WO1998043975A1 - Pyrolopyrrolone derivatives - Google Patents

Abstract

The present invention relates to therapeutically active bicyclic compounds, processes for the manufacture of said compounds, pharmaceutical formulations containing said compounds and the use of said compounds in treatment and prophylaxis, particularly of viral infections, more particularly of infections caused by viruses which encode for a serine protease enzyme, especially viruses of the herpes family. Thus, according to one aspect of this invention, we provide a compound of general formula (I) wherein R represents H, substituted or unsubstituted C1-3 alkyl; R1 represents optionally substituted heteroaryl or fused heteroaryl with one to four heteroatoms, R5CO, R5NHCO, R5CS or R5NHCS wherein R5 may be substituted or unsubstituted and represents H, C1-6 alkyl, C1-6 alkenyl, C3-7 cylcoalkyl or fused cycloalkyl, heteroaryl or fused heteroaryl containing one to four heteroatoms, aryl or fused aryl, or arylC1-3alkyl; R2 represents R6-X- or R3CO, wherein R3 may be substituted or unsubstituted and represents (A), (B), (C), (D), (E), (F), (G), or (H), optionally including one or more further heteroatoms; R4 represents R6-X-; R6 is optionally substituted heterocyclic or fused heterocyclic with 1-4 heteroatoms, heteroaryl or fused heteroaryl with 1-4 heteroatoms, C3-10cycloalkyl or fused cycloalkyl, aryl or fused aryl; and X represents a linker group chosen from C=O, NHC=O, C(=O)C=O, CH=CHCO, CH2CO, CH2 or SO2; and salts and solvates thereof.

Description

PYROLOPYRROLONE DERIVATIVES

The present invention relates to therapeutically active bicyclic compounds, processes for the manufacture of said compounds, pharmaceutical formulations containing said compounds and the use of said compounds in chemotherapy. In particular, we have found a novel group of bicyclic compounds which are effective in the treatment and prophylaxis of viral infections, more particularly infections caused by viruses which encode for a serine protease enzyme, especially viruses of the Herpes family.

The Herpes family of viruses is responsible for a wide range of infectious diseases in several species especially chicken pox, shingles, retinitis, pneumonitis and keratitis in humans and diseases of the skin and mucosa, including keratitis in rabbits, herpetic encephalitis in mice, Herpes viruses include HSV1 and HSV2 (Herpes Simplex Virus type 1 and type 2), hCMV and mCMV (human and murine cytomegalovirus), VZV (varicella zoster virus), EBV (Epstein barr virus) HHV6 and HHV8 (human herpes viruses, types 6 and 8).

All herpes viruses encode a serine protease which is crucial for viral replication as it cleaves the assembly protein precursor during capsid maturation. Protease deficient mutants do not cleave this scaffold protein thus giving rise to immature virions. We have found that inhibitors of this protease can have a similar effect thus preventing formation of mature, infectious viral progeny in infected cells.

We now present a novel group of bicyclic compounds which are inhibitors of herpes virus proteases and are thus of benefit in the treatment, prophylaxis and suppression of virus infections caused by viruses of the Herpes family. The compounds show broad spectrum activity against herpes viruses including HSV 1 and 2, hCMV, mCMV and VZV.

Thus, according to one aspect of this invention, we provide a compound of the general formula (I) Thus, according to one aspect of this invention, we provide a compound of the general formula (I) o

and salts and solvates thereof, wherein:

R represents H or substituted or unsubstituted C,.₃ alkyl, preferably methyl;

Rn represents optionally substituted heteroaryl or fused heteroaryl with one to four heteroatoms, R₅CO, R₅NHCO, R₅CS or R₅NHCS wherein R₅ may be substituted or unsubstituted and represents H, C-|.₆ alkyl, C-,-₆ alkenyl, C₃.₇ cycloalkyl or fused cycloalkyl, heteroaryl or fused heteroaryl containing one to four heteroatoms, aryl or fused aryl, or arylC₁_₃alkyl;

R₂ represents R₆-X- or R₃CO, wherein R₃ may be substituted or unsubstituted and represents

optionally including one or more further heteroatoms;

R₄ represents R₆-X-;

R₆ is optionally substituted heterocyclic or fused heterocyclic with 1-4 heteroatoms, heteroaryl or fused heteroaryl with 1-4 hetero atoms, C₃_ ₁₀cycloalkyl or fused cycloalkyl, aryl or fused aryl; and X represents a linker group chosen from C=O, NHC=O, C(=O)C=O, CH=CHCO, CH₂CO, CH₂ or SO₂.

Where any of the groups R, R_{1 f} R₃ or R₆ as defined above are substituted, they are preferably singly or doubly substituted.

When used herein "alkyl" includes branched as well as straight chain alkyl.

Where stated, nitrogen atoms may be substituted by C^alkyl or -CO-C^alkyl groups and alkyl, aryl or heteroaryl carbon atoms may be substituted by halogen or hydroxy groups, C=O, O-C₁.₃haloalkyl, NH₂, NO₂, C^alkyl, C.,-₃alkoxy, C,. ₃hydroxyalkyl, fluorinated C.,.₃alkoxy, SO₂R₇, C(O)R₇, CO₂R₇, (CH₂)_nCO₂R₇, (CH₂)_nOC(O)NHR₇, (CH₂)_nC(O)NHR₇, (CH₂)_nNR₇R₈, CONHR₇, and

(CH₂)_nNHCOR₇ wherein n is an integer from 1 to 4 and R₇ and R₈ independently represent H, C₁.₃alkyl, C.,-₃haloalkyl, or together form a ring.

When used herein "aryl" includes aromatic groups having up to two rings, including phenyl and naphthyl, and arylalkyl, heteroaryl are to be read accordingly. Thus, heteroaryl includes aromatic groups having up to two rings containing one or more (e.g. 1-4) heteroatoms e.g. pyridine, quinoline thiadiazole, thiophene, benzoxazole and benzothiophene.

Suitable R-_\ groups include:

Q = CHoOH, <V CH₂0H, or

CH₂NHC0Me, CH₂NHCOMe,

Cl,

CH₂CH₂OH, OMe, CH₂CH₂NHCO e, F,

S0₂CHF₂

CH₂C0₂Et, OCF₃, CONHR, N0₂, CH₂NH₂ CONHR, or C0₂H CH₂NH₂ or C0₂H

D = NHCOMe,

OMe,

OH,

C0₂H or halogen

het = 5 or 6 membered

E = NHCOMe heteroaryl or fused heteroaryl OMe, OH, C0₂H containing 1-4 heteroatoms

Preferred Ri substituents include

optionaiiy substituted with one or more groups selected from halogen, NH₂, N0₂, C_halky!, C^alkoxy, C_Lahydroxyalkyl, fluoriπated C_Lgalkoxy, S0₂R₇, C0₂R₇, (CH₂)_nC0₂R_7l (CH₂)_nOC(0)NHR_7l CONHR₇, and (CH₂)_nNHCOR₇ wherein n is an integer from 1 to 4 and R₇ represents H, C,.₃alkyl₆, C^haloalkyl;

and b)

wherein A and B independently represent H, C alkyl or

A and B may be linked to form a saturated or unsaturated carbocyciic ring.

Where R₅ is arylC₁.₃alkyl, arylC^alkyl is preferred.

Preferably, R₃ represents

Preferred Rg substituents include

wherein Y represents a hetero atom such as O, S or N, wherein N may optionally be substituted e.g. by C alkyl or -CO-C^alkyl groups, W represents hydrogen or C₁₄ alkyl and Z represents hydrogen, halogen, C_halky!, C₃. ₆cycloalkyl, aryl, C^alkoxy, C_nH_2n+ιOC_mH_2m wherein n and m independently represent an integer of from 1 to 3, CF₃, O-haloC^alkyl, S-C.,.₃alkyl or S-haloC,_ ₃alkyl, O-aryl, C₂-₄alkenyl, and N(A)B wherein A and B are independently selected from H and C^ alkyl;

Preferred Z substituents include phenyl, OPhenyl, isopropyl, t-butyl, n-propoxy, SCF₃, OCF₃ and OCHF_2.

Formula (I) hereinbefore shows the relative stereochemistry of the chiral centres. Generally, we prefer a compound of formula (I) in which the hydrogens at the two ring fusion carbons are trans to one another and the hydrogen at the R- substituted carbon is cis to that at the adjacent ring fusion carbon. More specifically, we prefer a chiral compound of formula (I) in the cis-trans form with SRS absolute stereochemistry as shown below in formula (la), in which the hydrogens at the two ring fusion carbons are trans to one another and the hydrogen at the R- substituted carbon is cis to that at the adjacent ring fusion carbon. The absolute configuration is set out below:

Where compounds of formula (I) are able to form physiologically acceptable salts, these are included within the present invention. Suitable physiologically acceptable salts of the compounds of formula (I) include inorganic base salts such as alkali metal salts (for example sodium and potassium salts) and ammonium salts and organic base salts. Suitable organic base salts include amine salts such as trialkylamine (e.g. triethylamine), dialkylamine (e.g. dicyclohexylamine), optionally substituted benzylamine (e.g. phenylbenzylamine or p-bromobenzylamine), procaine, ethanolamine, diethanolamine, N- methyiglucosamine and tri(hydroxymethyl)methylamine salts and amino acid salts (e.g. lysine and arginine salts). Suitable inorganic and organic acid salts include the hydrochloride, trifluoroacetate and tartrate. Certain compounds of formula (I) embodying the invention have structures as follows:

(II)

Z= tert-butyl or PhO

(III)

(IV)

(V)

(V)b R1 =

N "

(VI)

(Vl)a R1 Z= /so-propyl, tert-butyl, CF₃, CF₃0, CF₃S, CH₃CH₂CH₂0, PhO

(Vl)b R1 Z= /sopropyl, CF₃0

(Vl)c R1= Z= /^'sopropyl, tert-butyl, CF₃0

(Vl)d R1= _/sopropyl, CF₃O

(VII) (VIII) Intermediates and processes described herein form further aspects of the invention.

The potential for compounds of formula (I) to inhibit activity may be demonstrated, for example, using the following enzyme and in vitro whole cell antiviral assays:

In vitro pNA assay of viral serine protease inhibitor activity

The hCMV serine protease used is a mutant of the 30K protease lacking the internal cleavage site (Ala142/Ala143) and which has been cloned in E.coli to produce active enzyme (hCMV δAla protease). IC₅₀ data for test compounds are determined both without preincubation and after preincubation of the enzyme with test inhibitor compound for 15 minutes in order to demonstrate time dependency. Test compounds are dissolved in DMSO, serially diluted and added at a range of concentrations (from 100μM - 0.195μM) to a reaction containing 0.5μM CMV δAla protease, 100mM HEPES pH7.5, 0.2mM EDTA, 10mM NaCI, 1 mM DTT, and 30% glycerol. The reaction mixture is pre- incubated at 32°C for 0 minutes or 15 minutes prior to addition of 4mM oligopeptide substrate (RESYVKA-pNA), and then analysed at 32°C in a BIO- TEK Bio Kinetics Reader EL340i.

The assay substrate is RESYVKA-pNA

RESYVK-pNA Ξ-≡

CMV Protease

O ,NO„ RESYVK + pNA ___≡ RESYVr 3

OH

H₂N

Me (λ max =405nm) The plate reader monitors production of pNA and calculates the reaction rates over 30 minutes. The rates are plotted against inhibitor concentration and IC₅₀ results determined at 0 and 15 minutes preincubation. Rate constant data (K_j) are obtained in a similar manner to IC₅₀ data without preincubation, with the exception that reaction rates are measured over 67.5 minutes, giving steady state rates. Curves are plotted of rate against inhibitor concentration and the K_| is calculated using a tight binding inhibition equation.

In vitro anti -viral assays

Enzyme Linked ImmunoSorbant Assay (ELISA) for HCMV

Tissue culture.

Compounds were formulated to 40mM in DMSO, then further diluted to 4 times the highest required final concentration in assay medium (bicarbonate-buffered Dulbecco's modification of Eagle's medium DMEM supplemented with 5% v/v foetal calf serum, 2mM w/v glutamine and antibiotics). 100μl of four test compounds were transferred to 3 wells each in the first row of a 96-well, plastic, tissue-culture plate, and serial doubling dilutions in assay medium were made down the plate to the penultimate row. 50μl of assay medium were added to all wells in the first column for each compound, and 50μl of HCMV strain Ad 169, diluted in medium to give a multiplicity of infection of 0.01 plaque-forming units per cell, were added to the wells in the remaining pairs of columns. Finally, 100μl of MRC-5 cell suspension at a concentration of 10⁵ cells per ml were added to each well. The plate was then incubated at 37°C in a humidified 5% CO₂ atmosphere for 7 days.

Dectection of Viral Antigens (7d.p.i)

The growth medium was removed and the cell monolayers were washed once with phosphate buffered saline (PBS). The cells were fixed by the addition 1 :1 acetone:methanol for 5 minutes and washed again with PBS. In order to inhibit subsequent non-specific binding of antibodies, 100μl of PBS containing 0.05% Tween 20 and 2% w/v skimmed milk powder (blocking buffer) were added to each well and the plate was incubated for 1 hour at 37°C . The blocking solution was removed and the cells were washed once with PBS/0.05% Tween 20 (ELISA wash). The primary, murine, monoclonal antibody, specific for HCMV glycoprotein B, was diluted to 2μg/ml in blocking buffer and 50μl were added to each well. Following incubation for 1.5 hours at 37°C, the primary antibody solution was removed and the plate was washed 3 times. Horseradish peroxidase labelled, rabbit anti-mouse, polyclonal antibody, pre-adsorbed on uninfected MRC-5 cells, was diluted 1/1 ,500 in blocking buffer and 50μl were added to each well. After further incubation at 37°C for 1.5 hours, the secondary antibody solution was removed and the plate was washed thoroughly 5 times and dried. 50μl of orthophenylenediamine/peroxide substrate in urea buffer were added to each well, and colour was allowed to develop at room temperature. The reaction was stopped by the addition of 25μl per well of "20%" H₂SO and the plate was read spectrophotometrically at 490nm.

The 50% inhibitory concentration (IC50) value of an active compound was calculated by regression analysis of the plot of concentration against percentage reduction in absorbance compared to drug-free virus controls.

On completion of the ELISA, the plate was washed, dried and the cells stained with 20% carbol fuchsin. An assessment of the lowest in-assay cytotoxic concentration of each compound was made by microscopic examination of both uninfected and infected cell monolayers.

Anti-viral plaque reduction whole cell assays

Human cytomegalovirus (hCMV) Monolayers are formed by seeding 24-well tissue culture plates with 10⁵ MRC-5 human fibroblasts per well suspended in Dulbecco's modification of Eagle's medium (DMEM) supplemented with 10% foetal calf serum (FCS), 1 % non- essential amino acid solution and 2mM L-glutamine. Following incubation at 37°C overnight in a 5% CO₂ atmosphere and subsequent removal of the growth medium, the cells are infected with 0.2ml of HCMV (strain AD169) suspension containing approximately 100 plaque-forming units, and maintained at 37°C for 1 hour prior to overlaying with DMEM containing 4% FCS and 1% carboxymethyl-cellulose. After 53 hours further incubation at 37°C in 5% CO₂, the original overlay is removed by aspiration and replaced by a similar overlay containing serial doubling dilutions of the test drug in the range 100-1.56μM, freshly prepared from a 40mM stock dissolved in dimethyl sulphoxide. Two further replacements with overlay containing freshly prepared dilutions of drug are carried out at 72 & 77 hours post infection. On day 6 the cell sheets are fixed with 10% formol saline and stained with 0.3% methylene blue. Plaques are counted microscopically, and the mean count of duplicate wells at each drug dilution are calculated as a percentage of the drug-free virus control wells. The 50% inhibitory concentration of the test compound is calculated by regression analysis of the curve of percentage plaque reduction against drug concentration.

Human simplex virus (HSV)

The assay for HSV is broadly similar to that for hCMV with the following modifications;

(a) Vero cells grown in DMEM supplemented with 2mM L-glutamine and 5%

FCS replace the MRC-5s. (b) The duration of the assay is reduced to 48 h.p.i. for both HSV 1 (strain

SC16) and HSV 2 (strain 186), with a delayed addition of drug dilutions in

DMEM + 2% FCS at 5.5 h.p.i.

(c) The cell monolayers are fixed using a 5% aqueous solution of glutaraldehyde and stained with carbol fuchsin.

Varicella Zoster Virus (VZV)

The assay for VZV is also broadly similar to that for hCMV with the following modifications; (a) MRC5 cells are infected with 0.2ml of VZV strain G31 at 37°C for 90min.

(b) The overlay does not contain carboxymethyl cellulose. Drug dilutions are added in the initial overlay at 90 minutes post-infection.

(c) The cell monolayers are fixed as for hCMV at 96 hours post infection.

Cytotoxicity assay

Vero cells

4,000 Vero cells suspended in 75μl of DMEM medium supplemented with 5% FCS are seeded into each well of a 96-well microplate. The cells are allowed to settle and adhere for 1 hour at 37°C, then 75μl per well of freshly prepared doubling dilutions of the compound from 500μM are added. Following 96 hours incubation at 37°C, 20μl of a 5mg/ml solution of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide (MTT) in phosphate buffered saline are added to each well. After a further 2 hour incubation at 37 °C, the supernatants are aspirated from the wells and 125μl of acidified isopropanol containing 0.5% SDS are added to each. The plate is maintained on a shaking incubator for 20 minutes then read spectrophotometrically at 590nm. The mean absorbances of replicate test wells are expressed as percentages of cell control well values, and are then plotted against drug concentration to allow calculation of the 50% toxic dose (CCID₅₀)

Compounds of formula (I) are of potential therapeutic benefit in the treatment and amelioration of the symptoms of many herpes virus diseases. Such diseases particularly include chicken pox and shingles (varicella and Herpes zoster viruses, respectively), keratitis in rabbits, herpetic encephalitis in mice, cutaneous herpes in guinea pigs, cold sores and genital herpes in humans (herpes simplex virus), retinitis, pneumonitis and keratitis in humans (hCMV), as well as diseases caused by Epstein Barr Virus (EBV), human herpes virus 6 (HHV 6), HHV 7 and HHV 8.

Compounds of the invention may also be useful for the treatment or prophylaxis of cardiovascular system diseases, such as thrombosis, arteriosclerosis and particularly restenosis, recurrent narrowing or occlusion of a coronary valve or vessel, in which CMV has been implicated.

As indicated above, compounds of formula (I) are useful in human or veterinary medicine, in particular as inhibitors of viral serine proteases, in the management of herpes family virus infections.

Thus, there is provided as a further aspect of the present invention a compound of formula (I) or a physiologically acceptable salt or solvate thereof for use in human or veterinary medicine, particularly in the treatment of conditions caused by viruses of the Herpes family, such as HSV or CMV infections. It will be appreciated that references herein to treatment extend to prophylaxis, prevention of recurrence and suppression of symptoms as well as the treatment of established conditions.

According to another aspect of the invention, there is provided the use of a compound of formula (I) or a physiologically acceptable salt or solvate thereof in the manufacture of a medicament for the treatment of conditions caused by viral infections, more particularly caused by viruses of the Herpes family, such as HSV or CMV infections.

In a further or alternative aspect there is provided a method for the treatment of a human or animal subject with a condition caused or mediated by a virus of the Herpes family, which method comprises administering to said human or animal subject an effective amount of a compound of formula (I) or a physiologically acceptable salt or solvate thereof.

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.

There is also provided according to the invention a process for preparation of such a pharmaceutical composition which comprises mixing the ingredients.

The compounds according to the invention may, for example, be formulated for oral, buccal, parenteral, topical or rectal administration.

Tablets and capsules for oral administration may contain conventional excipients such as binding agents, for example syrup, acacia, gelatin, sorbitol, tragacanth, mucilage of starch or polyvinyl pyrrolidone; fillers, for example, lactose, microcrystalline cellulose, sugar, maize- starch, calcium phosphate or sorbitol; lubricants, for example, magnesium stearate, stearic acid, talc, polyethylene glycol or silica; disintegrants, for example, potato starch, croscarmellose sodium or sodium starch glycollate; or wetting agents such as sodium lauryl sulphate. The tablets may be coated according to methods well known in the art. Oral liquid preparations may be in the form of, for example, aqueous or oily suspensions, solutions, emulsions, syrups or elixirs, or may be presented as a dry product for constitution with water or other suitable vehicle before use. Such liquid preparations may contain conventional additives such as suspending agents, for example, sorbitol syrup, methyl cellulose, glucose/sugar syrup, gelatin, hydroxymethyl cellulose, carboxymethyl cellulose, aluminium stearate gel or hydrogenated edible fats; emulsifying agents, for example, lecithin, sorbitan mono-oleate or acacia; non-aqueous vehicles (which may include edible oils), for example almond oil, fractionated coconut oil, oily esters, propylene glycol or ethyl alcohol; or preservatives, for example, methyl or propyl p-hydroxybenzoates or sorbic acid. The preparations may also contain buffer salts, flavouring, colouring and/or sweetening agents (e.g. mannitol) as appropriate.

For buccal administration the compositions may take the form of tablets or lozenges formulated in conventional manner.

The compound may also be formulated as suppositories, e.g. containing conventional suppository bases such as cocoa butter or other glycerides.

The compound according to the invention may also be formulated for parenteral administration by bolus injection or continuous infusion and may be presented in unit dose form, for instance as ampoules, vials, small volume infusions or pre- filled syringes, or in multi-dose containers with an added preservative. The compositions may take such forms as solutions, suspensions, or emulsions in aqueous or non-aqueous vehicles, and may contain formulatory agents such as anti-oxidants, buffers, antimicrobial agents and/or toxicity adjusting agents. Alternatively, the active ingredient may be in powder form for constitution with a suitable vehicle, e.g. sterile, pyrogen-free water, before use. The dry solid presentation may be prepared by filling a sterile powder aseptically into individual sterile containers or by filling a sterile solution aseptically into each container and freeze-drying. By topical administration as used herein, we include administration by insufflation and inhalation. Examples of various types of preparation for topical administration include ointments, creams, lotions, powders, pessaries, sprays, aerosols, capsules or cartridges for use in an inhaler or insufflator or drops (e.g. eye or nose drops).

Ointments and creams may, for example, be formulated with an aqueous or oily base with the addition of suitable thickening and/or gelling agents and/or solvents. Such bases may thus, for example, include water and/or an oil such as liquid paraffin or a vegetable oil such as arachis oil or castor oil or a solvent such as a polyethylene glycol. Thickening agents which may be used include soft paraffin, aluminium stearate, cetostearyl alcohol, polyethylene glycols, microcrystalline wax and beeswax.

Lotions may be formulated with an aqueous or oily base and will in general also contain one or more emulsifying agents, stabilising agents, dispersing agents, suspending agents or thickening agents.

Powders for external application may be formed with the aid of any suitable powder base, for example, talc, lactose or starch. Drops may be formulated with an aqueous or non-aqueous base also comprising one or more dispersing agents, solubilising agents or suspending agents.

Spray compositions may be formulated, for example, as aqueous solutions or suspensions or as aerosols delivered from pressurised packs, with the use of a suitable propellant, e.g. dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, 1 ,1 ,1 ,2,3,3,3-heptafluoropropane, 1 ,1 ,1 ,2- tetrafluorethane, carbon dioxide or other suitable gas.

Capsules and cartridges for use in an inhaler or insufflator, of for example gelatin, may be formulated containing a powder mix of a compound of the invention and a suitable powder base such as lactose or starch. The pharmaceutical compositions according to the invention may also be used in combination with other therapeutic agents, for example other anti-viral and/or antimicrobial agents.

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.

The combination 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.

Compounds of the invention may conveniently be administered in amounts of, for example, 0.01 to 50mg/kg body weight, suitably 0.05 to 25mg/kg body weight and preferably 1 to 25mg/kg body weight orally, one or more times a day. The precise dose will of course depend on the age and condition of the patient, the particular route of administration chosen, and the disease being treated.

The compounds of the formula (I) have useful duration of action.

The compounds of formula (I) and salts and solvates thereof may be prepared by the methodology described hereinafter, constituting a further aspect of this invention.

A process according to the invention for preparing a compound of formula (I) comprises: (i) condensation of a compound of formula (M) wherein R and R₁ are as defined above:

with a compound R₃COOH or R₃COY, where Y is a leaving group such as halogen e.g. chlorine and R₃ is as defined above; or

(ii) condensation of a compound of formula (N) wherein R and R₂ are as defined above:

with a compound R₅COOH, R₅COY, R₅NCO or R.,Y' wherein Y is a leaving group such as halogen, e.g. chlorine, Y' is a halogen such as bromine or fluorine, R-, is optionally substituted heteroaryl or fused heteroaryl with one to four heteroatoms and R₅ is as defined; or

(iii) preparation of a compound of formula (I) by introduction of a substituent R₄ as defined above, by treatment of the corresponding precursor compound

optionally following deprotection of the corresponding nitrogen-protected compound, with a compound R₆COOH, RgCOCOOH (both with suitable activation), R₆COY, R₆SO₂Y or R₆NCO wherein Y is a leaving group such as a halogen e.g. chlorine and R₆ is as defined above; or

(iv) converting one compound of the formula (I) into another compound of the formula (I); or

(v) purifying one diastereomer of the compound of formula (I) from its racemic mixture by e.g. one or more of the following methods: chromatography of a diastereomeric mixture, crystallisation or resolution using a chiral template;

and where desired or necessary converting a resultant free acid or base compound of formula I into a physiologically acceptable salt form or vice versa or converting one salt form into another physiologically acceptable salt form.

Process (i) and Process (ii)

The condensation reaction with R₃COOH is suitably carried out in the presence of a coupling agent such as O-benzotriazol-1-yl-N,N,N',N'-tetramethyluronium tetrafluoroborate (TBTU) used with 1-hydroxybenzotriazole (HOBT) in the presence of an organic base such as N,N-diisopropylethylamine (DIPEA) and a solvent such as dichloromethane, tetrahydrofuran or dimethylformamide at a temperature of suitably between 0°C and ambient. It will be appreciated that on introduction of R₅COOH, where the linker group X between the aromatic group R₆ and the nitrogen atom of the group R₃ represents NHC=O, it may be necessary to protect the nitrogen atom of the group X. It will also be appreciated that as an alternative to using R₅COOH and R₃COOH, acid derivatives such as the acid chloride, acid anhydride, or a mixed anhydride may be used as is the case with R₅COOH, where the mixed anhydride, formed using pivaloyl chloride and triethylamine, is preferred. Reaction conditions will be modified accordingly, for instance by inclusion of a base e.g. triethylamine, N,N-diisopropylethylamine lithium hexamethyldisilazide or sodium hydride. Suitable solvents include tetrahydrofuran and the acylation is carried out between -78°C and ambient temperature. Reaction with R₅NCO or the isocyanate R₅NCS may be carried out in the presence of a base such as sodium hydride in a solvent such as tetrahydrofuran at a temperature suitably between 0°C and ambient. In instances where chlorosulphonyl isocyanate is used, the base may be omitted.

Reaction with R.,Y' wherein Y' is bromine is the Goldberg variant of the Ullman reaction, ref: A. Greiner, Synthesis 1989 p.312, and is carried out in the presence of a copper catalyst such as copper(l)chloride in the presence of a base such as potassium carbonate in a high boiling inert solvent such as xylene at a suitably elevated temperature usually at reflux. The reaction is advantageously carried out in the presence of a solid-liquid phase transfer catalyst such as tris(3,6-dioxaheptyl)amine (TDA-1 ).

Reaction with R.,Y' wherein Y' is fluorine is carried out in the presence of a base such as sodium hydride or lithium hexamethyldisilazide in a suitable solvent such as tetrahydrofuran at a temperature suitably between -78°C and ambient depending on the base used. The reaction with R₃COY is carried out in the presence of an organic base such as triethylamine in a suitable solvent such dichloromethane at a temperature suitably between 0°C and ambient. The reaction with R₅COY is carried out in the presence of an organic base such as lithium hexamethyldisilazide in a suitable solvent such tetrahydrofuran at a temperature suitably between -78°C and 0°C.

Process (iii) Removal of an N-protecting group may be carried out in a conventional manner, for instance by treatment with acid such as trifluoroacetic acid (where the protecting group is t-butoxycarbonyl (Boc) or 4-methoxybenzyloxycarbonyl (Moz)) or hydrogenation over palladium on carbon in solvents such as ethyl acetate, tetrahydrofuran or isopropanol or by transfer hydrogenation (where the protecting group is benzyloxycarbonyl (Cbz)).

The condensation reaction with R₆COOH and RgCOCOOH is suitably carried out under similar conditions to those described for processes (i) and (ii). The reaction with R₆COY and R₆SO₂Y is carried out in the presence of an organic base such as triethylamine in a suitable solvent such dichloromethane at a temperature suitably between 0°C and ambient. Reaction with RgNCO is carried out in a suitable solvent such as acetonitrile at a temperature suitably between 0°C and ambient. The reaction may be modified for instance by the addition of base to neutralise the acid addition salt of the starting translactam.

Process (iv)

Examples of typical interconversions include reducing a NO₂ group to NH₂, and alkenyl group to alkyl.

Process (v)

Isolation of a single diastereomer may be achieved by conventional methods such as flash chromatography on silica gel; chiral chromatography (e.g. chiral HPLC) and crystallisation with a homochiral acid (e.g. tartaric acid) or base (e.g. norephedrine).

Physiologically acceptable base salts of the compounds of formula (I) may conveniently be prepared by treating a compound of formula (I) with a suitable base such as a bicarbonate, e.g. sodium bicarbonate, in the presence of a suitable solvent. Acid salts such as the hydrochloride, trifluoroacetate or tartrate may be prepared by treating a basic compound of formula (I) with the desired acid.

Preparation of intermediates

Certain intermediates useful in the preparation of compounds of formula (I) may be prepared using the following further processes:

(vi) Compounds of formula (M) above may be prepared by condensation of a compound of formula (P) wherein R₈ is a nitrogen protecting group,

R₅COY, R₅NCO, or R.,Y' wherein Y is a leaving group such as halogen, e.g. chlorine, Y' is a halogen such as bromine or fluorine, R₁ is optionally substituted heteroaryl or fused heteroaryl with one to four heteroatoms and R₅ is as defined above, in a process analogous to (ii) above, followed by deprotection.

(vii) N-protected compounds such as those of formula (XXIII) below may be prepared by condensation of a compound of formula (Q) wherein

P₂ and P₃ are different and represent nitrogen protecting groups o

II _^p N ³

(Q)

?7 sequentially with a base and then with a compound RY, wherein Y is a leaving group such as those noted above and R represents C,.₃ alkyl e.g. methyl. Further intermediates may then be formed by selective deprotection of P₂ and/or

P₃ Similarly, where P₂ is a nitrogen protecting group and R., is , as in formula (R) below,

N-protected compounds of formula (M) above may be prepared by condensation of a compound of formula (R) sequentially with a base and then with a compound RY, wherein Y is a leaving group such as those noted above and R represents C,_₃ alkyl e.g. methyl.

These reactions are suitably carried out in the presence of a strong base, such as LiHMDS, in the presence of a solvent such as tetrahydrofuran, at a reduced temperature such as -78° to 0°C.

(viii) Compounds of formula (N) above may be prepared by condensation of a compound of formula (S) wherein P is a nitrogen protecting group,

with a compound R₃COOH or R₃COY, where Y is a leaving group such as halogen e.g. chlorine and R₃ is as defined above, in a process analogous to (vi) followed by deprotection.

(xi) Nitrogen-protected intermediates useful in process (iii) above i.e. compounds of formula (XXXII) in Scheme 2 below, may be prepared according to Scheme 2 or by condensation of a compound of formula (M) with a nitrogen- protected L-proline of formula (T), or a nitrogen-protected L-azetidine carboxylic acid of formula (U), wherein P₃ is a nitrogen protecting group,

or an acid derivative thereof such as the acid chloride, in a process analogous to (i) above followed by deprotection.

Preparation of compounds of Formula (I)

In one example of this aspect of the invention, compounds of Formula (I) wherein R-, represents R₅CO as defined above may conveniently be prepared according to the methodology shown in the following general scheme 1 :

Scheme 1

0

HO-' // NH,

H₂N

(a)

t7* * O

R O o

N

N 2_ N

N

(XXIII) (d)

(XXII)

(e)

O

R O

R

NH n NCOR₅

N N

(XXIV) (f) (XXV)

(g)

O

R

R o

. NCOR-

R₃CO N

V H NCOR-

(XXVII) (h) N f

. /

(XXVI) Steps (a)

One possible route to the desired 2R.3S enantiomer of the intermediate (XX) is given below (steps p-u), wherein P is a N-protecting group, preferably Boc (t- butyloxycarbonyl), P₂ is another N-protecting group, preferably Cbz (benzyloxycarbonyl) and R₇ is suitably a C^ straight or branched alkyl group e.g. ethyl or t-butyl.

(S) diamiπobutyric acid

Step p The compounds of formula (XXXV)(S) are either known compounds or may be prepared in analogous manner to known compounds. The reaction is suitably carried out using PIFA (phenyi iodosyibis(trifiuoroacetate) and a base such as pyridine in an aqueous solvent, such as aqueous THF, dioxan or acetonitrile. This is the method of Stansfield, C.F. Organic Preparations and Procedures Int., 1990, 22(5), 593-603.

Step q This protection reaction may be carried out in a conventional manner. For instance it is suitably carried out in a water miscible solvent such as THF, DMF or dioxan using N-(benzyloxycarbonyloxy)succinimide, benzyloxycarbonyl chloride, or any suitable source of the benzyloxycarbonyl group, with pH adjustment to alkaline with sodium carbonate.

As an alternative, step q¹, the compound of formula (XXXVII) can be prepared in conventional manner from (S) diaminobutyric acid, the starting material shown in Schemes 1 and 2, herein.

Step r

This reaction is suitably carried out in two stages. Firstly, reacting at reduced temperature with N-methylmorpholine and then an alkyl chloroformate such as ethyl chloroformate, in an organic solvent such as DCM, dioxan or THF. Secondly, the intermediate product while in solution is reduced, suitably with sodium borohydride dissolved in a suitable solvent such as water, at reduced temperature, such as -20° to 10°C.

Step s This oxidation reaction may be suitably carried out in any suitable manner, for instance using oxalyl chloride in DMSO and methylene dichloride under nitrogen at reduced temperature, such as -30° to -70°C, followed by triethylamine. The intermediate (XXXIX) suitably is not isolated.

Step t

This reaction is suitably carried out using a Wittig reagent such as a triphenylphosphorane R₇O₂CH=PPh₃, or may also be carried out using a phosphonate in a Wadsworth-Emmons reaction.

Step u

This Michael addition reaction is suitably carried out using lithium bis(trimethylsilylamide) or other suitable strong base in a suitable organic solvent such as THF, ether or toluene, and preferably a complexing agent such as tetramethylethylenediamine is also present. Alternatively, for step (a), an achiral preparation may be employed and the mixture of enantiomers (XX) may be resolved so that the required 2R,3S- enantiomer is brought through step (b) and the following steps chirally. Any suitable resolving agent , preferably (+) di-p-toluoyl-tartaric acid ((+)-DPTTA) followed by typically two recrystallisations suitably from ethanol, is used to give the 2R,3S-enantiomer as the tartrate, (XX)2R,3S.

(XX)2R,3S

As a further alternative, the racemic mixture (XX) may be processed through steps (b) to (h) and the enantiomeric separation carried out at a later stage, for example at step (h) as described further below.

Step (b)

Deprotection carried out with addition of acid such as trifluoroacetic acid followed by cyclocondensation with an alkyl Grignard reagent, suitably 'ButylMgCI in a solvent, suitably tetrahydrofuran, at a temperature between -20°C and ambient.

Step (c)

Reprotection is carried out in a conventional manner. When P₃ is Boc, this is suitably achieved with Boc₂O in a solvent suitably tetrahydrofuran at a suitable temperature such as -78°C in the presence of a base such as lithium hexamethyldisilazide or sodium hydride.

Step (d) Alkylation reaction, where R is other than hydrogen, which may be carried out by treating sequentially with a base and then with a compound RY, wherein Y is a leaving group such as a halogen and R is C,_₃ alkyl. This reaction may suitably be carried out using methyl iodide when it is desired to introduce a methyl group as the R substituent in Formula (I), as is depicted in scheme 1. Where R represents hydrogen, this step is omitted. The following steps (e) to (h) are then carried out as described.

It has been found that where the alkylation reaction is carried out with Mel, in tetrahydrofuran as solvent at -78°C, a predominance of the desired α-methyl stereoisomer over the β-methyl results, in a ratio of approximately 50:1.

Step (e)

The deprotection is carried out by conventional means, e.g. by addition of acid such as trifluoroacetic acid.

Step (f) Where R., is R₅CO, the R₁ group may be incorporated by acylation carried out using the acyl chloride R₅COCI and lithium hexamethyldisilazide at a reduced temperature, suitably -78 °C in a solvent, suitably tetrahydrofuran. Alternatively, it may be carried out using the preformed mixed anhydride generated by treatment of the acid R₅COOH with an appropriate acid chloride such as pivaloyl chloride in a solvent, suitably tetrahydrofuran.. The reaction is carried out under similar conditions to those using the acid chloride. Where R., is R₅NHCO, the reaction to incorporate R₁ may be carried out with the appropriate isocyanate, R₅NCO, in the presence of less than 1 mol equivalent of a suitable base such as sodium hydride, preferably 0.3mol equivalents, in a solvent suitably tetrahydrofuran at ambient temperature.

Step (g)

Deprotection, which can be carried out by conventional means, e.g. where P₂ is Cbz by hydrogenation in the presence of a palladium catalyst in a suitable solvent such as propan-2-ol. The product is preferably isolated as a salt, such as the hydrochloride salt.

Step (h) R₃CO is R₂, as set out in formula (I). The group R₃CO may be introduced in one step, by use of any of the procedures described for processes (i), (ii) and (iv) above.

Alternatively, when a racemic mixture of the compound of formula (XXVI) is used, the process may involve two steps. The first, which represents a further aspect of the invention, is the introduction of an L-prolyl or L-azetidyl group suitably N-protected with groups such Boc or Cbz, followed by separation of the resulting diastereomeric mixture. The second step involves deprotection and introduction of the group R (as discussed in process (iv) above), onto the pyrrolidine nitrogen of the desired diastereoisomer to give the required group R₃ in compound of formula (XXVII).

In a second example of this aspect of the invention, compounds of formula (I) wherein R., represents optionally substituted heteroaryl or fused heteroaryl may conveniently be prepared according to the methodology shown in the following general scheme 2, in which steps corresponding to those discussed above in relation to scheme 1 are correspondingly labelled:

Scheme 2

0

o

(c)

N

N

(XXII)

(d)

(XXVIII)

(j)

(k)

.CF, 3C-.O--„2H' (XXXIII)

(XXXIV) Step Q)

The acylation may be carried out according to the procedure described for process (i) above using the N-protected L-proline, its acid chloride, anhydride or mixed anhydride. When a racemic mixture of the compound of formula (XXVIII) is used, the resulting diastereomeric mixture is advantageously separated before proceeding to the next stage. Corresponding azetidyl compounds are produced by introduction of the N-protected L-azetidine carboxylic acid, acid chloride, acid anhydride, etc., in a similar manner.

Step (k)

This protecting group exchange is carried out according to the procedure in Step g but in this case in the presence of Boc₂O (P₃) to effect reprotection of the free proline or azetidine nitrogen. This step is necessary in order to overcome the difficulty which may otherwise arise from hydrogenolysis of a Cbz-protecting group in the case where R-i is a sulphur containing heteroaromatic group.

Step (I)

This coupling reaction may be carried out according to the procedure described for process (ii) starting with the appropriate haloheteroaromatic compound, R.,Y' and using for example the Goldberg variant of the Ullman Reaction.

Step (m)

This reaction may be carried out according to the procedure described for process (iii) above. For example, in the case of compounds of formula (VI), the reaction may be carried out using the appropriate para-substituted phenyl isocyanate in the presence of a base, suitably triethylamine, to quench the acid addition salt of the compound of formula (XXXIII) in a solvent suitably acetonitrile at temperature suitably between 0°C and ambient.

It will be appreciated that the compounds of formula (XXXV) below may also be used for the synthesis of compounds of formula I, specifically compounds of formula (XXXIV), by protection of the pyrrolidine nitrogen with an acid labile protecting group, such as 4-methoxybenzyloxycarbonyl (Moz), introduction of the group R-,, deprotection by conventional means and introduction of the group R₃CO as described above.

(XXXV)

The compound of formula (XXXV) may be obtained, for example, by deprotection of the compound of formula (XXVIII) using the procedure of Step (e) described above.

In a third example of this aspect of the invention, compounds of formula (I) wherein R represents methyl may conveniently be prepared according to the methodology shown in the following general scheme 3.

Scheme 3

L-methionine

C

(w)|

(XLIV)

OSi(alkyl)₃

CH, (y)

OC₂H₅

H CH,. * CO 2,C^,2H' '5

^CH ₃\^∞₂C₂H₆

CBZ NH,

P₂.V NHCOCF₃

(z)

(XLVII) (XLVI)

(2R, 3S)

(zz) alpha / beta

(XLVIII)

Step (n) This is a conventional protection reaction which, in the case when P₁ represents BOC, may be performed by reacting with (BOC)₂O in the presence of base (e.g. NaOH) in a polar solvent system such as dioxan/water.

Step (o)

This conversion may be performed on treatment with ammonium bicarbonate in the presence of a suitable solvent such as pyridine/DMF and in the presence of (BOC)₂O or suitable equivalent.

Step (v)

This is a conventional protection reaction which, in the case when P₂ represents CBZ, may be performed by reaction with nBuLi followed by CBZ-CI in the presence of an inert solvent such as THF below -50 °C.

Step (w)

This reaction may be performed in two stages, firstly by treatment with RX where RX is a compound (e.g. Mel, benzyl iodide or Me₂SO₄) capable of converting sulphur in the SMe moiety to sulphonium in a suitable solvent, e.g. propanone or acetonitrile. Generally R will represent alkyl or aralkyl and X will represent halide, especially iodide, or sulphate. Protection of the amide is convenient, although not essential, for this reaction. Secondly, the ring closure reaction may be performed by treatment with Dowex 2 x 8 400 mesh OH^' resin in a suitable solvent, e.g. MeCN. Alternatively, the ring closure may be performed by treatment with potassium carbonate or diisopropylamine in a suitable solvent, e.g. MeCN. Step (x)

Following deprotection, which may be performed in a conventional manner, for example, a BOC protecting group may be removed by treatment with HCI, e.g. in dioxan, the amine may be treated with a trifluoroacetic acid alkyl ester (e.g. the methyl ester) or trifluoroacetic anhydride in the presence of a suitable base e.g. N-methylmorpholine, then addition of a reducing agent e.g. lithium borohydride, followed by treatment with acetyl chloride in the presence of an alkyl alcohol e.g. methanol solvent, gives the ether (XLV).

Step (y)

The reaction of compounds of formula (XLV) and the methyl ketenesilylacetal takes place in the presence of a Lewis acid e.g. boron trifluoride diethyl etherate and an inert solvent e.g. dichloromethane or MeCN. The group "alkyl" in OSi(alkyl)₃ generally represents C.,-₆alkyl, . suitably methyl, isopropyl or t- butyldimethyl. Preferred OSi(alkyl)₃ is OSi(i-Pr)₃. These conditions give the desired α isomer in (α:β = 1 :1 ). The use of variants in which OC₂H₅ is replaced by OSi(alkyl)₃ is also envisaged.

Step (z)

This deprotection reaction will take place on treatment with base, such as potassium carbonate.

Step (zz)

This ring closure reaction may be performed on treatment with an alkyl Grignard reagent (e.g. t-butylmagnesium choride) in an inert solvent such as THF in the presence of tetramethylethylenediamine at a temperature of -20°C to 25°C. Compounds of formula (XLVIII) are a subset of compounds of formula (XXIV) in scheme 1 and can be carried forward through steps (f), (g) and (h) of that scheme to give compounds of formula (I).

As mentioned above, this route will give a mixture of the α and β anomers of (XLVI) in a ratio of 1 :1. As an alternative to the synthesis shown in Scheme 3, the starting material may be D, L-methionine and the racemic R,S compound of formula (XLIV) thus produced may be separated by dynamic resolution with a homochiral preparation of an appropriate chiral acid, e.g. (-) di-p-toluoyl-tartaric acid, to give the desired S isomer (90% yield).

o

TFA CH₂CI₂

^NHP1 „ NH, - NH₂

2 ~ N ► 2 - N

^/ H H

(-)Di-0,0'-p-tolulyl- .1/2 (-) DPTTA

(RS) XLIV (RS) L-Tartaric acid (S)

reprotect r

^ HP,

(S) XLIV

In a further aspect, the present invention provides the use of a compound of formula (XXII) in the synthesis of a compound of formula (I). Further, the present invention provides a method of making a compound of formula (I) comprising alkylating a compound of Formula (XXII) at the carbon atom adjacent to the lactam carbonyl group.

(XXII) In a further aspect, the present invention provides a compound of formula (L)

( )

wherein R is C^alkyl, P₃ is a protecting group such as CBZ and R₉ is C.,..₃ alkyl, and its use in the preparation of compounds of formula (I) in which R is C,.₃ alkyl. An example of such a compound is the compound of formula (XLVII) and an example of its use in the preparation of compounds of formula (I) can be seen in Scheme 3 above.

In a further aspect, the present invention provides a compound of formula (XXIII)

which compound may find use in the synthesis of compounds of Formula (I). The use of a compound of formula (XXIII) in the synthesis of a compound of formula (I) is also provided. Further, the present invention provides a method of making a compound of formula (I) comprising removal of the protecting group P₂ in the compound of formula (XXIII) followed by acylation at nitrogen of the pyrrolidine ring to introduce the group R₂. Further, the present invention provides a method of making a compound of formula (I) comprising removal of the protecting group, P₃ in the compound of formula (XXIII) followed by reaction at the nitrogen of the lactam ring to introduce the group R.,.

ABBREVIATIONS

Boc t-butyloxycarbonyl CBZ Benzyloxycarbonyl

DCM Dichloromethane

(Boc)₂O Di-tert-butyldicarbonate

Et₃N Triethylamine

THF Tetrahydrofuran

TFA Trifluoroacetic Acid

LiHMDS Lithium bis (trimethylsilyl)amide

DMF Dimethylformamide

TBTU O-(1 H-Benzotriazol-1-yl)-N,N,N',N'- tetramethyluronium tetrafluoroborate

HOBT 1 -Hydroxybenzotriazole

DIPEA N,N-Diisopropylethylamine

Moz 4-Methoxybenzyloxycarbonyl

TDA-1 Tris[2-(2-methoxyethoxy)ethyl]amine

IMS Industrial methylated spirit

Dansyl 5-dimethylamino-naphthalene-1-sulfonyl

The following non-limiting examples illustrate the present invention.

Examples

Compounds according to Formula (I) have been synthesised and tested in the enzyme and whole cell in vitro assays discussed above. The results of these tests are presented below. The compounds are listed in Table 1.

Table 1

cc-z φ

Table 1 (Cont.)

Biological Data

1. The compounds of Examples 1-17 were tested in the viral serine protease enzyme inhibition assay, whole cell hCMV in vitro plaque reduction assay and Vero cell cytotoxicity assay as described earlier. The data is shown in Table 2.

Table 2

E = Elisa assay In addition, the compound of Example 1 in Table 2 was active in the plaque reduction assay against HSV-1 and HSV-2 in Vero cells (IC₅₀ = 18 and 12.5μM, respectively), and the compound of Example 2 was active in this assay against HSV-2 with an IC₅₀= 3μM, and the compound of Example 19 was active in the HCMV plaque reduction assay with an IC₅₀ (μM) of >50.

The compounds of Examples 18-26 were also tested in the in vitro HCMV serine protease enzyme inhibition assay to find their Ki(nM). IC₅₀ (μM) against the HCMV virus (Elisa assay) were determined after 48h incubation and also these compounds and the compounds of examples 11-17 were tested in the plaque reduction assay against HSV-2, HSV-1 and VZV. These results and Vero cell cytotoxicity results are set out in Table 3.

Table 3

pi = Plaque Assay

Other compounds of formula (II) to (VI) have been synthesised and tested in the enzyme and in vitro assays and the results are shown below in Table 4.

Table 4

Four further compounds according to the invention have been synthesised and tested in the HCMV in vitro assay (plaque reduction) and HCMV protease assay as well as being checked for cytotoxicity in Vero cells. The results are set out below in Table 5:

Table 5

Synthetic Examples

A. Preparation of Intermediates

Intermediate 1

1 -{3-[(Benzyloxy-carbonyl)-amino]-1 -hydroxymethyl-propyl}-carbamic acid, tert- butyl ester

A solution of compound Nα-BOC,Nγ-CBZ-2,4-Diaminobutyric acid (3.198g) in tetrahydrofuran (44ml, dry) was cooled to -10°C under nitrogen, 4- methylmorpholine (1.0ml) was added followed by ethylchloroformate (0.868ml).

After stirring for 8 mins sodium borohydride (1.03g) was added in one portion followed by methanol (88ml) over a period of .11 mins at 0°C. The mixture was stirred at ca 0°C for an additional 11 mins before 1M hydrochloric acid (18ml) was added. The mixture was evaporated under reduced pressure and the aqueous residue was extracted with ethyl acetate. The organic layer was separated and washed with 1 M hydrochloric acid, water, saturated aqueous sodium bicarbonate solution and water, then dried (magnesium sulphate), evaporated under reduced pressure and some of the residue (1.8g from 2.87g) was purified by chromatography (Merck 7734) using cyclohexane:ethylacetate

(3:2) as eluent to give the title compound (1.6g) : t.l.c. (1 :1 cyclohexane : ethyl acetate) Rf 0.23 ir (CHBr₃) 3432, 1704cm-¹.

Intermediate 2 6-Benzyloxycarbonylamino-4-ferf-butoxycarbonylamino-hex-2E-enoic acid ethyl ester

A solution of dimethyl sulfoxide (6.82ml) in dry dichloromethane (135ml) was stirred under N2 and cooled (dry ice/acetone) to - 72°C. Oxalyl chloride (7.4ml) was added dropwise over 10 minutes (temp kept in the range - 60 - 65°C) and the reaction was stirred for 15 minutes. A solution of the alcohol, Intermediate 1 , (12.6g) in dry dichloromethane (135ml) was added over 20 minutes (temp kept in the range -60→-63°C) and the reaction mixture then stirred for 20 minutes by which time the temperature had risen to -52°C. Triethylamine (53.7ml) was added dropwise over 10 minutes followed by the immediate addition of the Wittig reagent (19.3g). The cooling bath was removed and the internal temperature allowed to rise to 17°C. The reaction mixture was poured into ether (400ml) and brine (400ml). The organic phase was separated and the aqueous phase extracted with ether (2x100ml). The combined organic phases were dried (MgSO4) and evaporated under reduced pressure to give a tan oil (36.22g). This was purified by flash column chromatography (Merck 9385 silica eluting with 40% ethyl acetate in cyclohexane) to give the title compound (15.71g) as an oil:

¹H nmr (CDCI₃): δ 7.40-7.30 (5H, m), 6.86 (1 H, dd), 5.93 (1 H, dd), 5.42-5.28 (1 H, br), 5.12 (2H, ABq), 4.72-4.60 (1 H, m), 4.50-4.32 (1 H, m), 4.19 (2H, q), 3.60-3.30 (1 H, m), 3.15-2.98 (1 H, m), 2.00-1.80 (1 H, m), 1.65-1.50 (1 H, m), 1.45 (9H, s) and 1.28 (3H, t), Rf 0.45 (2:3 ethyl acetate /cyclohexane)

Intermediate 3 rel-(2R,3S)-3-tert-Butoxycarbonylamino-2-ethoxycarbonylmethyl-pyrrolidine-1- carboxylic acid benzyl ester

Intermediate 2 (12.2g) was suspended in dry toluene (175ml) with stirring under N2- Tetramethylethylenediamine (1.1 ml) was added followed by lithium bis- (trimethylsilyl)amide (1.0M in hexanes, 7.6ml). On completion of the addition a solution had formed. The reaction mixture was stirred for 15 minutes and then poured into ethyl acetate (300ml) and saturated aqueous ammonium chloride (300ml). The organic phase was separated and the aqueous phase extracted with ethyl acetate (2x50ml). The combined organic extracts were washed with brine (150ml) and the aqueous phase extracted with ethyl acetate (2x25ml). The combined organic extracts were dried (MgSO4) and evaporated under reduced pressure to give a tan oil (12.86g) which was filtered through a plug of silica gel using ethyl acetate/cyclohexane (2/3) as eluant to give a crude mixture including title compound (10.74g) as an oil. This oil was purified further by flash column chromatography on silica gel. Elution with ethyl acetate/cyclohexane (2/3) gave the title compound, as a solid (8.49g, 69.7%). A small sample of the title compound was crystallised from ether to give a white solid: ¹H nmr (CDCI₃): δ 7.40-7.30 (5H, m), 5.12 (2H, s), 4.72-4.53 (1 H, m), 4.20-3.95 (4H, m), 3.65- 3.40 (2H, m), 2.95-2.65 (1H, m), 2.60-2.40 (1H, m), 2.25-2.10 (1 H, m), 1.92-1.75 (1 H, m), 1.40 (9H, s) and 1.30-1.15 (3H, m). Rf 0.8 (1 :1 , ethyl acetate/cyclohexane) Intermediate 4 trans-3-Amino-2-ethoxycarbonylmethyl-pyrrolidine-1 -carboxylic acid benzyl ester

To the Boc-protected Intermediate 3 (246.6gm, 1eq, 0.607mol) was added trifluoroacetic acid (25eq, 15.18mol, 1731gm, 1169ml) at room temperature. After stirring for one hour the solution was evaporated and the residue azeotroped twice with toluene (300ml). The resulting oil was dissolved in ethyl acetate (2500ml) and washed with 2M sodium hydroxide (1x800ml + 3x300ml), water and brine, dried (MgSO₄) and evaporated to give the title compound as a golden oil, 168.9gm, after high vacuum. Mass spec 613 [2M+H]\ 307 [MH]⁺.

Intermediate 5

(2R,3S)-3-amino-2-ethoxycarbonylmethyl-pyrrolidine-1 -carboxylic acid benzyl ester (2S,3S)-bis-(4-methyl-O-benzyloxy)-succinate salt.

A solution of Intermediate 4 (168.9gm, 0.55mol) was dissolved in ethanol (2500ml) and was added to a solution of (+) di-O-para-toluyl-D-tartaric acid (ex Fluka) (213gm, 0.55mol) in ethanol (2500ml) and the solution allowed to stand overnight. The resulting solid was collected by filtration and washed with ethanol and then recrystallised from boiling ethanol (~3500ml) to give the title compound as a white solid, 93.7gm. A further recrystallisation gave a white solid, mp 184-185°C. Chiral HPLC (Chiracel OJ column, eluent system ethanol:heptane; 3:7; flow rate = 1 ml/min, , λ = 215nm). Retention time = 7.7 min, 97.5 %ee.

Intermediate 6

(2R,3S)-3-amino-2-ethoxycarbonylmethyl-pyrrolidine-1 -carboxylic acid benzyl ester. The Intermediate 5 (131.8 gm, 190mmol) was suspended in an 1:1 mixture of water: ethyl acetate (1500ml), and solid potassium carbonate added (63gm, 457mmol). After fifteen minutes the phases were separated and the aqueous phase extracted with ethyl acetate (3x200ml). The organic portions were combined and washed with water and brine, dried (MgSO₄) and evaporated to give the title compound as a colourless oil, 57.8gm. Mass spec 613 (100%) [2M+H]\ 307(83%) [MH]⁺. [α]_D = -11.3° (c=1.33, MeOH) Intermediate 7

(3aS,6aR)-5-Oxo-hexahydro-pyrrolo[3,2-b]pyrrole-1 -carboxylic acid benzyl ester

To the Intermediate 6 (52.9gm, 173mmol) in tetrahydrofuran (550ml) in an ice- salt bath was added a solution of tert butyl magnesium chloride (554ml of a 1 M solution in tetrahydrofuran, 554mmol), keeping the temperature <1°C. The mixture was warmed to room temperature over 1hr 15min, then quenched with saturated ammonium chloride whilst cooling in an ice bath. The phases were separated and the aqueous phase extracted with ethyl acetate. The combined organics were washed with water and brine, dried (MgSO₄) and evaporated and to give the title compound as a cream solid, 43.5gm. A portion of the solid was purified by flash column chromatography over silica gel ( Merck 9385) using ethyl acetate as the eluting solvent, to give the title compound as a white solid, mp 157-159°C.

[α]_D = -68.4° (c=1.28, MeOH)

Intermediate 8

(3aR,6aS)-2-Qxo-hexahydro-pyrrolo[3,2-b]pyrrole-1 ,4-dicarboxylic acid 4-benzyl ester 1 -tert-butyl ester

To the Intermediate 7 (43.2gm, 166mmol) in tetrahydrofuran (1200ml) at -72°C under a nitrogen blanket was added lithium bis (trimethylsilyl)amide (216ml of a 1 M solution in tetrahydrofuran, 216mmol) dropwise, keeping the temperature <- 71 °C. After ten minutes a solution of di-tert-butyldicarbonate (54.3 gm, 249mmol) in tetrahydrofuran (350ml) was added, keeping the temperature <- 71 °C. The reaction was stirred at -73°C for two and a half hours and then quenched with saturated ammonium chloride. Then the mixture was allowed to warm to room temperature, water was added and the phases separated. The aqueous phase was extracted with ethyl acetate and the combined organics were washed with water and brine, dried (MgSO₄) and evaporated to give the title compound as an orange-red semi solid. Pure material could be obtained as a pale cream solid by trituration under diethyl ether. Yield 42.5gm. A portion of the solid was recrystallised from boiling diethyl ether to give the title compound as a white solid, mp 101 -103°C. [α]_D = -45.6° (c=1.13, MeOH). Intermediate 9

(3S,3aR,6aS)-3-Methyl-2-oxo-hexahydro-pyrrolo[3,2-b]pyrrole-1 ,4-dicarboxylic acid 4-benzyl ester 1 -tert-butyl ester

The Intermediate 8 (606mg, 1eq, 1.68mmol) was dissolved in tetrahydrofuran (6ml) and cooled, under nitrogen, to -75°C. Lithium hexamethyldisilazide (1.3 eq, 2.2ml of a 1 M solution in tetrahydrofuran) was added, keeping the temperature below -70°C. After 10 minutes methyl iodide was added (17eq, 28.9mmol, 1.8ml). After stirring for a further 45 minutes the reaction was quenched with saturated aqueous ammonium chloride and then allowed to warm to room temperature. Water was added and then the aqueous phase was extracted with ethyl acetate, and the combined organic phase was washed with water and brine, dried (MgSO₄) and evaporated to give a golden oil. Purification by flash column chromatography over silica gel ( Merck 9385) using cyclohexane:ethyl acetate (3:1 )as the eluting solvent system, afforded the title compound as a white foam, 526mg. Mass spec 275 [M-100+H]⁺ [α]_D = -88.6° (c=1.1 , MeOH).

Intermediate 10

(3aS,6S,6aR)-6-Methyl-5-oxo-hexahydro-pyrrolo[3,2-b]pyrrole-1 -carboxylic acid benzyl ester

To the Intermediate 9, (486mg, 1eq, 1.3mmol) was added trifluoroacetic acid (60eq, 6ml) and the mixture stirred at room temperature for 40 minutes, then evaporated to give a brown oil. This was dissolved in ethyl acetate (6ml) and washed with saturated sodium bicarbonate solution (2x3ml), water (3ml) and brine (3ml), dried (MgSO₄) and evaporated to give the title compound as a pale beige solid 340mg. The solid was recrystallised from boiling diethyl ether to give a white solid (202mg), mp 112-113°C.

Chiral HPLC (Chiral Pak464 column, eluent system propan-2-ol:heptane; 2:25; flow rate = 1 ml/min). Retention time = 20.91 min, 96.00 %ee.

Intermediate 10 (Alternative synthesis) (3aS,6S,6aR)-6-Methyl-5-oxo-hexahydro-pyrrolo[3,2-b]pyrrole-1 -carboxylic acid benzyl ester and (3aS,6R,6aR)-6-Methyl-5-oxp-hexahydro-pyrrolo[3,2-b]pyrrole- 1 -carboxylic acid benzyl ester t-Butylmagnesium chloride (1 M in tetrahydrofuran) 2.2ml, 3eq) was added over 15 min to a stirred solution of the Intermediate 59 (230mg, 0.72mmol) in dry tetrahydrofuran (1.5ml) under nitrogen at -15°C. After stirring under nitrogen at - 15°C for 1 h and -10°C for 1 h tic indicated consumption of starting material. Dilute hydrochloric acid (2M, 2ml) was added over 5min with the temperature rising to 0°C. The mixture was then diluted with ethyl acetate (4ml) and more dilute hydrochloric acid (2M, 2ml) and the layers were separated. The aqueous layer was re-extracted with ethyl acetate (2x4ml). All the ethyl acetate solutions were combined, washed with saturated brine (4ml) and dried (MgSO₄). Solvent removal in vacuo gave a yellow oil which crystallised on standing (200mg). This was dissolved in dichloromethane (2ml) and silica gel (0.4g) was added. Solvent removal in vacuo afforded a yellow solid which was applied to a silica gel column (6g) which was eluted with an ethyl acetate:cyclohexane (3:1 ) mixture. Concentration in vacuo of the fractions with an RF 0.18 gave a white solid (110mg 50%) whose nmr was consistant for a mixture of title compounds (3aS,6S,6aR) and (3aS,6R,6aR).

HPLC: Retention time = 7.57min, 62.1 % , 9.44min, 37.9%, using Whelk- 01 , 10% Isopropanol 89.5% heptane 0.5% Acetic Acid, flow: 1ml min ^"1uv detection:

The above solid was purified by preparative HPLC on a chiral pak AD column with heptane:ethanol 70:30 as eluent, flow: 10ml min -1 , uv detection to give

The title compound (3aS,6S,6aR) a white solid (30mg, 15.8%) ¹H nmr (CDCI₃): δ 7.41-7.28 (m,5H); 6.40 (bd, 1 H);5.19-5.06 (m,2H); 3.88

(m,1 H); 3.72-3.60 (m, 2H); 3.59-3.42 (m,2H); 3.05-2.70 (bdm, 1 H); 2.21 (m,1 H);

1.95-1.78 (m,1 H); 1.10 (bd, 3H)

Mass spec, thermospray: +ve m/e; 275 = [MH]⁺ ,549 = [2M.H]⁺

The (3aS,6R,6aR) isomer, a white solid (40mg, 21 %)

¹H nmr (CDCI₃): δ (7.28 - 7.40 (m, 5H); 6.69 (bs, 1 H); 5.12 (s, 2H); 3.883.67

(m,2H); 3.32- 3.20 (m,1 H); 3.05-2.95 (t, 1 H); 2.52 (bd, 1 H); 2.18 (m, 1 H); 1.96-

1.66 (m,1 H); 1.33 (bd, 3H)

Mass spec, thermospray: +ve m/e; 275 = [MH]⁺, 549 = [2M.H]⁺ Intermediate 11

(3aS,6S,6aR)-4-Cyclopropanecarbonyl-6-methyl-5-oxo-hexahydro-pyrrolo[3,2- b]pyrrole-1 -carboxylic acid benzyl ester

To a solution of the Intermediate 10, (992mg, 1eq, 2.75mmol) in dry THF (7ml) at -78°C under nitrogen was added lithium hexamethyldisilazide (3.3ml of a 1M solution in tetrahydrofuran, 1.2eq, 3.3mmol), keeping the temperature below - 70°C. The solution was kept at -78°C. for 10 mins, then at 0°C for 10 mins, recooled to -78°C and cyclopropanecarbonyl chloride (0.75ml, 3eq, 8.25mmol) was added and the reaction mixture was the stirred at -78°C. for 55 minutes. The reaction mixture was quenched with saturated aqueous ammonium chloride (25ml) and then allowed to warm to room temperature. Water was added (20ml) and then the aqueous phase was extracted with ethyl acetate (100ml), and the combined organic phase was washed with water (30ml) and brine(30ml), dried (MgSO₄) and evaporated to give a yellow oil. Purification by flash column chromatography over silica gel ( Merck 9385) using cyclohexane:ethyl acetate (4:1 )as the eluting solvent system afforded the title compound as a pale yellow gum, 334mg. Mass Spec: 343 [MH]⁺

Intermediate 12

(3aS,6S,6aR)-1-Cyclopropanecarbonyl-3-methyl-hexahydro-pyrrolo[3,2-b]pyrrol-

2-one hydrochloride

A solution of Intermediate 11 , (330mg, 0.96mmol) in isopropanol (30ml) was added to the catalyst (119mg, 10% palladium on activated carbon with 50% water, Degussa type E101 NE/W) under nitrogen and the resulting mixture stirred vigorously under an atmosphere of hydrogen for 2.75hours. The catalyst was filtered off under an atmosphere of nitrogen and a 1 M solution of hydrogen chloride in diethyl ether (1 ml, 1eq, 1 mmol) was added to the filtrate. Evaporation of the solvent gave the title compound as a colourless gum, 175mg. Mass Spec: 209 [MH]⁺

Intermediate 13

2S-(4-Cyclopropanecarbonyl-6S-methyl-5-oxo-hexahydro-(3aS,6aR)-pyrrolo[3,2- b]pyrrole-1-carbonyl)-pyrrolidine-1 -carboxylic acid benzyl ester and 2S-(4- Cyclopropanecarbonyl-6R-methyl-5-oxo-hexahydro-(3aR,6aS)-pyrrolo[3,2- b]pyrrole-1-carbonyl)-pyrrolidine-1 -carboxylic acid benzyl ester To a solution of carbobenzyloxy-L-proline (0.171g,0.69mmoi,1.1eq.) in DMF (3mL), stirred at room temperature, was added a solution of TBTU (0.221 g, 0.69mmol, 1.1 eq.) in DMF:MeCN (0.5mL:0.5mL) and a solution of HOBT (0.094g, 0.70mmol,1.1 eq.) in DMF: MeCN (0.5mL:0.5mL). After 15 min. the Intermediate 12 (racemic) (0.152g,0.62mmpl,1 eq.) in DMF (0.5mL) and diisopropylethylamine (0.215mL,1.23 mmol,2 eq.) were added. The reaction mixture was allowed to stir at room temperature for 16 hours and was then diluted with dichloromethane (15mL) and water (15mL). The aqueous layer was re-extracted with dichloromethane (15mL) and the combined organic extracts were then washed with 2N HCI (15mL) , water (15mL) and sat. NaHCO₃ solution (15mL). After drying over MgSO₄ the solvent was evaporated in vacuo to give an orange oil. The product was purified by flash column chromatography over silica gel (Merck 9385) using ethyl acetate: cyclohexane to give as the less polar(6S,3aS,6aR diastereoismer) the title compound, as a white solid (89mg). Mass Spec: 440 [MH]⁺

Circular dichroism λ_max199.8 nm, dE 4.82; λ_max216.6 nm, dE -15.8; λ_max238.6 nm, dE 17.8; HPLC (Intersil M column ODS2, eluent system A (H₂O, 0.1% H₃PO₄) B (95% MeCN/H₂O, 0.1% H₃PO₄):Gradient 0%B 2mins, 0%-100% B 40mins,100% B 10mins; flow rate = 1ml/min, λ = 215nm). Retention time =24.9 min., 100%.

The more polar 6R,3aR,6aS diastereoismer, was obtained as a white foam (61 mg,22%). Mass Spec: 440 [MH]⁺

Circular dichroism λ_max214.6 nm, dE 26.8; λ_max238.6 nm, dE -17.0;

HPLC (Same system as less polar isomer) : Retention time = 24.1 min. ,92%,

24.9 min., 4.5%.

Intermediate 14

(3S,3aR,6aS)-1-Cyclopropanecarbonyl-3-methyl-4-(pyrrolidine-2S-carbonyl)- hexahydro-pyrrolo[3,2-b]pyrrol-2-one hydrochloride

The Intermediate 13 (0.893g,2.03mmol) was added to the catalyst (0.419g, 10% palladium on activated carbon with 50% water, Degussa type E101 NE/W) as a solution in isopropanol (160mL). Warming was required for all material to dissolve. A 1.0M solution of HCI in ether (2.3mL,2.3mmol) was also added. The reaction mixture was stirred vigorously under an atmosphere of hydrogen for 5 hours with further catalyst (0.4g) added after 1.5 hours to complete the reaction. The catalyst was filtered off and a 1.OM solution of HCI in ether (0.2 mL) was added to the filtrate. Evaporation of the filtrate in vacuo afforded the title compound, as a white foam (0.721 g). It was used in the preparation of Examples 1-10 without further purification. Mass Spec: 306 [MH]⁺

Intermediate 15 (3aS,6S,6aR)-4-(cis-2,3-Dimethyl-cyclopropanecarbonyl)-6-methyl-5-oxo- hexahydro-pyrrolo[3,2-b1pyrrole-1 -carboxylic acid benzyl ester

To a solution of c/s,c/s-bis(2,3-dimethylcyclopropanecarboxylic) acid ( 678mg, 5.95 mmol ) in anhydrous tetrahydrofuran ( 20 mL ) stirred under nitrogen at - 13° was added methanesulphonyl chloride ( 250μL, 3.23 mmol ) followed by a solution of triethylamine ( 1.5mL, 10.8mmol ) in anhydrous tetrahydrofuran ( 10mL ) dropwise over 15 min at -12 to -14°. The resulting suspension was stirred below -11° for 1h and then allowed to warm up to 20° over 2.5h. Solid was filtered off and bed-washed with ether and the combined filtrates were evaporated. The residue was partitioned between ether ( 2x 50mL ) and ice-cold saturated aqueous sodium bicarbonate solution ( 25mL ) . The combined organic phases were washed with water ( 20mL ) and saturated brine ( 20mL ), dried over magnesium sulphate and evaporated to give c/s,c/s-bis(2,3-dimethyl- cyclopropanecarboxylic) anhydride ( 543mg ) as an oil which crystallised. [MNH₄] ⁺ 228.

To a solution of the Intermediate 10 ( 600mg, 2.19 mmol ) in anhydrous tetrahydrofuran ( 8 mL ) stirred under nitrogen in an ice and IMS bath was added 1 M lithium hexamethyldisilazide ( 2.4mL; 2.4 mmol ) slowly. The yellow solution was stirred for a further 20 min and then a solution of c/s,c/s-bis(2,3- dimethyl-cyclopropanecarboxylic) anhydride ( 510mg, 2.42mmol ) in anhydrous tetrahydrofuran ( 7 mL ) was added slowly. The resulting solution was stirred for 1h under nitrogen in an ice and IMS bath and then poured into saturated aqueous ammonium chloride solution ( 50 mL ). The solution was extracted with ethyl acetate ( 2x 50mL ) and the combined organic phases were washed sequentially with saturated aqueous sodium bicarbonate solution ( 20mL ), water ( 20mL ) and saturated brine ( 20mL ), dried over magnesium sulphate and evaporated to give an oil which crystallised ( 1.09g ) . The crude product was purified on a 10g Bond Elute Silica cartridge eluting with cyclohexane / dichloromethane to give the the title compound, ( 785mg). LCMS: 371 [MH]⁺, Retention time = 3.45 min, using the same conditions as in Intermediate 27

Intermediate 16

(3S,3aR,6aS)-1-(cis-2,3-Dimethyl-cyclopropanecarbonyl)-3-methyl-hexahydro- pyrrolo[3,2-b]pyrrol-2-one hydrochloride

A solution of the Intermediate 15 ( 890mg, 2.4 mmol ) in isopropanol ( 70mL ) containing 1 M ethereal HCI ( 2.5mL , 2.5 mmol ) was hydrogenated in the presence of the catalyst (0.2g, 10% palladium on activated carbon with 50% water, Degussa type E101 NE/W) for 1.5h. The catalyst was filtered off , 1 M ethereal H Cl ( 0.2mL; 0.2mmol) was added and the solvent was evaporated off. Toluene ( 30mL ) was added and evaporated, and the resulting solid was triturated under ether ( 40mL ) . Ether ( 50mL ) was added and evaporated off to give the title compound (670mg) as a white solid.

LCMS: 237 [MH]⁺, Retention time =2.03 min, using the same conditions as in Intermediate 27

Intermediate 17

2S-[4-( cis-2,3-Dimethyl-cyclopropanecarbonyl)-6S-methyl 5-oxo-hexahydro- (3aS,6aR)-pyrrolo[3,2-b]pyrrole-1 -carbonyl]-pyrrolidine-1 -carboxylic acid benzyl ester and 2S-[4-( cis-2,3-Dimethyl-cyclopropanecarbonyl)-6R-methyl-5-oxo-hexahydro- (3aR,6aS)-pyrrolo[3,2-b]pyrrole-1 -carbonyl]-pyrrolidine-1 -carboxylic acid benzyl ester.

To a stirred mixture of carbobenzyloxy-L-proline (613mg, 2.46mmol, 1.5eq) in acetonitrile (6mL) and dimethylformamide (O.δmL) were added TBTU (790mg, 2.46mmol, 1.5eq) and HOBT (333mg, 2.46mmol 1.5eq) at room temperature. After 16 minutes a solution of the racemic form of Intermediate 16 (452mg, 1eq) and di-isopropylethylamine (0.714mL, 4.1 mmol, 2.5eq) in dimethylformamide (1 mL) and acetonitrile (2mL) was added. The mixture was stirred for 20 minutes and then left to stand overnight (16hr) before it was poured into ethyl acetate (60mL) and washed with water (2 x 30mL) and brine (30mL), dried (MgSO₄) and evaporated to leave a yellow gum. Purification by flash column chromatography over silica gel ( Merck 9385) using cyclohexane:ethyl acetate (1 :1) as the eluting solvent system gave 288mg of title compound 3aS,6aR, 6S as a white foam. Mass spec 468 [MH]⁺.

Circular dichroism λ_max199.8 nm, dE 3.89; λ_max221.6 nm, dE -17.90; λ_max242.6 nm, dE 16.00; and 253mg of the 3aR,6aS.6R isomer as a white foam.

Mass spec 468 [MH]⁺.

Circular dichroism λ_max218.2 nm, dE 26.50; λ_max243.0 nm, dE -15.10;

Intermediate 18

(3S,3aR,6aS)-1-(cis-2,3-Dimethyl-cyclopropanecarbonyl)-3-methyl-4- (pyrrolidine-2S-carbonyl)-hexahydro-pyrrolo[3,2-b]pyrrol-2-one hydrochloride

The Intermediate 17, (134mg, 0.29mmol) was dissolved in isopropanol (8mL) and added to the catalyst (20mg, 10% palladium on activated carbon with 50% water, Degussa type E101 NE/W) under nitrogen. Then the mixture was stirred vigorously under an atmosphere of hydrogen. After 1 hour 40min more catalyst (5mg) was added. The mixture was stirred in a hydrogen atmosphere for a further 1 hour before the catalyst was removed by filtration through glass fibre filters and washed with isopropanol. The filtrate and washings were combined and treated with 1 M hydrogen chloride in ether (0.3mL, 0.3mmol, 1.05eq). The mixture was evaporated to leave title compound as a white solid, 102mg. Mass spec 334 [MH]⁺.

Intermediate 19

(3S,3aR,6aS)-3-Methyl-2-oxo-hexahydro-pyrrolo[3,2-b]pyrrole-1 -carboxylic acid tert-butyl ester hydrochloride

Prepared in a similar manner to Intermediate 14 from Intermediate 9. The title compound , was obtained as a white solid in 93% yield and was used in the next reaction without further purification. Mass Spec. : 141 [M-Boc]⁺, 241 [MH]⁺, 481 [2MH]⁺ Intermediate 20 (3S,3aR,6aS)-3-Methyl-hexahydro-pyrrolo[3,2-b]pyrrol-2-one hydrochloride

To the Intermediate 19, (8.61g, 31.1 mmol) in dry dichloromethane (20mL) was added trifluoroacetic acid (20mL, 260mmol, 8 eq.). The solution was stirred at room temperature for 2 hours. The solvents were then evaporated and the resulting brown gum was azeotroped with toluene (4X 50 mL). The title compound , was obtained in quantitative yield as a brown gum which was used in the next reaction without further purification. Mass Spec: 141 [MH]⁺ parent amine.

¹H nmr (d⁶-DMSO) : δ 9.18-8.80 (2H,brd.), 7.30-7.08 (1 H,m), 3.92-3.20 (4H,m), 2.60 (1H,m), 2.26-2.10 (1 H,m), 1.75 (1 H,m), 1.09 (3H,d) ppm.

Intermediate 21 (3aS,6S,6aR)-6-Methyl-5-oxo-hexahydro-pyrrolo[3,2-b]pyrrole-1 -carboxylic acid 4-methoxy-benzyl ester

Intermediate 20 (12.01g, 31.1 mmol of trans-lactam + 57 mmol TFA) and triethylamine (11.2 mL, 80.43 mmol, 2.6 eq. w.r.t. trans-lactam) were dissolved in water (40mL). A solution of 2-(4-methoxybenzyloxycarbonyloxyimino)-2- phenylacetonitrile (MOZ-ON) (7.45g,31 mmol, 0.8 eq.) in 1 ,4-dioxane (60mL) was added to the stirred mixture. After 18 hours it was diluted with water (140mL) and extracted with ethyl acetate (2 X 100mL). The aqueous phase was acidified to pH2 with solid citric acid and then saturated with solid sodium chloride. It was then further extracted with ethyl acetate (3 X 60mL). The combined organic extracts were washed with water (50mL) and sat. brine (50mL), dried over MgSO₄ and the solvent was evaporated to give a brown oil. Trituration under ether caused precipitation of a white solid which was collected by filtration washed with ether and dried in vacuo at room temperature to give the title compound (5.37g).

LCMS: 305.2 [MH]⁺, Retention time = 3.76 min, using the same conditions as in Intermediate 27

Intermediate 22 (3aS,6S,6aR)-4-Benzothiazol-2-yl-6-methyl-5-oxo-hexahydro-pyrrolo[3,2- b]pyrrole-1 -carboxylic acid 4-methoxy-benzyl ester To the Intermediate 21 , (1.04g,3.42mmol,1eq.) was added 2- bromobenzothiazole² (1.17g,5.47mmol,1.6eq.), potassium carbonate (0.768g,5.56 mmol, 1.6 eq.), copper (I) chloride (0.345g,3.49mmol,1eq.), TDA-1 (0.330mL,1.03 mmol,0.3eq.) and xylene (55mL). The reaction mixture was refluxed under a Dean-Stark head for 7.5 hours. After leaving to cool, the brown solids were filtered off and washed with ethyl acetate (30mL). The combined filtrate and washings were washed with water (100mL) and brine (100mL) and dried over MgSO₄. The solvent was evaporated to give a brown gum (1.99g). The title compound, was purified by flash column chromatography over silica gel (Merck 9385) using cyclohexane:ether and was obtained as a white solid (0.95g).

Mass Spec. : 438 [MH]⁺

Ref 2. M.P. Doyle, J.F. Dellaria, B.J.Siegfried, J. Org. Chem., 42, 1977, 2426- 2430.

Intermediate 23

(3S,3aR,6aS)-1-Benzothiazol-2-yl-3-methyl-hexahydro-pyrrolor3,2-b]pyrrol-2- one trifluoroacetate

To the Intermediate 22, (850mg, 2.194mmol) was added trifluoroacetic acid (20mL) at room temperature. After 20 minutes the solvent was evaporated in vacuo to give, after trituration under diethyl ether, the title compound as a tan solid, (643 mg). Mass spec 274 [MH]⁺.

Intermediate 24

2S-(4-Benzothiazol-2-yl-6S-methyl-5-oxo-hexahydro-(3aS,6aR)-pyrrolor3,2- b]pyrrole-1-carbonyl)-pyrrolidine-1 -carboxylic acid tert-butyl ester and 2S-(4- Benzothiazol-2-yl-6R-methyl-5-oxo-hexahydro-(3aR,6aS)-pyrrolo[3,2-b]pyrrole- 1-carbonyl)-pyrrolidine-1 -carboxylic acid tert-butyl ester.

To a stirred mixture of N-tert Boc-L-proline (165mg, 0.77mmol, 2eq) in acetonitrile (3mL) and dimethylformamide (0.6mL) were added TBTU (246mg, 0.77mmol, 2eq) and HOBT (104mg, 0.77mmol 2eq) at room temperature. After 40 minutes a solution of the racemic form of Intermediate 23 (230mg, 1eq) and di-isopropylethylamine (0.267mL, 1.53mmol, 4eq) in dimethylformamide (O.δmL) was added. The mixture was stirred for 2 hours and then left to stand overnight (14.75hr) before it was poured into ethyl acetate (30mL) and washed with water (2 x 15mL) and brine (15mL), dried (MgSO₄) and evaporated to leave a pale yellow gum. Purification by flash column chromatography over silica gel( Merck 9385) using cyclohexane:ethyl acetate (2:1 )as the eluting solvent system gave 52mg of the title compound (3aS,6aR,6S diastereoismer) as a white solid. Mass spec 471 [MH]⁺, 371 [M-Bocf.

Circular dichroism λ_max197.4 nm, dE 6.72; λ_max208.0 nm, dE -2.39; λ_max215.0 nm, dE 3.67; λ_max230.6 nm, dE -5.72; λ_max2.48.2 nm, dE 3.44; HPLC (Intersil M column ODS2, eluent system A (H₂O, 0.1 % H₃PO₄) B (95% MeCN/H₂O, 0.1% H₃PO₄):Gradient 0%B 2mins, 0%-100% B 40mins,100% B 10mins; flow rate = 1 ml/min, , λ = 215nm). Retention time =31.44min 100%. contains none of the other isomer, checked by spiking the sample with the 3aR,6aS,6R diastereoismer). and 49mg of the (3aR,6aS,6R diastereoismer) as a pale yellow solid. Mass spec 471 [MH]⁺, 371 [M-Boc]⁺. Circular dichroism λ_max205.0 nm, dE 18.90; λ_max216.4nm, dE 3.24; λ_max228.4 nm, dE 7.48; λ_max247.8 nm, dE -3.40;

HPLC (Same system as less polar isomer) : Retention time = 31.03min 98% (the peak has a small shoulder at 31.4min, which when spiked with the 3aS,6aR,6S diastereoismer becomes more marked, so probably contains a small amount of the other isomer).

Intermediate 24(Alternative synthesis)

2S-(4-Benzothiazol-2-yl-6S-methyl-5-oxo-hexahydro-(3aS,6aR)-pyrrolo[3,2- b]pyrrole-1-carbonyl)-pyrrolidine-1 -carboxylic acid tert-butyl ester

A mixture of Intermediate 27 (40.4mg, 0.12mmol, 1eq), copper (1 ) chloride (12mg, 0.12mmol, 1eq), potassium carbonate (27.5mg, 0.2mmol, 1.66eq), TDA- 1 (11.1μL, 0.03mmol, 0.29eq) and 2-bromobenzothiazole (45mg, 0.21 mmol, 1.75eq) in p-xylene (25mL) was stirred and refluxed under nitrogen. The oil bath temperature was ca. 170°C. After 4 hr 30 min the mixture was left to stand at room temperature overnight before it was reheated and stirred at reflux for a further 5 hr 15 min, and then allowed to cool to room temperature overnight. Then the mixture was filtered and the residue washed with ethyl acetate (10mL). The filtrate and washings were combined and washed with 1 M hydrochloric acid (10mL), water (7.5mL) and brine (7.5mL), dried (MgSO₄) and evaporated to leave a yellow solid. The crude material was purified by preparative plate chromatography using (WhatmanPK6F silica gel 60A plate) eluting with ethyl acetate/cyclohexane(1 :1 ) to give the title compound (3aS,6aR,6S diastereoismer) as a white solid, 26.2mg. Mass spec 471 [MH]⁺, 371 [M-Boc]⁺.

Circular dichroism spectra superimposed with that obtained in method above.

Intermediate 25

(3S,3aR,6aS)-4-(1-Benzyloxycarbonyl-pyrrolidine-2S-carbonyl)-3-methyl-2-oxo- hexahydro-pyrrolo[3,2-b]pyrrole-1 -carboxylic acid tert-butyl ester

To a stirred suspension of carbobenzoxy - L - proline ( 8g, 32.1 mmol ), TBTU ( 10.3g, 32.1 mmol ) and 1-hydroxybenzotriazole ( 4.5g, 32.1 mmol ) in anhydrous acetonitrile (65mL) was added diisopropylethylamine ( 10.6 mL, 61 mmol ) and the resulting solution was stirred for 20 min. The Intermediate 19 hydrochloride salt ( 7.45g, 27 mmol ) was added and acetonitrile ( 35 mL ) and anhydrous DMF (30mL ) were added to give a clear solution which was stirred at 20° for 24h. The solution was poured into ethyl acetate ( 500mL ) and the solution was washed with water ( 500mL ). The aqueous phase was back-extracted with ethyl acetate ( 300mL ) and the combined organic phases were washed sequentially with 2M HCI ( 3x 200 mL ), water ( 2x 200mL ) and saturated brine ( 200mL), dried over magnesium sulphate and evaporated to give a gum (18.5g ). Purification by flash chromatography over silica gel (Merck 9385) eluting with a 2:1 and then 3:1 mixture of ethyl acetate and cyclohexane gave the title compound ( 11.26g) as a foam, R_f 0.31 in EtOAc/ cyclohexane 4:1. LCMS: 472 [MH]⁺, Retention time = 3.01 min, using the same conditions as in Intermediate 27

Intermediate 26

2S-(6S-Methyl-5-oxo-hexahydro-(3aS,6aR)-pyrrolo[3,2-b]pyrrole-1-carbonyl)- pyrrolidine-1 -carboxylic acid benzyl ester

To a solution of the Intermediate 25 ( 11.2g, 23.8 mmol ) in dichloromethane ( 60 mL ) was added trifluoroacetic acid ( 20 mL ). The solution was stirred in a cold water bath for 45 min and was then concentrated in vacuo . Toluene ( 100 mL ) was added and evaporated off and the resulting gum was dissolved in ethyl acetate ( 250mL ) . The solution was washed sequentially with saturated aqueous sodium bicarbonate solution ( 2x 100mL ), water (2x 100mL ) and saturated brine ( 100mL ), dried over magnesium sulphate and evaporated to a foam. The aqueous washes were combined and extracted with ethyl acetate ( 3x 300mL ) and the organic extracts were combined, washed with saturated brine ( 50mL ), dried over magnesium sulphate and evaporated to give a crisp foam. The batches were combined to give the title compound ( 8.68g). R_f 0.12 in ethyl acetate / toluene / acetic acid 25:5:1

LCMS: 372 [MH]⁺, Retention time = 2.45min, using the same conditions as in Intermediate 30

Intermediate 27 2S-(6S-Methyl-5-oxo-hexahydro-(3aS,6aR)-pyrrolo[3,2-b]pyrrole-1-carbonyl)- pyrrolidine-1 -carboxylic acid tert-butyl ester

A solution of Intermediate 26 ( 8.0g, 21.5 mmol ) and di-tert-butyldicarbonate ( 6.0g, 27.5g ) in isopropanol ( 350mL ) was hydrogenated in the presence of the catalyst ( 2g, 10% palladium on activated carbon with 50% water, Degussa type E101 NE/W) for 5h . The catalyst was filtered off and the filtrate was evaporated to a white foam. Crystallisation from toluene ( 35 mL ) and cyclohexane ( 40 mL ) gave the title compound ( 5.6g) . A second crop ( 0.97g) was obtained by evaporating the mother liquor and crystallising the residue from toluene ( 10mL ) and cyclohexane ( 10mL ). R_f 0.12 in ethyl acetate / toluene / acetic acid 25:5:1 LCMS: 338 [MH]⁺, Retention time = 2.25min, using the same conditions as in Intermediate 30.

Intermediate 28

(3S,3aR,6aS)- 4-[1-(5-Dimethylamino-naphthalene-1-sulfonyl)-pyrrolidine-2S- carbonyl]-3-methyl-2-oxo-hexahydro-pyrrolo[3,2-b]pyrrole-1 -carboxylic acid tert- butyl ester and (3R,3aS,6aR)-4-[1-(5-Dimethylamino-naphthalene-1-sulfonyl)- pyrrolidine-2S-carbonyl]-3-methyl-2-oxo-hexahydro-pyrrolo[3,2-b]pyrrole-1- carboxylic acid tert-butyl ester.

The above Intermediate was prepared in a similar manner to Intermediate 13 by reacting the racemic form of Intermediate 19 with dansyl -L- proline. The less polar (3S,3aR,6aS) diastereomer, was obtained as a yellow foam.

Mass Spec. : 571 [MH]⁺, 417 [M-Boc]⁺ and the more polar (3R,3aS,6aR)diastereomer, was also obtained as a yellow foam. Mass Spec. : 117, 571 [MH]⁺, 417 [M-Bocf Intermediate 29

(3S,3aR,6aS)- 4-[1 -(5-Dimethylamino-naphthalene-1 -sulfonyl)-pyrrolidine-2S- carbonyl]-3-methyl-hexahydro-pyrrolo[3,2-b]pyrrol-2-one The above Intermediate was prepared in a similar manner to Intermediate 10 from intermediate 28

The product was obtained as a yellow solid. Mass Spec. : 471 [MH]⁺

Intermediate 30

2S-(4-Benzothiazol-2-yl-6S-methyl-5-oxo-hexahydro-(3aS,6aR)-pyrrolo[3,2- b]pyrrole-1-carbonyl)-azetidine-1 -carboxylic acid tert-butyl ester

The above Intermediate was prepared in a similar manner to Intermediate 13 by reacting the Intermediate 23 with N-tert-butyloxycarbonyI-L-azetidine-2- carboxylic acid ¹.

LCMS: 457 [MH]⁺, Retention time =3.21 min, using (Gilson Supelcosil LC ABZplus column, eluent system A (H₂O, 0.1% Formic acid, 10mmol Ammonium Acetate) B (MeCN, 0.05% Formic acid):Gradient 100%A 0.7mins, 100%A-100% B 3.5mins,100% B 3.5mins, 100%-0%B 0.3mins; flow rate = 1 ml/min)

Ref 1. Rodney L. Johnson, G. Rajakumar, Kuo-Long Yu, Ram K Mishra, J. Med. Chem., 1986,29,10,2104-2107.

Intermediate 31

2S-[4-(cis-2,3-Dimethyl-cyclopropanecarbonyl)-6S-methyl-5-oxo-hexahydro- (3aS,6aR)-(pyrrolo[3,2-b]pyrrole-1 -carbonyl]-azetidine-1 -carboxylic acid tert- butyl ester The above Intermediate was prepared in a similar manner to Intermediate 13 by reacting the Intermediate 16 with N-tert-butyloxycarbonyl-L-azetidine-2- carboxylic acid ¹.

LCMS: 420 [MH]⁺, 364 [M -tBu]⁺, 320 [M-Boc]⁺, Retention time = 3.09 min., using the same conditions as in Intermediate 30 Intermediate 32 2-Bromo-benzothiazole-6-carboxylic acid ethyl ester

Tert-butyl nitrite (1.2mL,10.09 mmol, 1.5 eq) and copper (II) bromide (1.84g,8.26 mmol,1.2eq) were stirred in dry acetonitrile (30 mL) under an atmosphere of nitrogen. 2-Amino-benzothiazole-6-carboxylic acid ethyl ester (1.53g,6.88 mmol,1 eq) was added portionwise to the stirred solution over 25 min. After 21 hours the reaction mixture was filtered to remove copper salts and the filtrate was poured into 2N HCI (75 mL) and extracted with diethyl ether (75 mL then 2x40 mL). The combined organic extracts were then washed with 2N HCI (40 mL), brine (40 mL), dried (MgSO₄) and the solvent evaporated in vacuo to give the title compound an orange solid (1.63g). This was used to prepare intermediate 33 without further purification. Mass Spec. : 286,288 [MH]⁺

Intermediate 33 2-Bromo-6-hydroxymethyl-benzothiazole

To the Intermediate 32, (1.70g,5.94 mmol,1 eq) in water: dioxan (20 mL:20 mL), stirred at -2°C (ice/salt bath) was added NaBH₄ (1.8g,47.5 mmol, 8 eq.), portionwise over 25 min. so there was no rise in temperture. After stirring at 0°C for 3 hours and then at room temperature for 19 hours, the reaction mixture was carefully acidified with 2N HCI whilst cooling (ice bath) so that the temperature did not rise above 10°C. The aqueous phase was then extracted with ethyl acetate (X3) and the solvent was evaporated to give a yellow solid (2.34g). The product was purified by flash column chromatography over silica gel ( Merck 9385) using cyclohexane:ethyl acetate as the eluting solvent system. The title compound, was isolated as a yellow solid (0.248g). Mass Spec: 244,246 [MH]⁺

Intermediate 34

2-Bromo-6-(tert-butyl-diphenyl-silanyloxymethyl)-benzothiazole

To a solution of Intermediate 30 (0.120g,0.492 mmol, 1 eq.) in anhydrous DMF (5 mL) was added imidazole (0.671 g,9.86 mmol, 20 eq) and tert- butylchlorodiphenylsilane (0.256 mL,0.983 mmol,2 eq). The reaction mixture was stirred at room temperature under nitrogen for 3 days and the solvent was then evaporated. The residue was partitioned between diethyl ether (30 mL) and water (15 mL). The aqueous phase was then re-extracted with diethyl ether (20 mL) and the combined organic phases were washed with 1 N HCI (20 mL), water (15 mL), sat. NaHCO₃ solution (15 mL), brine (15 mL) and dried (MgSO₄). The solvent was evaporated to give a yellow liquid (1.45g) which was purified by flash column chromatography over silica gel ( Merck 9385) using cyclohexane: diethyl ether as the eluting solvent system to give the title compound, as a colourless gum (0.177g). Mass Spec: 484 [MH]⁺

Intermediate 35

2S-{4-[6-(tert-Butyl-diphenyl-silanyloxymethyl)-benzothiazol-2-yl1-6S-methyl-5- oxo-hexahydro-(3aS,6aR)-pyrrolo[3,2-b]pyrrole-1-carbonyl}-pyrrolidine-1- carboxylic acid tert-butyl ester

Prepared in a similar manner to Intermediate 22 by reacting the Intermediate 27 with Intermediate 34 to give the title compound as a pale yellow foam. Mass Spec: 639 [MH-BOC]⁺, 739 [MH]⁺

Intermediate 36 2-Bromo-4-methoxy-benzothiazole.

Prepared in a similar manner to Intermediate 32 starting from 2-amino-4- methoxy-benzothiazole, to give the title compound as a yellow solid. ¹H nmr (CDCI₃): δ 7.37 (2H, m),(1 H, m), 4.04 (3H, s). Mass spec 244, 246 [MH]⁺.

Intermediate 37

2S-[4-(4-Methoxy-benzothiazol-2-yl)-6S-methyl-5-oxo-hexahydro-(3aS,6aR)- pyrrolo[3,2-b]pyrrole-1-carbonyl]-pyrrolidine-1 -carboxylic acid tert-butyl ester

Prepared in a similar manner to Intermediate 22 by reacting the Intermediate 27 with Intermediate 36 to give the title compound, as a white solid. ¹H nmr (CDCI₃): δ 7.41 (1 H, d); 7.27 (1 H, d); 6.90 (1 H, d); 4.43 (1 H, m); 4.02 (3H, s); 4.29-4.07 (2H, m), 3.95-3.69 (2H, m);3.68-3.36 (3H, m); 3.19 (1 H, m); 2.40 (1 H, m); 2.20 and 1.90 (4H, 2m); 1.45 (9H, d); 1.18 (3H, m). Mass spec 501 [MH]⁺, 401 [M-Boc].

Intermediate 38 2-Bromo-5-chloro-benzothiazole

A solution of bromine (0.95mL, 18.5mmol, 3.6eq) in chloroform (10mL) was added dropwise over a period of 1hr 50min to a stirred suspension of 5-chloro-2- mercaptobenzothiazole (1.02g, 5.05mmol) in chloroform (10mL) at room temperature. The mixture was stirred for 30min before it was added slowly to water (50mL) and stirred for a further 20min. Then the mixture was flltered to remove a cream solid. The organic phase was dried (MgSO₄) and evaporated to leave a brown solid. The brown solid was dissolved in ether (10mL) and filtered. The residue was washed with ether (10mL). The filtrate and washings were combined and evaporated to give the title compound as a pale yellow/brown solid, 1.04g.

¹H nmr (CDCI₃): δ 8.00 (1 H, d) 4-H; 7.83 (1 H, d) 7-H; 7.42(1 H, dd)5-H. Mass spec 248, 250 [MH]⁺.

Intermediate 39

2S-[4-(5-Chloro-benzothiazol-2-yl)-6S-methyl-5-oxo-hexahydro-(3aS,6aR)- pyrrolo[3,2-b]pyrrole-1-carbonyl]-pyrrolidine-1 -carboxylic acid tert-butyl ester

Prepared in a similar manner to Intermediate 22 by reacting the Intermediate 27 with Intermediate 38 to give the title compound as a white solid. ¹H nmr (CDCI₃): δ 7.78 (1 H, d); 7.72 (1 H, d); 7.28 (1 H, dd); 4.44 (2H, m); 4.13 (1 H, m); 3.99-3.70 (2H, m); 68-3.36 (3H, m); 3.12 (1 H, m); 2.37 (1 H, m); 2.20 and 1.91 (4H, 2m); 1.45 (9H, d); 1.20 (3H, m). Mass spec 505 [MH]⁺, 405 [M-Boc]. Intermediate 40 2-Bromo-5-methoxy-thiazolo[5,4-b]pyridine,

Prepared in a similar manner to Intermediate 32, starting from 2-amino-5- methoxy-thiazolo[5,4-b]pyridine compound(ex Maybridge) to give the title compound as a yellow solid. mass spec:[MH]⁺ = 247

Η nmr(CDCI₃): d 8.05(d, 1 H); 6.85(d, 1H); 3.0(s, 3H);

Intermediate 41

2S-[4-(5-Methoxy-thiazolo[5,4-b]pyridin-2-yl)-6S-methyl-5-oxo-hexahydro- (3aS,6aR)-pyrrolo[3,2-b]pyrrole-1 -carbonyl]-pyrrolidine-1 -carboxylic acid tert- butyl ester

Prepared in a similar manner to Intermediate 22 by reacting the Intermediate 27 with Intermediate 40 to give the title compound as a glassy solid, mass spec:[MH]⁺ = 502

Η nmr(CDCI₃): d 7.9(d, 1 H); 6.8(d,1 H); 4.4(m, 2H); 4.0(s, 3H); 4.2-3.4(m 6H); 3.1 (m, 1 H); 2.4-1.8(m,5H); 1.4(m, 9H); 1.2(d, 3H)

Intermediate 42 2-Bromo-5-methoxy-benzothiazole

Prepared in a similar manner to Intermediate 38 GW421721X (R95/170), starting from 5-methoxy-2-mercaptobenzothiazo!e (ex Lancaster), to give after purification by flash column chromatography the title compound as a white solid. ¹H nmr (CDCI₃): δ 7.65 (1 H, d) 7-H; 7.47 (1 H, d) 4-H; 7.06 (1 H, dd) 6-H; 3.88 (3H, s) OCH₃. Mass spec 244, 246 [MH]⁺.

Intermediate 43

2S-[4-(5-Methoxy-benzothiazol-2-yl)-6S-methyl-5-oxo-hexahydro-(3aS,6aR)- pyrrolo[3,2-b]pyrrole-1-carbonyl]-pyrrolidine-1 -carboxylic acid tert-butyl ester Prepared in a similar manner to Intermediate 22 by reacting the Intermediate 27 with Intermediate 42 to give the title compound as an off-white foam. ¹H nmr (CDCI₃): δ 7.66 (1 H, d); 7.30 (1 H, d); 6.95 (1 H, dd); 4.48 (1 H, m); 4.42 (1 H, m); 4.13 (1 H, m); 3.88 (3H, s); 3.94-3.69 (2H, m); 3.65-3.48 (2H, m); 3.45 (1 H, m); 3.14 (1 H, m); 2.38 (1 H, m); 2.20 and 1.90 (4H, 2m); 1.45 (9H, d); 1.20 (3H, m). Mass spec 501 [MH]⁺, 401 [M - Boc group]*.

Intermediate 44 2S-[4-(4-Chloro-benzothiazol-2-yl)-6S-methyl-5-oxo-hexahydro-(3aS,6aR)- pyrrolo[3,2-b]pyrrole-1-carbonyl]-pyrrolidine-1 -carboxylic acid tert-butyl ester

Prepared in a similar manner to Intermediate 22 by reacting the Intermediate 27 with 2-bromo-4- chlorobenzothiazole¹, to give the title compound as a white solid.

¹H nmr (CDCI₃): δ 7.71 (1 H, d) and 7.46 (1 H, d); 7.23 (1 H, t) 6'-H; 4.44 (1.5H, m) and 4.17 (1.5H, m); 4.01-3.70 (2H, m); 3.69-3.36 (3H, m); 3.21 (1 H, m) 6-H; 2.40 (1 H, m) 6-H; 2.32- 1.78 (4H, m); 1.46 (9H, s); 1.20 (3H, m). Mass spec 505 [MH]⁺, 405 [M - Boc group]*.

Ref 1. R.C. Elderfield and F.W.Short J. Org Chem. 1953, 18, 1092-1103.

Intermediate 45

2-B romo-6-nitro-benzoth iazole

Prepared in a similar manner to Intermediate 38 starting from 2-mercapto-6- nitrobenzothiazole (ex Acros), to give the title compound, as an orange solid. ¹H nmr (CDCI₃): δ 8.88 (1 H, d); 8.38 (1 H, dd); 8.11 (1 H, d). GC/Mass spec 258, 260 [MH]⁺, at 17.33 min, 98.5%.

Intermediate 46

2S-[6S-Methyl-4-(6-nitro-benzothiazol-2-yl)-5-oxo-hexahydro-(3aS,6aR)- pyrrolo[3,2-b]pyrrole-1-carbonyl]-pyrrolidine-1 -carboxylic acid tert-butyl ester Prepared in a similar manner to Intermediate 22 by reacting the Intermediate 27 with Intermediate 45 to give the title compound as a yellow solid. ¹H nmr (CDCI₃): δ 8.72 (1 H, d); 8.30 (1 H, dd); 7.82 (1 H, d); 4.53- 4.37 (1.5H, m) and 4.29- 4.06 (1.5H, m); 4.04-3.73 (2H, m) 3a-H and 5-H; 3.69- 3.36 (3H, m); 3.14 (1 H, m); 2.42 (1 H, m); 2.32- 1.83 (4H, m); 1.45 (9H, d); 1.24 (3H, m). Mass spec 516 [MH]⁺, 416 [M - Boc group]⁺.

Intermediate 47 2-Bromo-6-fluoro-benzothiazole

Prepared in a similar manner to Intermediate 32, starting from 2-amino-6- fluorobenzothiazole (ex Aldrich) to give the title compound as an orange, fluffy solid.

¹H nmr (CDCI₃): δ 7.94 (1 H, dd) 4-H; 7.41 (1 H, dd) 7-H; 7.24 (1 H, td) 5-H. Mass spec 234, 232 [MH]⁺.

Intermediate 48

2S-[4-(6-Fluoro-benzothiazol-2-yl)-6S-methyl-5-oxo-hexahydro-(3aS,6aR)- pyrrolo[3,2-b]pyrrole-1-carbonyl]-pyrrolidine-1 -carboxylic acid tert-butyl ester

Prepared in a similar manner to Intermediate 22 by reacting the Intermediate 27 with Intermediate 47 to give the title compound as an off-white foam.

¹H nmr (CDCI₃): δ 7.72 (1 H, dd); 7.50 (1 H, dd); 7.16 (1 H, td); 4.52-4.36 (2H, m);

4.14 (1H, m); 3.97-3.71 (2H, m); 3.66-3.36 (3H, m); 3.12 (1H, m); 2.38 (1H, m) 6-H; 2.34-2.04 (2H, m) and 1.91 (2H, m); 1.45 (9H, d); 1.20 (3H, dd).

Mass spec 489 [MH]⁺, 389 [M - Boc group]*.

Intermediate 49 2-Bromo-4-methoxy-7-methyl-benzothiazole

Prepared in a similar manner to Intermediate 32 starting from 2-amino-4- methoxy-7-methylbenzothiazole¹, to give the title compound, as a pale yellow solid.

¹H nmr (CDCI₃): δ 7.13 (1 H, d); 6.83 (1 H, d); 4.01 (3H, s); 2.44 (3H, s). Mass spec 258, 260 [MH]⁺. Ref 1. J. Weinstock, D.E. Gaitanofoulous, O.D. Stringer, R.G. Franz, J.P.Hiefle, L.B. Kinter, W.A. Mann, K. E. Flaim and G. Gessner. J.Med.Chem. 1987, 30, 1166-1176

Intermediate 50

2S-[4-(4-Methoxy-7-methyl-benzothiazol-2-yl)-6S-methyl-5-oxo-hexahydro- (3aS,6aR)-pyrrolo[3,2-b]pyrrole-1 -carbonyl]-pyrrolidine-1 -carboxylic acid tert- butyl ester Prepared in a similar manner to Intermediate 22 by reacting the Intermediate 27 with Intermediate 49 to give the title compound as a yellow solid.

¹H nmr (CDCI₃): δ 7.05 (1 H, d); 6.83 (1 H, d); 4.47 (1 H, m); 4.39 (1 H, m); 4.18 (1 H, m); 4.00 (3H, s); 3.94-3.71 (2H, m); 3.67-3.36 (3H, m); 3.20 (1 H, m); 2.48 (3H, s); 2.50-2.02 (3H, m); 1.90 (2H, m); 1.45 (9H, d); 1.19 (3H, dd). Mass spec 515 [MH]⁺, 415 [M - Boc group]*.

Intermediate 51

2S-[4-(Bicyclor4.1.0]hept-3-ene-7-carbonyl)-6S-methyl-5-oxo-hexahydro- (3aS,6aR)-pyrrolo[3,2-b]pyrrole-1-carbonyl]-pyrrolidine-1 -carboxylic acid tert- butyl ester

To a solution of (1a,6a,7a)-bicyclo[4.1.0]hept-3-ene-7-carboxylic acid¹ ( 103mg, 0.75mmol) anhydrous dichloromethane ( 4 mL) stirred under nitrogen was added anhydrous N,N-dimethylformamide (10 ml ) followed by oxalyl chloride ( 100 mL , 1.14 mmol) over 3 min. The solution was stirred for a further 1h and was then concentrated under reduced pressure. Toluene (3 x 3 mL ) was added and evaporated off and the resulting liquid was dried under vacuum for 1 h. A solution of this acid chloride in anhydrous tetrahydrofuran (2 mL) was added to a solution of the Boc proline trans-lactam (169 mg, 0.5 mmol ) Intermediate 27 in anhydrous tetrahydrofuran (4 mL) which had been stirred with lithium hexamethyldisilazide ( 0.55 mL of a 1 M solution in tetrahydrofuran, 0.55 mmol ) under nitrogen in an ice/ IMS bath for 15 min. The resulting solution was stirred in an ice/ IMS bath for 2h. It was then poured into saturated ammonium chloride solution ( 10 mL ) and extracted twice with ethyl acetate ( 25mL, 10 mL ). The combined organic phases were washed with saturated sodium bicarbonate solution (10 mL ), water (10 mL), and saturated brine ( 10 mL ) dried over magnesium sulphate and evaporated. The crude mixture was purified using a Bond Elute^® silica column eluting with dichloromethane, chloroform, ether, 1 :1 cyclohexane : ethyl acetate and ethyl acetate to give a white solid foam (177 mg, 76% ). H nmr (CDCI₃): δ 5.6 - 5.5 ( m), 4.5 - 4.0 ( m), 3.8 - 3.25 ( m), 2.72 ( m) , 2.55 - 2.35 ( m), 2.3 - 1.8 ( m), 1.7 - 1.5 ( m), 1.46 ( s), 1.40 ( s), 1.25 ( m), 1.12 (3H, dd). Mass spec 458 [MH]*. Ref. 1 H Musso and U. Bietham , Chem Ber, 1964, 97, 2282

Intermediate 52

(3aS,6S,6aR)-4-Thiazol-2-yl-6-methyl-5-oxo-hexahydro-pyrrolo[3,2-b]pyrrole-1- carboxylic acid 4-methoxy-benzyl ester

To the Intermediate 21 , (500mg, 1.643mmol, 1eq.) was added 2-bromothiazole (Aldrich) (0.296mL, 3.286mmol, 2.0eq.), potassium carbonate (453mg, 3.286mmol, 2.0 eq.), copper (I) chloride (163mg,1.643mmol,1eq.), TDA-1 (0.161mL,0.493 mmol, 0.3eq.) and xylene (30mL). The reaction mixture was refluxed under a Dean-Stark head for 6 hours. After leaving to cool, the brown solids were filtered off and washed with ethyl acetate (30mL). The combined filtrate and washings were washed with 0.1 M HCI aq. (100mL) and brine (100mL) and dried over MgSO₄. The solvent was evaporated to give a brown gum which was purified by flash column chromatography over silica gel (Merck 9385) using cyclohexane:ethyl acetate 4:1. The title compound was obtained as a colourless gum (225mg). ¹H nmr (CDCI₃): d 7.45 (1 H, d); 7.30 (2H, m); 7.00 (1 H, d); 6.90 (2H, m); 4.95- 5.20 (2H, m); 3.85-4.20 (2H, m); 3.80 (3H, s); 3.10-3.80 (2H, m); 3.00-3.40 (1H, 2 broad signals); 2.86-3.00 (1 H, m); 2.10-2.39 (1 H, m); 1.02-1.35 (3H, broad m).

Intermediate 53 rel-(3S,3aR,6aS)-3-Methyl-1-thiazol-2-yl-hexahydro-pyrrolo[3,2-b]pyrrol-2-one trifluoroacetate

To the Intermediate 52, (214mg, 0.552mmol) was added trifluoroacetic acid

(10mL) at room temperature. After 20 minutes the solvent was evaporated in vacuo to give, after trituration under diethyl ether, the title compound as a colourless solid, (186 mg). ¹H nmr (CDCI₃): d 7.45 (1 H, d); 7.06 (1 H, d); 6.5-7.0 (1 H, broad); 4.22-4.39 (1 H, m); 3.45-3.96 (4H, m); 3.04-3.26 (2H, m); 2.20-2.46 (1 H, m); 1.35 (3H, s).

Intermediate 54

2S-(6S-Methyl-5-oxo-4-thiazol-2-yl-hexahydro-(3aS,6aR)-pyrrolo[3,2-b]pyrrole-1- carbonyl)-pyrrolidine-1 -carboxylic acid tert-butyl ester and 2S-(6R-Methyl-5-oxo- 4-thiazol-2-yl-hexahydro-(3aR,6aS)-pyrrolo[3,2-b]pyrrole-1-carbonyl)-pyrrolidine- 1 -carboxylic acid tert-butyl ester

To a solution of (S)-N-tert-butyloxycarbonyl proline (127 mg, 590 mmol, 1.1 eq.) in anhydrous DMF (2 mL) stirred at room temperature was added a solution of TBTU (189 mg, 589 mmol) in 1 :1 acetonitrile:DMF (2 mL) and HOBT (80mg). The mixture was stirred at room temperature for 30 min, then the (racemic) amine trifluoroacetate (180 mg, 534 mmol) was added followed by diisopropylethylamine (138 mg, 2 eq., 186 mL). The mixture was stirred at room temperature overnight. It was then partitioned between ethyl acetate and water. The organic phase was separated, dried over anhydrous magnesium sulphate and filtered. The filtrate was evaporated under reduced pressure and the resulting gum was purified by preparative plate chromatography (Whatman

PK6F silica gel 60A plate) eluting with 1 :1 ethyl acetate:cyclohexane to give the title compound (3aS, 6aR, 6S diastereomer) as an off-white solid (54 mg, 21.7%) as the less polar component (Rf 0.44 in ethyl acetate). ¹H nmr (CDCI₃): d 7.45 (1 H, d); 7.00 (1 H, d); 4.35-4.51 (2H, m); 3.96-4.30 (2H, m); 3.65-3.95 (2H, m); 3.48-3.65 (2H, m); 2.95-3.10 (1 H, m); 1.80-2.42 (5H, m); 1.45 (9H, s); 1.10-1.25 (3H, m). Mass spectrum 421 [MH]*,

The more polar 3aR, 6aS, 6R diastereomer was obtained as an off-white solid (55 mg, 22.1 %)

(Rf 0.39 in ethyl acetate).

¹H nmr (CDCI₃): d 7.45 (1H, m); 7.00 (1H, m); 4.30-4.44 (1H, m); 3.82-4.28 (3H, m); 3.32-3.78 (4H, m); 2.90-3.15 (1 H, d); 1.80-2.40 (5H, m); 1.40 (9H, s), 1.20-

1.35 (3H, m). Mass spectrum 421 [MH]*. Intermediate 55

(3S,3aR,6aS)-3-Methyl-4-(pyrrolidine-2S-carbonyl)-1-thiazol-2-yl-hexahydro- pyrrolo[3,2-b]pyrrol-2-one trifluoroacetate.

To the Intermediate 54 (54 mg, 128mmol) was added trifluoroacetic acid (5 mL). After 1 hour, the solvent was removed under reduced pressure; the resulting gum was taken up in diethyl ether and azeotroped. Trituration under diethyl ether afforded the title compound as a tan solid (45 mg, 80%). ¹H nmr (CDCI₃): δ 7.45 (1H, d); 7.02 (1 H, d); 4.8-5.0 (1H, broad); 3.98-4.30 (2H, m); 3.70-3.95 (2H, m); 3.25-3.65 (3H, m); 3.05-3.20 (1 H, m); 2. 0-2.65 (5H, broad m , overlapping with HOD signal); 1.18 (3H, d). Mass spectrum 321 [MH]*.

Intermediate 56

(3S)-2-Oxo-3-(2,2,2-trifluoroacetylamino)-pyrrolidine-1 -carboxylic acid benzyl ester

Trifluoroacetic acid (35ml, 0.45M) was added over 2 min to a stirred solution of the racemic lactam (SR)-3-tert-Butoxycarbonylamino-2-oxo-pyrrolidine-1- carboxylic acid benzyl ester (Ref: International patent application published under No. WO97/36903) (20.03g 59.9mmol) in dichloromethane (100ml) at room temperature under nitrogen. After 4h tic indicated absence of starting material so the mixture was concentrated in vacuo. The residue was partitioned between dichloromethane (350ml) and acqueous potassium carbonate (10% w/v, 350ml). The acqueous layer was re-extracted with dichloromethane (350ml). The organic solutions were combined, washed with saturated brine (250ml) dried (MgSO₄) and filtered into a suspension of (-)-di-p-toluoyl-L-tartaric acid monohydrate (12.31g, 30.5mmol) in dichloromethane (300ml). The resulting solution was concentrated in vacuo to give a white solid which was suspended in tetrahydrofuran (500ml) under nitrogen. 3,5-Dichlorosalicylaldehyde (0.5g, 2.6mmol) was added and the mixture was stirred at room temperature under nitrogen for 14 days monitoring derivitised aliquots by hplc The solid was filtered off, washed with tetrahydrofuran and partitioned between ethyl acetate (500ml) and aqueous potassium carbonate (1 M, 200ml). The residue was dissolved in dichloromethane (50ml) and methanol (10ml). Methyl trifluoroacetate (18.5ml, 0.183M) and 4-methylmorpholine (6.1 ml, 55.4mmol) were added and the resulting mixture was stirred at room temperature under nitrogen for 20h. After concentrating in vacuo the residue was partitioned between dichloromethane (150ml) and dilute hydrochloric acid (1M, 50ml). The organic layer was washed with dilute hydrochloric acid (1 m, 50ml) and saturated brine (50ml) dried (Na₂SO₄) and concentrated in vacuo to give a yellow solid. This was recrystallised from an ethyl acetate/cyclohexane mixture (1 :1 , 116ml) to give the title compound (4g, 20.3%) as a cream solid. HPLC: Retention time = 11.04min 99.5%, using Chiralpak AD 85% Heptane/15% Ethanol, Flow: 1 ml min-1 , uv 215nm.

¹H nmr (CDCI₃): δ 7.39 (m, 5H); 7.16 (d,1 H); 5.28 (s, 2H); 4.55 (m, 1 H); 3.91 (dd, 1 H); 3.65 (m, 1 H); 2.72 (m, 1 H); 1.95 (m, 1-H).

Intermediate 57

(3S)-2-Methoxy-3-(2,2,2-trifluoroacetylamino)pyrrolidine-1 -carboxylic acid benzyl ester

Lithium borohydride (2M in tetrahydrofuran; 6.2ml, 12.4mmol) was added over 5 min, to a stirred suspension of the Intermediate 56, (4g, 12.1 mmol) in dry tetrahydrofuran (16ml) under nitrogen at -20°C. After stirring, under nitrogen at - 20°C for 2h the mixture was added over 20 min to a stirred solution of acetyl chloride (1.4ml, 19.7mmol) in methanol (16ml) at 18°C under nitrogen. After stirring under nitrogen at 18°C for 1h this mixture was added over 15 min to a stirred solution of potassium hydrogen carbonate (2g, 20mmol) in water (8ml) at 21 °C, 30 min later a solution of sodium chloride in water (5% w/v, 20ml) was added and stirring was continued for 15 min. The layers were separated and the aqueous layer was extracted with t-butyl methyl ether (2x12ml). The combined organic solutions were washed with brine (5% w/v, 8ml), dried (Na₂SO₄) and concentrated in vacuo. The residue was purified by column chromatography on silica gel (150g) with dichloromethane:ether (4:1 ) as eluent. Concentration in vacuo of the fractions with an RF = 0.57 gave a white solid (1.4g, 33.4%).

¹H nmr (400 MHz; DMSO, 100°C): δ 9.1-9.03 (bs, 1 H); 7.39-7.28 (m, 5H); 5.13 (d, J=3Hz; 2H, ); 5.11 (d, J=4.5 Hz; 1 H ); 4.16 - 4.07 (m, 1 H); 3.47 - 3.31 (m, 2H); 3.29 (s, 3H ); 2.35 - 2.06 (m, 2H ) Mass spec [MNH4]* = 364

HPLC: Retention time = 8.28min, 98.1% using the same conditions as in Example 56.

Concentration in vacuo of the fractions with an RF = 0.40 gave a white solid (1.38g, 32.9%)

¹H nmr (400 MHz; DMSO, 100°C): δ 9.38 - 9.27 (bs, 1-H, ), 7.42 - 7.28 (m; 5-H, ) 5.14 (s;2-H, ), 4.99 (s; 1-H, d), 4.14 (t, J=6HZ; 1-H, ), 3.71 - 3.61 (m; 1-H, ) 3.49 - 3.37 (m; 1-H, ), 3.29 (s; 3-H, ) 2.30 - 2.18 (m; 1-H, ), 1.98 - 1.90 (m;1-H, ) Mass spec [MNH4]+ = 364

HPLC: Retention time = 4.95 min 95.2%, using the same conditions as in Example 56.

Intermediate 58

(2R,3S)-2-(1-Ethoxycarbonyl-ethyl)-3-(2,2,2-trifluoro-acetylamino)-pyrrolidine-1- carboxylic acid benzyl ester Boron trifluoride diethyl etherate (0.6ml, 4.73mmol) was added over 10 min to a stirred mixture of the Intermediate 57 (0.79g, 2.28mmol) and ethyl trimethylsilyl methylketene acetal (1.2g, 6.88mmol) in dry dichloromethane (7.5ml) at 0°C under nitrogen. After 2.5h the mixture was poured into saturated aqueous sodium bicarbonate (19ml) and dichloromethane (11ml). The layers were separated and the aqueous layer was re-extracted with dichloromethane (19ml). The organic solutions were combined, washed with saturated brine (20ml), dried (MgSO₄) and concentrated in vacuo. The residue was purified by column chromatography on silica gel (70g) with a graded elution of dichloromethane:ether from (9:1 ) to (4:1 ). Concentration of the appropriate fractions gave the title compound as a colourless oil (380mg, 26.4%).

Tic silica/dichloromethane:Ether (9:1 ) Rf 0.31

¹H nmr (CDCI₃): δ 7.4-7.3 (m,5H); 5.14 (d,J=3Hz, 2H); 4.52 (m,1 H);

4.4 - 4.0 (3H,d); 3.8 - 3.55 (bs, 1H ); 3.55 - 3.37 (m, 1H); 3.37 - 2.70 (1-H);

2.50 - 2.17 (m 1 H ); 2.0 - 1.82 (m, 1 H); 1.28 - 1.10 (m, 6H) Mass spec thermospray t,+ve m/e: 417 = [MNH4]* HPLC: Retention time = 18.36 min, 54.5%; 19.64min, 45.5% using a OA Chiral Column, eluent heptane:ethanol (98:2), flow: 1 ml min ^"1 , detection: uv 215nM,

Intermediate 59

(2R,3S)-3-Amino-2-(1 -ethoxycarbonyl-ethyl)-pyrrolidine-1 -carboxylic acid benzyl ester

A solution of potassium carbonate in water (1 M, 5ml) was added to a stirred solution of the Intermediate 58 (360mg, 0.85mmol) in ethanol (5ml) and the resulting mixture was stirred at room temperature for 24h whereupon the indicated consumption of starting material. The mixture was extracted with ethyl acetate (3x5ml) and the combined extracts were washed with saturated brine (5ml). After drying over magnesium sulphate, solvent removal in vacuo gave a yellow oil. This was dissolved in chloroform (10ml) and re-dried over magnesium sulphate, sovent removal in vacuo followed by vacuum drying afforded a pale yellow oil (230mg, 83%).

¹H nmr (CDCI₃): δ 7.45 - 7.19 (m, 5-H); 5.23 - 5.00 (bd, 2H); 4.2 - 3.94 (m, 2H); 3.94 - 2.71 (m, 4H); 2.71 - 2.31 (bs, 3H); 2.2 - 1.95 (m 1 H); 1.82 - 1.58 (bs, 1 H); 1.30 - 0.98 (m, 6H) Mass spec thermospray: +ve m/e 321 = [MH*], 641 = 2M.H]*

B. Preparation of Examples

Example 1

(3S,3aR,6aS)-1 -Cyclopropanecarbonyl-4-[1 -(5-dimethylamino-naphthalene-1 - sulfonyl)-pyrrolidine-2S-carbonyl]-3-methyl-hexahydro-pyrrolo[3,2-b]pyrrol-2-one

The Intermediate 14 (57mg,233mmol,1eq.) was dissolved in MeCN (4 mL). To the clear solution was added triethylamine (80mL,574mmol,2.5eq.) and a solution of dansyl chloride (69mg,256mmol,1.1eq.) in MeCN (1 mL). The reaction mixture was stirred at room temperature for 1.5 hours. Isopropanol (10mL, 128mmol) was added and the solution was evaporated to dryness. The residue was purified by flash column chromatography silica gel (Merck 9385) and eluted with cyclohexane:ethyl acetate to give the title compound as a yellow/ green foam (15.4mg).

¹H nmr (CDCI₃): δ 8.54 (1 H,d), 8.42 (1 H,d), 8.29 (1 H,d), 7.55 (2H,dd), 7.18 (1 H,d), 4.75 (1 H,dd), 4.26 (1 H,t), 3.89-3.20 (6H,m), 3.00-2.84 (7H,m), 2.83-2.68 (1 H,m), 2.32-1.80 (5H,m), 1.28-0.94 (7H,m) ppm. Mass Spec. 539 [MH]*¹ 471 [M-COcyclopropyl]*

Circular dichroism λ_max200.8 nm, dE 0.01 ; λ_max217.8 nm, dE -23.80; λ_max238.4 nm, dE 15.10;

(3S,3aR,6aS)-1-Cyclopropanecarbonyl-4-[1-(5-dimethylamino-naphthalene-1- sulfonyl)-pyrrolidine-2S-carbonyl]-3-methyl-hexahydro-pyrrolo[3,2-b]pyrrol-2-one (Alternative Synthesis)

The title compound was prepared in a similar manner to Intermediate 11 from intermediate 29

Mass Spec: 539 [MH]*¹ 471 [M-COcyclopropyl]*

Example 2

2S-(4-Cyclopropanecarbonyl-6S-methyl-5-oxo-hexahydro-(3aS,6aR)-pyrrolo[3,2- b]pyrrole-1 -carbonyl)-pyrrolidine-1 -carboxylic acid (4-isopropyl-phenyl)-amide

To a solution of Intermediate 14 (0.03g,0.09mmol) in dry acetonitrile(2ml) was added dry triethylamine(0.03ml,0.22mmol) followed by addition of 4- isopropylphenyl isocyanate(0.022g,0.135mmol). The reaction mixture was left standing at room temperatue overnight, then quenched with propan-2-ol (0.1 ml). It was then evaporated to dryness and purified using preparative plate chromatography (WhatmanPKβF silica gel 60A plate) eluting with ethyl acetate/cyclohexane(2:1 ) to give the title compound as an amorphous solid, mass spec: [MH]* = 467. Η nmr(CDCI₃): δ 7.25-7.1 (m,4H); 6.2(s,1H); 4.6(m,1 H); 4.45(t,1 H); 3.9- 3.4(m,5H); 3.3(m,1 H); 3.0-2.7(m,3H); 2.4- 1.9(m,5H);1.22(d,6H); 1.19(d,3H); 1.1-0.9(m,4H)

Example 3-5,7 The above Examples were prepared in a similar manner to Example 2 from Intermediate 14. Example 3

2S-(4-Cyclopropanecarbonyl-6S-methyl-5-oxo-hexahydro-(3aS,6aR)-pyrrolo[3,2- b]pyrrole-1 -carbonyl)-pyrrolidine-1 -carboxylic acid (4-tert-butyl-phenyl)-amide

Mass spec:[MH]* = 481

Η nmr(CDCI₃): δ 7.25(m,4H); 6.2(s,1 H); 4.65(m,1 H); 4.48(t,1 H); 3.85-

3.45(m,5H); 3.3(m,1 H); 2.9(m,1 H); 2.7(m,1 H); 2.4-1.85(m,5H); 1.3(s,9H);

1.19(d,3H); 1.2-0.95(m,4H).

Example 4

2S-(4-Cyclopropanecarbonyl-6S-methyl-5-oxo-hexahydro-(3aS,6aR)-pyrrolo[3,2- b]pyrrole-1 -carbonyl)-pyrrolidine-1 -carboxylic acid (4-trifluoromethoxy-phenyl)- amide

Mass spec:[MH]* = 509

'H nmr(CDCI₃): δ 7.4(d,2H); 7.1 (d,2H); 6.4(s,1 H); 4.6(m,1 H); 4.4(t,1 H); 3.9- 3.4(m,5H); 3.3(m,1 H); 2.9(m,1 H); 2.75(m,1 H); 2.4-1.9(m,5H); 1.2(d,3H); 1.2- 0.9(m,4H). Circular dichroism λ_max204.2 nm, dE -9.45; λ_max218.6 nm, dE -20.48; λ_max239.0 nm, dE 24.70.

Example 5

2S-(4-Cyclopropanecarbonyl-6S-methyl-5-oxo-hexahydro-(3aS,6aR)-pyrrolo[3,2- b]pyrrole-1 -carbonyl)-pyrrolidine-1 -carboxylic acid (4-trifluoromethylsulfanyl- phenyl)-amide

Mass spec:[MH]* = 525

Η nmr(CDCI₃): δ 7.55(d,2H); 7.45(d,2H); 6.4(s,1 H); 4.65(m,1 H); 4.4(t,1 H); 3.9-

3.5(m,5H); 3.3(m,1 H); 2.95(m,1 H); 2.75(m,1 H); 2.4-1.9(m,5H); 1.15(d,3H); 1.2- 0.9(m,4H).

Example 6

2S-(4-Cyclopropanecarbonyl-6S-methyl-5-oxo-hexahydro-(3aS,6aR)-pyrrolo[3,2- b]pyrrole-1 -carbonyl)-pyrrolidine-1 -carboxylic acid (4-propoxy-phenyl)-amide

To a stirred solution of di-tert-butyldicarbonate(0.0195g,0.09mmol) in dry acetonitrile(3ml) was added 2,4-dimethylamino pyridine(0.001g,0.009mmol) and a solution of 4-n-propoxyphenyl amine(0.0128g,0.085mmol) in acetonitrile(2ml). The reaction mixture was stirred at room temperature for 10mins before a solution of Intermediate 14 (0.03g,0.09mmol) in acetonitrile(2ml) and triethyl amine(0.012ml,0.09mmol) was added. The reaction mixture was stirred at room temperature overnight.then quenched with isopropanol(O.l ml). The solvent was evaporated under reduced pressure and the residue was partitioned between ethyl acetate(IOml) and water(IOml). The organic phase was washed with saturated sodium hydrogen carbonate(IOml), dried over MgSO4 and evaporated to dryness. HPLC purification using (Gilson Supelcosil LC ABZplus column, eluent system A (H₂O, 0.1% Formic acid) B (MeCN, 0.05% Formic acid):Gradient 0%B 1.45mins, 0%-95% B 20mins,95% B 10mins, 100%B 2mins; flow rate = 4ml/min) gave the title compound as an amorphous solid. Mass spec:[MH]^* = 483 'H nmr(CDCI₃): δ 7.2(d,2H);6.8(d,2H); 6.1 (s,1 H); 4.65(m,1 H); 4.45(t,1 H); 3.9(t,2H); 3.8-3.4(m,5H); 3.3(m,1 H); 2.95(m,1 H); 2.7(m,1 H); 2.4-1.9(m,5H); 1.8(q,2H); 1.15(d,3H); 1.0(t,3H); 1.2-0.9(m,4H).

Example 7 2S-(4-Cyclopropanecarbonyl-6S-methyl-5-oxo-hexahydro-(3aS,6aR)-pyrrolo[3,2- b]pyrrole-1 -carbonyl)-pyrrolidine-1 -carboxylic acid (4-trifluoromethyl-phenyl)- amide

Mass spec:[MH]* = 493 'H nmr(CDCI₃): δ 7.5(m,4H); 6.4(s,1 H); 4.65(m,1 H); 4.4(t,1 H); 3.9-3.5(m,5H); 3.3(m,1H); 2.95(m,1 H); 2.7(m,1H); 2.4-1.9(m,5H); 1.15(d,3H); 1.2-0.9(m,4H).

Example 8

(3S,3aR,6aS)-1 -Cyclopropanecarbonyl-3-methyl-4-[1 -(4-phenoxy- benzenesulfonyl)-pyrrolidine-2S-carbonyl]-hexahydro-pyrrolo[3,2-b]pyrrol-2-one

To a solution of the Intermediate 14 (30.7mg,89.8μmol,1eq) in dry MeCN (2mL) was added triethylamine (29μL,207μmol,2.3 eq). To this solution was added a solution of 4-phenoxybenzenesulfonylchloride³ (28.2mg,105μmol,1.2 eq) in dry MeCN (1mL). The reaction mixture was stirred at room temperature for 3 hours. Isopropanol (10μL) was added to quench unreacted sulfonyl chloride. The mixture was evaporated to dryness, re-dissolved in dichloromethane (15mL) and washed with water (15mL), sat. NaHCO₃ solution (15mL) and water (15mL). After drying over MgSO₄ the solvent was evaporated in vacuo to give a white solid. This residue was purified by flash column chromatography over silica gel (Merck 9385) eluted with cyclohexane:ethyl acetate to give the title compound as a white solid (33.9mg).

¹H nmr (CDCI₃) : δ 7.96-7.78 (2H,m), 7.50-7.33 (2H,m), 7.32-7.13 (1 H,m), 7.14- 6.98 (4H,m), 4.62-4.45 (1 H,m), 4.27 (1 H,t), 3.91-3.62 (2H,m), 3.60-3.23 (4H,m), 3.00-2.86 (1 H,m), 2.78 (1 H,m), 2.24-1.74 (5H,m), 1.22-0.78 (7H, m) ppm. Mass Spec: 538 [MH]* Circular dichroism λ_max196.6 nm, dE 0.65; λ_max215.4 nm, dE -22.30; λ_max237.8 nm, dE 9.70; ; λ_max254.2 nm, dE -2.08;

Ref 3 : CM. Suter, JACS, 1931, 53,1112-1116 and C.S Marvel, P.D. Caesar, JACS, 1951 , 73, 1097-1099.

Example 9 1-[2S-(4-Cyclopropanecarbonyl-6S-methyl-5-oxo-hexahydro-(3aS,6aR)- pyrrolo[3,2-b]pyrrole-1-carbonyl)-pyrrolidin-1-yl]-2-(4-isopropyl-phenyl)-ethane- 1 ,2-dione

To a solution of p-isopropylphenylglyoxylic acid⁴ (22 mg,114μmol,1.2 eq) in DMF (1mL), stirred at room temperature, was added a solution of TBTU (34.5 mg,107μmol,1.2eq) in MeCN (0.5mL) and a solution of HOBT (15 mg,112μmol,1.2eq) in DMF (0.5mL). After 20 min. the Intermediate 14 (31.9mg, 93μmol,1eq) in DMF (0.5mL) and diisopropylethylamine (33μL,187μmol,2 eq) were added. The reaction mixture was allowed to stir at room temperature for 24 hours and was then diluted with dichloromethane (15 mL) and water (15mL). The aqueous layer was re-extracted with dichloromethane (15mL) and the combined organic extracts were then washed with 2N HCI (15mL), water (15mL), sat. NaHCO₃ solution (15mL) and water (15mL). After drying over MgSO₄ the solvent was evaporated to give an orange oil. This residue was purified by flash column chromatography over silica gel (Merck 9385) and eluted with cyclohexane:ethyl acetate to give the title compound as an off-white foam (26mg). ¹H nmr (CDCI₃) : δ 8.09-7.90 (2H,m), 7.43-7.30 (2H,m), 4.85-4.60 (1 H,m), 4.47

(1 H,t), 4.07-3.30 (6H,m), 3.14-2.65 (3H,m), 2.40-1.85 (5H,m), 1.40-0.68 (13H,m) ppm. Mass Spec. : 480 [MH]*

Circular dichroism λ_max204.4 nm, dE -2.38; λ_max217.8 nm, dE -15.40; λ_max238.2 nm, dE 16.90;

Ref 4 : E.T. Stiller, P.A. Diassi, D. Gerschutz, D.Meikle, J. Moetz, P.A. Principe, S.D. Levine, J.Med.Chem., 1972, 15, 10, 1029-1032.

Example 10

(3S,3aR,6aS)-1-Cyclopropanecarbonyl-3-methyl-4-[1-(4-phenoxy-benzoyl)- pyrrolidine-2S-carbonyl]-hexahydro-pyrrolo[3,2-b]pyrrol-2-one

To a solution of 4-phenoxybenzoic acid(0.025g, 0.117mmol) in dry dimethylformamide (1 ml) was added a solution of HOBT(0.013g, 0.099mmol) in DMF(1ml) and TBTU(0.035g, 0.11 mmol) in acetonitrile(1 ml). After stirring at room temperature for 20mins a solution of Intermediate 14 (0.03g,0.09mmol) in dry DMF was added followed by diisopropylethylamine(0.035ml,0.2mmol). After stirring at room temperature overnight the reaction was incomplete therefore more 4-phenoxybenzoic acid(0.012g,0.056mmol), HOBT(0.006g,0.045mmol) and TBTU(0.018g,0.055mmol) were added. The reaction was complete after 5hours at room temperature. The mixture was diluted with dichloromethane(IOml) and water(IOml), the organic and aqueous layers were separated and the latter extracted with dichloromethane(2x10ml), the combined organic extracts washed with 2N HCI(10ml),water(10ml), saturated NaHCO₃ solution(IOml), again with water(10ml),dried over MgSO₄, flltered and evaporated to dryness. The residue was purified by preparative plate chromatography using (WhatmanPKβF silica gel 60A plate) eluting with ethyl acetate/cyclohexane(1 :1 ) to give the pure title compound as an amorphous solid. Mass spec:[MH]* = 502 'H nmr(CDCI₃): δ 7.6(d,2H);7.45(m,2H); 7.2(m,1 H); 7.0(m,4H); 4.7(t,1 H); 4.6(t,1 H); 3.95-3.5(m,5H); 3.35(m,1 H); 2.95(m,1 H); 2.7(m,1 H); 2.4- 1.8(m,5H);1.15(d,3H); 1.3-0.9(m,4H). Circular dichroism λ_max214.6 nm, dE -21.70; λ_max238.6 nm, dE 22.30.

Example 11 2S-[4-(cis-2,3-Dimethyl-cyclopropanecarbonyl)-6S-methyl-5-oxo-hexahydro- (3aS,6aR)-pyrrolo[3,2-b]pyrrole-1 -carbonyl]-pyrrolidine-1 -carboxylic acid (4- trifluoromethoxy-phenyl)-amide

The Intermediate 18 (46mg, 0.12mmol) was dissolved in acetonitrile (1 mL) and 4-(trifluoromethoxy)phenyl isocyanate (28.15μL, 0.19mmol, 1.5eq) added, followed by triethylamine (43.4μL, 0.31 mmol, 2.5eq). The mixture was left to stand at room temperature overnight (20hr 40min) before it was evaporated and purified using preparative plate chromatography (WhatmanPK6F silica gel 60A plate) eluting with diethyl ether to give the title compound as a white solid, 34mg. H nmr (CDCI₃): δ 7.36 (2H, d); 7.12 (2H, d); 6.32 (1 H, bs); 4.63 (1 H, dd); 4.40 (1 H, t); 3.78 (1 H, m); 3.69 (2H, m); 3.54 (2H, m); 3.26 (1 H, m); 2.84 (1 H, t); 2.77 (1 H, m); 2.28 (2H, m); 2.03 (3H, m); 1.60 (2H, m); 1.19 (9H, m). Mass spec 537 [MH]*, 334 [M-4-(trifluoromethoxy)phenyl isocyanate group]*.

Circular dichroism λ_max208.0 nm, dE -8.90; λ_max221.4 nm, dE -20.9; λ_max242.0 nm, dE 23.40.

Example 12 2S-[4-(cis-2,3-Dimethyl-cyclopropanecarbonyl)-6S-methyl-5-oxo-hexahydro-

(3aS,6aR)-pyrrolo[3,2-b]pyrrole-1 -carbonyl]-pyrrolidine-1 -carboxylic acid (4- isopropyl-phenyl)-amide

The above example 12 was prepared in a similar manner to Example 11 from intermediate 18 and 4-(isopropyl)phenyl isocyanate give the title compound as a white solid.

¹H nmr (CDCI₃): δ 7.24 (2H, d); 7.12 (2H, d); 6.20 (1 H, bs); 4.63 (1 H, dd); 4.45

(1 H, t); 3.77 (1 H, m); 3.66 (2H, m); 3.52 (2H, m); 3.25 (1 H, m); 2.84 (2H, m);

2.75 (1 H, m); 2.27 (2H, m); 2.00 (3H, m); 1.60 (2H, m); 1.20 (15H, m). Mass spec 495 [MH]*, 334 [M-4-(isopropyl)phenyl isocyanate group]*.

Example 13

2S-(4-Benzothiazol-2-yl-6S-methyl-5-oxo-hexahydro-(3aS,6aR)-pyrrolo[3,2- b]pyrrole-1 -carbonyl)-pyrrolidine-1 -carboxylic acid (4-trifluoromethoxy-phenyl)- amide The Intermediate 24 (20.7mg, 44μmol) was treated as Example 14, but 4- (trifluoromethoxy)phenyl isocyanate (10.1 μL, 66.7μmol, 1.5eq) was used and the crude mixture was purified using a Bond Elute^® silica column eluting with ethyl acetate / cyclohexane 1 :1 to give the title compound as a white solid, 22.5mg.

¹H nmr (CDCI₃): δ 7.81 (2H, m); 7.43 (1 H, td); 7.38 (2H, m); 7.31 (1 H, t); 7.13 (2H, d); 6.34 (1H, s); 4.68 (1 H, m); 4.56 (1H, t); 4.13 (1 H, m); 3.93 (1H, m); 3.80 (1 H, dd); 3.70 (1 H, m); 3.54 (1 H, q); 3.44 (1 H, m); 3.15 (1 H, m); 2.40 (2H, m); 2.24 (1 H, m); 2.12 (1 H, m); 2.00 (1 H, m); 1.20 (3H, d). Mass spec 574 [MH]*, 371 [M-4-(trifluoromethoxy)phenyl isocyanate group]*.

Example 14

2S-(4-Benzothiazol-2-yl-6S-methyl-5-oxo-hexahydro-(3aS,6aR)-pyrrolo[3,2- b]pyrrole-1 -carbonyl)-pyrrolidine-1 -carboxylic acid (4-isopropyl-phenyl)-amide

To the Intermediate 24, (64mg, 136μmol) was added trifluoroacetic acid (315μL, 4.09mmol, 30eq) at room temperature. After 10 minutes the solution was azeotroped with toluene (1mL x 2) to leave a yellow gum (78mg). The gum was dissolved in acetonitrile (2mL) and 4-(isopropyl)phenyl isocyanate (30μL, 188μmol, 1.37eq) added, followed by triethytamine (47.5μL, 341μmol, 2.5eq). The mixture was left to stand at room temperature for 4 hours before it was directly purified using preparative plate chromatography (WhatmanPK6F silica gel 60A plate) eluting with ethyl acetate to give the title compound as an off- white solid, 62mg.

Circular dichroism λ_max198.0 nm, dE -11.0; λ_max212.4 nm, dE 8.47; λ_max228.0 nm, dE -4.01 ; λ_max239.0 nm, dE 2.94; λ_max248.2 nm, dE 0.47; λ_max257.2 nm, dE

4.78.

¹H nmr (CDCI₃): δ 7.80 (2H, m); 7.44 (1 H, td); 7.31 (1 H, td); 7.25 (2H, m); 7.14 (2H, m); 6.23 (1 H, s); 4.70 (1 H, m); 4.62 (1 H, t); 4.13 (1 H, td); 3.93 (1 H, m); 3.80 (1 H, dd); 3.70 (1 H, m); 3.54 (1H, q); 3.45 (1H, m); 3.14 (1H, m); 2.86 (1 H, m); 2.39 (2H, m); 2.25 (1 H, m); 2.11 (1 H, m); 2.00 (1 H, m); 1.21 (9H, d). Mass spec 532 [MH]*, 371 [M-4-(isopropyl)phenyl isocyanate group]*.

Example 15

2S-(4-Benzothiazol-2-yl-6S-methyl-5-oxo-hexahydro-(3aS,6aR)-pyrrolo[3,2- b]pyrrole-1 -carbonyl)-azetidine-1 -carboxylic acid (4-isopropyl-phenyl)-amide. To Intermediate 30 (67.4 mg, 31.1 mmol) in dry dichloromethane (2mL) was added trifluoroacetic acid (0.5ml). The solution was stirred at room temperature for 0.5 hours. The solvents were then evaporated and the resulting brown gum was azeotroped with toluene (X3). The product was obtained in quantitative yield as a brown foam, which was used in the next step without further purification. The brown foam (83mg,148 mmol of trans-lactam + 0.12mmol TFA) was dissolved in acetonitrile (3mL). Triethylamine (61 μL, 443μmol, 3eq) and 4- (isopropyl)phenyl isocyanate (38μL, 238μmol, 1.6eq) were added to the solution and the reaction mixture was stirred at room temperature for 4 hours. The reaction mixture was quenched (drop of formic acid) and the solvent was evaporated in vacuo. The residue was taken up in CH₂CI₂ (5mL) and washed with water (5mL) and brine (5mL). After drying (MgSO₄), the solvent was evaporated to give a pale yellow residue. This residue was purified by Bond Elute^® silica cartridge eluting with ethyl acetate/cyclohexane(1 :1 ) to give the title compound as a white solid, (58.1 mg). Mass Spec. : 357, 518 [MH]⁺.

HPLC (Intersil M column ODS2, eluent system A (H₂O, 0.1 % H₃PO₄) B (95% MeCN/H₂O, 0.1% H₃PO₄):Gradient 0%B 2mins, 0%-100% B 40mins,100% B 10mins; flow rate = 1 ml/min, , λ = 215nm). Retention time =32 min., 96%.

¹H nmr (CDCI₃) : δ 7.81 (2H,t), 7.69 (1 H, br.s), 7.45 (1 H,t), 7.38-7.23 (3H,m), 7.12 (2H,d), 4.88 (1 H,t), 4.23-4.01 (3H,m), 3.97-3.73 (3H,m), 3.62-3.45 (1 H,m), 3.26-3.12 (1 H,m), 2.92-2.76 (1 H,m), 2.62-2.27(3H,m), 1.35-1.13 (9H,m) ppm.

Example 16

2S-[4-(cis-2,3-Dimethyl-cyclopropanecarbonyl)-6S-methyl-5-oxo-hexahydro- (3aS,6aR)-pyrrolo[3,2-b3pyrrole-1-carbonyl]-azetidine-1 -carboxylic acid (4- isopropyl-phenyl)-amide

Example 16 was prepared in a similar manner to Example 15 from Intermediate

31.

LCMS: 481 [MH]⁺, Retention time = 3.36 min, using the same conditions as in

Intermediate 27

¹H nmr (CDCI₃) : δ 7.78 (1 H,br.s), 7.29 (2H, d), 7.12 (2H,d), 4.84 (1 H,t), 4.15- 3.26 (7H,m), 2.93-2.74 (3H,m), 2.59-2.40 (2H,m), 2.12-1.92 (1 H,m), 1.75-1.50

(2H,m),1.29-1.10 (15H,m) ppm. Example 17

2S-[4-(6-Hydroxymethyl-benzothiazol-2-yl)-6S-methyl-5-oxo-hexahydro- (3aS,6aR)-pyrrolo[3,2-b]pyrrole-1 -carbonyl]-pyrrolidine-1 -carboxylic acid (4- isopropyl-phenyl)-amide

To a solution of trans-lactam Intermediate 35 (0.125g,0.169 mmol) in dry 1 ,4- dioxane (1mL) was added a 4.0M solution of HCI in 1 ,4-dioxane (0.5 mL). The reaction mixture was allowed to stir at room temperature under an atmosphere of nitrogen for ca. 68 hours. The reaction mixture was then evaporated to dryness and azeotroped with toluene (X3) to give a cream residue which was used in the next step without further purification. The crude mixture was treated in a similar manner to Intermediate 18 in Example 11 to give the title compound as an off-white solid. LCMS: 562 [MH]⁺, Retention time = 3.16 min, using the same conditions as in Intermediate 27.

¹H nmr (CDCI₃) : δ 7.85-7.66 (2H,m), 7.42 (1 H,dd), 7.33-7.19 (2H,m), 7.19-7.10 (2H,d), 6.23 (1 H, s), 4.78 (2H,s), 4.72-4.52 (2H,m), 4.18-3.00 (7H,m), 2.92-2.77 (1 H,m), 2.45-1.90 (6H,m), 1.37-1.05 (9H,m) ppm.

Example 18

2S-[4-(4-Methoxy-benzothiazol-2-yl)-6S-methyl-5-oxo-hexahydro-(3aS,6aR)- pyrrolo[3,2-b]pyrrole-1 -carbonyl]-pyrrolidine-1 -carboxylic acid (4-isopropyl- phenyl)-amide

To the Intermediate 37 (29mg, 57μmol)was added trifluoroacetic acid (240μL, 3.12mmol, 53eq) at room temperature. After 11 minutes the solution was azeotroped with toluene (1mL x 2) to leave a yellow gum (35mg). The gum was dissolved in acetonitrile (1 mL) and 4-(isopropyl)phenyl isocyanate (10.8μL, 67μmol, 1.18eq) added, followed by triethylamine (20.2μL, 145μmol, 2.5eq). The mixture was left to stand at room temperature for 17 hours before it was evaporated to leave a yellow gum. Purification using a Bond Elute^® silica column eluting with diethyl ether gave the title compound as a white solid, 29.4mg. ¹H nmr (CDCI₃): δ 7.42 (1 H, d); 7.25 (3H, m); 7.13 (2H, d); 6.90 (1 H, d); 6.20 (1 H, s); 4.68 (1H, m); 4.59 (1 H, t); 4.20 (1H, td); 4.02 (3H, s); 3.80 (3H, m); 3.49 (2H, m); 3.16 (1 H, m); 2.85 (1 H, m); 2.38 (2H, m); 2.25 (1 H, m); 2.08 (2H, m); 1.20 (9H, m). Mass spec 562 [MH]*, 401 [M-4-(isopropyl)phenyl isocyanate group]*. Example 19

2S-[4-(5-Chloro-benzothiazol-2-yl)-6S-methyl-5-oxo-hexahydro-(3aS,6aR)- pyrrolo[3,2-b]pyrrole-1 -carbonyl]-pyrrolidine-1 -carboxylic acid (4-isopropyl- phenyl)-amide

Prepared in a similar manner to Example18, .from Intermediate 39 to give the title compound as a white solid.

¹H nmr (CDCI₃): δ 7.78 (1 H, d); 7.71 (1 H, d); 7.27 (3H, m); 7.02 (2H, d); 6.33 (1H, s); 4.67 (1H, m); 4.59 (1H, t); 4.10 (1H, m); 3.91 (1H, m); 3.74 (1 H, dd); 3.67(1 H, m); 3.52 (1 H, q); 3.43 (1 H, m); 3.07 (1 H, m); 2.84 (1 H, m); 2.36 (2H, m); 2.27 (1 H, m); 2.03 (2H, m); 1.19 (9H, d). Mass spec 566 [MH]*, 405 [M-4-(isopropyl)phenyl isocyanate group]^*.

Example 20

2S-[4-(5-Methoxy-thiazolo[5,4-b]pyridin-2-yl)-6S-methyl-5-oxo-hexahydro-

(3aS,6aR)-pyrrolo[3,2-b]pyrrole-1 -carbonyl]-pyrrolidine-1 -carboxylic acid (4- isopropyl-phenyl)-amide.

Prepared in a similar manner to Example18 from Intermediate 41 to give the title compound.

Mass spec:[MH]* = 563

Η nmr(CDCI₃): δ 7.9(d,1 H); 7.25(d, 2H); 7.1 (d, 2H); 6.8(d, 1 H); 6.2(s, 1 H); 4.7-

4.5(m, 2H); 4.2-3.3(m, 6H); 4.0(s, 3H); 3.1(m,1 H); 2.8(m, 1 H); 2.5-1.9(m, 5H);

1.2(d,9H).

Example 21

2S-[4-(Bicyclo[4.1.0]hept-3-ene-7-carbonyl)-6S-methyl-5-oxo-hexahydro- (3aS,6aR)-pyrrolo[3,2-b]pyrrole-1 -carbonyl]-pyrrolidine-1 -carboxylic acid (4- isopropyl-phenyl)-amide

A solution of the Intermediate 51 ( 162 mg, 0.35mmol ) in anhydrous dichloromethane ( 3 mL ) and trifluoroacetic acid ( 1 mL ) was stirred at 21 ° for 45 min. The solution was evaporated to dryness, toluene ( 2x 5 mL ) was added and evaporated off and the residue ( 216 mg ) was dissolved in anhydrous acetonitrile (4 mL ). Triethylamine (100 mL, 0.72mmol ) and 4- isopropylphenylisocyanate ( 80mL, 0.5 mmol ) were added and the solution was stirred at 21 ° for 16h. It was poured into ethyl acetate (25 ml) and the solution was washed with water (10 ml) , 2N hydrochloric acid (10 ml), water (10 ml), and saturated brine (10 ml) , dried over magnesium sulphate and evaporated. The product was purified on two 2g Bond Elute^® silica columns eluting with dichloromethane, chloroform, 1 :1 cyclohexane : ether, 1 :1 cyclohexane : ethyl acetate (3x) and ethyl acetate (2x) . Evaporation of the second and third cyclohexane : ethyl acetate and first ethyl acetate fractions gave the title compound ( 104 mg, 57% ) as a pale cream solid.

LCMS: 519 [MH]⁺, Retention time = 5.17 min., using the same conditions as in Intermediate 30 HPLC (Intersil M column ODS2, eluent system A (H₂O, 0.1% H₃PO₄) B (95%

MeCN/H₂O, 0.1 % H₃PO₄):Gradient 0%B 2mins, 0%-100% B 40mins,100% B 10mins; flow rate = 1ml/min, , λ = 215nm). Retention time =30.3 min 95.8%. ¹H nmr (CDCI₃): δ 7.28 (2H, d); 7.12 (2H, d); 6,17 (1 H, s); 5.53 (2H, s); 4.64 (1 H, dd); 4.43 (1 H, t); 3.82 - 3.62 (4H, m); 3.58 - 3.46 (2H, m); 2.85 (1 H, dt); 2.72 (1 H, dt), 2.5 - 1.8 ( m); 1.7 - 1.5 ( m); 1.21 (6H, d); 1.13 (3H, d).

Example 22 rel-2S-[4-(4-Chloro-benzothiazol-2-yl)-6S-methyl-5-oxo-hexahydro-(3aS,6aR)- pyrrolo[3,2-b]pyrrole-1-carbonyl]-pyrrolidine-1 -carboxylic acid (4-isopropyl- phenyl)-amide

Prepared in a similar manner to Example18, from Intermediate 44 to give the title compound as a pale yellow solid.

¹H nmr (CDCI₃): δ 7.70 (1 H, dd) and 7.45 (1 H, dd); 7.25 (3H, m) and 7.13 (2H, d); 6.21 (1 H, s); 4.69 (1 H, dd); 4.59 (1 H, t); 4.16 (1 H, m); 3.94 (1 H, dm); 3.80 (1 H, dd); 3.69 (1 H, m); 3.53 (1 H, dd); 3.45 (1 H, m); 3.19 (1 H, m); 2.85 (1 H, m);

2.51-2.19 (3H, m); 2.19-1.86 (2H, m); 1.22 (9H, m).

Mass spec 566 [MH]*, 407 [M - 4-(isopropyl)phenyl isocyanate group]*.

Example 23 2S-[6S-Methyl-4-(6-nitro-benzothiazol-2-yl)-5-oxo-hexahydro-(3aS,6aR)- pyrrolo[3,2-b] pyrrole- 1 -carbonyl]-pyrrolidine-1 -carboxylic acid (4-isopropyl- phenyl)-amide

Prepared in a similar manner to Example18, from Intermediate 46 to give the title compound as a yellow solid. ¹H nmr (CDCI₃): δ 8.74 (1 H, d); 8.31 (1 H, dd); 7.83 (1 H, d); 7.24 (2H, d) and 7.13 (2H, d); 6.28 (1 H, s); 4.65 (2H, m); 4.13 (1 H, m); 3.94 (1 H, dm); 3.79 (1 H, dd); 3.69 (1H, m); 3.53 (1H, m); 3.47 (1H, m); 3.11 <1H, m); 2.84 (1H, m); 2.49-2.18 (3H, m); 2.18-1.92 (2H, m); 1.20 (9H, m). Mass spec 577 [MH]*, 416 [M - 4-(isopropyl)phenyl isocyanate group]*.

Example 24

2S-[4-(6-Fluoro-benzothiazol-2-yl)-6S-methyl-5-oxo-hexahydro-(3aS,6aR)- pyrrolo[3,2-b]pyrrole-1-carbonyl]-pyrrolidine-1 -carboxylic acid (4-isopropyl- phenyl)-amide

Prepared in a similar manner to Example18, from Intermediate 48 to give the title compound as a white solid.

¹H nmr (CDCI₃): δ 7.72 (1 H, dd); 7.49 (1 H, dd; 7.24 (2H, d) and 7.15 (3H, m);

6.23 (1 H, s); 4.68 (1 H, dd); 4.61 (1 H, t); 4.10 (1 H, td); 3.91 (1 H, m); 3.77 (1 H, dd); 3.69 (1 H, m); 3.52 (1 H, dd); 3.43 (1 H, m); 3.09 (1 H, m); 2.84 (1 H, m); 2.47-

2.18 (3H, m; 2.18-1.92 (2H, m); 1.20 (9H, d).

Mass spec 550 [MH]*, 389 [M - 4-(isopropyl)phenyl isocyanate group]*.

Example 25

2S-[4-(5-Methoxy-benzothiazol-2-yl)-6S-methyl-5-oxo-hexahydro-(3aS,6aR)- pyrrolo[3,2-b]pyrrole-1 -carbonyl]-pyrrolidine-1 -carboxylic acid (4-isopropyl- phenyl)-amide

Prepared in a similar manner to Example18, from Intermediate 43 to give the title compound as a cream solid.

¹H nmr (CDCI₃): δ 7.66 (1 H, d); 7.31 (1 H, d); 7.25 (2H, d) and 7.13 (2H, d); 6.95

(1 H, dd); 6.27 (1 H, s); 4.68 (1 H, dd); 4.60 (1 H, t); 4.09 (1 H, m); 3.88 (3H, s);

3.94 (1 H, m); 3.76 (1 H, dd); 3.68 (1 H, dm); 3.52 (1 H, dd); 3.43 (1 H, m); 3.11 (1 H, m); 2.84 (1 H, m); 2.46-2.19 (3H, m); 2.18-1.92 (2H, m); 1.20 (9H, m).

Mass spec 562 [MH]*, 401 [M - 4-(isopropyl)phenyl isocyanate group]*.

Example 26 2S-[4-(4-Methoxy-7-methyl-benzothiazol-2-yl)-6S-methyl-5-oxo-hexahydro- (3aS,6aR)-pyrrolo[3,2-b]pyrrole-1 -carbonyl]-pyrrolidine-1 -carboxylic acid (4- isopropyl-phenyl)-amide

Prepared in a similar manner to Example18, from Intermediate 50 to give the title compound as a cream solid.

¹H nmr (CDCI₃): δ 7.25 (2H, d) and 7.13 (2H, d); 7.04 (1 H, d); 6.82 (1 H, d); 6.22 (1 H, s); 4.68 (1 H, dd); 4.58 (1 H, t); 4.19 (1 H, td); 4.00 (3H, s); 3.89 (1 H, dm); 3.78 (1 H, dd); 3.68 (1 H, m); 3.52 (1 H, q); 3.44 (1 H, m); 3.15 (1 H, m); 2.84 (1 H, m); 2.47 (3H, s); 2.46-2.19 (3H, m); 2.19-1.91 (2H, m); 1.20 (9H, m). Mass spec 576 [MH]*, 415 [M - 4-(isopropyl)phenyl isocyanate group]*.

Example 27

(3S,3aR,6aS)-1 -Cyclopropanecarbonyl-3-methyl-4-[1 -(4-propoxy- benzenesulfonyl)-pyrrolidine-2S-carbonyl]-hexahydro-pyrrolo[3,2-b]pyrrol-2-one

Similarly prepared to Example 8 from Intermediate 14, and 4-n-propoxysulfonyl chloride ⁵ to give the title compound, as a beige solid.

HPLC: Retention time =28.86 min. 100%, using the same conditions as in

Example 21. ¹H nmr (CDCI₃) : δ 7.80 (2H,d), 6.97(2H,d), 4.52(1 H, dd,), 4.28 (1 H,t), 3.98

(2H,t), 3.84 (1 H,m), 3.72 (1 H,m), 3.55 (1 H,m), 3.44-3.28 (3H,m), 2.95 (1 H,m),

2.78 (1H,m), 2.16-2.02 (3H,m), 1.92 (1H,m), 1.82 (2H,m),1.78 (1H,m), 1.18

(3H,d), 1.04 (3H,t), 1.20-0.96 (4H,m).

Mass Spec. 438 [MH]*.

Ref 5. R. V. Hoffman.Org. Syn, (Vol.60), 121-126.

Example 28

(3S,3aR,6aS)-4-[1-(4-tert-Butyl-benzoyl)-pyrrolidine-2S-carbonyl]-1- cyclopropanecarbonyl-3-methyl-hexahydro-pyrrolo[3,2-b]pyrrol-2-one

Similarly prepared to Example 1 from Intermediate 14, and 4-tert-butylbenzoyl chloride to give the title compound, as a off-white solid.

HPLC: Retention time = 28.087min. 100%, using the same conditions as in

Example 21. Mass Spec. 466 [MH]*. Η nmr (CDCI₃): δ 7.55 (2H, d); 7.40 (2H, d); 4.7(1 H, t); 4.6 (1 H, t); 3.9-3.6 (5H, m); 3.35(1 H, m); 2.95(1 H, m); 2.75 (1 H, m); 2.4-1.8 (5H, m); 1.3(9H, s); 1.2 (3H, d); 1.3-0.9 (4H, m).

Example 29

(3S,3aR,6aS)-4-[1-(5-Dimethylamino-naphthalene-1-sulfonyl)-pyrrolidine-2S- carbonyl]-3-methyl-1-thiazol-2-yl-hexahydro-pyrrolo[3,2-b]pyrrol-2-one

Prepared in a similar manner to Example18 by reacting 2-bromothiazole, with Intermediate 29 to give the title compound as a yellow solid.

¹H nmr (CDCI₃): δ 8.56 (1 H, d), 8.44 (1 H, d), 8.31 (1 H, d), 7.56 (2H, m) and 7.19

(1 H, d); 7.45 (1 H, d) and 7.02 (1 H, d); 4.78 (1 H, dd); 4.37 (1 H, t); 4.02 (1 H, m);

3.91 (H, dm); 3.57 (2H, m); 3.46 (1 H, m); 3.37 (1 H, m); 3.01 (1 H, m); 2.88 (6H, s); 2.37- 2.09 (3H, m); 2.07- 1.84 (2H, m); 1.14 (3H, d). Mass spec 554 [MH]*.

Example 30

(3S,3aR,6aS)-4-[1-(5-Dimethylamino-naphthalene-1-sulfonyl)-pyrrolidine-2S- carbonyl]-3-methyl-2-oxo-hexahydro-pyrrolo[3,2-b]pyrrole-1 -carboxylic acid (4- acetyl-phenyl)-amide

Prepared in a similar manner to Example31 by reacting Intermediate 29 with 4- acetylphenyl isocyanate to give the title compound .

¹H nmr (CDCI₃): δ 10.18 ( br), 8.54 (1 H, d), 8.42 (1 H, d), 8.28 (1 H, d), 7.98 -7.9 (3H, m), 7.63 -7.45 (5H, m), 7.19 (1 H, d), 6.65 (1 H, d), 4.76 (1 H, dd), 4.31 (1 H, t), 4.10 (1 H, m), 3.95 -3.76 (2H, m), 3.6 -3.4 (4H, m), 2.88 (6H, s), 2.57(3H, 2s),

2.4 -1.8 (7H, m), 1.18 (3H, dd)

LCMS: 632.1 [MH]⁺, Retention time = 4.83 min., using the same conditions as in Intermediate 30 HPLC: Retention time = 28.5 min, 89.5 %, using the same conditions as in

Example 21.

Example 31 (3S,3aR,6aS)-4-[1-(5-Dimethylamino-naphthalene-1-sulfonyl)-pyrrolidine-2S- carbonyl]-3-methyl-2-oxo-hexahydro-pyrrolo[3,2-b]pyrrole-1 -carboxylic acid benzylamide

Intermediate 29 (38.4mg, O.Oδmmol) was dissolved in tetrahydrofuran (3.5mL, anhydrous) at room temperature under nitrogen. Sodium hydride (1.8mg, 0.04mmol, 0.55eq, 60% dispersion in mineral oil) was added to the stirred solution, followed after 20 minutes by benzyl isocyanate (25.25μL, 0.2mmol, 2.5eq). After the mixture had been stirred for 1 hour 25 minutes saturated aqueous ammonium chloride (4mL) was added. The mixture was diluted with water (2mL) and extracted with ethyl acetate (2 x 4mL). The combined organic phase was washed with saturated brine (4mL), dried (MgSO₄) and evaporated to leave a bright yellow / green gum (64mg). Purification by preparative plate chromatography, eluted and extracted with ethyl acetate, gave the title compound as a bright yellow / green foam, 42.5mg (86%).

¹H nmr (CDCI₃): δ 8.54 (1 H, d), 8.42 (1 H, d), 8.29 (1 H, d), 7.54 (2H, m) and 7.18 (1 H, d); 8.22 (1 H, t); 7.33 (5H, m); 4.73 (1 H, m); 4.49 (2H, d); 4.27 (1 H, t); 3.88- 3.66 (2H, m); 3.63- 3.40 (2H, m); 3.38 (1 H, dd); 3.23 (1 H, m); 2.87 (6H, s); 2.78 (1 H, m) 6-H; 2.34- 2.06 (3H, m); 2.05- 1.80 (2H, m); 1.11 (3H, d). Mass spec 604 [MH]*, 471 [M - benzyl isocyanate group]*. HPLC: Retention time = 29.03 min, 100%, using the same conditions as in Example 21.

Example 32

2S-(4-Cyclopropanecarbonyl-6S-methyl-5-oxo-hexahydro-(3aS,6aR)-pyrrolo[3,2- b]pyrrole-1 -carbonyl)-pyrrolidine-1 -carboxylic acid (4-phenoxy-phenyl)-amide

Similarly prepared to Example 2 from Intermediate 14, and 4-phenoxyphenyl isocyanate to give the title compound, as a off-white solid.

HPLC: Retention time = 27.751 min. 92%, using the same conditions as in

Example 21.

Mass Spec. 517 [MH]*. 'H nmr(CDCI₃): δ 7.35 (6H, m); 7.05 (1 H, t); 6.95(2H, d); 6.2 (1 H, s) 4.65 (1 H, m); 4.45(1 H, t); 3.95-3.5 (5H, m); 3.3(1 H, t); 2.95(1 H, m); 2.7(1 H, m); 2.4-1.9 (5H, m); 1.2 (3H, d); 1.3-1.0 (4H, m).

Example 33

2S-(6S-Methyl-5-oxo-4-thiazol-2-yl-hexahydro-(3aS,6aR)-pyrrolo[3,2-b]pyrrole-1- carbonyl)-pyrrolidine-1 -carboxylic acid (4-isopropyl-phenyl)-amide

To the intermediate 55 (15 mg, 35mmol) in anhydrous acetonitrile (1 mL) was added with stirring 4-/sopropylphenyl isocyanate (6.6 mg, 1.2 eq.) followed by diisopropylethylamine (4.5 mg, 6.1 mL). The mixture was stirred at room temperature overnight then taken up in ethyl acetate and washed with water.

The organic phase was separated, dried over anhydrous magnesium sulphate and filtered. The filtrate was evaporated under reduced pressure to give a yellow gum which was purified by preparative plate chromatography (Whatman

PK6F silica gel 60A plate) eluting with 1 :1 ethyl acetate:cyclohexane to give the product as an off-white solid (16 mg, 96%).

¹H nmr (CDCl₃): δ 7.44 (1 H, d); 7.12 and 7.25 (4H, pseudo-ABq); 7.01 (1 H, d);

6.29 (1 H, s); 4.63-4.70 (1 H, m); 4.52-4.62 (1 H, m); 3.97-4.12 (1 H, m); 3.80-3.96 (1 H, m); 3.60-3.80 (2H, m); 3.45-3.55 (1 H, m); 3.35-3.45 (1 H, m), 2.92-3.03 (1 H, m); 2.79-2.92 (1 H, m); 1.95-2.40 (5H, m); 1.13-1.30 (9H, m).

Mass spectrum 482 [MH]*, 321 [MH-ArNCO]*

Retention time = 27.20 min, using the same conditions as in intermediate 27.

Example 34

2S-(6S-Methyl-5-oxo-4-thiazol-2-yl-hexahydro-(3aS,6aR)-pyrrolo[3,2-b]pyrrole-1- carbonyl)-pyrrolidine-1 -carboxylic acid (4-tert-butyl-phenyl)-amide

Similarly prepared to Example 33 from Intermediate 55, and 4-tert-butylphenyl isocyanate to give the title compound, as a off-white solid. ¹H nmr (CDCI₃): d 7.44 (1 H, d); 7.22-7.35 (4H, pseudo-ABq); 7.01 (1 H, d); 6.31 (1 H, s); 4.64-4.71 (1H, m); 4.52-4.64 (1 H, m); 3.97-4.15 (1H, m); 3.80-3.97 (1H, m); 3.62-3.80 (2H, m); 3.34-3.59 (2H, m), 2.91-3.03 (1 H, m); 1.92-2.42 (5H, m); 1.27 (9H,s); 1.17 (3H, d). Mass spectrum 496 [MH]*, 321 [MH-ArNCO]*.

Retention time = 28.55 min, using the same conditions as in intermediate 27.

Claims

Claims

A compound of the general formula (I)

(relative stereochemistry indicated) wherein:

R represents H, substituted or unsubstituted C_1-3 alkyl;

R-, represents optionally substituted heteroaryl or fused heteroaryl with one to four heteroatoms, R₅CO, R₅NHCO, R₅CS or R₅NHCS wherein R₅ may be substituted or unsubstituted and represents H, C^ alkyl, C,_₆ alkenyl, C₃-₇ cycloalkyl or fused cycloalkyl, heteroaryl or fused heteroaryl containing one to four heteroatoms, aryl or fused aryl, or arylC,_₃alkyl;

R₂ represents R₆-X- or R₃CO, wherein R₃ may be substituted or unsubstituted and represents

optionally including one or more further heteroatoms;

R₄ represents R₆-X-; R₆ is optionally substituted heterocyclic or fused heterocyclic with 1-4 heteroatoms, heteroaryl or fused heteroaryl with 1-4 hetero atoms, C₃_ ₁₀cycloalkyl or fused cycloalkyl, aryl or fused aryl; and

X represents a linker group chosen from C=O, NHC=O, C(=O)C=O, CH=CHCO, CH₂CO, CH₂ or SO₂;

and salts and solvates thereof.

2. A compound according to claim 1 having SRS absolute stereochemistry as shown in formula (la)

3. A compound according to claim 1 or claim 2 wherein R₆ represents

wherein the aryl rings are optionally further substituted with one or more heteroatoms, Y represents a hetero atom such as O, S or N, wherein N is optionally further substituted, W represents hydrogen or C_1-3 alkyl and Z represents hydrogen, halogen, C^╧ü alkyl, aryl, C,-₆ alkoxy, C_nH_2n+╬╣OC_mH_2m wherein n and m are independently selected integers 1-3, CF₃, O-haloC₁.₃alkyl, S-C-^alkyl, S-haloC-,-₃alkyl, O-aryl, C₂-₄ alkenyl or N(A)B wherein A and B are independently selected from H and C^ alkyl.

4. A compound according to any one of claims 1-3 wherein R represents methyl.

5. A compound according to any preceding claim wherein R₁ represents

^c<┬░> < in which A and B independently represent H or C_1- alkyl or A and B together form a saturated or unsaturated carbocyclic ring.

6. A compound according to claim 5 wherein A and B each represent methyl.

7. A compound according to any one of claims 1-4 wherein R₁ represents

8. A compound according to any preceding claim wherein R₃ represents

9. A compound according to any preceding claim wherein R₆-X- represents

wherein Z represents i-Pr, t-Bu, O-n-Pr, OCF₃, OCHF₂ , OPhenyl or SCF₃

10. The use of a compound of formula (XXII)

(XXII) wherein P₂ and P₃ are nitrogen protecting groups such as benzyloxycarbonyl (Cbz) or t-butyloxycarbonyl (Boc), in the synthesis of a compound according to claim 1.

11. A compound of formula (XXIII)

wherein R is a substituted or unsubstituted C^ alkyl group, or an N-protected derivative thereof.

12. The use of a compound according to claim 11 in the synthesis of a compound according to claim 1.

13. A compound of formula (L)

wherein R is C₁.₃alkyl, or a carboxylic acid ester thereof, e.g. a C^alkyl ester or an N-protected derivative thereof.

14. The use of a compound according to claim 13 in the synthesis of a compound according to claim 1.

15. A compound of formula (L) which has absolute SRS stereochemistry as shown:

16. A Compound according to any one of claims 1-8 or a physiologically acceptable salt or solvate thereof for use in human or veterinary medicine.

17. A Compound according to any one of claims 1 -9 or 16, or a physiologically acceptable salt or solvate thereof for use in the treatment or prophylaxis of a condition caused by a virus of the herpes family.

18. A Compound according to claim 17 wherein the virus of the herpes family is varicella zoster virus, herpes simplex virus 1 or 2, cytomegalovirus, Epstein Barr Virus, human herpes virus 6 (HHV 6), HHV 7 or HHV 8.

19. A Compound according to any one of claims 1-9, 15, 17 or 18, or a physiologically acceptable salt or solvate thereof, for use in the treatment or prophylaxis of a cardiovascular system disease.

20. A Compound according to claim 19, wherein the cardiovascular system disease is thrombosis, arteriosclerosis, restenosis, or recurrent narrowing or occlusion of a coronary valve or vessel.

21. Use of a compound of formula (I) or formula (la), or a physiologically acceptable salt or solvate thereof, in the manufacture of a medicament for the treatment of a condition caused a by herpes family virus infection.

22. A method for the treatment of a human or animal subject with a condition caused or mediated by a virus of the herpes family, which method comprises administering to said human or animal subject an effective amount of a compound of formula (I) or formula (la), or a physiologically acceptable salt or solvate thereof.

23. A method of prophylaxis or suppression of the symptoms of a condition caused or mediated by a virus of the herpes family, which method comprises administering to a human or animal subject an effective amount of a compound of formula (I) or formula (la), or a physiologically acceptable salt or solvate thereof.

24. A method according to claim 22 or 23 wherein the virus of the herpes family is varicella zoster virus, herpes simplex virus 1 or 2, cytomegalovirus, Epstein Barr Virus, human herpes virus 6 (HHV 6), HHV 7 or HHV 8.

25. A method according to any one of claims 22 to 24, wherein the condition is a cardiovascular system disease.

26. A method according to claim 25, wherein the cardiovascular system disease is thrombosis, arteriosclerosis, restenosis, or recurrent narrowing or occlusion of a coronary valve or vessel.

27. A pharmaceutical composition for use in therapy or prophylaxis, comprising a compound of formula (I) or formula (la), or a physiologically acceptable salt or solvate thereof, in admixture with one or more physiologically acceptable diluents or carriers.

28. A combination for administration together or separately, sequentially or simultaneously in separate or combined pharmaceutical formulations, said combination comprising a compound of formula (I) or formula (la), or a physiologically acceptable salt or solvate thereof, together with another therapeutically active agent.