`
`WORLD INTELLECTUAL PROPERTY ORGANIZATION
`International Bureau
`
`
`
`INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT)
`
`(51) International Patent Classification 6;
`
`A61K 31/495
`
`(11) International Publication Number:
`
`WO 97/03675
`
`(43) International Publication Date:
`
`6 February 1997 (06.02.97)
`
`(21) International Application Number:
`
`PCT/EP96/03024
`
`(22) International Filing Date:
`
`11 July 1996 (11.07.96)
`
`(30) Priority Data:
`9514464.8
`
`14 July 1995 (14.07.95)
`
`GB
`
`(71) Applicant (for all designated States except US): LABORA-
`TORRE GLAXO WELLCOME S.A.
`[FR/FR]; 43, me
`Vineuse, F-75116 Paris (FR).
`
`(72) Inventor; and
`(75) Inventor/Applicant (for US only): DAUGAN,Alain, Claude-
`Marie [FR/FR]; Laboratoire Glaxo Wellcome S.A., Centre
`de Recherches, Z.A. de Courtaboeuf, 25, avenue de Quebec,
`F-91940 Les Ulis (FR).
`
`(74) Agents: FILLER, Wendy, Anneetal.; Glaxo Wellcome House,
`Berkeley Avenue, Greenford, Middlesex UB6 ONN (GB).
`
`GB,
`LS, LT, LU, LV,
`
`MD, MG, MK, MN,
`SD, SE, SG,SI, SK,
`SZ, UG), Eurasian
`VN, ARIPO patent (KE, LS, MW, SD,
`patent (AM, AZ, BY, KG, KZ, MD, RU, TJ, TM), European
`patent (AT, BE, CH, DE.
`, ES, FI, FR, GB, GR,IE,IT,
`LU, MC, NL, PT, SE),
`I patent (BF, BJ, CF, CG, CI,
`CM, GA, GN, ML, MR, NE, SN, TD, TG).
`
`Published
`With international search report.
`
`(54) Title; USE OF CGMP-PHOSPHODIESTERASE INHIBITORS TO TREAT IMPOTENCE
`
`(57) Abstract
`
`
`
`The
`
`(J)
`formula
`of
`compounds
`of
`use
`12aR)-2,3,6,7,12,12a-hexahydro-2-methyl-
`(6R,
`6-(3,4-methylenedioxyphenyl)-pyrazino[2’,1’:
`6,1 ]pyrido[3,4-b]indole-i ,4-dione,
`(3S,
`12aR)-2,3,6,7, 12, 12a-hexahydro-2,3-dimethyl-
`6-(3,4-methylenedioxyphenyl)-pyrazino[2’,1°:
`6,1 ]pytido[3,4-bJindole-1,4-dione, and physiologically
`acceptable salts and solvates thereof,
`in the treatment
`of impotence.
`
`6R,
`
`ATI 1005-0001
`
`ATI v. ICOS
`
`IPR2018-01183
`
`ATI 1005-0001
`
`ATI v. ICOS
`IPR2018-01183
`
`
`
`FOR THE PURPOSES OF INFORMATION ONLY
`
`Codes used to identify States party to the PCT on the front pages of pamphlets publishing international
`applications under the PCT.
`
`Viet Nam
`
`United Kingdom
`Georgia
`Guinea
`Greece
`Hungary
`Treland
`Italy
`Japan
`Kenya
`Kyrgystan
`Democratic People’s Republic
`of Korea
`Republic of Korea
`Kazakhstan
`Liechtenstein
`Sri Lanka
`Liberia
`Lithuania
`Luxembourg
`Latvia
`Monaco
`Republic of Moldova
`Madagascar
`Mali
`Mongolia
`Mauritania
`
`AM
`AT
`AU
`BB
`BE
`BF
`BG
`BJ
`BR
`BY
`CA
`CF
`CG
`CH
`cl
`CM
`CN
`cs
`CZ
`DE
`DK
`EE
`ES
`FI
`FR
`GA
`
`Armenia
`Austria
`Australia
`Barbados
`Belgium
`Burkina Faso
`Bulgaria
`Benin
`Brazil
`Belarus
`Canada
`Central African Republic
`Congo
`Switzerland
`Cote d'Ivoire
`Cameroon
`China
`Czechoslovakia
`Czech Republic
`Germany
`Denmark
`Estonia
`Spain
`Finland
`France
`Gabon
`
`ATI 1005-0002
`
`Malawi
`Mexico
`Niger
`Netherlands
`Norway
`New Zealand
`Poland
`Portugal
`Romania
`Russian Federation
`Sudan
`Sweden
`Singapore
`Slovenia
`Slovakia
`Senegal
`Swaziland
`Chad
`Togo
`Tajikistan
`Trinidad and Tobago
`Ukraine
`Uganda
`United States of America
`Uzbekistan
`
`ATI 1005-0002
`
`
`
`WO 97/03675
`
`PCT/EP96/03024
`
`USE OF CGMP-PHOSPHODIESTERASE INHIBITORS TO TREAT IMPOTENCE
`
`This invention relates to the use of tetracyclic derivatives which are potent
`and selective inhibitors of cyclic guanosine 3',5'-monophosphate specific
`phosphodiesterase (cGMPspecific PDE)in the treatment of impotence.
`
`Impotence can be defined as a lack of power, in the male, to copulate and
`mayinvolve aninability to achieve penile erection or ejaculation, or both. More
`specifically, erectile impotence or dysfunction may be defined as an inability to
`obtain or sustain an erection adequate for intercourse.
`Its prevalence is claimed
`to be between 2 and 7% of the human male population, increasing with age, up
`to 50 years, and between 18 and 75% between 55 and 80 years of age.
`
`Reports of well-controlled clinical trials in man are few and the efficacy of
`orally administered drugs is low. Although many different drugs have been
`shownto induce penile erection, they are only effective after direct injection into
`the penis, e.g. intraurethrally or intracavernosally(i.c.), and are not approvedfor
`erectile dysfunction. Current medical treatment is based on thei.c. injection of
`vasoactive
`substances
`and
`good
`results
`have
`been
`claimed with
`phenoxybenzamine, phentolamine, papaverine and prostaglandin E,, either
`alone or in combination; however, pain, priapism and fibrosis of the penis are
`associated with the ic. administration of some of these agents. Potassium
`channel openers (KCO) and vasoactive intestinal polypeptide (VIP) have also
`been shown to be active i.c., but cost and stability issues could limit
`developmentof the latter. An alternative to the i.c. route is the use of glyceryl
`trinitrate (GTN) patches applied to the penis, which has been shown to be
`effective but producesside-effects in both patient and partner.
`
`As a general alternative to pharmacological intervention, a variety of penile
`prostheses has been used to assist achievement of an erection. The short term
`success rate is good, but problems with infection and ischaemia, especially in
`diabetic men, make this type of treatment a final option rather than first-line
`therapy.
`The compoundsofthe invention are potentinhibitors of cyclic guanosine 3',5'-
`monophosphate phosphodiesterases (CGMP PDEs). GB 9514464.8, whichis
`the priority document for the present application describes the syntheses of the
`compoundsofthe invention andtheirutility in impotence. WO95/19978. which
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`was unpublished at the priority date of the present application, also describes
`the syntheses of the compoundsofthe invention and theirutility in other
`diseases associated with inhibition of CGMP PDEs. The compounds may be
`represented by the following general formula(I):
`
`(i)
`
`and salts and solvates (e.g. hydrates) thereof, in which:
`’ RO represents hydrogen, halogen or C4.6 alkyl;
`R1 represents hydrogen, C4.galkyl, C2, alkenyl, C2, alkynyl, haloC4galkyl,
`C3-gcycloalkyl, C3_gcycloalkylC 4_3alkyl, aryiC4_3alkyl or heteroarylC4-_3alkyl;
`R2 represents an optionally substituted monocyclic aromatic ring selected
`from benzene, thiophene, furan and pyridine or an optionally substituted bicyclic
`
`OD
`
`attached to the rest of the molecule via one of the benzene
`ring
`ring carbon atoms and wherein the fused ring A is a 5- or 6-memberedring
`which may be saturatedorpartially orfully unsaturated and comprises carbon
`atomsand optionally one or two heteroatoms selected from oxygen, sulphur and
`nitrogen; and
`R® represents hydrogenor C,. alkyl, or R' and R® together represent a 3- or
`4- membered alkyl or alkenyl chain.
`
`Suitable individual compoundsofthe invention for use in the treatment of
`erectile dysfunction include:
`Cis-2,3,6,7,12,12a-hexahydro-2-(4-pyridylmethy!)-6-(3,4-methylenedioxyphenyl)-
`pyrazinof[2', 1'
`: 6,1]pyrido[3,4-blindole-1 ,4-dione;
`Cis-2,3,6,7,12, 12a-hexahydro-6-(2,3-dihydrobenzo[b]furan-5-yl)-2-methyl-
`pyrazino[2',1':6, 1]pyrido[3,4-b]indole -1,4-dione;
`Cis-2,3,6,7,12,12a-hexahydro-6-(5-bromo-2-thienyl)-2-methyl-
`pyrazino[2'1':6, 1]pyrido[3,4-bJindole -1,4-dione;
`
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`Cis-2,3,6,7,12,12a-hexahydro-2-butyl-6-(4-methylphenyl)-
`pyrazino[2‘, 1':6, 1 ]pyrido[3,4-blindole -1,4-dione;
`(6R, 12aR)-2,3,6,7,12, 12a-Hexahydro-2-isopropyl-6-(3,4-methylenedioxyphenyl)-
`pyrazino[2',1':6,1]pyrido[3,4-b]indole -1,4-dione;
`(6R, 12aR)-2,3,6,7,12,12a-Hexahydro-2-cyclopentyl-6-(3,4-
`methylenedioxyphenyl)-pyrazino[2’, 1:6, 1]pyrido[3,4-b]indole -1,4-dione;
`(6R, 12aR)-2,3,6,7,12,12a-Hexahydro-2-cyclopropylmethyl-6-(4-methoxyphenyl)-
`pyrazino[2’1':6, 1]pyrido[3,4-b]indole -1,4-dione;
`(6R, 12aR)-2,3,6,7,12, 12a-Hexahydro-6-(3-chloro-4-methoxypheny!)-2-methyl-
`pyrazino[2', 1':6, 1]pyrido[3,4-blindole -1,4-dione;
`(6R,12aR)-2,3,6,7, 12, 12a-Hexahydro-2-methyl-6-(3,4-methylenedioxyphenyl)-
`pyrazino[2',1':6, 1]pyrido[3,4-bjindole-1 4-dione:
`(6R, 12aR)-2,3,6,7,12, 12a-Hexahydro-6-(3,4-methylenedioxyphenyl)-
`pyrazino[2', 1': 6,1] pyrido [3,4-b] indole-1,4-dione;
`(5aR, 12R, 14aS)-1,2,3,5,6,11,12,14a-Octahydro-12-(3,4-
`methylenedioxyphenyl)-pyrrolo[1",2" : 4'5"Jpyrazino[2’,1'
`: 6, 1]pyrido[3,4-
`bjindole-5-1 ,4-dione;
`Cis-2,3,6,7,12,12a-hexahydro-2-cyclopropyl-6-(3,4-methylenedioxypheny!)-
`pyrazino[2'1:6, 1]pyrido[3,4-bjindole -1,4-dione;
`(3S, 6R,12aR)-2,3,6,7,12,12a-hexahydro-3-methyl-6-(3,4-
`methylenedioxyphenyl)-pyrazino[2’, 1':6, 1]pyrido[3,4-bJindole -1,4-dione;
`
`and physiologically acceptable salts and solvates (e.g. hydrates) thereof.
`
`The specific compoundsof the invention are:
`
`(6R, 12aR)-2,3,6,7,12,12a-hexahydro-2-methyl-6-(3,4-methylenedioxyphenyl)-
`pyrazino[2',1':6, 1]pyrido[3,4-bJindole -1,4-dione (Compound A); and
`(3S, 6R, 12aR)-2,3,6,7,12, 12a-hexahydro-2,3-dimethy!-6-(3,4-
`methylenedioxypheny/)-pyrazino[2',1'
`: 6, 1]pyrido[3,4-bJindole-1 ,4-dione
`(Compound B);
`
`and physiologically acceptable saits and solvates (e.g. hydrates) thereof.
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`Unexpectedly, it has now been found that compoundsof formula (I), and in
`particular compounds A and B, are useful
`in the treatment of erectile
`dysfunction. Furthermore the compounds may be administered orally, thereby
`
`ATI 1005-0005
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`4
`
`Thus the
`obviating the disadvantages associated with i.c. administration.
`present
`invention concerns the use of compounds of formula (1), and in
`particular compounds A andB,or a pharmaceutically acceptable sait thereof, or
`a pharmaceutical composition containing either entity, for the manufacture of a
`medicament for the curative or prophylactic treatment of erectile dysfunction in a
`male animal, including man.
`
`The pharmaceutically acceptable salts of the compoundsof formula(I), and in
`particular compounds A and B which contain a basic centre are acid addition
`salts formed with pharmaceutically acceptable acids. Examples include the
`hydrochloride, hydrobromide, sulphate or bisulphate, phosphate or hydrogen
`phosphate, acetate, benzoate, succinate, fumarate, maleate,
`lactate, citrate,
`tartrate,
`gluconate,
`methanesulphonaie,
`benzenesulphonate
`and
`p-toluenesulphonate salts. Compounds of
`formula
`(!),
`and in particular
`compounds A and B canalso provide pharmaceutically acceptable metalsalts,
`in particular alkali metal salts, with bases. Examples include the sodium and
`potassium salts.
`
`it has been shown that compounds of the present invention are potent and
`selective inhibitors of CGMP specific PDE.
`It has now been surprisingly found
`that human corpus cavernosum contains three distinct PDE enzymes. The
`predominant PDEhasfurther surprisingly been found to be cGMP PDE. Asa
`consequence of the selective PDE V inhibition exhibited by compounds of the
`present invention, the subject compounds can elevate cGMP levels, which in
`turn can mediate relaxation of the corpus cavernosum tissue and consequent
`penile erection.
`
`Although the compounds of the invention are envisaged primarily for the
`treatment of erectile dysfunction or male sexual dysfunction, they may also be
`useful
`for
`the treatment of female sexual dysfunction including orgasmic
`dysfunction relatedto clitoral disturbances.
`
`in man, oral administration of the compoundsof the invention is
`Generally,
`the preferred route, being the most convenient and avoiding the disadvantages
`associated with i.c. administration.
`In circumstances where the recipient suffers
`from a swallowing disorder or from impairment of drug absorption after oral
`administration, the drug may be administered parenterally, e.g. sublingually or
`buccally.
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`For administration to man in the curative or prophylactic treatment of the
`disorders identified above, oral dosages of a compound of formula (I), and in
`particular compounds A and B will generally be in the range of from 0.5-800mg
`daily for an average adult patient (70kg). Thus for a typical adult patient,
`individual tablets or capsules contain from 0.2-400mgof active compound,ina
`suitable pharmaceutically acceptable vehicle or carrier, for administration in
`single or multiple doses, once or several times per day. Dosages for buccal or
`sublingual administration will typically be within the range of from 0.1-400 mg
`per single dose as required.
`In practice the physician will determine the actual
`dosing regimen which will be most suitable for an individual patient and it will
`vary with the age, weight and response of the particular patient. The above
`dosages are exemplary of the average case but
`there can be individual
`instances in which higher or lower dosage ranges may be merited, and such are
`within the scope of this invention.
`
`For human use, compoundsofformula (1), and in particular compounds A and
`B can be administered alone, but will generally be administered in admixture
`with a pharmaceutical carrier selected with regard to the intended route of
`administration and standard pharmaceutical
`practice.
`For example,
`the
`compound may be administered orally, buccally or sublingually,
`in the form of
`tablets containing excipients such as starch or lactose, or in capsules or ovules
`either alone or
`in admixture with excipients, or in the form of elixirs or
`suspensions containing flavouring or colouring agents. Suchliquid preparations
`may be prepared with pharmaceutically acceptable additives
`such as
`Suspending agents (e.g. methylcellulose, a semi-synthetic glyceride such as
`witepsol or mixtures of glycerides such as a mixture of apricot kernel oil and
`PEG-6 esters or mixtures of PEG-8 and caprylic/capric glycerides).
`
`For veterinary use, a compoundof formula(I), and in particular compound A
`or B or a non-toxic salt thereof is administered as a suitably acceptable
`formulation in accordance with normal veterinary practice and the veterinary
`surgeonwill determine the dosing regimen and route of administration which will
`be most appropriate for a particular male animal.
`
`Thus the invention includes a pharmaceutical composition for the curative or
`prophylactic treatment of erectile dysfunction in a male animal, including man,
`comprising a compound of formula (1), and in particular compound A or B, ora
`pharmaceutically acceptable salt
`thereof,
`together with a pharmaceutically
`acceptable diluent or carrier.
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`6
`
`There is further provided a process for the preparation of a pharmaceutical
`composition for the curative or prophylactic treatment of erectile dysfunction in a
`male animal, including man, comprising formulating a compound of formula (I),
`and in particular compound A or B, or a pharmaceutically acceptable salt
`thereof, with a pharmaceutically acceptable diluent or carrier.
`
`including
`The invention also provides a method of treating a male animal,
`man,to cure or preventerectile dysfunction which comprisestreating said male
`animalwith an effective amount of a compound of formula (I), and in particular
`compound A or B, or a pharmaceutically acceptable salt
`thereof, or a
`pharmaceutical composition containing eitherentity.
`
`Moreover, the invention includes the use of a compoundof formula (I), and in
`particular compound A or B, or a pharmaceutically acceptable salt thereof, or a
`pharmaceutical composition containing either entity, for the manufacture of a
`medicament for the curative or prophylactic treatment of erectile dysfunction in a
`male animal, including man.
`
`A compound of formula(I), and in particular compound A or B, may also be
`used in combination with other therapeutic agents which may be useful in the
`treatment of erectile dysfunction substantially as hereinbefore described. The
`invention thus provides,
`in another aspect, a combination of a compound of
`formula (1), and in particular compound A or B together with another
`therapeutically active agent.
`
`The combination referred to above may conveniently be presented for use in
`the form of a pharmaceutical formulation and thus pharmaceutical compositions
`comprising a combination as defined above together with a pharmaceutically
`acceptablediluent or carrier comprise a further aspectof the invention.
`
`Theindividual components of such a combination may also be administered
`either sequentially or simultaneously in separate pharmaceutical formulations.
`
`Appropriate doses of known therapeutic agents for use in combination with a
`compoundofthe invention will be readily appreciated by those skilled in the art.
`
`The compoundsof the invention may be prepared by any suitable method
`known in the art or by the following process which forms part of the present
`invention. The process has been previously substantially described in the
`priority documentof the present invention GB9514464.8, and in WO95/1 9978.
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`7
`
`Thus, a process for preparing a compound offormula(I) comprises treating a
`compoundof formula(Il)
`
`R°
`
`|
`
`i
`
`(Il)
`
`ho
`
`A
`
`Oo
`
`is a halogen
`(in which Alk represents C4_galkyl, e.g. methyl or ethyl and Hal
`atom, e.g. chlorine) with a primary amine RINH9 in a suitable solvent such as
`an alcohol (e.g. methanol or ethanol) or a mixture of solvents, conveniently at a
`temperature of from 20°C to reflux (e.g. at about 50°C).
`
`A compound of formula (Il) may conveniently be prepared by treating a
`compound of formula (III) with a compoundof formula (IV)
`
`ne
`
`O
`
`OAIk
`
`NH
`
`pz
`
`ill)
`
`3
`
`N
`
`H
`
`0
`
`sal
`a
`
`HY
`
`(vy)
`
`R
`
`in a suitable solvent such as a halogenated hydrocarbon (e.g.trichioromethane
`or dichloromethane), or an ether (e.g.
`tetrahydrofuran), preferably in the
`presence of a base such as an organic amine (e.g. a trialkylamine such as
`triethylamine) or an alkali metal carbonate or bicarbonate (e.g. NaHCO3). The
`reaction may conveniently be effected at a temperature of from -20°C to +20°C
`(e.g. at about O°C).
`
`A compoundof formula (|) may also be prepared from a compound of formula
`(lll) in a two-step procedure via a compoundof formula (I!) isolated without
`purification.
`
`Compounds of formula (i) may be prepared asindividual enantiomers in two
`steps from the appropriate enantiomer of formula(Ill) or as mixtures (e.g.
`racemates) of either pairs of cis or trans isomers from the correspondong
`mixtures of either pairs of cis or trans isomers of formula(Ill).
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`Individual enantiomers of the compounds of the invention may be prepared
`from racemates by resolution using methods knownin the art for the separation
`of racemic mixturesinto their constituent enantiomers, for example using HPLC
`(high performanceliquid chromatography) on a chiral column such as Hypersil
`naphthylurea.
`
`A compoundofformula(Ill) may conveniently be prepared from a tryptophan
`alkyl ester of formula (V)
`
`R°
`
`O
`
`NH,
`
`OAIk
`
`(V)
`
`|
`
`N
`H
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`(where Alk is as previously defined) or a salt thereof (e.g. the hydrochloride salt)
`with an aldehyde R2CHO. The reaction may conveniently be effected in a
`Suitable solvent such as a halogenated hydrocarbon (e.g. dichloromethane) or
`an aromatic hydrocarbon (e.g. toluene) in the presence of an acid such as
`trifluoroacetic acid.
`The reaction may conveniently be carried out at a
`temperature of from -20°Cto reflux to provide a compoundof formula (III) in one
`step. The reaction may also be carried out in a solvent such as an aromatic
`hydrocarbon (e.g. benzene or toluene) under reflux, optionally using a Dean-
`Stark apparatus to trap the water produced.
`
`The reaction provides a mixture of cis and trans isomers which may be either
`individual enantiomers or racemates of pairs of cis or trans isomers depending
`upon whether racemic or enantiomerically pure tryptophan alkyl ester was used
`as the starting material.
`Individualcis or trans enantiomers may conveniently be
`separated
`from mixtures
`thereof
`by
`fractional
`crystallisation
`or
`by
`chromatography(e.g. flash column chromatography) using appropriate solvents
`and eluents.
`Similarly, pairs of cis and-trans isomers may be separated by
`chromatography (e.g. flash column chromatography) using appropriate eluents.
`An optically pure trans isomer may also be converted to an optically pure cis
`isomerusing suitable epimerisation procedures. One such procedure comprises
`treating the trans isomeror a mixture (e.g. 1: 1 mixture) of cis and trans isomers
`with methanolic or aqueous hydrogen chloride at a temperature of from 0°C to
`the refluxing temperature of the solution. The mixture may then be subjected to
`chromatography (e.g. flash column chromatography) to separate the resulting
`diastereoisomers, or in the procedure utilising aqueous hydrogen chloride the
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`desired cis isomerprecipitates out as the hydrochloride salt which may then be
`isolated byfiltration.
`
`The pharmaceutically acceptable acid addition salts of a compound of
`formula(|), and in particular compound A or B which contain a basic centre may
`be prepared in a conventional manner. For example, a solution of the free base
`may be treated with a suitable acid, either neat or in a suitable solution, and the
`resulting salt isolated either by filtration or by evaporation under vacuum of the
`reaction solvent.
`Pharmaceutically acceptable base addition salts may be
`obtained in an analogous mannerby treating a solution of compound A or B
`with a suitable base. Both types of salt may be formed orinterconverted using
`ion-exchange resin techniques.
`
`Compounds of the invention may be isolated in association with solvent
`moleculesby crystallisation from or evaporation of an appropriate solvent.
`
`The syntheses of compounds A and B and of the intermediates for use
`therein are illustrated by the following examples. The examples have been
`previously described in
`the priority document of
`the instant
`invention
`GB9514464.8, and the corresponding Intermediate or Example numbers therein
`are shownin parentheses‘next to the current Intermediate or Example number.
`
`In the Examples section hereinafter the following abbreviations are used:
`
`10
`
`15
`
`20
`
`MeOH (methanol) and EtOH (ethanol),
`
`
`bjindole-3-carboxy!
`is
`iso
`
`25
`
`30
`
`To a stirred solution of D-tryptophan methyl ester (11 g) and piperonal (7.9 g) in
`anhydrous CH2Clo (400 mL) cooled at 0°C was added dropwise trifluoroacetic
`acid (7.7 mL) and the solution was allowed to react at ambient temperature.
`After 4 days, the yellow solution was diluted with CHaClo (200 mL) and washed
`with a saturated aqueous solution of NaHCO3, then with water (3x200 mL) and
`dried over NagSQq. The organic layer was evaporated under reduced pressure
`and the residue containing the two geometric isomers was purified by flash
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`ATI 1005-0011
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`
`chromatographyeluting with dichioromethane/ethyl acetate (97/3) to give as the
`firsr eluting productthetitlecompound (6.5 g)
`
`m.p. : 154°C
`
`Intermediate2(83)
`
`1
`
`R)-M
`
`11
`
`4-tetrahydro-2-(2-chloropropionyl)-1-(3.4-
`
`methylenedi
`
`nvi)-9H-pvri
`
`4-bJindole-3-carboxyla
`
`To a solution of (R)-(+)-2-chloropropionic acid (191 pl, 2.2 mmol) in anhydrous
`dichloromethane (30 mL), was added dicyclohexylcarbodiimide
`(0.45 g,
`2.2. mol). intermediate 1 (0,7 g, 2 mmol) was then added and the mixture was
`stirred at
`room temperature for 20 hours. The formed precipitate of
`dicyclohexylurea was removedbyfiltration, the filtrate was evaporated in vacuo
`and the crude product was purified by flash chromatography eluting with
`toluene/ethyl acetate : 95/5. The oily compound obtained was then crystallised
`from ether/hexane to givethetitlecompound as paleyellow crystals (0.74 g)
`
`10
`
`15
`
`m.p. : 126-128°C.
`
`ei?
`
`(6R,12aR)-2,3,6.7.12,12a-Hexahydro-2-methyl-6-(3.4-methylenedioxyphenyl)-
`ino[2'.1':6, Tpyri
`-bJindole
`-1,4-di
`
`20
`
`25
`
`30
`
`in
`(0.5 g) and NaHCO3 (0.14 g)
`a) To a stirred solution of intermediate 1
`anhydrous CHCl3 (20 mL) was added dropwise chloroacetyl chloride (0.27 mL)
`at 0°C. The resulting mixture was stirred for 1 hour at the same temperature
`and diluted with CHCI3 (20 mL). Water (10 mL) was then added dropwise with
`Stirring to the mixture, followed by a saturated solution of NaHCO3. The organic
`layer was washed with water until neutrality and dried over NagSO,. After
`evaporation of the solvent under reduced pressure, (6R.12aR)-methyl 1.2,3.4-
`ro-2-chior.
`|-1-(3,4-methyl
`i
`I)-9H-pyrido[3.4-blindole-
`3-carboxylate was obtained as an oil which wascrystallised from etherto give a
`solid (0.38 g, m.p.
`: 233°C) which was used without further purification in the
`next step.
`
`the chloroacetyl intermediate (0.37 g) in MeOH
`b) To a stirred suspension of
`(20 mL) was added at room temperature a solution of methylamine (33% in
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`EtOH) (0.4 mL) and the resulting mixture was heated at 50°C under No for
`16 hours. The solvent was removed under reduced pressure and the residue
`was dissolved in CH2Cl9 (50 mL). After washing with water (3x20 mL), drying
`over NagSO, and evaporating to dryness, the residue waspurified by flash
`chromatographyeluting with CHoClo/MeOH (99/1) and recrystallised from 2-
`propanolto give thetitlecompound as white crystals (0.22 g)
`
`m.p. : 302-303°C.
`
`Analysis for Co9H19N30a:
`
`Caiculated:C,67.86;H,4.92:N, 10.79;
`
`10
`
`Found:C,67.77;H,4.92:N,10.74%.
`
`[o]20°, = +71.0° (C=1.00; CHC!3):
`
`15
`
`20
`
`
`
`To a stirred solution of intermediate 2 (0.3 g, 0.68 mmol) in THF (30 mL) was
`added at room temperature a solution of methylamine (33 % in EtOH) (0.68 mL)
`and the resulting solution was treated at reflux under No for 6 days. The solvent
`was removed underreduced pressure and the residue was dissolved in CHCl.
`(50 mL). After washing with water
`(2,25 mL), drying over NajSO, and
`evaporating to dryness, the crude product waspurified by flash chromatography
`eluting with dichioromethane/methanol : 99/1. The oily residue obtained was
`crystallised from methanol to give the title compound as white crystals (40 mg)
`m.p. : 307-309°C.
`
`25
`
`Analysis for Co3Ho4N3Q0z,:
`
`Calculated :
`
`C, 68.47 ; H, 5.25; N, 10.42 ;
`
`Found:
`
`C, 68.35; H, 5.33; N, 10.42%.
`
`[o1]20°, = + 65.2° (c = 1.15 ; CHCl).
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`
`The following compound wassimilarly prepared:
`
`Example3
`
`(3S, 6R,12aR)-2,3.6.7.12,12a-Hexahydro-3-methyl-6-(3.4-
`methylenedioxyphenyl)-pyrazino[2'.1':6, 1ipyrido[3.4-blindole -1,4-dione as white
`crystals using ammonia as the base.
`
`m.p. : 319-321°C.
`
`Analysis for Co9H49N30,:
`
`Calculated : C, 67.86 ; H, 4.92; N, 10.79;
`
`Found:
`
`C, 67.86; H, 5.17; N, 10.72%.
`
`10
`
`[a]20°, = + 107° (c = 1; pyridine).
`
`CompoundsA and B have beenincluded in pharmacy formulations and details
`of such formulations are given below.
`
`15
`
`TABLETS FOR ORAL ADMINISTRATION
`
`A.
`
`Dir
`
`mpressi
`
`mg/tablet
`
`
`50.0
`Active ingredient
`Crospovidone USNF
`Magnesium Stearate Ph Eur
`
` Anhydrous Lactose
`
`The active ingredient was sieved and blended with the excipients. The
`resultant mix was compressedinto tablets.
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`ATI 1005-0014
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`Active ingredient
`
`
`
`
`
`Magnesium Stearate Ph Eur
`
`
`Colloidal Silicon Dioxide
`
`Crospovidone
`
`Sodium Lauryl Sulphate
`
`Microcrystalline Cellulose USNF
`
`The active ingredient was sieved and blended with the excipients. The
`resultant mix was compressedinto tablets.
`
`B.
`
`WET GRANULATION
`
`Active ingredient
`
`Polyvinyl pyrollidone
`
`Polyethylene glycol
`
`Polysorbate 80
`
`Microcrystalline Cellulose USNF
`
`Magnesium Stearate Ph Eur
`
`Croscarmellose Sodium
`
`Colloidal Silicon Dioxide
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`ATI 1005-0015
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`
`The polyvinyl pyrollidone, polyethylene glycol and polysorbate 80 were
`dissolved in water. The resultant solution was used to granulate the
`active ingredient. After drying the granules were screened, then extruded
`at elevated temperatures and pressures. The extrudate was milled
`and/or screened then was blended with the microcrystalline cellulose,
`croscarmellose sodium,colloidal silicon dioxide and magnesium stearate.
`The resultant mix was compressedinto tablets.
`
`mg/tablet
`
`Magnesium Stearate BP
`
`Active ingredient
`
`Polysorbate 80
`
`Lactose Ph Eur
`
`Starch BP
`
`Pregelatinised Maize Starch BP
`
`The active ingredient was sieved and blended with the lactose, starch
`and pregelatinised maize starch. The polysorbate 80 was dissolved in
`purified water. Suitable volumes of the polysorbate 80 solution were
`added and the powders were granulated. After drying, the granules were
`screened and blended with the magnesium stearate. The granules were
`then compressedinto tablets.
`
`Tablets of other strengths may be prepared byaltering the ratio of active
`ingredient to the other excipients.
`
`FILM COATED TABLETS
`
`The aforementioned tablet formulations werefilm coated.
`
`5
`
`10
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`ATI 1005-0016
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`1
`
`to 100.0*
`
`Opadry whiteT
`
`Purified water Ph Eur
`
`13.2
`
`* The water did not appearin the final product. The maximum theoretical weight
`of solids applied during coating was 20mg/tablet.
`
`t Opadry white is a proprietary material obtainable from Colorcon Limited, UK
`which contains hydroxypropyl methylcellulose,titanium dioxide andtriacetin.
`
`5
`
`The tablets were film coated using the coating suspension in conventionalfilm
`coating equipment.
`
`CAPSULES
`
`10
`
`Active ingredient
`
`Lactose
`
`Polyvinyl pyrollidone
`
`Magnesium Stearate
`Sodium Lauryl Sulphate
`
`The active ingredient was sieved and blended with the excipients. The mix was
`filled into size No. 1 hard gelatin capsules using suitable equipment.
`
`Active ingredient
`
`Microcrystalline Cellulose
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`ATI 1005-0017
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`
`Crospovidone
`
`Magnesium Stearate
`
`The active ingredient was sieved and blended with the excipients. The mix was
`filled into size No. 1 hard gelatin capsules using suitable equipment.
`
`Other doses may be prepared by altering the ratio of active ingredient to
`excipient, the fill weight and if necessary changing the capsule size.
`
`
`
` Active ingredient
`
` Labrafil M1944CS
`to 1.0 ml
`
`mg/capsule
`
`50.0
`
`The active ingredient was sieved and blended with the Labrafil. The suspension
`wasfilled into soft gelatin capsules using appropriate equipment.
`
`10
`
`15
`
`20
`
`Inhibitory effect on cGMP-PDE
`cGMP-PDEactivity of compoundsof the present invention was measured using
`a one-step assay adapted from Wells at al. (Wells, J. N., Baird, C. E., Wu, Y. J.
`and Hardman, J. G., Biochim. Biophys. Acta 384, 430 (1975)). The reaction
`medium contained 50mM Tris-HCl,pH 7.5, 5mM Mg-acetate, 250ug/m! 5’-
`Nucleotidase, 1mM EGTA and 0.15uM 8-[H*]-cGMP. The enzyme used was a
`human recombinant PDE V (ICOS, Seattle USA).
`
`Compoundsof the invention were dissolved in DMSOfinally present at 2% in
`the assay. The incubation time was 30 minutes during which the total substrate
`conversion did not exceed 30%.
`
`examined were determined from
`the compounds
`for
`ICs) values
`The
`concentration-response curves using typically concentrations ranging from 10nM
`to 10M. Tests against other PDE enzymes using standard methodology also
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`
`showed that compounds of the invention are highly selective for the cGMP
`specific PDE enzyme.
`
`-cGMP level measurements
`Rat aortic smooth muscle cells (RSMC) prepared according to Chamley et al. in
`Cell Tissue Res. 177, 503 - 522 (1977) were used between the 10th and 25th
`passage at confluence in 24-well culture dishes. Culture media was aspirated
`and replaced with PBS (0.5m!) containing the compound tested at
`the
`appropriate concentration. After 30 minutes at 37°C, particulates guanylate
`cyclase was stimulated by addition of ANF (100nM) for 10 minutes. At the end
`of incubation, the medium was withdrawn and two extractions were performed
`by addition of 65% ethanol (0.25ml). The two ethanolic extracts were pooled
`and evaporated until dryness, using a Speed-vac system.
`c-GMP was
`
`
`
`
`measured by—scintillationafter acetylation proximity immunoassay
`
`15
`(AMERSHAM).
`
`10
`
`The compounds according to the present invention were typically found to
`exhibit an IC,, value of less than 500nM, and an EC,value of less than 5.
`In
`vitro test data for representative compoundsof the invention is given in following
`Table 1:
`
`Table 1
`
`Example No.
`
`1
`
`
`
`
`
`0.2
`
`
`
`
`
`The above data demonstrates the ability of the subject compounds of the
`invention to inhibit cGMP PDE, and hencetheirutility in the treatment of erectile
`dysfunction substantially as hereinbefore described.
`
`20
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`1. Use of a compoundof formula(I):
`
`CLAIMS
`
`(!)
`
`and salts and solvates (e.g. hydrates) thereof, in which:
`R®° represents hydrogen, halogen or C4_6 alkyl;
`R1 represents hydroge