GB2346142A - Pyrimidinones useful for treating benign prostatic hyperplasia - Google Patents

Abstract

Compounds of the formula: <EMI ID=1.1 HE=50 WI=112 LX=532 LY=660 TI=CF> <PC>wherein each X is independently hydrogen, halogen, cyano, C<SB>1</SB>-C<SB>4</SB> alkyl, C<SB>1</SB>-C<SB>4</SB> alkoxy, (CH<SB>2</SB>)<SB>0-3</SB>OCF<SB>3</SB>, or (CH<SB>2</SB>)<SB>0-3</SB>CF<SB>3</SB> ; and<BR> q is an integer from, 0 to 5;<BR> or a pharmaceutically acceptable salt thereof, is useful for treating benign prostatic hyperplasia.

Description

TITLE OF THE INVENTION DERIVATIVES OF 4- (3, 4-DIFLUOROPHENYL)-5- METHOXYCARBONYL-3, 4-DIHYDROPYRIMIDIN-2-ONE AS ALPHA 1A ADRENOCEPTOR ANTAGONISTS FIELD OF THE INVENTION This invention relates to 3- (4-substituted phenyl-4-hydroxy-piperidin I-yl) propylamine derivatives of4- (3, 4-difluorophenyl)-5-methoxycarbonyl-3, 4 dihydropyrimidin-2-one and pharmaceutically acceptable salts thereof, and their synthesis. More particularly, the compounds of the present invention are alpha I a adrenoceptor antagonists useful for treating benign prostatic hyperplasia (BPH).

References are made throughout this application to various publications, the disclosures of which are hereby incorporated by reference in their entireties, in order to more fully describe the state of the art to which this invention pertains.

BACKGROUND OF THE INVENTION Human adrenergic receptors are integral membrane proteins which have been classified into two broad classes, the alpha and the beta adrenergic receptors. Both types mediate the action of the peripheral sympathetic nervous system upon binding of catecholamines, norepinephrine and epinephrine.

Norepinephrine is produced by adrenergic nerve endings, while epinephrine is produced by the adrenal medulla. The binding affinity of adrenergic receptors for these compounds forms one basis of the classification: alpha receptors bind norepinephrine more strongly than epinephrine and much more strongly than the synthetic compound isoproterenol. The binding affinity of these hormones is reversed for the beta receptors. In many tissues, the functional responses, such as smooth muscle contraction, induced by alpha receptor activation are opposed to responses induced by beta receptor binding.

Subsequently, the functional distinction between alpha and beta receptors was further highlighted and refined by the pharmacological characterization of these receptors from various animal and tissue sources. As a result, alpha and beta adrenergic receptors were further subdivided into alpha I, alpha 2, ssl, and 32 subtypes. Functional differences between alpha I and alpha 2 receptors have been recognized, and compounds which exhibit selective binding between these two subtypes have been developed.

For a general background on the alpha adrenergic receptors, the reader's attention is directed to Robert R. Ruffolo, Jr., Molecular Biology, Biochemistry and Pharmacology. (Progress in Basic and Clinical Pharmacolo series, Karger, 1991), wherein the basis of alpha 1/alpha 2 subclassification, the molecular biology, signal transduction (G-protein interaction and location of the significant site for this and ligand binding activity away from the 3'-terminus of alpha adrenergic receptors), agonist structure-activity relationships, receptor functions, and therapeutic applications for compounds exhibiting alphaadrenergic receptor affinity was explored.

The cloning, sequencing and expression of alpha receptor subtypes from animal tissues has led to the subclassification of the alpha 1 receptors into alpha ld (formerly known as alpha la or la/ld), alpha lb and alpha la (formerly known as alpha I c) subtypes. Each alpha 1 receptor subtype exhibits its own pharmacologic and tissue specificities. The designation"alpha 1 a"is the appellation recently approved by the IUPHAR Nomenclature Committee for the previously designated "alpha lc"cloned subtype as outlined in the 1995 Receptor and Ion Channel Nomenclature Supplement (Watson and Girdlestone, 1995). The designation alpha la is used throughout this application to refer to this subtype. At the same time, the receptor formerly designated alpha I a was renamed alpha 1 d. The new nomenclature is used throughout this application. Stable cell lines expressing these alpha 1 receptor subtypes are referred to herein; however, these cell lines were deposited with the American Type Culture Collection (ATCC) under the old nomenclature. For a review of the classification of alpha 1 adrenoceptor subtypes, see, Michel et al., Naunyn Schmiedeberg's Arch. Pharmacol. (1995), 352: 1-10.

The differences in the alpha adrenergic receptor subtypes have relevance in pathophysiologic conditions. Benign prostatic hyperplasia, also known as benign prostatic hypertrophy or BPH, is an illness typically affecting men over fifty years of age, increasing in severity with increasing age. The symptoms of the condition include, but are not limited to, increased difficulty in urination and sexual dysfunction. These symptoms are induced by enlargement, or hyperplasia, of the prostate gland. As the prostate increases in size, it impinges on free-flow of fluids through the male urethra. Concomitantly, the increased noradrenergic innervation of the enlarged prostate leads to an increased adrenergic tone of the bladder neck and urethra, further restricting the flow of urine through the urethra.

In benign prostatic hyperplasia, the male hormone 5alpha- dihydrotestosterone has been identified as the principal culprit. The continual production of 5a-dihydrotestosterone by the male testes induces incremental growth of the prostate gland throughout the life of the male. Beyond the age of about fifty years, in many men, this enlarged gland begins to obstruct the urethra with the pathologic symptoms noted above.

The elucidation of the mechanism summarized above has resulted in the recent development of effective agents to control, and in many cases reverse, the pernicious advance of BPH. In the foreront of these agents is Merck & Co., Inc.'s product PROSCARX (finasteride). The effect of this compound is to inhibit the enzyme testosterone 5-a reductase, which converts testosterone into 5adihydrotesterone, resulting in a reduced rate of prostatic enlargement, and often reduction in prostatic mass.

The development of such agents as PROSCAR bodes well for the long-term control of BPH. However, as may be appreciated from the lengthy development of the syndrome, its reversal also is not immediate. In the interim, those males suffering with BPH continue to suffer, and may in fact lose hope that the agents are working sufficiently rapidly.

In response to this problem, one solution is to identify pharmaceutically active compounds which complement slower-acting therapeutics by providing acute relief. Agents which induce relaxation of the lower urinary tract tissue, by binding to alpha 1 adrenergic receptors, thus reducing the increased adrenergic tone due to the disease, would be good candidates for this activity. Thus, one such agent is alfuzosin, which is reporte in EP 0 204597 to induce urination in cases of prostatic hyperplasia. Likewise, in WO 92/00073, the selective ability of the R (+) enantiomer of terazosin to bind to adrenergic receptors of the alpha 1 subtype was reported. In addition, in WO 92/16213, combinations of 5a-reductase inhibitory compounds and alphal-adrenergic receptor blockers (terazosin, doxazosin, prazosin, bunazosin, indoramin, alfuzosin) were disclosed. However, no information as to the alpha I d, alpha 1 b, or alpha 1 a subtype specificity of these compounds was provided as this data and its relevancy to the treatment of BPH was not known. Current therapy for BPH uses existing non-selective alpha 1 antagonists such as prazosin (Minipress, Pfizer), Terazosin (Hytrin, Abbott) or doxazosin mesylate (Cardura, Pfizer). These non-selective antagonists suffer from side effects related to antagonism of the alpha ld and alpha lb receptors in the peripheral vasculature, e. g., hypotension and syncope.

The relatively recent cloning of the human alpha la adrenergic receptor (ATCC CRL 11140) and the use of a screening assay utilizing the cloned human alpha 1 a receptor has enabled identification of compounds which specifically interact with the human alpha 1 a adrenergic receptor. For further description, see WO 94/08040 and WO 94/10989. As disclosed in the instant patent disclosure, a cloned human alpha 1 a adrenergic receptor and a method for identifying compounds which bind the human alpha la receptor has made possible the identification of selective human alpha 1 a adrenergic receptor antagonists useful for treating BPH.

Several classes of compounds have been disclosed to be selective alpha 1 a adrenergic receptor antagonists useful for treating BPH. For example, WO 96/14846, WO 97/17969 and WO 97/42956 each disclose certain dihydropyrimidine derivatives (e. g., certain tetrahydro-2-oxo-pyrimidine derivatives) which are selective antagonists for the human alpha 1 a receptor and useful for treatment of BPH, impotency, cardiac arrhythmia, and other diseases where antagonism of the alpha 1 a receptor may be useful. In particular, Example 37 of WO 96/14846 discloses 3-{N- [3- (4- (4-fluorophenyl)-4-hydroxypiperidinyl-1-yl) propyl]} carboxamido-5- methoxycarbonyl-6-methyl-4- (3, 4-di fluorophenyl)-2-oxo-1, 2,3,4-tetrahydropyrimidine hydrochloride.

Other alpha la selective antagonists are disclosed in: WO 94/22829, which discloses, e. g., certain 4- (un) substituted phenyl1,4-dihydropyridine derivatives which are described as potent, selective alpha 1 a antagonists with weak calcium channel antagonistic activity and which are further described to be anticipated as useful for treating BPH) ; WO 96/40135, which discloses, inter alia, certain phenylpiperidinyl alkyl saccharin derivatives and their use as selective alpha la antagonists; US 5661163 and WO 96/40136, which disclose, inter alia, piperidinyland piperazinyl-alkyl-substituted phenyl acetamides; and EP 748840, which discloses, inter alia, certain arylpiperazinylpropyl substituted pyrimidinediones useful as alpha 1 adrenoceptor antagonists.

Still other alpha 1 a selective antagonist compounds are disclosed in WO 98/57632, WO 98/57638, WO 98/57639, WO 98/57640, WO 98/57641 and WO 98/57642.

The instant patent disclosure discloses novel dihydropyrimidinone compounds which selectively bind to the human alpha I a receptor. These compounds are further tested for binding to other human alpha 1 receptor subtypes, as well as counterscreened against other types of receptors (e. g., alpha 2), thus defining the specificity of the compounds of the present invention for the human alpha 1 a adrenergic receptor.

It is an object of the present invention to identify compounds which bind to the alpha la adrenergic receptor. It is a further object of the invention to identify compounds which act as antagonists of the alpha 1 a adrenergic receptor. It is another object of the invention to identify alpha la adrenergic receptor antagonist compounds which are useful agents for treating BPH in animals, preferably mammals, especially humans. Still another object of the invention is to identify alpha la adrenergic receptor antagonists which are useful for relaxing lower urinary tract tissue in animals, preferably mammals, especially humans.

The compounds of the present invention are alpha 1 a adrenergic receptor antagonists. Thus, the compounds of the present invention are useful for treating BPH in mammals. Additionally, it has been found that the alpha 1 a adrenergic receptor antagonists of the present invention are also useful for relaxing lower urinary tract tissue in mammals.

SUMMARY OF THE INVENTION The present invention provides 3- (4-substituted phenyl-4 hydroxypiperidin-1-yl) propylamine derivatives of 4- (3, 4-difluorophenyl)-5methoxycarbonyl-3,4-dihydropyrimidin-2-Qne and pharmaceutically acceptable salts thereof for the treatment of urinary obstruction caused by benign prostatic hyperplasia (BPH). The compounds antagonize the human alpha 1 a adrenergic receptor at nanomolar and subnanomolar concentrations while exhibiting lower affinity for the alpha ld and alpha lb human adrenergic receptors and many other G-protein coupled receptors. This invention has the advantage over non-selective alpha 1 adrenoceptor antagonists of reduced side effects related to peripheral adrenergic blockade. Such side effects include hypotension, syncope, lethargy, etc.

More particularly, the present invention is a compound of formula (I) :

wherein each X is independently hydrogen, halogen, cyano, C1-C4 alkyl, C1-C4 alkoxy, (CH2) 0-30CF3, or (CH2) 0-3CF3 ; and q is an integer from 0 to 5 ; or a pharmaceutically acceptable salt thereof.

The present invention also includes pharmaceutical compositions, methods of preparing pharmaceutical compositions, and methods of treatment.

These and other embodiments, aspects and features of the present invention are either further described in or will be apparent from the ensuing description, examples and appended claims.

DETAILED DESCRIPTION OF THE INVENTION The present invention includes dihydropyrimidinone compounds of Formula (I) above. These compounds and their pharmaceutically acceptable salts are useful as alpha la antagonists.

In a first embodiment, the present invention is a compound of Formula (I), wherein the compound is a (+)-enantiomer; and all of the variables are as originally defined; or a pharmaceutically acceptable salt thereof.

In a second embodiment, the present invention is a compound of Formula (1), wherein the compound is a (-)-enantiomer; and all of the variables are as originally defined; or a pharmaceutically acceptable salt thereof.

In a third embodiment, the present invention is a compound of Formul wherein each X is independently hydrogen, fluorine, cyano, methyl, ethyl, OCF3, or CF3 ; and q is an integer from 0 to 3; or a pharmaceutically acceptable salt thereof.

In a class of the invention is a compound of Formula (II) :

wherein X is as originally defined; or a pharmaceutically acceptable salt thereof.

In one aspect of the foregoing class, the compound is a compound of Formula (II), wherein each X is independently hydrogen, fluorine, cyano, methyl, ethyl, OCF3, or CF3; or a pharmaceutically acceptable salt thereof.

Exemplifying the invention is a compound of Formula:

or a pharmaceutically acceptable salt thereof.

The present invention also includes a pharmaceutical composition comprising a therapeutically effective amount of any of the compounds described above and a pharmaceutically acceptable carrier. In one embodiment is a pharmaceutical composition made by combining any of the compounds described above and a pharmaceutically acceptable carrier. The present invention further includes a process for making a pharmaceutical composition comprising combining any of the compounds described above and a pharmaceutically acceptable carrier.

The present invention further includes a pharmaceutical composition as described in the preceding paragraph further comprising a therapeutically effective amount of a testosterone 5-alpha reductase inhibitor. In one embodiment, the testosterone 5-alpha reductase inhibitor is a type 1, a type 2, both a type 1 and a type 2 (i. e., a three component combination comprising any of the compounds described above combined with both a type 1 testosterone 5-alpha reductase inhibitor and a type 2 testosterone 5-alpha reductase inhibitor), or a dual type 1 and type 2 testosterone 5alpha reductase inhibitor. In another embodiment, the testosterone 5-alpha reductase inhibitor is a type 2 testosterone 5-alpha reductase inhibitor. The testosterone 5-alpha reductase inhibitor is suitably finasteride.

The present invention also includes a method of treating benign prostatic hyperplasia in a subject in need thereof which comprises administering to the subject a therapeutically effective amount of any of the compounds (or any of the compositions) described above. In one embodiment of the method of treating BPH, the compound (or composition) does not cause a fall in blood pressure at dosages effective to alleviate BPH. In another embodiment of the method of treating BPH, the compound is administered in combination with a testosterone 5-alpha reductase inhibitor. A suitable testosterone 5-alpha reductase inhibitor for use in the method is finasteride.

The present invention also includes a method of inhibiting contraction of prostate tissue or relaxing lower urinary tract tissue in a subject in need thereof which comprises administering to the subject a therapeutically effective amount of any of the compounds (or any of the compositions) described above. In one embodiment of the method of inhibiting contraction of prostate tissue or relaxing lower urinary tract tissue, the compound (or composition) additionally does not cause a fall in blood pressure at dosages effective to inhibit contraction of prostate tissue. In another embodiment, the compound is administered in combination with a testosterone 5-alpha reductase inhibitor; the testosterone 5-alpha reductase inhibitor is suitably finasteride.

The present invention also includes a method of treating a disease which is susceptible to treatment by antagqnism of the alpha la receptor which comprises administering to a subject in need thereof an amount of any of the compounds described above effective to treat the disease. Diseases which are susceptible to treatment by antagonism of the alpha 1 a receptor include, but are not limited to, BPH, high intraocular pressure, high cholesterol, impotency, sympathetically mediated pain, migraine (see Vatz, Headache (1997), 37 : 107-108) and cardiac arrhythmia.

The present invention also includes the use of any of the compounds described above in the preparation of a medicament for: a) treating benign prostatic hyperplasia; b) relaxing lower urinary tract tissue; or c) inhibiting contraction of prostate tissue; in a subject in need thereof.

The present invention further includes the use of any of the alpha 1 a antagonist compounds described above and a 5-alpha reductase inhibitor for the manufacture of a medicament for: a) treating benign prostatic hyperplasia; b) relaxing lower urinary tract tissue; or c) inhibiting contraction of prostate tissue which comprises an effective amount of the alpha 1 a antagonist compound and an effective amount of 5-alpha reductase inhibitor, together or separately.

As used herein, the term"C 1-C4 alkyl"means linear or branched chain alkyl groups having from 1 to 4 carbon atoms and includes n-, iso-, sec-and t-butyl, n-and isopropyl, ethyl and methyl.

The term"C 1-C4 alkoxy"means an-0-alkyl group wherein alkyl is Ci -C4 alkyl."C1-C4 alkoxy"includes methoxy, ethoxy, n-propoxy, isopropoxy, nbutoxy, isobutoxy, tert-butoxy, and sec-butoxy.

The term"halogen" (which may alternatively be referred to as"halo") refers to fluorine, chlorine, bromine and iodine (alternatively, fluoro, chloro, bromo, and iodo).

The term"aryl"refers to phenyl and substituted phenyl.

The term"substituted"includes mono-and poly-substitution by a named substituent to the extent such single and multiple substitution is chemically allowed.

It is understood that the definition of a variable (e. g., X) at a particular location in a molecule is independent of its definition at other locations in that molecule. Thus, for example, when q is 2, the X substituent may be fluoro at one position on the phenyl ring, and may be hydrogen, cyano, halo, etc. at the other position.

It is understood that substituents and substitution patterns on the compounds of the instant invention can be selected by one of ordinary skill in the art to provide compounds that are chemically stable and that can be readily synthesized by the methods set forth below and, when viewed in the light of this disclosure, by techniques known in the art. Where multiple substituent moieties are disclosed or claimed, the substituted compound can be independently substituted by one or more of the disclosed or claimed substituent moieties, singly or plurally.

Representative compounds of the present invention exhibit selectivity for the human alpha 1 a adrenergic receptor. One implication of this selectivity is that these compounds display selectivity for lowering intraurethral pressure without substantially affecting diastolic blood pressure.

Representative compounds of this invention display submicromolar affinity for the human alpha la adrenergic receptor subtype while displaying lower affinity for the human alpha ld and alpha lb adrenergic receptor subtypes, and many other G-protein coupled human receptors. Particular representative compounds of this invention exhibit nanomolar and subnanomolar affinity for the human alpha I a adrenergic receptor subtype while displaying at least about 10 fold lower affinity for the human alpha ld and alpha lb adrenergic receptor subtypes, and many other Gprotein coupled human receptors (e. g., serotonin, dopamine, alpha 2 adrenergic, beta adrenergic or muscarinic receptors). Still other compounds of this invention exhibit nanomolar and subnanomolar affinity for the human alpha 1 a adrenergic receptor subtype while displaying at least about 20 fold lower affinity for the human alpha 1 d and alpha lb adrenergic receptor subtypes, in addition to exhibiting selectivity over other G-protein coupled human receptors (e. g., serotonin, dopamine, alpha 2 adrenergic, beta adrenergic or muscarinic receptors).

These compounds are administered in dosages effective to antagonize the alpha 1 a receptor where such treatment is needed; e. g., treatment of BPH. For use in medicine, the salts of the compounds of this invention refer to non-toxic "pharmaceutically acceptable salts."Other salts may, however, be useful in the preparation of the compounds according to the invention or in the prepartion of their pharmaceutically acceptable salts. Suitable pharmaceutically acceptable salts of the compounds of this invention include acid addition salts which may, for example, be formed by mixing a solution of the compound according to the invention with a solution of a pharmaceutically acceptable acid such as hydrochloric acid, sulfuric acid, fumaric acid, maleic acid, succinic acid, acetic acid, benzoic acid, citric acid, tartaric acid, carbonic acid or phosphoric acid. Furthermore, where the compounds of the invention carry an acidic moiety, suitable pharmaceutically acceptable salts thereof may include alkali metal salts, e. g. sodium or potassium salts; alkaline earth metal salts, e. g. calcium or magnesium salts; and salts formed with suitable organic ligands, e. g. quaternary ammonium salts. Thus, representative pharmaceutically acceptable salts include the following: Acetate, benzenesulfonate, benzoate, bicarbonate, bisulfate, bitartrate, borate, camsylate, carbonate, clavulanate, citrate, dihydrochloride, edetate, edisylate, estolate, esylate, fumarate, gluceptate, gluconate, glutamate, glycollylarsanilate, hexylresorcinate hydrobromide, hydrochloride, hydroxynaphthoate, hydroiodide, isothionate, lactate, lactobionate, laurate, malate, maleate, mandelate, mesylate, methylbromide, methylnitrate, methylsulfate, mucate, napsylate, nitrate, nmethylglucamine ammonium salt, oleate, palmitate, pantothenate, phosphate/diphosphate, polygalacturonate, salicylate, stearate, sulfate, subacetate, succinate, tannate, tartrate, tosylate and valerate.

Compounds of this invention are used to reduce the acute symptoms of BPH. Thus, compounds of this invention may be used alone or in combination with more long-term anti-BPH therapeutics, such as testosterone 5-a reductase inhibitors, including PROSCARX (finasteride). Aside from their utility as anti-BPH agents, these compounds may be used to induce highly tissue-specific, localized alpha 1 a adrenergic receptor blockade whenever this is desired. Effects of this blockade include reduction of intra-ocular pressure, control of cardiac arrhythmias, and possibly a host of alpha 1 a receptor mediated central nervous system events.

The present invention includes within its scope prodrugs of the compounds of this invention. In general, such prodrugs will be functional derivatives of the compounds of this invention which are readily convertible in vivo into the required compound. Thus, in the methods of treatment of the present invention, the term"administering"shall encompass the treatment of the various conditions described with the compound specifically disclosed or with a compound which may not be specifically disclosed, but which converts to the specified compound in vivo after administration to the patient Conventional procedures for the selection and preparation of suitable prodrug derivatives are described, for example, in Design of Prodrugs, ed. H. Bundgaard, Elsevier, 1985.

The present invention further includes metabolites of the compounds of the present invention. Metabolites include active species produced upon introduction of compounds of this invention into the biological milieu.

Where the compounds according to the invention have at least one chiral center, they may accordingly exist as enantiomers. Where the compounds according to the invention possess two or more chiral centers, they may additionally exist as diastereoisomers. It is to be understood that all such isomers and mixtures thereof are encompassed within the scope of the present invention. Furthermore, some of the crystalline forms for compounds of the present invention may exist as polymorphs and as such are intended to be included in the present invention. In addition, some of the compounds of the present invention may form solvates with water (i. e., hydrates) or common organic solvents. Such solvates are also encompassed within the scope of this invention.

The term"selective alpha 1 a adrenergic receptor antagonist,"as used herein, refers to an alpha 1 a antagonist compound which is at least about ten fold selective for the human alpha 1 a adrenergic receptor as compared to the human alpha lb, alpha Id, alpha 2a, alpha 2b and alpha 2c adrenergic receptors.

The term"lower urinary tract tissue,"as used herein, refers to and includes, but is not limited to, prostatic smooth muscle, the prostatic capsule, the urethra and the bladder neck.

The term"subject,"as used herein refers to an animal, preferably a mammal, most preferably a human, who has been the object of treatment, observation or experiment.

The term"therapeutically effective amount"as used herein means that amount of active compound or pharmaceutical agent that elicits the biological or medicinal response in a tissue, system, animal or human that is being sought by a researcher, veterinarian, medical doctor or other clinician, which includes alleviation of the symptoms of the disease being treated.

The present invention includes pharmaceutical compositions comprising one or more compounds of this invention in association with a pharmaceutically acceptable carrier. Preferably these compositions are in unit dosage forms such as tablets, pills, capsules, powders, granules, sterile parenteral solutions or suspensions, metered aerosol or liquid sprays, drops, ampoules, auto-injector devices or suppositories; for oral, parenteral, intranasal, sublingual or rectal administration, or for administration by inhalation or insufflation. Alternatively, the compositions may be presented in a form suitable for once-weekly or once-monthly administration; for example, an insoluble salt of the active compound, such as the decanoate salt, may be adapted to provide a depot preparation for intramuscular injection. For preparing solid compositions such as tablets, the principal active ingredient is mixed with a pharmaceutical carrier, e. g. conventional tableting ingredients such as corn starch, lactose, sucrose, sorbitol, talc, stearic acid, magnesium stearate, dicalcium phosphate or gums, and other pharmaceutical diluent, e. g. water, to form a solid preformulation composition containing a homogeneous mixture of a compound of the present invention, or a pharmaceutically acceptable salt there outer dosage component, the latter being in the form of an envelope over the former.

The two components can be separated by an enteric layer which serves to resist disintegration in the stomach and permits the inner component to pass intact into the duodenum or to be delayed in release. A variety of materials can be used for such enteric layers or coatings, such materials including a number of polymeric acids and mixtures of polymeric acids with such materials as shellac, cetyl alcohol and cellulose acetate.

As used herein, the term"composition"encompasses a product comprising the specified ingredients in the specified amounts, as well as any product which results, directly or indirectly, from combination of the specified ingredients in the specified amounts.

The liquid forms in which the novel compositions of the present invention may be incorporated for administration orally or by injection include aqueous solutions, suitably flavoured syrups, aqueous or oil suspensions, and flavored emulsions with edible oils such as cottonseed oil, sesame oil, coconut oil or peanut oil, as well as elixirs and similar pharmaceutical vehicles. Suitable dispersing or suspending agents for aqueous suspensions include synthetic and natural gums such as tragacanth, acacia, alginate, dextran, sodium carboxymethylcellulose, methylcellulose, polyvinyl-pyrrolidone or gelatin.

Where the processes for the preparation of the compounds according to the invention give rise to mixtures of stereoisomers, these isomers may be separated by conventional techniques such as preparative chromatography. The compounds may be prepared in racemic form, or individual enantiomers may be prepared either by enantiospecific synthesis or by resolution. The compounds may, for example, be resolved into their component enantiomers by standard techniques, such as the formation of diastereomeric pairs by salt formation with an optically active acid, such as (-)-di-p-toluoyl-d-tartaric acid and/or (+)-di-p-toluoyl-l-tartaric acid followed by fractional crystallization and regeneration of the free base. The compounds may also be resolved by formation of diastereomeric esters or amides, followed by chromatographic separation and removal of the chiral auxiliary. Alternatively, the compounds may be resolved using a chiral HPLC column.

During any of the processes for preparation of the compounds of the present invention, it may be necessary and/or desirable to protect sensitive or reactive groups on any of the molecules concerned. This may be achieved by means of conventional protecting groups, such as those described in Protective Groups in Organic Chemistry, ed. J. F. W. McOmie, Plenum Press, 1973; and T. W. Greene & P. G. M. Wuts, Protective Groups in Organic Synthesis, John Wiley & Sons, 1991.

The protecting groups may be removed at a convenient subsequent stage using methods known in the art.

The specificity of binding of compounds showing affinity for the alpha 1 a receptor is shown by comparing affinity to membranes obtained from transfected cell lines that express the alpha la receptor and membranes from cell lines or tissues known to express other types of alpha (e. g., alpha Id, alpha lb) or beta adrenergic receptors. Expression of the cloned human alpha 1 d, alpha 1 b, and alpha 1 a receptors and comparison of their binding properties with known selective antagonists provides a rational way for selection of compounds and discovery of new compounds with predictable pharmacological activities. Antagonism by these compounds of the human alpha 1 a adrenergic receptor subtype may be functionally demonstrated in anesthetized animals. These compounds may be used to increase urine flow without exhibiting hypotensive effects.

The ability of compounds of the present invention to specifically bind to the alpha la receptor makes them useful for the treatment of BPH. The specificity of binding of compounds showing affinity for the alpha la receptor is compared against the binding affinities to other types of alpha or beta adrenergic receptors. The human alpha adrenergic receptor of the 1 a subtype was recently identified, cloned and expressed as described in PCT International Application Publication Nos.

W094/08040, published 14 April 1994 and WO 94/21660, published 29 September 1994. The cloned human alpha 1 a receptor, when expressed in mammalian cell lines, is used to discover ligands that bind to the receptor and alter its function. Expression of the cloned human alpha Id, alpha I b, and alpha 1 a receptors and comparison of their binding properties with known selective antagonists provides a rational way for selection of compounds and discovery of new compounds with predictable pharmacological activities.

Compounds of this invention exhibiting human alpha 1 a adrenergic receptor antagonism may further be defined by counterscreening. This is accomplished according to methods known in the art using other receptors responsible for mediating diverse biological functions. (See e. ., PCT International Application Publication No. W094/10989, published 26 May 1994; US 5403847, issued April 4, 1995]. Compounds which are both selective amongst the various human alphal adrenergic receptor subtypes and which have low affinity for other receptors, such as the alpha 2 adrenergic receptors, the B-adrenergic receptors, the muscarinic receptors, the serotonin receptors, the histamine receptors, and others are particularly preferred.

The absence of these non-specific activities may be confirmed by using cloned and expressed receptors in an analogous fashion to the method disclosed herein for identifying compounds which have high affinity for the various human alphal adrenergic receptors. Furthermore, functional biological tests are used to confirm the effects of identified compounds as alpha la adrenergic receptor antagonists.

The present invention also has the objective of providing suitable topical, oral, systemic and parenteral pharmaceutical formulations for use in the novel methods of treatment of the present invention. The compositions containing compounds of this invention as the active ingredient for use in the specific antagonism of human alpha 1 a adrenergic receptors can be administered in a wide variety of therapeutic dosage forms in conventional vehicles for systemic administration. For example, the compounds can be administered in such oral dosage forms as tablets, capsules (each including timed release and sustained release formulations), pills, powders, granules, elixirs, tinctures, solutions, suspensions, syrups and emulsions, or by injection. Likewise, they may also be administered in intravenous (both bolus and infusion), intraperitoneal, subcutaneous, topical with or without occlusion, or intramuscular form, all using forms well known to those of ordinary skill in the pharmaceutical arts. An effective but non-toxic amount of the compound desired can be employed as an alpha la antagonistic agent.

Advantageously, compounds of the present invention may be administered in a single daily dose, or the total daily dosage may be administered in divided doses of two, three or four times daily. Furthermore, compounds for the present invention can be administered in intranasal form via topical use of suitable intranasal vehicles, or via transdermal routes, using those forms of transdermal skin patches well known to those of ordinary skill in that art. To be administered in the form of a transdermal delivery system, the dosage administration will, of course, be continuous rather than intermittent throughout the dosage regimen.

The dosage regimen utilizing the compounds of the present invention is selected in accordance with a variety of factors including type, species, age, weight, sex and medical condition of the patient; the severity of the condition to be treated; the route of administration; the renal and hepatic function of the patient; and the particular compound thereof employed. A physician or veterinarian of ordinary skill can readily determine and prescribe the effective amount of the drug required to prevent, counter or arrest the progress of the condition. Optimal precision in achieving concentration of drug within the range that yields efficacy without toxicity requires a regimen based on the kinetics of the drug's availability to target sites. This involves a consideration of the distribution, equilibrium, and elimination of a drug.

In the methods of the present invention, the compounds herein described in detail can form the active ingredient, and are typically administered in admixture with suitable pharmaceutical diluents, excipients or carriers (collectively referred to herein as"carrier"materials) suitably selected with respect to the intended form of administration, that is, oral tablets, capsules, elixirs, syrups and the like, and consistent with conventional pharmaceutical practices.

For instance, for oral administration in the form of a tablet or capsule, the active drug component can be combined with an oral, non-toxic pharmaceutically acceptable inert carrier such as ethanol, glycerol, water and the like. Moreover, when desired or necessary, suitable binders, lubricants, disintegrating agents and coloring agents can also be incorporated into the mixture. Suitable binders include, without limitation, starch, gelatin, natural sugars such as glucose or beta-lactose, corn sweeteners, natural and synthetic gums such as acacia, tragacanth or sodium alginate, carboxymethylcellulose, polyethylene glycol, waxes and the like. Lubricants used in these dosage forms include, without limitation, sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride and the like. Disintegrators include, without limitation, starch, methyl cellulose, agar, bentonite, xanthan gum and the like.

The liquid forms in suitably flavored suspending or dispersing agents such as the synthetic and natural gums, for example, tragacanth, acacia, methylcellulose and the like. Other dispersing agents which may be employed include glycerin and the like. For parenteral administration, sterile suspensions and solutions are desired. Isotonic preparations which generally contain suitable preservatives are employed when intravenous administration-is desired.

The compounds of the present invention can also be administered in the form of liposome delivery systems, such as small unilamellar vesicles, large unilamellar vesicles and multilamellar vesicles. Liposomes can be formed from a variety of phospholipids, such as cholesterol, stearylamine or phosphatidylcholines.

Compounds of the present invention may also be delivered by the use of monoclonal antibodies as individual carriers to which the compound molecules are coupled. The compounds of the present invention may also be coupled with soluble polymers as targetable drug carriers. Such polymers can include polyvinyl-pyrrolidone, pyran copolymer, polyhydroxypropylmethacrylamidephenol, polyhydroxy-ethylaspartamidephenol, or polyethyleneoxidepolylysine substituted with palmitoyl residues. Furthermore, the compounds of the present invention may be coupled to a class of biodegradable polymers useful in achieving controlled release of a drug, for example, polylactic acid, polyepsilon caprolactone, polyhydroxy butyric acid, polyorthoesters, polyacetals, polydihydro-pyrans, polycyanoacrylates and cross-linked or amphipathic block copolymers of hydrogels.

Compounds of this invention may be administered in any of the foregoing compositions and according to dosage regimens established in the art whenever specific blockade of the human alpha 1 a adrenergic receptor is required.

The daily dosage of the products may be varied over a wide range; e. g., from about 0.01 to about 1000 mg per adult human per day. For oral administration, the compositions are preferably provided in the form of tablets containing 0.01,0.05, 0.1,0.5,1.0,2.5,5.0,10.0,15.0,25.0,50.0 and 100 milligrams of the active ingredient for the symptomatic adjustment of the dosage to the patient to be treated.

A medicament typically contains from about 0.01 mg to about 500 mg of the active ingredient, preferably, from about 1 mg to about 100 mg of active ingredient. An effective amount of the drug is ordinarily supplied at a dosage level of from about 0.0002 mg/kg to about 20 mg/kg of body weight per day. Preferably, the range is from about 0.001 to about 10 mg/kg of body weight per day, and especially from about 0.001 mglkg to about 7 mg/kg of body weight per day. The compounds may be administered on a regimen of 1 to 4 times per day.

Compounds of this patent disclosure may be used alone at appropriate dosages defined by routine testing in order to obtain optimal antagonism of the human alpha 1 a adrenergic receptor while minimizing any potential toxicity. In addition, coadministration or sequential administration of other agents which alleviate the effects of BPH is desirable. Thus, in one embodiment, this invention is administration of compounds of this invention and a human testosterone 5-a reductase inhibitor.

Included with this embodiment are inhibitors of 5-alpha reductase isoenzyme 2.

Many such compounds are now well known in the art and include such compounds as PROSCAR@, (also known as finasteride, a 4-Aza-steroid ; see US 4377584 and 4760071, for example). In addition to PROSCAR (D, which is principally active in prostatic tissue due to its selectivity for human 5-a reductase isozyme 2, combinations of compounds which are specifically active in inhibiting testosterone 5-alpha reductase isozyme 1 and compounds which act as dual inhibitors of both isozymes 1 and 2, are useful in combination with compounds of this invention. Compounds that are active as 5a-reductase inhibitors have been described in W093/23420, EP 0572166; WO 93/23050 ; W093/23038,; W093/23048; W093/23041 ; W093/23040 ; W093/23039 ; W093/23376 ; W093/23419, EP 0572165; W093/23051.

The dosages of the alpha 1 a adrenergic receptor and testosterone 5- alpha reductase inhibitors are adjusted when combined to achieve desired effects.

As those skilled in the art will appreciate, dosages of the 5-alpha reductase inhibitor and the alpha 1 a adrenergic receptor antagonist may be independently optimized and combined to achieve a synergistic result wherein the pathology is reduced more than it would be if either agent were used alone. In accordance with the method of the present invention, the individual components of the combination can be administered separately at different times during the course of therapy or concurrently in divided or single combination forms. The instant invention is therefore to be understood as embracing all such regimes of simultaneous or alternating treatment and the term"administering"is to be interpreted accordingly.

Thus, in one embodiment of the present invention, a method of treating BPH is provided which comprises administering to a subject in need of treatment any of the compounds of the present invention in combination with finasteride effective to treat BPH. The dosage of finasteride administered to the subject is from about 0.01 mg per subject per day to about 50 mg per subject per day in combination with an alpha 1 a antagonist. In one aspect, the dosage of finasteride in the combination is from about 0.2 mg per subject per day to about 10 mg per subject per day, and, in another aspect, from about 1 to about 7 mg per subject to day (e. g., about 5 mg per subject per day).

For the treatment of benign prostatic hyperplasia, compounds of this invention exhibiting alpha la adrenergic receptor blockade can be combined with a therapeutically effective amount of a 5a-reductase 2 inhibitor, such as finasteride, in addition to a 5a-reductase 1 inhibitor, such as 4,7 -dimethyl-4-aza-Sa-cholestan-3one, in a single oral, systemic, or parenteral pharmaceutical dosage formulation.

Alternatively, a combined therapy can be employed wherein the alpha 1 a adrenergic receptor antagonist and the 5a-reductase 1 or 2 inhibitor are administered in separate oral, systemic, or parenteral dosage formulations. See, e. g., US 4377584 and US 4760071 which describe dosages and formulations for 5a-reductase inhibitors.

Abbreviations used in the instant specification, particularly the Schemes and Examples, are as follows: Boc or BOC = t-butyloxycarbonyl DMF = N, N-dimethylformamide DMSO = dimethylsulfoxide EDTA = ethylenediamine tetraacetic acid EtOAc = ethyl acetate Et20 = diethyl ether LDA = lithium diisopropyl amide NMR = nuclear magnetic resonance TEA = triethylamine THF = tetrahydrofuran The compounds of the present invention can be readily prepared according to the following reaction schemes and example, or modifications thereof, using readily available starting materials, reagents and conventional synthesis procedures. In these reactions, it is also possible to make use of variants which are themselves known to those of ordinary skill in this art, but are not mentioned in greater detail. Furthermore, other methods for preparing compounds of the invention will be readily apparent to the person of ordinary skill in the art in light of the following reaction schemes and examples. Unless otherwise indicated, all variables are as defined above.

Many of the compounds of the present invention can be prepared via Schemes 1 and 2 shown below. Scheme 1 shows a method for preparing the "activated"dihydropyrimidinone precursor. 3,4-Difluorobenzaldehyde, urea, and methyl 3,3-dimethoxypropionate are contacte in the presence of copper oxide and boron trifluoride diethyl etherate to form a mixture 1 of the enantiomers of 4- (3, 4difluorophenyl)-2-oxo-1, 2,3,4-tetrahydro-pyrimidine-5-carboxylic acid methyl ester. I can be used directly in subsequent preparative steps or, as illustrated in Scheme 1, is resolved (e. g., by chiral chromatography) to obtain the (+)-enantiomer 2, which is deprotonated with a strong base (for example, LDA) and then treated with a THF solution of p-nitrophenylchloroformate to produce stable, isolable"activated" dihydropyrimidinone 3.

Scheme 2 provides a method for preparing the amine component incorporated into the compounds of the invention. An aryl Grignard reagent is reacted with a Boc-protected piperidone to provide tertiary alcohol 4, which is deprotected to give 5, which is treated with TEA, then with a nitrogen-protected 3-halopropylamine (e. g., N-tert-butoxycarbonyl-3-bromo-propylamine), and then with HCI to obtain 6.

Amine 6 is then coupled with activated compound 3 to form 7.

SCHEME 1

F Me. MeCycQ2Me BF3 Et20, CuO js H2N NH2 + o Met O H 0 F / Solution , 4 H 1 O H 2 O'N L LDA LIA (p-N02Ph) OCOCI F(N 2 02Me N, 4 0 N ON H (+) 3 SCHEME 2

The following Examples further describe and illustrate the invention and its practice and are not to be construed as limiting the scope or spirit of the invention.

EXAMPLE 1 (+)-4- (3, 4-Difluorophenyl)-3- {3- [4- (4-fluorophenyl)-4-hydroxypiperidin-1-yl]- propcarbamoYl}-2-oxo-1, 2, 3, 4-tetrahvdro-pyrimidine-5-carboxylic acid methyl ester

Step A. 1- (3-Aminopropyl)-4- (4-fluorophenyl)-piperidin-4-ol

A solution of 4- (4-fluorophenyl)-4-piperidinol (2.3g, 10 mmole) (Sigma) and N-tert-butoxycarbonyl-3-bromopropylamine (2.5g, 10.4 mmole) in DMF (20 mL) was treated with TEA (2.5 g, 25 mmole) and stirred at room temperature, for 24 hours. The reaction was poured into saturated sodium bicarbonate solution (150 mL) and the mixture extracted with ethyl acetate (3 X 150 mL). The combined extracts were dried over anhydrous magnesium sulfate, filtered and concentrated at reduced pressure to give an oil which was chromatographed on silica gel eluting with methanol/methylene chloride to give 2.2 g of the Boc protected intermediate. This material was dissolved in ethyl acetate (100 mL), cooled to 0 C, and hydrogen chloride gas was bubbled through the solution until saturated (10 min). The solution was stirred at 0 C (20 min) and then concentrated in vacuo to give the title product as a white solid.

1H NMR (CD30D) : 8 7.6-7.45 (m, 2H), 7.15-7.05 (m, 2H), 3.65-3.40 (m, 4H), 3.3-3.25 (m, 2H), 3.15-3.05 (t, J = 8.2 Hz, 2H), 2.55-2.4 (m, 2H), 2.30-2.15 (m, 2H), 2.05-1.8 (m, 2H) StepB. 4- (3, 4-Difluorophenyl)-2-oxo-1, 2,3,4-tetrahydro-pyrimidine-5-carboxylic acid methyl ester

To a solution of methyl 3,3-dimethoxypropionate (284 mL, 2.0 mmol) in 4.0 mL THF was added urea (180 mg, 3. 0 mmol), 3,4-difluorobenzaldehyde (284 mg, 2.0 mmol), acetic acid (12.0 mL, 0.20 mmol), copper oxide (27.0 mg, 0.20 mmol), and boron trifluoride diethyl etherate (0.320 mL, 2.6 mmol). The solution was heated to 65 C under argon for 24 h, poured onto saturated sodium bicarbonate, and extracted with ethyl acetate. The combined organic layers were washed with saturated sodium chloride, dried with magnesium sulfate, and concentrated in vacuo.

The crude material was swished with dichloromethane to give the mixture of enantiomers. The enantiomers were resolved on a chiral 250 X 46 mm OD column with a mobile phase of 3: 1 hexane: ethyl acetate, a flow rate of 1 ml/min, and detection at 280 nm.

IH NMR (DMSO, 300 MHz) 8 9.29 (d, 1H, J= 5.37Hz), 7.75 (s, 1H), 7.50-7.35 (m, 1H), 7.30-7.20 (m, 2H), 7.15-7.05 (m, 1H), 5.155 (d, 1H, J= 2.68Hz), 3.565 (s, 3H).

Step C. (+) -4- (3, 4-Difluorophenyl)-2-oxo-1,2,3,4-tetrahydro-3,5-dicarboxylic acid 5methylester-3-p-nitrophenylester

To a-78 C solution of 0. 50g (1.87 mmol) (+)-4- (3, 4-difluorophenyl) 2-oxo-1, 2,3,4-tetrahydro-pyrimidine-5-carboxylic acid methyl ester in 20 ml THF was added 1.1 mL (2.2 mmol, 2M solution in THF/heptane/ethylbenzene) of lithium diisopropylamide. The reaction mixture was stirred at-78 C for 15 min., then tranferred via cannula to a-78 C solution of 0. 47g (2.3 mmol) 4nitrophenylchloroformate in 10 mL THF. After 40 min at-78 C, the reaction mixture was diluted with 200 ml EtOAc, washed with 200 ml each of saturated sodium bicarbonate solution, 5% aqueous K2CO3 solution, water, and brine. The organic layer was dried over Na2SO4, filtered and concentrated. Purification by flash chromatography (3.5x12 cm silica gel, linear gradient 5-10% acetonelCH2Cl2) provided 0. 5g of (+)-4- (3, 4-difluorophenyl)-2-oxo-1, 2,3,4-tetrahydropyrimidine-3,5dicarboxylic acid 5-methylester-3-p-nitrophenylester. lH NMR (CDCI3, 300 MHz) 8 8.29 (br d, 2H, J= 8. 8 Hz); 7.91 (br d, 1H, J= 5.8 Hz); 7.44 (d, 1H, J= 5.8 Hz); 7.36 (br d, 2H, J = 8.9 Hz); 7.31-7.10 (m, 3H) ; 6.28 (s, 1H) ; 3.78 (s, 3H).

StepD. (+) -4- (3, 4-Difluorophenyl)-3- {3- [4- (4-fluorophenyl)-4-hydroxypiperidin-l- yl]-propylcarbamoyl}-2-oxo-1, 2,3,4-tetrahydro-pyrimidine-5-carboxylic acid methyl ester To a solution of 1- (3-aminopropyl)-4- (4-fluorophenyl)-piperidin-4-ol dihydrochloride (0.214 mmol, 66.0 mg) and triethylamine (30.0 uL, 0.642 mmol) in dimethylformamide (2.0 mL) under argon was added (+)-4- (3, 4-Difluorophenyl)-2- oxo-1, 2,3,4-tetrahydropyrimidine-3,5-dicarbox-ylic acid 5-methylester-3-p- nitrophenylester (0. 208 mmol, 90.0 mg). The solution was stirred for 2h, poured onto saturated sodium bicarbonate, and extracted with ethyl acetate. The combined organic layers were washed with saturated sodium chloride, dried with magnesium sulfate, and concentrated in vacuo. The crude material was passed through silica (10% methanol, dichloromethane) to give the title product.

1H NMR (CDC13, 300 MHz) 8 8.94-8.01 (m, 1H) ; 7.50-7.00 (m, 7H); 6.60 (s, 1H) ; 3.72 (s, 1H) ; 3.42-3.32 (m, 2H) ; 2.63-2.43 (m, 4H); 2.27-2.19 (m, 2H); 1.85-1.58 (m, 6H).

Analysis: Calcd. for C21H29F3N405'1. 70 H20 C, 56.18; H, 5.66; N, 9.71 Found: C, 56.16; H, 5.70; N, 10.55 EXAMPLE 2 As a specific embodiment of an oral composition, 100 mg of the compound of Example 1 is formulated with sufficient finely divided lactose to provide a total amount of 580 to 590 mg to fill a size O hard gel capsule.

EXAMPLE 3 Screening assay : Alpha 1 a Adrenergic Receptor Binding Membranes prepared from the stably transfected human alpha la cell line (ATCC CRL 11140) were used to identify compounds that bind to the human alpha 1 a adrenergic receptor. These competition binding reactions (total volume = 200 ul) contained 50 mM Tris-HCI pH. 7.4,5 mM EDTA, 150 mM NaCI, 100 pM [125 I]-HEAT, membranes prepared from the alpha la cell line and increasing amounts of unlabeled ligand. Reactions wre incubated at room temperature for one hour with shaking. Reactions were filtered onto Whatman GFIC glass fiber filters with a Inotec 96 well cell harvester. Filters were washed three times with ice cold buffer and bound radioactivity was determined (Ki).

EXAMPLE 4 Selective Binding assays Membranes prepared from stably transfected human alpha I d and alpha lb cell lines (ATCC CRL 11138 and CRL 11139, respectively) were used to identify compounds that selectively bind to the human alpha I a adrenergic receptor.

These competition binding reactions (total volume = 200 ul) contained 50 mM Tris HCI pH. 7.4,5 mM EDTA, 150 mM NaCl, 100 pM [125 I]-HEAT, membranes prepared from cell lines transfected with the respective alpha 1 subtype expression plasmid and increasing amounts of unlabeled ligand. Reactions were incubated at room temperature for one hour with shaking. Reactions were filtered onto Whatman GF/C glass fiber filters with a Inotec 96 well cell harvester. Filters were washed three times with ice cold buffer and bound radioactivity was determined (Ki).

The compound of the present invention prepared in Example 1 was found to have alpha 1 a Ki value of about 0.12 nM as determined via the screening assay described in Example 3. The compound was further found to be about 729-fold more selective in binding to the alpha la receptor versus binding to the alpha lb receptor and 1900-fold

EXAMPLE 6 EXEMPLARY COUNTERSCREENS 1. Assay Title: Dopamine D2, D3, D4 in vitro screen Objective of the Assay : The objective of this assay is to eliminate agents which specifically affect binding of [3H] spiperone to cells expressing human dopamine receptors D2, D3 or D4.

Method: Modified from VanTol et al., Nature (1991), 350: 610-613.

Frozen pellets containing specific dopamine receptor subtypes stably expressed in clonal cell lines are lysed in 2 ml lysing buffer (lOmM Tris-HCI/5mM Mg, pH 7.4). Pellets obtained after centrifuging these membranes (15'at 24,450 rpm) are resuspended in 50mM Tris-HCl pH 7.4 containing EDTA, MgCl2, KCI, NaCl, CaCl2 and ascorbate to give a 1 Mg/mL suspension. The assay is initiated by adding 50-75 gg membranes in a total volume of 500 RI containing 0.2 nM [3H]-spiperone.

Non-specific binding is defined using 10 uM apomorphine. The assay is terminated after a 2 hour incubation at room temperature by rapid filtration over GF/B filters presoaked in 0.3% PEI, using 50mM Tris-HCI pH 7.4.

2. Assay Title : Serotonin5HTla Obiective of the Assay The objective of this assay i$ to eliminate agents which specifically affect binding to cloned human 5HTla receptor Method: Modified from Schelegel and Peroutka, Biochemical Pharmacology (1986), 35: 1943-1949.

Mammalian cells expressing cloned human 5HTla receptors are lysed in ice-cold 5 mM Tris-HCI, 2 mM EDTA (pH 7.4) and homogenized with a polytron homogenizer. The homogenate is centrifuged at 1000Xg for 30', and then the supernatant is centrifuged again at 38, OOOXg for 30'. The binding assay contains 0.25 nM [3H] 8-OH-DPAT (8-hydroxy-2-dipropylamino-1, 2,3,4-tetrahydronaphthalene) in 50 mM Tris-HCI, 4 mM CaCl2 and lmg/ml ascorbate. Non-specific binding is defined using 10 pM propranolol. The assay is terminated after a 1 hour incubation at room temperature by rapid filtration over GF/Cfilters EXAMPLE 7 EXEMPLARY FUNCTIONAL ASSAYS In order to confirm the specificity of compounds for the human alpha I a adrenergic receptor and to define the biological activity of the compounds, the following functional tests may be performed: 1. In vitro Rat, Dog and Human Prostate and Dog Urethra Taconic Farms Sprague-Dawley male rats, weighing 250-400 grams are sacrificed by cervical dislocation under anesthesia (methohexital; 50 mg/kg, i. p.).

An incision is made into the lower abdomen to remove the ventral lobes of the prostate. Each prostate removed from a mongrel dog is cut into 6-8 pieces longitudinally along the urethra opening and stored in ice-cold oxygenated Krebs solution overnight before use if necessary. Dog urethra proximal to prostate is cut into approximately 5 mm rings, the rings are then cut open for contractile measurement of circular muscles. Human prostate chips from transurethral surgery of benign prostate hyperplasia are also stored overnight in ice-cold Krebs solution if needed.

The tissue is placed in a Petri dish containing oxygenated Krebs solution [NaCI, 118 mM; KCI, 4.7 mM; CaCl2, 2.5 mM; KH2PO4, 1.2 mM; MgS04, 1.2 mM; NaHC03, 2.0 mM ; dextrose, 11 mM] warmed to 37 C. Excess lipid material and connective tissue are carefully removed. Tissue segments are attached to glass tissue holders with 4-0 surgical silk and placed in a 5 ml jacketed tissue bath containing Krebs buffer at 37 C, bubbled with 5% C02/95% 02. The tissues are connected to a Statham-Gould force transducer; 1 gram (rat, human) or 1.5 gram (dog) of tension is applied and the tissues are allowed to equilibrate for one hour.

Contractions are recorded on a Hewlett-Packard 7700 series strip chart recorder.

After a single priming dose of 3 uM (for rat), 10 uM (for dog) and 20 uM (for human) of phenylephrine, a cumulative concentration response curve to an agonist is generated; the tissues are washed every 10 minutes for one hour. Vehicle or antagonist is added to the bath and allowed to incubate for one hour, then another cumulative concentration response curve to the agonist is generated.

ECso values are calculated for each group using GraphPad Inplot software. pA2 (-log Kb) values were obtained from Schild plot when three or more concentrations were tested. When less than three concentrations of antagonist are tested, Kb values are calculated according to the following formula Kb =L x-1 where x is the ratio of ECso of agonist in the presence and absence of antagonist and [B] is the antagonist concentration.

2. Measurement of Intra-Urethral Pressure in Anesthetized Dog PURPOSE: Benign prostatic hyperplasia causes a decreased urine flow rate that may be produced by both passive physical obstruction of the prostatic urethra from increased prostate mass as well as active obstruction due to prostatic contraction.

Alpha adrenergic receptor antagonists such as prazosin and terazosin prevent active prostatic contraction, thus improve urine flow rate and provide symptomatic relief in man. However, these are non-selective alpha 1 receptor antagonists which also have pronounced vascular effects. Because we have identified the alpha la receptor subtype as the predominent subtype in the human prostate, it is now possible to specifically target this receptor to inhibit prostatic contraction without concomitant changes in the vasculature. The following model is used to measure adrenergically mediated changes in intra-urethral pressure and arterial pressure in anesthetized dogs in order to evaluate the efficacy and potency of selective alpha adrenergic receptor antagonists. The goals are to: 1) identify the alpha-1 receptor subtypes responsible for prostatic/urethral contraction and vascular responses, and 2) use this model to evaluate novel selective alpha adrenergic antagonists. Novel and standard alpha adrenergic antagonists may be evaluated in this manner.

METHODS: Male mongrel dogs (7-12 kg) are used in this study. The dogs are anesthetized with pentobarbital sodium (35 mg/kg, i. v. plus 4 mg/kg/hr iv infusion).

An endotracheal tube is inserted and the animal ventilated with room air using a Harvard instruments positive displacement large animal ventilator. Catheters (PE 240 or 260) are placed in the aorta via the femoral artery and vena cava via the femoral veins (2 catheters, one in each vein) for the measurement of arterial pressure and the administration of drugs, respectively. A supra-pubic incision-1/2 inch lateral to the penis is made to expose the urethers, bladder and urethra. The urethers are ligated and cannulated so that urine flows freely into beakers. The dome of the bladder is retracted to facilitate dissection of the proximal and distal urethra.

Umbilical tape is passed beneath the urethra at the bladder neck and another piece of umbilical tape is placed under the distal urethra approximately 1-2 cm distal to the prostate. The bladder is incised and a Millar micro-tip pressure transducer is advanced into the urethra. The bladder incision is sutured with 2-0 or 3-0 silk (pursestring suture) to hold the transducer. The tip of the transducer is placed in the prostatic urethra and the position of the Millar catheter is verified by gently squeezing the prostate and noting the large change in urethral pressure.

Phenylephrine, an alpha 1 adrenergic agonist, is administered (0.1-100 ug/kg, iv; 0.05 ml/kg volume) in order to construct dose response curves for changes in intra-urethral and arterial pressure. Following administration of increasing doses of an alpha adrenergic antagonist (or vehicle), the effects of phenylephrine on arterial pressure and intra-urethral pressure are re-evaluated. Four or five phenylephrine dose-response curves are generated in each animal (one control, three or four doses of antagonist or vehicle). The relative antagonist potency on phenylephrine induced changes in arterial and intra-urethral pressure are determined by Schild analysis. The family of averaged curves are fit simultaneously (using ALLFIT software package) with a four paramenter logistic equation constraining the slope, minimum response, and maximum response to be constant among curves. The dose ratios for the antagonist doses (rightward shift in the dose-response curves from control) are calculated as the ratio of the EDso's for the respective curves. These dose-ratios are then used to construct a Schild plot and the Kb (expressed as ug/kg, iv) determined.

The Kb (dose of antagonist causing a 2-fold rightward shift of the phenylephrine dose-response curve) is used to compare the relative potency of the antagonists on inhibiting phenylephrine responses for intra-urethral and arterial pressure. The relative selectivity is calculated as the ratio of arterial pressure and intra-urethral pressure Kb's. Effects of the alpha I antagonists on baseline arterial pressure are also monitored. Comparison of the relative antagonist potency on changes in arterial pressure and intra-urethral pressure provide insight as to whether the alpha receptor subtype responsible for increasing intra-urethral pressure is also present in the systemic vasculature. According to this method, one is able to confirm the selectivity of alpha 1 a adrenergic receptor antagonists that prevent the increase in intra-urethral pressure to phenylephrine without any activity at the vasculature.

While the foregoing specification teaches the principles of the present invention, with examples provided for the purpose of illustration, the practice of the invention encompasses all of the usual variations, adaptations and/or modifications that come within the scope of the following claims.

Claims (23)

1. WHAT IS CLAIMED IS: I. A compound of formula :

   wherein each X is independently hydrogen, halogen, cyano, C 1-C4 alkyl, C 1-C4 alkoxy, (CH2) 0-30CF3, or (CH2) 0-3CF3 ; and q is an integer from 0 to 5; or a pharmaceutically acceptable salt thereof.
2. 2. The compound according to claim 1, wherein the compound is a (+)-enantiomer ; or a pharmaceutically acceptable salt thereof.
3. 3. The compound according to claim 1, wherein the compound is a (-)-enantiomer ; or a pharmaceutically acceptable salt thereof.
4. 4. The compound according to claim 1, wherein each X is independently hydrogen, fluorine, cyano, methyl, ethyl, OCF3, or CF3 ; and q is an integer from 0 to 3; or a pharmaceutically acceptable salt thereof.
5. 5. The compound according to claim 1, having the formula :

   or a pharmaceutically acceptable salt thereof.
6. 6. The compound according to claim 5, wherein each X is independently hydrogen, fluorine, cyano, methyl, ethyl, OCF3, or CF3 ; or a pharmaceutically acceptable salt thereof.
7. 7. The compound according to claim 6, which is

   or a pharmaceutically acceptable salt thereof.
8. 8. A pharmaceutical composition comprising the compound according to claim 1 and a pharmaceutically acceptable carrier.
9. 9. A pharmaceutical composition made by combining the compound according to claim 1 and a pharmaceutically acceptable carrier.
10. 10. A process for making a pharmaceutical composition comprising combining the compound according to claim 1 and a pharmaceutically acceptable carrier.
11. 11. The composition according to claim 8 further comprising a testosterone 5-alpha reductase inhibitor.
12. 12. The composition according to claim 11, wherein the testosterone 5-alpha reductase inhibitor is a type 1, a type 2, both a type 1 and a type 2, or a dual type 1 and type 2 testosterone 5-alpha reductase inhibitor.
13. 13. The composition according to claim 12, wherein the testosterone 5-alpha reductase inhibitor is a type 2 testosterone 5-alpha reductase inhibitor.
14. 14. The composition according to claim 13, wherein the testosterone 5-alpha reductase inhibitor is finasteride.
15. 15. A method of treating benign prostatic hyperplasia in a subject in need thereof which comprises administering to the subject a therapeutically effective amount of the compound according to claim 1.
16. 16. The method according to claim 15, wherein the compound does not cause a fall in blood pressure at dosages effective to alleviate benign prostatic hyperlasia.
17. 17. The method according to claim 16, wherein the compound is administered in combination with a testosterone 5-alpha reductase inhibitor.
18. 18. The method according to claim 17, wherein the testosterone 5alpha reductase inhibitor is finasteride.
19. 19. A method of treating benign prostatic hyperplasia in a subject in need thereof which comprises administering a therapeutically effective amount of the composition according to claim 8.
20. 20. The method according to claim 19, wherein the composition further comprises a therapeutically effective amount of a testosterone 5-alpha reductase inhibitor.
21. 21. A method of relaxing lower urinary tract tissue in a subject in need thereof which comprises administering to the subject a therapeutically effective amount of the compound according to claim 1.
22. 22. The method according to claim 21, wherein the compound is administered in combination with a testosterone 5-alpha reductase inhibitor.
23. 23. The method according to claim 22, wherein the testosterone 5alpha reductase inhibitor is finasteride.