors prior to testing for actual inotropic activity.
Rings of rat aorta (endothelium removed) were prepared for the measurement of isometric force in isolated tissue organ chambers essentially as previously described by T. J. Rimele et al in the Journal of Pharmacol. Exo. Ther. 245:102-111 (1988). The experimental portion of the protocol began with the addition of methylene blue (1.times.10.sup.-5 M) and propranolol (1.times.10.sup.-6 M) to each organ chamber to inhibit basal cGMP accumulation due to soluble guanylate cyclase and beta-adrenoceptors. Phenylephrine (1.times.10.sup.-7 M) was then added and the rings were allowed to obtain a stable contractile response after which time, the test compound was added in a cumulative fashion. The relaxation induced by each concentration of the test compound was expressed as a percentage of the maximal relaxation produced by nitroprusside (1.times.10.sup.-4 M). The results were graphically represented as a plot of the percentage relaxation vs. the negative log of the molar concentration of the test compound. The IC.sub.50 (concentration of test compound which produced a relaxation equivalent to 50% of the maximal relaxation induced by nitroprusside) was determined for each tissue. The IC.sub.50 for the compound of Example 2 was 23 micromolar with the maximal response being 97% at the highest dose tested (100 micromolar).
Inotropic effects were evaluated in barbiturate anesthetized dogs by differentiating left intraventricular pressure. This procedure was carried out essentially as described by M. K. Grizzel et al in the FASEB Journal, Vol. 3, page 1039, abstract 4728 (1989). Purpose breed mongrel dogs (14-20 kg) of either sex were anesthetized with a mixture of sodium pentobarbital (15 mg/kg) and sodium barbital (300 mg/kg) i.v., intubated with a cuffed endotracheal tube and ventilated with a respirator (Harvard Apparatus, model 613, South Natick, Mass.) with room air (22 rpm,10-12 ml/kg/stroke). A 5F pressure transducer (Millar Instruments, Mikkro-tip. Houston, Tex.) was inserted through the right carotid artery into the left ventricle to monitor intraventricular pressure. The left ventricular pressure signal was differentiated (using a 100 Hz low pass differential amplifier, Gould Inc., Cleveland, Ohio) to obtain its maximal rate of rise (+dP/dt), and used to trigger a biotach amplifier to record heart rate. Cardiac output was determined via thermodilution with a Spectramed computer (Starcom , Oxnard, Calif.) and a 5F Swan Ganz catheter which was inserted into the right jugular vein and positioned in the pulmonary artery. The femoral artery was cannulated for monitoring arterial blood pressure with a pressure transducer (Micron model MP15D, Simi Valley, Calif.) A lead II electrocardiogram was measured using subcutaneous electrodes. Following surgery and instrumentation, each dog was placed in the left lateral decubitus position for the remainder of the experiment, and allowed to stabilize for 45-60 min before starting the experimental protocol. Rectal temperature was monitored and maintained at 37.degree.-38.degree. C. with a heating pad (Baxter Health Care model K20, McGaw Park, Ill.) All variables were recorded using a Gould 3800S physiograph.
Isoproterenol (0.1-0.5 .mu.g/kg) was injected i.v. into the cephalic vein at 10 min intervals except for when the test drug infusion was begun. Four of these initial injections were made to establish the baseline response. Ten min after the forth isoproterenol injection an infusion of test compound was started at 0.01 micromol/kg/10 min after 10 min of test drug infusion an isoproterenol injection was made and the dose rate of test compound was increased. This process was continued up to a dose of test compound ranging from 300 to 10,000 micromol/kg total cumulative dose. Inhibition of the isoproterenol responses on contractility (dP/dt), heart rate and diastolic blood pressure were determined at each dose of test compound. The inotropic effect of each compound was determined by comparing the level of dP/dt at the end of each 10 min period to that of the level of dP/dt just prior to the infusion of test compound. The ED.sub.50 s were determined by a 2 point interpolation of the responses obtained that were just below and above 50% inhibition of the isoproterenol response or a 50% increase in dP/dt. Data are expressed in nanomol/kg.
Inotropic effects of the compounds were determined by changes in the baseline dP/dt whereas the beta-blocking effects of the compounds were determined by quanitating inhibition of the dP/dt response to isoproterenol. The compound of the invention of Example 2 showed an inotropic ED.sub.50 of 40 nanomoles/kg, compared to a baseline established prior to drug infusion and an ED.sub.50 for inhibition of the isoproterenol response of 55 nanomoles/kg. Further, as opposed to many prior inotropic agents which show partial beta agonism whereby the inotropic effects can be prevented by infusion of atenolol, the compound of formula (I) produced in Example 2 showed inotropic effects at higher doses which were not blocked by atenolol. In addition, many other inotropic agents whose inotropic effects at higher doses are not prevented by atenolol are phosphodiesterase inhibitors which do not have beta blocking properties.
This test is carried out as generally described by T.P. Kenakin et al in Journal of Cardiovascular Pharmacology 101, 658-666 (1987) and in the Journal of Pharmacology and Experimental Therapeutics, Vol. 213, 406-413 (1980).
Male Hartley guinea-pigs (300-400 grams) were sacrificed by cervical dislocation or carbon dioxide asphyxiation. The hearts were immediately removed and placed in oxygenated Krebs-Henseleit buffer (composition (millimolar): Na.sup.+ 143, K.sup.+ 5.9, Ca.sup.++ 1.25, Mg.sup.++ 1.2, Cl.sup.- 128, HCO.sub.3.sup.- 25, SO.sub.4.sup.-- 1.2, H.sub.2 PO.sub.4.sup.- 1.0, and D-glucose 10). Left atria were dissected away from the remainder of the heart and mounted on holders against platinum punctate electrodes. The mounted atria were placed in tissue baths maintained at 31.degree. C. and oxygenated with 95% O.sub.2 -5% CO.sub.2 under 1.0 gram resting tension. The atria were stimulated through the punctate electrode and an external platimum electrode at the threshold voltage plus thirty percent, one Hertz frequency and five to ten milliseconds duration. Contractions were detected with a force displacement transducer and recorded on a physiograph.
The atria were allowed to equilibrate for at least one-half hr before the experimental compounds were added to the tissue baths. Propranolol (1.0 micromolar) and phentolamine (1.0 micromolar) were added to the buffer solution in the tissue baths to eliminate any effects of endogenous catecholamine release. Propranolol and phentolamine were added at least thirty min prior to the addition of the test compounds. During the equilibration period, the buffer solution was removed and replaced frequently. Phentolamine and propranolol were immediately re-introduced to the tissue baths after refilling with buffer.
Direct effects of the test compounds on the force of atrial contraction were observed and recorded after addition of the compounds to the tissue bath. Test compounds were added in concentrations from 1.0 to 100 micromolar in ten-fold increments (1.0, 10, 100 micromolar) with an additional concentration of 300 micromolar. Atria were exposed to each concentration of the test compounds until a constant response was observed. After a constant response was observed with the highest concentration (or five min in the absence of a response), forskolin was added in the presence of the test compound. Forskolin was added in ten-fold increments from 0.1 to 100 micromolar. Responses to the test compounds and forskolin were expressed as a percentage of the maximal response to forskolin. EC.sub.50 values for the test compounds were calculated as the concentration of the compound necessary to produce an inotropic response half that of the maximal response produced by the test compound. For the compound produced in Example 2 the maximal response was 36% at 100 micromolar with an ED.sub.50 of 13 micromolar concentration.
Because of its receptor density rat brain cortices were used as the source of membrane vesicles to be used in the receptor binding assays. Freshly excised cortexes were homogenized in 20 volumes (w/v) 50 mM TRIS HCl Buffer (pH 7.5), with a glass/Teflon homogenizer following the procedure previously described by T.J. Rimele in J. Pharmacol. Exp. Ther. 239: 1-8, 1986. Beta-1 adrenoceptor by M.H. Randall, et al, J. Med. Chem. 20: 1090-1094, 1977, and J. Homberger, et al in Mol. Pharmacol. 20: 453-469, 1981. The incubation mixture consisted of ; 26 .mu.l of 50 mM TRIS/HCl, 10 mM MgCl.sub.2 pH 7.6 buffer, 25 .mu.l of test drug or 10-6 pindolol to define nonspecific binding, 100 .mu.l of [I-125]-Pindolol (2200 Ci/mM) at a final concentration of 10-9, and 100 .mu.l of brain cortical membranes. The mixture was incubated at RT (22.degree. C.) for two hr in the dark. The reaction was stopped by filtration of the mixture through buffer soaked glass fiber membranes (GF/B) using an cell harvesting device (Skatron Inc.). The radioactivity in each filter containing the trapped membrane particles was counted with a gamma counter. The value for non-specific binding in each assay was subtracted from total binding to give a value for specific binding. All specific binding values obtained in the presence of test compounds were expressed as the percentage of specific binding displaced by the individual agents. The resultant values were plotted on a log plot of concentration of test compound vs. percentage of displacement and an IC.sub.50 value (drug concentration which produces 50% displacement) determined. Values obtained by this analysis are then reported as the negative log of the IC.sub.50 (pIC.sub.50). The compound of Example 2 showed a pIC.sub.50 of 7.7.
Compounds of the invention of formula (I) may be used in the treatment of CHF in a manner similar to the use of beta-adrenergic blocking agents and (.+-.)-inotropic agents. After suffering a heart attack, one therapy which may be used is administration of a beta-blocker, such as atenolol to lessen oxygen consumption for the damaged heart muscle. However, there is often a negative inotropic action associated with beta-blockers whereby one may consider use of a positive inotropic agent. The usage of compounds of the invention may thus be correlated to the desire to manifest both beta-blocking and positive inotropic actions in a patient.
The compounds of the invention of formula (I) can be administered orally, topically or parenterally, e.g. rectal or i.v., of which the preferred route is oral. The compounds may be admixed with conventional tableting aids, diluents, excepients as known in the art to form tablets, capsules, powders, elixirs, liquids or suspensions as known in the pharmaceutical art. For administration to humans, the compounds of the invention may be administered in an amount of about 0.1 to 5 mg/kg about 1-4 times per day. The particular dosage will depend on the activity of the specific compound chosen and the severity of the physiological condition being treated. The projected dosage can be determined by correlation of test results in pharmacological tests for known positive inotropic agents such as milronone to those for compounds of formula (I).
In the following examples and throughout the specification, the following abbreviations may be used: g (grams),; mg (milligrams); 1 (liters); ml (milliliters); M (molar); mM (millimolar); i.v. (intraveneous); Hz (Hertz); dP/dt (change in pressure per time period); mol (moles); DMF (N,N-dimethylformamide); DMSO (dimethylsulfoxide); TFA (trifluoroacetic acid): RT (room temperature); EtOAc (ethyl acetate); min (minutes); hr (hours); m.p. (melting point); NMR (nuclear magnetic resonance); s (singlet); d (doublet); t (triplet); q(quartet); m (multiplet); and TLC (thin layer chromatography).
Unless otherwise indicated, all temperatures are expressed in .degree. C. (degrees Centigrade), pressures in mmHg (millimeters of mercury), NMR data in delta units and all references to ether are to diethyl ether.
US Referenced Citations (3)
Foreign Referenced Citations (4)
| Number |
Date |
Country |
| 39892 |
Nov 1981 |
EPX |
| 178189 |
Apr 1986 |
EPX |
| 236624 |
Sep 1987 |
EPX |
| 259835 |
Mar 1988 |
EPX |
Non-Patent Literature Citations (2)
| Entry |
| Slater, R. A. et al., J. Med, Chem. 31, p. 345-351 (1988). |
| Curran, W. V. et al., J. Med. Chem., vol. 17, No. 3, p. 273-281 (1974). |
Divisions (1)
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Date |
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| Parent |
565297 |
Aug 1990 |
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Continuation in Parts (1)
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Date |
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| Parent |
411065 |
Sep 1989 |
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Patent Grant
5153209
