HCV INHIBITING MACROCYCLIC PHOSPHONATES AND AMIDOPHOSPHATES

Technical field This invention relates to macrocyclic compounds having inhibitory activity on the NS3 serine protease of HCV. It further concerns compositions comprising these compounds as active ingredients as well as processes for preparing these compounds and compositions.

Background of the invention

Hepatitis C virus (HCV) is the leading cause of chronic liver disease worldwide and has become a focus of considerable medical research. HCV is a member of the Flaviviridae family of viruses in the hepacivirus genus, and is closely related to the flavivirus genus, which includes a number of viruses implicated in human disease, such as dengue virus and yellow fever virus, and to the animal pestivirus family, which includes bovine viral diarrhea virus (BVDV). The genome of HCV comprises both 5' and 3' untranslated regions that adopt RNA secondary structures, and a central open reading frame that encodes a single polyprotein. The polyprotein encodes ten gene products, which are generated from the precursor polyprotein by an orchestrated series of co- and posttranslational endoproteo lytic cleavages mediated by both host and viral proteases. The viral structural proteins include the core nucleocapsid protein, and two envelope glycoproteins El and E2. The non- structural (NS) proteins encode some essential viral enzymatic functions (helicase, polymerase, protease), as well as proteins of unknown function. Replication of the viral genome is mediated by an RNA- dependent RNA polymerase, encoded by non-structural protein 5b (NS5B). In addition to the polymerase, the viral helicase and protease functions, both encoded in the bifunctional NS3 protein, have been shown to be essential for replication of HCV RNA. In addition to the NS3 serine protease, HCV also encodes a metalloproteinase in the NS2 region.

Following the initial acute infection, a majority of infected individuals develop chronic hepatitis because HCV replicates preferentially in hepatocytes but is not directly cytopathic. In particular, the lack of a vigorous T-lymphocyte response and the high propensity of the virus to mutate appear to promote a high rate of chronic infection. Chronic hepatitis can progress to liver fibrosis leading to cirrhosis, end- stage liver disease, and HCC (hepatocellular carcinoma), making it the leading cause of liver transplantations .

There are 6 major HCV genotypes and more than 50 subtypes, which are differently distributed geographically. HCV type 1 is the predominant genotype in Europe and the US. The extensive genetic heterogeneity of HCV has important diagnostic and clinical implications, perhaps explaining difficulties in vaccine development and the lack of response to current therapy.

Transmission of HCV can occur through contact with contaminated blood or blood products, for example following blood transfusion or intravenous drug use. The introduction of diagnostic tests used in blood screening has led to a downward trend in post-transfusion HCV incidence. However, given the slow progression to the end-stage liver disease, the existing infections will continue to present a serious medical and economic burden for decades.

Current HCV therapies are based on (pegylated) interferon-alpha (IFN-α) in combination with ribavirin. This combination therapy yields a sustained viro logic response in more than 40% of patients infected by genotype 1 viruses and about 80% of those infected by genotypes 2 and 3. Beside the limited efficacy on HCV type 1, this combination therapy has significant side effects and is poorly tolerated in many patients. Major side effects include influenza- like symptoms, hematologic abnormalities, and neuropsychiatric symptoms. Hence there is a need for more effective, convenient and better-tolerated treatments.

A number of similar HCV protease inhibitors have been disclosed in the academic and patent literature. The sustained administration of HCV protease inhibitors usually leads to the selection of resistant HCV mutants, so called drug escape mutants. These have characteristic mutations in the HCV protease genome, notably D 168V, D 168 Y and/or A 165 S. Accordingly, there is a need for additional drugs with different resistance patterns to provide failing patients with treatment options. Such drugs may find use in combination therapy, which is expected to become the norm in the future, even for first line treatment.

Experience with HIV drugs, in particular with HIV protease inhibitors, has taught that sub-optimal pharmacokinetics and complex dosing regimes quickly result in inadvertent compliance failures. This in turn means that the 24 hour trough concentration (minimum plasma concentration) for the respective drugs in an HIV regime frequently falls below the IC90 or ED90 threshold for large parts of the day. It is considered that a 24 hour trough level of at least the IC ₅₀ , and more realistically, the IC90 or ED ₉₀ , is essential to slow down the development of drug escape mutants.

Achieving the necessary pharmacokinetics and drug metabolism to allow such trough levels provides a stringent challenge to drug design. Known HCV protease inhibitors, with multiple peptide bonds, pose additional pharmacokinetic hurdles to effective dosage regimes.

There is a need for HCV inhibitors that may overcome the disadvantages of current HCV therapy such as side effects, limited efficacy, the emerging of resistance, and compliance failures.

The present invention concerns inhibitors of HCV replication that exhibit at least one improved property in view of the compounds of the prior art compounds. In particular, the inhibitors of the present invention are superior in one or more of the following pharmacological related properties, i.e. potency, decreased cytotoxicity, improved pharmacokinetics, improved resistance profile, acceptable dosage and pill burden.

Brief description of the invention

The present invention concerns inhibitors of HCV replication, which can be represented by formula (I):

including the stereochemical^ isomeric forms thereof, wherein each dashed line (represented by ) represents an optional double bond;

X is N, CH and where X bears a double bond it is C;

R ¹ is aryl or a saturated, a partially unsaturated or completely unsaturated 5 or 6 membered monocyclic or 9 to 12 membered bicyclic heterocyclic ring system wherein said ring system contains one nitrogen, and optionally one to three additional heteroatoms selected from the group consisting of oxygen, sulfur and nitrogen, and wherein the remaining ring members are carbon atoms; wherein said ring system may be optionally substituted on any carbon or nitrogen ring atom with one, two, three, or four substituents each independently selected from C3-7cyclo- alkyl, aryl, Het, -C(=O)-NR ^4a R ^4b , -C(=O)R ⁶ , -C(=O)OR ^5a , and Chalky! optionally

substituted with C ₃ - ₇ cycloalkyl, aryl, Het, -C(=O)NR ^4a R ^4b , -NR ^4a R ^4b , -C(=O)R ⁶ , -NR ^4a C(=O)R ⁶ , -NR ^4a SO _p R ⁷ , -SO _P R ⁷ , -SO _p NR ^4a R ^4b , -C(=O)OR ⁵ , or -NR ^4a C(=O)OR ^5a ; and wherein the substituents on any carbon atom of the heterocyclic ring may also be selected from -OR ⁷ , -SR ⁷ , halo, polyhalo-Ci_6alkyl, oxo, thio, cyano, nitro, azido, -NR ^4a R ^4b , -NR ^4a C(=O)R ⁶ , -NR ^4a SO ₂ R ⁷ , -SO ₂ R ⁷ , -SO ₂ NR ^4a R ^4b , -C(=O)OH, and -NR ^4a C(=O)OR ^5a ; or

R ¹ is a group of formula

R ² is -C(=O)-NH-P(=O)(OR ^8a )(R ^8b ), or -P(=O)(OR ^8a )(R ^8b ); R ³ is hydrogen, and where X is C or CH, R ³ may also be Ci_ ₆ alkyl; n is 3, 4, 5, or 6; p is 1 or 2; each R ^4a and R ^4b are, independently, hydrogen, C _3-7 Cy cloalkyl, aryl, Het, Ci_ ₆ alkyl optionally substituted with halo, Ci_4alkoxy, cyano, polyhaloCi_4alkoxy, C ₃ _ ₇ cycloalkyl, aryl, or with Het; or R ^4a and R ^4b taken together with the nitrogen atom to which they are attached form pyrrolidinyl, piperidinyl, piperazinyl, 4-Ci_ ₆ alkylpiperazinyl, 4-Ci_ ₆ alkylcarbonyl-piperazinyl, and morpholinyl; wherein the morpholinyl and piperidinyl groups may be optionally substituted with one or with two Ci_ ₆ alkyl radicals; R ⁵ is hydrogen; C ₂ _ ₆ alkenyl; Het; C ₃ _ ₇ cycloalkyl optionally substituted with Ci_ ₆ alkyl; or Ci_ ₆ alkyl optionally substituted with C ₃ _ ₇ cycloalkyl, aryl or Het; R ^5a is C ₂ _ ₆ alkenyl, C ₃ _ ₇ cycloalkyl, Het, or Ci_ ₆ alkyl optionally substituted with

C ₃ - ₇ cycloalkyl, aryl or with Het; R ⁶ is hydrogen, Ci_ ₆ alkyl, C ₃ _ ₇ cycloalkyl, or aryl; R ⁷ is hydrogen; polyhaloCi-βalkyl; aryl; Het; C ₃ - ₇ cycloalkyl optionally substituted with

Ci_ ₆ alkyl; or Ci_ ₆ alkyl optionally substituted with C ₃ _ ₇ cycloalkyl, aryl or Het; R ^8a is hydrogen, C^aUcyl, C ₂ _ ₆ alkenyl, C ₃ - ₇ cycloalkyl, aryl, or Ci_ ₆ alkyl optionally substituted with C ₃ _ ₇ cycloalkyl or aryl; R ^8b is R ^8b' , OR ^8b' or NHR ^8b' ; R ^8b is Ci_ ₆ alkyl, C ₂ _ ₆ alkenyl, C ₃ _ ₇ cycloalkyl, aryl, or Ci_ ₆ alkyl optionally substituted with C ₃ _ ₇ cycloalkyl or with aryl; E is NR ⁹ or when X is N then E is NR ⁹ or CR ^10a R ^10b ;

R ⁹ is hydrogen, C _h alky!, Ci_ ₆ alkoxyCi_ ₆ alkyl, or C ₃ - ₇ cycloalkyl; R ^1Oa and R ^1Ob are independently hydrogen or Ci_ ₆ alkyl, or R ^1Oa and I the carbon atom to which they are attached form C ₃ _ ₇ cycloalkyl;

R ¹¹ is hydrogen; aryl; Het; C ₃ - ₇ cycloalkyl; Ci_ ₆ alkyl optionally substituted with C ₃ _ ₇ Cycloalkyl, aryl or with Het; halo; polyhaloCi-βalkyl; hydroxy; Ci_ ₆ alkoxy; polyhaloCi-βalkoxy; Ci_6alkoxyCi_6alkyl; carboxyl; Ci_6alkylcarbonyl; Ci_6alkoxy- carbonyl; cyano; nitro; amino; mono- or diCi_6alkylamino; amino-carbonyl; mono- or diCi_ ₆ alkylaminocarbonyl;

R ¹² is hydrogen; Ci_6alkyl; halo; polyhaloCi-βalkyl; hydroxy; Ci_6alkoxy; polyhalo- Ci_ ₆ alkoxy; Ci_ ₆ alkoxyCi_ ₆ alkyl; carboxyl; Ci_ ₆ alkylcarbonyl; Ci_ ₆ alkoxycarbonyl; cyano; nitro; amino; mono- or diCi_6alkylamino; amino-carbonyl; mono- or diC i _ ₆ alkylaminocarbonyl; R ¹³ is hydrogen; Ci_6alkyl; halo; polyhaloCi-βalkyl; polyhaloCi-βalkoxy; hydroxy;

Ci_6alkoxy; cyano; aryl as a group or part of a group is phenyl, naphthyl, indanyl, or 1,2,3,4-tetrahydro- naphthyl, each of which may be optionally substituted with one, two or three substituents selected from halo, Ci_6alkyl, polyhaloCi-βalkyl, hydroxy, Ci_6alkoxy, polyhaloCi-βalkoxy, Ci_6alkoxyCi_6alkyl, carboxyl, Ci_6alkylcarbonyl, Ci_6alkoxy- carbonyl, cyano, nitro, amino, mono- or diCi_6alkylamino, amino-carbonyl, mono- or diCi_ ₆ alkylaminocarbonyl, azido, mercapto, C ₃ - ₇ cycloalkyl, phenyl, pyridyl, thiazolyl, pyrazolyl, pyrrolidinyl, piperidinyl, piperazinyl, 4-Ci_ ₆ alkylpiperazinyl, 4-Ci_ ₆ alkylcarbonyl-piperazinyl, and morpholinyl; wherein the morpholinyl and piperidinyl groups may be optionally substituted with one or with two Ci_ ₆ alkyl radicals; and the phenyl, pyridyl, thiazolyl, pyrazolyl groups may be optionally substituted with 1, 2 or 3 substituents each independently selected from Ci_6alkyl, Ci_6alkoxy, halo, amino, mono- or diCi_6alkylamino; Het as a group or part of a group is a 5 or 6 membered saturated, partially unsaturated or completely unsaturated heterocyclic ring containing 1 to 4 heteroatoms each independently selected from nitrogen, oxygen and sulfur, said heterocyclic ring being optionally condensed with a benzene ring, and wherein the group Het as a whole may be optionally substituted with one, two or three substituents each independently selected from the group consisting of halo, Chalky!, polyhalo- Ci_6alkyl, hydroxy, Ci_6alkoxy, polyhaloCi-βalkoxy, Ci_6alkoxyCi_6alkyl, carboxyl,

Ci_ ₆ alkylcarbonyl, Ci_ ₆ alkoxycarbonyl, cyano, nitro, amino, mono- or diCi_ ₆ alkyl- amino, aminocarbonyl, mono- or diCi_ ₆ alkylaminocarbonyl, C ₃ _ ₇ Cycloalkyl, phenyl, pyridyl, thiazolyl, pyrazolyl, pyrrolidinyl, piperidinyl, piperazinyl, 4-Ci_ ₆ alkylpiperazinyl, 4-Ci_ ₆ alkylcarbonyl-piperazinyl, and morpholinyl; wherein the morpholinyl and piperidinyl groups may be optionally substituted with one or with two Ci_6alkyl radicals; and the phenyl, pyridyl, thiazolyl, pyrazolyl groups may be optionally substituted with 1 , 2 or 3 substituents each independently selected from Ci_6alkyl, Ci_6alkoxy, halo, amino, mono- or diCi_6alkylamino;

or the iV-oxides, pharmaceutically acceptable addition salts, or pharmaceutically acceptable solvates thereof.

The invention relates to the compounds of formula (I) per se, and the iV-oxides, pharmaceutically acceptable addition salts, and stereochemically isomeric forms thereof, for use as a medicament. The invention further relates to pharmaceutical compositions comprising the aforementioned compounds for administration to a subject suffering from HCV infection. The pharmaceutical compositions may comprise combinations of the aforementioned compounds with other anti-HCV agents.

The invention also relates to the use of a compound of formula (I), an JV-oxide, a pharmaceutically acceptable addition salt, or stereochemically isomeric form thereof, for the manufacture of a medicament for inhibiting HCV replication. Or the invention relates to a method of inhibiting HCV replication in a warm-blooded animal, said method comprising the administration of an effective amount of a compound of formula (I), an JV-oxide, a pharmaceutically acceptable addition salt, or stereochemically isomeric form thereof.

Detailed description of the invention As used in the foregoing and hereinafter, the following definitions apply unless otherwise noted.

The term halo is generic to fluoro, chloro, bromo and iodo.

The term "polyhaloCi-βalkyl" as a group or part of a group, e.g. in polyhaloCi-βalkoxy, is defined as mono- or polyhalo substituted Ci_6alkyl, in particular Ci_6alkyl substituted with up to one, two, three, four, five, six, or more halo atoms, such as methyl or ethyl with one or more fluoro atoms, for example, difluoromethyl, trifluoromethyl, trifluoro- ethyl. Preferred is trifluoromethyl. Also included are perfluoroCi-βalkyl groups, which are Ci_6alkyl groups wherein all hydrogen atoms are replaced by fluoro atoms, e.g. pentafluoroethyl. In case more than one halogen atom is attached to an alkyl group within the definition of polyhaloCi-βalkyl, the halogen atoms may be the same or different.

As used herein "Ci_ ₄ alkyl" 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 for example methyl, ethyl, 1 -propyl, 2-propyl, 1 -butyl, 2-butyl, 2-methyl-l -propyl; "Ci_ ₆ alkyl" encompasses Ci_ ₄ alkyl radicals and the higher homologues thereof having 5 or 6 carbon atoms such as, for example, 1-pentyl, 2-pentyl, 3-pentyl, 1-hexyl, 2-hexyl,

2-methyl-l -butyl, 2-methyl-l-pentyl, 2-ethyl-l -butyl, 3-methyl-2-pentyl, and the like. Of interest amongst Ci_ ₆ alkyl is Ci_ ₄ alkyl.

The term "C ₂ - ₆ alkenyl" as a group or part of a group defines straight and branched chained hydrocarbon radicals having saturated carbon-carbon bonds and at least one double bond, and having from 2 to 6 carbon atoms, such as, for example, ethenyl (or vinyl), 1-propenyl, 2-propenyl (or allyl), 1-butenyl, 2-butenyl, 3-butenyl, 2-methyl-2-propenyl, 2-pentenyl, 3-pentenyl, 2-hexenyl, 3-hexenyl, 4-hexenyl, 2-methyl-2-butenyl, 2-methyl-2-pentenyl and the like. Of interest amongst C ₂ - ₆ alkenyl is C ₂ - ₄ alkenyl.

The term "C2-6alkynyl" as a group or part of a group defines straight and branched chained hydrocarbon radicals having saturated carbon-carbon bonds and at least one triple bond, and having from 2 to 6 carbon atoms, such as, for example, ethynyl, 1-propynyl, 2-propynyl, 1-butynyl, 2-butynyl, 3-butynyl, 2-pentynyl, 3-pentynyl, 2-hexynyl, 3-hexynyl and the like. Of interest amongst C ₂ -βalkynyl is C ₂ - ₄ alkynyl.

C _{3 y} Cycloalkyl is generic to cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and cycloheptyl. C _{3 7} cycloalkyl when substituted on aryl or Het in particular is cyclopropyl.

C _{1 6} alkanediyl defines bivalent straight and branched chain saturated hydrocarbon radicals having from 1 to 6 carbon atoms such as, for example, methylene, ethylene, 1,3-propanediyl, 1 ,4-butanediyl, 1 ,2-propanediyl, 2,3-butanediyl, 1,5-pentanediyl, 1,6-hexanediyl and the like. Of interest amongst C _{1 6} alkanediyl is C _{1 4} alkanediyl.

Ci_6alkoxy means Ci_6alkyloxy wherein Ci_6alkyl is as defined above and is bonded to an oxygen atom, i.e. -O-Ci_ ₆ alkyl. Of interest amongst Ci-βalkoxy are methoxy, ethoxy and propoxy.

As used herein before, the term (=0) or oxo forms a carbonyl moiety when attached to a carbon atom, a sulfoxide moiety when attached to a sulfur atom and a sulfonyl moiety when two of said terms are attached to a sulfur atom. Whenever a ring or ring system is substituted with an oxo group, the carbon atom to which the oxo is linked is a saturated carbon.

The radical Het is a heterocycle as specified in this specification and claims. Examples of Het comprise, for example, pyrrolidinyl, piperidinyl, morpholinyl, piperazinyl, pyrrolyl, pyrazolyl, imidazolyl, oxazolyl, isoxazolyl, thiazinolyl, isothiazinolyl, thiazolyl, isothiazolyl, oxadiazolyl, thiadiazolyl, triazolyl (including 1,2,3-triazolyl,

1,2,4-triazolyl), tetrazolyl, furanyl, thienyl, pyridyl, pyrimidyl, pyridazinyl, pyrazolyl-triazinyl, or any of such heterocycles condensed with a benzene ring, such as indolyl, indazolyl (in particular lH-indazolyl), indolinyl, quinolinyl, tetrahydro- quinolinyl (in particular 1,2,3,4-tetrahydroquinolinyl), isoquinolinyl, tetrahydroiso- quinolinyl (in particular 1,2,3,4-tetrahydroisoquinolinyl), quinazolinyl, quinoxalinyl, cinnolinyl, phthalazinyl, benzimidazolyl, benzoxazolyl, benzisoxazolyl, benzothiazinolyl, benzisothiazinolyl, benzothiazolyl, benzoxadiazolyl, benzothia- diazolyl, benzo-l,2,3-triazolyl, benzo-l,2,4-triazolyl, benzotetrazolyl, benzo furanyl, benzothienyl, benzopyrazolyl, and the like. Of interest amongst the Het radicals are those which are non-saturated, in particular those having an aromatic character. Of further interest are those Het radicals that are monocyclic. Of further interest are those Het radicals having one or two nitrogens.

Each of the Het or R ¹ radicals mentioned in the previous and the following paragraph may be optionally substituted with the number and kind of substituents mentioned in the definitions of the compounds of formula (I) or any of the subgroups of compounds of formula (I). Some of the Het or R ¹ radicals mentioned in this and the following paragraph may be substituted with one, two or three hydroxy substituents. Such hydroxy substituted rings may occur as their tautomeric forms bearing keto groups. For example a 3-hydroxypyridazine moiety can occur in its tautomeric form

2H-pyridazin-3-one. Some examples keto-substituted ηet or R ¹ radicals are 1 ,3-dihydro-benzimidazol-2-one, 1 ,3-dihydro-indol-2-one, lH-indole-2,3-dione, lH-benzo[<i]isoxazole, lH-benzo[<i]isothiazole, lH-quinolin-2-one, lH-quinolin-4-one, lH-quinazolin-4-one, 9H-carbazole, and lH-quinazolin-2-one. Where ηet is piperazinyl, it preferably is substituted in its 4-position by a substituent linked to the 4-nitrogen with a carbon atom, e.g. 4-Ci_ ₆ alkyl, 4-polyhaloCi_ ₆ alkyl, Ci_ ₆ alkoxy- Ci_ ₆ alkyl, Ci_ ₆ alkylcarbonyl, C ₃ _ ₇ Cycloalkyl.

R ¹ can be a saturated, a partially unsaturated or completely unsaturated 5 or 6 membered monocyclic or 9 to 12 membered bicyclic heterocyclic ring system as specified in this specification and claims. Examples of said monocyclic or bicyclic ring system comprise for example, any of the rings mentioned in the previous paragraph as examples of the radical ηet and additionally any of the monocyclic heterocycles mentioned in the previous paragraph condensed with pyridyl or pyrimidinyl such as, for example, pyrrolopyridine (in particular lH-pyrrolo[2,3-δ]pyridine, lH-pyrrolo[2, 3-c]- pyridine), naphtyridine (in particular 1,8-naphtyridine), imidazopyridine (in particular lH-imidazo[4, 5-c]pyridine, lH-imidazo[4,5-δ]pyridine), pyridopyrimidine, purine (in particular 7H-purine) and the like.

Interesting Het or R ¹ radicals comprise, for example pyrrolidinyl, piperidinyl, morpholinyl, piperazinyl, pyrrolyl, pyrazolyl, imidazolyl, oxazolyl, isoxazolyl, thiazolyl, isothiazolyl, oxadiazolyl, thiadiazolyl, triazolyl (including 1,2,3-triazolyl, 1,2,4-triazolyl), tetrazolyl, furanyl, thienyl, pyridyl, pyrimidyl, pyridazinyl, pyrazolyl, triazinyl, or any of such heterocycles condensed with a benzene ring, such as indolyl, indazolyl (in particular lH-indazolyl), indolinyl, quinolinyl, tetrahydroquinolinyl (in particular 1,2,3,4-tetrahydroquinolinyl), isoquinolinyl, tetrahydroisoquinolinyl (in particular 1,2,3,4-tetrahydroisoquinolinyl), quinazolinyl, phthalazinyl, benzimidazolyl, benzoxazolyl, benzisoxazolyl, benzothiazolyl, benzoxadiazolyl, benzo thiadiazolyl, benzofuranyl, benzothienyl.

Where Het is pyrrolidinyl, piperidinyl, morpholinyl, piperazinyl, 4-substituted piperazinyl, these radicals preferably are linked via their nitrogen atom (i.e. 1 -pyrrolidinyl, 1 -piperidinyl, 4-morpholinyl, 1 -piperazinyl, 4-substituted piperazin-1-yl).

Each "aryl" is as specified above and preferably is phenyl substituted with the substituents specified above. This applies equally to arylCi-βalkyl, which in particular can be arylmethyl, e.g. benzyl.

It should be noted that the radical positions on any molecular moiety used in the definitions may be anywhere on such moiety as long as it is chemically stable.

Radicals used in the definitions of the variables include all possible isomers unless otherwise indicated. For instance pyridyl includes 2-pyridyl, 3 -pyridyl and 4-pyridyl; pentyl includes 1-pentyl, 2-pentyl and 3-pentyl.

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

Whenever used hereinafter, the term "compounds of formula (I)", or "the present compounds" or similar terms, it is meant to include the compounds of formula (I), their iV-oxides, pharmaceutically acceptable addition salts, and stereochemical^ isomeric forms. One embodiment comprises the compounds of formula (I) or any subgroup of compounds of formula (I) specified herein, and the pharmaceutically acceptable addition salts and the possible stereoisomeric forms thereof.

The compounds of formula (I) have several centers of chirality and exist as stereo chemically isomeric forms. The term "stereochemically isomeric forms" as used herein defines all the possible compounds made up of the same atoms bound by the same sequence of bonds but having different three-dimensional structures which are not interchangeable, which the compounds of formula (I) may possess.

With reference to the instances where (R) or (S) is used to designate the absolute configuration of a chiral atom within a substituent, the designation is done taking into consideration the whole compound and not the substituent in isolation.

Unless otherwise mentioned or indicated, the chemical designation of a compound encompasses the mixture of all possible stereochemically isomeric forms, which said compound may possess. Said mixture may contain all diastereomers and/or enantiomers of the basic molecular structure of said compound. All stereochemically isomeric forms of the compounds of the present invention both in pure form or mixed with each other are intended to be embraced within the scope of the present invention.

Pure stereoisomeric forms of the compounds and intermediates as mentioned herein are defined as isomers substantially free of other enantiomeric or diastereomeric forms of the same basic molecular structure of said compounds or intermediates. In particular, the term "stereoisomerically pure" concerns compounds or intermediates having a stereoisomeric excess of at least 80% (i.e. minimum 90% of one isomer and maximum 10% of the other possible isomers) up to a stereoisomeric excess of 100% (i.e. 100% of one isomer and none of the other), more in particular, compounds or intermediates having a stereoisomeric excess of 90% up to 100%, even more in particular having a stereoisomeric excess of 94% up to 100% and most in particular having a stereoisomeric excess of 97% up to 100%. The terms "enantiomerically pure" and "diastereomerically pure" should be understood in a similar way, but then having regard to the enantiomeric excess, and the diastereomeric excess, respectively, of the mixture in question.

Pure stereoisomeric forms of the compounds and intermediates of this invention may be obtained by the application of art-known procedures. For instance, enantiomers may be separated from each other by the selective crystallization of their diastereomeric salts with optically active acids or bases. Examples thereof are tartaric acid, dibenzoyl-tartaric acid, ditoluoyltartaric acid and camphosulfonic acid. Alternatively, enantiomers may be separated by chromatographic techniques using chiral stationary phases. 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 is synthesized by stereospecific methods of preparation. These methods will advantageously employ enantiomerically pure starting materials.

The diastereomeric racemates of the compounds of formula (I) can be obtained separately by conventional methods. Appropriate physical separation methods that may advantageously be employed are, for example, selective crystallization and chromatography, e.g. column chromatography.

For some of the compounds of formula (I), the iV-oxides, the pharmaceutically acceptable addition salts, and solvates thereof, and the intermediates used in the preparation thereof, the absolute stereochemical configuration was not experimentally determined. A person skilled in the art is able to determine the absolute configuration of such compounds using art-known methods such as, for example, X-ray diffraction.

The present invention is also intended to include all isotopes of atoms occurring on the present compounds. Isotopes include those atoms having the same atomic number but different mass numbers. By way of general example and without limitation, isotopes of hydrogen include tritium and deuterium. Isotopes of carbon include C- 13 and C- 14.

The pharmaceutically acceptable addition salts comprise the therapeutically active non-toxic acid and base addition salt forms of the compounds of formula (I). The pharmaceutically acceptable acid addition salts can conveniently be obtained by treating the base form with such appropriate acid. Appropriate acids comprise, for example, inorganic acids such as hydrohalic acids, e.g. hydrochloric or hydrobromic acid, sulfuric, nitric, phosphoric and the like acids; or organic acids such as, for example, acetic, propanoic, hydroxyacetic, lactic, pyruvic, oxalic (i.e. ethanedioic), malonic, succinic (i.e. butanedioic acid), maleic, fumaric, malic (i.e. hydroxyl- butanedioic acid), tartaric, citric, methanesulfonic, ethanesulfonic, benzenesulfonic, /?-toluenesulfonic, cyclamic, salicylic, /^-aminosalicylic, pamoic and the like acids. Conversely said salt forms can be converted by treatment with an appropriate base into the free base form.

The compounds of formula (I) containing an acidic proton may also be converted into their non-toxic metal or amine addition salt forms by treatment with appropriate organic and inorganic bases. Appropriate base 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. the

benzathine, JV-methyl-D-glucamine, hydrabamine salts, and salts with amino acids such as, for example, arginine, lysine and the like.

The term addition salts also is meant to comprise the solvates, which the compounds of formula (I) as well as the salts thereof, are able to form. Such solvates are for example hydrates, alcoholates, e.g. ethanolates, propanolates, and the like.

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

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

As mentioned above, the compounds of formula (I) have several asymmetric centers. In order to more efficiently refer to each of these asymmetric centers, the numbering system as indicated in the following structural formula will be used.

Asymmetric centers are present at positions 1, 4 and 6 of the macrocycle as well as at the carbon atom 3' in the 5-membered ring, at carbon atom 2' when the R substituent is Ci-βalkyl, and at carbon atom 1 ' when X is CH. Each of these asymmetric centers can occur in their R or S configuration.

When X is N, the stereochemistry at position 1 preferably corresponds to that of an L-amino acid configuration, i.e. that of L-proline as shown below.

When X is CH, the 2 carbonyl groups substituted at positions 1 ' and 5' of the cyclopentane ring preferably are in a trans configuration. The carbonyl substituent at position 5 ' preferably is in that configuration that corresponds to an L-proline configuration.

The carbonyl groups substituted at positions 1 ' and 5 ' preferably are as depicted below in the structure of the following formula (I-a2).

The compounds of formula (I) include a cyclopropyl group as represented in the structural fragment below:

wherein C ₇ represents the carbon at position 7 and carbons at position 4 and 6 are asymmetric carbon atoms of the cyclopropane ring. The presence of these two asymmetric centers means that the compounds can exist as mixtures of diastereomers,

such as the diastereomers of compounds of formula (I) wherein the carbon at position 7 is configured either cis to the carbonyl or cis to the amide as shown below.

C7 cis to carbonyl C7 cis to amide

One embodiment concerns compounds of formula I wherein the carbon at position 7 is configured cis to the carbonyl. Another embodiment concerns compounds of formula (I) wherein the configuration at the carbon at position 4 is R. A specific subgroup of compounds of formula (I) is that wherein the carbon at position 7 is configured cis to the carbonyl and wherein the configuration at the carbon at position 4 is R.

According one embodiment the cyclopropyl group (C4-C5-C6) is linked to a group R ² that is a phosphonate group -P(=O)(OR ^8a )(R ^8b ). According to this embodiment, the carbon at position 7 is configured in a cis relationship either to the phosphonate or to the amide as presented in the structural fragment below:

C7 cis to phosphonate

C7 cis to amide

C7 cis to phosphonate C7 cis to amide

One embodiment concerns compounds of formula (I) wherein the carbon at position 7 is configured cis to the phosphonate. Another embodiment concerns compounds of formula (I) wherein the configuration at the carbon at position 4 is S. A specific subgroup of compounds of formula (I) are those wherein the carbon at position 7 is configures cis to the phosphonate and wherein the configuration at the carbon at position 4 is S.

The compounds of formula (I) may include a proline residue (when X is N) or a cyclopentyl or cyclopentenyl residue (when X is CH or C). Preferred are the compounds of formula (I) wherein the substituent at the 1 (or 5') position and the substituent -O-R ¹ (at position 3') are in a trans configuration. Of particular interest are the compounds of formula (I) wherein position 1 has the configuration corresponding to L-proline and the -O-R ¹ substituent is in a trans configuration in respect of position 1. Preferably the compounds of formula (I) have the stereochemistry as indicated in the structure of formula (I-b) as depicted below:

One embodiment of the present invention concerns compounds of formula (I) or of formulae (I-al), (I-a2), (I-b), or of any subgroup of compounds of formula (I), wherein one or more of the following conditions apply: (a) R ³ is hydrogen;

(b) X is nitrogen;

(c) E is NR ⁵ ;

(d) a double bond is present between carbon atoms 7 and 8.

One embodiment of the present invention concerns compounds of formula (I) or of formulae (I-a), (I-b), or of any subgroup of compounds of formula (I), wherein one or more of the following conditions apply: (a) R ³ is hydrogen;

(b) X is CH;

(c) E is NR ³ ;

(d) a double bond is present between carbon atoms 7 and 8.

A further embodiment of the present invention concerns compounds of formula (I) or of formulae (I-al), (I-a2), (I-b), or of any subgroup of compounds of formula (I), wherein one or more of the following conditions apply:

(a) R is hydrogen;

(b) X is CH;

(c) E is NR ⁵ , wherein R ⁵ is as defined above, particularly R ⁵ is hydrogen or Ci_ ₆ alkyl;

(d) a double bond is present between carbon atoms 7 and 8.

Particular subgroups of compounds of formula (I) are those represented by the following structural formulas:

(I-c) (I-d)

Amongst the compounds of formula (I-c) or (I-d), those having the stereochemical configuration of the compounds of formulae (I-al), (I-a2), and (I-b) are of particular interest.

The double bond between carbon atoms 7 and 8 in the compounds of formula (I), or in any subgroup of compounds of formula (I), may be in a cis or in a trans configuration. Preferably the double bond between carbon atoms 7 and 8 is in a cis configuration, as depicted in formulae (I-c) and (I-d).

A double bond between carbon atoms 1 ' and 2' may be present in the compounds of formula (I), or in any subgroup of compounds of formula (I), as depicted in formula (I-e) below.

Yet another particular subgroup of compounds of formula (I) are those represented by the following structural formulae:

(I-f) (i-g) (I-h)

Amongst the compounds of formulae (I-f), (I-g) or (I-h), those having the stereochemical configuration of the compounds of formulae (I-al), (I-a2), and (I-b) are of particular interest.

In (I-al), (I-a2), (I-b), (I-c), (I-d), (I-e), (I-f), (I-g) and (I-h), where applicable, E, X, n, R ¹ , R ² , and R ³ are as specified in the definitions of the compounds of formula (I) or in any of the subgroups of compounds of formula (I) specified herein.

It is to be understood that the above defined subgroups of compounds of formulae (I-a), (I-b), (I-c), (I-d) or (I-e), as well as any other subgroup defined herein, are meant to also comprise any JV-oxides, addition salts, and stereochemically isomeric forms of such compounds.

When n is 2, the moiety -CH ₂ - bracketed by "n" corresponds to ethanediyl in the compounds of formula (I) or in any subgroup of compounds of formula (I). When n is 3, the moiety -CH ₂ - bracketed by "n" corresponds to propanediyl in the compounds of formula (I) or in any subgroup of compounds of formula (I). When n is 4, the moiety

-CH ₂ - bracketed by "n" corresponds to butanediyl in the compounds of formula (I) or in any subgroup of compounds of formula (I). When n is 5, the moiety -CH ₂ - bracketed by "n" corresponds to pentanediyl in the compounds of formula (I) or in any subgroup of compounds of formula (I). When n is 6, the moiety -CH ₂ - bracketed by "n" corresponds to hexanediyl in the compounds of formula (I) or in any subgroup of compounds of formula (I). Particular subgroups of the compounds of formula (I) are those compounds wherein n is 4 or 5.

One embodiment of the invention concerns compounds of formula (I), or any of the subgroups of compounds of formula (I), wherein R ⁴ is -C(=O)-NH-P(=O)(OR ^8a )(R ^8b ); in particular wherein R ^4a is Ci_ ₆ alkyl, especially ethyl or isopropyl and R ^4b is OR ^4b and R ^4b is d-βalkyl, such as ethyl or isopropyl.

A further embodiment of the invention are compounds of formula (I), or any of the subgroups of compounds of formula (I), wherein R ⁴ is -P(=O)(OR ^8a )(R ^8b ); in particular wherein R ^4a is d-βalkyl, especially ethyl or isopropyl and R ^4b is OR ^4b and R ^4b is Ci_ ₆ alkyl, especially ethyl or isopropyl.

Further embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein

(a) R ⁹ is hydrogen;

(b) R ⁹ is Ci_ ₆ alkyl;

(c) R ⁹ is Ci_6alkoxyCi_6alkyl or C _3-7 Cy cloalkyl.

Preferred embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ⁹ is hydrogen, or d-βalkyl, more preferably hydrogen or methyl.

Still a further embodiment concerns compounds of formula (I), (I-e) or any subgroup of compounds of formula (I) wherein R ^1Oa and R ^10b independently are hydrogen or Ci_ ₆ alkyl, e.g. methyl. Preferably R ^1Oa is hydrogen and R ^10b is methyl, or more preferably R ^1Oa and R ^10b are both hydrogen.

Further subgroups of the compounds of formula (I) are those compounds of formula (I), or any subgroup of compounds of formula (I) specified herein, wherein R ¹ is phenyl, naphthyl, pyridyl, pyridazinyl, triazolyl, tetrazolyl, quinolinyl, isoquinolinyl, quinazolinyl, pyrimidinyl, [l,8]naphthyridinyl, indolinyl, 1,2,3,4-tetrahydroquinolinyl, 1,2,3,4-tetrahydroisoquinolinyl; all optionally substituted with one, two or three

substituents selected from those mentioned in relation to R ¹ in the definitions of the compounds of formula (I) or any of the subgroups of compounds of formula (I).

Other subgroups of the compounds of formula (I) are those compounds of formula (I), or any subgroup of compounds of formula (I) specified herein, wherein

(a) R ¹ is phenyl, naphtyl (such as naphth-1-yl, or naphth-2-yl), quinolinyl (in particular quinolin-4-yl), isoquinolinyl (in particular isoquinolin-1-yl), quinazolinyl (in particular quinazolin-4-yl), pyridyl (in particular 3-pyridyl), pyrimidinyl (in particular pyrimidin-4-yl), pyridazinyl (in particular pyridazin-3-yl and pyridazin-2-yl), [l,8]naphtyridinyl (in particular [ 1 ,8]naphthyridin-4-yl);

(b) R ¹ is triazolyl (in particular triazol-1-yl, triazol-2-yl), tetrazolyl (in particular tetrazol-1-yl, tetrazol-2-yl), 6-oxo-pyridazin-l-yl, pyrazolyl (in particular pyrazol- 1 -yl), or imidazolyl (in particular imidazol- 1 -yl, imidazol-2-yl);

(c) R ¹ is a heterocycle selected from

wherein each of the above mentioned R ¹ radicals in (a), (b) or (c) may be optionally substituted with one, two or three substituents selected from those mentioned in relation to R ¹ in the definitions of the compounds of formula (I) or any of the subgroups of compounds of formula (I); or

(d) R ¹ is a group

wherein one or more of R , 11 , R , 12 and R , 13 are as specified above; or wherein in (a-l) R 11

R ¹² and R ¹³ are as follows:

R ¹¹ is hydrogen; aryl; Het; C ₃ - ₇ cycloalkyl; Ci_ ₆ alkyl optionally substituted with aryl or with Het; halo; polyhaloCi-βalkyl; hydroxy; Ci_6alkoxy; Ci_6alkoxy- Ci_ ₆ alkyl; carboxyl; Ci_ ₆ alkylcarbonyl; Ci_ ₆ alkoxycarbonyl; cyano; amino; mono- or diCi_6alkylamino; or R ¹¹ is aryl; Het; Ci_6alkyl optionally substituted with aryl or with Het; halo; polyhaloCi-βalkyl; Ci_ ₆ alkoxy; Ci_ ₆ alkoxyCi_ ₆ alkyl; carboxyl; Ci_6alkoxycarbonyl; cyano; amino; mono- or diCi_6alkylamino; or R ¹¹ is aryl; Het; Ci_ ₆ alkyl; halo; Ci_ ₆ alkoxy; Ci_ ₆ alkoxycarbonyl; or or R ¹¹ is aryl; Het; halo (e.g. chloro); Ci_6alkoxy (e.g. methoxy); Ci_6alkoxycarbonyl (e.g. methoxy carbonyl) ;

R ¹² is hydrogen; Ci_ ₆ alkyl; halo; polyhaloCi-βalkyl; hydroxy; Ci_ ₆ alkoxy; Ci_ ₆ alkoxyCi_ ₆ alkyl; carboxyl; Ci_ ₆ alkoxycarbonyl; cyano; or R ¹² is hydrogen; Ci_6alkyl; halo (e.g. chloro); Ci_6alkoxy (e.g. methoxy);

R ¹³ is hydrogen; Ci_6alkyl; halo (e.g. chloro); Ci_6alkoxy (e.g. methoxy); cyano; or R ¹³ is hydrogen.

In the previous paragraph, aryl and Het are as specified above or herein after, in particular aryl is phenyl optionally substituted with Ci_ ₆ alkoxy (e.g. with methoxy, ethoxy or isopropoxy); and Het in particular is pyridyl or pyrimidinyl.

Embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ¹ is

Embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R is a radical (a-1) or (a-2) wherein

R , 11 is aryl; Het;

R . 12 is hydrogen; Ci_ ₆ alkyl; halo; polyhaloCi-βalkyl; hydroxy; Ci_ ₆ alkoxy; Ci_ ₆ alkoxyCi_ ₆ alkyl; carboxyl; Ci_ ₆ alkoxycarbonyl; cyano; or R ¹² is hydrogen; Ci_6alkyl; halo (e.g. chloro); Ci_6alkoxy (e.g. methoxy);

R ¹³ is hydrogen; Ci_6alkyl; halo (e.g. chloro); Ci_6alkoxy (e.g. methoxy); cyano; or R ¹³ is hydrogen.

Embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R is a radical (a-1) or (a-2) wherein

R ^{1 !} is phenyl; pyridyl; each optionally substituted with one or two radicals, or optionally substituted with one radical selected from halo, Ci_6alkyl, polyhalo- d-βalkyl, hydroxy, Ci_6alkoxy, carboxyl, Ci_6alkylcarbonyl, Ci_6alkoxycarbonyl, cyano, nitro, amino, mono- or diCi_6alkylamino, amino carbonyl, mono- or diCi_ ₆ alkylaminocarbonyl, phenyl, pyridyl, thiazolyl, pyrazolyl, pyrrolidinyl, piperidinyl, piperazinyl, 4-Ci_ ₆ alkylpiperazinyl, 4-Ci_ ₆ alkylcarbonyl-piperazinyl, and morpholinyl; wherein the morpholinyl and piperidinyl groups may be optionally substituted with one or with two Ci_ ₆ alkyl radicals; and the phenyl, pyridyl, thiazolyl, pyrazolyl groups may be optionally substituted with 1, 2 or 3 substituents each independently selected from Ci_6alkyl, Ci_6alkoxy, halo, amino, mono- or diCi_6alkylamino; and R ¹² and R ¹³ are as specified herein; or wherein

R ¹ is a radical (a-1) or (a-2) wherein

R ¹¹ is phenyl; pyridyl; each optionally substituted with one or two radicals, or optionally substituted with one radical selected from halo, Ci_6alkyl, polyhalo- Ci_ ₆ alkyl, hydroxy, Ci_ ₆ alkoxy, carboxyl, Ci_ ₆ alkylcarbonyl, Ci_ ₆ alkoxycarbonyl, cyano, amino, mono- or diCi_6alkylamino, amino carbonyl, mono- or diCi_6alkyl- aminocarbonyl, phenyl, pyridyl, thiazolyl.

Other subgroups of the compounds of formula (I) are those compounds of formula (I), or any subgroup of compounds of formula (I) specified herein, wherein R ¹ is a radical (a-1) or (a-2) wherein R ¹¹ is phenyl, pyrrolyl (in particular pyrrol- 1-yl), pyridyl (in particular 3-pyridyl), pyrimidinyl (in particular pyrimidin-4-yl), pyridazinyl (in particular pyridazin-3-yl and pyridazin-2-yl), 6-oxo-pyridazin-l-yl, triazolyl (in particular 1,2,3-triazolyl, 1,2,4-triazolyl, more in particular l,2,3-triazol-2-yl, l,2,4-triazol-3-yl), tetrazolyl (in particular tetrazol-1-yl, tetrazol-2-yl), pyrazolyl (in particular pyrazol-1-yl, pyrazol-3-yl), imidazolyl (in particular imidazol-1-yl, imidazol-2-yl), thiazolyl (in particular thiazol-2-yl), pyrrolidinyl (in particular pyrrolidin-1-yl), piperidinyl (in particular piperidin-1-yl), piperazinyl (in particular 1 -piperazinyl), 4-Ci_ ₆ alkyl- piperazinyl (in particular 4-Ci_ ₆ alkylpiperazin-l-yl, more in particular 4-methyl- piperazin-1-yl), furanyl (in particular furan-2-yl), thienyl (in particular thien-3-yl), morpholinyl (in particular morpholin-4-yl); all optionally substituted with one or two substituents selected from Ci_ ₆ alkyl, polyhaloCi-βalkyl, or Ci_ ₆ alkoxycarbonyl.

Further subgroups of the compounds of formula (I) are those compounds of formula (I), or any subgroup of compounds of formula (I) specified herein, wherein R ¹ is a radical (a-1) or (a-2) wherein R ¹¹ is thiazol-2-yl substituted with one or two Ci_ ₆ alkyl, such as methyl, ethyl, isopropyl or tert-butyl; in particular wherein R ¹¹ is selected from the following structures:

Embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of form miula (I) wherein R ¹ is a radical (a-1) or (a-2) wherein R ¹¹ is as specified above, R ¹² is hydrogen, halo, nitro, amino, carboxyl, C^alkyl, , CC i_6alkoxy, CCii__ ₆₆ aaUlk<ylcarbonyl, Ci_ ₆ alkoxycarbonyl, polyhaloCi-βalkyl, cyano; and R ¹³ is as specified above.

Embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ¹ is quinolinyl (in particular quinolin-4-yl), isoquinolinyl (in particular isoquinolin-1-yl), quinazolinyl (in particular quinazolin- 4-yl), or pyrimidinyl (in particular pyrimidin-4-yl), either of which is, independently, optionally mono, di, or tri substituted with d-βalkyl, Ci-βalkoxy, nitro, hydroxy, halo, trifluoromethyl, -NR ^4a R ^4b , -C(=O)NR ^4a R ^4b , C ₃ - ₇ cycloalkyl, aryl, Het, -C(=O)OH, or -C(=O)OR ^5a ; wherein aryl or Het are each, independently, optionally substituted with halo, Ci-βalkyl, Ci-βalkoxy, amino, mono- or diCi_6alkylamino, pyrrolidinyl, piperidinyl, piperazinyl, 4-Ci_ ₆ alkylpiperazinyl (e.g.4-methylpiperazinyl), thiomorpholinyl or morpholinyl; and wherein the morpholinyl, thiomorpholinyl and piperidinyl groups may optionally substituted with one or two Ci-Cβalkyl radicals.

Embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ¹ is quinolinyl (in particular quinolin-4-yl), isoquinolinyl (in particular isoquinolin-1-yl), quinazolinyl (in particular quinazolin-4-yl), or pyrimidinyl (in particular pyrimidin-4-yl), either of which is, independently, optionally mono, di, or tri substituted with methyl, ethyl, isopropyl, tert-butyl, methoxy, trifluoromethyl, trifluoromethoxy, fluoro, chloro, bromo, -NR ^4a R ^4b , -C(=O)NR ^4a R ^4b , phenyl, methoxyphenyl, cyanophenyl, halophenyl, pyridyl, Ci- ₄ alkylpyridyl, pyrimidinyl, piperidinyl, morpholinyl, piperazinyl, Ci-4alkyl-piperazinyl, pyrrolidinyl, pyrazolyl, Ci-βalkyl-pyrazolyl, thiazolyl, Ci-βalkylthiazolyl, cyclopropylthiazolyl, or mono- or diCi_6alkyl-aminothiazolyl; and

wherein the morpholinyl, thiomorpholinyl and piperidinyl groups may optionally be substituted with one or two Ci-Cβalkyl (in particular one or two methyl) radicals.

Embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ¹ is quinolinyl, optionally substituted with 1, 2, 3 or 4 (or with 1, 2 or 3) substituents selected from those mentioned as possible substituents on the monocyclic or bicyclic ring systems of R ¹ , as specified in the definitions of the compounds of formula (I) or of any of the subgroups of compounds of formula (I).

Specific embodiments of the invention are those compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ¹ is

(d-1) a radical of formula

(d-2) a radical of formula

(d-3) a radical of formula

(d-4) a radical of formula

or in particular, (d-4-a) a radical of formula

(d-5) a radical of formula

or in particular, (d-5 -a) a radical of formula

or of formula

wherein in radicals (d-1) - (d-5), as well as in (d-4-a) and (d-5-a): each R ^la , R ^lb , R ^lb' , R ^ld , R ^ld' , R ^le , R ^lf are independently any of the substituents selected from those mentioned as possible substituents on the monocyclic or bicyclic ring systems of R , as specified in the definitions of the compounds of formula (I) or of any of the subgroups of compounds of formula (I);

or, in particular, wherein in radicals (d-1) - (d-5), as well as in (d-4-a) and (d-5-a):

R ^lb and R ^lb' may, independently, be hydrogen, Ci_ ₆ alkyl, Ci- ₆ alkoxy, -NR ^4a R ^4b (in particular amino or mono- or diCi_6alkylamino), -C(=O)NR ^4a R ^4b , (in particular aminocarbonyl or mono- or diCi_ ₆ alkylaminocarbonyl), nitro, hydroxy, halo, trifluoromethyl, -C(=O)OH, or -C(=O)OR ^5a (in particular wherein R ^5a is Ci_ ₆ alkyl);

wherein each R ^4a , R ^4b , R ^5a mentioned above or hereinafter independently is as defined in the definitions of the compounds of formula (I) or of any of the subgroups of compounds of formula (I);

or, in particular, wherein in radicals (d-1) - (d-5), as well as in (d-4-a) and (d-5-a): R ^la is hydrogen, Ci_6alkyl, Ci_6alkoxy, Ci_6alkylthio, monoCi-βalkylamino, amino, C ₃ - ₇ cycloalkyl, aryl, or Het;

more specifically R ^la is Ci_ ₆ alkoxy, aryl or Het; of interest are embodiments wherein R ^la is methoxy, ethoxy, propoxy, phenyl, pyridyl, thiazolyl, pyrazolyl, each substituted as specified in the definitions of the compounds of formula (I) or of any of the subgroups of the compounds of formula (I); in specific embodiments said aryl or Het may each, independently, optionally substituted with Ci_ ₆ alkyl, Ci_ ₆ alkoxy, amino, mono- or diCi_ ₆ alkylamino, pyrrolidinyl, piperidinyl, morpholinyl, piperazinyl, 4-Ci- ₆ alkylpiperazinyl; and wherein the morpholinyl, and piperidinyl groups may optionally substituted with one or two Ci_ ₆ alkyl radicals; and in particular R ^la can be a radical Het; wherein Het may include pyrrolidinyl, piperidinyl, morpholinyl, piperazinyl, 4-Ci_ ₆ alkylpiperazinyl; and wherein the morpholinyl, thiomorpholinyl and piperidinyl groups may optionally substituted with one or two Ci_6alkyl radicals;

Embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ^la is a radical

or, in particular, wherein R ^la is selected from the group consisting of:

(q-l) (q-2) (q-3) (q-4)

wherein, where possible a nitrogen may bear an R ^lc substituent or a link to the remainder of the molecule; each R ^lc is any of the R ¹ substituents may be selected from those mentioned as possible substituents on the monocyclic or bicyclic ring systems of R ¹ , as specified in the definitions of the compounds of formula (I) or of any of the subgroups of compounds of formula (I);

specifically each R ^lc may be hydrogen, halo, Ci_ ₆ alkyl, Ci_ ₆ alkoxy, polyhaloCi-βalkyl (in particular trifluoromethyl), -NR ^4a R ^4b (in particular amino or mono- or diCi_ ₆ alkyl- amino), -C(=O)NR ^4a R ^4b , (in particular aminocarbonyl or mono- or diCi_6alkylamino- carbonyl), nitro, hydroxy, -C(=O)OH, or -C(=O)OR ^5a (in particular wherein R ^5a is

Ci_ ₆ alkyl), phenyl, pyridyl, thiazolyl, pyrazolyl, pyrrolidinyl, piperidinyl, morpholinyl, piperazinyl, 4-Ci_ ₆ alkylpiperazinyl (in particular 4-methylpiperazinyl); and wherein the morpholinyl and piperidinyl groups may optionally substituted with one or two

Ci-βalkyl radicals;

more specifically each R ^lc may be hydrogen, halo, Ci_ ₆ alkyl, amino, or mono- or di-Ci-βalkylamino, pyrrolidinyl, piperidinyl, morpholinyl, piperazinyl, 4-Ci_ ₆ alkyl- piperazinyl; and wherein the morpholinyl and piperidinyl groups may optionally substituted with one or two Ci-βalkyl radicals; and the phenyl, pyridyl, thiazolyl, pyrazolyl groups may be optionally substituted with 1, 2 or 3 (in particular with 1 or 2) substituents each independently selected from Ci_6alkyl, Ci_6alkoxy, halo, amino, mono- or diCi_6alkylamino;

more specifically each R ^lc may be hydrogen, halo, Ci_ ₆ alkyl, amino, or mono- or di-Ci-βalkylamino, pyrrolidinyl, piperidinyl, morpholinyl, piperazinyl, 4-Ci_ ₆ alkyl- piperazinyl; and wherein the morpholinyl and piperidinyl groups may optionally substituted with one or two Ci_6alkyl radicals;

and where R ^lc is substituted on a nitrogen atom, it preferably is a carbon containing substituent that is connected to the nitrogen via a carbon atom or one of its carbon atoms;

specifically each R ^ld and R ^ld independently may be hydrogen, Ci_ ₆ alkyl, Ci_ ₆ alkoxy, or halo;

or more specifically each R ^ld in (d-3) may be hydrogen, C^aUcyl, Ci_ ₆ alkoxy or halo;

specifically R ^le may be hydrogen, Ci_ ₆ alkyl, amino, mono- or diCi_ ₆ alkylamino, pyrrolidinyl, piperidinyl, morpholinyl, piperazinyl, 4-Ci_ ₆ alkylpiperazinyl (in particular

4-methylpiperazinyl); and wherein the morpholinyl and piperidinyl groups may optionally substituted with one or two Ci-βalkyl radicals;

preferably each R ^lb is Ci_6alkoxy, more preferably methoxy;

specifically R ^lf may be hydrogen, Ci_ ₆ alkyl (in particular methyl, ethyl, 1-propyl, 2-propyl, 1 -butyl, or 2-butyl, more in particular 2-propyl), amino, mono- or diCi_ ₆ alkyl- amino, pyrrolidinyl, piperidinyl, piperazinyl, 4-Ci_ ₆ alkylpiperazinyl (in particular 4-methyl-piperazinyl), or morpholinyl.

Specific embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ¹ is 7-methoxy-2-phenyl- quinolin-4-yl; or wherein R ¹ is a group

(d-5-al) or (d-6a),

wherein in (d-5-al) or (d-6a), R . If , r R _> lb , r R _> lb' are as defined herein; and in particular wherein:

R ^lf is methyl, ethyl, 1-propyl, 2-propyl, 1 -butyl, or 2-butyl, more in particular wherein

R . if is 2-propyl; R , 1b' is Ci_6aloxy, in particular Ci_4aloxy, more in particular methoxy, ethoxy,

1-propoxy, 2-propoxy; specifically wherein R Ib' is methoxy;

R , 1b is Ci_ ₆ alkyl (in particular methyl, ethyl, 1-propyl, 2-propyl, 1 -butyl, or 2-butyl, more in particular methyl).

Embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ¹ is

(e) isoquinolinyl (in particular 1-isoquinolinyl), optionally substituted with 1, 2, 3 or 4 (or with 1 , 2 or 3) substituents selected from those mentioned as possible substituents on the monocyclic or bicyclic ring systems of R ¹ , as specified in the definitions of the compounds of formula (I) or of any of the subgroups of compounds of formula (I).

Specific such embodiments are those wherein R ¹ is (e-1) a radical of formula:

or in particular (e-1 -a) a radical of formula:

wherein R ^9a , R ^9b , R ^9c independently form one another are any of the substituents selected from those mentioned as possible substituents on the monocyclic or bicyclic ring systems of R ¹ , as specified in the definitions of the compounds of formula (I) or of any of the subgroups of compounds of formula (I); in particular

R ^9a may have the same meanings as R ^la as specified above; in particular it may be aryl or Het, either of which is optionally substituted with any of the radicals mentioned as substituents of aryl or of Het as specified definitions of the compounds of formula (I) or of any of the subgroups of compounds of formula (I) (including the number of substituents); specifically said aryl or Het may be substituted with 1 , 2 or 3 (in particular with one) radical or radicals R ¹⁰ ; wherein said R ¹⁰ is any of the radicals mentioned as substituents of aryl or Het as specified definitions of the compounds of formula (I) or of any of the subgroups of compounds of formula (I) as defined above; or in particular R ¹⁰ is hydrogen, C^alkyl, C3_7Cycloalkyl, phenyl, pyridyl, thiazolyl, pyrazolyl, amino optionally mono or disubstituted with Ci_ ₆ alkyl, or aminocarbonyl or mono- or diCi_ ₆ alkylaminocarbonyl; wherein Het also includes pyrrolidinyl, piperidinyl, piperazinyl, 4-Ci_ ₆ alkylpiperazinyl (e.g. 4-methylpiperazinyl), or morpholinyl; and wherein the morpholinyl, or piperidinyl groups may optionally be substituted with one or two Ci_6alkyl radicals; and the phenyl, pyridyl, thiazolyl, pyrazolyl groups may be optionally substituted with 1, 2 or 3 (in particular with 1 or 2) substituents each independently selected from Ci_ ₆ alkyl, Ci_6alkoxy, halo, amino, mono- or diCi_6alkylamino;

R ^9b may have the same meanings as R ^lb as specified above; in particular it may be hydrogen, Chalky!, C3-7cycloalkyl, aryl, Het, halo (e.g. bromo, chloro or fluoro);

R ^9c may have the same meanings as R ^lc as specified above; in particular it may be is hydrogen or Ci_6alkoxy.

In particular R ^9a in the isoquinolinyl radical specified under (e-1) or (1-e-a) includes phenyl, pyridyl, thiazolyl, oxazolyl or pyrazolyl either of which is optionally substituted with R ¹⁰ as defined above, in particular optionally substituted whith an R ¹⁰ which may be hydrogen, Ci_ ₆ alkyl (e.g. methyl, ethyl, isopropyl, tert-butyl), amino, pyrrolidinyl, piperidinyl, piperazinyl, 4-Ci_ ₆ alkylpiperazinyl (e.g. 4-methylpiperazinyl), or morpholinyl, Ci-βalkylamino, (Ci_6alkyl)2amino, aminocarbonyl, or mono- or diCi-βalkylaminocarbonyl; and wherein the morpholinyl, and piperidinyl groups may optionally substituted with one or two Ci_6alkyl radicals.

Preferably R ^9a in the isoquinolinyl radical specified under (e-1) or (e-1 -a) includes any of radicals (q), (q'), (q'-l), (q-1), (q-2), (q-3), (q-4) specified above as well as:

wherein each R ¹⁰ is any of the radicals mentioned as substituents of Het as specified in the definitions of the compounds of formula (I) or of any of the subgroups of compounds of formula (I); or in particular R ¹⁰ is as defined above; especially R ¹⁰ is hydrogen, Ci_ ₆ alkyl (e.g. methyl, ethyl, isopropyl, tert-butyl), amino, pyrrolidinyl, piperidinyl, piperazinyl, 4-Ci_ ₆ alkylpiperazinyl (e.g. 4-methylpiperazinyl), morpholinyl, Ci_6alkylamino, (Ci-ealkyFhamino, aminocarbonyl, or mono- or diCi_6alkylamino- carbonyl; and wherein the morpholine and piperidine may optionally substituted with one or two Ci_6alkyl radicals.

Also preferably R ^9a in the isoquinolinyl radical specified under (e-1) or (e-1 -a) includes:

wherein each R ¹⁰ is as defined above, and especially is hydrogen, halo, Ci-βalkyl (e.g. methyl, ethyl, isopropyl, tert-butyl), amino, pyrrolidinyl, piperidinyl, piperazinyl,

4-Ci- ₆ alkylpiperazinyl (e.g. 4-methylpiperazinyl), morpholinyl, Ci-βalkylamino,

(Ci- ₆ alkyl) ₂ amino, aminocarbonyl, or mono- or diCi-ealkylaminocarbonyl; and wherein the morpholinyl, and piperidinyl groups may optionally substituted with one or two Ci_6alkyl radicals.

R ^9b in the isoquinolinyl radical specified under (e-2) may be hydrogen, C^alkyl, halo (e.g. bromo, chloro or fluoro), especially hydrogen or bromo.

R ^9b in the isoquinolinyl radical specified under (e-2) may be hydrogen or Ci_ ₆ alkoxy (e.g. methoxy).

Embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ¹ is

(e-2) (e-3)

wherein R ^9b is hydrogen or halo (e.g. bromo) and R ^9c is hydrogen or Ci_6alkoxy (e.g. methoxy).

Embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ¹ is

(e-4)

wherein R ^9a is as defined in any of the groups or subgroups of compounds of formula (I); and R ^9b is hydrogen, halo, or trifluoromethyl.

Further preferred embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ¹ is:

(e-4)

wherein R , 9a ^a is methoxy, ethoxy or propoxy; and

R ^9b is hydrogen, fluoro, bromo, chloro, iodo, methyl, ethyl, propyl, or trifluoromethyl.

Further embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ¹ is:

(e-5)

wherein R , 9b is hydrogen, halo, or trifluoromethyl.

Embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ¹ is

(f) quinazolinyl (in particular quinazolin-4-yl), optionally substituted with 1, 2, 3 or 4 (or with 1 , 2 or 3) substituents selected from those mentioned as possible substituents on the monocyclic or bicyclic ring systems of R ¹ , as specified in the definitions of the compounds of formula (I) or of any of the subgroups of compounds of formula (I). Quinazoline embodiments of R ¹ include (f-1) a radical :

or in particular (f-1 -a) a radical

wherein R ^9a , R ^9b and R ^9c have the meanings stated above in relation to R ¹ being isoquinolinyl (such as in radicals (e-1), (e-l-a), etc ).

wherein specifically R ^9a is C ₃ _ ₇ Cycloalkyl, aryl or Het, any of which is optionally substituted with one, two or three (in particular with one) R ¹⁰ ; wherein

R ¹⁰ is hydrogen, d-βalkyl, C3-7cycloalkyl, phenyl, pyridyl, thiazolyl, pyrazolyl, pyrrolidinyl, piperidinyl, piperazinyl, 4-methylpiperazinyl, thiomorpholinyl or morpholinyl, aminocarbonyl, mono or di Ci- ₆ alkylaminocarbonyl; wherein the piperidinyl, morpholinyl may be optionally substituted with one or two Ci_ ₆ alkyl radicals; and the phenyl, pyridyl, thiazolyl, pyrazolyl groups may be optionally substituted with 1 , 2 or 3 (or with 1 or 2) substituents each independently selected from Ci_6alkyl, Ci_6alkoxy, halo, amino, mono- or diCi_6alkylamino (in particular selected from Ci_ ₆ alkyl);

R ^9b is hydrogen, halogen, Ci-βalkyl (preferably methyl), C ₃ - ₇ cycloalkyl, aryl, Het, halo (in particular bromo, chloro or fluoro); R ^9c is hydrogen or Ci-βalkoxy.

Favoured embodiments of R ^9a for quinazo lines include aryl or Het, especially wherein R ^9a is phenyl, pyridyl, thiazolyl, oxazolyl or pyrazolyl either of which is optionally substituted with one, two or three (in particular with one) R ¹⁰ as defined.

Embodiments of R ¹⁰ for quinazo line include is hydrogen, methyl, ethyl, isopropyl, tert-butyl, methoxy, halo (including dihalo, such as difluoro), pyrrolidinyl, piperidinyl, piperazinyl, 4-Ci_ ₆ alkylpiperazinyl (e.g. 4-methylpiperazinyl) or morpholinyl, Ci-βalkylamino, (Ci-ealkyFhamino, amino carbonyl, mono or diCi-βalkylamino-carbonyl, or C ₃ _ ₇ Cycloalkyl (in particular cyclopropyl).

Preferably R ^9a in the quinazo IyI radical specified under (f-1) or (f-l-a) includes any of radicals (q), (q'), (q'-l), (q-1), (q-2), (q-3), (q-4), (q-5), (q-6), (q-7), (q-8) specified above; wherein R ¹⁰ in these radicals is as defined above or in particular is hydrogen, Ci_ ₆ alkyl (such as methyl, ethyl, isopropyl, tert-butyl), pyrrolidinyl, piperidinyl, piperazinyl,

4-Ci_ ₆ alkylpiperazinyl, N-methylpiperazinyl or morpholinyl, Ci_ ₆ alkylamino, (Ci_ ₆ alkyl) ₂ amino or amino carbonyl, mono or diCi_ ₆ alkylaminocarbonyl.

R > 9a ^a for quinazo lines may include

wherein R , 10 is hydrogen, halogen, Ci_ ₆ alkyl (such as methyl, ethyl, isopropyl, tert-butyl, Ci_6alkylamino, (Ci_6alkyl)2amino, Ci_6alkylamido, morpholinyl or piperidin-1-yl, the morpholinyl and piperidinyl being optionally substituted with one or two Ci-βalkyl groups.

Additional R ^9a embodiments for quinazolines include phenyl substited with one or two R ¹⁰ groups such as is hydrogen, methyl, ethyl, isopropyl, tert-butyl, methoxy, saturated monocyclic amino, Ci-6alkylamino, (Ci-ealkyThamino or aminocarbonyl, mono- and diCi_ ₆ alkylaminocarbonyl or halo (in particular fluoro).

Embodiments of R ^9b for quinazolines include hydrogen, Ci-βalkyl (in particular methyl), halo (e.g. bromo, chloro or fluoro) especially wherein R ^9b is hydrogen or bromo.

Embodiments of R ^9c for quinazolines include hydrogen or Ci_6alkoxy (in particular methoxy).

Specific embodiments of the compounds of formula (I) or any of the subgroups of compounds of formula (I) are those wherein R ¹ is :

(f-2) (f-3)

wherein R ¹⁰ and R ^9c are as specified above and in particular and R ^9c is hydrogen or Ci_6alkoxy (e.g. methoxy).

Embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ¹ is

(g-i) wherein R , 9a ^a is as defined in any of the groups or subgroups of compounds of formula (I), preferably R ^9a is p-methoxyphenyl or p-fluoromethyl; and R ^9b is hydrogen, methyl, halo, or trifluoromethyl.

Further preferred embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ¹ is:

wherein R ^9a is methoxy, ethoxy or propoxy; and

R , 9b is hydrogen, fluoro, bromo, chloro, iodo, methyl, ethyl, propyl, or trifluoromethyl.

Further embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ¹ is:

wherein R , 9b is hydrogen, halo, or trifluoromethyl.

A further embodiment of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein p is 2.

Further embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein each R ^4a and R ^4b are, independently, hydrogen, C ₃ - ₇ cycloalkyl, aryl, Ci_ ₆ alkyl optionally substituted with Ci_ ₄ alkoxy, cyano, phenyl; or R ^4a and R ^4b taken together with the nitrogen atom to which they are attached form pyrrolidinyl, piperidinyl, piperazinyl, 4-Ci_ ₆ alkylpiperazinyl, 4-Ci_ ₆ alkylcarbonyl-piperazinyl, and morpholinyl; wherein the morpholinyl and piperidinyl groups may be optionally substituted with one or with two Ci_ ₆ alkyl radicals; or wherein each R ^4a and R ^4b are, independently, hydrogen, Ci_ ₆ alkyl optionally substituted with Ci_4alkoxy, phenyl.

Further embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ⁵ is hydrogen; or Ci_ ₆ alkyl optionally substituted with aryl or Het; or R ⁵ is hydrogen; or Ci_ ₆ alkyl optionally substituted with phenyl.

Further embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein R ^5a is Ci_ ₆ alkyl optionally substituted with aryl or Het; or R ^5a is Ci_ ₆ alkyl.

Further embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein

R ⁶ is hydrogen, C _h alky!, or aryl; or R ⁶ is hydrogen, C^alkyl, or phenyl; or R ⁶ is

Ci_ ₆ alkyl.

Further embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein

R ⁷ is trifluoromethyl; aryl; Het; or Ci_ ₆ alkyl optionally substituted with aryl or Het; or

R ⁷ is trifluoromethyl; aryl; or Ci_ ₆ alkyl optionally substituted with aryl; or R ⁷ is trifluoromethyl; phenyl; or Ci_ ₆ alkyl optionally substituted with phenyl; or

R ⁷ is Ci_ ₆ alkyl.

A further embodiment of the invention are compounds of formula (I), or any of the subgroups of compounds of formula (I), wherein R ² is -C(=O)-NH-P(=O)(OR ^8a )(R ^8b ), in particular wherein R ^8a is Ci_ ₆ alkyl, especially ethyl or isopropyl and R ^8b is OR ^8b and R is Ci_ ₆ alkyl, such as ethyl or isopropyl.

A further embodiment of the invention are compounds of formula (I), or any of the subgroups of compounds of formula (I), wherein R ² is -P(=O)(OR ^8a )(R ^8b ), in particular wherein R ^8a is C _1-6 alkyl, especially ethyl or isopropyl and R ^8b is OR ^8b' and R ^8b' is Ci_ ₆ alkyl, especially ethyl or isopropyl.

Further embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein

(a) R ³ is hydrogen;

(b) R ³ is d-βalkyl, preferably methyl.

Further embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein

(a) R ⁹ is hydrogen; Ci_ ₆ alkyl; Ci_ ₆ alkoxyCi_ ₆ alkyl; or C ₃ _ ₇ Cycloalkyl

(b) R ⁹ is hydrogen or Ci_ ₆ alkyl; (c) R ⁹ is hydrogen.

Embodiments of the invention are compounds of formula (I) or any of the subgroups of compounds of formula (I) wherein

(a) X is N, C (X being linked via a double bond) or CH (X being linked via a single bond) and R ³ is hydrogen;

(b) X is C (X being linked via a double bond) and R is Ci_6alkyl, preferably methyl.

The compounds of formula (I) consist of three building blocks Pl, P2, P3, which are each delimited by a curved line. The building block Pl further contains a Pl ' tail. The linking of building blocks Pl with P2, and optionally Pl with Pl ', involves forming an amide bond. The linking of building blocks P3 with P2 involves an acylation when P2 is a pyrrolidine ring. The linking of blocks Pl and P3 involves double bond formation. The linking of building blocks Pl, Pl ', P2 and P3 to prepare compounds of formula (I) can be done in any given sequence. One of the steps involves a cyclization whereby the macrocycle is formed. Compounds of formula (I-j) can be prepared from compound of formula (I-i) by a reduction of the double bond, e.g. with hydrogen in the presence of a noble metal catalyst such as Rh, Pd or Pt.

(I-i) (i-j)

The synthesis procedures described hereinafter are meant to be applicable for as well the racemates, stereochemically pure intermediates or end products, or any stereoisomeric mixtures. The racemates or stereochemical mixtures may be separated into stereoisomeric forms at any stage of the synthesis procedures. In one embodiment, the intermediates and end products have the stereochemistry specified above in the compounds of formula (I-b).

In one embodiment, compounds (I-i) are prepared by first forming the amide bond between P2 and Pl, coupling the P3 moiety to P2, and subsequent forming the double bond linkage between P3 and Pl with concomitant cyclization to the macrocycle.

In a preferred embodiment, compounds (I) wherein the bond between C ₇ and Cs is a double bond, which are compounds of formula (I-i), as defined above, may be prepared as outlined in the following reaction scheme:

Formation of the macrocycle can be carried out via an olefin metathesis reaction in the presence of a suitable metal catalyst such as e.g. the Ru-based catalyst reported by

Miller, S.J., Blackwell, H.E., Grubbs, R.H. J. Am. Chem. Soc. 118, (1996), 9606-9614; Kingsbury, J. S., Harrity, J. P. A., Bonitatebus, P. J., Hoveyda, A. H., J. Am. Chem. Soc. 121, (1999), 791-799; and Huang et al, J. Am. Chem. Soc. 121, (1999),

2674-2678; for example a Hoveyda-Grubbs catalyst.

Air-stable ruthenium catalysts such as bis(tricyclohexylphosphine)-3-phenyl-lH-inden- 1-ylidene ruthenium chloride (Neolyst Ml ^® ) or bis(tricyclohexylphosphine)- [(phenylthio)methylene]ruthenium (IV) dichloride can be used. Other catalysts that can be used are Grubbs first and second generation catalysts, i.e. Benzylidene- bis(tricyclohexylphosphine)dichlororuthenium and ( 1 ,3-bis-(2,4,6-trimethylphenyl)- 2-imidazolidinylidene)dichloro(phenylmethylene)-(tricyclohex ylphosphine)ruthenium, respectively. Of particular interest are the Hoveyda-Grubbs first and second generation catalysts, which are dichloro(o-isopropoxyphenylmethylene)(tricyclohexylphosphine )- ruthenium(II) and 1 ,3-bis-(2,4,6-trimethylphenyl)-2-imidazolidinylidene)dichlor o(o- isopropoxyphenylmethylene)ruthenium respectively. Also other catalysts containing other transition metals such as Mo can be used for this reaction.

The metathesis reactions may be conducted in a suitable solvent such as for example ethers, e.g. THF, dioxane; halogenated hydrocarbons, e.g. dichoromethane, CHCI3, 1 ,2-dichloroethane and the like, hydrocarbons, e.g. toluene. In a preferred embodiment, the metathesis reaction is conducted in toluene. These reactions are conducted at increased temperatures under nitrogen atmosphere.

Compounds of formula (I) wherein the link between C7 and C8 in the macrocycle is a single bond, i.e. compounds of formula (I-j), can be prepared from the compounds of formula (I-i) by a reduction of the C7-C8 double bond in the compounds of formula (I-i). This reduction may be conducted by catalytic hydrogenation with hydrogen in the presence of a noble metal catalyst such as, for example, Pt, Pd, Rh, Ru or Raney nickel. Of interest is Rh on alumina. The hydrogenation reaction preferably is conducted in a solvent such as, e.g. an alcohol such as methanol, ethanol, or an ether such as THF, or mixtures thereof. Water can also be added to these solvents or solvent mixtures.

The R ² group can be connected to the Pl building block at any stage of the synthesis, i.e. before or after the cyclization, or before or after the cyclization and reduction as described herein above. In the compounds of formula (I) wherein R ² represents

-P(=O)(OR ^8a )(R ^8b ), this group is preferably introduced when building up the Pl moiety as is described hereinafter.

The compounds of formula (I) wherein R ² represents -C(=O)-NH-P(=O)(OR ^8a )(R ^8b ), said compounds being represented by formula (I-d-1), can be prepared by linking the R ² group to Pl by forming an amide bond between both moieties. Intermediate (2a) can be coupled with the phosphoramidate (2b) by an amide forming reaction such as any of the procedures for the formation of an amide bond described hereinafter. In particular, (2a) may be treated with a coupling agent in an appropriate solvent, preferably in the presence of a base, followed by reaction with phosphoramidate (2b), preferably after reacting (2a) with the coupling agent. Preferably, the base is added to the phosphoramidate (2b) prior to reaction with (2a) or the activated derivative of (2a), a useful base to this purpose being an alkalimetal hydride such as sodium hydride.

Intermediate (2a) can also be converted into an activated form, e.g. an activated form of general formula G-CO-Z, wherein Z represents halo, or the rest of an active ester, e.g. Z is an aryloxy group such as phenoxy, /?-nitrophenoxy, pentafluorophenoxy, trichloro- phenoxy, pentachlorophenoxy and the like; or Z can be the rest of a mixed anhydride. In one embodiment, G-CO-Z is an acid chloride (G-CO-Cl) or a mixed acid anhydride (G-CO-O-CO-R or G-CO-O-CO-OR, R in the latter being e.g. d_ ₄ alkyl, such as methyl, ethyl, propyl, i.propyl, butyl, t.butyl, i.butyl, or benzyl). The activated form G-CO-Z is reacted with the desired (2b). The coupling agent, solvent and base may be as described hereinafter in the general description of the preparation of amide bonds.

The activation of the carboxylic acid in (2a) as described in the above reactions may lead to an internal cyclization reaction to an azalactone intermediate of formula

wherein R ¹ , R ³ , n are as specified above and wherein the stereogenic centers may have the stereochemical configuration as specified above, for example as in (I-a) or (I-b). The intermediates (2a- 1) can be isolated from the reaction mixture, using conventional methodology, and the isolated intermediate (2a-l) is then reacted with (2b), or the reaction mixture containing (2a- 1) can be reacted further with (2b) without isolation of (2a- 1). In one embodiment, where the reaction with the coupling agent is conducted in a water-immiscible solvent, the reaction mixture containing (2a- 1) may be washed with water or with slightly basic water in order to remove all water-soluble side products. The thus obtained washed solution may then be reacted with (2b) without additional purification steps. The isolation of intermediates (2a- 1) on the other hand may provide certain advantages in that the isolated product, after optional further purification, may be reacted with (2b), giving rise to less side products and an easier work-up of the reaction.

The compounds of formula (2a) can be obtained from the corresponding esters (3), wherin R is a Ci_ ₄ alkyl group such as methyl or ethyl, using standard hydrolysis procedures, e.g. with an alkali metal hydroxide such as LiOH or NaOH in an aqueous medium.

(2a) (3)

The compounds of formula (I) wherein E is NH, said compounds being represented by (1-1), can also be prepared by removal of a protecting group PG, from a corresponding nitrogen-protected intermediate (3a), as in the following reaction scheme. The protecting group PG in particular is any of the nitrogen protecting groups mentioned hereinafter and can be removed using procedures also mentioned hereinafter:

(3a) (M)

The starting materials (3a) in the above reaction can be prepared following the procedures for the preparation of compounds of formula (I), but using intermediates wherein the group R ⁹ is PG.

The compounds of formula (I) can also be prepared by reacting an intermediate (4a) with intermediates (4b) as outlined in the following reaction scheme wherein the various radicals have the meanings specified above:

Y in (4b) represents hydroxy or a leaving group such as a halide, e.g. bromide or chloride, or an arylsulfonyl group, e.g. mesylate, triflate or tosylate and the like.

In one embodiment, the reaction of (4a) with (4b) is an O-arylation reaction and Y represents a leaving group. This reaction can be conducted following the procedures described by E. M. Smith et al. (J. Med. Chem. (1988), 31, 875-885). In particular, this reaction is conducted in the presence of a base, preferably a strong base, in a reaction- inert solvent, e.g. one of the solvents mentioned for the formation of an amide bond.

In a particular embodiment, starting material (4a) is reacted with (4b) in the presence of a base which is strong enough to detract a hydrogen from the hydroxy group, for example an alkali of alkaline metal hydride such as LiH or sodium hydride, or alkali

metal alkoxide such as sodium or potassium methoxide or ethoxide, potassium te/t-butoxide, in a reaction inert solvent like a dipolar aprotic solvent, e.g. DMA, DMF and the like. The resulting alcoholate is reacted with the arylating agent (4b), wherein Y is a suitable leaving group as mentioned above. The conversion of (4a) to (I) using this type of O-arylation reaction does not change the stereochemical configuration at the carbon bearing the hydroxy or -O-R ¹ group.

Alternatively, the reaction of (4a) with (4b) can also be conducted via a Mitsunobu reaction (Mitsunobu, 1981, Synthesis, January, 1-28; Rano et al., Tetrahedron Lett., 1995, 36, 22, 3779-3792; Krchnak et al., Tetrahedron Lett, 1995, 36, 5, 6193-6196; Richter et al., Tetrahedron Lett., 1994, 35, 27, 4705-4706). This reaction comprises treatment of intermediate (4a) with (4b) wherein Y is hydroxy, in the presence of triphenylphosphine and an activating agent such as a dialkyl azocarboxylate, e.g. diethyl azodicarboxylate (DEAD), diisopropyl azodicarboxylate (DIAD) or the like. The Mitsunobu reaction changes the stereochemical configuration at the carbon bearing the hydroxy or -O-R ¹ group. The compounds of formula (I) wherein R ¹ is a group of formula

i.e. compounds of formula (I-k), can also be prepared by reacting an intermediate (4a) with an amine (4c) in the presence of a carbamate forming reagent as outlined in the following reaction scheme wherein the various radicals have the meanings specified above:

The reaction of intermediates (4a) with the carbamate forming reagent is conducted in the same solvents and bases as those used for the amide bond formation as described hereinafter.

Carbamate forming reactions may be conducted using a variety of methods, in particular by reaction of amines with alkyl chloro formates; by reaction of alcohols with carbamoyl chlorides or isocyanates; via reactions involving metal complexes or acyl transfer agents. See for example, Greene, T. W. and Wuts, P. G. M., "Protective Groups in Organic Synthesis"; 1999; Wiley and Sons, p. 309-348. Carbon monoxide and certain metal catalysts can be used to synthesize carbamates from several starting compounds, including amines. Metals such as palladium, iridium, uranium, and platinum may be used as catalysts. Methods using carbon dioxide for synthesis of carbamates that have been also been reported, can also be used (see for example, Yoshida, Y., et al, Bull. Chem. Soc . Japan 1989, 62, 1534; and Aresta, M., et al, Tetrahedron, 1991, 47, 9489).

One approach for the preparation of carbamates involves the use of intermediates

wherein Q is leaving group such as halo, in particular chloro or bromo, or a group used in active esters for amide bond formation, such as those mentioned above, for example phenoxy or substituted phenoxy such as/?-chloro and/?-nitrophenoxy, trichloro- phenoxy, pentachlorophenoxy, N-hydroxy-succinimidyl, and the like. Intermediates (4d) can be derived from alcohols (4a) and phosgene, thus forming a chloro formate, or by transferring the chloro in the latter to intermediates (4e) which are intermediates of formula (4d) wherein Q is Q ¹ . In this and the following reaction procedures, Q ¹ represents any of the active ester moieties such as those mentioned above. Intermediates (4d) are reacted with (4c), obtaining compounds (I-k).

Intermediates (4e), which are intermediates (4d) wherein Q is Q ¹ , can also be prepared by reacting the alcohol (4a) with carbonates Q'-CO-Q ¹ such as e.g. bisphenol, bis-(substituted phenol) or bis N-hydroxy-succinimidyl carbonates:

The above reactions to prepare reagents (4e) may be conducted in the presence of the bases and solvents mentioned hereinafter for the synthesis of amide bonds, in particular triethylamine and dichloromethane.

Alternatively, in order to prepare the compounds of formula (I), first an amide bond between building blocks P2 and Pl is formed, followed by coupling of the P3 building block to the Pl moiety in P1-P2, and a subsequent carbamate or ester bond formation between P3 and the P2 moiety in P2-P1-P3 with concomitant ring closure.

Yet another alternative synthetic methodology is the formation of an amide bond between building blocks P2 and P3, followed by the coupling of building block Pl to the P3 moiety in P3-P2, and a last amide bond formation between Pl and P2 in P1-P3-P2 with concomitant ring closure.

Building blocks Pl and P3 can be linked and the thus formed P1-P3 block can be coupled to building block P2 and the thus forming sequence P1-P2-P3 subsequently cyclized, by forming carbamate or ester amide bonds.

Building blocks Pl and P3 in any of the previous approaches can be linked via double bond formation, e.g. by the olefin metathesis reaction described hereinafter, or a Wittig type reaction. If desired, the thus formed double bond can be reduced, similarly as described above for the conversion of (I-i) to (I-j). The double bond can also be reduced at a later stage, i.e. after addition of a third building block, or after formation of the macrocycle. Building blocks P2 and Pl are linked by amide bond formation and P3 and P2 are linked by carbamate or ester formation.

The tail Pl ' can be bonded to the Pl building block at any stage of the synthesis of the compounds of formula (I), for example before or after coupling the building blocks P2 and Pl; before or after coupling the P3 building block to Pl; or before or after ring closure.

The individual building blocks can first be prepared and subsequently coupled together or alternatively, precursors of the building blocks can be coupled together and modified at a later stage to the desired molecular composition.

The functionalities in each of the building blocks may be protected to avoid side reactions.

The formation of amide bonds can be carried out using standard procedures such as those used for coupling amino acids in peptide synthesis. The latter involves the dehydrative coupling of a carboxyl group of one reactant with an amino group of the other reactant to form a linking amide bond. The amide bond formation may be performed by reacting the starting materials in the presence of a coupling agent or by converting the carboxyl functionality into an active form such as an active ester, mixed anhydride or a carboxyl acid chloride or bromide. General descriptions of such coupling reactions and the reagents used therein can be found in general textbooks on peptide chemistry, for example, M. Bodanszky, "Peptide Chemistry", 2nd rev. ed., Springer- Verlag, Berlin, Germany, (1993).

Examples of coupling reactions with amide bond formation include the azide method, mixed carbonic-carboxylic acid anhydride (isobutyl chloroformate) method, the carbodiimide (dicyclohexylcarbodiimide, diisopropylcarbodiimide, or water-soluble carbodiimide such as λ/-ethyl-N'-[(3-dimethylamino)propyl]carbodiimide) method, the active ester method (e.g. /?-nitrophenyl,/?-chlorophenyl, trichlorophenyl, pentachloro- phenyl, pentafluorophenyl, JV-hydroxysuccinic imido and the like esters), the

Woodward reagent K-method, the 1,1-carbonyldiimidazole (CDI or N,N'-carbonyl- diimidazole) method, the phosphorus reagents or oxidation-reduction methods. Some of these methods can be enhanced by adding suitable catalysts, e.g. in the carbodiimide method by adding 1-hydroxybenzotriazole, DBU (l,8-diazabicyclo[5.4.0]undec-7-ene), or 4-DMAP. Further coupling agents are (benzotriazol-l-yloxy)tris-(dimethylamino) phosphonium hexafluorophosphate, either by itself or in the presence of 1-hydroxybenzotriazole or 4-DMAP; or 2-(lH-benzotriazol-l-yl)-λ/,λ/,λf',λf'-tetra-methyluroni um tetrafluoroborate, or O-(7-azabenzotriazol- 1 -yl)-λ/,λ/,λf',λf'-tetramethyluronium

hexafluorophosphate. These coupling reactions can be performed in either solution (liquid phase) or solid phase.

A preferred amide bond formation is performed employing N-ethyloxycarbonyl- 2-ethyloxy- 1 ,2-dihydroquinoline (EEDQ) or N-isobutyloxy-carbonyl-2-isobutyloxy- 1 ,2-dihydroquinoline (IIDQ). Unlike the classical anhydride procedure, EEDQ and IIDQ do not require base nor low reaction temperatures. Typically, the procedure involves reacting equimolar amounts of the carboxyl and amine components in an organic solvent (a wide variety of solvents can be used). Then EEDQ or IIDQ is added in excess and the mixture is allowed to stir at room temperature.

The coupling reactions preferably are conducted in an inert solvent, such as halogenated hydrocarbons, e.g. dichloromethane, chloroform, dipolar aprotic solvents such as acetonitrile, dimethylformamide, dimethylacetamide, DMSO, HMPT, ethers such as tetrahydrofuran (THF).

In many instances the coupling reactions are done in the presence of a suitable base such as a tertiary amine, e.g. triethylamine, diisopropylethylamine (DIPEA), JV-methyl-morpholine, JV-methylpyrrolidine, 4-DMAP or l,8-diazabicycle[5.4.0]undec-7-ene (DBU). The reaction temperature may range between 0 ⁰ C and 50 ⁰ C and the reaction time may range between 15 min and 24 h.

The functional groups in the building blocks that are linked together may be protected to avoid formation of undesired bonds. Appropriate protecting groups that can be used are listed for example in Greene, "Protective Groups in Organic Chemistry", John Wiley & Sons, New York (1999) and "The Peptides: Analysis, Synthesis, Biology", Vol. 3, Academic Press, New York (1987).

Carboxyl groups can be protected as an ester that can be cleaved off to give the carboxylic acid. Protecting groups that can be used include 1) alkyl esters such as methyl, trimethylsilyl and tert-butyi; T) arylalkyl esters such as benzyl and substituted benzyl; or 3) esters that can be cleaved by a mild base or mild reductive means such as trichloroethyl and phenacyl esters.

Amino groups can be protected by a variety of N-protecting groups, such as:

1) acyl groups such as formyl, trifluoroacetyl, phthalyl, and/?-toluenesulfonyl;

2) aromatic carbamate groups such as benzyloxycarbonyl (Cbz or Z) and substituted benzyloxycarbonyls, and 9-fluorenylmethyloxycarbonyl (Fmoc);

3) aliphatic carbamate groups such as te/t-butyloxycarbonyl (Boc), ethoxycarbonyl, diisopropylmethoxy-carbonyl, and allyloxycarbonyl;

4) cyclic alkyl carbamate groups such as cyclopentyloxycarbonyl and adamantly- oxycarbonyl; 5) alkyl groups such as triphenylmethyl, benzyl or substituted benzyl such as 4-methoxybenzyl;

6) trialkylsilyl such as trimethylsilyl or t.Bu dimethylsilyl; and

7) thiol containing groups such as phenylthiocarbonyl and dithiasuccinoyl.

Interesting amino protecting groups are Boc and Fmoc.

Preferably the amino protecting group is cleaved off prior to the next coupling step. Removal of N-protecting groups can be done following art-known procedures. When the Boc group is used, the methods of choice are trifluoroacetic acid, neat or in dichloromethane, or HCl in dioxane or in ethyl acetate. The resulting ammonium salt is then neutralized either prior to the coupling or in situ with basic solutions such as aqueous buffers, or tertiary amines in dichloromethane or acetonitrile or dimethyl- formamide. When the Fmoc group is used, the reagents of choice are piperidine or substituted piperidine in dimethylformamide, but any secondary amine can be used. The deprotection is carried out at a temperature between 0 ⁰ C and room temperature, usually around 15-25 ⁰ C, or 20-22 ⁰ C.

Other functional groups that can interfere in the coupling reactions of the building blocks may also be protected. For example hydroxyl groups may be protected as benzyl or substituted benzyl ethers, e.g. 4-methoxybenzyl ether, benzoyl or substituted benzoyl esters, e.g. 4-nitrobenzoyl ester, or with trialkylsilyl goups (e.g. trimethylsilyl or tert-buty ldimethylsily 1) .

Further amino groups may be protected by protecting groups that can be cleaved off selectively. For example, when Boc is used as the α-amino protecting group, the following side chain protecting groups are suitable: p-toluenesulfonyl (tosyl) moieties can be used to protect further amino groups; benzyl (Bn) ethers can be used to protect hydroxy groups; and benzyl esters can be used to protect further carboxyl groups. Or when Fmoc is chosen for the α-amino protection, usually tert-butyl based protecting groups are acceptable. For instance, Boc can be used for further amino groups; tert-butyl ethers for hydroxyl groups; and tert-butyl esters for further carboxyl groups.

Any of the protecting groups may be removed at any stage of the synthesis procedure but preferably, the protecting groups of any of the functionalities not involved in the reaction steps are removed after completion of the build-up of the macrocycle.

Removal of the protecting groups can be done in whatever manner is dictated by the choice of protecting groups, which manners are well known to those skilled in the art.

The intermediates of formula (Ia) wherein X is N and E is N-R ⁹ , said intermediates being represented by formula (Ia-I), may be prepared starting from intermediates (5a) which are reacted with an alkenamine (5b) in the presence of a carbonyl introducing agent as outlined in the following reaction scheme.

Carbonyl (CO) introducing agents include phosgene, or phosgene derivatives such as carbonyl diimidazole (CDI), and the like. In one embodiment (5a) is reacted with the CO introducing agent in the presence of a suitable base and a solvent, which can be the bases and solvents used in the amide forming reactions as described above. In a particular embodiment, the base is a hydrogencarbonate, e.g. NaHCOs, or a tertiary amine such as triethylamine and the like, and the solvent is an ether or halogenated hydrocarbon, e.g. THF, CH ₂ Cl ₂ , CHCI3, and the like. Thereafter, the amine (5b) is added thereby obtaining intermediates (Ia-I) as in the above scheme. An alternative route using similar reaction conditions involves first reacting the CO introducing agent with the alkenamine (5b) and then reacting the thus formed intermediate with (5a).

The intermediates (Ia-I) can alternatively be prepared as follows:

deprotection

PG ¹ is an O-protecting group, which can be any of the groups mentioned herein and in particular is a benzoyl or substituted benzoyl group such as 4-nitrobenzoyl.

Intermediates (6a) are reacted with a carbamate forming reagent derived from alkenyl (5b) and this reaction yields intermediates (6c). These are deprotected, in particular using the reaction conditions mentioned above. For example where PG ¹ is benzoyl or substituted benzoyl this group is removed by reaction with a an alkali metal hydroxide (LiOH, NaOH, KOH), in particular where PG ¹ is 4-nitrobenzoyl, with LiOH, in an aqueous medium comprising water and a water-soluble organic solvent such as an alkanol (methanol, ethanol) and THF. The resulting alcohol (6d) is reacted with intermediates (4b) or (4c) as described above for the reaction of (4a) with (4b) or (4c), and this reaction results in intermediates (Ia).

Carbamate forming reactions may be conducted using a variety of methods, in particular by reaction of amines with alkyl chloro formates; by reaction of alcohols with carbamoyl chlorides or isocyanates; via reactions involving metal complexes or acyl transfer agents. See for example, Greene, T. W. and Wuts, P. G. M., "Protective Groups in Organic Synthesis"; 1999; Wiley and Sons, p. 309-348. Carbon monoxide and certain metal catalysts can be used to synthesize carbamates from several starting compounds, including amines. Metals such as palladium, iridium, uranium, and platinum may be used as catalysts. Methods using carbon dioxide for synthesis of carbamates that have been also been reported, can also be used (see for example, Yoshida, Y., et al, Bull. Chem. Soc . Japan 1989, 62, 1534; and Aresta, M., et al, Tetrahedron, 1991, 47, 9489).

One approach for the preparation of carbamates is by using a reagent

wherein W is leaving group such as halo, in particular chloro and bromo, or a group used in active esters for amide bond formation, such as those mentioned above, for example phenoxy or substituted phenoxy such as p. chloro and p.nitrophenoxy, trichlorophenoxy, pentachlorophenoxy, N-hydroxy-succinimidyl, and the like. Reagent (7) can be formed from alkene amine (5b) and phosgene thus forming an alkenyl chloro formate or by transferring the chloro in the latter to ragents (7) wherein W is W ¹ , the latter being any of the active ester moieties such as those mentioned above, hereafter referred to as reagents (7a). Reagents (7) are reacted with (5a) or (6a), obtaining ( 1 a- 1 ) or (6c) .

The reagents (7a) can also be prepared by reacting alkene amines (5b) with carbonates W'-CO-W ¹ such as e.g. bisphenol, bis-(substituted phenol) or bis N-hydroxy-succinimidyl carbonates :

The reagents (7a) may also be prepared from chloroformates Cl-CO-W ¹ as follows

(5b)

The above reactions to prepare reagents (7a) may be conducted in the presence of a suitable base and in a reaction- inert solvent such as the bases and solvents mentioned above for the synthesis of amide bonds, in particular triethylamine and dichloromethane .

The intermediates of formula (Ia) wherein X is C, said intermediates being represented by formula (la-2), may be prepared by an amide forming reaction starting from an intermediate (8a) which are reacted with an alkene amine (5b) as shown in the

following reaction scheme, using reaction conditions for preparing esters such as the reaction conditions as those described above for the reaction of (4a) with (4e).

The intermediates (Ia-I) can alternatively be prepared as follows:

PG ¹ is an O-protecting group as described above. The same reaction conditions as described above may be used: ester formation as for the reaction of (4a) with (4e), removal of PG ¹ as in the description of the protecting groups and introduction of R ¹ as in the reactions of (4a) with (4b).

The intermediates of formula (2a) may be prepared by first cyclizing the open amide (9a) to a macrocyclic amide (9b), which in turn is converted to (2a) as follows:

O-R ¹ is as specified above and PG ² is a carboxyl protecting group, e.g. one of the carboxyl protecting groups mentioned above, in particular a Ci_ ₄ alkyl or benzyl ester, e.g. a methyl, ethyl or t.butyl ester. The reaction of (9a) to (9b) is a metathesis reaction and is conducted as described above. The group PG ² is removed following procedures also described above. Where PG ² is a Ci_4alkyl ester, it is removed by alkaline hydrolysis, e.g. with NaOH or preferably LiOH, in an aqueous solvent, e.g. a C i_ ₄ alkano I/water mixture. A benzyl group can be removed by catalytic hydrogenation.

In an alternative synthesis, intermediates (2a) can be prepared as follows:

The PG ¹ group is selected such that it is selectively cleavable towards PG ² . PG ² may be e.g. methyl or ethyl esters, which can be removed by treatment with an alkali metal hydroxide in an aqueous medium, in which case PG ¹ e.g. is t.butyl or benzyl. PG ² may be t.butyl esters removable under weakly acidic conditions or PG ¹ may be a benzyl ether removable with strong acid or by catalytic hydrogenation, in the latter two cases PG ¹ e.g. is a benzoic ester such as a 4-nitrobenzoic ester.

First, intermediates (10a) are cyclized to the macrocyclic esters (10b), the latter are deprotected by removal of the PG ¹ group to (10c), which are reacted with intermediates (4b) or (4c) to intermediates (9b), followed by removal of carboxyl protecting group PG ² , which yields intermediates (2a) The cyclization, deprotection of PG ¹ and PG ² and the coupling with (4b) or (4c) are as described above.

The R ² groups can be introduced at any stage of the synthesis, either as the last step as described above, or earlier, before the macrocycle formation. In the following scheme the group R ² being -C(=O)-NH-P(=O)(OR ^8a )(R ^8b ) (which is as specified above):

In the above scheme, PG ² is as defined above and L ¹ is a P3 group o

_' '" ^{' E} (b), wherein n is as defined above and where X is N, L ¹ may also be a nitrogen-protecting group (PG, as defined above) and where X is C, L ¹ may also be a

group -COOPG ^a , wherein the group PG ^a is a carboxyl protecting group as PG , but wherei nn l PG ^2a is selectively cleavable towards PG ² . In one embodiment PG ^2a is t.butyl and PG ² is methyl or ethyl.

The intermediates (1 Ic) wherein L ¹ represents a group (b) correspond to the intermediates (Ia) and may be processed further as specified above.

Coupling of Pl and P2 building blocks

The Pl and P2 building blocks are linked using an amide forming reaction following the procedures described above. The Pl building block may have a carboxyl protecting group PG ² (as in (12b)) or may already be linked to Pl' group (as in (12c)). L ² is a N-protecting group (PG), or a group (b), as specified above. L ³ is hydroxy, -OPG ¹ or a group -O-R ¹ as specified above. Where in any of the following reaction schemes L ³ is hydroxy, prior to each reaction step, it may be protected as a group -OPG ¹ and, if desired, subsequently deprotected back to a free hydroxy function. Similarly as described above, the hydroxy function may be converted to a group -O-R ¹ .

In the procedure of the above scheme, R ² preferably is -C(=O)-NH-P(=O)(OR ^8a )(R ^8b ). In a first step a cyclopropyl amino acid (12b) or (12c) is coupled to the acid function of the P2 building block (12a) with the formation of an amide linkage, following the procedures described above. Intermediates (12d) or (12e) are obtained. Where in the latter L ² is a group (b), the resulting products are P3-P2-P1 sequences encompassing some of the intermediates (1 Ic) in the previous reaction scheme. Removal of the acid protecting group in (12d), using the appropriate conditions for the protecting group

used, followed by coupling with a phosphoramidate NH ₂ -P(=O)(OR ^8a )(R ⁸ K ^B ) (2b) as described above, again yields the intermediates (12e), wherein -COR ² are carbonyl phosphoramidate groups. Where L ² is a N-protecting group, it can be removed yielding intermediates (5a) or (6a). In one embodiment, PG in this reaction is a BOC group and PG ² is methyl or ethyl. Where additionally L ³ is hydroxy, the starting material (12a) is Boc-L-hydroxyproline. In a particular embodiment, PG is BOC, PG ² is methyl or ethyl and L ³ Is -O-R ¹ .

In one embodiment, L ² is a group (b) and these reactions involve coupling Pl to P2-P3, which results in the intermediates (Ia-I) or (Ia) mentioned above. In another embodiment, L ² is a N-protecting group PG, which is as specified above, and the coupling reaction results in intermediates (12d-l) or (12e-l), from which the group PG can be removed, using reaction conditions mentioned above, obtaining intermediates (12-f) or respectively (12g), which encompass intermediates (5a) and (6a) as specified above:

In one embodiment, the group L ³ in the above schemes represents a group -O-PG ¹ which can be introduced on a starting material (12a) wherein L is hydroxy. In this instance PG ¹ is chosen such that it is selectively cleavable towards group L ² being PG.

In a similar way, P2 building blocks wherein X is C, which are cyclopentane or cyclopentene derivatives, can be linked to Pl building blocks as outlined in the following scheme wherein R ² , R ³ , L ³ , PG ² and PG ^2a are carboxyl protecting groups. PG ^2a typically is chosen such that it is selectively cleavable towards group PG ² . Removal of the PG ^2a group in (13c) yields intermediates (8a), which can be reacted with (5b) as described above.

In one particular embodiment, where X is C, R ³ is H, and where X and the carbon bearing R ³ are linked by a single bond (P2 being a cyclopentane moiety), PG ^2a and L ³ a bond and the P2 building block is represented by formula:

Bicyclic acid (14a) is reacted with (12b) or (12c) similar as described above to (14b) and (14c) respectively, wherein the lactone is opened giving intermediates (14c) and (14e). The lactones can be opened using ester hydrolysis procedures, for example usin^ basic conditions such as an alkali metal hydroxide, e.g. NaOH, KOH, in particular

Intermediates (14c) and (14e) can be processed further as described hereinafter.

Coupling of P3 and P2 building blocks

For P2 building blocks that have a pyrrolidine moiety, the P3 and P2 or P3 and P2-P1 building blocks are linked using a carbamate forming reaction following the procedures described above for the coupling of (5 a) with (5b). A general procedure for coupling P2 blocks having a pyrrolidine moiety is represented in the following reaction scheme wherein L ³ is as specified above and L ⁴ is a group -O-PG ² , a group

In one embodiment L ⁴ in (15a) is a group -OPG ² , the PG ² group may be removed and the resulting acid coupled with cyclopropyl amino acids (12a) or (12b), yielding intermediates (12d) or (12e) wherein L ² is a radical (d) or (e).

A general procedure for coupling P3 blocks with a P2 block or a with a P2-P1 block wherein the P2 is a cyclopentane or cyclopentene is shown in the following scheme.

The reactions in the above two schemes are conducted using the same procedures as described above for the reactions of (5 a) or (8a) with (5b) and in particular the above

reactions wherein L ⁴ is a group (d) or (e) correspond to the reactions of (5a) or (8a) with (5b), described above.

The building blocks Pl, Pl ', P2 and P3 used in the preparation of the compounds of formula (I) can be prepared starting from art-known intermediates. A number of such syntheses are described hereafter in more detail.

Synthesis of P2 building blocks

The P2 building blocks contain either a pyrrolidine, a cyclopentane, or a cyclopentene moiety substituted with a group -O-R ¹ .

P2 building blocks containing a pyrrolidine moiety can be derived from commercially available hydroxy proline.

The preparation of P ² building blocks that contain a cylopentane ring may be performed as shown in the scheme below.

The bicyclic acid (17b) can be prepared, for example, from 3,4-bis(methoxy- carbonyl)-cyclopentanone (17a), as described by Rosenquist et al. in Acta Chem. Scand. 46 (1992) 1127-1129. A first step in this procedure involves the reduction of the keto group with a reducing agent like sodium borohydride in a solvent such as methanol, followed by hydrolysis of the esters and finally ring closure to the bicyclic lactone (17b) using lactone forming procedures, in particular by using acetic anhydride in the presence of a weak base such as pyridine. The carboxylic acid functionality in (17b) can then be protected by introducing an appropriate carboxyl protecting group,

such as a group PG ² , which is as specified above, thus providing bicyclic ester (17c). The group PG ² in particular is acid-labile such as a t.butyl group and is introduced e.g. by treatment with isobutene in the presence of a Lewis acid or with di-tert-buty\ dicarbonate in the presence of a base such as a tertiary amine like dimethylamino- pyridine or triethylamine in a solvent like dichloromethane. Lactone opening of (17c) using reaction conditions described above, in particular with lithium hydroxide, yields the acid (17d), which can be used further in coupling reactions with Pl building blocks. The free acid in (17d) may also be protected, preferably with an acid protecting group PG ^2a that is selectively cleavable towards PG ² , and the hydroxy function may be converted to a group -OPG ¹ or to a group -O-R ¹ . The products obtained upon removal of the group PG ² are intermediates (17g) and (17i) which correspond to intermediates (13a) or (16a) specified above.

Intermediates with specific stereochemistry may be prepared by resolving the intermediates in the above reaction sequence. For example, (17b) may be resolved following art-known procedures, e.g. by salt form action with an optically active base or by chiral chromatography, and the resulting stereoisomers may be processed further as described above. The OH and COOH groups in (17d) are in cis position. Trans analogs can be prepared by inverting the stereochemistry at the carbon bearing the OH function by using specific reagents in the reactions introducing OPG ¹ or OR ¹ that invert the stereochemistry, such as, e.g. by applying a Mitsunobu reaction.

In one embodiment, the intermediates (17d) are coupled to Pl blocks (12b) or (12c), which coupling reactions correspond to the coupling of (13a) or (16a) with the same Pl blocks, using the same conditions. Subsequent introduction of a -O-R^substituent as described above followed by removal of the acid protection group PG ² yields intermediates (8a-l), which are a subclass of the intermediates (8a), or part of the intermediates (16a). The reaction products of the PG ² removal can be further coupled to a P3 building block. In one embodiment PG ² in (17d) is t.butyl which can be removed under acidic conditions, e.g. with trifluoroacetic acid.

An unsaturated P2 building block, i.e. a cyclopentene ring, may be prepared as illustrated in the scheme below.

A bromination-elimination reaction of 3,4-bis(methoxycarbonyl)cyclopentanone (17a) as described by Dolby et al. in J. Org. Chem. 36 (1971) 1277-1285 followed by reduction of the keto functionality with a reducting agent like sodium borohydride provides the cyclopentenol (19a). Selective ester hydrolysis using for example lithium hydroxide in a solvent like a mixture of dioxane and water, provides the hydroxy substituted monoester cyclopentenol (19b).

An unsaturated P2 building block wherein R ³ can also be other than hydrogen, may be prepared as shown in the scheme below.

(2Og) (2Oh) (2Oi)

Oxidation of commercially available 3-methyl-3-buten-l-ol (20a), in particular by an oxidizing agent like pyridinium chlorochromate, yields (20b), which is converted to the corresponding methyl ester, e.g. by treatment with acetyl chloride in methanol, followed by a bromination reaction with bromine yielding the α-bromo ester (20c). The latter can then be condensed with the alkenyl ester (2Oe), obtained from (2Od) by an ester forming reaction. The ester in (2Oe) preferably is a t.butyl ester which can be

prepared from the corresponding commercially available acid (2Od), e.g. by treatment with di-tert-buty{ dicarbonate in the presence of a base like dimethylaminopyridine. Intermediate (2Oe) is treated with a base such as lithium diisopropyl amide in a solvent like tetrahydrofuran, and reacted with (20c) to give the alkenyl diester (2Of). Cyclisation of (2Of) by an olefin metathesis reaction, performed as described above, provides cyclopentene derivative (2Og). Stereoselective epoxidation of (2Og) can be carried out using the Jacobsen asymmetric epoxidation method to obtain epoxide (2Oh). Finally, an epoxide opening reaction under basic conditions, e.g. by addition of a base, in particular DBN (l,5-diazabicyclo-[4.3.0]non-5-ene), yields the alcohol (2Oi). Optionally, the double bond in intermediate (2Oi) can be reduced, for example by catalytic hydrogenation using a catalyst like palladium on carbon, yielding the corresponding cyclopentane compound. The t.butyl ester may be removed to the corresponding acid, which subsequently is coupled to a Pl building block.

The -O-R ¹ group can be introduced on the pyrrolidine, cyclopentane or cyclopentene rings at any convenient stage of the synthesis of the compounds according to the present invention. One approach is to first introduce the R ¹ group to the said rings and subsequently add the other desired building blocks, i.e. Pl (optionally with the Pl ' tail) and P3, followed by the macrocycle formation. Another approach is to couple the building blocks P2, bearing no -O-R ¹ substituent, with each Pl and P3, and to add the -O-R ¹ group either before or after the macrocycle formation. In the latter procedure, the P2 moieties have a hydroxy group, which may be protected by a hydroxy protecting group PG ¹ .

The -O-R ¹ -groups can be introduced on building blocks P2 by reacting hydroxy substituted intermediates (21a) or (21b) with intermediates (4b) as described above for the synthesis of (I) starting from (4a). These reactions are represented in the schemes below, wherein L ² is as specified above and L ⁵ and L ^5a independently from one another, represent hydroxy, a carboxyl protecting group -OPG ² or -OPG ^2a , or L ⁵ may also represent a Pl group such as a group (d) or (e) as specified above, or L ^5a may also represent a P3 group such as a group (b) as specified above The groups PG ² and PG ^2a are as specified above. Where the groups L ⁵ and L ^5a are PG ² or PG ^2a , they are chosen such that each group is selectively cleavable towards the other. For example, one of L ⁵ and L ^5a may be a methyl or ethyl group and the other a benzyl or t.butyl group.

In one embodiment in (21a), L ² is PG and L ⁵ is -OPG ² , or in (2Id), L ^5a is -OPG ² and L ⁵ is -OPG ^Z and the PG ^Z groups are removed as described above.

(21b-1 ) (21 c)

In another embodiment the group L ² is BOC, L ⁵ is hydroxy and the starting material (21a) is the commercially available BOC-hydroxyproline, or any other stereoisomeric form thereof, e.g. BOC-L-hydroxyproline, in particular the trans isomer of the latter. Where L ⁵ in (21b) is a carboxyl-protecting group, it may be removed following procedures described above to (21c). In still another embodiment PG in (2Ib-I) is Boc and PG ² is a lower alkyl ester, in particular a methyl or ethyl ester. Hydrolysis of the latter ester to the acid can be done by standard procedures, e.g. acid hydrolysis with hydrochloric acid in methanol or with an alkali metal hydroxide such as NaOH, in particular with LiOH. In another embodiment, hydroxy substituted cyclopentane or cyclopentene analogs (2Id) are converted to (2Ie), which, where L ⁵ and L ^5a are -OPG ²

or -OPG ^a , may be converted to the corresponding acids (2If) by removal of the group PG ² . Removal of PG ^2a in (2Ie-I) leads to similar intermediates.

The intermediates (4b) are art-known compounds or can be prepared following art-known methods using known starting materials.

Intermediates (4b), which are quinoline derivatives, may be prepared as shown in the scheme below. Such intermediates (4b) for example are those wherein R ¹ is a radical (d-1), (d-2), (d-3), (d-4), (d-4-a), (d-5) or (d-5-a) as specified above

Friedel-Craft acylation of a 3 -methoxy aniline (22a), available either commercially or via art-known procedures, using an acylating agent such as acetyl chloride or the like, in the presence of one or more Lewis acids such as boron trichloride or aluminium trichloride, in a solvent like dichloromethane, provides (22b). Coupling of (22b) with 4-isopropyl-thiazole-2-carboxylic acid (22c), preferably under basic conditions, such as in pyridine, in the presence of an activating agent for the carboxylate group, for instance POCI3, followed by ring closure and dehydration under basic conditions like potassium tert-butoxide in tert-butanol yields quinoline derivative (4b-l). The latter can be converted to (4b-2) wherein LG is a leaving group, e.g. by reaction of (4b- 1) with a halogenating agent, for example phosphoryl chloride or the like, or by reaction of (4b-l) with an arylsulfonyl chloride, e.g. with tosyl chloride.

Substituted anilines (22a) are available commercially or may be prepared from a suitable substituted benzoic acid (23 a), which is reacted with diphenylphosphorylazide at increased temperature and subsequently treated with a Ci_4alkanol, in particular t.butanol, affording Ci_4alkoxycarbonylamines such as compound (23b). Deprotection of compound (23b) yields substituted anilines (22a).

(23a) (23b) (22a)

Alternatively, substituted anilines (22a) may be prepared from the corresponding substituted nitrobenzenes by reducing the latter with elemental zinc, tin or iron in the presence of an acid.

A variety of carboxylic acids with the general structure (22c) can be used in the above synthesis. These acids are available either commercially or can be prepared via art-known procedures. An example of the preparation of 2-(substituted)aminocarboxy- aminothiazole derivatives (22c- 1), following the procedure described by Berdikhina et al. in Chem. Heterocycl. Compd. (Engl. Transl.) (1991), 427-433, is shown the following reaction scheme which illustrates the preparation of 2-carboxy-4-isopropyl-

Ethyl thiooxamate (24a) is reacted with the β-bromoketone (24b) to form the thiazolyl carboxylic acid ester (24c) which is hydro lyzed to the corresponding acid (22c- 1). The ethyl ester in these intermediates may be replaced by other carboxyl protecting groups PG ² , as defined above. In the above scheme R ^lf is as defined above and in particular is Ci_ ₄ alkyl, more in particular i.propyl.

The bromoketone (24b) may be prepared from 3-methyl-butan-2-one (MIK) with a sililating agent (such as TMSCl) in the presence of a suitable base (in particular LiHMDS) and bromine.

Intermediates (22b) having a methoxy substituent, said intermediates being represented by formula (22b- 1), may be prepared as described by Brown et al. J. Med. Chem. 1989, 32, 807- 826, or as outlined in the following scheme.

Starting materials ethyl acetylacetate and ethoxymethylene malononitrile, which are commercially available, are reacted in the presence of a suitable base, such as sodium ethoxide, and a solvent, such as ethanol and the like. This reaction affords intermediate (25a). The latter is hydrolyzed, e.g. with a base such as an alkali metal hydroxide, e.g. NaOH or LiOH, in a suitable solvent such as ethano I/water to produce (25b). Decarboxylation of intermediate (25b) to intermediate (25 c) is performed at increased temperature, preferably in the presence of a basic solvent such as quinoline. Methylation of intermediate (25c), in particular with a methylating agent such as MeI in the presence of a suitable base (e.g. K2CO3) in a suitable solvent (such as DMF and the like) yields (25d). The latter is reacted with a Grignard reagent such as MeMgBr in the presence of a suitable solvent (e.g. THF), followed by hydrolysis, for instance with aqueous HCl, affording intermediate (22b- 1).

The synthesis of further carboxylic acids (22c), in particular of substituted amino thiazole carboxylic acids (22c-2) is illustrated herebelow:

Thiourea (26c) with various substituents R ^4a , which in particular are d^alkyl, can be formed by reaction of the appropriate amine (26a) with tert-butylisothiocyanate in the presence of a base like diisopropylethylamine in a solvent like dichloromethane followed by removal of the tert-bvXy\ group under acidic conditions. Subsequent condensation of thiourea derivative (26c) with 3-bromopyruvic acid provides the thiazole carboxylic acid (22c-2).

Intermediates (4b) that are isoquinoline derivatives can be prepared using art-known procedures. For example, US 2005/0143316 provides diverse methods for the synthesis of isoquino lines as R'-OH or R'-LG intermediates. Methodology for the synthesis of such isoquinolines has been described by N. Briet et al., Tetrahedron, 2002, 5761 and is shown below, wherein R ^la , R ^lb and R ^lb are substituents on the isoquinoline moiety having the meanings defined herein for the substituents on the Regroup.

Cinnamic acid derivatives (27b) are converted to 1-chloroisoquino lines in a three-step process. The resulting chloroisoquinolines can be subsequently coupled to hydroxypyrrolidine, hydroxycyclopentane or hydro xycyclopentene derivatives as described herein. In a first step, the carboxyl group in cinnamic acid (27b) is activated, for example by treatment with a Ci_ ₆ alkyl (in particular methyl or ethyl) chloroformate in the presence of a base. The resulting mixed anhydride is then treated with sodium azide yielding the acyl azide (27c). Several other methods are available for the formation of acylazides from carboxylic acids, for example the carboxylic acid can be treated with diphenylphosphorylazide (DPPA) in an aprotic solvent such as methylene chloride, in the presence of a base. In a next step the acyl azide (27c) is converted to the corresponding isoquinolone (27d) by heating the acylazide, in a high boiling solvent such as diphenylether. The starting cinnamic acid derivatives are commercially available or can be obtained from the corresponding benzaldehydes (27a) by direct condensation with malonic acids or derivatives thereof, or by employing a Wittig reaction. The intermediate isoquino lones (21 ά) can be converted to the corresponding 1-chloro-isoquino lines by treatment with a halogenating agent such as phosphorous oxychloride.

Regroups which are isoquino lines can also be prepared following procedures as described in K. Hirao, R. Tsuchiya, Y. Yano, H. Tsue, Heterocycles 42(1) 1996, 415-422.

An alternative method for the synthesis of the isoquino line ring system is the Pomeranz-Fritsh procedure. This method begins with the conversion of a benzaldehyde derivative (28a) to a functionalized imine (28b), which then is converted to an isoquinoline ring system by treatment with acid at elevated temperature. This method is particularly useful for preparing isoquinoline intermediates that are substituted at the C8 position indicated by the asterisk. The intermediate isoquino lines (28c) can be converted to the corresponding 1-chloroquino lines (28e) in a two-step process. The first step comprises the formation of an isoquinoline N-oxide (28d) by treatment of isoquinoline (28c) with a peroxide such as meta-chloroperbenzoic acid in an appropriate solvent such as dichloromethane. Intermediate (28d) is converted to the corresponding 1-chloroisoquinoline by treatment with a halogenating agent such as phosphorous oxychloride.

Another method for the synthesis of the isoquinoline ring system is shown in the scheme below. ^a

In this process the anion form of ortho-alkylbenzamide derivative (29a) is obtained by treatment with a strong base such as tert-butyl lithium in a solvent such as THF and is subsequently condensed with a nitrile derivative, yielding isoquinoline (29b). The latter can be converted to the corresponding 1-chloroisoquinoline by the methods described above. R' and R" in (29a) are alkyl groups, in particular Ci_ ₄ alkyl groups, e.g. methyl or ethyl.

The following scheme shows an additional method for the synthesis of isoquino lines.

Intermediate (29a) is deprotonated using a strong base as described above. R' and R" are as specified above. The resulting intermediate anion is condensed with an ester (30a), obtaining ketone intermediate (30b). In a subsequent reaction the latter intermediate (30b) is reacted with ammonia or an ammonium salt, e.g. ammonium acetate, at elevated temperature, resulting in the formation of isoquino lone (29b).

Yet an additional method for the preparation of isoquino lines is illustrated in the following reaction scheme.

In the first step of this process an ortho-alkylarylimine derivative (31a) is subjected to deprotonation conditions (e.g. sec-butyl lithium, THF) and the resulting anion is condensed with an activated carboxylic acid derivative such as a Weinreb amide (31b). The resulting keto imine (31c) is converted to the isoquino line (3Id) by condensation with ammonium acetate at elevated temperatures. The thus obtained isoquino lines can be converted to the corresponding 1-chloroisoquino lines by the methods described herein.

The isoquino lines described herein, either as such or incorporated onto the hydroxyl-pyrrolidine, hydroxycyclopentane or hydroxycyclopentane moieties in the compounds of formula (I) or in any of the intermediates mentioned herein, can be further functionalized. An example of such functionalization is illustrated herebelow.

The above scheme shows the conversion of a l-chloro-6-fluoro-isoquinoline to the corresponding l-chloro-6-Ci_6alkoxy-isoquinoline moiety (32b), by treatment of (32a) with a sodium or potassium alkoxide in an alcohol solvent from which the alkoxide is derived. L ⁶ in the above scheme represents halo or a group

R represents Ci_6alkyl and LG is a leaving group. In one embodiment LG is fluoro. L ⁷ and L ⁸ represent various substituents that can be linked at these positions of the P2 moiety, in particular groups such as OL ⁵ , or L ⁸ may be a Pl group and L ⁷ a P3 group, or L ⁷ and L ⁸ taken together may form the remainder of the macrocyclic ring system of the compounds of formula (I).

The following scheme provides an example for the modification of isoquino lines by Suzuki reactions. These couplings can be employed to functionalize an isoquinoline at each position of the ring system provided said ring is suitably activated or functionalized, as for example with chloro.

(33e) (33f)

This sequence begins with 1-chloroisoquinoline (33a) which upon treatment with a peroxide such as metachloroperbenzoic acid is converted to the corresponding N-oxide (33b). The latter intermediate is converted to the corresponding 1,3-dichloro- isoquinoline (33c) by treatment with a halogenating agent, e.g. phosphorous oxychloride. Intermediate (33c) can be coupled with an intermediate (33d), wherein L ⁶ is a group PG where X is N, or L ⁶ is a group -COOPG ² where X is C, using methods described herein for introducing -O-R'-groups, to provide intermediate (33e). Intermediate (33e) is derivatized using a Suzuki coupling with an aryl boronic acid, in the presence of a palladium catalyst and a base, in a solvent such as THF, toluene or a dipolar aprotic solvent such as DMF, to provide the C3-arylisoquinoline intermediate (15f). Heteroarylboronic acids can also be employed in this coupling process to provide C3 -heteroarylisoquino lines.

Suzuki couplings of isoquino lines systems with aryl or heteroaryl groups can also be employed at a later synthesis stage in the preparation of compounds of formula (I). The isoquinoline ring systems can also be functionalized by employing other palladium catalyzed reactions, such as the Heck, Sonogashira or Stille couplings as illustrated for example in US 2005/1043316.

Synthesis of Pl building blocks

The cyclopropane amino acid used in the preparation of the Pl fragment is commercially available or can be prepared using art-known procedures.

The amino-vinyl-cyclopropyl ethyl ester (12b) may be obtained according to the procedure described in WO 00/09543 or as illustrated in the following scheme, wherein PG ² is a carboxyl protecting group as specified above:

(12b-1) (12b)

Treatment of commercially available or easily obtainable imine (34a) with 1,4-dihalo-butene in presence of a base produces (34b), which after hydrolysis yields cyclopropyl amino acid (12b), having the allyl substituent syn to the carboxyl group. Resolution of the enantiomeric mixture (12b) results in (12b-l). The resolution is performed using art-known procedures such as enzymatic separation; crystallization with a chiral acid; or chemical derivatization; or by chiral column chromatography. Intermediates (12b) or (12b-l) may be coupled to the appropriate proline derivatives as described above.

Introduction of a N-protecting group PG and removal of PG ² results in cyclopropyl amino acids (35s) which are converted to the amides (12c-l) or esters (12c-2), which are subgroups of the intermediates (12c), as outlined in the following reaction scheme, wherein R ^{2 a} , R ^{2 b} and PG are as specified above.

The reaction of (35 a) with (2b) can be performed following the procedures described above for the preparation of amidophosphates. This reaction yields intermediates (35b) or (35 c) from which the amino protecting group is removed by standard methods such as those described above. This in turn results in the desired intermediate (12c-l). Starting materials (35 a) may be prepared from the above mentioned intermediates (12b) by first introducing a N-protecting group PG and subsequent removal of the group PG ² .

Pl building blocks useful for the preparation of compounds according to general formula (I) wherein A is -C(=O)NH-P(=O)(OR ^8a )(R ^8b ), or -P(=O)(OR ^8a )(R ^8b ) a phosphonate can be prepared following procedures described in WO 2006/020276. In particular compounds of formula (I) wherein A is -P(=O)(OR ^8a )(R ^8b ) can be prepared as follows:

Starting material 32i is reacted with a base, in particular with CsOH, preferably in the presence of a phase transfer catalyst such as triethylbenzylammonium chloride, and 32j is added forming a cyclopropyl ring with a vinyl side chain, i.e. cyclopropyl phosphonate 32k. The phenyl-CH= protecting group is removed under acidic conditions (e.g. HCl in dichloromethane) yielding 321. The latter can be resolved in its stereoisomers using art-known methodolgy, e.g. by formation of a salt with an optically active acid, for example with dibenzoyl-L-tartaric acid, which after removal of the tartaric acid derivative yields 32m. Analogues other that the ethyl phophonates can be prepared from starting materials 32i having ester groups other than ethyl. The starting materials 32i are known materials or can easily be prepared using art-known methods.

Intermediates (12c-l) or (12c-2) in turn may be coupled to the appropriate pro line, cyclopentyl or cyclopenenyl derivatives (P2 moieties) as described above.

Synthesis of the P3 building blocks

The P3 building blocks are available commercially or can be generated according to methodologies known to the skilled in the art. One of these methodologies is shown in the scheme below and uses monoacylated amines, such as trifluoroacetamide or a Boc-protected amine.

In the above scheme, R together with the CO group forms a N-protecting group, in particular R is t-butoxy, trifluoromethyl; R ⁵ and n are as defined above and LG is a leaving group, in particular halogen, e.g. chloro or bromo.

Coupling of the appropriate P3 building block to P2-P or P2 moieties have been described above. Coupling of a P3 building block to Pl or P1-P2 moieties can be achieved via formation of a double bond, such as a Wittig synthesis or preferably by an olefin metathesis reaction as described herein above.

Compounds of formula (I) may be converted into each other following art-known functional group transformation reactions. For example, amino groups may be N-alkylated, nitro groups reduced to amino groups, a halo atom may be exchanged for another halo.

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, benzenecarbo- peroxoic acid or halo substituted benzenecarboperoxoic acid, e.g. 3-chlorobenzene- carboperoxoic acid, peroxoalkanoic acids, e.g. peroxoacetic acid, alkylhydroperoxides, e.g. tert-bvXy\ 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.

Pure stereochemically isomeric forms of the compounds of formula (I) may be obtained by the application of art-known procedures. Diastereomers may be separated by physical methods such as selective crystallization and chromatographic techniques, e.g., counter-current distribution, liquid chromatography and the like.

The compounds of formula (I) may be obtained as racemic mixtures of enantiomers which can be separated from one another following art-known resolution procedures. The racemic compounds of formula (I) that are sufficiently basic or acidic may be converted into the corresponding diastereomeric salt forms by reaction with a suitable chiral acid, respectively chiral base. Said diastereomeric salt forms are subsequently separated, for example, by selective or fractional crystallization and the enantiomers are liberated therefrom by alkali or acid. An alternative manner of separating the enantiomeric forms of the compounds of formula (I) involves liquid chromatography, in particular 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 may be synthesized by stereospecifϊc methods of preparation. These methods may advantageously employ enantiomerically pure starting materials.

In a further aspect, the present invention concerns a pharmaceutical composition comprising a therapeutically effective amount of a compound of formula (I) as specified herein, or a compound of any of the subgroups of compounds of formula (I) as specified herein, and a pharmaceutically acceptable carrier. A therapeutically effective amount in this context is an amount sufficient to act in a prophylactic way against, to stabilize or to reduce viral infection, and in particular HCV viral infection, in infected subjects or subjects being at risk of being infected. In still a further aspect, this invention relates to a process of preparing a pharmaceutical composition as specified herein, which comprises intimately mixing a pharmaceutically acceptable carrier with a therapeutically effective amount of a compound of formula (I), as specified herein, or of a compound of any of the subgroups of compounds of formula (I) as specified herein.

Therefore, the compounds of the present invention or any subgroup thereof 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 addition salt form or metal complex, 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, percutaneous Iy, 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, 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 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.

The compounds of the present invention may also be administered via oral 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, a solution being preferred. Any system developed for the delivery of solutions, suspensions or dry powders via oral inhalation or insufflation are suitable for the administration of the present compounds.

Thus, the present invention also provides a pharmaceutical composition adapted for administration by inhalation or insufflation through the mouth comprising a compound of formula (I) and a pharmaceutically acceptable carrier. Preferably, the compounds of the present invention are administered via inhalation of a solution in nebulized or aerosolized doses.

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, suppositories, powder packets, wafers, injectable solutions or suspensions and the like, and segregated multiples thereof.

The compounds of formula (I) show antiviral properties. Viral infections and their associated diseases treatable using the compounds and methods of the present invention include those infections brought on by HCV and other pathogenic flaviviruses such as Yellow fever, Dengue fever (types 1-4), St. Louis encephalitis, Japanese encephalitis, Murray valley encephalitis, West Nile virus and Kunjin virus. The diseases associated with HCV include progressive liver fibrosis, inflammation and necrosis leading to cirrhosis, end-stage liver disease, and HCC; and for the other pathogenic flaviruses the diseases include yellow fever, dengue fever, hemorraghic fever and encephalitis. A number of the compounds of this invention moreover are active against mutated strains of HCV. Additionally, many of the compounds of this invention show a favorable pharmacokinetic profile and have attractive properties in terms of bioavailabilty, including an acceptable half-life, AUC (area under the curve) and peak values and lacking unfavourable phenomena such as insufficient quick onset and tissue retention.

The in vitro antiviral activity against HCV of the compounds of formula (I) can be tested in a cellular HCV replicon system based on Lohmann et al. (1999) Science

285:110-113, with the further modifications described by Krieger et al. (2001) Journal of Virology 75: 4614-4624 (incorporated herein by reference), which is further exemplified in the examples section. This model, while not a complete infection model for HCV, is widely accepted as the most robust and efficient model of autonomous HCV RNA replication currently available. Compounds exhibiting anti-HCV activity in this cellular model are considered as candidates for further development in the treatment of HCV infections in mammals. It will be appreciated that it is important to distinguish between compounds that specifically interfere with HCV functions from those that exert cytotoxic or cytostatic effects in the HCV replicon model, and as a consequence cause a decrease in HCV RNA or linked reporter enzyme concentration. Assays are known in the field for the evaluation of cellular cytotoxicity based for example on the activity of mitochondrial enzymes using fluorogenic redox dyes such as resazurin. Furthermore, cellular counter screens exist for the evaluation of non-selective inhibition of linked reporter gene activity, such as firefly luciferase. Appropriate cell types can be equipped by stable transfection with a luciferase reporter gene whose expression is dependent on a constitutively active gene promoter, and such cells can be used as a counter-screen to eliminate non-selective inhibitors.

Due to their antiviral properties, particularly their anti-HCV properties, the compounds of formula (I) or any subgroup thereof, iV-oxides, pharmaceutically acceptable addition salts, and stereochemically isomeric forms, are useful in the treatment of individuals infected with a virus, particularly a virus that is HCV , and for the prophylaxis of viral infections, in particular HCV infections. In general, the compounds of the present invention may be useful in the treatment of warm-blooded animals infected with viruses, in particular flaviviruses such as HCV.

The compounds of the present invention or any subgroup thereof may therefore be used as a medicine. Said use as a medicine or method of treatment comprises the systemic administration to virally infected subjects or to subjects susceptible to viral infections of an amount effective to combat the conditions associated with the viral infection, in particular HCV infection.

The present invention also relates to the use of the present compounds or any subgroup thereof in the manufacture of a medicament for the treatment or the prevention of a viral infection, particularly HCV infection.

The present invention furthermore relates to a method of treating a warm-blooded animal infected by a virus, or being at risk of infection by a virus, in particular by

HCV, said method comprising the administration of an anti- virally effective amount of a compound of formula (I), as specified herein, or of a compound of any of the subgroups of compounds of formula (I), as specified herein.

In general it is contemplated that an antiviral effective daily amount would be from

0.01 mg/kg to 500 mg/kg body weight, or from 0.1 mg/kg to 50 mg/kg body weight, or from 0.5 mg/kg to 5 mg/kg body weight. It may be appropriate to administer the required dose as two, three, four or more sub-doses at appropriate intervals throughout the day. Said sub-doses may be formulated as unit dosage forms, for example, containing 1 to 1000 mg, and in particular 5 to 200 mg of active ingredient per unit dosage form.

The invention also relates to a combination of a compound of formula (I), including a stereoisomeric form thereof, a pharmaceutically acceptable salt, or a pharmaceutically acceptable solvate thereof, and another antiviral compound, in particular another anti-HCV compound. The term "combination" may relate to a product containing (a) a compound of formula (I), as specified above, and (b) optionally another anti-HCV

compound, as a combined preparation for simultaneous, separate or sequential use in treatment of HCV infections.

Anti-HCV compounds that can be used in such combinations include agents selected from an HCV polymerase inhibitor, an HCV protease inhibitor, an inhibitor of another target in the HCV life cycle, and an immunomodulatory agent, and combinations thereof. HCV polymerase inhibitors include, NM283 (valopicitabine), R803, JTK- 109, JTK-003, HCV-371, HCV-086, HCV-796 and R-1479. Inhibitors of HCV proteases (NS2-NS3 inhibitors and NS3-NS4A inhibitors) include the compounds of WO 02/18369 (see, e.g., page 273, lines 9-22 and page 274, line 4 to page 276, line 11); BILN-2061, VX-950, GS-9132 (ACH-806), SCH-503034, and SCH-6. Further agents that can be used are those disclosed in WO 98/17679, WO 00/056331 (Vertex); WO 98/22496 (Roche); WO 99/07734, (Boehringer Ingelheim ), WO 2005/073216, WO 2005/073195 (Medivir) and structurally similar agents.

Inhibitors of other targets in the HCV life cycle, including NS3 helicase; metallo-protease inhibitors; antisense oligonucleotide inhibitors, such as ISIS- 14803, AVI-4065 and the like; siRNA's such as SIRPLEX- 140-N and the like; vector-encoded short hairpin RNA (shRNA); DNAzymes; HCV specific ribozymes such as heptazyme, RPI.13919 and the like; entry inhibitors such as HepeX-C, HuMax-HepC and the like; alpha glucosidase inhibitors such as celgosivir, UT-231B and the like; KPE-02003002; and BIVN 401.

Immunomodulatory agents include, natural and recombinant interferon isoform compounds, including α-interferon, β-interferon, γ-interferon, ω-interferon and the like, such as Intron A®, Roferon-A®, Canferon-A300®, Advaferon®, Infergen®, Humoferon®, Sumiferon MP®, Alfaferone®, IFN-beta®, Feron® and the like; polyethylene glycol derivatized (pegylated) interferon compounds, such as PEG interferon-α-2a (Pegasys®), PEG interferon-α-2b (PEG-Intron®), pegylated IFN-α-conl and the like; long acting formulations and derivatizations of interferon compounds such as the albumin- fused interferon albuferon α and the like; compounds that stimulate the synthesis of interferon in cells, such as resiquimod and the like; interleukins; compounds that enhance the development of type 1 helper T cell response, such as SCV-07 and the like; TOLL-like receptor agonists such as CpG-IOlOl (actilon), isatoribine and the like; thymosin α-1; ANA-245; ANA-246; histamine dihydrochloride; propagermanium; tetrachlorodecaoxide; ampligen; IMP-321; KRN-7000; antibodies, such as civacir, XTL-6865, and the like; and prophylactic and therapeutic vaccines such as InnoVac C, HCV E1E2/MF59, and the like.

Other antiviral agents include, ribavirin, amantadine, viramidine, nitazoxanide; telbivudine; NOV-205; taribavirin; inhibitors of internal ribosome entry; broad-spectrum viral inhibitors, such as IMPDH inhibitors, and mycophenolic acid and derivatives thereof, and including, but not limited to VX-950, merimepodib (VX-497), VX-148, and/or VX-944); or combinations of any of the above.

Particular agents for use in said combinations include interferon-α (IFN-α), pegylated interferon-α or ribavirin, as well as therapeutics based on antibodies targeted against HCV epitopes, small interfering RNA (Si RNA), ribozymes, DNAzymes, antisense RNA, small molecule antagonists of for instance NS3 protease, NS3 helicase and NS5B polymerase.

In another aspect there are provided combinations of a compound of formula (I) as specified herein and an anti-HIV compound. The latter preferably are those HIV inhibitors that have a positive effect on drug metabolism and/or pharmacokinetics that improve bioavailabilty. An example of such an HIV inhibitor is ritonavir. As such, this invention further provides a combination comprising (a) an HCV NS3/4a protease inhibitor of formula (I) or a pharmaceutically acceptable salt thereof; and (b) ritonavir or a pharmaceutically acceptable salt thereof. The compound ritonavir, its pharmaceutically acceptable salts, and methods for its preparation are described in WO 94/14436. US 6,037,157, and references cited therein: US 5,484,801, US 08/402,690, WO95/07696, and WO95/09614, disclose preferred dosage forms of ritonavir. One embodiment relates to a combination comprising (a) an HCV NS3/4a protease inhibitor of formula (I) or a pharmaceutically acceptable salt thereof; and (b) ritonavir or a pharmaceutically acceptable salt thereof; optionally comprising an additional anti-HCV compound selected from the compounds mentioned above.

The invention also concerns a process for preparing a combination as described herein, comprising the step of combining a compound of formula (I), as specified above, and another agent, such as an antiviral, including an anti-HCV or anti-HIV agent, in particular those mentioned above.

The said combinations may find use in the manufacture of a medicament for treating HCV infection, or another pathogenic flavi- or pestivirus, in a mammal infected with therewith, said combination in particular comprising a compound of formula (I), as specified above and interferon-α (IFN-α), pegylated interferon-α, or ribavirin. Or the invention provides a method of treating a mammal, in particular a human, infected with

HCV, or another pathogenic flavi- or pestivirus, comprising the administration to said mammal of an effective amount of a combination as specified herein. In particular, said treating comprises the systemic administration of the said combination and an effective amount is such amount that is effective in treating the clinical conditions associated with HCV infection.

In one embodiment the above-mentioned combinations are formulated in the form of a pharmaceutical composition that includes the active ingredients described above and a carrier, as described above. Each of the active ingredients may be formulated separately and the formulations may be co-administered, or one formulation containing both and if desired further active ingredients may be provided. In the former instance, the combinations may also be formulated as a combined preparation for simultaneous, separate or sequential use in HCV therapy. The said composition may take any of the forms described above. In one embodiment, both ingredients are formulated in one dosage form such as a fixed dosage combination. In a particular embodiment, the present invention provides a pharmaceutical composition comprising (a) a therapeutically effective amount of a compound of formula (I), including a stereoisomeric form thereof, or a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable solvate thereof, and (b) a therapeutically effective amount of ritonavir or a pharmaceutically acceptable salt thereof, and (c) a carrier.

The individual components of the combinations of the present invention can be administered separately at different times during the course of therapy or concurrently in divided or single combination forms. The present invention is meant to embrace all such regimes of simultaneous or alternating treatment and the term "administering" is to be interpreted accordingly. In a preferred embodiment, the separate dosage forms are administered simultaneously.

In one embodiment, the combinations of the present invention contains an amount of ritonavir, or a pharmaceutically acceptable salt thereof, that is sufficient to clinically improve the bioavailability of the HCV NS3/4a protease inhibitor of formula (I) relative to the bioavailability when said HCV NS3/4a protease inhibitor of formula (I) is administered alone. Or, the combinations of the present invention contains an amount of ritonavir, or a pharmaceutically acceptable salt thereof, which is sufficient to increase at least one of the pharmacokinetic variables of the HCV NS3/4a protease inhibitor of formula (I) selected from ti/2, Cm _1n , C _max , C _ss , AUC at 12 hours, or AUC at 24 hours, relative to said at least one pharmacokinetic variable when the HCV NS3/4a protease inhibitor of formula (I) is administered alone.

The combinations of this invention can be administered to humans in dosage ranges specific for each component comprised in said combinations, e.g the compound of formula (I) as specified above, and ritonavir or a pharmaceutically acceptable salt, may have dosage levels in the range of 0.02 to 5.0 g/day.

The weight ratio of the compound of formula (I) to ritonavir may be in the range of from about 30:1 to about 1 :15, or about 15: 1 to about 1 : 10, or about 15: 1 to about 1 : 1, or about 10: lto about 1 : 1, or about 8: 1 to about 1 : 1, or about 1 : 5 to 1 : 1 to about 5 : 1 , or about 3 : 1 to about 1 : 1 , or about 2:1 to 1 :1. The compound formula (I) and ritonavir may be co-administered once or twice a day, preferably orally, wherein the amount of the compound of formula (I) per dose is from about 1 to about 2500 mg, or about 50 to about 1500 mg, or about 100 to about 1000 mg, or about 200 to about 600 mg, or about 100 to about 400 mg; and the amount of ritonavir per dose is from 1 to about 2500 mg, or about 50 to about 1500 mg, or about 100 to about 800 mg, or about 100 to about 400 mg, or 40 to about 100 mg of ritonavir.

Examples

The following examples are intended to illustrate the present invention and not to limit it thereto.

Example 1 : Synthesis of 4-hydroxy-2-(4-isopropylthiazole-2-yl)-7-methoxy-8- methylquinoline (6)

Step 1 : synthesis of A/-(fer^butyloxycarbonyl)-3-methoxy-2-methylaniline (2) Triethylamine (42.4 niL, 302 mmol) was added to a suspension of 3-methoxy- 2-methylbenzoic acid (45.6 g, 274 mmol) in dry toluene (800 mL). A clear solution was obtained. Then, dppa (65.4 mL, 302 mmol) in toluene (100 mL) was slowly added.

After 1 h at room temperature, the reaction mixture was successively heated at 50 ⁰ C for 0.5 h, at 70 ⁰ C for 0.5 h then at 100 ⁰ C for 1 h. To this solution, t-BuOH (30.5 g, 411 mmol) in toluene (40 mL) was added at 100 ⁰ C and the resulting mixture was refluxed for 7h. The solution was cooled to room temperature then successively washed with water, 0.5 N HCl, 0.5 N NaOH and brine, dried (Na ₂ SO ₄ ), and evaporated to give 67 g of the target product: m/z = 237 (M) ⁺ .

Step 2: synthesis of 3-methoxy-2-methylaniline (3)

TFA (40.7 mL, 548 mmol) was added to a solution of jV-(teτt-butyloxycarbonyl)- 3-methoxy-2-methylaniline, in dichloromethane (500 mL). After 2 h at room temperature, TFA (40.7 mL, 548 mmol) was added and the resulting mixture was stirred at room temperature overnight. Then, volatiles were evaporated. The residue was triturated with toluene (100 mL) and diisopropylether (250 mL), filtered off and washed with diisopropyl ether (100 mL) to give 56.3 g of the title product as a TFA salt: m/z = 138 (M+H) ⁺ . The TFA salt was transformed to the free aniline by treatment with NaHCO ₃ .

Step 3: synthesis of (2-amino-4-methoxy-3-methylphenyl)(methyl)ketone (4)

A solution Of BCl ₃ (1.0 M, 200 mL, 200 mmol) in CH ₂ Cl ₂ was slowly added under nitrogen to a solution of 3-methoxy-2-methylaniline (26.0 g, 190 mmol) in xylene (400 mL). The temperature was monitored during the addition and was kept below 10 ⁰ C. The reaction mixture was stirred at 5°C for 0.5 h. Then, dry acetonitrile (13 mL, 246 mmol) was added at 5°C. After 0.5 h at 5°C, the solution was transferred into a dropping funnel and slowly added at 5°C to a suspension OfAlCl ₃ (26.7 g, 200 mmol) in CH ₂ Cl ₂ (150 mL). After 45 min at 5°C, the reaction mixture was heated at 70 ⁰ C under a nitrogen stream. After evaporation Of CH ₂ Cl ₂ , the temperature of the reaction mixture reached 65°C. After 12 h at 65°C, the reaction mixture was cooled at 0 ⁰ C, poured onto ice (300 g), and slowly heated to reflux for 7h. After 2 days at room temperature, 6 N NaOH (50 mL) was added. The pH of the resulting solution was 2-3. The xylene layer was decanted. The organic layer was extracted with CH ₂ Cl ₂ . The xylene and CH ₂ Cl ₂ layers were combined, successively washed with water, IN NaOH, and brine, dried (Na ₂ SO ₄ ) and evaporated. The residue was triturated in diisopropyl

ether at O ⁰ C, filtered off and washed with diisopropylether to give 13.6 g (40 %) of the title product as a yellowish solid: m/z = 180 (M+H) ⁺ .

Step 4: synthesis of 2'-[[(4-isopropylthiazole-2-yl)(oxo)methyllaminol-4'-methoxy -3'- methylacetophenone (5)

A solution of (2-amino-4-methoxy-3-methylphenyl)(methyl)ketone (18.6 g, 104 mmol) in dioxane (50 rnL) was added under nitrogen to a suspension of 4-isopropylthiazole- 2-carbonyl chloride in dioxane (250 mL). After 2 h at room temperature, the reaction mixture was concentrated to dryness. Then, the residue was partitioned between an aqueous solution of NaHCOs and AcOEt, organic layer was washed with brine, dried (Na ₂ SO ₄ ), and evaporated. The residue was triturated in diisopropyl ether, filtered off and washed with diisopropyl ether to give 30.8 g (90 %) of the title product 5.

Step 5: synthesis of 4-hydroxy-2-(4-isopropylthiazole-2-yl)-7-methoxy-8-methyl- quinoline (6)

Potassium tert-butoxide (21.8 g, 195 mmol) was added to a suspension of 2'-[[(4-iso- propylthiazo le-2-yl)(oxo)methyl] amino] -4 ' -methoxy-3 ' -methylacetophenone (5, 30.8 g, 92.7 mmol) in tert-butanol. The resulting reaction mixture was heated at 100 ⁰ C overnight. Then, the reaction mixture was cooled at room temperature and diluted with ether (100 mL). The precipitate was filtered off and washed with Et ₂ O to give a powder (fraction A). The mother liquor was concentrated in vacuo, triturated in ether, filtered off, and washed with ether to give a powder (fraction 2). Fractions 1 and 2 were mixed and poured into water (250 mL). The pH of the resulting solution was adjusted to 6-7 (control with pH paper) with HCl IN. The precipitate was filtered off, washed with water and dried. Then, the solid was triturated in diisopropyl ether, filtered off and dried to give 26 g (88%) of the title product 6 as a brownish solid: m/z = 315 (M+H) ⁺ .

Example 2: Synthesis of l-{[2-(Hex-5-enyl-methyl-carbamoyl)-4-hydroxy- cyclopentanecarbonyll-amino|-2-vinyl-cyclopropanecarboxylic acid ethyl ester (13)

Step 1

Sodium hydride (1.05 eq) was slowly added at O ⁰ C to a solution of JV-methyltrifluoro- acetamide (25 g) in DMF (140 rnL). The mixture was stirred for Ih at room temperature under nitrogen. Then, a solution of bromohexene (32,1 g) in DMF (25 mL) was added dropwise and the mixture was heated to 70 ⁰ C for 12 hours. The reaction mixture was poured on water (200 mL) and extracted with ether (4 x 50 mL), dried (MgSO ₄ ), filtered and evaporated to give 35 g of the target product 7 as a yellowish oil which was used without further purification in the next step.

Step 2: Synthesis of (hex-5-enyl)(methyl)amine (8)

A solution of potassium hydroxide (187.7 g) in water (130 mL) was added dropwise to a solution of 7 (35 g) in methanol (200 mL). The mixture was stirred at room temperature for 12 hours. Then, the reaction mixture was poured on water (100 mL) and extracted with ether (4 x 50 mL), dried (MgSO ₄ ), filtered and the ether was distilled under atmospheric pressure. The resulting oil was purified by distillation under vacuum (13 mm Hg pressure, 50 ⁰ C) to give 7,4 g (34 %) of the title product 8 as a colourless oil: ¹ H-NMR (CDCl ₃ ): δ 5.8 (m, IH), 5 (ddd, J= 17.2 Hz, 3.5 Hz, 1.8 Hz, IH), 4.95 (m, IH), 2.5 (t, J= 7.0 Hz, 2H), 2.43 (s, 3H), 2.08 (q, J= 7.0 Hz, 2H), 1.4 (m, 4H), 1.3 (br s, IH).

Step 3

3-Oxo-2-oxa-bicyclo[2.2.1]heptane-5-carboxylic acid 9 (500 mg, 3.2 mmol) in 4 mlDMF was added at 0 ⁰ C to HATU (1.34 g, 3.52 mmol) and JV-methylhex-5- enylamine (435 mg, 3.84 mmol) in DMF (3 mL), followed by DIPEA. After stirring for 40 min at 0 ⁰ C, the mixture was stirred at room temperature for 5 h. Then, the solvent was evaporated, the residue dissolved in EtOAc (70 mL) and washed with saturated NaHCO ₃ (10 mL). The aqueous layer was extracted with EtOAc (2 x 25 mL). The organic phases were combined, washed with saturated NaCl (20 mL), dried (Na ₂ SO ₄ ), and evaporated. Purification by flash chromatography (EtO Ac/petroleum ether, 2:1) afforded 550 mg (68%) of the target product 10 as a colorless oil: m/z = 252 (M+H) ⁺ .

Step 4

A solution of LiOH (105 mg in 4 mlof water) was added at 0 ⁰ C to the lactone amide 10. After Ih, the conversion was completed (HPLC). The mixture was acidified to pH 2 - 3 with IN HCl, extracted with AcOEt, dried (MgSO ₄ ), evaporated, co-evaporated with toluene several times, and dried under high vacuum overnight to give 520 mg (88%) of the target product 11: m/z = 270 (M+H) ⁺ .

Step 5

The l-(amino)-2-(vinyl)cyclopropanecarboxylic acid ethyl ester hydrochloride 12 (4.92 g, 31.7 mmol) and HATU (12.6 g, 33.2 mmol) were added to 11 (8.14 g, 30.2 mmol). The mixture was cooled in an ice bath under argon, and then DMF (100 mL) and DIPEA (12.5 mL, 11.5 mmol) were successively added. After 30 min at 0 ⁰ C, the solution was stirred at room temperature for an additional 3 h. Then, the reaction mixture was partitioned between EtOAc and water, washed successively with 0.5 N HCl (20 mL) and saturated NaCl (2 x 20 mL), and dried (Na ₂ SO ₄ ). Purification by flash chromatography (AcOEt/CH ₂ Cl ₂ /Petroleum ether, 1 :1 :1) afforded 7.41 g (60%) of the target product 13 as a colorless oil: m/z = 407 (M+H) ⁺ .

Example 3: Preparation of {17-[2-(4-isopropylthiazole-2-yl)-7-methoxy-8- methylquinolin-4-yloxy]- 13-methyl-2 J4-dioxo-3 J 3-diazatricyclo[ 13.3.0.0 ⁴ ' ⁶ ]octadec-

Step 1

DIAD (1.02 niL, 5.17 mmol) was added at -15°C under nitrogen atmosphere to a solution of 13 (1.5 g, 3.69 mmol), quinoline 6 (1.39 g, 4.43 mmol) and triphenyl- phosphine (1.26 g, 4.80 mmol) in dry THF (40 mL). After 4.5 h, at -15°C, the reaction mixture was partitioned between ice-cold water and AcOEt, dried (Na ₂ SO ₄ ) and evaporated. The crude material was purified by flash column chromatography (gradient of petroleum AcOEt/CH ₂ Cl ₂ , 1 :9 to 2:8) to give 1.45 g (56 %) of the target product 14: m/z = 782 (M+H) ⁺ ; TLC (EtOAc): Rf= 0.35.

Step 2

A solution of 14 (1.07 g, 1.524 mmol) and Hoveyda-Grubbs 1 ^st generation catalyst (33 mg, 0.03 eq) in dried and degassed 1 ,2-dichloroethane (900 mL) was heated at

75°C under nitrogen for 12 h. Then, the solvent was evaporated and the residue purified by silica gel chromatography (25% EtOAc in CH ₂ Cl ₂ ). 620 mg (60%) of pure macrocycle 15 were obtained, m/z = 674 (M+H) ⁺ . ¹ H NMR (CDCl ₃ ): 1.18-1.39 (m,

12H), 1.59 (m, IH), 1.70-2.08 (m, 5H), 2.28 (m, IH), 2.38 (m, IH), 2.62 (m, 2H), 2.68 (s, 3H), 2.83 (m, IH), 3.06 (s, 3H), 3.19 (sept, J= 6.7 Hz, IH), 3.36 (m, IH), 3.83 (m, IH), 3.97 (s, 3H), 4.09 (m, 2H), 4.65 (td, J= 4 Hz, 14 Hz, IH), 5.19 (dd, J= 4 Hz, 10 Hz, IH), 5.31 (m, IH), 5.65 (td, J= 4 Hz, 8 Hz, IH), 7.00 (s, IH), 7.18 (s, IH), 7.46 (d, J= 9 Hz, IH), 7.48 (s, IH), 8.03 (d, J= 9 Hz, IH).

Step 3

A solution of lithium hydroxide (1.65 g, 38.53 mmol) in water (15 rnL) was added to a stirred solution of ester 15 (620 mg, 0.920 mmol) in THF (30 mL) and MeOH (20 mL). After 16 h at room temperature, the reaction mixture was quenched with NH ₄ Cl sat., concentrated under reduced pressure, acidified to pH 3 with HCl IN and extracted with CH ₂ Cl ₂ , dried (MgSO ₄ ) and evaporated to give 560 mg (88%) of carboxylic acid 16. m/z = 647 (M+H) ⁺ . ¹ H NMR (CDCl ₃ ): 1.11-1.40 (m, 8H), 1.42-1.57 (m, 2H), 1.74 (m, 2H), 1.88-2.00 (m, 2H), 2.13 (m, IH), 2.28 (m, IH), 2.40 (m, IH), 2.59 (m, 2H), 2.67 (s, 3H), 2.81 (m, IH), 2.97 (s, 3H), 3.19 (m, IH), 3.31 (m, IH), 3.71 (m, IH), 3.96 (s, 3H), 4.56 (dt, J= 4 Hz, 12 Hz, IH), 5.23 (m, 2H), 5.66 (m, IH), 7.01 (s, IH), 7.10 (s, IH), 7.22 (d, J= 10 Hz, IH), 7.45 (s, IH), 8.00 (d, J= 10 Hz, IH).

Step 4 A solution of 17-[2-(4-isopropylthiazole-2-yl)-7-methoxy-8-methylquinolin- 4-yloxy]- 13-methyl-2,14-dioxo-3,13-diazatricyclo[13.3.0.04,6]octadec- 7-ene-4-carboxylic acid 16 (138.3 mg, 0.214 mmol) prepared according to the procedure described above, and carbonyldiimidazole (96.9 mg, 0.598 mmol) in dry THF (5 mL) was stirred at reflux under nitrogen for 2h. The reaction mixture was cooled down at room temperature and concentrated under reduced pressure. The residue was partitioned between EtOAc and HCl 1 N, the organic layer was washed with brine, dried (Na2SO ₄ ) and evaporated. Then the solid was triturated in i-Pr ether to get 16' as a white powder: m/z = 629 (M+H) ⁺ . ¹ H NMR (CDCl ₃ ): 0.99-1.00 (m, IH), 1.20-1.35 (m, 2H), 1.39 (d, J = 6.9 Hz, 6H), 1.55-1.7 (m, IH), 1.9-2 (m, 2H), 2.15-2.25 (m, 2H), 2.3-2.60 (m, 4H), 2.68 (s, 3H), 2.71-2.82 (m, IH), 2.82-2.9 (m, IH), 3.08 (s, 3H), 3.1-3.2 (m, IH), 3.4-3.5 (m, IH), 3.65-3.71 (m, IH), 3.91 (s, 3H), 4.28-4.4 (m, IH), 5.32-5.46 (m, 2H), 5.85-5.95 (m, IH), 7.00 (s, IH), 7.22 (d, J= 9.2 Hz, IH), 7.45 (s, IH), 8.09 (d, J= 9.2 Hz, IH).

Step 5

Phosphoramidate (144 mg, 0.784 mmol) and DBU (490 μL, 2.86 mmol) were successively added to a stirred solution of intermediate 16' (360 mg, 0.574 mmol) in dry THF (10 mL). The resulting mixture was heated to reflux. After 2h, the solution

was cooled down to room temperature, then concentrated under reduced pressure. The residue was purified by column chromatography to give the target product 17: m/z = 754 (M+H) ⁺ .

Example 5

Step l

To a solution of 9 (857 mg, 5.5 mmol), in DMF (14 ml) and DCM (25 ml) at room temperature, was added 12 (1.15 g, 6.0 mmol), HATU (2.29 g, 6.0 mmol) and DIPEA (3.82 ml, 22 mmol). The reaction was stirred under N2-atmosphere at ambient temperature for 1 h. LC/MS analysis showed complete conversion, and the reaction mixture was concentrated in vacuo. The residue was re-dissolved in DCM (100 ml) and 0.1 M HCl (aqueous) and the layers separated. The organic phase was washed with NaHCO ₃ (aqueous) and brine, dried (MgSO ₄ ) and filtered. Removal of the solvent in vacuo afforded the target compound 18 (1.6 g, 99%). LC/MS (Method A): t _R =2.46 min, >95%, m/z (ESI ⁺ ) = 294(MH ⁺ )

Step 2 To a solution of 18 (800 mg, 2.73 mmol) in water (15 ml) in a 20 ml microwave reaction vessel was added DIPEA (1.2 ml, 6.8 mmol) and a stir bar. The reaction vessel was sealed and the immiscible slurry was shaken vigorously before insertion in the microwave cavity. After 1 min of pre-stirring, the reaction was irradiated for 40 min to a set temperature of 100 ⁰ C. After cooling to 40 ⁰ C, the transparent solution was concentrated in vacuo, and the residual brown oil co-evaporated 3 times with acetonitrile to remove any residual water. The crude product 19, in the form of a DIPEA salt, was immediately taken forward to the next step. LC/MS (Method A): t _R =1.29 min, >95%, m/z (ESI ⁺ )= 312(MH ⁺ ).

Step 3

The crude compound 19 (5.5 mmol) was dissolved in DCM (50 ml) and DMF (14 ml) followed by addition of HATU (2.09 g, 5.5 mmol), 8 (678 mg, 6.0 mmol) and DIPEA (3.08 ml, 17.5 mmol) at room temperature. The reaction was stirred at ambient temperature for 1 h. LC/MS analysis showed complete conversion and the reaction mixture was concentrated in vacuo. The residue was re-dissolved in ethyl acetate

(100 ml) and the organic layer washed with 0.1 M HCl (aqueous), K2CO3 (aqueous) and brine, dried (MgSO ₄ ) and filtered. Evaporation of the solvent in vacuo gave an oil which was purified by flash chromatography (Silica, ethyl acetate/methanol) to afford the target compound 20 (1.65 g, 74%). TLC (Silica): methanol/ethyl acetate 5:95, R _f =0.5; LC/MS (Method A): t _R =3.44 min, >95%, m/z (ESI ⁺ )= 407(MH ⁺ ).

Step 4

To a stirred solution of the l-{[2-(Hex-5-enylmethylcarbamoyl)-4-hydroxycyclo- pentanecarbonyl]amino}-2-vinylcyclopropane carboxylic acid ethyl ester (20, 1.91 g, 4.70 mmol) and N-ethyldiisopropylamine (2.46 ml, 14.1 mmol) in dichloromethane

(20 ml) at 0 ⁰ C was added chloromethyl ethyl ether (0.65 ml, 7.05 mmol). After stirring at rt over night the reaction mixture was cooled to 0 ⁰ C and more N-ethyldiisopropylamine (0.82 ml, 4.7 mmol) and chloromethyl ethyl ether (0.22 ml, 2.4 mmol) was added, then stirred additional 16 h at rt. The reaction mixture was then directly applied on a silica gel column and eluted using stepwise gradient elution (ethyl acetate in hexane 50-100 %). Concentration of the appropriate fractions gave compound 21 as a slight yellow syrup (1.83 g, 84 %). LR-MS: Calcd for C ₂₅ H ₄ IN ₂ O ₆ : 465. Found: 465 [M+H].

Step 5

A degassed solution of l-{[4-Ethoxymethoxy-2-(hex-5-enyl-methyl-carbamoyl)- cyclopentanecarbonyl] -amino }-2-vinyl-cyclopropanecarboxylic acid ethyl ester (21, 1.83 g, 3.93 mmol) in dichloroethane (1.8 L, stored over 4A molecular sieves before use) was added Howeyda-Grubbs 1 ^st generation catalyst (0.165 g, 0.27 mmol), then shortly degassed and stirred at approximately 85 ⁰ C bath temperature overnight

(monitored by LC-MS). The reaction mixture was then allowed to cool somewhat after which solid phase catalyst scavenger (1.3 g, MP-TMT, Argonaut Technologies) and stirred additional 1.5 h, then filtered and concentrated. Flash chromatography of the residue (YMC-GEL silica) using stepwise gradient elution (ethyl acetate in hexane, 50-100 %) and concentration of the appropriate fractions gave the title compound as a brown syrup which crystallized upon standing (1.33 g, 77 %, purity approx. 90 %). This material was crystallized from 4:1 ethyl acetate (40 ml) giving a brown solid 22 (0.79 g, 1.8 mmol) and chromatography of the mother liquor gave additional product 22 (0.36 g, 0.81 mmol). LR-MS: Calcd for C ₂₃ H ₃₇ N ₂ O ₆ : 437. Found: 437 [M+H].

Step 6. Preparation of 17-hydroxy-13-methyl-2,14-dioxo-3,13-diaza-tricyclo- ri3.3.0.0 ⁴ ' ⁶ loctadec-7-ene-4-carboxyric acid ethyl ester (23)

To a stirred solution of 17-Ethoxymethoxy-13-methyl-2,14-dioxo-3,13-diazatricyclo- [13.3.0.0 ⁴ ' ⁶ ]octadec-7-ene-4-carboxylic acid ethyl ester (22, 0.083 g, 0.19 mmol) in 1 :1 :1 THF/methano I/water at rt was added concentrated hydrochloric acid (0.325 ml). The reaction mixture was monitored by TLC (9:1 ethyl acetate/methanol) and after 3 h; more hydrochloric acid (0.2 ml) was added. After 2 more hours the reaction mixture was neutralized using sodium hydrogen carbonate (s) (approx. 0.5 g). The reaction mixture was concentrated into Vi the volume, then partitioned between aqueous 10 % citric acid (10 ml) and dichloromethane (5 ml). The water layer was washed with dichloromethane (4 x 5 ml) and the combined organic layers were dried (Na ₂ SO ₄ ), filtered and concentrated. Flash chromatography of the residue using stepwise gradient elution (methanol in ethyl acetate 5 to 10 %) followed by concentration and drying of the appropriate fractions gave a colorless foam (0.027 g, 38 %).

Step 7: Preparation of 2-Pyridin-2-yl-5-trifluoromethyl-phenylamine (24).

A screw cap tube was charged with 2-tributyltinpyridine (1.4 eq), prepared from 2-bromopyridine and tributyltin hydride according to the procedure described in example 19-1, o-bromoaniline (200 mg, 1 eq), Pd(dba) ₂ (10-14 mg, 2 mol%), CuI (20-mg, 10 mol%), and PPh ₃ (40 mg, 15 mol%). The mixture was degassed and back-filled with argon. Dry diethyl ether (5 ml) was added, and the reaction mixture was heated at 120 ⁰ C for 4h in a microwave oven. The reaction mixture was cooled to room temperature, stirred with saturated aqueous KF (3 ml) for 3h, and filtered. The solid was discarded after washing with ethyl acetate (three times). The liquid was poured into H ₂ O and extracted with ethyl acetate. The combined organic layer was washed with H ₂ O and brine, dried over MgSO ₄ , and filtered and the solvent was removed in vacuo. The residue was purified by column chromatography on silica (ethyl acetate/petroleum ether as eluent) to afford the title compound as a white solid (60 mg, 38%). M ⁺ 239.

Step 8. 13-Methyl-2,14-dioxo-17-(2-pyridin-2-yl-5-trifluoromethylphe nyl carbamoyl- oxy)-3 J3-diaza-tricyclo[13.3.0.0 ⁴ ' ⁶ ]octadec-7-ene-4-carboxyric acid ethyl ester (25). 17-hydroxy- 13-methyl-2, 14-dioxo-3, 13-diaza-tricyclo[ 13.3.0.0 ⁴ ' ⁶ ]octadec-7-ene- 4-carboxylic acid ethyl ester (23, 20 mg, 0.053 mmol) was dissolved in dry DCE and of sodium bicarbonate (20 mg) was added, followed by 2 ml of phosgene solution in

toluene (20%). The reaction mixture was stirred at room temperature for 2-3 h (full conversion to chloroimidate according to LC-MS). The reaction mixture was then concentrated by rotary evaporation and dried from excess of phosgene in high vacuum (1.5 h). The dry reaction mixture was transferred into a "microwave" vial (2-5 mL), mixed with dry DCE (3-4 mL), 2-pyridin-2-yl-5-trifluoromethyl-phenylamine (24, 2 eq), potassium carbonate (9 mg, 1.5 eq), powdered molecular sieves (4A, 5-10 mg) and heated by microwave at 100 ⁰ C for 45 min. The reaction mixture was passed through a short pad of silica (eluent DCM, then 10% methanol in DCM). The resulting fractions containing the desired carbamate were combined, concentrated by rotary evaporation and purified by column chromatography on YMC silica (15 g, ethyl acetate/petroleum ether 1 :3 to remove excess of aniline, followed by dichloromethane and then 2% methanol in dichloromethane) to give the title compound 25 as a powder.

Step 9. Preparation of 13-methyl-2,14-dioxo-17-(2-pyridin-2-yl-5-trifluoromethyl- phenyl carbamoyloxy)-3, 13-diaza-tricyclo[ 13.3.0.0 ⁴ ' ⁶ loctadec-7-ene-4-carboxylic acid (26).

A solution of 13-methyl-2,14-dioxo-17-(2-pyridin-2-yl-5-trifluoromethylphe nyl carbamoyloxy)-3, 13-diaza-tricyclo[ 13.3.0.0 ⁴ ' ⁶ ]octadec-7-ene-4-carboxylic acid ethyl ester (25, 217 mg, 0.34 mmol) and LiOH. H ₂ O (1 eq) in THF (5 mL) methanol (5 mL) water (5 mL) IM LiOH (10 ml) was stirred at room temperature for 4 days. The reaction mixture was then concentrated into approximately 1/3 of the volume, diluted with water (30 mL) and acidified to approx. pH 4 using aqueous 10 % citric acid (60 mL), then washed with ethyl acetate (3 x 50 mL). The combined organic layers were washed with brine (1 x 100 mL), then dried (Na ₂ SO ₄ ), filtered and concentrated. Column chromatography of the residue using 9:1 ethyl acetate/methanol as eluent gave the title compound as a yellowish powder.

Step 10: Preparation of (2-pyridin-2-yl-5-trifluoromethylphenyl)carbamic acid 4- [(dimethoxyphosphoryl)aminocarbonyll-13-methyl-2,14-dioxo-3J 3-diaza-tricyclo- ri3.3.0.0 ⁴ ' ⁶ loctadec-7-en-17-yl ester (28).

To a solution of 13-methyl-2,14-dioxo-17-(2-pyridin-2-yl-5-trifluoromethylphe nyl carbamoyloxy)-3, 13-diaza-tricyclo[ 13.3.0.0 ⁴ ' ⁶ ]octadec-7-ene-4-carboxylic acid (26, 1.60 mmol) in dichloromethane (20 mL) at room temperature was added N-ethyl-N'- (3-dimethylaminopropyl)carbodiimide x HCl (0.74 g, 1.65 mmol), then stirred for 2.5 h after which TLC (9:1 ethyl acetate/methanol, stained using ammoniummolybdate- cerium sulfate in aq. 10% sulfuric acid) and LC-MS indicated complete conversion of the acid into the product. The reaction mixture was then diluted with dichloromethane (20 mL), washed with water (3 x 20 mL), then dried (Na ₂ SO ₄ ) filtered and concentrated

into a foamy syrup of intermediate 27 which was used without further purification in the next step.

Phosphoramidate (144 mg, 0.784 mmol) and DBU (490 μL, 2.86 mmol) are successively added to a stirred solution of intermediate 27 (0.574 mmol) in dry THF (10 mL). The resulting mixture is heated to reflux. After 2h, the solution is cooled down to room temperature, then concentrated under reduced pressure. The residue is purified by column chromatography to give the target product 28 as a white powder.

Example 6: Activity of compounds of formula (I) Replicon assay

The compounds of formula (I) were examined for activity in the inhibition of HCV RNA replication in a cellular assay. The assay demonstrated that the compounds of formula (I) exhibited activity against HCV replicons functional in a cell culture. The cellular assay was based on a bicistronic expression construct, as described by

Lohmann et al. (1999) Science vol. 285 pp. 110-113 with modifications described by Krieger et al. (2001) Journal of Virology 75: 4614-4624, in a multi-target screening strategy. In essence, the method was as follows. The assay utilized the stably transfected cell line Huh-7 luc/neo (hereafter referred to as Huh-Luc). This cell line harbors an RNA encoding a bicistronic expression construct comprising the wild type NS3-NS5B regions of HCV type Ib translated from an Internal Ribosome Entry Site (IRES) from encephalomyocarditis virus (EMCV), preceded by a reporter portion (FfL-luciferase), and a selectable marker portion (neo ^R , neomycine phosphotransferase). The construct is bordered by 5' and 3' NTRs (non-translated regions) from HCV type Ib. Continued culture of the replicon cells in the presence of G418 (neo ^R ) is dependent on the replication of the HCV RNA. The stably transfected replicon cells that express HCV RNA, which replicates autonomously and to high levels, encoding inter alia luciferase, are used for screening the antiviral compounds.

The replicon cells were plated in 384 well plates in the presence of the test and control compounds which were added in various concentrations. Following an incubation of three days, HCV replication was measured by assaying luciferase activity (using standard luciferase assay substrates and reagents and a Perkin Elmer ViewLux ^Tm ultraHTS microplate imager). Replicon cells in the control cultures have high luciferase expression in the absence of any inhibitor. The inhibitory activity of the compound on luciferase activity was monitored on the Huh-Luc cells, enabling a dose-response curve for each test compound. EC50 values were then calculated, which value represents the

amount of the compound required to decrease by 50% the level of detected luciferase activity, or more specifically, the ability of the genetically linked HCV replicon RNA to replicate.

Inhibition assay

The aim of this in vitro assay was to measure the inhibition of HCV NS3/4A protease complexes by the compounds of the present invention. This assay provides an indication of how effective compounds of the present invention would be in inhibiting

HCV NS3/4A proteolytic activity.

The inhibition of full-length hepatitis C NS3 protease enzyme was measured essentially as described in Poliakov, 2002 Prot Expression & Purification 25 363 371. Briefly, the hydrolysis of a depsipeptide substrate, Ac-DED(Edans)EEAbuψ[COO]ASK(Dabcyl)- NH ₂ (AnaSpec, San Jose, USA), was measured spectrofluorometrically in the presence of a peptide cofactor, KKGS VVIVGPJVLSGK (Ake Engstrόm, Department of

Medical Biochemistry and Microbiology, Uppsala University, Sweden). [Landro, 1997 #Biochem 36 9340-9348]. The enzyme (1 nM) was incubated in 50 mM HEPES, pH 7.5, 10 mM DTT, 40% glycerol, 0.1% n-octyl-D-glucoside, with 25 μM NS4A cofactor and inhibitor at 30 ⁰ C for 10 min, whereupon the reaction was initiated by addition of 0.5 μM substrate. Inhibitors were dissolved in DMSO, sonicated for 30 sec. and vortexed. The solutions were stored at - 20 ⁰ C between measurements.

The final concentration of DMSO in the assay sample was adjusted to 3.3%. The rate of hydrolysis was corrected for inner filter effects according to published procedures. [Liu, 1999 Analytical Biochemistry 267 331-335]. Ki values were estimated by non-linear regression analysis (GraFit, Erithacus Software, Staines, MX, UK), using a model for competitive inhibition and a fixed value for Km (0.15 μM). A minimum of two replicates was performed for all measurements.