WO2008092862A1

WO2008092862A1 - Bicyclic derivatives as ep4 agonists

Info

Publication number: WO2008092862A1
Application number: PCT/EP2008/051045
Authority: WO
Inventors: Joannes Theodorus Maria Linders; Guy Rosalia Eugeen Van Lommen; Louis Jozef Elisabeth Van Der Veken; Robert Jozef Maria Hendrickx; Jean Pierre Frans Van Wauwe; Jeroen Marcel Maria Roger Aerssens; Mansoor D'lavari; Sorin-Vasile Filip; Kenneth Andrew Lowerson; Petr Vladimirivich Davidenko; Mikhail Viktorovich Reynov
Original assignee: Janssen Pharmaceutica N.V.
Priority date: 2007-01-30
Filing date: 2008-01-29
Publication date: 2008-08-07

Abstract

The present invention relates to a compound of formula (I) including any stereochemical isomeric form thereof, wherein A represents a 5 or 6-membered aromatic optionally substituted heterocycle containing 1, 2 or 3 heteroatoms; ring E represents a partially saturated or aromatic 5-membered heterocycle wherein the dotted lines represent an optional double bond and wherein B, C and D each independently represent CH2, CH, N, NH, S or O and F represents N or C, provided that the 5-membered ring contains 1, 2 or 3 heteroatoms; X represents a direct bond or C1-4alkanediyl; Y represents N or CH; R1 represents hydrogen or fluoro; R2 represents hydrogen, halo, cyano, C1-6alkyl, C1-6alkyloxy, C1-6alkylcarbonyl or C1-6alkylcarbonylamino; R3 represents hydrogen, halo, C1-6alkyl, C1-6alkyloxy, cyano, nitro, amino or mono-or di(C1-6alkyl)amino; n represents an integer of value 1, 2 or 3; a N-oxide thereof, a pharmaceutically acceptable salt thereof or a solvate thereof. The claimed compounds are useful for the treatment of a disease by activating the EP4 receptor.

Description

BICYCLIC DERIVATIVES AS EP4 AGONISTS

Field of the invention The present invention concerns bicyclic derivatives having EP4 receptor agonistic properties. The invention further relates to methods for their preparation and pharmaceutical compositions comprising them. The invention also relates to the use of said compounds for the manufacture of a medicament for the prevention or the treatment of a disease by activating the EP4 receptor.

Background prior art

WO2005/047268 relates to substituted pyrimidine compositions that are capable of modulating the activity of receptors of the NGFI-B family.

WO2006/030031 relates to thieno -pyridine and thieno -pyrimidine derivatives that are positive allosteric modulators of metabotropic receptors-subtype 2.

WO2005/116010 relates to phenyl or pyridyl derivatives having EP4 agonistic activity.

EP 1544202 relates to herbicidal compounds.

US2006/128729 describes bicyclic pyrazole derivatives for the treatment of diseases associated with cellular proliferation, diseases related to glycosidase expression or inflammatory conditions.

EP675124 relates to purine derivatives as anti- inflammatory agents.

US2006084650 discloses (pyrazolyl)(imidazopyrimidinyl)amines as kinase inhibitors.

Description of the invention The compounds of the invention differ from the prior art compounds in structure, in their pharmacological activity and/or pharmacological potency.

One aspect of the present invention relates to a compound of formula

including any stereochemical^ isomeric form thereof, wherein A represents a 5 or 6-membered aromatic heterocycle containing 1 , 2 or 3 heteroatoms, each heteroatom independently being selected from O, N or S; said heterocycle optionally being substituted with 1 , 2 or 3 R⁴ substituents; ring E represents a partially saturated or aromatic 5-membered heterocycle wherein the dotted lines represent an optional double bond and wherein B, C and D each independently represent CH₂, CH, N, NH, S or O and F represents N or C, provided that the 5-membered ring contains 1, 2 or 3 heteroatoms;

X represents a direct bond or Ci_4alkanediyl;

Y represents N or CH; R¹ represents hydrogen or fluoro;

R² represents hydrogen, halo, cyano, C^alkyl, Ci_6alkyloxy, Ci_6alkylcarbonyl or

C i -βalky lcarbony lamino ;

R³ represents hydrogen, halo, C^alkyl, Ci_6alkyloxy, cyano, nitro, amino or mono-or di(C i _6alkyl)amino ; R⁴ represents halo; hydroxyl; carboxyl; Ci_6alkyl optionally substituted with one or two substituents, each substituent independently selected from cyano, carboxyl or

Ci_₆alkyloxycarbonyl; polyhaloCi-βalkyl; Ci_₆alkyloxycarbonyl; polyhaloCi-βalkyloxy;

C₂-₆alkenyl optionally substituted with one or two substituents, each substituent independently selected from cyano, carboxyl or Ci_₆alkyloxycarbonyl; cyano; nitro; amino; mono-or di(Ci_6alkyl)amino; Ci_6alkyloxy optionally substituted with at least one substituent, in particular one, two or three substituents, each substituent independently selected from hydroxyl, halo, cyano, carboxyl, Ci_₆alkyloxycarbonyl or

NR⁵R⁶;

R⁵ and R⁶ each independently represent hydrogen, Ci_₆alkyl, C₂-₆alkenyl, C₃-₆cycloalkyl; or

RR⁵⁵ aanndd I R⁶ together with the nitrogen atom to which they are attached form a radical of formula

_{with A} representing O, NR⁷, CR⁸R⁹ or S; R⁷ represents hydrogen, Ci_₆alkyl, C₂-₆alkenyl, Ci_₆alkylcarbonyl, Ci_₆alkyloxycarbonyl, arylCi-βalkyl or aryl;

R⁸ and R⁹ each independently represent hydrogen, Ci_₆alkyloxy, halo, amino, mono-or di(Ci_₆alkyl)amino, C_halky!, Ci_₆alkyloxycarbonylCi_₆alkyl or C₂-₆alkenyl; or R⁸ and R⁹ together with the carbon atom to which they are attached represent C(=O) or a 5, 6 or 7-membered saturated heterocyclic ring containing 1 or 2 oxygen atoms; n represents an integer of value 1, 2 or 3; p and q each independently represent an integer of value 0, 1, 2 or 3; aryl represents phenyl or phenyl substituted with at least one substituent, in particular one, two or three substituents, each substituent independently selected from halo, Ci_6alkyl, Ci_6alkyloxy, cyano, nitro, amino or mono-or di(Ci_6alkyl)amino; provided that R³ may only be other than hydrogen if at least one of R¹ or R² is other than hydrogen; a JV-oxide thereof, a pharmaceutically acceptable salt thereof or a solvate thereof.

The present invention also relates to the use of a compound of formula (I) for the manufacture of a medicament for preventing or treating a disease by activating the EP4 receptor, in particular for treating a disease by activating the EP4 receptor, in particular for preventing or treating, in particular for treating, a disease associated with loss of bone mass.

As used hereinbefore or hereinafter Ci_4alkyl as a group or part of a group defines straight or branched chain saturated hydrocarbon radicals having from 1 to 4 carbon atoms such as methyl, ethyl, propyl, 1-methylethyl, butyl; Ci_₆alkyl as a group or part of a group defines straight or branched chain saturated hydrocarbon radicals having from 1 to 6 carbon atoms such as the group defined for Ci_4alkyl and pentyl, hexyl, 2-methylbutyl and the like; Ci_4alkanediyl defines straight or branched chain saturated bivalent hydrocarbon radicals having from 1 to 4 carbon atoms such as methylene, 1 ,2-ethanediyl or 1 ,2-ethylidene, 1,3-propanediyl or 1,3-propylidene, 1 ,4-butanediyl or 1 ,4-butylidene and the like; C2-6alkenyl as a group or part of a group defines straight or branched chain hydrocarbon radicals having from 2 to 6 carbon atoms and having 1 double bond such as ethenyl, propenyl, butenyl, pentenyl, hexenyl, 3-methylbutenyl and the like; C_{3 6}cycloalkyl is generic to cyclopropyl, cyclobutyl, cyclopentyl and cyclohexyl.

The term halo is generic to fluoro, chloro, bromo and iodo. As used hereinbefore or hereinafter, polyhaloCi-βalkyl as a group or part of a group is defined as mono- or polyhalosubstituted Ci_6alkyl, for example methyl substituted with one or more fluoro atoms, for example, difluoromethyl or trifluoromethyl, 1,1-difluoro-ethyl or 1,1- difluoro-2,2,2-trifluoroethyl and the like. In case more than one halogen atoms are attached to a Ci_₆alkyl group within the definition of polyhaloCi-βalkyl, they may be the same or different. Particular examples of 5, 6 or 7-membered saturated heterocyclic rings comprising 1 or 2 oxygen atoms are tetrahydrofuranyl, dioxolanyl, dihydrooxazolyl, isoxazolidinyl, oxadiazolidinyl, dioxanyl, morpholinyl, dioxepanyl.

Particular examples of 5 or 6-membered aromatic heterocycles comprising 1 , 2 or 3 heteroatoms each independently being selected from O, N or S are pyrrolyl, furyl, thienyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, pyrazolyl, triazolyl, thiadiazolyl, oxadiazolyl, tetrazolyl, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, triazinyl, pyranyl.

When any variable occurs more than one time in any constituent, each definition is independent.

For therapeutic use, salts of the compounds of formula (I) are those wherein the counterion is pharmaceutically acceptable. However, salts of acids and bases which are non-pharmaceutically acceptable may also find use, for example, in the preparation or purification of a pharmaceutically acceptable compound. All salts, whether pharmaceutically acceptable or not are included within the ambit of the present invention.

The pharmaceutically acceptable salts as mentioned hereinbefore or hereinafter are meant to comprise the therapeutically active non-toxic acid addition salt forms which the compounds of formula (I) are able to form. The latter can conveniently be obtained by treating the base form with such appropriate acids as inorganic acids, for example, hydrohalic acids, e.g. hydrochloric, hydrobromic and the like; sulfuric acid; nitric acid; phosphoric acid and the like; or organic acids, for example, acetic, propanoic, hydroxy- acetic, 2-hydroxypropanoic, 2-oxopropanoic, oxalic, malonic, succinic, maleic, fumaric, malic, tartaric, 2-hydroxy-l,2,3-propanetricarboxylic, methanesulfonic, ethanesulfonic, benzenesulfonic, 4-methylbenzenesulfonic, cyclohexanesulfonic, 2-hydroxybenzoic, 4-amino-2-hydroxybenzoic and the like acids. Conversely the salt form can be converted by treatment with alkali into the free base form.

The compounds of formula (I) containing acidic protons may be converted into their therapeutically active non-toxic metal or amine addition salt forms by treatment with appropriate organic and inorganic bases. The pharmaceutically acceptable salts as mentioned hereinbefore or hereinafter are meant to also comprise the therapeutically active non-toxic metal or amine addition salt forms (base addition salt forms) which the compounds of formula (I) are able to form. Appropriate base addition salt forms comprise, for example, the ammonium salts, the alkali and earth alkaline metal salts, e.g. the lithium, sodium, potassium, magnesium, calcium salts and the like, salts with organic bases, e.g. primary, secondary and tertiary aliphatic and aromatic amines such as methylamine, ethylamine, propylamine, isopropylamine, the four butylamine isomers, dimethylamine, diethylamine, diethanolamine, dipropylamine, diisopropylamine, di-n-butylamine, pyrrolidine, piperidine, morpholine, trimethylamine, triethylamine, tripropylamine, quinuclidine, pyridine, quinoline and isoquinoline, the benzathine, TV-methyl-D-glucamine, 2-amino-2-(hydroxymethyl)-l,3- propanediol, hydrabamine salts, and salts with amino acids such as, for example, arginine, lysine and the like.

Conversely the salt form can be converted by treatment with acid into the free acid form.

The term salt also comprises the quaternary ammonium salts (quaternary amines) which the compounds of formula (I) are able to form by reaction between a basic nitrogen of a compound of formula (I) and an appropriate quaternizing agent, such as, for example, an optionally substituted Ci_6alkylhalide, arylhalide, Ci_6alkylcarbonylhalide, arylcarbonylhalide, or arylCi-βalkylhalide, e.g. methyliodide or benzyliodide. Other reactants with good leaving groups may also be used, such as for example Ci_6alkyl trifluoromethanesulfonates, Ci_6alkyl methanesulfonates, and Ci_6alkyl/?-toluenesulfonates. A quaternary amine has a positively charged nitrogen. Pharmaceutically acceptable counterions include chloro, bromo, iodo, trifluoroacetate, acetate, triflate, sulfate, sulfonate. The counterion of choice can be introduced using ion exchange resins.

The term solvate comprises the hydrates and solvent addition forms which the compounds of formula (I) are able to form, as well as the salts thereof. Examples of such forms are e.g. hydrates, alcoholates and the like.

The iV-oxide forms of the present compounds are meant to comprise the compounds of formula (I) wherein one or several tertiary nitrogen atoms are oxidized to the so-called iV-oxide.

It will be appreciated that some of the compounds of formula (I) and their TV-oxides, salts, and solvates may contain one or more centers of chirality and exist as stereochemically isomeric forms. The term "stereochemically isomeric forms" as used hereinbefore or hereinafter defines all the possible stereoisomeric forms which the compounds of formula (I), and their TV-oxides, salts, or solvates may possess. Unless otherwise mentioned or indicated, the chemical designation of compounds denotes the mixture of all possible stereochemically isomeric forms, said mixtures containing all diastereomers and enantiomers of the basic molecular structure as well as each of the individual isomeric forms of formula (I) and their JV-oxides or salts, substantially free, i.e. associated with less than 10%, preferably less than 5%, in particular less than 2% and most preferably less than 1% of the other isomers. Thus, when a compound of formula (I) is for instance specified as (E), this means that the compound is substantially free of the (Z) isomer.

In particular, stereogenic centers may have the R- or S-configuration; substituents on bivalent cyclic (partially) saturated radicals may have either the cis- or trans- configuration. Compounds encompassing double bonds can have an E (entgegen) or Z (zusammen) -stereochemistry at said double bond. The terms cis, trans, R, S, E and Z are well known to a person skilled in the art.

Stereochemically isomeric forms of the compounds of formula (I) are obviously intended to be embraced within the scope of this invention.

Following CAS-nomenclature conventions, when two stereogenic centers of known absolute configuration are present in a molecule, an R or S descriptor is assigned (based on Cahn-Ingold-Prelog sequence rule) to the lowest-numbered chiral center, the reference center. The configuration of the second stereogenic center is indicated using relative descriptors [R*,R* ] or [i?*,i_>*], where the first R* is always specified as the reference center and [R*,R*] indicates centers with the same chirality and [i?*,i_>*] indicates centers of unlike chirality. For example, if the lowest-numbered chiral center in the molecule has an S configuration and the second center is R, the stereo descriptor would be specified as S-[R*, S*]. If "α" and "β" are used : the position of the highest priority substituent on the asymmetric carbon atom in the ring system having the lowest ring number, is arbitrarily always in the "α" position of the mean plane determined by the ring system. The position of the highest priority substituent on the other asymmetric carbon atom in the ring system relative to the position of the highest priority substituent on the reference atom is denominated "α", if it is on the same side of the mean plane determined by the ring system, or "β", if it is on the other side of the mean plane determined by the ring system. The compounds of (I) may be synthesized in the form of racemic mixtures of enantiomers which can be separated from one another following art-known resolution procedures. The racemic compounds of formula (I) may be converted into the corresponding diastereomeric salt forms by reaction with a suitable chiral acid. Said diastereomeric salt forms are subsequently separated, for example, by selective or fractional crystallization and the enantiomers are liberated therefrom by alkali. An alternative manner of separating the enantiomeric forms of the compounds of formula (I) involves liquid chromatography using a chiral stationary phase. Said pure stereochemically isomeric forms may also be derived from the corresponding pure stereochemically isomeric forms of the appropriate starting materials, provided that the reaction occurs stereospecifically. Preferably if a specific stereoisomer is desired, said compound will be synthesized by stereospecifϊc methods of preparation. These methods will advantageously employ enantiomerically pure starting materials.

Some of the compounds of formula (I) may also exist in their tautomeric form. Such forms although not explicitly indicated in the above formula (I) are intended to be included within the scope of the present invention.

Whenever used hereinafter, the term "compounds of formula (I)" or any subgroup thereof, is meant to also include their JV-oxide forms, their salts, their stereochemically isomeric forms and their solvates. Of special interest are those compounds of formula (I) which are stereochemically pure.

Whenever used hereinbefore or hereinafter that substituents can be selected each independently out of a list of numerous definitions, all possible combinations are intended which are chemically possible.

A first interesting embodiment of the present invention are those compounds of formula (I) having the following formula

including any stereochemically isomeric form thereof, wherein A represents a 5 or 6-membered aromatic heterocycle containing 1 , 2 or 3 heteroatoms, each heteroatom independently being selected from O, N or S; said heterocycle optionally being substituted with 1 , 2 or 3 R⁴ substituents; ring E represents a partially saturated or aromatic 5-membered heterocycle wherein the dotted lines represent an optional double bond and wherein B, C and D each independently represent CH₂, CH, N, NH, S or O provided that the 5-membered ring contains 1 or 2 heteroatoms;

X represents a direct bond or Ci_4alkanediyl;

R¹ represents hydrogen or fluoro; R² represents hydrogen, halo, cyano, Ci_6alkyl, Ci_6alkyloxy, Ci_6alkylcarbonyl or

C i -βalky lcarbony lamino ;

R represents hydrogen, halo, Ci_6alkyl, Ci_6alkyloxy, cyano, nitro, amino or mono-or di(C i _₆alkyl)amino ;

R⁴ represents halo; hydroxyl; carboxyl; Ci_6alkyl optionally substituted with one or two substituents, each substituent independently selected from cyano, carboxyl or

C₂-₆alkenyl optionally substituted with one or two substituents, each substituent independently selected from cyano, carboxyl or Ci_₆alkyloxycarbonyl; cyano; nitro; amino; mono-or di(Ci_6alkyl)amino; Ci_6alkyloxy optionally substituted with at least one substituent, each substituent independently selected from hydroxyl, halo, cyano, carboxyl, Ci_₆alkyloxycarbonyl or NR⁵R⁶;

R⁵ and R⁶ each independently represent hydrogen, Ci_₆alkyl, C₂-₆alkenyl,

C₃-₆cycloalkyl; or

_{with A} representing O, NR⁷, CR⁸R⁹ or S;

R⁷ represents hydrogen, Ci_₆alkyl, C₂-₆alkenyl, Ci_₆alkylcarbonyl, Ci_₆alkyloxy- carbonyl, arylCi-βalkyl or aryl;

R⁸ and R⁹ each independently represent hydrogen, Ci_₆alkyloxy, halo, amino, mono-or di(Ci_₆alkyl)amino, C_halky!, Ci-ealkyloxycarbonylCi-βalkyl or C₂-₆alkenyl; or

R⁸ and R⁹ together with the carbon atom to which they are attached represent C(=O) or a 5, 6 or 7-membered saturated heterocyclic ring containing 1 or 2 oxygen atoms; n represents an integer of value 1, 2 or 3; p and q each independently represent an integer of value 0, 1, 2 or 3; aryl represents phenyl or phenyl substituted with at least one substituent, each substituent independently selected from halo, Ci_6alkyl, Ci_6alkyloxy, cyano, nitro, amino or mono-or di(Ci_6alkyl)amino; provided that R³ may only be other than hydrogen if at least one of R¹ or R² is other than hydrogen; a N-oxide thereof, a pharmaceutically acceptable salt thereof or a solvate thereof.

A second interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein X represents a direct bond.

A third interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein X represents Ci_4alkanediyl, in particular CH₂.

A fourth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein R¹ represents hydrogen or wherein R¹ represents fluoro.

A fifth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein R² represents hydrogen, halo, cyano, Ci_₆alkylcarbonyl, Ci_6alkyloxy or Ci_6alkylcarbonylamino; in particular hydrogen, halo, cyano, Ci_₆alkylcarbonyl or Ci_₆alkyloxy; more in particular hydrogen, halo, cyano or Ci_₆alkyloxy.

A sixth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein R³ represents hydrogen, halo or Ci_₆alkyloxy; more in particular hydrogen.

A seventh interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein R⁴ represents halo; hydroxyl; carboxyl; Ci_₆alkyl optionally substituted with one or two substituents, in particular one substituent, each substituent independently selected from cyano, carboxyl or Ci_₆alkyloxycarbonyl; polyhalo- Ci_₆alkyl; Ci_₆alkyloxycarbonyl; polyhaloCi-βalkyloxy; C₂-₆alkenyl optionally substituted with one or two substituents, in particular one substituent, each substituent independently selected from cyano, carboxyl or Ci_₆alkyloxycarbonyl; cyano; nitro; amino; mono-or di(Ci_6alkyl)amino; Ci_6alkyloxy optionally substituted with at least one substituent, in particular one, two or three substituents, more in particular one or two substituents, each substituent independently selected from hydroxyl, halo, cyano, carboxyl, Ci_₆alkyloxycarbonyl or NR⁵R⁶; wherein R⁵ and R⁶ each independently represent hydrogen, d-βalkyl, C₂-₆alkenyl, C₃-₆cycloalkyl; or R⁵ and R⁶ together with the nitrogen atom to which they are attached form a radical of formula

h A representing O, NR⁷, CH₂ or S; in particular R⁴ represents halo; hydroxyl; Ci_₆alkyl; Ci_₆alkyloxycarbonyl; C₂-₆alkenyl substituted with carboxyl; cyano; Ci_6alkyloxy optionally substituted with cyano, carboxyl, Ci_ βalkyloxycarbonyl or NR⁵R⁶; wherein R⁵ and R⁶ each independently represent hydrogen, Ci_₆alkyl, C₂-₆alkenyl, C₃_₆Cycloalkyl; or R⁵ and R⁶ together with the nitrogen atom to which they are attached form a radical of formula

in particular R⁴ represents halo; hydroxyl; Ci_₆alkyl; Ci_₆alkyloxycarbonyl; C₂-₆alkenyl substituted with carboxyl; cyano; Ci_6alkyloxy optionally substituted with cyano, carboxyl,

— N A (_a_i) Ci_₆alkyloxycarbonyl or a radical of formula ^ — ' with A representing O,

CH₂ or S; even more in particular R⁴ represents halo; hydroxyl; d-βalkyl; d-βalkyloxycarbonyl; cyano; C₂-6alkenyl substituted with carboxyl; Ci_6alkyloxy optionally substituted with cyano, Ci_₆alkyloxycarbonyl or a radical of formula

— N A _(a-i)

^ — ' with A representing O or CH₂; still more in particular R represents halo; Ci_₆alkyloxy optionally substituted with cyano, Ci_₆alkyloxycarbonyl or a radical

— N A (_a-i) of formula ^ — ' with A representing O or CH₂; cyano; hydroxyl; Ci_₆alkyl; C₂-₆alkenyl substituted with carboxyl; yet even more in particular R⁴ represents halo; Ci_6alkyloxy optionally substituted with cyano, Ci_6alkyloxycarbonyl

— N A (_a-i) or a radical of formula ^ — ' with A representing O or CH₂; cyano; hydroxyl; or Ci_6alkyl. Preferably, R⁴ represents halo. An eighth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein A is an unsubstituted heterocycle or wherein A is a heterocycle substituted with 1 R⁴.

A ninth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein n represents 1.

A tenth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein p represents 1, 2 or 3, in particular 1 or 2, more in particular 1.

An eleventh interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein q represents 1, 2 or 3, in particular 1 or 2, more in particular 1.

A twelfth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein R¹ is fluoro and R² is hydrogen.

A thirteenth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein R¹ is hydrogen and R² is halo, cyano, Ci_₆alkyl, Ci_₆alkyloxy, Ci_ βalkylcarbonyl or Ci_₆alkylcarbonylamino; more in particular R¹ is hydrogen and R² represents halo, cyano, Ci_6alkylcarbonyl, Ci_6alkyloxy or Ci_6alkylcarbonylamino.

A fourteenth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein R¹ is fluoro and R² is halo, cyano, Ci_6alkyl, Ci_6alkyloxy, Ci_ βalkylcarbonyl or Ci_6alkylcarbonylamino; more in particular R¹ is fluoro and R² represents halo, cyano, Ci_6alkylcarbonyl, Ci_6alkyloxy or Ci_6alkylcarbonylamino.

A fifteenth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein R¹ and R² are both hydrogen. A sixteenth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein A is an optionally substituted pyridyl or optionally substituted thienyl, in particular optionally substituted 3-pyridyl, optionally substituted 4-pyridyl or optionally substituted 2-thienyl, more in particular unsubstituted 3-pyridyl, unsubstituted 4-pyridyl or unsubstituted 2-thienyl.

A seventeenth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein Y represents N.

An eighteenth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein Y represents CH.

A nineteenth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein F represents C and the dotted line attached to F represents a bond.

A twentieth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein F represents N and the dotted line attached to F does not represents a bond.

A twenty first interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein B, C and D each independently represent CH₂, CH, N, NH or O.

A twenty second interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein the dotted lines at B, C and D in ring E do not represent a bond.

A twenty third interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein ring E does not represent pyrazole. A twenty fourth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein ring E contains 1 or 2 heteroatoms.

A twenty fifth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein R⁸ and R⁹ each independently represent hydrogen, Ci_₆alkyloxy, halo, amino, mono-or di(Ci_₆alkyl)amino, Ci_₆alkyl, Ci_₆alkyloxycarbonylCi_₆alkyl or C₂-₆alkenyl.

A twenty sixth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein the compound of formula (I) is a compound having the following formula

including any stereochemically isomeric form thereof, a JV-oxide thereof, a pharmaceutically acceptable salt thereof or a solvate thereof.

A twenty seventh interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein the compound of formula (I) is a compound having the following formula

including any stereochemically isomeric form thereof, a JV-oxide thereof, a pharmaceutically acceptable salt thereof or a solvate thereof. A twenty eighth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein the compound of formula (I) is a compound having the following formula

A twenty ninth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein the compound of formula (I) is a compound having the following formula

A thirtieth interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein the compound of formula (I) is a compound having the following formula

including any stereochemically isomeric form thereof, a JV-oxide thereof, a pharmaceutically acceptable salt thereof or a solvate thereof. A thirty first interesting embodiment of the present invention are those compounds of formula (I) or any subgroup thereof as mentioned hereinbefore as interesting embodiment wherein the compound of formula (I) is a compound having the following formula

including any stereochemically isomeric form thereof, a N-oxide thereof, a pharmaceutically acceptable salt thereof or a solvate thereof.

A thirty second interesting embodiment of the present invention are those compounds of formula (1-1), (I-l-a) or (I-l-b) wherein one or more, preferably all, of the following restrictions apply : a) X represents a direct bond or CH₂; b) R² represents hydrogen, halo, cyano, Ci_6alkylcarbonyl, Ci_6alkyloxy or Ci_6alkylcarbonylamino; c) R represents hydrogen, halo or Ci_6alkyloxy; d) R⁴ represents halo; e) n represents 1 ; f) A represents optionally substituted pyridyl, or thienyl; g) Y represents N.

A thirty third interesting embodiment of the present invention are those compounds of formula (I) wherein the compound is selected from the group consisting of

4-(2,4-difluoro-phenoxy)-2-pyridin-4-yl-thieno[3,2-d]pyrimidine; 4-(4-chloro-2-fluoro-phenoxy)-2-pyridin-4-yl-thieno[3,2-d]pyrimidine; 4-(2-pyridin-4- yl-thieno[3,2-d]pyrimidin-4-yloxy)-benzonitrile;

2-pyridin-4-yl-4-(2,4,6-trifluoro-phenoxy)-thieno[3,2-d]pyrimidine;

3-fluoro-4-(2-pyridin-4-yl-thieno[3,2-d]pyrimidin-4-yloxy)-benzonitrile;

4-(2-pyridin-3-yl-thieno[3,2-d]pyrimidin-4-yloxy)-benzonitrile; 4-[2-(2-chloro-pyridin-4-yl)-thieno[3,2-d]pyrimidin-4-yloxy]-3-fluoro-benzonitrile; a JV-oxide thereof, a pharmaceutically acceptable salt thereof or a solvate thereof. In general, compounds of formula (I) can be prepared by reacting an intermediate of formula (II) wherein Wi represents a suitable leaving group, such as for example halo, e.g. chloro and the like, or -O-S(=O)₂-CF₃, with an intermediate of formula (III) in the presence of a suitable base, such as for example K2CO3 or sodium hydride, and a suitable solvent, such as for example acetonitrile or dimethylsulfoxide.

Depending on the ring E moiety, it may be preferred to protect the intermediate of formula (II) before reaction with an intermediate of formula (III), for instance when ring E represents an imidazolyl moiety as in the compounds of formula (I-l-c). Therefore, compounds of formula (I-l-c) can be prepared by reacting in a first step (a), an intermediate of formula (II-a) with an intermediate of formula P-W₂ wherein P represents a suitable protective group, such as for example benzyl or Ci_4alkyloxy- Ci_4alkyl and wherein W₂ represents a suitable leaving group, such as for example halo, e.g. chloro and the like, in the presence of a suitable base, such as for example K2CO3 or sodium hydride, and a suitable solvent, such as for example acetonitrile or dimethylsulfoxide. The product obtained in step (a), is then reacted in a next step (b) with an intermediate of formula (III) in the presence of a suitable base, such as for example K2CO3 or sodium hydride, and a suitable solvent, such as for example acetonitrile or dimethylsulfoxide. The product obtained in step (b) is then deprotected in a next step (c) by reaction with a suitable acid, such as for example hydrochloric acid, in the presence of a suitable solvent, such as for example an alcohol, e.g. methanol, or acetonitrile.

(Il-a)

(III)

(b)

(I-l-c)

The compounds of formula (I) may further be prepared by converting compounds of formula (I) into each other according to art-known group transformation reactions.

The compounds of formula (I) may be converted to the corresponding JV-oxide forms following art-known procedures for converting a trivalent nitrogen into its iV-oxide form. Said JV-oxidation reaction may generally be carried out by reacting the starting material of formula (I) with an appropriate organic or inorganic peroxide. Appropriate inorganic peroxides comprise, for example, hydrogen peroxide, alkali metal or earth alkaline metal peroxides, e.g. sodium peroxide, potassium peroxide; appropriate organic peroxides may comprise peroxy acids such as, for example, benzenecarboper- oxoic acid or halo substituted benzenecarboperoxoic acid, e.g. 3-chlorobenzenecarbo- peroxoic acid, peroxoalkanoic acids, e.g. peroxoacetic acid, alkylhydroperoxides, e.g. tert.butyl hydro-peroxide. Suitable solvents are, for example, water, lower alcohols, e.g. ethanol and the like, hydrocarbons, e.g. toluene, ketones, e.g. 2-butanone, halogenated hydrocarbons, e.g. dichloromethane, and mixtures of such solvents.

Compounds of formula (I) wherein R⁴ represents methoxy, can be converted into a compound of formula (I) wherein R⁴ represents hydroxyl, by reaction with BBr₃ in the presence of a suitable solvent, such as for example dichloromethane. Compounds of formula (I) wherein R⁴ represents hydroxyl, can be converted into a compound of formula (I) wherein R⁴ represents Chalky Io xy optionally substituted with cyano, halo, carboxyl, Ci_₆alkyloxycarbonyl or NR⁵R⁶, by reaction with W₃-Ci_ βalkyl-R⁴ wherein W3 represents a suitable leaving group, such as for example halo, e.g. chloro, and R⁴ represents hydrogen, cyano, halo, carboxyl, Ci_6alkyloxycarbonyl or NR⁵R⁶, in the presence of a suitable base, such as for example K2CO3, and a suitable solvent, such as for example acetonitrile, Λ/,Λ/-dimethylformamide or an alcohol, e.g. butanol.

Compounds of formula (I) wherein R⁴ represents Ci_6alkyloxy substituted with halo, can be converted into a compound of formula (I) wherein R⁴ represents Ci_6alkyloxy substituted with NR⁵R⁶, by reaction with HNR⁵R⁶ in the presence of a suitable base, such as for example K2CO3, optionally in the presence of a catalytic amount of KI, and in the presence of a suitable solvent, such as for example acetone or 4-methyl-2- pentanone (MIK).

Compounds of formula (I) wherein R⁴ represents Ci_6alkyloxy substituted with NR⁵R⁶ wherein NR⁵R⁶ represents a radical of formula (a) wherein A represents NH, can be converted into a compound of formula (I) wherein R⁴ represents Ci_6alkyloxy substituted with NR⁵R⁶ wherein NR⁵R⁶ represents a radical of formula (a) wherein A represents N-Q=O)-C₁ -βalkyl, by reaction with a suitable corresponding ester, such as for example acetyl acetate, in the presence of a suitable solvent, such as for example dichloromethane .

Compounds of formula (I) wherein R⁴ represents Ci_6alkyloxy substituted with NR⁵R⁶ wherein NR⁵R⁶ represents a radical of formula (a) wherein A represents N-Q=O)-O- Ci_6alkyl, can be converted into a compound of formula (I) wherein R⁴ represents Ci_ βalkyloxy substituted with NR⁵R⁶ wherein NR⁵R⁶ represents a radical of formula (a) wherein A represents NH, by reaction with a suitable acid, such as for example trifluoroacetic acid, in the presence of a suitable solvent, such as for example dichloromethane .

Compounds of formula (I) wherein R⁴ represents halo, can be converted into a compound of formula (I) wherein R⁴ represents C₂-₆alkenyl optionally substituted with cyano or Ci_₆alkyloxycarbonyl, by reaction with the appropriate C₂-₆alkenyl derivative in the presence of a suitable catalyst, such as for example palladium acetate, a suitable ligand, such as for example l,3-bis(diphenylphosphino)propane, a suitable base, such as for example triethylamine, and a suitable solvent, such as for example tetrahydrofuran. Compounds of formula (I) wherein R 4 represents C₂-₆alkenyl substituted with Ci_₆alkyloxycarbonyl can be converted into a compound of formula (I) wherein R > 4 represents C2-6alkenyl substituted with carboxyl, by reaction with a suitable base, such as for example sodium hydroxide, and a suitable solvent, such as dioxane.

Compounds of formula (I) wherein R⁴ represents halo, can also be converted into a compound of formula (I) wherein R⁴ represents Ci_₆alkyloxycarbonyl, e.g. CH₃-O-C(=O)-, by reaction with CO and Ci_6alkylOH, e.g. methanol, in the presence of potassium acetate, palladium acetate, a suitable ligand, such as for example l,3-bis(diphenylphosphino)propane, and a suitable solvent, such as for example tetrahydrofuran.

Compounds of formula (I) wherein R⁴ represents nitro, can be converted into a compound of formula (I) wherein R⁴ represents amino, by hydrogenation in the presence of a suitable catalyst, such as for example palladium on charcoal, a suitable catalyst poison, such as for example a thiophene solution, and a suitable solvent, such as for example tetrahydrofuran.

The compounds of formula (I) and some of the intermediates in the present invention may contain an asymmetric carbon atom. Pure stereochemically isomeric forms of said compounds and said intermediates can be obtained by the application of art-known procedures. For example, diastereoisomers can be separated by physical methods such as selective crystallization or chromatographic techniques, e.g. counter current distribution, chiral liquid chromatography and the like methods. Enantiomers can be obtained from racemic mixtures by first converting said racemic mixtures with suitable resolving agents such as, for example, chiral acids, to mixtures of diastereomeric salts or compounds; then physically separating said mixtures of diastereomeric salts or compounds by, for example, selective crystallization or chromatographic techniques, e.g. liquid chromatography and the like methods; and finally converting said separated diastereomeric salts or compounds into the corresponding enantiomers. Pure stereochemically isomeric forms may also be obtained from the pure stereochemically isomeric forms of the appropriate intermediates and starting materials, provided that the intervening reactions occur stereospecifically. An alternative manner of separating the enantiomeric forms of the compounds of formula (I) and intermediates involves liquid chromatography or SCF (Super Critical Fluid) chromatography, in particular using a chiral stationary phase.

Some of the intermediates and starting materials are known compounds and may be commercially available or may be prepared according to art-known procedures.

Intermediates of formula (II) wherein Wi represents chloro, said intermediates being represented by formula (II-b), can be prepared by reacting an intermediate of formula (IV) with SOCl₂ or phosphoric trichloride optionally in the presence of a suitable solvent, such as for example chloroform, N,Λ/-dimethylformamide or N,Λ/-dimethylbenzeneamine.

(IV) (^π-b)

Intermediates of formula (II) wherein Wi represents -O-S(=O)₂-CF₃, said intermediates being represented by formula (II-c), can be prepared by reacting an intermediate of formula (IV) with O(S(=O)2-CF3)2 in the presence of a suitable base, such as for example iV,jV-diisopropylethanamine, and a suitable solvent, such as for example dichloromethane .

(IV) (II-c)

Intermediates of formula (IV) wherein Y represents N, said intermediates being represented by formula (IV-a), can be prepared by cyclizing an intermediate of formula (V) in the presence of an aqueous ammoniak solution or NH₄OH or NH₃ in acetonitrile or an alcohol, e.g. methanol.

(V) (IV-a) Intermediates of formula (IV-a) can also be prepared by reacting an intermediate of formula (VI) in the presence of a suitable base, such as for example K2CO3, NaOCH₃ or l,8-diazabicyclo[5.4.0]undecene-7 (DBU), and a suitable solvent, such as for example water, alcohol, e.g. ethanol or isopropanol, chloroform.

(VI) (IV-a)

Intermediates of formula (IV-a) can also be prepared by reacting an intermediate of formula (XIV) with an intermediate of formula (XIII) in the presence of a suitable base such as potassium t-butoxide, and a suitable solvent, such as for example tetrahydrofuran.

Intermediates of formula (IV-b) can be prepared by reacting an intermediate of formula (VII) with an intermediate of formula (VIII) in the presence of a suitable base, such as for example K2CO3, and a suitable solvent, such as for example water.

(VII)

(TV-b)

Intermediates of formula (IV-c) can be prepared by reacting an intermediate of formula (XII) with an intermediate of formula (XIII) in the presence of lithium diisopropylamide (LDA) and a suitable solvent, such as for example tetrahydrofuran.

(IV-c)

Intermediates of formula (III) and (V) are known in the art or can be prepared according to art-known reactions. For example intermediates of formula (V) can be prepared by reacting an intermediate of formula (IX) with an intermediate of formula (X) wherein W₄ represents a suitable leaving group, such as for example halo, e.g. chloro and the like, in the presence of a suitable solvent, such as for example acetone or pyridine, and optionally a suitable base, such as for example pyridine or 4-dimethylaminopyridine.

Intermediates of formula (VI) can be prepared by reacting an intermediate of formula (XI) with an intermediate of formula (X) in the presence of a suitable base, such as for example N,N-dimethyl-4-pyridine amine, and a suitable solvent, such as for example pyridine.

Intermediates of formula (VI) can also be prepared from the corresponding carboxylic acid derivative of formula (VF) by reaction with ethyl chloro formate and NH3 in the presence of a suitable base, such as for example triethylamine, in a suitable solvent, such as for example acetonitrile.

(VI') (VI)

Intermediates of formula (VF) can be prepared by hydrolysis of an intermediate of formula (V) in the presence of a suitable base, such as for example KOH, and a suitable solvent, such as for example an alcohol, e.g. ethanol.

Pharmacological part

The compounds of formula (I) and any subgroup thereof show EP4 agonist properties, in particular selective EP4 agonist properties. EP4 is one of the four subtype receptors (EPl, EP2, EP3 EP4) of prostaglandine E2. Prostaglandins are arachidonic acid metabolites that are synthetized by the cyclo- oxygenase pathway. A major cyclo-oxygenase product is prostaglandin E₂ (PGE₂), which participates in a broad range of biological activities, such as smooth muscle relaxation, vasodilation, fever, inflammatory pain, enteric mucus secretion, renal regulation and bone formation. Mechanistically, PGE₂ can exert agonistic activities on four G-protein-coupled receptor subtypes, which are termed EPl, EP2, EP3 and EP4. Each of these receptor subtypes has a distinct pharmacological signature based on their ligand preference and coupling to intracellular signalling pathways. The EPl and EP3 receptors are coupled to calcium metabolism and inhibition of cyclic adenosine 5 -monophosphate (cAMP) via GqZG₁ G-proteins. In contrast, EP2 and EP4 are linked to the stimulation of adenylyl cyclase and increased cAMP synthesis via G_s G-proteins. A considerable number of PGE₂ effects are linked to its activation of the EP4 receptor, as evidenced by experiments using EP4-deficient mice and specific EP4 agonists or antagonists. Thus, EP4-deficient mice produce a reduced vasodepressor response following PGE₂ infusion, exhibit decreased contact hypersensitivity and show reduced incidence and intensity of disease in the collagen antibody- induced arthritis model, while they have increased colitis induced by dextran sulphate treatment. Furthermore, the absence of EP4 decreases bone mass and impaired fracture healing in aged male mice, whilst PGE₂ administration fails to induce bone formation in EP4-deficient mice. In rats, administration of EP4 selective agonists suppresses dextran sulphate colitis, restores bone mass and strength in both normal and aged, ovariectomized animals, reduces indomethacin- induced small intestinal ulceration, attenuates endotoxin/galactosamine-induced liver injury, reduces mercury chloride-evoked acute kidney failure and attenuated pain responses in Freund's complete adjuvant-induced joint inflammation. In glaucomatous Cynomolgus monkeys, topical application of an EP4 agonist reduces the increased intraocular pressure. In line with these findings, EP4 antagonists block the bone anabolic effects of PGE₂ in rats. Unfortunately, the use of PGE₂ in human therapy is compromised by its inability to discern EP receptor subtypes, which leads to a number of untoward effects, including nausea, emesis, diarrhea and hypotension.

Rather, based on the abovementioned observations, the use of selective EP4 agonists is believed to be of therapeutic interest, particularly for the treatment of diseases associated with loss of bone mass (primary and secondary osteoporosis, bone fracture, metastatic bone disease, rheumatoid arthritis, osteoarthritis, periodontitis, osteogenesis imperfecta, hypercalcemia), diseases associated with liver injury and acute hepatitis, renal failure and nephritis, ulcerative colitis, Crohn's disease, stomatitis, gastritis, ocular hypertension, glaucoma, neuropathic pain, bone pain, Reflex Sympathetic Dystrophy syndrome (RSD) also known as Complex Regional Pain Syndrome (CRPS)).

Due to their EP4 agonistic activity, the compounds of formula (I), their JV-oxides, pharmaceutically acceptable salts, or solvates are useful for the treatment or prevention, in particular for the treatment, of a disease by activating the EP4 receptor.

In view of the above-described pharmacological properties, the compounds of formula (I), their JV-oxides, pharmaceutically acceptable salts or solvates, may be used as a medicine. In particular, the present compounds can be used for the manufacture of a medicament for treating or preventing a disease by activating the EP4 receptor, in particular for treating a disease by activating the EP4 receptor. More in particular, the compounds of the invention can be used for the manufacture of a medicament for treating or preventing, preferably treating, a disease associated with loss of bone mass (primary and secondary osteoporosis, bone fracture, metastatic bone disease, rheumatoid arthritis, osteoarthritis, periodontitis, osteogenesis imperfecta, hypercalcemia), diseases associated with liver injury and acute hepatitis, renal failure and nephritis, ulcerative colitis, Crohn's disease, stomatitis, gastritis, ocular hypertension, glaucoma, neuropathic pain, bone pain, Reflex Sympathetic Dystrophy syndrome (RSD) also known as Complex Regional Pain Syndrome (CRPS)).

In view of the utility of the compounds of formula (I), there is provided a method of treating a warm-blooded mammal, including a human, suffering from or a method of preventing a warm-blooded mammal, including a human, to suffer from a disease by activating the EP4 receptor, in particular a method of treating a warm-blooded mammal, including a human, suffering from a disease by activating the EP4 receptor. Said methods comprise the administration of an effective amount of a compound of formula (I), a JV-oxide form thereof, a pharmaceutically acceptable salt thereof or a solvate thereof, to a warm-blooded mammal, including a human.

The present invention also provides compositions for preventing or treating a disease by activating the EP4 receptor, in particular for treating a disease by activating the EP4 receptor. Said compositions comprise a therapeutically effective amount of a compound of formula (I), a JV-oxide form thereof, a pharmaceutically acceptable salt thereof or a solvate thereof, and a pharmaceutically acceptable carrier or diluent. The compounds of the present invention may be formulated into various pharmaceutical forms for administration purposes. As appropriate compositions there may be cited all compositions usually employed for systemically administering drugs. To prepare the pharmaceutical compositions of this invention, an effective amount of the particular compound, optionally in salt form, as the active ingredient is combined in intimate admixture with a pharmaceutically acceptable carrier, which carrier may take a wide variety of forms depending on the form of preparation desired for administration. These pharmaceutical compositions are desirable in unitary dosage form suitable, particularly, for administration orally, rectally, percutaneously, or by parenteral injection. For example, in preparing the compositions in oral dosage form, any of the usual pharmaceutical media may be employed such as, for example, water, glycols, oils, alcohols and the like in the case of oral liquid preparations such as suspensions, syrups, elixirs, emulsions and solutions; or solid carriers such as starches, sugars, kaolin, diluents, lubricants, binders, disintegrating agents and the like in the case of powders, pills, capsules, and tablets. Because of their ease in administration, tablets and capsules represent the most advantageous oral dosage unit forms, in which case solid pharmaceutical carriers are obviously employed. For parenteral compositions, the carrier will usually comprise sterile water, at least in large part, though other ingredients, for example, to aid solubility, may be included. Injectable solutions, for example, may be prepared in which the carrier comprises saline solution, glucose solution or a mixture of saline and glucose solution. Injectable suspensions may also be prepared in which case appropriate liquid carriers, suspending agents and the like may be employed. Also included are solid form preparations, which are intended to be converted, shortly before use, to liquid form preparations. In the compositions suitable for percutaneous administration, the carrier optionally comprises a penetration enhancing agent and/or a suitable wetting agent, optionally combined with suitable additives of any nature in minor proportions, which additives do not introduce a significant deleterious effect on the skin. Said additives may facilitate the administration to the skin and/or may be helpful for preparing the desired compositions. These compositions may be administered in various ways, e.g., as a transdermal patch, as a spot-on, as an ointment.

The compounds of the present invention may also be administered via inhalation or insufflation by means of methods and formulations employed in the art for administration via this way. Thus, in general the compounds of the present invention may be administered to the lungs in the form of a solution, a suspension or a dry powder. Any system developed for the delivery of solutions, suspensions or dry powders via oral or nasal inhalation or insufflation are suitable for the administration of the present compounds.

The compounds of the present invention may also be topically administered in the form of drops, in particular eye drops. Said eye drops may be in the form of a solution or a suspension. Any system developed for the delivery of solutions or suspensions as eye drops are suitable for the administration of the present compounds.

It is especially advantageous to formulate the aforementioned pharmaceutical compositions in unit dosage form for ease of administration and uniformity of dosage. Unit dosage form as used herein refers to physically discrete units suitable as unitary dosages, each unit containing a predetermined quantity of active ingredient calculated to produce the desired therapeutic effect in association with the required pharmaceutical carrier. Examples of such unit dosage forms are tablets (including scored or coated tablets), capsules, pills, powder packets, wafers, suppositories, injectable solutions or suspensions and the like, and segregated multiples thereof.

The exact dosage and frequency of administration depends on the particular compound of formula (I) used, the particular condition being treated, the severity of the condition being treated, the age, weight, sex, extent of disorder and general physical condition of the particular patient as well as other medication the individual may be taking, as is well known to those skilled in the art. Furthermore, it is evident that said effective daily amount may be lowered or increased depending on the response of the treated subject and/or depending on the evaluation of the physician prescribing the compounds of the instant invention.

Depending on the mode of administration, the pharmaceutical composition will preferably comprise from 0.05 to 99 % by weight, more preferably from 0.1 to 70 % by weight, even more preferably from 0.1 to 50 % by weight of the active ingredient, and, from 1 to 99.95 % by weight, more preferably from 30 to 99.9 % by weight, even more preferably from 50 to 99.9 % by weight of a pharmaceutically acceptable carrier, all percentages being based on the total weight of the composition.

The following examples are intended to illustrate the present invention.

Experimental Part

Hereinafter "DMF" means N,Λ/-dimethylformamide, "DIPE" means diisopropyl ether, "DCM" means dichloromethane and "DMSO" means dimethylsulfoxide. Where solutions were "dried," they were generally dried over a drying agent such as Na₂SO₄ or MgSO₄. Where mixtures, solutions, and extracts were "concentrated", they were typically concentrated on a rotary evaporator under reduced pressure.

A. Preparation of the intermediate compounds

Example Al a) Preparation of intermediate 1

A mixture of 3-[(4-pyridinylcarbonyl)amino]-2-thiophenecarboxylic acid, methyl ester (0.057 mol) in NH₄OH (300 ml) was stirred at 150 ⁰C for 4 hours in a sealed vial. Then the reaction mixture was cooled and concentrated to half the original volume. The precipitate was filtered off. Yield: 1.8 g of intermediate 1. The filtrate could be neutralized with a 1 N HCl aqueous solution to obtain more product (1.2 g).

b) Preparation of intermediate 2

A mixture of intermediate 1 (0.013 mol) in phosphoric trichloride (50 ml) was stirred and refluxed for 3 hours. Then the solvent was evaporated. The residue was dissolved in H₂O and this solution was neutralized with an aqueous Na₂CO₃ solution. The mixture was extracted with ethyl acetate. The separated organic layer was dried, filtered and the solvent was evaporated. The residue was triturated under DIPE. The precipitate was filtered off. Yield : 2.2 g of intermediate 2. Example A2 a) Preparation of intermediate 15

3-[(3-pyridinylcarbonyl)amino]-2-thiophenecarboxylic acid, methyl ester (40 g, 0.15 mol) in a saturated aqueous NH₃ solution (350 ml) was heated in an autoclave at 120 ⁰C for 6 hours. Then the mixture was cooled. The precipitate was filtered off, washed with H₂O and dried. Yield: 9 g of intermediate 15 (26 %).

b) Preparation of intermediate 16

SOCl₂ (9.45 ml, 5 equivalent) was added dropwise to a solution of intermediate 15 (5 g, 0.0167 mol) in CHCl₃ (100 ml). Then DMF (8.4 ml) was added dropwise and the reaction mixture was heated under reflux for 3 hours. Then the mixture was cooled and the solvent was concentrated. Toluene (200 ml) was added to the residue and the mixture was concentrated again. The residue was slurried with DCM. The precipitate was filtered off, washed with DCM and dried. Yield: 3 g of intermediate 16 (73 %).

Example A3 a-1) Preparation of intermediate 3

2-Chloro-4-pyridinecarbonyl chloride (0.018 mol) in acetone (20 ml) was added dropwise to a stirring mixture of 3-amino-2-thiophenecarboxylic acid methyl ester (0.016 mol) in acetone (130 ml) and pyridine (10 ml). The reaction mixture was stirred overnight and then the solvent was evaporated. The residue was dissolved in DCM. The solution was washed with H₂O, dried, filtered and the filtrate's solvent was evaporated. The residue was triturated under DIPE. The precipitate was filtered off, yielding 3.47 g of intermediate 3.

a-2) Preparation of intermediate 17

β-Chloro-S-pyridinecarbonyl chloride (0.02 mol) in acetone (20 ml) was added dropwise to a stirring mixture of 3-amino-2-thiophenecarboxylic acid methyl ester (0.016 mol) in acetone (80 ml) and pyridine (10 ml) and the mixture was stirred overnight. The solvent was evaporated. The residue was dissolved in DCM, washed, dried, filtered and the filtrate's solvent was evaporated. The residue was triturated under DIPE/CH3OH. The precipitate was filtered off, yielding 3.9 g of intermediate 17.

b-1) Preparation of intermediate 4

A mixture of intermediate 3 (0.01 mol) and NH₃ in CH₃OH (200 ml) was stirred for 48 hours at 100 ⁰C. The reaction mixture was cooled and the solvent was evaporated. The residue was triturated under DIPE/CH3OH. The precipitate was filtered off. Yield: 2 g of intermediate 4 (66 %).

b-2) Preparation of intermediate 18

A mixture of intermediate 17 (0.013 mol) and NH₃ in CH₃OH (200 ml) was stirred for 48 hours at 90 ⁰C. The reaction mixture was cooled and the solvent was evaporated. The residue was triturated under DIPE/CH3OH. The precipitate was filtered off, yielding 2.6 g of intermediate 18 (76 %).

C-I) Preparation of intermediate 5

Phosphoric trichloride (0.011 mol) was added dropwise to a stirring mixture of intermediate 4 (0.0076 mol) in DMF (150 ml) and was stirred for 3 hours. Then the solvent was evaporated and the residue was dissolved in DCM. This solution was washed, dried, filtered and the filtrate's solvent was evaporated. The residue was triturated under DIPE. The precipitate was filtered off. Yield : 0.5 g of intermediate 5.

c-2) Preparation of intermediate 19

Phosphoric trichloride (0.02 mol) was added dropwise to a stirring mixture of intermediate 18 (0.01 mol) in DMF (150 ml) and was stirred for 3 hours. The reaction mixture was poured out in H₂O and stirred for 30 minutes. The precipitate was filtered off, yielding 2.1 g of intermediate 19.

Example A4 a-1) Preparation of intermediate 6

3-[(2-thienylcarbonyl)amino]-2-thiophenecarboxylic acid, methyl ester (max. 0.064 mol) was reacted in NH₄OH (400 ml) for 4 hours at 150 ⁰C. The reaction mixture was concentrated by partial evaporation of the solvent. The concentrate was acidified to pH = ± 1. The resulting precipitate was filtered off and was then triturated under DIPE/methanol 9/1. The precipitate was filtered off again and dried. Yield: 10 g of intermediate 6.

a-2) Preparation of intermediate 13

Intermediate 13 was prepared according to A4.a-1, except that the reaction mixture was stirred for 5 hours at 140 ⁰C. Yield: Intermediate 13 (98 %).

b-1) Preparation of intermediate 7

Intermediate 6 (0.021 mol) was dissolved in DMF (100 ml). Phosphoric trichloride (5 ml) was added drop wise. The reaction mixture was stirred for 2 hours at room temperature. The mixture was poured out into ice-water and the resulting precipitate was filtered off and dried. Yield: 4.5 g of intermediate 7 (90 %).

b-2) Preparation of intermediate 14

Intermediate 13 (0.008 mol) was suspended in DMF (50 ml). Phosphoric trichloride (1.5 g) was added and the reaction mixture was heated to 60 ⁰C. More phosphoric trichloride (3.5 g) was added and when the reaction was finished, the mixture was cooled to room temperature. The mixture was poured out on ice. The precipitate was filtered off and dried. Yield: 2.5 g of intermediate 14 (95 %).

Example A5 a) Preparation of intermediate 8

2-Amino-3-thiophenecarboxylic acid methyl ester (0.02 mol) was dissolved in acetone (80 ml). Then pyridine (3.2 g) was added, followed by dropwise addition of 4- pyridinecarbonyl chloride, hydrochloride (0.02 mol) in acetone (20 ml). The reaction mixture was stirred overnight. The solvent was evaporated and the residue was dissolved in DCM (200 ml). The organic solution was washed with 1 N HCl, 1 N NaOH and 2 x 50 ml of an aqueous NaCl solution and was then dried (MgSO₄), filtered and the solvent was evaporated. Yield: 3.1 g of intermediate 8 (50 %).

b) Preparation of intermediate 9

Intermediate 8 (2.7 g; 0.012 mol) was reacted in NH₄OH (50 ml) for 4 hours at 150 ⁰C. The reaction mixture was concentrated by partial evaporation of the solvent. The concentrate was acidified to pH = ± 1. The resulting precipitate was filtered off and was then triturated under DIPE/methanol 9/1. The precipitate was filtered off again and dried. Yield: 2.3 g of intermediate 9 (80 %). c) Preparation of intermediate 10

A mixture of intermediate 9 (0.003 mol) in phosphoric trichloride (25 ml) was stirred and refluxed for 4 hours. Then the solvent was evaporated and the residue was dissolved in DCM. The organic solution was washed with an aqueous NaHCOs solution and with brine, and was then dried (MgSO₄), filtered and the solvent was evaporated. Yield: 0.6 g of intermediate 10 (90 %).

Example A6 a) Preparation of intermediate 11

Tetrahydro-S-oxo^-thiophenecarboxylic acid, methyl ester (0.006 mol) was added drop wise to a solution of 4-pyridinecarboximidamide, monohydrochloride (0.006 mol) and K2CO3 (0.006 mol) in H₂O (100 ml). The reaction mixture was stirred for one hour at 60 ⁰C. The mixture was treated with 1 N HCl to get a neutral pH. The resulting precipitate was filtered off and dried. Yield: 1.2 g of intermediate 11 (85 %).

b) Preparation of intermediate 12

Intermediate 11 (0.005 mol) was dissolved in phosphoric trichloride (25 ml). The reaction mixture was stirred and refluxed for 3 hours. Then the solvent was evaporated and the residue was dissolved in DCM. This organic solution was washed with an aqueous NaHCOs solution and with brine, was then dried, filtered and the solvent was evaporated. Yield: 1.0 g of intermediate 12 (80 %).

B. Preparation of the final compounds

Example Bl a-1) Preparation of compound 22

A mixture of intermediate 2 (0.0004 mol), 3-fluoro-4-hydroxybenzonitrile (0.0005 mol) and K2CO3 (0.150 g) in acetonitrile (10 ml) was stirred overnight at 90 ⁰C. The reaction mixture was cooled and was then washed with a 10 % aqueous NaOH solution. This mixture was extracted with DCM. The separated organic layer was washed, dried, filtered and the solvent was evaporated. Yield: 0.116 g of compound 22.

a-2) Preparation of compound 25

Compound 25 was prepared from intermediate 2 according to Bl. a-1, but after the extraction with DCM, the separated organic layer was dried over Extrelut. The solvent was evaporated and the residue was triturated under DIPE/ethylacetate. The precipitate was filtered off and dried. Yield: 0.096 g of compound 25. Example B2 a-1) Preparation of compound 20

A mixture of intermediate 2 (0.0004 mol), 4-chloro-2-fluorophenol (0.0012 mol) and K2CO3 (0.165 g) in acetonitrile (2.5 ml) was stirred at 160 ⁰C in the microwave for 40 minutes. Then the mixture was stirred in a 10 % aqueous NaOH solution (10 ml). The precipitate was filtered off and was stirred in ethyl acetate. The precipitate was filtered off again and dried. Yield: 0.067 g of compound 20.

a-2) Preparation of compound 15

A mixture of intermediate 2 (0.2 g, 0.00051 mol), 4-hydroxybenzonitrile (0.29 g, 0.00243 mol, 3) and K₂CO₃ (0.335 g, 0.00243 mol) in acetonitrile (5 ml) was microwaved in a microwave tube for 35 minutes at 160 ⁰C. Then the reaction mixture was poured in a NaOH solution (10 %) and the mixture was stirred for 10 minutes. The precipitate was filtered off, washed with H₂O and dried. If necessary the product can be further purified by column chromatography (eluent: ethylacetate/hexane in varying proportions). Yield: Compound 15. a-3) Preparation of compound 9

Compound 9 was prepared from intermediate 16 according to B2.a-2.

Example B3 a) Preparation of compound 19

A mixture of intermediate 2 (0.0004 mol), 2,4-difluorophenol (0.0012 mol) and K₂CO₃ (0.165 g) in acetonitrile (10 ml) was stirred overnight at 90 ⁰C. The reaction mixture was cooled and was then washed with a 10 % aqueous NaOH solution. This mixture was extracted with DCM. The separated organic layer was dried over Extrelut and the filtrate's solvent was evaporated. The residue was triturated under DIPE/ethyl acetate. The precipitate was filtered off and dried. Yield: 0.034 g of compound 19.

b) Preparation of compound 35

A mixture of compound 19 (0.0021 mol) and 3-chlorobenzenecarboperoxoic acid (0.0027 mol) in DCM (75 ml) was stirred for 3 hours at room temperature. Then, the reaction mixture was washed with a 2 M NaOH solution. The organic layer was separated, washed, dried, filtered and the solvent was evaporated. Yield: 0.7 g of compound 35 (93 %).

Example B4 a) Preparation of compound 26

Reaction under N₂ flow. A mixture of NaH (0.00208 mol) in DMSO (4 ml) was stirred for 1 hour at 60 ⁰C. The mixture was cooled to room temperature and 4- (hydroxymethyl)-benzonitrile (0.0008 mol) was added. This mixture was stirred for 15 minutes. Then intermediate 2 (0.0008 mol) was added and the reaction mixture was stirred overnight. H₂O (10 ml) was added to the reaction mixture and stirred for 10 minutes. The precipitate was filtered off and was purified by column chromatography (Supelco) over silica gel (eluent: DCM/CH₃OH 99/1). The product fractions were collected and the solvent was evaporated. Yield : 0.055 g of compound 26.

Example B5 a-1) Preparation of compound 34

A mixture of intermediate 5 (0.00035 mol), 3-fluoro-4-hydroxybenzonitrile (0.00035 mol) and K₂CO₃ (0.0005 mol) in acetonitrile (10 ml) was stirred overnight at 90 ⁰C. The reaction mixture was cooled and then washed with a 2 N aqueous NaOH solution. The mixture was extracted with DCM. The separated organic layer was dried, filtered and the solvent was evaporated. The residue was triturated under CH3OH and the precipitate was filtered off. Yield : 0.072 g of compound 34. a-2) Preparation of compound 29

Compound 29 was prepared from intermediate 19 according to B5.a-1. Yield: Compound 29 (56 %).

Example B6 a-1) Preparation of compound 5

Intermediate 7 (0.0006 mol) was dissolved in acetonitrile (5 ml). K2CO3 (0.2 g) and 3- fluoro-4-hydroxybenzonitrile (0.1 g; ± 0.001 mol) were added and the reaction mixture was stirred and refluxed for one hour. Then the mixture was diluted with DCM and washed with 1 N NaOH. The organic layer was separated, dried, filtered and the solvent was evaporated. The residue was crystallized from 2-propanol, filtered off and dried. Yield: 0.040 g of compound 5 (20 %).

a-2) Preparation of compound 6

A mixture of intermediate 14 (0.003 mol), 2,4-difiuorophenol (0.86 g) and K₂CO₃ (1.5 g) in acetonitrile (15 ml) was stirred for 4 hours at 90 ⁰C. The reaction mixture was poured out into H₂O. DCM (50 ml) was added and the layers were separated. The separated organic layer was washed with 1 N NaOH (25 ml) and with 1 N HCl (25 ml), and was then dried, filtered and the solvent was evaporated. The residue was crystallized from DIPE. The precipitate was filtered off and dried. Yield: 0.75 g of compound 6 (50 %).

Example B7 a) Preparation of compound 38

Intermediate 10 (0.0004 mol) was dissolved in acetonitrile (10 ml). K₂CO₃ (0.200 g) and 4-hydroxybenzonitrile (0.15 g; ± 0.0013 mol) were added. The reaction mixture was stirred and refluxed for one hour. Then the mixture was cooled to room temperature, diluted with DCM and washed with 1 N NaOH. The organic layer was separated, dried (MgSO₄), filtered and the solvent was evaporated. The residue was crystallized from 2-propanol, filtered off and dried. Yield: 0.070 g of compound 38 (60 %).

Example B8 a) Preparation of compound 37

A mixture of intermediate 12 (0.0004 mol), 3-fluoro-4-hydroxybenzonitrile (0.001 mol) and K₂CO₃ (0.200 g) in acetonitrile (10 ml) was stirred and refluxed for 3 hours. The reaction mixture was cooled and was then diluted with DCM. The mixture was extracted with 1 N NaOH and with brine. The separated organic layer was dried, filtered and the solvent evaporated. The residue was crystallized from 2-propanol. The precipitate was filtered off and dried. If necessary, this fraction was again recrystallized from 2-propanol, filtered off and dried. Yield: 0.050 g of compound 37 (30 %).

The following compounds of formula (I), as depicted in Table 1 and 2, were prepared by analogy to the above examples (Ex. No.).

Table 1

db = direct bond

Table 2

C. Analytical Part

The mass of present compounds was recorded with LCMS (liquid chromatography mass spectrometry). The methods used are described below.

LCMS conditions

General procedure A

The HPLC measurement was performed using an Alliance HT 2790 (Waters) system comprising a quaternary pump with degasser, an autosampler, a column oven (set at 40 °C, unless otherwise indicated), a diode-array detector (DAD) and a column as specified in the respective methods below. Flow from the column was split to a MS spectrometer. The MS detector was configured with an electrospray ionization source. Mass spectra were acquired by scanning from 100 to 1000 in 1 second using a dwell time of 0.1 second. The capillary needle voltage was 3 kV and the source temperature was maintained at 140 ⁰C. Nitrogen was used as the nebulizer gas. Data acquisition was performed with a Waters-Micromass MassLynx-Openlynx data system.

General procedure B

The LC measurement was performed using an Acquity UPLC (Waters) system comprising a binary pump, a sample organizer, a column heater (set at 55 °C), a diode- array detector (DAD) and a column as specified in the respective methods below. Flow from the column was split to a MS spectrometer. The MS detector was configured with an electrospray ionization source. Mass spectra were acquired by scanning from 100 to 1000 in 0.18 seconds using a dwell time of 0.02 seconds. The capillary needle voltage was 3.5 kV and the source temperature was maintained at 140 ⁰C. Nitrogen was used as the nebulizer gas. Data acquisition was performed with a Waters-Micromass MassLynx-Openlynx data system.

Method 1 In addition to general procedure A: Reversed phase HPLC was carried out on an Xterra MS Cl 8 column (3.5 μm, 4.6 x 100 mm) with a flow rate of 1.6 ml/min. Three mobile phases (mobile phase A: 95% 25 mM ammoniumacetate + 5 % acetonitrile; mobile phase B: acetonitrile; mobile phase C: methanol) were employed to run a gradient condition from 100 % A to 1 % A, 49 % B and 50 % C in 6.5 minutes, to 1 % A and 99 % B in 1 minute and hold these conditions for 1 minute and reequilibrate with 100 % A for 1.5 minutes. An injection volume of 10 μl was used. Cone voltage was 10 V for positive ionization mode and 20 V for negative ionization mode.

Method 2

In addition to general procedure B: Reversed phase UPLC (Ultra Performance Liquid Chromatography) was carried out on a bridged ethylsiloxane/silica hybrid (BEH) Cl 8 column (1.7 μm, 2.1 x 50 mm; Waters Acquity) with a flow rate of 0.8 ml/min. Two mobile phases (mobile phase A: 0.1 % formic acid in H2θ/methanol 95/5; mobile phase B: methanol) were used to run a gradient condition from 95 % A and 5 % B to 5 % A and 95 % B in 1.3 minutes and hold for 0.2 minutes. An injection volume of 0.5 μl was used. Cone voltage was 10 V for positive ionization mode and 20 V for negative ionization mode.

Table 3 : LCMS data (R(t) means retention time in minutes; MH(+) means the protonated mass of the compound; procedure refers to the method used for LCMS).

D. Pharmacological example

Stimulation of cAMP-production in response to activation of the human EP4 receptor

Functional activity of the test compounds was assessed by measuring their potency to stimulate cAMP production upon activation of the human EP4 (hEP4) receptor through homogenous time resolved fluorescence (HTRF) assays. HEK293 cells stably transfected with hEP4 (Euroscreen, Belgium) were grown up to 80-90% confluence in T175 Falcon flasks in Dulbecco's modified Eagle's medium (DMEM) (Invitrogen) supplemented with 100 IU/ml penicillin G, 100 μg/ml streptomycin sulphate, 1 mM sodium pyruvate, 300 μg/ml L-glutamine and 10% heat inactivated foetal calf serum (Biochrom AG) in a humidified atmosphere of 5%CO₂ at 37°C. Cells were washed with 5 ml PBS containing EDTA 0.04% and detached from culture flasks with 3 ml PBS containing EDTA 0.04%, resuspended in prewarmed culture medium and centrifuged at 500 g for 5 minutes. The pellet was resuspended in freezing medium (culture medium with 20% heat inactivated foetal calf serum and 10% DMSO) to a concentration of 10⁷ cells/ml and frozen at -80⁰C. The experiments were performed with the cAMP Dynamic HTRF kit (CIS bio international, France), used according to the supplier's instructions. Specifically, cells were thawed rapidly by warming up the vials in a warm water bath at 37°C. The thawed cell suspension (2 ml; 10⁷ cells/ml) was transferred to a 50 ml Falcon tube and for each vial, 10 ml prewarmed culture medium was added. The falcon tube was centrifuged at 1,500 RPM for 5 minutes. The pellet was washed by adding 15 ml HBSS (per vial) and centrifuged at 1,500 RPM for another 5 minutes. The final pellet was resuspended in stimulation buffer (HBSS Ix, IBMX ImM, Hepes 5mM, MgCl₂ 1OmM, BSA 0.1%, pH 7.4). The suspension was counted in a nucleocounter and further diluted in stimulation buffer at a concentration of 500,000 cells/ml. The cells were seeded out in a MW384 COSTAR 3710 with the compounds using a Multidrop 384 at a density of 10,000 cells/well in 20 μl.

To test for agonistic activity, the cells were incubated for 30 minutes at room temperature in the dark in the presence of different concentrations of the compounds diluted in stimulation buffer in a final volume of 30 μl/well. The final concentration of DMSO (whenever needed to dissolve the compounds) did not exceed 1% (v/v) and was also included in the corresponding control samples. Reaction was stopped by adding 10 μl cAMP-d2 conjugate and subsequently 10 μl of anti-cAMP with the Multidrop. After equilibration of the reaction mixtures for 1 to 24 hours in dark at room temperature, fluorescence was measured at 665 nm and 620 nm using a Discovery microplate fluorescence counter (Perkin Elmer) and the signal ratio of 665 nm / 620 nm was calculated. The signal ratios of the test compounds were expressed relative to the signal ratios of the DMSO control (maximal signal ratio, no stimulation of cAMP) and prostaglandin E2 (PGE2) (minimal signal ratio, maximal stimulation of cAMP). From the dose response curves generated for each test compound, the dose at which 50% of the maximal stimulation of cAMP level is observed (EC50) and the level of stimulation reached of the test compound compared to PGE2 was calculated. Table 4 below discloses pECso values (= -log(ECso) values).

Table 4

Selectivity of the compounds for EP4 compared to EP2 can be demonstrated with the above test by also measuring stimulation of cAMP production upon activation of the human EP2 (hEP2) receptor by the present compounds. Selectivity of the compounds for EP4 can also be demonstrated by determining whether the compounds have activity on the EPl receptor, for instance by [Ca²⁺J₁ measurements in response to activation or inhibition of the monkey EPl receptor as follows : The antagonistic and agonistic effect of the test compounds on intracellular Ca²⁺ concentrations ([Ca²⁺J₁) was measured in a fluorescent based assay, using the calcium assay kit (Molecular Devices, Crawley, England). HEK293 cells stably transfected with monkey EPl receptor were cultured in T 175 Falcon flasks in Dulbecco's modified Eagle's medium (DMEM) (Invitrogen) supplemented with 100 IU/ml penicillin G, 100 μg/ml streptomycin sulphate, 1 mM sodium pyruvate, 300 μg/ml L-glutamine and 10% heat inactivated foetal calf serum (Biochrom AG) in a humidified atmosphere of 5%CO₂ at 37°C. Before the experiments, the cells were grown on 384-well (black wall/transparent bottom) plates from Greiner for 1 day until they reached confluency. The cells were loaded with loading buffer supplied by the kit supplemented with 10 mM probenecid and 0.1% fatty acid free bovine serum albumine, adjusted to pH 7.4 with 1 M Hepes-acid, for 90 minutes at 37°C in a CO₂ incubator. Ca²⁺ signals were measured in a Fluorometric Imaging Plate Reader (FLIPR, from Molecular Devices). To test antagonistic activity the loaded cells were preincubated with the compounds for 30 minutes at room temperature before starting the experiment in the FLIPR, where 100 nM of the reference agonist prostaglandin E2 (PGE2) was added. To test agonistic activity the compounds were added to the loaded cells during the measurement in the FLIPR where 1000 nM PGE2 was used as the reference agonist. In the FLIPR changes in relative fluorescence units were recorded in function of time. The final concentration of DMSO (whenever needed to dissolve the compounds) did not exceed 1% (v/v) and was also included in the corresponding control samples. ZD6416 was used as the reference antagonist. The peak fluorescence (maximum signal between 1 and 50 sec) was considered as the relevant signal. Concentration response curves were constructed based on peak fluorescence for each concentration of test drug. For antagonistic activity the peak fluorescence of the test compounds were expressed relative to the peak fluorescence of the DMSO control (minimal signal, maximal inhibition of calcium release) and 100 nM PGE2 (maximal signal, minimal inhibition of calcium release). From the inhibition curves generated for each test compound, the dose at which 50% of the maximal inhibition of calcium release is observed (IC₅₀ or PIC50 = -log(IC₅₀) values) and the level of inhibition reached of the test compound was calculated. For agonistic activity the peak fluorescence of the test compounds were expressed relative to the peak fluorescence of the DMSO control (minimal signal, minimal calcium release) and 1000 nM PGE2 (maximal signal, maximal calcium release). From the curves generated for each test compound, the dose at which 50% of the maximal stimulation of calcium release is observed (EC50 or pECso = -log(ECso) values) and the level of stimulation reached of the test compound was calculated. PEC50 (agonism) and pICso (antagonism) values for the tested compounds were <5.

Claims

1. A compound of formula

including any stereochemically isomeric form thereof, wherein

A represents a 5 or 6-membered aromatic heterocycle containing 1 , 2 or 3 heteroatoms, each heteroatom independently being selected from O, N or S; said heterocycle optionally being substituted with 1 , 2 or 3 R⁴ substituents; ring E represents a partially saturated or aromatic 5-membered heterocycle wherein the dotted lines represent an optional double bond and wherein B, C and D each independently represent CH₂, CH, N, NH, S or O and F represents N or C, provided that the 5-membered ring contains 1, 2 or 3 heteroatoms;

X represents a direct bond or Ci_4alkanediyl;

Y represents N or CH; R¹ represents hydrogen or fluoro;

C i -βalky lcarbony lamino ;

NR⁵R⁶;

R⁵ and R⁶ together with the nitrogen atom to which they are attached form a radical of formula

_{with A} representing O, NR⁷, CR⁸R⁹ or S;

R⁷ represents hydrogen, Ci_₆alkyl, C₂-₆alkenyl, Ci_₆alkylcarbonyl,

Ci_₆alkyloxycarbonyl, arylCi-βalkyl or aryl;

R⁸ and R⁹ each independently represent hydrogen, Ci_₆alkyloxy, halo, amino, mono-or di(Ci_₆alkyl)amino, Ci_₆alkyl, Ci_₆alkyloxycarbonylCi_₆alkyl or C₂-₆alkenyl; or

R⁸ and R⁹ together with the carbon atom to which they are attached represent C(=O) or a 5, 6 or 7-membered saturated heterocyclic ring containing 1 or 2 oxygen atoms; n represents an integer of value 1, 2 or 3; p and q each independently represent an integer of value 0, 1, 2 or 3; aryl represents phenyl or phenyl substituted with at least one substituent, in particular one, two or three substituents, each substituent independently selected from halo,

Ci_6alkyl, Ci_6alkyloxy, cyano, nitro, amino or mono-or di(Ci_6alkyl)amino; provided that R³ may only be other than hydrogen if at least one of R¹ or R² is other than hydrogen; a N-oxide thereof, a pharmaceutically acceptable salt thereof or a solvate thereof.

2. The compound according to claim 1 wherein the compound has the following formula

including any stereochemically isomeric form thereof; a JV-oxide thereof, a pharmaceutically acceptable salt thereof or a solvate thereof.

3. The compound according to claim 2 wherein ring E represents a partially saturated or aromatic 5-membered heterocycle wherein the dotted lines represent an optional double bond and wherein B, C and D each independently represent CH₂, CH, N,

NH, S or O provided that the 5-membered ring contains 1 or 2 heteroatoms.

4. The compound according to claim 2 or 3 wherein the compound has the following formula

5. The compound according to claim 2 or 3 wherein the compound has the following formula

6. The compound according to claim 2 or 3 wherein the compound has the following formula

7. The compound according to claim 1 wherein the compound has the following formula

8. The compound according to claim 1 wherein the compound has the following formula

9. The compound according to claim 1, 2 or 3 wherein B, C and D each independently represent CH₂, CH, N, NH or O.

10. The compound according to any one of the preceding claims wherein X represents a direct bond.

11. The compound according to any one of claims 1 to 10 wherein X represents Ci_

4alkanediyl.

12. The compound according to any one of the preceding claims herein R¹ represents hydrogen.

13. The compound according to any one of claims 1 to 11 wherein R¹ represents fluoro.

14. The compound according to any one of the preceding claims wherein R² represents hydrogen, halo, cyano, Ci_₆alkylcarbonyl, Ci_₆alkyloxy or Ci_₆alkylcarbonyl- amino.

15. The compound according to any one of the preceding claims wherein R³ represents hydrogen, halo or Ci_6alkyloxy.

16. The compound according to any one of the preceding claims wherein A is an unsubstituted heterocycle.

17. The compound according to any one of claims 1 to 15 wherein A is a heterocycle substituted with 1 R⁴.

18. The compound according to claim 16 or 17 wherein the heterocycle is pyridyl or thienyl.

19. The compound according to any one of the preceding claims wherein R⁴ represents halo.

20. The compound according to any one of the preceding claims wherein n represents

1.

21. The compound according to any one of claims 1 to 11, 13 to 20 wherein R¹ is fluoro and R² is hydrogen.

22. The compound according to any one of claims 1 to 12, 15 to 20 wherein R¹ is hydrogen and R² is halo, cyano, Ci_₆alkyl, Ci_₆alkyloxy, Ci_₆alkylcarbonyl or Ci_6alkylcarbonylamino.

23. The compound according to any one of claims 1 to 11, 13, 15 to 20 wherein R¹ is fluoro and R² is halo, cyano, Ci_6alkyl, Ci_6alkyloxy, Ci_6alkylcarbonyl or

C i _6alkylcarbonylamino .

24. The compound according to any one of claims 1 to 12, 14 to 20 wherein R¹ and R² are both hydrogen.

25. The compound according to any one of the preceding claims wherein ring E contains 1 or 2 heteroatoms.

26. The compound according to any one of the preceding claims wherein R⁸ and R⁹ each independently represent hydrogen, Ci_₆alkyloxy, halo, amino, mono-or di(Ci_ ₆alkyl)amino, C^alkyl, Ci-ealkyloxycarbonylCi-βalkyl or C₂-₆alkenyl.

27. The compound according to any one of claims 1 to 5 wherein X represents a direct bond or CH₂; R² represents hydrogen, halo, cyano, Ci_6alkylcarbonyl, Ci_6alkyloxy or Ci_6alkylcarbonylamino; R³ represents hydrogen, halo or Ci_6alkyloxy; R⁴ represents halo; n represents 1; A represents optionally substituted pyridyl, or thienyl.

28. The compound according to claim 1 wherein the compound is selected from the group consisting of

4-(2,4-difluoro-phenoxy)-2-pyridin-4-yl-thieno[3,2-d]pyrimidine;

4-(4-chloro-2-fluoro-phenoxy)-2-pyridin-4-yl-thieno[3,2-d]pyrimidine; 4-(2-pyridin-4- yl-thieno[3,2-d]pyrimidin-4-yloxy)-benzonitrile; 2-pyridin-4-yl-4-(2,4,6-trifluoro-phenoxy)-thieno[3,2-d]pyrimidine;

3-fluoro-4-(2-pyridin-4-yl-thieno[3,2-d]pyrimidin-4-yloxy)-benzonitrile;

4-(2-pyridin-3-yl-thieno[3,2-d]pyrimidin-4-yloxy)-benzonitrile;

4-[2-(2-chloro-pyridin-4-yl)-thieno[3,2-d]pyrimidin-4-yloxy]-3-fluoro-benzonitrile; a JV-oxide thereof, a pharmaceutically acceptable salt thereof or a solvate thereof.

29. The compound according to any one of the preceding claims for use as a medicine.

30. The compound according to any one of claims 1 to 28 for use as a medicine for the treatment of a disease by activating the EP4 receptor.

31. The compound according to any one of claims 1 to 28 for the treatment of primary osteoporosis, secondary osteoporosis, bone fracture, metastatic bone disease, rheumatoid arthritis, osteoarthritis, periodontitis, osteogenesis imperfecta, hypercalcemia, a disease associated with liver injury and acute hepatitis, renal failure and nephritis, ulcerative colitis, Crohn's disease, stomatitis, gastritis, ocular hypertension, glaucoma, neuropathic pain, bone pain, Reflex Sympathetic Dystrophy syndrome (RSD) also known as Complex Regional Pain Syndrome (CRPS).

32. A pharmaceutical composition comprising a pharmaceutically acceptable carrier, and as active ingredient a therapeutically effective amount of a compound as claimed in any one of claims 1 to 28.

33. A process of preparing a composition as claimed in claim 32 characterized in that a pharmaceutically acceptable carrier is intimately mixed with a therapeutically effective amount of a compound as claimed in any one of claims 1 to 28.

34. Use of a compound as claimed in any one of claims 1 to 28 for the manufacture of a medicament for treating a disease by activating the EP4 receptor.

35. Use of a compound for the manufacture of a medicament for treating primary osteoporosis, secondary osteoporosis, bone fracture, metastatic bone disease, rheumatoid arthritis, osteoarthritis, periodontitis, osteogenesis imperfecta, hypercalcemia, a disease associated with liver injury and acute hepatitis, renal failure and nephritis, ulcerative colitis, Crohn's disease, stomatitis, gastritis, ocular hypertension, glaucoma, neuropathic pain, bone pain, Reflex Sympathetic Dystrophy syndrome (RSD) also known as Complex Regional Pain Syndrome

(CRPS), wherein the compound is a compound as claimed in any one of claims 1 to 28.

36. A process of preparing a compound as defined in claim 1 characterized by a) reacting an intermediate of formula (II) wherein Wi represents a suitable leaving group with an intermediate of formula (III) in the presence of a suitable base and a suitable solvent,

with the variables as defined in claim 1 ; b) reacting in a first step (a), an intermediate of formula (II-a) with an intermediate of formula P-W₂ wherein P represents a suitable protective group and wherein W₂ represents a suitable leaving group, in the presence of a suitable base and a suitable solvent, followed by reaction (step b) with an intermediate of formula (III) in the presence of a suitable base and a suitable solvent, further followed by deprotection (step c) by reaction with a suitable acid in the presence of a suitable solvent,

(Il-a)

(III)

(b)

(I-l-c) with the variables as defined in claim 1 ;

or, if desired, converting compounds of formula (I) into each other following art- known transformations, and further, if desired, converting the compounds of formula (I), into a therapeutically active non-toxic acid addition salt by treatment with an acid, or into a therapeutically active non-toxic base addition salt by treatment with a base, or conversely, converting the acid addition salt form into the free base by treatment with alkali, or converting the base addition salt into the free acid by treatment with acid; or, if desired, preparing stereochemically isomeric forms, quaternary amines, solvates or JV-oxide forms thereof.