WO2009037556A1

WO2009037556A1 - New agents for the treatment of the low urinary tract dysfunctions

Info

Publication number: WO2009037556A1
Application number: PCT/IB2008/002443
Authority: WO
Inventors: Giancarlo Santus; Piero Del Soldato; Anna Sparatore
Original assignee: CTG PHARMA Srl
Current assignee: CTG PHARMA Srl
Priority date: 2007-09-21
Filing date: 2008-09-19
Publication date: 2009-03-26
Anticipated expiration: 2010-03-21
Also published as: ITMI20071827A1

Abstract

The present invention relates to novel compounds that are derivatives of 5 -phosphodiesterase inhibitors that comprise in their formula a polysulfurated group and that are useful for treating dysfunctions of low urinary tract such as incontinence, benign prostatic hyperplasia and erectile dysfunction.

Description

TITLE OF THE INVENTION

"New agents for the treatment of the low urinary- tract dysfunctions" .

Background of the invention

This invention relates to new compounds that are derivatives of 5-phosphodiesterase inhibitors (PDE-5) for the treatment of the low urinary tract dysfunctions (LUTS) including incontinence, benign prostatic hyperplasia (BPH) and erectile dysfunction (ED) .

Although none of these conditions are fatal, they affect overall quality of life of the patients and constitute the basis for more serious pathologies. Urinary incontinence is defined as the involuntary and uncontrolled loss of urine in conditions socially impossible .

Urinary incontinence can be of different types: urge incontinence (urgency and frequency) , stress incontinence and mixed urinary incontinence. Urgency-frequency is the increased micturition frequency with a strong desire to void. Several can be the causes neurological and not, but all leading to an increased bladder contraction, that is commonly defined as vesical hyperactivity or detrusor hyperreflexia depending on its neurogenic or not neurogenic origin.

Benign prostatic hyperplasia is an hormone- dependent process that begins at 30-35 years and continues in a variable way up to 70 years. This hormone-dependent process requires dihydro- testosterone (DHT) that through a series of molecular interactions exerts an influence on the prostatic cells. The androgenic activity of dihydrotestosterone depends on its linkage with the high affinity receptor for androgens present in the cellular nucleus and its affinity is five times superior to the testosterone one. Dihydrotestosterone is the main metabolite of testosterone and both in prostate and skin it is converted to dihydrotestosterone by 5- alpha reductases .

Looking at the natural evolution of benign prostatic hyperplasia versus the prostatic volume, it is evident that volume increases with the increase of age . Tiresome symptoms are LUTS that according to progression of the disease can interfere with sleep/waking rhythm of the patients and also with daily activities and therefore to compromise the quality of life. These symptoms get worse with age.

The principal factors for benign prostatic hyperplasia are: an increase of the specific prostatic antigen (PSA) , an increase of the prostatic volume and the age; secondary symptoms to be considered are the urine flow rate and the increase of LUTS.

Benign hyperplasia and prostate carcinoma appear often with overlapping and sometimes hazy symptoms; therefore every "prostatic" patient must be always evaluated keeping in mind these pathologies .

Erectile dysfunction (ED) is the persistent and consistent inability to achieve and/or maintain penile erection sufficient to have a satisfactory sexual intercourse. ED is a widely diffused disease in male population and it is treatable in 95% of the subjects .

Even if psychological causes play a fundamental role in sexual functionality and in the desire mechanisms, the major part of erection's disturbances has a physical background. It must be remembered that in the male non diabetic population the incidence of erectile dysfunction is approximately of 13% with variations of 2.5% at 40 years of age and 25-20% at 65 years. In diabetic subjects the same problems is more severe and ED is present in 15% of cases between 20 and 40 years of age to reach 40-60% between 50 and 70 years.

Despite the incidence of erectile dysfunction in diabetic male population and the negative impact on quality of life this anomaly does not receive yet from the diabetologists and the patients the same attentions reserved to the other complications. The phosphodiesterases inhibitors act by inhibiting in selective way the PD-5 enzymes present in the various tissues, such as penile tissue, platelets, vascular tissue, smooth muscle tissue etc. The activity of phosphodiesterase-5 inhibitors on these tissues has achieved therapeutic successes in patients suffering of low urinary tract dysfunctions, pulmonary hypertension, etc.

The inhibitors of phosphodiesterases-5 act also on erectile tissue causing the relaxation of penile muscles, and subsequent vasodilatation leading to penis erection.

Drugs that are inhibitors of PDE-5 include sildenafil (Viagra ) , vardenafil (Levitra ) , tadalafil (Cialis ) , udenafil (Zydena ) , avanafil .

Recently it has been observed that oxidative stress conditions play a relevant role in the pathological process above described. It has been also observed that a gaseous compound such as hydrogen sulphide (H₂S) plays an active role in such redox processes although the physiological and physio-pathological characteristics remain to be defined.

Summary of the invention

This invention relates to new hybrid derivatives of phosphodiesterases inhibitors comprising in their formula a polysulfurated group and that are useful for treating diseases and pathological conditions of the low urinary tract. The results have been surprising because not only the safety was dramatically improved but also efficacy was sometimes increased as compared with known inhibitors of phosphodiesterases. The polysulfurated groups object of the present invention contain 2 or more atom of sulphur. The polysulfurated groups of the present invention linked to compounds inhibitors of phosphodiesterases are organic thiosulfonates or dithiole-thione derivatives such as 5- (4- hydroxyphenyl) -3H-1, 2-dithiol-3-thione, or 4- (4- hydroxyphenyl ) -3H-1, 2-dithiol-3-thione, or 4- (3- thioxo-3H-l,2-dithiol-4-yl) benzoic acid, or 4- (3- thioxo-3H-l,2-dithiol-5-yl) benzoic acid, 1,3-dithiol- 2~thione~5-carboxylic acid, 3-thioxo-3H-l, 2-dithiole- 5-carboxylic acid, 3-thioxo-3H-l, 2-dithiole-4- carboxylic acid, or linear dithiole-thiones or trithiocarbonates . This invention also relates to processes for preparing these compounds and to pharmaceutical compositions containing these compounds . Description of the invention

An object of the present invention are new compounds inhibitors of phosphodiesterases of general formula:

A-Y-W (I) wherein

A is selected from the group comprising the residues of PDE- 5 inhibitors having formula ( I I or II I or IV or V) :

(I I I )

wherein X is a group capable to link to ~Y o ~W, selected from a group comprising -C00~; -O- ; -CONH-;

-OCO-; -0C00-; -CO-; - (C_p) alkyl-COO- ; - (C₅) alkyl-0-; - (C_p) alkyl-CONH-; - (C_p) alkyl-0C0~; - (C_p) alkyl-0C00-; - (C_p) alkyl-C0~; wherein (C_p) alkyl is straight or branched and p is 1-10;

Y is zero; - (C_n- ) alkyl-, - (C_n- ) alkyl-CO- , -0- (C_n-) alkyl -0-, -00C- (C_n- ) alkyl-COO- ; -0- (C_n- ) alkyl- , -HN- (C_n')alkyl-, ~OOC- (C_n- ) alkyl- ; ~ (C_n. ) alkyl-O-CO- (C_n..) alkyl-; ~ (C_n. ) alkyl-CO-O- (C_n.0 alkyl- wherein (C_n-) alkyl and (C_n. <) alkyl are straight or branched, and n' and n' ' , the same or different to each other, are 0-10;

W is a polysulfurated group containing 2 or more atoms of sulphur, selected from the group comprising an organic thiosulfonate moiety or a dithiole-thione derivative cyclic or linear or trithiocarbonates : more in particular, as a further preferred embodiment, W is an organic thiosulfonate moiety having formula:

-S-SO₂-R (VI) wherein -S-SO₂-R is linked to A-Y-; R is a straight or branched alkyl , such as methyl , ethyl , propyl ; alkenyl, alkinyl ; alkylaryl, alkenylaryl, alkinylaryl ; arylalkyl, arylalkenyl, arylalkinyl; or cycloalkyl, cycloalkenyl , cycloalkinyl ; or aromatic and/or heterocyclic ring, all substituted or unsubstituted; or more in particular, as a further preferred embodiment, W is a dithiole-thione derivative having the following formula:

(VII) or (VIII)

wherein

Z is S (sulphur) and at least 1 Z is C=S (thione) , m is 0, 1-10;

T is :

~OOC- ; or

wherein

Rl is -H; -COOH; -NH₂; -OH; -SH;

R2 is hydrogen; -COOH; alkyl, alkenyl, alkynyl ; aryl ; fluoro, chloro, bromo; hydroxyl , alkyloxy, alkenyloxy, aryloxy, acyloxy; amino, alkylamino, arylamino; thio; cyano; nitro; acyl; amido; and a 5 or 6-merabered aromatic or non-aromatic ring containing 0, 1, or 2 heteroatoms selected from nitrogen, oxygen, or sulphur; and related salts.

As a further preferred embodiment of the compounds of general formula (I) of the present invention (Cjalkyl, (C_n. ) alky1 and (C_n-.) alky1 are (CHa)_nA , (CH₂) nA_' , (CH₂) nA_" respectively, wherein nA, nA' and nA' ' , the same or different to each other, are 1-10, such as that more preferably Y is selected from the group comprising - (CH₂) _ΠA' - , ~ (CH₂) nA_' -CO- , -O- (CH₂) _IA' -O- , ~00C- (CH₂) _ΠA- -COO-; -O- (CH₂) ^. -, -HN- (CHa)_nA_'-, -00C- (CH₂)HA.-; - (CH₂) ^- -0-C0- (CH₂) nA- -; - (CH₂) _ΠA. -CO-O- (CH₂) nA_"- wherein nA, nA' and nA' ' , the same or different to each other, are 1-10.

A further preferred embodiment of the derivative compounds of phosphodiesterases inhibitors according to the present invention are the compounds of general formula (I) wherein the group -Y-W is selected from the group comprising thiosulfonate moieties derived from the corresponding precursors having formula: S- (2-carboxyethyl) methanthiosulfonate, S- (2- aminoethyl) methanthiosulfonate and S- (2 -hydroxyethyl) methanthiosulfonate .

A further preferred embodiment of the derivative compounds of phosphodiesterases inhibitors compounds according to the present invention, are the compounds of general formula (I) wherein the polysulfurated group W is selected from the group comprising dithiole-thione derivatives of the corresponding precursors having formula: 5- (p-hydroxyphenyl) -3H- 1, 2-dithiol-3-thione, 1, 3-dithiol-2-thione-5- carboxylic acid, 3-thioxo-3H-l, 2-dithiole-5- carboxylic acid, 3-thioxo-3H-l, 2-dithiole-4- carboxylic acid.

The compounds inhibitors of phosphodiesterases, in particular those selected from the group comprising sildenafil, vardenafil, tadalafil, udenafil, avanafilo and the moiety containing the polysulfurated group can be linked via different linking groups such as esters, amides, imides, sulfonamides, azo groups, carbamates, carbonates, anhydrides, acetals, thioacetals, etc. The polysulfurated group, i.e. the thiosulfonate moiety or dithiol-thionic derivative, can be also directly linked by an ionic bond to the compound inhibitor of phosphodiesterases as salt when Y=O .

Bi-functional linkers (Y) , known to the expert in the field, (such as ethyl, propyl, or butyl diols; diamines; hydroxy amines; etc.) can be optionally present when they are necessary to link the drug (phosphodiesterases inhibitor) to the polysulfurated group .

As a further object of the present invention are the preferred compounds according to general formula (I) , such as : 4- (5-thioxo-5H-l, 2-dithiol-3-yl) phenyl 2- (4- (4- ethoxy-3 - (5-methyl-4-oxo-7-propyl-1,4-dihydroimidazo [1, 5-jf] [1,2,4] triazin-2 -yl) phenylsulfonyl) piperazin- 1-yl) acetate

4- (5-thioxo-5H-l, 2 -di thiol- 3 -yl) phenyl 2-(2-(2-(3-l- methyl - 7 -oxo- 3 -propyl - 6 , 7 -dihydro- lH-pyrazole [4,3- d] pyrimidin-5-yl) -4-propoxyphenylsulf onamido) pyrrolidin-1-yl) acetate .

Ester of (6R, 12aR) -2,3, 6, 7, 12, 12a-esahydro-2-hydroxy- 6- (3, 4-methylendioxyphenyl) -pyrazino [2' , 1' : 6, l]pyrido [3,4-b] indole-l,4-dione with 4- (3-thioxo-3H-l, 2- dithiol-4-yl) benzoic acid.

Ester of (6R, 12aR) -2, 3, 6, 7, 12, 12a-esahydro-2-hydroxy- 6- (3 ,4-methylendioxyphenyl) -pyrazino [2' , 1' : 6, l]pyrido [3 , 4-b] indole-1, 4-dione with 3-methansulfonyl sulfanyl propionic acid.

4- (5-thioxo-5H-l,2-dithiol-3-yl)phenyl (6R,12aR) 2 , 3 , 6, 7, 12 , 12a-esahydro-6- (3 , 4-methylendioxyphenyl) -pyrazino [2' , 1' :6, l]pyrido [3, 4-b] indole-1, 4-dione-2- acetate

It is a further objective of the present

invention compounds of general formula (I) wherein A

is a residue of a PDE-5 inhibitor having formula IX:

(IX) W is a dithiole-thione derivative of formula (VIII) :

~T-(CH₂)rr^-z_χ ^z—(CH₂)m—CH₃

^Z (VIII)

X, Y e m have the meaning as above reported.

When the compounds include at least one

asymmetric carbon atom, the products can be used in

racemic mixture or in form of single enantiomer.

In the present invention the parent compound

is considered in its original form or in a proper

modification to allow the chemical manipulation with the polysulfurated group.

It is a further object of the present invention

the pharmaceutical acceptable salts of compounds of

formula (I) , such as for example salts with alkaline metals and alkaline earth metals, non-toxic amines and aminoacids, inorganic acids such as hydrochloric acid, phosphoric acid, etc. or organic acids such as fumaric acid, citric acid, tartaric acid, etc.

Salts of organic thiosulfonates such as, for example, S- (2-carboxyethyl)methanthiosulfonate, S-

(2-aminoethyl) methanthiosulfonate with the different derivatives inhibitors of PDE-5 above described, are also part of the present invention. Salts of dithiolthiones such as, for example, 1 , 3-dithiol-2- thione-5-carboxylic acid, 3-thioxo-3H-l, 2-dithiole-5- carboxylic acid, 3-thioxo-3H-l, 2-dithiole-4- carboxylic acid with the different inhibitor of PDE-5 compounds above described are also part of the present invention.

According to the present invention it has been found that it is possible to link an organic polysulfurated group to an inhibitor of the phosphodiesterases for treating the low urinary tract dysfunctions. The resulting compounds have good bioavailability, increased safety and maintain good efficacy.

The main advantages of the compounds of the present invention are related to their biological activity for the treatment of patients with dysfunctions of the low urinary tract such as incontinence, benign prostatic hyperplasia, erectile dysfunction.

Surprisingly it has been found that the compounds of the present invention, having general formula (I) and the preferred compounds as above described, are active also treating cardiovascular diseases such as angina, hypertension, stroke, atherosclerosis; gastrointestinal diseases, such as irritable bowl syndrome (IBD) ; diabetic and not diabetic gastropathy; diseases where cytoprotection is important such us for ulcer healing and prevention.

It has been further found that the ester of the sildenafil analogue with 5- (p-hydroxyphenyl) -3H-1, 2- dithiol-3-thione having formula:

is active for the treatment of the low urinary tract dysfunctions such as incontinence, benign prostatic hypertrophy. Further object of the present invention are pharmaceutical compositions that contain at least one compound of the above mentioned compounds derivatives of phosphodiesterases inhibitors (according to the present invention as for general formula (I) and the preferred compounds as described above) including salts thereof, as an active ingredient, moreover, as a further object of the present invention, in combination with pharmaceutically acceptable adjuvant (s) or carrier (s) .

It is a further object of the present invention the use of the derivatives of phosphodiesterases inhibitors as for general formula (I) , and the preferred compounds as described above, as a medicament .

As a further object of the present invention is the use of compounds according to the present invention, as for general formula (I) and the preferred compounds as described above, for the preparation of a pharmaceutical composition, and therefore the corresponding method, for treating low urinary tract dysfunctions, such as incontinence, benign prostatic hyperplasia, erectile dysfunctions, also in combination with other agents such as for example, alpha blockers such as terazosin, doxazosin, tamsulosin etc., 5-alpha reductase inhibitors such as finasteride, etc.

Accordingly it is a further object of the present invention a method of treating low urinary tract dysfunctions such as incontinence, benign prostatic hyperplasia and erectile dysfunction, which comprises administering to a patient suffering from such diseases a therapeutically effective amount of a compound according to the present invention as for general formula (I) , and the preferred compounds as described above.

The compounds of the present invention can be administered in the form of any pharmaceutical formulation, the nature of which will depend upon the route of administration and the nature of the disease to be treated. These pharmaceutical compositions can be prepared by conventional methods, using compatible, pharmaceutically acceptable excipients or vehicles .

Examples of such compositions include capsules, tablets, syrups, powders and granulates for the preparation of extemporaneous solutions, injectable preparations, rectal, nasal, ocular, vaginal etc. A preferred route of administration is the oral route.

It is a further object of the present invention the process of the synthesis of the above said derivatives of phosphodiesterases inhibitors

(according to the present invention as for general formula (I) , and the preferred compounds as described above) including salts thereof, said process comprising at least a reaction between the corresponding precursor of an organic thiosulfonate or dithiol-thione or a trithiocarbonate, moiety W or

Y-W, in particular selected from the group comprising: S- (2-carboxyethyl) methanethiosulfonate, S- (2-aminoethyl) methanethiosulfonate and S- (2- hydroxyethyl) methanethiosulfonate or 5- (4- hydroxyphenyl) -3H-1, 2-dithiol-3-thione, 4- (4- hydroxyphenyl ) -3H-1, 2-dithiol-3-thione, or 4- (3- thioxo-3H-l, 2-dithiol-4-yl) benzoic acid, or 4- (3- thioxo-3H-l, 2-dithiol-5-yl) benzoic acid, 1,3-dithiol- 2-thione-5-carboxylic acid, 3-thioxo-3H-l , 2-dithiole- 5-carboxylic acid, 3-thioxo-3H-l, 2-dithiole-4- carboxylic acid and a derivative of a PDE-5 inhibitor or a corresponding precursor of a PDE-5 inhibitor modified with Y wherein the PDE-5 inhibitor is selected in the group comprising vardenafil, udenafil, tadalafil, avanafil and wherein W and Y have the above said meaning . The following non-limitative examples further describe and enable an ordinary skilled in the art to make and use the invention.

EXAMPLE 1. Synthesis of 4- (5-th.ioxo-5.H-l , 2-dithiol-3- yl) phenyl 2- (4- (4-ethoxy-3- (5-methyl-4-oxo-7-propyl- 1, 4-dihydroimidazo [1, 5-f] [1,2,4] triazin-2-yl) phenylsulfonyl) piperazin-1-yl) acetate .

Step 2: Preparation of 5- (4-hydroxyphenyl) -3H-1, 2- dithiole-3 -thione To 280 mtnol of sulphur, 40 mmol of anethole in 20 ml of dimethylacetamide were added. After heating at 145 ⁰C for 6 hours, 2.5 g of anethole dithiolethione (ADT) were obtained. The product, washed with ether, was crystallized by ethyl acetate: melting point 110-111 ⁰C. Then 1.5g of ADT were mixed with 7.5g of pyridine hydrochloride and the mixture was heated for 25 minutes at 215 ⁰C. After cooling, IN HCl in excess was added and the precipitate was filtered, washed and crystallized from ethanol . The obtained compound, 5- (p- hydroxyphenyl ) -3H-1, 2-dithiol-3-thione, melted at 191-192 ⁰C.

Step 2: Synthesis of 2- (4- (4-ethoxy-3- (5-methyl-4- oxo-7-propyl-1, 4-dihydroimidazo [1, 5-f] [1,2,4] triazin-2-yl) phenylsulfonyl) piperazin-1-yl) acetic acid a) Synthesis of 2-butyrylaminopropionic acid DL-alanine (4.061 g) are dissolved in NaOH 4N (34.2 ml, 3 eq.) . Butyrylchloride (1.1 eq.) is slowly added to this solution under stirring and keeping it cold. The ice bath is removed and the pH is corrected to 1.5 with cone. HCl. The product is extracted with ethyl acetate and the organic phases, dried on anhydrous Na₂SO₄, are evaporated to dryness. A pale yellow oil, that crystallizes after treatment with petrol ether, is obtained. After washing with ether and re-crystallization with a mixture of ethyl acetate and petrol ether 7 g of product are obtained. b) Synthesis of 2-ethoxybenzamidine hydrochloride NH₄Cl (1.26 g, 23.52 mmoles) is introduced in a 100 ml round bottom flask and suspended in 16 ml of anhydrous toluene under argon atmosphere . The mixture is cooled in an ice bath and 2N Al (CH₃) ₃ (12 ml) in toluene are added dropwise, keeping under stirring for 30' at room temperature. 2-Ethoxybenzonitrile (Ig) is added and the reaction is refluxed with stirring for 9 hours. Silica (0.8g) and then methanol (8 ml) are added to the mixture of reaction to eliminate the excess of Al (CH₃) ₃. The suspension is diluted with 8 ml of dichloromethane and then filtered washing the obtained solid several times with methanol .

The filtered solution is acidified with cone. HCl and evaporated to dryness. The obtained residue is treated with 12 ml of a mixture of dichlorometane/methanol (9:1), filtered and the organic solution evaporated to dryness . The residue is washed with ether. c) Synthesis of 2- (2-ethoxyphenyl) -5-methyl-7-propyl- 3ff[5,lf] [1,2,4] triazinone

2-butyrylaminopropionic acid (633 mg) , dimethylaminopyridine (DMAP, 16 mg) , pyridine (1 ml) in 2.7 ml of tetrahydrofurane (THF) are introduced in a 25 ml flask and heated at 55 ⁰C. Ethylchloro- oxalate (0.88 ml) is added and the mixture is refluxed for 2 hours. After cooling, iced water is added to the mixture and the enol ester is extracted with ethylacetate .

Separately, in a flask cooled on an ice bath, a mixture of 800 mg of 2-ethoxybenzamidine hydrocloride (prepared in step b) and 3.2 ml of methanol are added slowly with 0.19 ml of hydrazine hydrate in methanol. The ice bath is removed and the mixture is maintained under stirring for 10 minutes. The previously prepared enolester in 0.5 ml of methanol is added and the reaction is refluxed for 4 hours. The 1,2,4- triazin-5 (4H) one is obtained and used without isolating it.

In a 50 ml flask are introduced 3.98 mmoles of 1, 2, 4-triazin-5 (4H) one in 6 ml of acetic acid and 0.34 ml of POCl₃ and the mixture was heated at 110 ⁰C for 2 hours. The acetic acid is removed at reduced pressure and methylene chloride and water are added, adjusting the pH to 7 with 2N NaOH. The aqueous solution is extracted with methylene chloride, the organic phase is dried on Na₂SO₄, filtered and evaporated to dryness. A red orange oil, that crystallizes after adding acetone, is obtained, d) Synthesis of 2- (4- (4-ethoxy-3- (5-methyl-4-oxo-7- propyl-l,4-dihydroimidazo [1,5-f] [1,2,4] triazin-2-yl) phenylsulfonil) piperazin-1-yl) acetic acid

210 mg of the compound prepared in step c) (0.67 mmoles) are dissolved in 2 ml of dichloromethane and after cooling, 0.5 ml of chlorosulfonic acid are added. The reaction mixture is stirred at room temperature, under nitrogen for 3 hours. After adding cold water, the organic phase is separated, dried on anhydrous Na₂SO₄ and evaporated to dryness. The obtained solid (195 mg, 0.54 mmoles) is dissolved in 4 ml o£ dichloromethane, triethylamine (0.08 ml, 1 eq) is added and subsequently 1-ethoxycarbonylmethyl- piperazina (0.09 ml, 1 eq) is added. The reaction mixture is stirred for 3 hours at room temperature under nitrogen. At the end of the reaction, the organic phase is washed with cold water, dried on anhydrous Na₂SO₄ and evaporated to dryness and the crude product is purified on silica gel column, eluting with CH₂Cl₂ containing 0.5-1% of methanol.

The obtained compound (210 mg, 80.4 mmoles) is suspended in 2.9 ml of NaOH IN and 1.5 ml of ethanol heating at 90 ⁰C for 2 hours. At the end of the reaction, ethanol is evaporated, the aqueous phase is acidified with 2N HCl up to pH 2.3 and extracted with a mixture of chloroform/methanol (9:1) obtaining a white solid that, after washing with ether, melts at 222-224 ⁰C

Step 3: Synthesis of 4- (5-thioxo-5H-l, 2-dithiol-3- yl) phenyl 2- (4- (4-ethoxy-3- (5-methyl-4-oxo-7-propyl- 1,4-dihydroimidazo [1,5-f] [1,2,4] triazin-2-yl) phenylsulfonyl) piperazin-1-yl) acetate

A solution of dicyclohexylcarbodiimide (DCC) )

(71.8 mg; 1.2 eq.) in dichlorometane is added dropwise to a suspension of the compound described in step 2 (142 mg; 0.2738 mmoles), 5- (4-hydroxyphenyl) -

3H-l,2-dithiol-3-thione (62 mg; 1 eq) prepared as above described (step 1) and dimethylaminopyridine (DMAP, 7 mg) in 15 ml of CH₂Cl₂. The reaction was performed at room temperature, stirring under nitrogen for 5 hours. At the end of the reaction, the dicyclohexylurea (DCU) was removed by filtration. The solution was evaporated to dryness and the residue was chromatographed on a silica gel column, eluting with a mixture of CH₂Cl₂/methanol (99:1) . An orange- coloured solid is obtained which, after washing with a mixture ether/ethanol (8:2), melts at 165-168 ⁰C. EXAMPLE 2. Synthesis of ester of (6R,12aR)- 2, 3, 6, 7, 12, 12a-esahydro-2 -hydroxy-6- (3,4- methylendioxyphenyl) -pyrazino [2 ' ,1' :6,l]pyrido [3,4-b] indole-1, 4 -dione with 4- (3-thioxo-3H-l, 2- dithiol-4-yl) benzoic acid. Step 1:

Thionyl chloride (SOCl₂; 2.4 ml) is added to a suspension of D-tryptophan (3g; 14.7 mmoles) in 20 ml methanol, stirring under nitrogen atmosphere in ice bath. The solution is then refluxed (~68°C) for 1.5 hour. Methanol is evaporated and 20 ml of tert-butyl- methyl ether (MTBE) are added. The solution is stirred in ice bath for 1 hour, then it is filtered and the product is washed with cold MTBE. The obtained white solid is dried under vacuum at 60⁰C. Steps 2 :

Piperonyl alcohol (4.08 g, 0.026 mmoles) , aluminium isopropoxide 98% and toluene (45 ml) are added in a round bottom flask under nitrogen atmosphere . The mixture is heated to -110⁰C and after addition of 1.6 g of p-formaldehyde, refluxed for 2 hours. Thereafter the mixture is cooled and 60 ml of 1 M NaOH are added. The 2 phases are separated; the organic phase is dried with Na₂SO₄, filtered and evaporated maintaining the bath temperature at 3O⁰C. The product is purified by chromatography (silica, cyclohexane/ethyl acetate, 90:10). Step 3:

D-triptophan metyl ester hydrochloride (2.8 g; 0.011 mmoles) is suspended in 25 ml isopropanol, under nitrogen atmosphere, at room temperature. A solution of piperonal prepared in step 2 (1.750 g, 0.012 moles) in 2 ml isopropanol is added and the reaction is maintained overnight at 66-70 ⁰C. The mixture is filtered, washed with cold isopropanol and dried. The tetrahydro-beta-cis-carboline hydrochloride is obtained. Step 4:

Triethylamine (2.4 ml) is added to a suspension of the tetrahydro-beta-cis-carboline hydrochloride (2.6 g; 6.7 mmoles) , prepared in step 3, in 14 ml of anhydrous tetrahydrofurane (THF) and the temperature is cooled to 0⁰C under nitrogen atmosphere. Then, chloroacetyl chloride (1.047 g; 0.72 ml) in 2 ml anhydrous THF is added dropwise. After 1 hour the reaction mixture is evaporated, ethyl acetate is added and the organic solution is washed with water in a separator funnel . The organic phase is dried with Na₂SO₄, filtered and evaporated. The product is washed with cold isopropanol . Step 5:

Hydroxylamine (NH₂OH) hydrochloride (49 mg; 0.7 mmoles), H₂O (0.15 ml) and diisopropylethylamine (0.12 ml; 0.7 mmoles) are added to a solution of the chloroacetyl carboline, prepared in step IV (150 mg; 0.35 mmoles) in 1 ml anhydrous THF. The mixture is heated at 45 ⁰C for 24 hours. After filtration a white product is obtained.

Step 6: Preparation of 4- (3-thioxo-3H-l, 2-dithiol-4- yl)benzoic acid

4-Isopropylbenzoic acid (1.0 g; 6.09 mmol) was suspended in 25 ml of ethanol and to this suspension 0.140 g of cone. H₂SO₄ were added. The reaction was performed at 100° C, stirring for 9 hours. The solution was evaporated to dryness and the residue was dissolved in CH₂Cl₂. The organic phase was washed with a saturated NaHCO₃ solution and then with brine, dried on anhydrous sodium sulphate and finally- evaporated to dryness to obtain an oily colourless product. This product (ethyl 4-isopropyl benzoate,940 mg; 5.16 mmol) was added dropwise to stirred melted sulfur (1.2 g) at 146°C and the reaction mixture was stirred at 220 °C for 24 hours. The temperature is lowered to 110 ^°C and 3 ml of toluene and 7 ml of acetone were added. After stirring the reaction mixture at r.t. for 4 h, unreacted sulphur was filtered and the obtained solution was evaporated to dryness. The residue was purified by column chromatography on silica gel, eluting with CH₂Cl₂- cyclohexane (6:4) to give a compound with m.p. 157.5- 159.5°C. Finally a suspension of this compound (100 mg, 0.35 mmol) in 4.5 ml of acetic acid and 0.72 ml of 9M H₂SO₄ was stirred at 100⁰C for 4 hours. After cooling, the solution was diluted with water and extracted with a mixture of CH₂Cl₂-methanol (9:1). The organic phase was dried on anhydrous sodium sulphate and evaporated to dryness and the residue was washed with ether and CH₂Cl₂ to obtain a yellow- orange solid, 4- (3-thioxo-3H-l, 2-dithiol-4-yl) benzoic acid, with m.p. 240-245 ^°C. Step 7 :

The product prepared in step V (80 mg, 0.2 mmoles) dissolved in anhydrous DMF is added to a solution of the product prepared in step VI (52 mg; 0.2 mmoles), N-hydroxybenzotriazole (HOBT 34 mg; 0.22 mmoles) and 1- (3-dimethylaminoisopropyl) -3-ethyl-carbodiimide hydrochloride (EDAC 42 mg; 0.22 mmoles) in anhydrous DMF (1 ml) . The reaction is stirred at r.t. for 24 hours, then the solution is evaporated, CH₂Cl₂ is added to the residue and the solution is washed in a separator funnel with sodium bicarbonate. The organic phase is dried with Na₂SO₄, filtered and evaporated. The product is chromatographed on silica eluting with CH₂Cl₂/Me0H (99.5:0.5). The product has mp 210-220⁰C with decomposition.

EXAMPLE 3. Synthesis of 4- (5-thioxo-5H-l, 2-dithiol-3- yl) phenyl (6R, 12aR) -2,3, 6, 7, 12, 12a-esahydro-6- (3,4- methylendioxyphenyl) -pyrazino[2' ,1' :6,l]pyrido [3 , 4-b] indole-1, 4-dione-2-acetate

The synthesis is the same of example 2 for steps from 1 to 4. Step 5:

Triethylamine (0.38 ml, 2.5 mmoles) is added to a solution of the chloroacetyl carboline (Ig; 2.3 mmoles) , prepared in step 4 of example 2 and glycine methyl ester hydrochloride 99% (379 mg; 3 mmoles) in DMF (0.4 ml) and the mixture is stirred under nitrogen atmosphere at 78⁰C. After 4 hours the mixture is cooled and after evaporation and the residue is dissolved in CH₂Cl₂ and washed with water. The product is chromatographed on silica, eluting with CH₂Cl₂/Me0H mixture in gradient .

The obtained ester is stirred with IN NaOH (8.8 ml) and methanol (4.4 ml) at 40⁰C for 2 hours for the hydrolysis. After treatment with IN HCl, the acid precipitates and it is filtered and washed. Step 6:

1- (3-dimethylaminoisopropyl) -3-ethyl-carbodiimide hydrochloride (EDAC 232 mg; 1.21 mmoles) is added to a suspension of the compound prepared in step V (480 mg; 1.1 mmoles), the compound prepared in example 1 step 1 (269 mg; 1.18 mmoles), and a catalytic amount of 4-dimethylaminopyridine (DMAP, 13 mg) in CHCl₃ (10 ml) . The reaction is stirred at r.t. for 4 hours. After evaporation, the residue is dissolved in CH₂Cl₂ and washed with cold water, sodium bicarbonate, NaOH and finally with water again. The organic phase is dried with Na₂SO₄, filtered and evaporated. The product is chromatographed on silica eluting with cyclohexane/ethyl acetate (1:1) and then washed with ethyl ether. It has a mp (166)-182°C with decomposition .

EXAMPLE 4. Biological determinations of cyclic nucleotides (cGMP, cAMP) and glutathione. cGMP, cAMP and glutathione were measured in normal rats and in rats treated with buthionione sulphoxime (BSO) to induce oxidative stress adding BSO, (30 mmol/L/giorno) to the water of the diet for 7 days. The control group received tap water. The compound of example 1 (5 mg/kg/day) alone or in combination with BSO as administered orally for 7 days. cGMP, cAMP and total glutathione were measured in several tissues (prostate, urhetra corpus cavernosum with specific kits ELISA kits (581021 and 581001 respectively; Cayman Chemical Co.).

a) Effect on cGMP

b) Effect on cAMP

c) Effect on glutathione

The products of the present invention are good inhibitors of phosphodiesterases and they are capable to restore the glutathione levels compromised by BSO- induced oxidative stress. EXAMPLE 5. Effect on proliferation

The products of the present invention are potent inhibitors of human prostatic cells proliferation by evaluation with the test of bromodeoxyuridine (BRDU) incorporation following the method reported in Carroll PR et al J Urol. 1993 Feb; 149 (2) :403-7. "Cell proliferation in prostatic adenocarcinoma: in vitro measurement by 5- bromodeoxyuridine incorporation and proliferating cell nuclear antigen expression" .

The results for compound 1 are reported in the table where it is possible to see that the product is active at low concentrations

Treatment BRDU (%)

Control 38

Example 1 - 1OnM 26

Example 1 - 10OnM 10

Claims

CLAIMS :

1. Compounds of general formula :

A-Y-W (I) wherein

A is selected from the group comprising the residues of PDE-5 inhibitors having formula (II or III or IV or V) :

( ID

(III )

(IV)

(V) wherein X is a group capable to link to ~Y o ~W, selected from a group comprising -C00~; -0~; -CONH-; -OCO-; -OCOO-; -CO-;

Y is zero; ~ (C_n. ) alkyl-, ~ (C_n. ) alkyl-CO- , -0- (Cn_')alkyl-O-, -00C- (C_n. ) alkyl-COO- ; -0- (C_n. ) alkyl- , -HN- (C_nO alkyl-, -00C- (C_n- ) alkyl- ; - (C_n- ) alkyl-0-C0- (C_n-) alkyl-; - (C_n. ) alkyl-CO-0- (C_n.. ) alkyl- wherein (C_n.) alkyl and (C_n-.) alkyl are straight or branched, and n' and n' ' , the same or different to each other, are 0-10;

W is a polysulfurated group containing 2 or more atoms of sulphur, selected from the group comprising an organic thiosulfonate moiety or a dithiole-thione derivative cyclic or linear or trithiocarbonates and salts thereof.

2. Compounds of general formula (I) according to claim 1 wherein X is a group capable to link to ~Y o ~W, selected from a group comprising - (C_p) alkyl-COO~ ; - (Cp)alkyl-O~; - (C_p) alkyl-CONH-; - (C_p) alkyl-OCO~;

(Cp)alkyl-OCOO-; - (C_p) alkyl -CO-; wherein (C_p) alkyl is straight or branched and p is 1-10.

3. Compounds of general formula (I) according to claim 1 wherein W is an organic thiosulfonate moiety having formula : -S-SO₂-R (VI) wherein -S-SO₂-R is linked to A-Y-; R is a straight or branched alkyl, selected from the group consisting of methyl, ethyl, propyl; alkenyl, alkinyl; alkylaryl, alkenylaryl, alkinylaryl; arylalkyl, arylalkenyl, arylalkinyl; or cycloalkyl, cycloalkenyl , cycloalkinyl ; or aromatic and/or heterocyclic ring, all substituted or unsubstituted.

4. Compounds of general formula (I) according to claim 1 wherein W is a dithiole-thione derivative having formula:

(VII) or (VIII)

wherein

Z is S (sulphur) and at least 1 Z is C=S (thione) , m is 0, 1-10;

T is:

~00C- or

wherein

Rl is -H; -COOH; -NH₂; -OH; -SH;

R2 is hydrogen; -COOH; alkyl , alkenyl, alkynyl ; aryl ; fluoro, chloro, bromo; hydroxyl , alkyloxy, alkenyloxy, aryloxy, acyloxy; amino, alkylamino, arylamino; thio; cyano; nitro; acyl ; amido; and a 5 or 6-membered aromatic or non-aromatic ring containing 0, 1, or 2 heteroatoms selected from nitrogen, oxygen, or sulphur.

5. Compounds of general formula (I) according to claim 1 wherein W is a trithiocarbonate.

6. Salts of compounds of general formula (I) according to claim 1, wherein said salts are pharmaceutical acceptable salts of compounds of formula (I) .

7. Salts according to claim 6, said salts comprising salts of compounds of formula (I) with alkaline metals and alkaline earth metals, non-toxic amines and aminoacids, inorganic acids comprising hydrochloric acid, phosphoric acid or organic acids comprising fumaric acid, citric acid, tartaric acid.

8. Compound of general formula (I) according to claim 1 , selected from the group comprising:

4- (5-thioxo-5H-l,2-dithiol-3-yl)phenyl 2- (4- (4- ethoxy-3- (5-methyl-4 -oxo-7-propyl -1, 4- dihydroimidazo [1,5-Jf] [1,2,4] triazin-2-yl) phenyl sulfonyl) piperazin-1-yl) acetate;

4- (5-thioxo-5H-l, 2 -dithiol-3-yl) phenyl 2- (2- (2- (3-1- methyl-7-oxo-3-propyl-6, 7-dihydro-lH-pyrazole [4,3- d] pyrimidin-5-yl) -4- propoxyphenylsulfonamido) pyrrolidin-1-yl) acetate; ester of (6R, 12aR) -2 , 3 , 6, 7, 12 , 12a-esahydro-2 -hydroxy-

6- (3,4-methylendioxyphenyl) - pyrazino [2' , 1' :6, l]pyrido [3,4-b] indole-1, 4-dione with

4- (3-thioxo-3H-l,2-dithiol-4-yl) benzoic acid; ester of (6R, 12aR) -2 , 3 , 6, 7, 12, 12a~esahydro-2-hydroxy- 6- (3 , 4-methylendioxyphenyl) - pyrazino [2' , 1' : 6, l]pyrido [3,4-b] indole-1, 4-dione with 3-methansulfonyl sulfanyl propionic acid;

9. Compound of general formula (I) according to claim 2 which is 4- (5-thioxo-5H-l, 2-dithiol-3- yl) phenyl (6R, 12aR) -2,3, 6, 7, 12, 12a-esahydro-6- (3,4- methylendioxyphenyl) -pyrazino [2' ,1' :6,l]pyrido

[3,4-b] indole-1, 4-dione-2 -acetate.

10. Pharmaceutical composition comprising at least one compound of general formula (I) according to claims 1-9 as an active ingredient and optionally one or more pharmaceutically acceptable adjuvant (s) or carrier (s) .

11. Compound of general formula (I) according to claims 1-9 for use as a medicament.

12. Use of a compound of general formula (I) according to claims 1-9 for the manufacture of a medicament for treating low urinary tract dysfunctions comprising incontinence, benign prostatic hyperplasia and erectile dysfunction.

13. Use of the ester of sildenafil analogue with 5- (p-hydroxyphenyl) -3H-1, 2-dithiol-3-thione having formula:

for the manufacture of a medicament for the treatment of low urinary tract dysfunctions comprising incontinence, benign prostatic hyperplasia.

14. Use of a compound of general formula (I) according to claims 1-9 for the manufacture of a pharmaceutical composition for treating diseases of low urinary tract in combination with other agents.

15. Process for the synthesis of compounds of general formula (I) according to claims 1-9, comprising at least a reaction between the corresponding precursor of an organic thiosulfonate or dithiol-thione or trithiocarbonate, moiety W or Y- W, and a derivative compound of 5-phosphodiesterases inhibitors or a corresponding precursor of a phosphodiesterases inhibitor modified with Y, wherein W and Y have the meaning according to claim 1.

16. A method of treating low urinary tract dysfunctions comprising incontinence, benign prostatic hyperplasia and erectile dysfunction, which comprises administering to a patient suffering from such diseases a therapeutically effective amount of a compound selected from the group comprising:

4- (5-thioxo-5H-l,2-dithiol-3-yl)phenyl 2- (4- (4- ethoxy-3- (5-methyl-4-oxo-7-propyl-l, 4- dihydroimidazo [1, 5-Jf] [1,2,4] triazin-2-yl) phenyl sulfonyl) piperazin-1-yl) acetate;

4- (5-thioxo-5H-l,2-dithiol-3-yl)phenyl 2- (2- (2- (3 -1-methyl-7-oxo-3 -propyl-6 , 7-dihydro-IH- pyrazole [4 , 3 -d] pyrimidin-5-yl) -4- propoxyphenylsulfonamido) pyrrolidin-1-yl) acetate; ester of (6R, 12aR) -2 , 3 , 6, 7, 12 , 12a-esahydro-2- hydroxy-6- (3 , 4-methylendioxyphenyl) - pyrazino [2 ' , 1' : 6, 1] pyrido [3 , 4-b] indole-1 , 4-dione with 4- (3-thioxo-3H-l,2-dithiol-4-yl) benzoic acid; ester of (6R, 12aR) -2 , 3 , 6, 7, 12 , 12a-esahydro-2- hydroxy-6- (3 , 4-methylendioxyphenyl) - pyrazino [2' , 1' :6, 1] pyrido [3, 4-b] indole-1, 4-dione with 3-methansulfonyl sulfanyl propionic acid; ester of sildenafil analogue with con 5- (p- hydroxyphenyl ) -3H-1, 2-dithiol-3-thione having formula :

4- (5-thioxo-5H-l, 2 -dithiol-3-yl) phenyl (6R,12aR) - 2,3, 6, 7, 12, 12a-esahydro-6- (3 , 4-methylendioxyphenyl) - pyrazino [2' , 1' :6, l]pyrido

[3,4-b] indole-1, 4-dione-2-acetate or a pharmaceutically-acceptable salt/s thereof.