EP1740582A1

EP1740582A1 - Acyl dihydro pyrrole derivatives as hcv inhibitors

Info

Publication number: EP1740582A1
Application number: EP05734941A
Authority: EP
Inventors: David GlaxoSmithKline HAIGH; Martin John GlaxoSmithKline SLATER
Original assignee: Glaxo Group Ltd
Current assignee: Glaxo Group Ltd
Priority date: 2004-04-22
Filing date: 2005-04-20
Publication date: 2007-01-10
Also published as: JP2007533693A; WO2005103045A1; GB0408995D0

Abstract

Novel anti-viral agents of Formula (I) in which: A represents hydroxy; D represents aryl or heteroaryl; E represents hydrogen, C1-6alkyl, aryl, heteroaryl or heterocyclyl; G represents hydrogen or C1-6alkyl optionally substituted by one or more substituents selected from halo, OR1, SR1, C(O)NR2R3, CO2H, C(O)R4, CO2R4, NR2R3, NHC(O)R4, NHCO2R4, NHC(O)NR5R6, SO2NR5R6, SO2R4, nitro, cyano, aryl, heteroaryl and heterocyclyl; R1 represents hydrogen, C1-6alkyl, arylalkyl, or heteroarylalkyl; R2 and R3 are independently selected from hydrogen, C1-6alkyl, aryl and heteroaryl; or R2 and R3 together with the nitrogen atom to which they are attached form a 5 or 6 membered saturated cyclic group; R4 is selected from the group consisting of C1-6alkyl, aryl, heteroaryl, arylalkyl, and heteroarylalkyl; R5 and R6 are independently selected from the group consisting of hydrogen, C1-6alkyl, aryl, heteroaryl, arylalkyl, and heteroarylalkyl; or R5 and R6 together with the nitrogen atom to which they are attached form a 5 or 6 membered saturated cyclic group; and J represents C1-6alkyl, heterocyclylalkyl, arylalkyl or heteroarylalkyl; and salts, solvates and esters thereof; provided that when A is esterified to form -OR where R is selected from straight or branched chain alkyl, aralkyl, aryloxyalkyl, or aryl, then R is other than tert-butyl; processes for their preparation, pharmaceutical compositions comprising them, and methods of using them in HCV treatment are provided.

Description

COMPOUNDS ACYL DIHYDRO PYRROLE DERIVATIVES AS HCV INHIBITORS FIELD OF THE INVENTION

The present invention relates to novel acyl dihydro pyrrole derivatives useful as anti-viral agents. Specifically, the present invention involves novel Hepatitis C virus (HCV) inhibitors.

BACKGROUND OF THE INVENTION

Ikeda et al, (1997) Journal of the Chemical Society, Perkin Transactions 1 : Organic and Bio-Organic Chemistry 22: 3339-3344 and Sato et al, (1995) Journal of the Chemical Society, Perkin Transactions 1 , 14:1801-1809 and Sato et al, (1994) Heterocycles 37(1 ): 245-248 disclose 4',5'-unsubstituted acyl pyrrolidine compounds useful as reagents in the regioselective synthesis of bridged azabicyclic compounds; no medical use was disclosed for the acyl pyrrolidine compounds.

Ikeda et al, (1996) Heterocycles 42(1): 155-158 and Confalone et al, (1988) Journal of Organic Chemistry 53(3): 482-487 and De artino et al, (1976) Farmaco, Ed. Sci. 31(11): 785-790 disclose 4',5'-unsubstituted acyl pyrrolidine compounds useful as reagents in the synthesis of tricyclic nitrogen-containing heterocycles; no medical use was disclosed for the acyl pyrrolidine compounds. Alig et al, (1992) Journal of Medicinal Chemistry 35(23): 4393-4407 discloses a 4',5'-unsubstituted acyl pyrrolidine compound useful as a reagent in the synthesis of non-peptide fibrinogen receptor antagonists; no medical use was disclosed for the acyl pyrrolidine compound.

Padwa et al, (1992) Journal of the American Chemical society 114(2): 593-601 discloses a 4',5'-unsubstituted acyl pyrrolidine compound useful as a reagent in the synthesis of azomethine ylides; no medical use was disclosed for the acyl pyrrolidine compound. Culbertson et al, (1990) Journal of Medicinal Chemistry 33(8): 2270-2275 and Crooks et al, (1979) Journal of the Chemical Society, Perkins Transactions 1 , 11 : 2719-2726 disclose 4',5'-unsubstituted acyl pyrrolidine compounds useful as reagents in the synthesis of 7-spiroamine quinolone and spiro[indan-2,2'-pyrrolidine] compounds respectively; no medical use was disclosed for the acyl pyrrolidine compounds.

WO2002/44168, WO96/33170 and EP505868A2 disclose 4',5'-unsubstituted acyl pyrrolidine compounds useful as intermediates in the synthesis of indolecarboxamide, N- aroylamino acid amide and N-acyl-α-amino acid derivatives respectively; no medical use was disclosed for the acyl pyrrolidine compounds.

De Caprariis et al, (1989) Journal of Heterocyclic Chemistry 26(4): 1023-1027 discloses 3 pyrrolidinedicarboxylic acid derivatives useful as intermediates in the synthesis of pyrrolo[1 ,4]benzodiazepine compounds; no medical use was disclosed for the pyrrolidinedicarboxylic acid derivatives. WO99/37304 discloses oxoazaheterocyclyl derivatives, especially piperazinone compounds, having Factor Xa inhibitory activity. These derivatives may include certain acyl pyrrolidine derivatives. There is no mention of HCV polymerase inhibitory activity for the disclosed compounds.

Infection with HCV is a major cause of human liver disease throughout the world. In the US, an estimated 4.5 million Americans are chronically infected with HCV. Although only 30% of acute infections are symptomatic, greater than 85% of infected individuals develop chronic, persistent infection. Treatment costs for HCV infection have been estimated at $5.46 billion for the US in 1997. Worldwide over 200 million people are estimated to be infected chronically. HCV infection is responsible for 40-60% of all chronic liver disease and 30% of all liver transplants. Chronic HCV infection accounts for 30% of all cirrhosis, end-stage liver disease, and liver cancer in the U.S. The CDC estimates that the number of deaths due to HCV will minimally increase to 38,000/year by the year 2010.

Due to the high degree of variability in the viral surface antigens, existence of multiple viral genotypes, and demonstrated specificity of immunity, the development of a successful vaccine in the near future is unlikely. Alpha-interferon (alone or in combination with ribavirin) has been widely used since its approval for treatment of chronic HCV infection. However, adverse side effects are commonly associated with this treatment: flu-like symptoms, leukopenia, thrombocytopenia, depression from interferon, as well as anemia induced by ribavirin (Lindsay, K.L. (1997) Hepatology 26 (suppl 1 ): 71S-77S). This therapy remains less effective against infections caused by HCV genotype 1 (which constitutes -75% of all HCV infections in the developed markets) compared to infections caused by the other 5 major HCV genotypes. Unfortunately, only -50-80% of the patients respond to this treatment (measured by a reduction in serum HCV RNA levels and normalization of liver enzymes) and, of those treated, 50-70% relapse within 6 months of cessation of treatment. Recently, with the introduction of pegylated interferon, both initial and sustained response rates have improved substantially, and combination treatment of Peg-IFN with ribavirin constitutes the gold standard for therapy. However, the side effects associated with combination therapy and the impaired response in patients with genotype 1 present opportunities for improvement in the management of this disease.

First identified by molecular cloning in 1989 (Choo, Q-L et al (1989) Science 244:359- 362), HCV is now widely accepted as the most common causative agent of post- transfusion non-A, non-B hepatitis (NANBH) (Kuo, G et al (1989) Science 244:362-364). Due to its genome structure and sequence homology, this virus was assigned as a new genus in the Flaviviridae family. Like the other members of the Flaviviridae, such as flaviviruses (e.g. yellow fever virus and Dengue virus types 1-4) and pestiviruses (e.g. bovine viral diarrhea virus, border disease virus, and classic swine fever virus) (Choo, Q-L et al (1989) Science 244:359-362; Miller, R.H. and R.H. Purcell (1990) Proc. Natl. Acad. Sci. USA 87:2057-2061), HCV is an enveloped virus containing a single strand RNA molecule of positive polarity. The HCV genome is approximately 9.6 kilobases (kb) with a long, highly conserved, noncapped 5' nontranslated region (NTR) of approximately 340 bases which functions as an internal ribosome entry site (IRES) (Wang CY et al 'An RNA pseudoknot is an essential structural element of the internal ribosome entry site located within the hepatitis C virus 5' noncoding region' RNA- A Publication of the RNA Society. 1 (5): 526-537, 1995 Jul.). This element is followed by a region which encodes a single long open reading frame (ORF) encoding a polypeptide of -3000 amino acids comprising both the structural and nonstructural viral proteins.

Upon entry into the cytoplasm of the cell, this RNA is directly translated into a polypeptide of -3000 amino acids comprising both the structural and nonstructural viral proteins. This large polypeptide is subsequently processed into the individual structural and nonstructural proteins by a combination of host and virally-encoded proteinases (Rice, CM. (1996) in B.N. Fields, D.M.Knipe and P.M. Howley (eds) Virology 2^nd Edition, p931- 960; Raven Press, N.Y.). Following the termination codon at the end of the long ORF, there is a 3' NTR which roughly consists of three regions: an - 40 base region which is poorly conserved among various genotypes, a variable length poly(U)/polypyrimidine tract, and a highly conserved 98 base element also called the "3' X-tail" (Kolykhalov, A. et al (1996) J. Virology 70:3363-3371 ; Tanaka, T. et al (1995) Biochem Biophys. Res. Commun. 215:744-749; Tanaka, T. et al (1996) J. Virology 70:3307-3312; Yamada, N. et al (1996) Virology 223:255-261). The 3' NTR is predicted to form a stable secondary structure which is essential for HCV growth in chimps and is believed to function in the initiation and regulation of viral RNA replication.

The NS5B protein (591 amino acids, 65 kDa) of HCV (Behrens, S.E. et al (1996) EMBO J. 15:12-22), encodes an RNA-dependent RNA polymerase (RdRp) activity and contains canonical motifs present in other RNA viral polymerases. The NS5B protein is fairly well conserved both intra-typically (-95-98% amino acid (aa) identity across 1b isolates) and inter-typically (-85% aa identity between genotype 1 a and 1 b isolates). The essentiality of the HCV NS5B RdRp activity for the generation of infectious progeny virions has been formally proven in chimpanzees (A. A. Kolykhalov et al.. (2000) Journal of Virology, 74(4), p.2046-2051). Thus, inhibition of NS5B RdRp activity (inhibition of RNA replication) is predicted to cure HCV infection.

Based on the foregoing, there exists a significant need to identify synthetic or biological compounds for their ability to inhibit HCV.

SUMMARY OF THE INVENTION The present invention involves acyl dihydro pyrrole compounds represented hereinbelow, pharmaceutical compositions comprising such compounds and use of the compounds in treating viral infection, especially HCV infection. DETAILED DESCRIPTION OF THE INVENTION

The present invention provides at least one chemical entity chosen from compounds of Formula (I) :

in which:

A represents hydroxy;

D represents aryl or heteroaryl;

E represents hydrogen, Cι-₆alkyl, aryl, heteroaryl or heterocyclyl;

G represents hydrogen or Cι-₆alkyl optionally substituted by one or more substituents selected from halo, OR¹, SR¹, C(O)NR²R³, CO₂H, C(O)R⁴, CO₂R⁴, NR²R³, NHC(O)R⁴, NHCO₂R⁴, NHC(O)NR⁵R⁶, SO₂NR⁵R⁶, SO₂R⁴, nitro, cyano, aryl, heteroaryl and heterocyclyl;

R represents hydrogen, Cι-₆alkyl, arylalkyl, or heteroarylalkyl;

R² and R³ are independently selected from hydrogen, C-ι-₆alkyl, aryl and heteroaryl; or R² and R³ together with the nitrogen atom to which they are attached form a 5 or 6 membered saturated cyclic group;

R⁴ is selected from the group consisting of Cι-₆alkyl, aryl, heteroaryl, arylalkyl, and heteroarylalkyl;

R⁵ and R⁶ are independently selected from the group consisting of hydrogen, Cι-₆alkyl, aryl, heteroaryl, arylalkyl, and heteroarylalkyl; or R⁵ and R⁶ together with the nitrogen atom to which they are attached form a 5 or 6 membered saturated cyclic group; and

J represents Cι-₆alkyl, heterocyclylalkyl, arylalkyl or heteroarylalkyl;

and salts, solvates and esters thereof; provided that when A is esterified to form -OR where R is selected from straight or branched chain alkyl, aralkyl, aryloxyalkyl, or aryl, then R is other than fe/t-butyl. There is provided as a further aspect of the present invention a compound of Formula (I) or a physiologically acceptable salt, solvate or ester thereof for use in human or veterinary medical therapy, particularly in the treatment or prophylaxis of viral infection, particularly HCV infection.

It will be appreciated that reference herein to therapy and/or treatment includes, but is not limited to prevention, retardation, prophylaxis, therapy and cure of the disease. It will further be appreciated that references herein to treatment or prophylaxis of HCV infection includes treatment or prophylaxis of HCV-associated disease such as liver fibrosis, cirrhosis and hepatocellular carcinoma.

According to another aspect of the invention, there is provided the use of a compound of Formula (I) or a physiologically acceptable salt, solvate or ester thereof in the manufacture of a medicament for the treatment and/or prophylaxis of viral infection, particularly HCV infection.

In a further or alternative aspect there is provided a method for the treatment of a human or animal subject with viral infection, particularly HCV infection, which method comprises administering to said human or animal subject an effective amount of a compound of Formula (I) or a physiologically acceptable salt, solvate or ester thereof.

It will be appreciated that the compounds of the present invention may contain one or more asymmetric carbon atoms and may exist in racemic, diastereoisomeric, and optically active forms. All of these racemic compounds, enantiomers and diastereoisomers are contemplated to be within the scope of the present invention.

In one aspect of the invention, the relative stereochemistry of compounds of Formula (I) is represented by Formula (Ip):

tive stereochemistry in which A, D, E, G and J are as defined above for Formula (I).

The following aspects of the invention are applicable in respect of each of Formulae I and Ip:

In one aspect of the invention, D represents optionally substituted phenyl; in another aspect terf-butylphenyl optionally further substituted; in a further aspect D represents para-tert-butylphenyl optionally further substituted, in one aspect mefø-substituted, by halo, Cι-₃alkyl or C^alkoxy, for example bromo, chloro, methyl or methoxy; in another aspect D represents mefa-methoxy-para-terf-butylphenyl (3-methoxy-4-te/f-butylphenyl).

In one aspect, E represents optionally substituted heteroaryl; in a further aspect E represents thiazolyl or isoxazolyl, each of which may be optionally substituted. In one aspect, E represents optionally substituted thiazolyl. In another aspect, E represents optionally substituted isoxazolyl. In another aspect, E represents 1 ,3-thiazol-2-yl or 5- methyl-isoxazol-3-yl .

In one aspect, G represents Cι-₆alkyl optionally substituted by OR¹, SR¹ or SO₂R⁴; in another aspect, G represents methyl optionally substituted by OR¹, SR¹ or SO₂R⁴.

In one aspect, R¹ represents C^alkyl; in another aspect, R¹ represents methyl, ethyl, or propyl; in a further aspect methyl or ethyl.

In one aspect, R⁴ represents C^alkyl; in another aspect, R⁴ represents methyl, ethyl, or propyl, especially methyl.

In one aspect, J represents C_halky!, arylalkyl or heteroarylalkyl. In another aspect, J represents C_halky! or heteroarylmethyl. In a further aspect, J represents isobutyl or 1 ,3- thiazol-4-ylmethyl. In one aspect, J represents isobutyl. In another aspect, J represents 1 ,3-thiazol-4-ylmethyl.

It is to be understood that the present invention covers all combinations of aspects of the invention described herein.

As used herein unless otherwise specified, "alkyl" refers to an optionally substituted hydrocarbon group. The alkyl hydrocarbon group may be linear, branched or cyclic, saturated or unsaturated. Where the alkyl group is linear or branched, examples of such groups include methyl, ethyl, n-propyl, 1 -methylethyl (isopropyl), n-butyl, isobutyl, sec- butyl, tert-butyl, n-pentyl, isopentyl, neopentyl or hexyl and the like. In one aspect, alkyl moieties are Cι-₄alkyl. Where the alkyl hydrocarbon group is unsaturated, it will be understood that there will be a minimum of 2 carbon atoms in the group, for example an alkenyl or alkynyl group. Where the alkyl hydrocarbon group is cyclic, it will be understood that there will be a minimum of 3 carbon atoms in the group. The cyclic alkyl group may be saturated or unsaturated, monocyclic or bridged bicyclic. Where the cyclic alkyl group is saturated, examples of such groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl and the like. Where the cyclic alkyl group is unsaturated, examples of such groups include cyclobutenyl, cyclopentenyl, cyclohexenyl and the like. In one aspect, the cyclic alkyl group has from 3 to 6 carbon atoms. Unless otherwise stated, optional substituents include C^alkyl, halo, OR¹, SR¹, C(O)NR²R³, C(O)R⁴, CO₂H, CO₂R⁴, NR²R³, NHC(O)R⁴, NHCO₂R⁴, NHC(O)NR⁵R⁶, SO₂NR⁵R⁶, SO₂R⁴, nitro, cyano, oxo, and heterocyclyl.

As used herein, the term "alkenyl" refers to a linear or branched hydrocarbon group containing one or more carbon-carbon double bonds. In one aspect the alkenyl group has from 2 to 6 carbon atoms. Examples of such groups include ethenyl, propenyl, butenyl, pentenyl or hexenyl and the like.

As used herein, the term "alkynyl" refers to a linear or branched hydrocarbon group containing one or more carbon-carbon triple bonds. In one aspect the alkynyl group has from 2 to 6 carbon atoms. Examples of such groups include ethynyl, propynyl, butynyl, pentynyl or hexynyl and the like.

As used herein, "alkoxy" (when used as a group or as part of a group) refers to an alkyl ether radical, in which the term "alkyl" is defined above. Examples of alkoxy as used herein include, but are not limited to; methoxy, ethoxy, n-propoxy, i-propoxy and the like.

As used herein, "aryl" refers to an optionally substituted aromatic group with at least one ring having a conjugated pi-electron system, containing up to two conjugated or fused ring systems. "Aryl" includes carbocyclic aryl and biaryl groups, all of which may be optionally substituted, for example phenyl, naphthyl or bi-phenyl. In one aspect, "aryl" moieties are unsubstituted, monosubstituted, disubstituted or trisubstituted phenyl. In another aspect, "aryl" substituents are selected from the group consisting of C_halky!, halo, OR¹, C(O)NR²R³, C(O)R⁴, CO₂H, CO₂R⁴, NR²R³, NHC(O)R⁴, NHCO₂R⁴, NHC(O)NR⁵R⁶, SO₂NR⁵R⁶, SO₂R⁴, nitro, cyano, oxo, heterocyclyl, CF₃, and NO₂.

As used herein, "arylalkyl" refers to an aryl group attached to the parent molecular moiety through an alkyl group.

As used herein, "heteroaryl" refers to an optionally substituted, 5 or 6 membered, aromatic group comprising one to four heteroatoms selected from N, O and S, with at least one ring having a conjugated pi-electron system, containing up to two conjugated or fused ring systems. In one aspect, "heteroaryl" moieties are unsubstituted, monosubstituted or disubstituted thiazolyl. In another aspect, "heteroaryl" moieties are unsubstituted, monosubstituted or disubstituted isoxazolyl. In another aspect, "heteroaryl" substituents are selected from the group consisting of C_halky!, halo, OR¹, C(O)NR²R³, C(O)R⁴, CO₂H, CO₂R⁴, NR²R³, NHC(O)R⁴, NHCO₂R⁴, NHC(O)NR⁵R⁶, SO₂NR⁵R⁶, SO₂R⁴, nitro, cyano, oxo, heterocyclyl, CF₃, and NO₂.

As used herein, "heteroarylalkyl" refers to a heteroaryl group attached to the parent molecular moiety through an alkyl group. As used herein, "heterocyclic" and "heterocyclyl" refer to an optionally substituted, 5 or 6 membered, saturated cyclic hydrocarbon group containing 1 or 2 heteroatoms selected from N, optionally substituted by hydrogen, Cι-₆alkyl, C(O)R⁴, SO₂R⁴, aryl or heteroaryl; O; and S, optionally substituted by one or two oxygen atoms.

As used herein, "heterocyclylalkyl" refers to a heterocyclyl group attached to the parent molecular moiety through an alkyl group

As used herein, the term "pharmaceutically acceptable" means a compound which is suitable for pharmaceutical use. Salts and solvates of compounds of the invention which are suitable for use in medicine are those in which the counterion or associated solvent is pharmaceutically acceptable. However, salts and solvates having non-pharmaceutically acceptable counterions or associated solvents are within the scope of the present invention, for example, for use as intermediates in the preparation of other compounds of the invention and their pharmaceutically acceptable salts and solvates.

In one aspect, chemical entities useful in the present invention may be at least one chemical entity selected from the group consisting of: re/-(2S,5R)-2-Isobutyl-1-(3-methoxy-4-te/ -butylbenzoyl)-4-ethoxymethyl-5-(1 ,3-thiazol-2- yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid; re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-te/ιf-butylbenzoyl)-4-methoxymethyl-5-(1 ,3-thiazol-

2-yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid; re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-terf-butylbenzoyl)-4-n-propyloxymethyl-5-(1 ,3- thiazol-2-yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid; re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-tert-butylbenzoyl)-4-methylthiomethyl-5-(1 ,3-thiazol-

2-yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid; re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-te/t-butylbenzoyl)-4-methylsulfonylmethyl-5-(1 ,3- thiazol-2-yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid; rel-(2S,5R)-1-(3-Methoxy-4-tert-butylbenzoyl)-4-(methoxymethyl)-5-(5-methylisoxazol-3- yl)-2-(1 ,3-thiazol-4-ylmethyl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid; and salts, solvates and esters, and individual enantiomers thereof.

In one aspect of the present invention, there are provided pharmaceutically acceptable salt complexes. The present invention also covers the pharmaceutically acceptable salts of the compounds of Formula (I). In one aspect, physiologically acceptable salts of the compounds of Formula (I) include acid salts, for example sodium, potassium, calcium, magnesium and tetraalkylammonium and the like, or mono- or di- basic salts with the appropriate acid for example organic carboxylic acids such as acetic, lactic, tartaric, malic, isethionic, lactobionic and succinic acids; organic sulfonic acids such as methanesulfonic, ethanesulfonic, benzenesulfonic and p-toluenesulfonic acids and inorganic acids such as hydrochloric, sulfuric, phosphoric and sulfamic acids and the like. In another aspect, the present invention also relates to pharmaceutically acceptable esters of the compounds of Formula (I), for example carboxylic acid esters -COOR, in which R is selected from straight or branched chain alkyl, for example n-propyl, n-butyl, alkoxyalkyl (e.g. methoxymethyl), aralkyl (e.g. benzyl), aryloxyalkyl (e.g. phenoxymethyl), aryl (e.g. phenyl optionally substituted by halogen, C_halky! or C^alkoxy or amino). Unless otherwise specified, in one aspect any alkyl moiety present in such esters contains 1 to 18 carbon atoms, particularly 1 to 4 carbon atoms. Any aryl moiety present in such esters comprises in one aspect a phenyl group.

In one aspect, the present invention relates to compounds of Formula (I) and salts and solvates thereof.

In another aspect, the present invention relates to compounds of Formula (I) and pharmaceutically acceptable salts and solvates thereof.

In one aspect, the present invention relates to solvates of the compounds of Formula (I), for example hydrates.

It will further be appreciated that certain compounds of the present invention may exist in different tautomeric forms. All tautomers are contemplated to be within the scope of the present invention.

Compounds of Formula (I) in which A is hydroxy may be prepared from a compound of Formula (II)

in which A is a protected hydroxy group, for example an alkoxy or silyloxy, for example tri-

(C.|-₄alkyl)-silyloxy group, and D, E, G and J are as defined above for Formula (I), by deprotection. Suitable protecting groups can be found, but are not restricted to, those found in T W Greene and P G M Wuts 'Protective Groups in Organic Synthesis', 3^rd Ed (1999), J Wiley and Sons.

For example when A is fe/if-butoxy, and D, E, G and J are as defined above for Formula (I), by treatment with an appropriate acid, for example trifluoroacetic acid. In one aspect, the reaction is carried out in a solvent, for example dichloromethane. In one aspect, the temperature is in the range 0 to 50°C, in another aspect 20 to 30°C. For example when A is allyloxy, and D, E, G and J are as defined above for Formula (I), by treatment with a suitable catalyst for example tetrakis(triphenylphosphine)palladium(0) and a suitable proton source, for example phenylsilane. The reaction is carried out in a suitable solvent, for example dichloromethane.

For example when A is tri(methyl)silyloxy, and D, E, G and J are as defined above for Formula (I), by treatment with a suitable fluoride source for example tetrabutylammonium fluoride. The reaction is carried out in a suitable solvent, for example THF.

Compounds of Formula (I) in which A is hydroxy or a protected form thereof may also be prepared by reaction of a compound of Formula (III)

in which A, E, G, and J are as defined above for Formula (I); with a suitable acylating agent, for example D-C(O)-hal, in which hal is a halo atom, for example chloro or bromo, and D is as defined above for Formula (I). In one aspect the reaction is carried out in a suitable solvent, for example dichloromethane, in the presence of a suitable base, for example triethylamine and thereafter removing any protecting group. Suitable protecting groups can be found, but are not restricted to, those found in T W Greene and P G M Wuts 'Protective Groups in Organic Synthesis', 3^rd Ed (1999), J Wiley and Sons.

Compounds of Formula (I) or (II) in which G represents optionally substituted alkyl, may be prepared by appropriate manipulation of a compound of Formula (I la)

in which A is hydroxy or an alkoxy or tri-(C₁-₄alkyl)-silyloxy group, D, E and J are as defined above for Formula (I), and L represents CO₂Y or COY in which Y represents hydrogen or alkyl.

For example, a compound of Formula (II) in which G represents hydroxyalkyl may be prepared by reduction of a compound of Formula (I la) in which L represents CO₂Y or COY and Y represents hydrogen or alkyl, using a suitable reducing agent, for example lithium borohydride, sodium borohydride, sodium triacetoxyborohydride, borane/dimethyl sulfide complex or lithium aluminium hydride, or suitable combinations thereof, in a suitable solvent or mixture thereof for example THF or methanol. For example, a compound of Formula (II) in which G represents hydroxyalkyl may also be prepared by reaction of a compound of formula (I la) in which L represents CO₂Y or COY and Y represents hydrogen or alkyl, with a suitable organometallic reagent, for example methylmagnesium bromide or methyl lithium, in a suitable solvent, for example THF.

For example, a compound of formula (II) in which G represents difluoroalkyl may be prepared by reaction of a compound of formula (lla) in which L represents COY and Y represents alkyl, with a suitable fluorinating agent, for example diethylaminosulfur trifluoride, in a suitable solvent, for example dichloromethane.

For example, a compound of formula (II) in which G represents alkenyl may be prepared by treatment of a compound of formula (lla) in which L represents COY and Y represents hydrogen or alkyl, with a phosphonium salt, for example phosphonium halide salts such as methyltriphenylphosphonium bromide, methyltriphenylphosphonium chloride, methoxymethyltriphenylphosphonium bromide or methoxymethyltriphenylphosphonium chloride, in the presence of a base, for example lithium bis(trimethylsilyl)amide, and in a suitable solvent, for example THF.

In a further aspect, a compound of Formula (II) may be prepared by appropriate manipulation of another compound of Formula (II). For example, a compound of Formula (II) in which G represents hydroxyalkyl may be converted into a compound of Formula (II) in which G represents optionally substituted alkyl, for example alkyl, haloalkyl or alkoxyalkyl.

For example, a compound of Formula (II) in which G represents alkoxyalkyl may be prepared by alkylation of a compound of formula (II) in which G represents hydroxyalkyl using a suitable base for example sodium hydride and a suitable alkylating agent such as an alkyl iodide, for example methylating using methyl iodide or ethylating using ethyl iodide. In one aspect the reaction is carried out in a suitable solvent, for example DMF.

For example, a compound of Formula (II) in which G represents haloalkyl may be prepared by halogenation of a compound of Formula (II) in which G represents hydroxyalkyl using a suitable halogenating agent, for example hydroxymethyl may be converted into fluoromethyl using a suitable fluorinating agent, for example diethylaminosulfur trifluoride, in a suitable solvent, for example dichloromethane.

For example, a compound of Formula (II) in which G represents cyanomethyl may be prepared by reacting a compound of Formula (II) in which G represents hydroxymethyl with, for example trifluoromethanesulfonic anhydride, and subsequently treating the product with a nucleophile, for example a cyanide salt such as tetrabutylammonium cyanide. A compound of Formula (II) in which G represents C^alkylthiomethyl can similarly be prepared using C^alkylthiolate as the nucleophile. For example, a compound of Formula (II) in which G represents may be prepared by oxidising a compound of Formula (II) in which G represents Ci- ₄alkylthiomethyl, using for example 3-chloroperbenzoic acid, in a suitable solvent, for example dichloromethane.

For example, a compound of Formula (II) in which G represents alkyl may be prepared by deoxygenation of a compound of Formula (II) in which G represents hydroxyalkyl. In one aspect, the deoxygenation is carried out in a two step process in which: Step(i) A compound of formula (II) in which G represents hydroxyalkyl is converted into a thionocarbonate by treatment with a suitable chloroformate for example 4-fluorophenyl thionochloroformate. In one aspect the reaction is carried out in a suitable solvent, for example dichloromethane, in the presence of a suitable base catalyst, for example 4-(N,N- dimethylamino)pyridine. Step(ii) The thionocarbonate from step(i) is treated with a suitable radical initiator, for example AIBN, and a suitable proton source, such as tris(trimethylsilyl)silane, in a suitable solvent, for example dioxan. In one aspect, the temperature is in the range 80 to 120°C.

For example, a compound of Formula (II) in which G represents an unsaturated alkyl group, for example 1-methylethenyl or 2-methoxyethenyl, may be converted into a compound of Formula (II) in which G represents a saturated alkyl group, for example isopropyl or 2-methoxyethyl, by hydrogenation in the presence of a suitable catalyst, for example palladium-on-carbon, and in a suitable solvent, for example ethanol. For example, a compound of Formula (II) in which G represents alkenyloxyalkyl or substituted alkenyloxyalkyl may be converted into a compound of Formula (II) in which G represents alkyloxyalkyl or substituted alkyloxyalkyl by hydrogenation in the presence of a suitable catalyst, for example palladium-on-carbon, and in a suitable solvent, for example ethanol. For example, allyloxyalkyl or substituted allyloxyalkyl may be converted into propyloxyalkyl or substituted. propyloxyalkyl.

A compound of Formula (lla) in which L represents CO₂Y in which Y represents hydrogen may be prepared from a compound of Formula (lla) in which L represents CO₂Y in which Y represents alkyl. For example, a compound of Formula (lla) in which L represents CO₂Me may be converted into a compound of Formula (lla) in which L represents CO₂H by hydrolysis, for example base catalysed hydrolysis using a suitable base such as sodium hydroxide in a suitable solvent such as methanol.

A compound of Formula (lla) in which L represents CO₂Y or COY in which Y represents hydrogen or alkyl may be prepared from a compound of Formula (Ilia) in which L represents CO₂Y or COY in which Y represents hydrogen or alkyl, and A, E, and J are as defined above for Formula (I); with a suitable acylating agent, for example D- C(O)-hal, in which hal is a halo atom, for example chloro or bromo, and D is as defined above for Formula (1). In one aspect the reaction is carried out in a suitable solvent, for example dichloromethane, in the presence of a suitable base, for example triethylamine.

In a further aspect, a compound of Formula (lla) in which L represents COY and Y represents hydrogen may be prepared in a two-stage process from a compound of Formula (lla) in which L represents CO₂Y and Y represents hydrogen or alkyl. In a first step, the compound of Formula (lla) in which L represents CO₂Y and Y represents hydrogen or alkyl is treated with a suitable reducing agent, for example, lithium aluminium hydride or sodium borohydride. In a second step, the resultant hydroxy group is oxidised with a suitable oxidising agent which may be selected from conventional oxidising reagents known in the art, for example an appropriate mixture of oxalyl chloride, dimethyl sulfoxide and triethylamine.

A compound of Formula (lla) in which A is hydroxy, may be converted to a compound of Formula (lla) in which A is an alkoxy or silyloxy group by standard hydroxy protecting techniques. Similarly, a compound of Formula (lla) in which A is an alkoxy or silyloxy group, may be converted to a compound of Formula (lla) in which A is hydroxy by standard deprotecting techniques. Suitable protecting groups can be found, but are not restricted to, those found in T W Greene and P G M Wuts 'Protective Groups in Organic Synthesis', 3^rd Ed (1999), J Wiley and Sons.

A compound of Formula la) may be prepared by reaction of a compound of Formula (IV)

in which E and J are as defined above for Formula (I) and A is as defined above for

Formula (II) with a compound of Formula (V) L

(V) in which L represents CO₂Y or COY in which Y represents hydrogen or alkyl. In one aspect, the reaction is carried out in a suitable solvent, for example THF or acetonitrile, optionally in the presence of a Lewis acid catalyst, such as lithium bromide or silver acetate, and a base, such as triethylamine, 1 ,8-diazabicyclo[5,4,0]undec-7-ene (DBU) or tetramethyl guanidine. Alternatively, the reaction is carried out in a suitable solvent, for example THF or acetonitrile, in the presence of an acid, such as acetic acid, or the reaction may be carried out by heating compounds of Formula (IV) and Formula (V) in a suitable solvent, for example toluene, xylene or acetonitrile in the absence of a catalyst.

Compounds of Formula (III) in which G represents optionally substituted alkyl may be prepared by appropriate manipulation of a compound of Formula (Ilia) after first optionally protecting the N-atom of the dihydropyrrole ring with a suitable N-protecting group, for example benzyloxycarbonyl (CBZ) or fe/t-butoxycarbonyl. For example, a compound of Formula (III) in which G represents hydroxyalkyl may be prepared by reduction of a compound of Formula (Ilia) in which L represents CO₂Y and Y represents alkyl, using a suitable reducing agent, for example lithium borohydride or sodium borohydride, in a suitable solvent for example THF. Where protected prior to manipulation, deprotection of the N-atom by standard procedures results in the compound of Formula (III). For example, when the N-protecting group is te/t-butoxycarbonyl, deprotection may be achieved by treatment with a suitable acid, for example trifluoroacetic acid.

In a similar manner to that described above in relation to compounds of Formula (II), a compound of Formula (III), in which G represents hydroxyalkyl and the N-atom is protected, may be converted into a compound of Formula (III) in which G represents optionally substituted alkyl, for example alkyl, haloalkyl or alkoxyalkyl and the N-atom is protected. Deprotection of the N-atom by standard procedures results in the new compound of Formula (III).

Compounds of Formula (IV) may be prepared by reaction of a compound of Formula (VI) in which J is as defined above for Formula (I) and A is as defined above for Formula (II) with a compound of Formula E-CHO in the presence of an optional drying agent, for example magnesium sulfate, in a suitable solvent, for example dichloromethane. Where an acid addition salt (such as a hydrochloride salt) of a compound of Formula (VI) is employed in place of a compound of Formula (VI), the reaction may be performed in the presence of a suitable base, for example triethylamine.

Compounds of Formula (VI) in which J is other than heteroarylalkyl, acid addition salts of compounds of Formula (VI) and E-CHO are known in the art or may be prepared by standard literature procedures. Compounds of Formula (VI) in which J is heteroarylalkyl, such as 1 ,3-thiazol-4-ylmethyl, and A is an alkoxy, benzyloxy or tri-(C₁-₄alkyl)-silyloxy group, may be prepared by treatment of a compound of Formula (VII) in which J is heteroarylalkyl, such as 1 ,3-thiazol-4-ylmethyl, and A is an alkoxy, benzyloxy or tri-(Cι-₄alkyl)-silyloxy group with an acid, for example 15% citric acid. Preferably, the reaction is carried out in a suitable solvent, for example THF.

Compounds of Formula (VII) may be prepared by reaction of a compound of Formula

in which A is an alkoxy, benzyloxy or tri-(C₁-₄alkyl)-silyloxy group with a compound of Formula J-hal in which J is heteroarylalkyl, such as 1,3-thiazol-4-ylmethyl, and hal is a halo atom, preferably chloro or bromo. Preferably, the reaction is carried out in the presence of a suitable base such as potassium ferf-butoxide. Preferably, the reaction is carried out in a suitable solvent, for example THF. Preferably the reaction is carried out at a temperature in the range -10 to 10°C, more preferably 0°C.

Compounds of Formula (VIII) and J-hal are known in the art or may be prepared by standard literature procedures.

Compounds of Formula (I) in which A is an ester may be prepared by esterification of a compound of Formula (I) in which A is hydroxy by standard literature procedures for esterification.

It will be appreciated that compounds of Formula (I), (II), (lla), (III) and/or (Ilia) which exist as diastereoisomers may optionally be separated by techniques well known in the art, for example by column chromatography.

It will be appreciated that racemic compounds of Formula (I), (II), (lla), (III) and/or (Ilia) may be optionally resolved into their individual enantiomers. Such resolutions may conveniently be accomplished by standard methods known in the art. For example, a racemic compound of Formula (I), (II), (lla), (III) and/or (Ilia) may be resolved by chiral preparative HPLC. Alternatively, racemic compounds of Formula (I), (II), (lla), (III) and/or (Ilia) which contain an appropriate acidic or basic group, such as a carboxylic acid group or amine group may be resolved by standard diastereoisomeric salt formation with a chiral base or acid reagent respectively as appropriate. Such techniques are well established in the art. It will also be appreciated that individual enantiomeric compounds of Formula (III) and/or (Ilia) may be prepared by general methods of asymmetric synthesis using, where appropriate, chiral auxiliaries or chiral catalytic reagents and additionally performing any suitable functional group interconversion step as hereinbefore described, including the addition or removal of any such chiral auxiliary. Such general methods of asymmetric synthesis are well known in the art and include, but are not restricted to, those described in "Asymmetric Synthesis," Academic Press, 1984 and/or "Chiral Auxiliaries and Ligands in Asymmetric Synthesis", Wiley, 1995. For example, suitable general chiral auxiliaries include chiral alcohols such as menthol or 1-phenylethanol; chiral oxazolidinones such as 4-benzyloxazolidin-2-one or 4-isopropyloxazolidin-2-one; chiral sultams such as camphor sultam; or chiral amines such as 1-phenylethylamine or 2-amino-2-phenylethanol. Suitable general chiral catalytic reagents include chiral basic amines and chiral ligands such as N-methylephedrine, 1-phenyl-2-(1-pyrrolidinyl)-1-propanol, 3-(dimethylamino)- 1 ,7,7-trimethylbicyclo[2.2.1]-heptan-2-ol, 3,4-bis(diphenylphosphanyl)-1-(phenylmethyl)- pyrrolidine, chinchonine, chinchonidine, sparteine, hydroquinine or quinine, BINAP or chiral bis(oxazoline) (BOX) ligands and derivatives, optionally in the presence of a metal salt, for example M_mX_x where M is silver, cobalt, zinc, titanium, magnesium, or manganese, and X is halide (for example chloride or bromide), acetate, trifluoroacetate, p- toluenesulfonate, trifluoromethylsulfonate, hexafluorophosphate or nitrate, and _m and _x are 1 , 2, 3 or 4, and optionally in the presence of a base, for example triethylamine. All of these chiral auxiliaries or chiral catalytic reagents are well described in the art. General illustrative examples of the preparation of various chiral pyrrolidines by asymmetric synthesis using chiral auxiliaries or chiral catalytic reagents include, but are not limited to, those described in Angew. Chem. Int. Ed., (2002), 41: 4236; Chem. Rev., (1998), 98: 863; J. Am. Chem. Soc, (2002), 124: 13400; J. Am. Chem. Soc, (2003), 125: 10175; Org. Lett., (2003), 5, 5043; Tetrahedron, (1995), 51 : 273; Tetrahedron: Asymm., (1995), 6: 2475; Tetrahedron: Asymm., (2001), 12: 1977; Tetrahedron: Asymm., (2002), 13: 2099 and Tet Lett., (1991), 41 : 5817.

With appropriate manipulation and protection of any chemical functionality, synthesis of compounds of Formula (I) is accomplished by methods analogous to those above and to those described in the Experimental section. Suitable protecting groups can be found, but are not restricted to, those found in T W Greene and P G M Wuts 'Protective Groups in Organic Synthesis', 3^rd Ed (1999), J Wiley and Sons.

ABBREVIATIONS

In describing the invention, chemical elements are identified in accordance with the Periodic Table of the Elements. Abbreviations and symbols utilized herein are in accordance with the common usage of such abbreviations and symbols by those skilled in the chemical arts. The following abbreviations are used herein: AIBN 2,2'-azo-bis-isobutyronitrile

BINAP 2,2'-bis(diphenylphosphino)-1 ,1'-binaphthyl

BOX bisoxazoline

CBZ benzyloxycarbonyl CDCI₃ deuterated chloroform

DBU 1 ,8-diazabicyclo[5,4,0]undec-7-ene

DCM dichloromethane

DMF N,N-dimethylformamide

DTT dithiothreitol EDTA ethylenediaminetetraacetic acid

GTP guanosine triphosphate

HCV Hepatitis C virus

HPLC High Performance Liquid Chromatography

ICAM intercellular adhesion molecule MgCI₂ magnesium chloride

MnCI₂ manganese II chloride

MS mass spectrometry

NaCI sodium chloride

NANBH non-A, non-B hepatitis Na₂SO₄ sodium sulfate

NMR Nuclear Magnetic Resonance spectroscopy nOe Nuclear Overhauser effect

NSAIDs non-steroidal anti-inflammatory drugs

PBS phosphate buffered saline SPA beads scintillation proximity beads

THF tetrahydrofuran

TLC thin layer chromatography

Tris-HCL 2-amino-2-(hydroxymethyl)-1 ,3-propanediol, hydrochloride

EXAMPLES

Intermediate 1

2-[N-(1,3-Thiazol-2-ylmethylene)amino]-4-methylpentanoic acid, terf-butyl ester

A stirred mixture of 2-amino-4-methyl-pentanoic acid fe/f-butyl ester, hydrochloride (5.00 g, 22.3 mmol), 1 ,3-thiazole-2-carboxaldehyde (2.53 g, 22.4 mmol) and triethylamine (3.10 L, 22.2 mmol) in dichloromethane (60 mL) was heated under reflux under nitrogen for 19 hours. The reaction mixture was allowed to cool to room temperature, washed twice with water, dried over Na₂SO₄ and evaporated to give the title compound as an oil. ¹H NMR (CDCI₃): δ 8.46 (s, 1 H), 7.94 (d, 1 H), 7.44 (dd, 1 H), 4.07 (dd, 1 H), 1.89-1.74 (m, 2H), 1.64-1.52 (m, 1 H), 1.48 (s, 9H), 0.96 (d, 3H) and 0.90 (d, 3H).

Intermediate 2 re/-(2S,5R)-2-lsobutyl-5-(1,3-thiazol-2-yl)-2,5-dihydro-1H-pyrrole-2,4-dicarboxylic acid, 2-terf-butyl

Racemic; Relative stereochemistry shown To a cooled (0°C) stirred solution of Intermediate 1 (0.250 g, 0.88 mmol) in anhydrous THF (5 mL) under nitrogen, was added triethylamine (0.123 mL, 0.88 mmol) followed by lithium bromide (0.077 g, 0.88 mmol) and methyl propiolate (0.08 mL, 0.90 mmol). The mixture was stirred at 0°C for 5 minutes, and then the ice/water bath was removed and stirring was continued at ambient temperature for 2 hours. Aqueous ammonium chloride was added with rapid stirring and the resulting mixture was extracted twice with ethyl acetate. The combined organic layers were dried over sodium sulfate and evaporated. The residue was purified by chromatography on silica gel using cyclohexane-ethyl acetate (8:2, v/v) as eluant to provide the title compound as an oil. MS calcd for (Cι₈H₂₆N₂O₄S + H)⁺: 367. MS (electrospray): Found (M+H)⁺ = 367.

Intermediate 3 re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-tert-butylbenzoyl)-5-(1,3-thiazol-2-yl)-2,5- dihydro-1H-pyrrole-2,4-dicarboxylic acid, 2-te/f-butyl ester, 4-methyl ester

Racemic; Relative stereochemistry shown

To a solution of Intermediate 2 (1.0 g, 2.73 mmol) in anhydrous dichloromethane (20 mL) under nitrogen, was added triethylamine (0.5 mL, 3.58 mmol) followed by 3-methoxy-4- te/if-butylbenzoyl chloride^* (0.79 g, 3.48 mmol) and the mixture was stirred overnight at room temperature. The reaction was quenched by the addition of water and extracted with dichloromethane. The organic fraction was evaporated and purified by silica gel chromatography, eluting with cyclohexane-ethyl acetate (9:1, v/v), to give the title compound as a solid. * Prepared from 3-methoxy-4-terf-butylbenzoic acid (J. Org. Chem. (1961), 26, 1732). MS calcd for (C₃₀H₄oN₂O₆S + H)⁺: 557. MS (electrospray): Found (M+H)⁺ = 557.

Intermediate 4 re/-(2S,5R)-2-Isobutyl-1-(3-methoxy-4-fert-butylbenzoyl)-4-hydroxymethyl-5-(1,3- thiazol-2-yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid, 2-ferf-butyl ester

Racemic; Relative stereochemistry shown

To a solution of Intermediate 3 (0.697 g, 1.25 mmol) in dry THF (50 mL) under nitrogen was added lithium aluminium hydride (1.25 mL of a 1 M solution in ether). After 20 mins, the reaction was quenched by the addition of sodium carbonate solution (1 M) and extracted with ethyl acetate. The organic fraction was evaporated and purified by silica gel column chromatography, eluting with a cyclohexane-ethyl acetate gradient (7:3 v/v then 6:4 v/v) to afford the title compound as a solid. MS calcd for (C₂₉H₄oN₂O₅S + H)⁺: 529. MS (electrospray): Found (M+H)⁺ = 529.

Intermediate 5 re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-tert-butylbenzoyl)-4-methoxymethyl-5-(1,3- thiazol-2-yl)-2,5-dihydro-1H-pyrrole-2-carboxylic acid, 2-ferf-butyl ester

Racemic; Relative stereochemistry shown

To a solution of Intermediate 4 (0.113 g, 0.21 mmol) in dry DMF (4 mL) at -10°C under nitrogen was added sodium hydride (9 mg, 60% dispersion in oil, 0.23 mmol). After 20 mins at this temperature, methyl iodide (0.027 mL, 0.43 mmol) was added, the mixture allowed to attain room temperature and stirred overnight. Methanol was added and the solvent removed in vacuo. The residue was extracted between water and ethyl acetate, the organic layer separated, dried over sodium sulfate and evaporated. Purification by silica gel chromatography using cyclohexane-ethyl acetate (9:1 v/v) as eluent afforded the title compound. ¹H NMR (CD₃OD): δ 7.51 (d, 1H), 7.32 (d, 1 H), 7.22 (d, 1 H), 6.84 (d, 1H), 6.50 (d, 1 H), 6.17 (d, 1 H), 5.89 (d, 1H), 3.62-3.75 (m, 5H), 3.23 (s, 3H), 2.56-2.68 (m, 1 H), 2.05-2.15 (m, 1 H), 1.70-1.85 (m, 1H), 1.60 (s, 9H), 1.34 (s, 9H) and 1.08 (2d, 6H).

Intermediate 6 re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-te/ -butylbenzoyl)-4-ethoxymethyl-5-(1,3- thiazol-2-y!)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid, 2-terf-butyl ester

Racemic; Relative stereochemistry shown

This was prepared in a similar manner to that described for Intermediate 5, replacing methyl iodide with ethyl iodide. The crude material was purified by chromatography on preparative TLC plates, using cyclohexane-ethyl acetate (6:4, v/v) as eluent to afford the title compound.

¹H NMR (CD₃OD): δ 7.48 (d, 1H), 7.28 (d, 1H), 7.19 (d, 1 H), 6.80 (d, 1H), 6.46 (s, 1 H),

6.14 (s, 1 H), 5.85 (d, 1 H), 3.6-3.75 (m, 2H), 3.67 (s, 3H), 3.25-3.41 (q, 2H), 2.55-2.65 (m, 1H), 2.02-2.12 (m, 1H), 1.70-1.85 (m, 1H), 1.57 (s, 9H), 1.31 (s, 9H) and 0.95-1.1 (m, 9H).

Intermediate 7 re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-terf-butylbenzoyl)-4-n-propyloxymethyl-5-(1,3- thiazol-2-yI)-2,5-dihydro-1H-pyrrole-2-carboxylic acid, 2-terf-butyl ester

Racemic; Relative stereochemistry shown

This was prepared in a similar manner to that described for Intermediate 5, replacing methyl iodide with n-propyl iodide. The crude material was purified by chromatography on preparative TLC plates, using cyclohexane-ethyl acetate (6:4, v/v) as eluent to afford the title compound. MS calcd for (C₃₂H₄₆N₂O₅S + H)⁺: 571. MS (electrospray): Found (M+H)⁺ = 571.

Intermediate 8 re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-fert-butylbenzoyI)-4-[(methanesuIfonyloxy)- methyl]-5-(1 ,3-thiazol-2-yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid, 2-terf-butyl ester

Racemic; Relative stereochemistry shown

Methanesulfonyl chloride (0.044 mL, 0.57 mmol) was diluted in dry dichloromethane (3 mL). To this solution was added triethylamine (0.08 mL, 0.57 mmol) followed by Intermediate 4 (0.3 g, 0.57 mmol) as a solution in dry dichloromethane (3 mL) and the mixture was stirred overnight at room temperature. Another equivalent each of methanesulfonyl chloride and triethylamine were added and after 1 hour the reaction was quenched with water (5 mL). The phases were separated using a hydrophobic frit and the solvent was evaporated to give the title compound. MS calcd for (C₃₀H₄₂N₂O₇S₂ + H)⁺: 607. MS (electrospray): Found (M+H)⁺ 607.

Intermediate 9 re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-tert-butylbenzoyl)-4-methylthiomethyl-5-(1,3- thiazol-2-yl)-2,5-dihydro-1H-pyrrole-2-carboxylic acid, 2-terf-butyl ester

Racemic; Relative stereochemistry shown

Sodium methanesulfide (0.045 g, 0.64 mmol) was dissolved in dry DMF (4 mL). To the solution was added Intermediate 8 (0.126 g, 0.21 mmol) as a solution in dry dichloromethane (0.5 mL) and the mixture was stirred over night at room temperature under nitrogen atmosphere. The reaction was quenched with brine (5 mL). The phases were separated and the aqueous layer extracted with dichloromethane (2 x 10 mL). The organic layers were combined and dried over Na₂SO , filtered and the solvent removed in vacuo. The resulting oil was purified by reverse phase HPLC on a C₁₈ column using a two-solvent gradient elution comprising (A) water containing 0.1% v/v formic acid and (B) acetonitrile-water (95:5 v/v) containing 0.05% v/v formic acid, with analysis of the fractions by electrospray mass spectroscopy. Further purification by analytical HPLC using a Thermo-Fluophase column (100 x 20 mm, particle size 5μ), eluting with aqueous acetonitrile (1 :1 v/v) and collecting the fraction with retention time 29 minutes gave the title compound as a solid.

MS calcd for (C₃₀H₄₂N₂O₄S₂ + H)⁺: 559.

MS (electrospray): Found (M+H)⁺ = 559.

Intermediate 10 re/^S.SRJ^-lsobutyl-I^S-methoxy^-tert-butylbenzoy ^-Kmethylsulfony -methyl]- 5-(1,3-thiazol-2-yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid, 2-tert-butyl ester

Racemic; Relative stereochemistry shown

Sodium methanesulfinic acid (0.065 g, 0.64 mmol) was dissolved in dry DMF (4 mL). To this solution was added Intermediate 8 (0.126 g, 0.21 mmol) as a solution in dry dichloromethane (0.5 mL), and the mixture was stirred overnight at room temperature under nitrogen atmosphere. The reaction was quenched with brine (5 mL), the phases were separated and the aqueous layer was extracted with dichloromethane (10 mL x 2). The organic layers were combined and dried over Na₂SO₄, filtered and reduced in vacuo. The resulting oil was purified by reverse phase HPLC on a Ciβ column using a two-solvent gradient elution comprising (A) water containing 0.1% v/v formic acid and (B) acetonitrile- water (95:5 v/v) containing 0.05% v/v formic acid, with analysis of the fractions by electrospray mass spectroscopy. Further purification by column chromatography over silica gel, eluting with cyclohexane/ethyl acetate (8:2, v/v), gave the title compound. MS calcd for (C₃o^₄₂^₂θ₆S₂ + H)⁺: 591. MS (electrospray): Found (M+H)⁺ 591.

Intermediate 11

Enantiomer A of re/-(2S,5R)-2-lsobutyl-1 -(S-methoxy^-terf-butylbenzoyl - ethoxymethyl-5-(1 ,3-thiazol-2-yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid, 2-tert- butyl ester

Chiral; Relative stereochemistry shown

The enantiomers of Intermediate 6 were separated by chiral HPLC using a Whelk O-1 column (2 inches x 20 cm, flow rate 70 mL/min), eluting with heptane-ethanol, (95:5 v/v). The second eluting enantiomer (retention time 10 mins) was collected to afford Enantiomer A.

Intermediate 12

2-[N-(Diphenylmethylene)amino]-3-(1,3-thiazol-4-yl)propanoic acid, terf-butyl ester

PartA

To a cooled (ice-bath) solution of 2-[N-(diphenylmethylene)amino]ethanoic acid, ferf-butyl ester (99.26 g, 336 mmol) in dry THF (600 mL) under an atmosphere of nitrogen, was added a 1 M solution of potassium terf-butoxide in THF (341 mL) dropwise (dropping funnel) over 1 hour. The dropping funnel was rinsed with further dry THF (150 mL) which was added to the reaction and the mixture was allowed to stir for a further 1 hour in the ice-bath. Part B

Independently during this time, 4-(chloromethyl)-1,3-thiazole hydrochloride (59.84 g, 352 mmol) was freshly converted to the free base as follows: The hydrochloride was mixed with dichloromethane (750 mL) and water (300 mL) and the pH of the aqueous layer was adjusted to pH 9 with solid sodium bicarbonate. The organic layer was separated, dried over sodium sulfate and carefully evaporated at 80 torr to give the free base. Part C The 4-(chloromethyl)-1 ,3-thiazole (formed in Part B) was dissolved in THF (400 mL) and added dropwise (dropping funnel) over 45 min to the reaction mixture from Part A, keeping the reaction at ice-bath temperature. Solid anhydrous lithium iodide (0.47 g, 3.5 mmol) was added directly to the reaction mixture 5 min after addition of the alkylating agent had started. The dropping funnel was rinsed with further dry THF (100 mL) which was added to the reaction. The reaction was allowed to warm to room temperature over 1 hour and was stirred at room temperature for an additional 2 hours before being partitioned between a mixture of saturated brine (1 L), water (500 mL) and ethyl acetate (1.5 L). The organic layer was separated and the aqueous layer re-extracted with further ethyl acetate (2 x 500 mL). The combined organic layers were dried over sodium sulfate and evaporated to give the title compound (141.5 g, crude) which was used without further purification in the following step.

¹H NMR (CDCI₃): δ 8.65 (d, 1 H), 7.55-7.62 (m, 2H), 7.2-7.55 (m, 6H), 7.05 (d, 1 H), 6.78- 6.87 (m, 2H), 4.36-4.41 (m, 1 H), 3.47-3.54 (m, 1 H), 3.36-3.44 (m, 1 H) and 1.44 (s, 9H). Intermediate 13

2-Amino-3-(1,3-thiazol-4-yl)propanoic acid, tert-butyl ester

A solution of Intermediate 12 (141.4 g, crude product) in THF (750 mL) under nitrogen was cooled in an ice-bath. A solution of citric acid in water (15% w/v, 750 mL) was added over 25 min with stirring. The mixture was allowed to warm to room temperature over 1 hour and stirred for a further 5 hours, left overnight and stirred for a further 2 hours. The THF was removed under reduced pressure at 25°C, and 1 M aqueous hydrochloric acid (375 mL) added. The mixture was extracted with diethyl ether (2 x 1 L) and the combined ether extracts back extracted with water (200 mL). The combined aqueous layers were extracted with further diethyl ether (500 mL). These ether layers were discarded. The aqueous layer was then carefully adjusted to pH 9.5 in the presence of fresh diethyl ether (1 L). This ether layer was separated and kept. Saturated brine (400 mL) was added to the aqueous layer and this layer was extracted with further diethyl ether (3 x 600 mL). The four ether extracts were combined and dried over sodium sulfate. Evaporation of the solvent gave the title compound, an oil.

¹H NMR (CDCI3): δ 8.77 (d, 1 H), 7.08 (d, 1 H), 3.77-3.85 (m, 1 H), 3.22-3.32 (m, 1 H), 3.02- 3.13 (m, 1 H) and 1.42 (s, 9H).

Intermediate 14 re/-(2S,5R)-5-(5-Methylisoxazol-3-yl)-2-(1,3-thiazol-4-ylmethyl)-2,5-dihydro-1H- pyrrole-2,4-dicarb n A mixture of Intermediate 13 (0.96 g, 4.20 mmol) and 5-methylisoxazol-3-carboxaldehyde (0.93 g, 8.37 mmol) in dichloromethane (15 mL) was heated at 40°C for 3h, allowed to cool to room temperature and left to stand overnight. The reaction mixture was evaporated under reduced pressure, to give a mixture of 2-[[N-(5-methylisoxazol-3- yl)methylene]amino]-3-(1 ,3-thiazol-4-yl)propanoic acid, tert-butyl ester and intermediate 13 (3:1 ratio). This mixture was dissolved in dry THF (15 mL), lithium bromide (0.355 g, 4.09 mmol) was added and the mixture stirred under nitrogen at 0°C. Triethylamine (0.57 mL, 4.09 mmol) was added followed by methyl propiolate (0.3 mL, 3.37 mmol) and the resulting mixture was stirred at 0°C for 20 min and allowed to warm to room temperature. After 2 hours, saturated ammonium chloride solution (40 mL) was added and the mixture was extracted with ethyl acetate. The organic layer was dried over magnesium sulfate and evaporated. The residue was purified by chromatography on silica gel using ethyl acetate-cyclohexane (1 :2 v/v) as eluent to give the title compound as an oil which solidified on standing. MS calcd for (C_1gH₂₃N₃O₅S + H)⁺: 406. MS found (electrospray): (M+H)⁺ = 406

Intermediate 15 re/-(2S,5R)-1-(3-Methoxy-4-tert-butylbenzoyl)-5-(5-methylisoxazol-3-yl)-2-(1,3-thiazol- 4-ylmethyl)-2,5-dihydro-1H-pyrrole-2,4-dicarboxylic acid, 2-ferf-butyl ester, 4-methyl ester

Racemic; Relative stereochemistry shown

A mixture of Intermediate 14 (0.974 g, 2.40 mmol), 3-methoxy-4-terf-butylbenzoyl chloride (0.652 g, 2.88 mmol) and triethylamine (0.4 mL, 2.87 mmol) was stirred at room temperature under nitrogen overnight. The mixture was washed with water and brine, dried over magnesium sulfate and evaporated. The residue was purified by chromatography on silica gel using cyclohexane-ethyl acetate (3:1 v/v) as eluent to give the title compound as a foam. MS calcd for (C₃₁H₃₇N₃O₇S + H)⁺: 596. MS found (electrospray): (M+H)⁺ = 596

Intermediate 16 re/-(2S,5R)-4-(Hydroxymethyl)-1-(3-methoxy-4-ferf-butylbenzoyl)-5-(5- methylisoxazol-3-yI)-2-(1,3-thiazol-4-ylmethyl)-2,5-dihydro-1 H-pyrroIe-2-carboxylic acid, terf-butyl ester

Racemic; Relative stereochemistry shown

A solution of Intermediate 15 (0.716 g, 1.20 mmol) in dry THF (13 mL) was stirred under nitrogen and cooled to -43°C. A 1.0 M solution of lithium aluminium hydride in diethyl ether (1.32 mL) was added dropwise. When addition was complete the mixture was warmed to -38°C and held at that temperature for 2.25 hours. The mixture was quenched with 1 M aqueous potassium carbonate solution (12 mL), allowed to warm to room temperature and extracted with ethyl acetate. Extracts were dried over magnesium sulfate and evaporated. The residue was purified by chromatography on silica gel using cyclohexane-ethyl acetate (initially 2:1 v/v then 1 :1 v/v) and methanol as eluents to give the title compound as a foam. MS calcd for (C₃₀H₃₇N₃O₆S + H)⁺: 568. MS found (electrospray): (M+H)⁺ = 568

Intermediate 17 re/-(2S,5R)-1-(3-Methoxy-4-tert-butylbenzoyl)-4-(methoxymethyl)-5-(5- methylisoxazol-3-yl)-2-(1,3-thiazol-4-ylmethyl)-2,5-dihydro-1H-pyrrole-2-carboxylic acid, terf-butyl ester

Racemic; Relative stereochemistry shown

A solution of Intermediate 16 (0.345 g, 0.61 mmol) in DMF (20 mL) was stirred under nitrogen and cooled to -10°C. A 60% dispersion of sodium hydride in mineral oil (40 mg, 1.00 mmol) was added and the resulting mixture was stirred below 0°C for 30 min. lodomethane (0.189 mL, 3.04 mmol) was added at -10°C and the mixture was allowed to warm to room temperature overnight. The mixture was quenched with methanol (15 mL) and evaporated under reduced pressure. The residue was partitioned between water and ethyl acetate. The organic layer was collected, washed with water and brine, dried over magnesium sulfate and evaporated. The residue was purified by chromatography on silica gel using cyclohexane-ethyl acetate (3:1 v/v) as eluent to give the title compound as a foam. MS calcd for (C₃.H₃₉N₃O₆S + H)⁺: 582. MS found (electrospray): (M+H)⁺ = 582

Example 1 re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-tert-butylbenzoyl)-4-ethoxymethyl-5-(1,3- thiazol-2-yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid Racemic; RReellaattiivvee sstereochemistry shown

A solution of Intermediate 6 (0.021 g, 0.038 mmol) in trifluoroacetic acid (2 mL) was stirred at 20°C for 3 hours. The mixture was evaporated and the residue was triturated with ether to give the title compound as a solid.

¹H NMR (CD₃OD): δ 7.65 (m, 1 H), 7.50 (m, 1 H), 7.25 (d, 1 H), 6.88 (d, 1 H), 6.45 (s, 1 H),

6.3 (s, 1 H), 5.9 (s, 1H), 3.82 (dd, 2H), 3.69 (s, 3H), 3.30-3.45 (m, 2H), 2.50-2.65 (m, 1 H),

2.15-2.25 (m, 1H), 1.75-1.90 (m, 1 H), 1.34 (s, 9H) and 0.95-1.15 (m, 9H). The carboxylic acid proton exchanges with solvent.

The relative stereochemistry was confirmed as (2S, 5R) by NMR nOe experiments.

Example 2 re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-tert-butylbenzoyl)-4-methoxymethyl-5-(1,3- thiazol-2-yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid

Racemic; Relative stereochemistry shown

The title compound was prepared in a similar manner to that described for Example 1 , replacing Intermediate 6 with Intermediate 5.

¹H NMR (CD₃OD): δ 7.64 (d, 1 H), 7.51 (d, 1 H), 7.25 (d, 1 H), 6.89 (d, 1H), 6.45 (s, 1 H), 6.28 (s, 1H), 5.91 (s, 1 H), 3.77 (d, 2H), 3.68 (s, 3H), 3.23 (s, 3H), 2.50-2.65 (m, 2H), 2.15- 2.30 (m, 1H), 1.75-1.90 (m, 1H), 1.34 (s, 9H) and 1.08 (2xd, 6H). Carboxylic acid proton assumed exchanged with solvent.

The relative stereochemistry was confirmed by NMR nOe experiments to be (2S, 5R).

Example 3 re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-tert-butylbenzoyl)-4-π-propyloxymethyl-5-(1,3- thiazol-2-yl)-2,5-dihydro-1H-pyrrole-2-carboxylic acid Racemic; Relative stereochemistry shown

The title compound was prepared in a similar manner to that described for Example 1 , replacing Intermediate 6 with Intermediate 7.

MS calcd for (C₂₈H₃₈N₂O₅S + H)⁺: 515.

MS (electrospray): Found (M+H)⁺ 515.

Example 4 re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-tert-butylbenzoyl)-4-methylthiomethyl-5-(1,3- thiazol-2-yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid

Racemic; Relative stereochemistry shown

The title compound was prepared in a similar manner to that described for Example 1 , replacing Intermediate 6 with Intermediate 9. MS calcd for (C₂₆H₃ N₂O₄S₂ + H)⁺: 503. MS (electrospray): Found (M+H)⁺ = 503.

Example 5 re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-fe/ -butylbenzoyl)-4-methylsulfonylmethyl-5- (1 ,3-thiazol-2-yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid

Racemic; Relative stereochemistry shown

The title compound was prepared in a similar manner to that described for Example 1 , replacing Intermediate 6 with Intermediate 10.

¹H NMR (CD₃OD): δ 7.65 (d, 1H), 7.51 (d, ΪH), 7.22 (d, 1 H), 6.83 (d, 1H), 6.49 (m, 2H), 6.21 (s, 1 H), 4.11 (d, 1H), 3.68 (s, 3H), 3.45 (d, 1 H), 2.99 (s, 3H), 2.50-2.63 (m, 1 H), 2.22- 2.34 (m, 1 H), 1.75-1.90 (m, 1 H), 1.32 (s, 9H) and 1.08 (2xd, 6H). Acid proton assumed to be exchanged with solvent.

Example 6

Enantiomer A of re/-(2S,5R)-2-lsobutyl-1 -(3-methoxy-4-ferf-butylbenzoyl)-4- ethoxymethyl-5-(1 ,3-thiazol-2-yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid

Chiral; Relative stereochemistry shown

A solution of Intermediate 11 (0.026 g, 0.047 mmol) in trifluoroacetic acid (2 mL) was stirred at 20°C for 3 hours. The mixture was evaporated and the residue was triturated with ether to give the title compound as a solid. ¹H NMR (CD₃OD): δ 7.61 (d, 1H), 7.47 (d, 1 H), 7.22 (d, 1 H), 6.86 (d, 1H), 6.43 (s, 1H), 6.26 (s, 1H), 5.87 (s, 1H), 3.80 (dd, 2H), 3.66 (s, 3H), 3.25-3.50 (m, 2H), 2.48-2.60 (m, 1 H), 2.15-2.25 (m, 1 H), 1.70-1.85 (m, 1 H), 1.31 (s, 9H) and 1.00-1.10 (m, 9H). Acid proton assumed to be exchanged with solvent.

Example 7 re/-(2S,5R)-1-(3-Methoxy-4-terf-butylbenzoyl)-4-(methoxymethyl)-5-(5- methylisoxazol-3-yl)-2-(1,3-thiazol-4-ylmethyl)-2,5-dihydro-1H-pyrrole-2-carboxylic acid

Racemic; Relative stereochemistry shown

A solution of Intermediate 17 (0.256 g, 0.44 mmol) in dichloromethane (3 mL) was treated with trifluoroacetic acid (3 mL) at room temperature overnight. The mixture was evaporated and the residue co-evaporated with toluene and triturated with methanol to give the title compound as a white solid. MS calcd for (C₂₇H₃ιN₃O_eS + H)⁺: 526. MS found (electrospray): (M+H)⁺ = 526

¹H NMR (CDCI₃): δ 8.88 (d, 1 H), 7.22 (d, 1 H), 7.14 (d, 1H), 6.66 (dd, 1H), 6.63 (d, 1H), 5.92 (s, 1 H), 5.60 (s, 1 H), 5.35 (s, 1 H), 4.28 (d, 1H), 3.72 (s, 3H), 3.87-3.60 (m, 3H), 3.10 (s, 3H), 2.24 (s, 3H) and 1.31 (s, 9H). Acid proton not seen.

The compounds according to the invention may be formulated for administration in any convenient way, and the invention therefore also includes within its scope pharmaceutical compositions for use in therapy, comprising a compound of formula (I) or a physiologically acceptable salt or solvate thereof in admixture with one or more physiologically acceptable diluents or carriers.

The compounds of the present invention can be administered by different routes including intravenous, intraperitoneal, subcutaneous, intramuscular, oral, topical, transdermal, or transmucosal administration. For systemic administration, oral administration is preferred. For oral administration, for example, the compounds can be formulated into conventional oral dosage forms such as capsules, tablets and liquid preparations such as syrups, elixirs and concentrated drops.

Alternatively, injection (parenteral administration) may be used, e.g., intramuscular, intravenous, intraperitoneal, and subcutaneous. For injection, the compounds of the invention are formulated in liquid solutions; for example, in physiologically compatible buffers or solutions, such as saline solution, Hank's solution, or Ringer's solution. In addition, the compounds may be formulated in solid form and redissolved or suspended immediately prior to use. Lyophilized forms can also be produced.

Systemic administration can also be by transmucosal or transdermal means. For transmucosal or transdermal administration, penetrants appropriate to the barrier to be permeated are used in the formulation. Such penetrants are generally known in the art, and include, for example, for transmucosal administration, bile salts and fusidic acid derivatives. In addition, detergents may be used to facilitate permeation. Transmucosal administration, for example, may be through nasal sprays, rectal suppositories, or vaginal suppositories.

For topical administration, the compounds of the invention can be formulated into ointments, salves, gels, or creams, as is generally known in the art.

The amounts of various compounds to be administered can be determined by standard procedures taking into account factors such as the compound (IC₅₀) potency, (EC₅₀) efficacy, and the biological half-life (of the compound), the age, size and weight of the patient, and the disease or disorder associated with the patient. The importance of these and other factors to be considered are known to those of ordinary skill in the art. Amounts administered also depend on the routes of administration and the degree of oral bioavailability. For example, for compounds with low oral bioavailability, relatively higher doses will have to be administered. Oral administration is a preferred method of administration of the present compounds.

In one aspect, the composition is in unit dosage form. For oral application, for example, a tablet, or capsule may be administered, for nasal application, a metered aerosol dose may be administered, for transdermal application, a topical formulation or patch may be administered and for transmucosal delivery, a buccal patch may be administered. In each case, dosing is such that the patient may administer a single dose.

Each dosage unit for oral administration contains, in one aspect, from 0.01 to 500 mg/Kg, in another aspect, from 0.1 to 50 mg/Kg, of a compound of Formula (I) or a pharmaceutically acceptable salt thereof, calculated as the free base. The daily dosage for parenteral, nasal, oral inhalation, transmucosal or transdermal routes contains in one aspect from 0.01 mg to 100 mg/Kg, of a compound of Formula (I). In one aspect, a topical formulation contains 0.01 to 5.0% of a compound of Formula (I). The active ingredient may be administered from 1 to 6 times per day, preferably once, sufficient to exhibit the desired activity, as is readily apparent to one skilled in the art.

Composition of Formula (I) and their pharmaceutically acceptable salts which are active when given orally can be formulated as syrups, tablets, capsules and lozenges. A syrup formulation will generally consist of a suspension or solution of the compound or salt in a liquid carrier for example, ethanol, peanut oil. olive oil, glycerine or water with a flavoring or coloring agent. Where the composition is in the form of a tablet, any pharmaceutical carrier routinely used for preparing solid formulations may be used. Examples of such carriers include magnesium stearate, terra alba, talc, gelatin, acacia, stearic acid, starch, lactose and sucrose. Where the composition is in the form of a capsule, any routine encapsulation is suitable, for example using the aforementioned carriers in a hard gelatin capsule shell. Where the composition is in the form of a soft gelatin shell capsule any pharmaceutical carrier routinely used for preparing dispersions or suspensions may be considered, for example aqueous gums, celluloses, silicates or oils, and are incorporated in a soft gelatin capsule shell.

Typical parenteral compositions consist of a solution or suspension of a compound or salt in a sterile aqueous or non-aqueous carrier optionally containing a parenterally acceptable oil, for example polyethylene glycol, polyvinylpyrrolidone, lecithin, arachis oil or sesame oil.

Typical compositions for inhalation are in the form of a solution, suspension or emulsion that may be administered as a dry powder or in the form of an aerosol using a conventional propellant such as dichlorodifluoromethane or trichlorofluoromethane. A typical suppository formulation comprises a compound of Formula (I) or a pharmaceutically acceptable salt thereof which is active when administered in this way, with a binding and/or lubricating agent, for example polymeric glycols, gelatins, cocoa- butter or other low melting vegetable waxes or fats or their synthetic analogs.

Typical dermal and transdermal formulations comprise a conventional aqueous or non- aqueous vehicle, for example a cream, ointment, lotion or paste or are in the form of a medicated plaster, patch or membrane.

No unacceptable toxicological effects are expected when compounds of the present invention are administered in accordance with the present invention.

ASSAY The potential for compounds of the invention to inhibit NS5B wildtype HCV polymerase activity may be demonstrated, for example, using the following in vitro assay:

In Vitro Detection of inhibitors of HCV RNA-dependent RNA Polymerase Activity

Incorporation of [³³P]-GMP into RNA was followed by absorption of the biotin labelled RNA polymer by streptavidin containing SPA beads. A synthetic template consisting of biotinylated 13mer-oligoG hybridised to polyrC was used as a homopolymer substrate.

Reaction Conditions were 0.5 μM [³³P]-GTP (0.2 Ci/mMol), 1 mM Dithiothreitol, 20 mM MgCI₂, 5mM MnCI₂, 20 mM Tris-HCI, pH7.5, 1.6 μg/mL polyC/0.256 μM biotinylated oligoG13, 10% glycerol, 0.01% NP-40, 0.2 u/μL RNasin and 50 mM NaCI.

HCV RNA Polymerase (Recombinant full-length NS5B (Lohmann et al, J. Virol. 71 (11), 1997, 8416 'Biochemical properties of hepatitis C virus NS5B RNA-dependent RNA polymerase and identification of amino acid sequence motifs essential for enzymatic activity') expressed in baculovirus and purified to homogeneity) was added to 10 nM final concentration.

5x concentrated assay buffer mix was prepared using 1 M MnCI₂ (0.25 mL), glycerol (4mL), 10% NP-40 (0.025 mL) and Water (7.225 mL), Total 10 mL.

2x concentrated enzyme buffer contained 1 M-Tris-HCI, pH7.5 (0.4 mL), 5M NaCI (0.2 mL), 1 M-MgCI₂ (0.4 mL), glycerol (1 mL), 10% NP-40 (10 μL), 1 M DTT (20 μL) and water (7.97 mL), Total 10 mL

Substrate Mix was prepared using 5x Concentrated assay Buffer mix (4μL), [³³P]-GTP (10 μCi/μL, 0.02μL), 25 μM GTP (0.4 μL), 0.4 u/μL RNasin (0.04 μL), 20 μg/mL polyrC/biotinylated-oligorG (1.6 μL), and Water (3.94 μL), Total 10 μL. Enzyme Mix was prepared by adding 1 mg/ml full-length NS5B polymerase (1.5 μL) to 2.811mL 2x-concentrated enzyme buffer.

The Assay was set up using compound (1μL), Substrate Mix (10 μL), and Enzyme Mix (added last to start reaction) (10 μL), Total 21 μL.

The reaction was performed in a U-bottomed, white, 96-well plate. The reaction was mixed on a plate-shaker, after addition of the Enzyme, and incubated for 1 h at 22°C. After this time, the reaction was stopped by addition of 40 μL 1.875 mg/ml streptavidin SPA beads in 0.1 M EDTA. The beads were incubated with the reaction mixture for 1h at 22°C after which 120 μL 0.1 M EDTA in PBS was added. The plate was sealed, mixed centrifuged and incorporated radioactivity determined by counting in a Trilux (Wallac) or Topcount (Packard) Scintillation Counter.

After subtraction of background levels without enzyme, any reduction in the amount of radioactivity incorporated in the presence of a compound, compared to that in the absence, was taken as a measure of the level of inhibition. Ten concentrations of compounds were tested in three- or fivefold dilutions. From the counts, percentage of inhibition at highest concentration tested or IC₅₀s for the compounds were calculated using Grafit3 or Grafit4 software packages.

The exemplified compounds all had an IC₅₀ of <50μM in the above-described assay. Accordingly, the compounds of the invention are of potential therapeutic benefit in the treatment and prophylaxis of HCV. Preferred compounds had an IC₅₀ of <10μM.

The pharmaceutical compositions according to the invention may also be used in combination with other therapeutic agents, for example immune therapies ((eg. Interferon, such as Interferon alfa-2a (Roferon-A; Hoffmann-La Roche), inteferon alpha-2b (Intron-A; Schering-Plough), interferon alfacon-1 (Infergen; Intermune), peginterferon alpha-2b (Peg- Intron; Schering-Plough) or peginterferon alpha-2a (Pegasys; Hoffmann-La Roche))), therapeutic vaccines, antifibrotic agents, anti-inflammatory agents such as corticosteroids or NSAIDs, bronchodilators such as beta-2 adrenergic agonists and xanthines (e.g. theophylline), mucolytic agents, anti-muscarinics, anti-leukotrienes, inhibitors of cell adhesion (e.g. ICAM antagonists), anti-oxidants (eg N-acetylcysteine), cytokine agonists, cytokine antagonists, lung surfactants and/or antimicrobial and anti-viral agents (eg ribavirin and amantidine). The compositions according to the invention may also be used in combination with gene replacement therapy.

The invention thus provides, in a further aspect, a combination comprising a compound of formula (I) or a physiologically acceptable salt or solvate thereof together with another therapeutically active agent, especially interferon and/or ribavirin. The combinations referred to above may conveniently be presented for use in the form of a pharmaceutical formulation and thus pharmaceutical formulations comprising a combination as defined above together with a pharmaceutically acceptable carrier thereof represent a further aspect of the invention.

The individual components of such combinations may be administered either sequentially or simultaneously in separate or combined pharmaceutical formulations. Appropriate doses of known therapeutic agents will be readily appreciated by those skilled in the art.

All publications, including but not limited to patents and patent applications cited in this specification are herein incorporated by reference as if each individual publication were specifically and individually indicated to be incorporated by reference as though fully set forth.

Claims

At least one chemical e s of Formula (I)

in which:

A represents hydroxy;

D represents aryl or heteroaryl;

E represents hydrogen, C_halky!, aryl, heteroaryl or heterocyclyl;

G represents hydrogen or dialkyl optionally substituted by one or more substituents selected from halo, OR¹, SR¹, C(O)NR²R³, CO₂H, C(O)R⁴, CO₂R⁴, NR²R³, NHC(O)R⁴, NHCO₂R⁴, NHC(O)NR⁵R⁶, SO₂NR⁵R⁶, SO₂R⁴, nitro, cyano, aryl, heteroaryl and heterocyclyl;

R¹ represents hydrogen, C^alkyl, arylalkyl, or heteroarylalkyl;

R² and R³ are independently selected from hydrogen, Cι-₆alkyl, aryl and heteroaryl; or R² and R³ together with the nitrogen atom to which they are attached form a 5 or 6 membered saturated cyclic group;

R⁵ and R⁶ are independently selected from the group consisting of hydrogen, d-ealkyl, aryl, heteroaryl, arylalkyl, and heteroarylalkyl; or R⁵ and R⁶ together with the nitrogen atom to which they are attached form a 5 or 6 membered saturated cyclic group; and

J represents C^alkyl, heterocyclylalkyl, arylalkyl or heteroarylalkyl;

and salts, solvates and esters thereof; provided that when A is esterified to form -OR where R is selected from straight or branched chain alkyl, aralkyl, aryloxyalkyl, or aryl, then R is other than ferf-butyl.

2. At least one chemical entity as claimed in claim 1 selected from the group consisting of: re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-terf-butylbenzoyl)-4-ethoxymethyl-5-(1 ,3-thiazol-2- yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid; re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-tert-butylbenzoyl)-4-methoxymethyl-5-(1 ,3-thiazol- 2-yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid; re/-(2S,5R)-2-lsobutyl-1 -(3-methoxy-4-terf-butylbenzoyl)-4-/7-propyloxymethyl-5-(1 ,3- thiazol-2-yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid; re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-tert-butylbenzoyl)-4-methylthiomethyl-5-(1 ,3-thiazol- 2-yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid; re/-(2S,5R)-2-lsobutyl-1-(3-methoxy-4-tert-butylbenzoyl)-4-methylsulfonylmethyl-5-(1 ,3- thiazol-2-yl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid; rel-(2S,5R)-1-(3-Methoxy-4-tert-butylbenzoyl)-4-(methoxymethyl)-5-(5-methylisoxazol-3- yl)-2-(1 ,3-thiazol-4-ylmethyl)-2,5-dihydro-1 H-pyrrole-2-carboxylic acid;

and salts, solvates and esters, and individual enantiomers thereof.

3. At least one chemical entity as claimed in claim 1 in which D represents optionally substituted phenyl.

4. At least one chemical entity as claimed in claim 3 in which D represents para-tert- butylphenyl optionally further substituted by halo, C_halky! or C^alkoxy.

5. At least one chemical entity as claimed in claim 1 in which E represents optionally substituted heteroaryl.

6. At least one chemical entity as claimed in claim 5 in which E represents thiazolyl or isoxazolyl, each of which may be optionally substituted.

7. At least one chemical entity as claimed in claim 1 in which G represents C_halky! optionally substituted by OR¹, SR¹, or SO₂R⁴.

8. At least one chemical entity as claimed in claim 7 in which G represents methyl optionally substituted by OR¹, SR¹ or SO₂R⁴.

9. At least one chemical entity as claimed in claim 7 or 8 in which R¹ represents C^ ₃alkyl.

10. At least one chemical entity as claimed in claim 7 in which R⁴ represents C_halky!.

11. At least one chemical entity as claimed in claim 1 in which J represents Cι-₆alkyl, arylalkyl or heteroarylalkyl.

12. At least one chemical entity chosen from compounds of Formula (I) as claimed in claim 1 and pharmaceutically acceptable salts and solvates thereof.

13. A method of treating or preventing viral infection which comprises administering to a subject in need thereof, an effective amount of at least one chemical entity chosen from compound

in which:

A represents hydroxy;

D represents aryl or heteroaryl;

E represents hydrogen, Cι-₆alkyl, aryl, heteroaryl or heterocyclyl;

R¹ represents hydrogen, C_halky!, arylalkyl, or heteroarylalkyl;

R² and R³ are independently selected from hydrogen, Ci-ealkyl, aryl and heteroaryl; or R² and R³ together with the nitrogen atom to which they are attached form a 5 or 6 membered saturated cyclic group;

R⁴ is selected from the group consisting of C_halky!, aryl, heteroaryl, arylalkyl, and heteroarylalkyl;

R⁵ and R⁶ are independently selected from the group consisting of hydrogen, C_halky!, aryl, heteroaryl, arylalkyl, and heteroarylalkyl; or R⁵ and R⁶ together with the nitrogen atom to which they are attached form a 5 or 6 membered saturated cyclic group; and

J represents C^ealkyl, heterocyclylalkyl, arylalkyl or heteroarylalkyl;

and salts, solvates and esters thereof; provided that when A is esterified to form -OR where R is selected from straight or branched chain alkyl, aralkyl, aryloxyalkyl, or aryl, then R is other than terf-butyl.

14. A method as claimed in claim 13 which involves inhibiting HCV replication.

15. A method as claimed in claim 13 in which the at least one chemical entity is administered in an oral dosage form.

16. At least one chemical en s of Formula (I)

in which: A represents hydroxy;

D represents aryl or heteroaryl;

E represents hydrogen, C^alkyl, aryl, heteroaryl or heterocyclyl;

G represents hydrogen or C-ι-₆alkyl optionally substituted by one or more substituents selected from halo, OR¹, SR¹, C(O)NR²R³, CO₂H, C(O)R⁴, CO₂R⁴, NR R³, NHC(O)R⁴, NHCO₂R⁴, NHC(O)NR⁵R⁶, SO₂NR⁵R⁶, SO₂R⁴, nitro, cyano, aryl, heteroaryl and heterocyclyl;

R represents hydrogen, C-ι-₆alkyl, arylalkyl, or heteroarylalkyl;

R² and R³ are independently selected from hydrogen, Cι._ealkyl, aryl and heteroaryl; or R² and R³ together with the nitrogen atom to which they are attached form a 5 or 6 membered saturated cyclic group;

R⁴ is selected from the group consisting of Cι_₆alkyl, aryl, heteroaryl, arylalkyl, and heteroarylalkyl;

R⁵ and R⁶ are independently selected from the group consisting of hydrogen, dialkyl, aryl, heteroaryl, arylalkyl, and heteroarylalkyl; or R⁵ and R⁶ together with the nitrogen atom to which they are attached form a 5 or 6 membered saturated cyclic group; and

J represents C-ι-₆alkyl, heterocyclylalkyl, arylalkyl or heteroarylalkyl; and salts, solvates and esters thereof; provided that when A is esterified to form -OR where R is selected from straight or branched chain alkyl, aralkyl, aryloxyalkyl, or aryl, then R is other than fetf-butyl;

for use in medical therapy.

17. At least one chemical entity as claimed in claim 16 in which the medical therapy is the treatment of viral infection.

18. At least one chemical entity as claimed in claim 17 in which the viral infection is HCV.

19. Use of at least one chemical entity chosen from compounds of Formula (I)

in which:

A represents hydroxy;

D represents aryl or heteroaryl;

E represents hydrogen, Cι-₆alkyl, aryl, heteroaryl or heterocyclyl;

G represents hydrogen or C_halky! optionally substituted by one or more substituents selected from halo, OR¹, SR¹, C(O)NR²R³, CO₂H, C(O)R⁴, CO₂R⁴, NR²R³, NHC(O)R⁴, NHCO₂R⁴, NHC(O)NR⁵R⁶, SO₂NR⁵R⁶, SO₂R⁴, nitro, cyano, aryl, heteroaryl and heterocyclyl;

R¹ represents hydrogen, C_halky!, arylalkyl, or heteroarylalkyl;

R² and R³ are independently selected from hydrogen, C₁.₆alkyl, aryl and heteroaryl; or R² and R³ together with the nitrogen atom to which they are attached form a 5 or 6 membered saturated cyclic group;

R⁴ is selected from the group consisting of Cι.₆alkyl, aryl, heteroaryl, arylalkyl, and heteroarylalkyl; R⁵ and R⁶ are independently selected from the group consisting of hydrogen, C,-₆alkyl, aryl, heteroaryl, arylalkyl, and heteroarylalkyl; or R⁵ and R⁶ together with the nitrogen atom to which they are attached form a 5 or 6 membered saturated cyclic group; and

J represents Cι-₆alkyl, heterocyclylalkyl, arylalkyl or heteroarylalkyl;

and salts, solvates and esters thereof; provided that when A is esterified to form -OR where R is selected from straight or branched chain alkyl, aralkyl, aryloxyalkyl, or aryl, then R is other than te/f-butyl;

in the manufacture of a medical for the treatment of viral infection.

20. Use as claimed in claim 19, in which the viral infection is HCV.

21. A pharmaceutical formulation comprising at least one chemical entity chosen from compounds of Formula (I) and salts, solvates and esters thereof as defined in claim 1 in conjunction with a pharmaceutically acceptable diluent or carrier.

22. A process for the preparation of a compound of Formula (I) as defined in claim 1 , comprising treatment of a comp

in which A is alkoxy, and D, E, G and J are as defined for Formula (I), with an acid.

23. A process as claimed in claim 22 in which A is fetf-butoxy.