WO2006085199A1 - Piperazine derivatives - Google Patents

Abstract

The present invention relates to a crystalline form of 5-{(1S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide, processes for the preparation thereof, compositions containing the crystalline form, and uses of the crystalline form. 5-{(1S)-2-[(2R)-4-Benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide is particularly suitable as a drug substance and inhibits the interaction of gp120 with CD4 and is therefore of use in the treatment of HIV, a retroviral infection genetically related to HIV, or AIDS.

Description

Piperazine Derivatives

The present invention relates to a crystalline form of 5-{(1S)-2-[(2R)-4-benzoyl-2- methyl-piperazin-i-yll-i-methyl^-oxo-ethoxyH-methoxy-pyridine-Σ-carboxylic acid methylamide, to pharmaceutical compositions thereof, and to processes for preparing such a crystalline form. The invention further relates to the use of the crystalline form and compositions thereof in the treatment of HIV, a retroviral infection genetically related to HIV, or AIDS.

The compound 5-{(1S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2- oxo-ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide, having the structural formula (I):

wherein Me is methyl, is useful in the treatment of HIV, such as HIV-1 , and genetically related retroviral infections (and the resulting acquired immune deficiency syndrome, AIDS).

If a compound is to be developed as a drug, it is important to provide a form of that compound (commonly known as a drug substance) which can be reliably prepared and purified on a large scale and which does not degrade on storage. Such characteristics are normally found in a drug substance which is crystalline and of high melting point; a high-melting point crystalline solid tends to be easy to purify by re-crystallisation and stable on storage.

A compound may exist in a continuum of solid states ranging from fully amorphous to fully crystalline. The term 'amorphous' refers to a state in which the material lacks long range order at the molecular level and, depending upon temperature, may exhibit the physical properties of a solid or a liquid. Typically such materials do not give distinctive X-ray diffraction patterns and, while exhibiting the properties of a solid, are more formally described as a liquid. Upon heating, a change from solid to liquid properties occurs which is characterised by a change of state, typically second order ('glass transition'). The term 'crystalline' refers to a solid phase in which the material has a regular ordered internal structure at the molecular level and gives a distinctive X-ray diffraction pattern with defined peaks. Such materials when heated sufficiently will also exhibit the properties of a liquid, but the change from solid to liquid is characterised by a phase change, typically first order ('melting point').

A compound may exist in unsolvated and solvated forms. The term 'solvate' is used herein to describe a molecular complex comprising the compound and one or more pharmaceutically acceptable solvent molecules, for example, ethanol. The term 'hydrate' is employed when said solvent is water.

A currently accepted classification system for organic hydrates is one that defines isolated site, channel, or metal-ion coordinated hydrates - see Polymorphism in Pharmaceutical Solids by K. R. Morris (Ed. H. G. Brittain, Marcel Dekker, 1995). Isolated site hydrates are ones in which the water molecules are isolated from direct contact with each other by intervening organic molecules. In channel hydrates, the water molecules lie in lattice channels where they are next to other water molecules. In metal-ion coordinated hydrates, the water molecules are bonded to the metal ion.

When the solvent or water is tightly bound, the complex will have a well-defined stoichiometry independent of humidity. However, when the solvent or water is weakly bound, as in channel solvates and hygroscopic compounds, the water/solvent content will be dependent on humidity and drying conditions. In such cases, non-stoichiometry will be the norm.

Unexpectedly, we have now found a crystalline form of 5-{(1S)-2-[(2R)-4-benzoyl- 2-methyl-piperazin-1-yl]-1-methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide. It is a crystalline solid with a high melting point. It crystallizes as laths, with no changes to crystallinity on dehydration. Furthermore, it has acceptable dissolution and solubility characteristics.

Crystal structure determination revealed the crystalline form of 5-{(1S)-2-[(2R)-4- benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2- carboxylic acid methylamide to be a channel hydrate, where the water molecules are loosely bound within channels running through the crystal lattice. As a consequence, the precise site occupancy level of the water molecules is sensitive to the ambient temperature and humidity conditions, ranging from anhydrous to fully hydrated. A combination of variable temperature and humidity Powder X-Ray Diffraction (PXRD), Dynamic Vapour Sorption (DVS), Differential Scanning Calorimetry (DSC) and Thermogravimetric Analysis (TGA) studies have shown that the gross crystalline structure is independent of water content. Accordingly, the PXRD pattern of the crystalline form of the invention may be used as an identifier which is independent of water content. The solid form data for the crystalline form of 5- {(1S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2-oxo-ethoxy}-4- methoxy-pyridine-2-carboxylic acid methylamide are described in more detail hereinbelow.

In one aspect, the present invention provides a crystalline form of 5-{(1S)-2-[(2R)- 4-benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2- carboxylic acid methylamide, which exhibits the following characteristic powder X ray diffraction peaks when measured using Cu Ka radiation. (Wavelength =

In one embodiment, the crystalline form is further characterised by the following characteristic powder X ray diffraction peaks when measured using Cu Ka radiation (Wavelength = 1.5406 A):

In another embodiment the crystalline form is further characterised by the following characteristic powder X ray diffraction peaks when measured using Cu Kg radiation (Wavelength = 1.5406 A):

In another embodiment the crystalline form is further characterised by the following characteristic powder X ray diffraction peaks when measured using Cu Kq radiation. (Wavelength = 1.5406 A):

25.9

28.6

29.2

In another embodiment the crystalline form is further characterised by a DSC trace which shows a sharp endotherm at 127⁰C.

The present invention also provides a process for preparing the crystalline form of the invention comprising crystallising the compound of formula (I)

from a solvent system comprising at least one ester and water.

In one embodiment the crystalline form of 5-{(1S)-2-[(2R)-4-benzoyl-2-methyl- piperazin-1 -yl]-1 -methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide has the following structural formula (II):

wherein Me is methyl and 0<n<1.

In one embodiment, the crystalline form of (5-{(1S)-2-[(2R)-4-benzoyl-2-methyl- piperazin-1 -yl]-1 -methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide, in particular the crystalline form of formula (II), may be prepared by crystallising the free base compound of formula (I) from a solvent system comprising at least one organic solvent. In one embodiment the solvent system comprises at least one organic solvent and water. The organic solvent may be a medium polarity organic solvent. Typically, the organic solvent is not anhydrous. The medium polarity organic solvent may be an ester. For example, the organic solvent may be n-butyl acetate or isopropyl acetate. In one embodiment, the organic solvent is n-butyl acetate. In one embodiment the solvent system comprises between 0.2-5% w/w water. In one embodiment, the solvent system comprises an ester, for example n-butyl acetate, and between 0.2-5% w/w water. In alternative embodiments, the solvent system comprises an ester, for example n-butyl acetate, and an amount of water selected from approximately 0.2 %w/w, approximately 0.5% w/w, approximately 1 % w/w, approximately 2% w/w, approximately 3% w/w, approximately 4% w/w and approximately 0.5% w/w water. In one embodiment, the solvent system comprises n-butyl acetate and approximately 1.5% w/w water.

The compound of formula (I) may be in the form of an amorphous solid prior to crystallisation. An amorphous form of the compound of formula (I) may comprise water, for example, between approximately 2-3% w/w water (e.g., see Figure 4).

In one embodiment, crystallisation of the compound of formula (I) in the form of an amorphous solid optionally comprising 2-3% w/w water may be performed using a solvent system comprising an ester, for example n-butyl acetate, and at least 0.2% w/w water (e.g., when butyl acetate is used at a level of 4 ml/g).

In another embodiment, crystallisation of an amorphous form of the compound of formula (I) may be carried out using pH adjustment. A compound of formula (I) may be suspended in water and an acid, for example hydrochloric acid, added until the solution is acidic, for example, less than pH 2, e.g, approximately pH 1. A suitable base, such as saturated sodium bicarbonate solution, then may be added to bring the solution to approximately pH 9. The resultant mixture optionally may be seeded to afford crystalline compound of formula (I).

Those skilled in the art also would realise that crystallisation of the compound of formula (I) may be carried out using an emulsion crystallisation procedure with water and an immiscible organic solvent. Thus, in another embodiment the compound of formula (I) may be crystallised by emulsion crystallisation from a solvent system comprising water and a water-immiscible organic solvent. The organic solvent may be an ester, such as n-butyl acetate or isopropyl acetate. In one embodiment the solvent is n-butyl acetate.

In another embodiment, crystallisation of the compound of formula (I) may be performed using an anti-solvent addition crystallisation procedure whereby a solution of the compound of formula (I) is added into (or alternatively has added to it) water, wherein the water acts as the anti-solvent. The compound of formula (I) may itself be prepared as described in WO 2005/016344 as well as the Examples and Preparations described hereinbelow.

The present invention includes all pharmaceutically acceptable isotopically- labelled forms of the crystalline form of the invention, including for example the crystalline form of formula (II). In an isotopically-labelled form, one or more atoms are replaced by an atom or atoms having the same atomic number, but an atomic mass or mass number different from the atomic mass or mass number which predominates in nature.

Suitable isotopes include isotopes of hydrogen, such as ²H and ³H; carbon, such as ¹¹C, ¹³C and ¹⁴C; nitrogen, such as ¹³N and ¹⁵N; oxygen, such as ¹⁵O, ¹⁷O and ¹⁸O; and sulphur, such as ³⁵S.

Certain isotopically-labelled compounds, such as those incorporating a radioactive isotope, are useful/in drug and/or substrate tissue distribution studies. The radioactive isotopes tritium, i.e. ³H, and carbon-14, i.e. ¹⁴C, are particularly useful for this purpose in view of their ease of incorporation and ready means of detection.

Substitution with heavier isotopes such as deuterium, i.e. ²H, may afford certain therapeutic advantages resulting from greater metabolic stability, for example, increased in vivo half-life or reduced dosage requirements, and hence may be preferred in some circumstances.

Substitution with positron emitting isotopes, such as ¹¹C, ¹⁸F, ¹⁵O and ¹³N, can be useful in Positron Emission Topography (PET) studies for examining substrate receptor occupancy.

Isotopically-labelled compounds generally may be prepared by conventional techniques known to those skilled in the art or by processes analogous to those described in the accompanying Examples and Preparations using an appropriate isotopically-labelled reagent in place of the non-labelled reagent previously employed.

Since 5-{(1 S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1 -yl]-1 -methyl-2-oxo-ethoxy}- 4-methoxy-pyridine-2-carboxylic acid methylamide inhibits the interaction of gp120 with CD4, it is therefore of use in the treatment of HIV, a retroviral infection genetically related to HIV, and AIDS. Accordingly, in another aspect the invention provides the crystalline form of the invention for use as a medicament.

In another aspect the invention provides the crystalline form of the invention for use in the treatment of HIV, a retroviral infection genetically related to HIV, or AIDS.

In another aspect the invention provides the use of the crystalline form of the invention for the manufacture of a medicament for the treatment of HIV, a retroviral infection genetically related to HIV, or AIDS.

In another aspect the invention provides a method of treatment of a mammal suffering from HIV, a retroviral infection genetically related to HIV, or AIDS which comprises treating said mammal with an effective amount of the crystalline form of the invention.

The crystalline form of the invention, also referred to herein as the compound of the invention, may be administered alone or in combination with one or more other drugs. Generally, the crystalline form of the invention will be administered as a formulation in association with one or more pharmaceutically acceptable excipients, diluents or carriers. The term "excipient" is used herein to describe any ingredient other than the compound of the invention or other active ingredient. The choice of excipient will to a large extent depend on factors such as the particular mode of administration, the effect of the excipient on solubility and stability, and the nature of the dosage form.

Pharmaceutical compositions suitable for the delivery of compound of the invention and methods for their preparation will be readily apparent to those skilled in the art. Such compositions and methods for their preparation may be found, for example, in 'Remington's Pharmaceutical Sciences', 19th Edition (Mack Publishing Company, 1995).

The compound of the invention may be administered orally. Oral administration may involve swallowing, so that the compound enters the gastrointestinal tract, or buccal or sublingual administration may be employed by which the compound enters the blood stream directly from the mouth. Formulations suitable for oral administration include solid formulations such as tablets, capsules containing particulates, liquids, or powders, lozenges (including liquid-filled), chews, multi- and nano-particulates, gels, solid solution, liposome, films (including muco-adhesive), ovules, sprays and liquid formulations.

Liquid formulations include suspensions, solutions, syrups and elixirs. Such formulations may be employed as fillers in soft or hard capsules and typically comprise a carrier, for example, water, ethanol, polyethylene glycol, propylene glycol, methylcellulose, or a suitable oil, and one or more emulsifying agents and/or suspending agents. Liquid formulations may also be prepared by the reconstitution of a solid, for example, from a sachet.

The compound of the invention may also be used in fast-dissolving, fast- disintegrating dosage forms such as those described in Expert Opinion in Therapeutic Patents, 11 (6), 981-986 by Liang and Chen (2001).

For tablet dosage forms, depending on dose, the drug may make up from 0.1 .wt% to 80 wt%, more typically from 1 wt% to 60 wt%, such as 5 wt% to 50 wt%, of the dosage form. In addition to the drug, tablets generally contain a disintegrant. Examples of disintegrants include sodium starch glycolate, sodium carboxymethyl cellulose, calcium carboxymethyl cellulose, croscarmellose sodium, crospovidone, polyvinylpyrrolidone, methyl cellulose, microcrystalline cellulose, lower alkyl-substituted hydroxypropyl cellulose, starch, pregelatinised starch and sodium alginate. Generally, the disintegrant will comprise from 0.1 wt% to 25 wt%, more typically from 0.5 wt% to 20 wt%, such as 1 wt% to 15 wt%, of the dosage form.

Binders are generally used to impart cohesive qualities to a tablet formulation. Suitable binders include microcrystalline cellulose, gelatin, sugars, polyethylene glycol, natural and synthetic gums, polyvinylpyrrolidone, pregelatinised starch, hydroxypropyl cellulose and hydroxypropyl methylcellulose. Tablets may also contain diluents, such as lactose (monohydrate, spray-dried monohydrate, anhydrous and the like), mannitol, xylitol, dextrose, sucrose, sorbitol, microcrystalline cellulose, starch, calcium carbonate and dibasic calcium phosphate dihydrate.

Tablets may also optionally comprise surface active agents, such as sodium lauryl sulfate and polysorbate 80, and glidants such as silicon dioxide and talc. When present, surface active agents may comprise from 0.2 wt% to 5 wt% of the tablet, and glidants may comprise from 0.2 wt% to 1 wt% of the tablet. Tablets also may contain lubricants such as magnesium stearate, calcium stearate, zinc stearate, sodium stearyl fumarate, and mixtures of magnesium stearate with sodium lauryl sulphate. Lubricants generally comprise from 0.25 wt% to 10 wt%, for example, from 0.5 wt% to 3 wt% of the tablet.

Other possible ingredients include anti-oxidants, colourants, flavours, preservatives and taste-masking agents.

Exemplary tablets contain up to about 80% drug, from about 10 wt% to about 90 wt% binder, from about 0 wt% to about 85 wt% diluent, from about 1 wt% to about 10 wt% disintegrant, and from about 0.25 wt% to about 10 wt% lubricant.

Tablet blends may be compressed directly or by roller to form tablets. Tablet blends or portions of blends may alternatively be wet-, dry-, or melt-granulated, melt congealed, or extruded before tabletting. The final formulation may comprise one or more layers, may be coated or uncoated, and may be encapsulated.

The formulation of tablets is discussed in "Pharmaceutical Dosage Forms: Tablets, Vol. 1", by H. Lieberman and L. Lachman, Marcel Dekker, N.Y., N.Y., 1980 (ISBN 0-8247-6918-X).

Solid formulations for oral administration may be formulated to be immediate and/or modified release. Modified release formulations include delayed-, sustained-, pulsed-, controlled-, targeted and programmed release.

Suitable modified release formulations for the purposes of the invention are described in US Patent No. 6,106,864. Details of other suitable release technologies such as high energy dispersions and osmotic and coated particles are to be found in Verma et a/, Pharmaceutical Technology On-line, 25(2), 1-14 (2001). The use of chewing gum to achieve controlled release is described in WO 00/35298.

The compound of the invention may also be administered directly into the blood stream, into muscle, or into an internal organ. Suitable means for parenteral administration include intravenous, intraarterial, intraperitoneal, intrathecal, intraventricular, intraurethral, intrasternal, intracranial, intramuscular and subcutaneous. Suitable devices for parenteral administration include needle (including microneedle) injectors, needle-free injectors and infusion techniques. Parenteral formulations are typically aqueous solutions which may contain excipients such as salts, carbohydrates and buffering agents (preferably to a pH of from 3 to 9), but, for some applications, they may be more suitably formulated as a sterile non-aqueous solution or as a dried form to be used in conjunction with a suitable vehicle such as sterile, pyrogen-free water.

The preparation of parenteral formulations under sterile conditions, for example, by reconstituting a lyophilised compound, may readily be accomplished using standard pharmaceutical techniques well known to those skilled in the art.

The solubility of the compound of the invention when used in the preparation of parenteral solutions may be enhanced by the use of appropriate formulation techniques, such as the incorporation of solubility-enhancing agents.

Formulations for parenteral administration may be formulated to be immediate and/or modified release. Modified release formulations include delayed-, sustained-, pulsed-, controlled-, targeted and programmed release. Thus compounds of the invention may be formulated as a solid, semi-solid, or thixotropic liquid for administration as an implanted depot providing modified release of the compound. Examples of such formulations include drug-coated stents and PGLA microspheres.

The compound of the invention also may be administered topically to the skin or mucosa, that is, dermally or transdermally. Typical formulations for this purpose include gels, hydrogels, lotions, solutions, creams, ointments, dusting powders, dressings, foams, films, skin patches, wafers, implants, sponges, fibres, bandages and microemulsions. Liposomes may also be used. Typical carriers include alcohol, water, mineral oil, liquid petrolatum, white petrolatum, glycerin, polyethylene glycol and propylene glycol. Penetration enhancers may be incorporated - see, for example, J Pharm Sci, 88 (10), 955-958 by Finnin and Morgan (October 1999).

Other means of topical administration include delivery by electroporation, iontophoresis, phonophoresis, sonophoresis and microneedle or needle-free (e.g. Powderject™, Bioject™, etc.) injection. Formulations for topical administration may be formulated to be immediate and/or modified release. Modified release formulations include delayed-, sustained-, pulsed-, controlled-, targeted and programmed release.

The compound of the invention also may be administered intranasally or by inhalation, typically in the form of a dry powder (either alone, as a mixture, for example, in a dry blend with lactose, or as a mixed component particle, for example, mixed with phospholipids, such as phosphatidylcholine) from a dry powder inhaler or as an aerosol spray from a pressurised container, pump, spray, atomiser (preferably an atomiser using electrohydrodynamics to produce a fine mist), or nebuliser, with or without the use of a suitable propellant, such as 1 ,1 ,1 ,2-tetrafluoroethane or 1 ,1 ,1 ,2,3,3,3-heptafluoropropane. For intranasal use, the powder may comprise a bioadhesive agent, for example, chitosan or cyclodextrin.

The pressurised container, pump, spray, atomizer, or nebuliser contains a solution or suspension of the compound of the invention comprising, for example, ethanol (optionally, aqueous ethanol) or a suitable alternative agent for dispersing, solubilising, or extending release of the compound, the propellant(s) as solvent and an optional surfactant, such as sorbitan trioleate, oleic acid, or an oligolactic acid.

Prior to use in a dry powder or suspension formulation, the drug product is micronised to a size suitable for delivery by inhalation (typically less than 5 microns). This may be achieved by any appropriate comminuting method, such as spiral jet milling, fluid bed jet milling, supercritical fluid processing to form nanoparticles, high pressure homogenisation, or spray drying.

Capsules (made, for example, from gelatin or HPMC), blisters and cartridges for use in an inhaler or insufflator may be formulated to contain a powder mix of the compound of the invention, a suitable powder base such as lactose or starch and a performance modifier such as /-leucine, mannitol, or magnesium stearate. The lactose may be anhydrous or in the form of the monohydrate, preferably the latter. Other suitable excipients include dextran, glucose, maltose, sorbitol, xylitol, fructose, sucrose and trehalose.

A suitable solution formulation for use in an atomiser using electrohydrodynamics to produce a fine mist may contain from 1μg to 20mg of the compound of the invention per actuation and the actuation volume may vary from 1 μl to 100μl. A typical formulation may comprise a compound of the invention, propylene glycol, sterile water, ethanol and sodium chloride. Alternative solvents which may be used instead of propylene glycol include glycerol and polyethylene glycol.

Suitable flavours, such as menthol and levomenthol, or sweeteners, such as saccharin or saccharin sodium, may be added to those formulations of the invention intended for inhaled/intranasal administration.

Formulations for inhaled/intranasal administration may be formulated to be immediate and/or modified release using, for example, poly(DL-lactic-coglycolic acid) (PGLA). Modified release formulations include delayed-, sustained-, pulsed-, controlled-, targeted and programmed release.

In the case of dry powder inhalers and aerosols, the dosage unit is determined by means of a valve which delivers a metered amount. Units in accordance with the invention are typically arranged to administer a metered dose or "puff' containing from 1 μg to 10mg of the compound of the invention. The overall daily dose will typically be in the range 1 μg to 200mg which may be administered in a single dose or, more usually, as divided doses throughout the day.

The compound of the invention may be administered rectally or vaginally, for example, in the form of a suppository, pessary, vaginal ring, or enema. Cocoa butter is a traditional suppository base, but various alternatives may be used as appropriate. As described hereinabove, the compound of the invention also may be applied topically to mucosa, such as vaginal and rectal mucosa. Typical formulations for this purpose include gels, creams, ointments, foams, wafers, implants and sponges.

Formulations for rectal/vaginal administration may be formulated to be immediate and/or modified release. Modified release formulations include delayed-, sustained-, pulsed-, controlled-, targeted and programmed release.

The compound of the invention may also be administered directly to the eye or ear, typically in the form of drops of a micronised suspension or solution in isotonic, pH-adjusted, sterile saline. Other formulations suitable for ocular and aural administration include ointments, biodegradable (e.g. absorbable gel sponges, collagen) and non-biodegradable (e.g. silicone) implants, wafers, lenses and particulate or vesicular systems, such as niosomes or liposomes. A polymer such as crossed-linked polyacrylic acid, polyvinylalcohol, hyaluronic acid, a cellulosic polymer, for example, hydroxypropylmethylcellulose, hydroxyethylcellulose, or methyl cellulose, or a heteropolysaccharide polymer, for example, gelan gum, may be incorporated together with a preservative, such as benzalkonium chloride. Such formulations may also be delivered by iontophoresis.

Formulations for ocular/aural administration may be formulated to be immediate and/or modified release. Modified release formulations include delayed-, sustained-, pulsed-, controlled-, targeted, or programmed release.

Inasmuch as it may desirable to administer the compound of the invention in combination with another therapeutic agent, for example, for the purpose of treating a particular disease or condition, it is within the scope of the present invention that two or more pharmaceutical compositions, at least one of which contains the compound of the invention, may conveniently be combined in the form of a kit suitable for coadministration of the compositions.

Thus, the kit of the invention may comprise two or more separate pharmaceutical compositions, at least one of which contains the compound of the invention, and means for separately retaining said compositions, such as a container, divided bottle, or divided foil packet. An example of such a kit is the familiar blister pack used for the packaging of tablets, capsules and the like.

The kit of the invention is particularly suitable for administering different dosage forms, for example, oral and parenteral, for administering the separate compositions at different dosage intervals, or for titrating the separate compositions against one another. To assist compliance, the kit typically comprises directions for administration and may be provided with a so-called memory aid.

For administration to human patients, having a weight of about 65 to 70kg, the total daily dose of the compound of the invention is typically in the range 1mg to 10,000mg, such as 10mg to 1 ,000mg, for example 25mg to 500mg, in particular 50mg to 400mg, depending, of course, on the mode of administration, the age, condition and weight of the patient, and will in any case be at the ultimate discretion of the physician. The total daily dose may be administered in single or divided doses. Accordingly in another aspect the invention provides a pharmaceutical composition including a crystalline form of the invention together with one or more pharmaceutically acceptable excipients, diluents or carriers.

The compound of the invention may be administered alone or as part of a combination therapy. Thus, included within the scope of the present invention are embodiments comprising co-administration of, and compositions which contain, in addition to the compound of the invention, one or more additional therapeutic agents. Such multiple drug regimens, often referred to as combination therapy, may be used in the treatment and prevention of infection by human immunodeficiency virus, HIV. The use of such combination therapy is especially pertinent with respect to the treatment and prevention of infection and multiplication of the human immunodeficiency virus, HIV, and related pathogenic retroviruses within a patient in need of treatment or one at risk of becoming such a patient. The ability of such retroviral pathogens to evolve within a relatively short period of time into strains resistant to any monotherapy which has been administered to said patient is well known in the literature. A recommended treatment for HIV is a combination drug treatment called Highly Active Anti- Retroviral Therapy, or HAART. HAART combines three or more HIV drugs. Thus, the methods of treatment and pharmaceutical compositions of the present invention may employ the compound of the invention in the form of monotherapy, but said methods and compositions may also be used in the form of combination therapy in which the compound of the invention is co-administered in combination with one or more additional therapeutic agents such as those described in detail further herein.

In a further embodiment of the invention, combinations of the present invention include treatment with the compound of the invention, and one or more additional therapeutic agents selected from the following: HIV protease inhibitors (PIs), including but not limited to amprenavir (141W94), CGP-73547, CGP-61755, DMP-450 (mozenavir), nelfinavir, ritonavir, saquinavir (invirase), lopinavir, TMC-126, atazanavir, palinavir, GS-3333, KN 1-413, KNI-272, LG-71350, CGP-61755, PD 173606, PD 177298, PD 178390, PD 178392, U-140690, ABT-378, DMP-450, AG-1776, MK-944, VX-478, indinavir, tipranavir, TMC-114, DPC-681 , DPC-684, fosamprenavir calcium (Lexiva), benzenesulfonamide derivatives disclosed in WO 03/053435, R-944, Ro-03-34649, VX-385, GS-224338, OPT-TL3, PL-100, PPL-100, SM-309515, AG-148, DG-35-VIII, DMP-850, GW-5950X, KNI-1039, L-756423, LB-71262, LP-130, RS-344, SE-063, UIC-94-003, Vb-19038, A-77003, BMS-182193, BMS-186318, SM-309515, JE-2147, GS-9005; non-nucleoside reverse transcriptase inhibitors (NNRTIs), including but not limited to: efavirenz, HBY-097, nevirapine, TMC-120 (dapivirine), TMC-125, etravirine, delavirdine, DPC-083, DPC-961 , capravirine, rilpivirine, 5-{[3,5-Diethyl- 1-(2-hydroxyethyl)-1/-/-pyrazol-4-yl]oxy}isophthalonitrile or pharmaceutically acceptable salts, solvates or derivatives thereof; 5-{[3-cyclopropyl-1-(2- hydroxyethyl)-5-methyl-1 /-/-pyrazol-4-yl]oxy}isophthalonitrile or pharmaceutically acceptable salts, solvates or derivatives thereof; GW-678248, GW-695634, MIV-150, calanolide, and tricyclic pyrimidinone derivatives as disclosed in WO 03/062238; nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs), including but not limited to abacavir, GS-840, lamivudine, adefovir dipivoxil, beta-fluoro-ddA, zalcitabine, didanosine, stavudine, zidovudine, tenofovir disoproxil fumarate, amdoxovir (DAPD), SPD-754, SPD-756, racivir, reverset (DPC-817), MIV-210 (FLG), beta-L-Fd4C (ACH-126443), MIV-310 (alovudine, FLT), dOTC, DAPD, entecavir, GS-7340, emtricitabine (FTC);

CCR5 antagonists, including but not limited to: TAK-779, SC-351125, SCH-C (ancriviroc), SCH-D (vicriviroc), maraviroc, PRO-140, aplaviroc, AMD-887 CMPD-167, methyl 1-eA7c/o-{8-[(3S)-3-(acetylamino)-3-(3-fluorophenyl)propyl]-8-azabicyclo- [3.2.1]oct-3-yl}-2-methyl-4,5,6,7-tetrahydro-1ry-imidazo[4,5-c]pyridine-5- carboxylate or pharmaceutically acceptable salts, solvates or derivatives thereof, methyl 3-eπc/o-{8-[(3S)-3-(acetamido)-3-(3-fluorophenyl)propyl]-8-azabicyclo- [3.2.1]oct-3-yl}-2-methyl-4,5,6,7-tetrahydro-3H-imidazo[4,5-c]pyridine-5- carboxylate or pharmaceutically acceptable salts, solvates or derivatives thereof, ethyl 1-endo-{8-[(3S)-3-(acetylamino)-3-(3-fluorophenyl)propyl]-8-azabicyclo- [S^.^oct-S-ylJ^-methyl^.δ.ey-tetrahydro-IH-imidazoμ.δ-cJpyridine-δ- carboxylate or pharmaceutically acceptable salts, solvates or derivatives thereof, and Λ/-{(1 S)-3-[3-enc/o-(5-isobutyryl-2-methyl-4,5,6,7-tetrahydro-1 W-imidazo[4,5- c]pyridin-1-yl)-8-azabicyclo[3.2.1]oct-8-yl]-1-(3-fluorophenyl)propyl}-acetamide) or pharmaceutically acceptable salts, solvates or derivatives thereof; entry and fusion inhibitors including, but not limited to, BMS-806, BMS-488043, and 4-{(1 S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1 -yl]-1 -methyl-2-oxo-ethoxy}-3- methoxy-N-methyl-benzamide, enfuvirtide (T-20), SP-01 A, T1249, PRO 542, PRO 140, AMD-3100, soluble CD4, compounds disclosed in JP 2003171381 , and compounds disclosed in JP 2003119137;

HIV integrase inhibitors, including but not limited to, L-000870810 GW-810781, 1 ,5-naphthyridine-3-carboxamide derivatives disclosed in WO 03/062204, compounds disclosed in WO 03/047564, compounds disclosed in WO 03/049690, and δ-hydroxypyrimidine^-carboxamide derivatives disclosed in WO 03/035076, MK-0518 (5-(1 ,1-dioxo-1 ,2-thiazinan-2-yl)-N- (4-fluorobenzyl)-8- hydroxy-1 ,6-naphthyridine-7-carboxamide disclosed in WO 03/016315); prenylation inhibitors including, but not limited to, HMG CoA reductase inhibitors, such as statins (e.g. atorvastatin); RNaseH inhibitors; maturation inhibitors which act by interfering with production of the HIV capsid protein; and compounds useful as anti-infectives.

There is also included within the scope the present invention, combinations of the compound of the invention, together with one or more additional therapeutic agents independently selected from the group consisting of proliferation inhibitors, e.g. hydroxyurea; immunomodulators, such as AD-439, AD-519, alpha interferon, AS-101 , bropirimine, acemannan, CL246,738, EL10, FP-21399, gamma interferon, granulocyte macrophage colony stimulating factor (e.g. sargramostim), IL-2, immune globulin intravenous, IMREG-1 , IMREG-2, imuthiol diethyl dithio carbamate, alpha-2 interferon, methionine-enkephalin, MTP-PE, remune, rCD4, recombinant soluble human CD4, interferon alfa-2, SK&F106528, soluble T4 thymopentin, tumor necrosis factor (TNF), tucaresol, recombinant human interferon beta, interferon alfa n-3; tachykinin receptor modulators (e.g. NK1 antagonists) and various forms of interferon or interferon derivatives; other chemokine receptor agonists/antagonists such as CXCR4 antagonists (e.g AMD070 and AMD3100) or CD4 antagonists (e.g. TNX-355); agents which substantially inhibit, disrupt or decrease viral transcription or RNA replication such as inhibitors of tat (transcriptional trans activator) or nef (negative regulatory factor); agents which substantially inhibit, disrupt or decrease translation of one or more proteins expressed by the virus (including, but not limited to, down regulation of protein expression or antagonism of one or more proteins) other than reverse transcriptase, such as Tat or Nef; agents which influence, in particular down regulate, CCR5 receptor expression; chemokines that induce CCR5 receptor internalisation such MIP-1α, MIP-1 β, RANTES and derivatives thereof; and other agents that inhibit viral infection or improve the condition or outcome of HIV-infected individuals through different mechanisms; agents which influence, in particular down regulate, CCR5 receptor expression; chemokines that induce CCR5 receptor internalisation such MIP-1α, MIP-1β, RANTES and derivatives thereof, including, but not limited to, immunosupressants, such as calcineurin inhibitors (e.g. tacrolimus and cyclosporin A); steroids; agents which interfere with cytokine production or signalling, such as Janus Kinase (JAK) inhibitors (e.g. JAK-3 inhibitors, including 3-{(3R,4R)-4-methyl-3-[methyl-(7H-pyrrolo[2,3-d]pyrimidin-4-yl)-amino]-piperidin- 1-yl}-3-oxo-propionitrile) and pharmaceutically acceptable salts, solvates or derivatives thereof; cytokine antibodies (e.g. antibodies that inhibit the interleukin- 2 (IL-2) receptor, including basiliximab and daclizumab); and agents which interfere with cell activation or cell cycling, such as rapamycin.

There is also included within the scope the present invention, combinations of the compound of the invention, together with one or more additional therapeutic agents which slow down the rate of metabolism of the compound of the invention, thereby leading to increased exposure in patients. Increasing the exposure in such a manner is known as boosting. This has the benefit of increasing the efficacy of the compound of the invention or reducing the dose required to achieve the same efficacy as an unboosted dose. Metabolism of the compound of the invention includes oxidative processes carried out by P450 (CYP450) enzymes, particularly CYP 3A4 and conjugation by UDP glucuronosyl transferase and sulphating enzymes. Thus, among the agents that may be used to increase the exposure of a patient to a compound of the present invention are those that can act as inhibitors of at least one isoform of the cytochrome P450 (CYP450) enzymes. The isoforms of CYP450 that may be beneficially inhibited include, but are not limited to, CYP1A2, CYP2D6, CYP2C9, CYP2C19 and CYP3A4. Suitable agents that may be used to inhibit CYP 3A4 include, but are not limited to, ritonavir, saquinavir or ketoconazole.

It will be appreciated by a person skilled in the art that a combination drug treatment, as described herein above, may comprise two or more compounds having the same, or different, mechanism of action. Thus, by way of illustration only, a combination may comprise the compound of the invention and: one or more NRTIs; one or more NRTIs and a Pl; one or more NRTIs and a CCR5 antagonist; one or more CCR5 antagonists; a Pl; a Pl and an NNRTI; an NNRTI; and so on.

In addition to the requirement of therapeutic efficacy, which may necessitate the use of therapeutic agents in addition to the compound of the invention, there may be additional rationales which compel or highly recommend the use of a combination of the compound of the invention with another therapeutic agent, such as in the treatment of diseases or conditions which directly result from or indirectly accompany the basic or underlying modulated disease or condition. For example, it may be necessary or at least desirable to treat Hepatitis C Virus (HCV), Hepatitis B Virus (HBV), Human Papillomavirus (HPV), opportunistic infections (including bacterial and fungal infections), neoplasms, and other conditions which may occur as the result of the immune-compromised state of the patient being treated. Other therapeutic agents may be used with the compounds of the invention, e.g., in order to provide immune stimulation or to treat pain and inflammation which may accompany HIV infection.

Accordingly, therapeutic agents for use in combination with the compound of the invention also include: interferons, pegylated interferons (e.g. peginterferon alfa-

2a and peginterferon alfa-2b), lamivudine, ribavirin and emtricitabine for the treatment of hepatitis; antifungals such as fluconazole, fosfluconazole, itraconazole, and voriconazole; antibacterials such as azithromycin and clarithromycin; interferons, daunorubicin, doxorubicin, and paclitaxel for the treatment of AIDS related Kaposi's sarcoma; and cidofovir, fomivirsen, foscarnet, ganciclovir and valcyte for the treatment of cytomegalovirus (CMV) retinitis.

In the above-described combinations, the compound of the invention may be administered, in terms of dosage forms, either separately or in conjunction with one or more other therapeutic agents; and in terms of their time of administration, either simultaneously or sequentially in any order. Thus, the administration of one component agent may be prior to, concurrent with, or subsequent to the administration of the other component agent(s).

Accordingly, in a further aspect the invention provides a pharmaceutical composition comprising a crystalline form of the invention and one or more additional therapeutic agents.

It is to be appreciated that all. references herein to treatment include curative, palliative and prophylactic treatment.

The following example and preparations illustrate the preparation of the crystalline form of 5-{(1S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2- oxo-ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide in which the following further abbreviations may be used: h = hour min = minute

RT means room temperature

LRMS = low resolution mass spectrum HRMS = high resolution mass spectrum APCI = atmospheric pressure chemical ionisation ESI = electrospray ionisation NMR = nuclear magnetic resonance HPLC = high-pressure liquid chromatography

[α]o = angle of optical rotation tic - thin layer chromatography Me = methyl

Unless otherwise stated, standard laboratory solvents were used in the following Examples and Preparations, which were not dried prior to use.

Example 1

Crystalline 5-((1 S)-2-IY2R)-4-Benzoyl-2-methyl-piperazin-1 -yli-1 -methyl-2-oxo- ethoxyM-methoxy-pyridine^-carboxylic acid methylamide

(S)-2-(4-Methoxy-6-methylcarbamoyl-pyridin-3-yloxy)-propionic acid hydrochloride (33 g, 0.11 mol), 1-hydroxybenzotriazole hydrate (20.0 g, 0.15 mol), Λ/-ethyldiisopropylamine (93 ml_, 0.53 mol) and 1-(3-dimethylaminopropyl)- 3-ethylcarbodiimide hydrochloride (58.8 g, 0.3 mol) were dissolved in N,N- dimethylacetamide (600 mL) and the mixture stirred at room temperature for two hours. A solution of (3R)-(3-methyl-piperazin-1-yl)-phenyl-methanone (J. Med. Chem. 43(23), 4499; 2000) (23.2 g, 0.11 mol) in Λ/,/V-dimethylacetamide (30 mL) was then added and the mixture stirred at room temperature for 48 hours. The reaction mixture was evaporated to dryness and the residue partitioned between ethyl acetate (400 mL) and saturated aqueous sodium bicarbonate solution (200 mL). The organic layer was separated and the aqueous layer extracted with additional ethyl acetate (4 X 100 mL). The combined organic extracts were dried (MgSO₄) and evaporated under reduced pressure to give an off-white solid. This was purified by column chromatography on silica gel, eluting with ethyl acetate:methanol, 90:10 to afford crude 5-{(1S)-2-[(2R)-4-benzoyl-2-methyl- piperazin-1 -yl]-1 -methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide as a white foam but still containing residual /V,Λ/-dimethylacetamide. The crude product was dissolved in ethyl acetate (400 mL) and washed with water (3 X 100 mL). The organic layer was dried (MgSO₄) and then evaporated under reduced pressure to give 5-{(1 S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1- yl]-1-methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide as an amorphous white solid. This was dissolved in the minimum amount of hot n- butyl acetate (100 mL), the solution filtered while hot and then allowed to cool to room temperature. After standing at room temperature for 120 hours the deposited crystalline material was filtered off, washed with a little cold π-butyl acetate and dried under vacuum at 65⁰C to give the title compound as a crystalline white solid, 15.22 g. LRMS (APCI⁺): m/z [M+H]⁺ 441

¹H NMR (400MHz, CD₃OD): δ : 8.05 (1H, s), 7.75 (1H, s), 7.50-7.40 (5H, m), 5.40 (1 H, m), 4.60-3.00 (8H, m), 3.95 (3H, br s), 2.90 (3H, s), 1.60-1.00 (6H, m) ppm. Found C, 60.91 ; H, 6.56; N, 12.31. C₂₃H₂₈N₄O₅. 0.75 H₂O requires C, 60.85; H, 6.55; N₁ 12.34%

PXRD, DSC, TGA and DVS data for the compound of Example 1 are provided hereinbelow.

Example 2 Crystalline 5-{(1S)-2-r(2R)-4-Benzoyl-2-methyl-piperazin-1-yll-1-methyl-2-oxo- ethoxy)-4-methoxy-pyridine-2-carboxylic acid methylamide

A solution of 5-{(1 S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2-oxo- ethoxy}-4-methoxy-pyridine-2-carboxylic acid methyl ester (0.2 g, 0.45 mmol) in a 33% w/w solution of methylamine in ethanol (3 ml) was heated in a Reacti-vial™ at 5O⁰C for 18 hours. After cooling to room temperature, solvent was evaporated under reduced pressure. The residue was purified by column chromatography on silica gel, eluting with dichloromethane:methanol:NH₃, 98:2:0.5 to 95:5:0.5, to afford 5-{(1S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2-oxo-ethoxy}- 4-methoxy-pyridine-2-carboxylic acid methylamide as an amorphous white solid, 0.18 g.

Found C, 61.65; H₁ 6.47; N, 12.41. C₂₃H₂₈N₄O₅. 0.1 mol CH₂CI₂ requires C,

61.80; H₁ 6.33; N₁ 12.48%

[α]_D -15.5° (1mg/ml in MeOH, 25⁰C, wavelength 589nM)

A sample of the amorphous 5-{(1S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1-yl]-1^j methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide was dissolved in the minimum amount of hot π-butyl acetate, the solution was filtered while hot and then allowed to cool to room temperature. After standing at room temperature for 120 hours the deposited crystalline material was filtered off, washed with a little cold π-butyl acetate and dried under vacuum at 65⁰C to give the title compound as a crystalline white solid. The LRMS and ¹H NMR data for the title compound of Example 2 were consistent with that of Example 1.

Example 3 Crystalline 5-((1S)-2-r(2R)-4-Benzoyl-2-methyl-piperazin-1-yl1-1-methyl-2-oxo- ethoxyH-methoxy-pyridine-2-carboxylic acid methylamide

(3/?)-(3-Methyl-piperazin-1-yl)-phenyl-methanone hydrochloride (570 g, 2.37 mol) was dissolved in water (1.71 L, 3 L/kg) and then potassium carbonate (655 g) was added with stirring. The resultant biphasic mixture was then extracted with dichloromethane (1.71 L) and the organic phase was dried over magnesium sulfate. The solvent was then removed under vacuum to give (3f?)-(3-methyl- piperazin-1-yl)-phenyl-methanone as an oil. This was weighed and the charges of reagents for the amide coupling were adjusted accordingly. To the (3f?)-(3- methyl-piperazin-1-yl)-phenyl-methanone was added dichloromethane (7.125 L), followed by (S)-2-(4-methoxy-6-methylcarbamoyl-pyridin-3-yloxy)-propionic acid (691 g, 2.21 mol) and then 1-hydroxybenzotriazole (388 g, 2.87 mol). 1-(3- Dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (1271 g, 6.63 mol) was then added to the reaction at room temperature under nitrogen. After 3 hours the reaction was deemed to be complete so the reaction was washed with water (3.56 L), and then 10% w/w aqueous citric acid (2 x 3.56 L). The organic phase was then washed with 0.5 M aqueous sodium bicarbonate solution (3.56 L) and then saturated brine (3.56 L). The organic phase was dried with magnesium sulfate and the solution was then concentrated under vacuum to give 5-{(1S)-2- [(2R)-4-benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2-oxo-ethoxy}-4-methoxy- pyridine-2-carboxylic acid methylamide as an oil/foam product (766 g, 1.74 mol, 79% yield). The product was then crystallised as follows. To a concentrated solution of (5-{(1S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2-oxo- ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide (903 g, 2.05 mol) in dichloromethane was added π-butyl acetate (2.0 L), then the solvent was removed under vacuum. To the resultant amorphous solid was added n-butyl acetate (4.0 L) followed by water (32.0 ml, 0.78% w/w) and then seed crystals of (5-{(1 S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1 -yl]-1 -methyl-2-oxo-ethoxy}-4- methoxy-pyridine-2-carboxylic acid methylamide (1.0 g). The resultant solution was then stirred at 18 ⁰C for 64 hours. Only a small amount of solid had crystallised out so more water (29.0 ml, 0.71 % w/w) was added and the slurry was then stirred at 22 ⁰C for a further 18 hours. The resultant slurry was then cooled to 5 ⁰C for 3 hours and then filtered under vacuum. The filter cake was washed with π-butyl acetate (1.0 L) containing water (30 ml) and then dried under vacuum at 60 ⁰C to give the title compound as a white crystalline solid (705 g, 1.6 mol, 78% yield).

The ¹H NMR, LRMS and optical rotation data for the title compound of Example 3 were consistent with that of Example 1.

Example 4

Crystalline 5-1(1 S)-2-r(2R)-4-Benzoyl-2-methyl-piperazin-1-yl1-1-methyl-2-oxo- ethoxy)-4-methoxy-pyridine-2-carboxylic acid methylamide

Amorphous 5-{(1 S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1 -yl]-1 -methyl-2-oxo- ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide (5.0 g, 11.4 mmol) was suspended in water, then concentrated hydrochloric acid was added until the pH was 1 as judged by indicator paper. To the resultant yellow solution was then added saturated sodium bicarbonate solution until the pH was 9 as judged by indicator paper. The resulting emulsion was seeded and stirred overnight. The resultant slurry was then filtered under vacuum and the product was dried overnight under vacuum at 50 ⁰C giving the title compound (3.7 g, 8.4 mmol, 74% yield).

The LRMS and ¹H NMR for the title compound of Example 4 were consistent with that of Example 1.

Preparation 1A δ-Benzyloxy^-methoxy-pyridine^-carboxylic acid methylamide

δ-Benzyloxy^-methoxy-pyridine^-carboxylic acid methyl ester (24.7 g, 90.4 mmol) (Tetrahedron Letters, 38, 1297 (1997)) was dissolved in a solution of methylamine in ethanol (3M, 82 mL) and the mixture heated in a bomb at 50 ⁰C for 18 hours. The cool reaction mixture was evaporated under reduced pressure to dryness and the residue purified by column chromatography on silica gel, eluting with dichloromethane:methanol, 97:3 to afford the title compound as a white solid, 21.9 g. LRMS (APCI⁺): m/z [MH]⁺ 273 Preparation 1 B δ-Benzyloxy^-methoxy-pyridine^-carboxylic acid methylamide

To a suspension of 5-benzyloxy-4-methoxy-pyridine-2-carboxylic acid methyl ester (1.62 kg, 5.94 mol) in methanol (8.12 L) at 32 ⁰C was added a solution of methylamine in water (40% w/w, 1.62 L). The resultant solution was then stirred at room temperature for 2 hours before seeding with a small quantity of 5- benzyloxy-4-methoxy-pyridine-2-carboxylic acid methylamide and then allowing to stir for 18 hours. The resultant slurry was cooled to 5 ⁰C and held at this temperature for 1.4 hours before filtration under vacuum. The product was dried under vacuum at 55 ⁰C overnight to give the title compound as an off-white solid

(1.33 kg, 4.88 mol, 82% yield).

¹H-NMR (CD₃OD, 300 MHz), δ: 2.94 (3 H, s), 3.97 (3 H, d), 5.22 (2 H, s), 7.32-

7.46 (5 H, m), 7.11 (1 H, s), 8.16 (1 H, s).

Preparation 2

5-Hvdroxy-4-methoxy-pyridine-2-carboxylic acid methylamide

A solution of 5-benzyloxy-4-methoxy-pyridine-2-carboxylic acid methylamide (10.8 g, 39.7 mmol) and 10 % w/w Pd on carbon (0.1 g) in methanol (100 mL) was hydrogenated (138 kPa H2) at room temperature for five hours. The reaction mixture was filtered through Arbocel^®, washing through with methanol (1OmL). The filtrate was concentrated in vacuo to afford the title compound as a white solid, 7.2 g.

LRMS (APCI⁺): m/z [M+H]⁺ 183; ¹H-NMR (CD₃OD, 300 MHz) δ: 2.94 (3 H, s), 3.98 (3 H, s), 4.81 (2 H, s), 7.68 (1 H, s), 8.02 (1 H, s).

Preparation 3A (S)-2-(4-Methoxy-6-methylcarbamoyl-pyridin-3-yloxy)-propionic acid methyl ester

δ-Hydroxy^-methoxy-pyridine^-carboxylic acid methylamide (13.1 g, 72 mmol), methyl (R)-lactate (8.23 g, 79 mmol) and triphenylphosphine (30.1 g, 15 mmol) were dissolved in THF (150 ml_) and the mixture cooled to 0 ⁰C under a nitrogen atmosphere. A solution of diisopropyl azodicarboxylate (21.2 mL, 108 mmol) in THF (50 mL) was then added dropwise over one hour, the reaction mixture allowed to warm to room temperature and stirred for an additional 18 hours. The reaction mixture was evaporated to dryness to afford crude (S)-2-(4-methoxy-6- methylcarbamoyl-pyridin-3-yloxy)-propionic acid methyl ester, which was used directly in Preparation 4.

Preparation 3B

(S)-2-(4-Methoxy-6-methylcarbamoyl-pyridin-3-yloxy)-propionic acid methyl ester

To a solution of methyl-(R)-lactate (486.0 g, 4.67 mol) in THF (2.0 L) was added 5-hydroxy-4-methoxy-pyridine-2-carboxylic acid methylamide (810.0 g, 4.45 mol) followed by THF (2.0 L). To this solution was added triphenylphosphine (1.28 kg, 4.89 mol) followed by THF (2.5 L) and then the reaction mixture was cooled to 5 ⁰C. A solution of diisopropylazodicarboxylate (989 g, 4.89 mol) in THF (0.8 L) was then added to the reaction whilst maintaining the temperature below 10 ⁰C. The addition line was washed with THF (0.8 L) and this was added to the reaction, which was then allowed warm to room temperature over 18 hours. The resultant slurry was then cooled to 5 ⁰C and stirred for 2.5 hours before collecting the solid by filtration under vacuum. The resultant solid was dried under vacuum at room temperature for 2 days to give the, title compound as a white crystalline solid (754.2 g, 2.81 mol, 63% yield) which could be further purified as follows. A suspension of (S)-2-(4-methoxy-6-methylcarbamoyl-pyridin-3-yloxy)-propionic acid methyl ester (743.8 g, 2.77 mol) in isopropanol (5.58 L) was heated to 80 ⁰C to give a hazy solution and then cooled to room temperature and stirred for 18 hours. The resultant slurry was cooled to 5 ⁰C and held at this temperature for 5 hours before filtration under vacuum. The product was dried under vacuum at 60 ⁰C overnight to give the title compound (676.4 g, 2.52 mol, 91 % yield).

mp 143 ⁰C; ¹H-NMR (d_s-DMSO, 300 MHz) δ: 1.53 (3 H, d), 2.80 (3 H, d), 3.69 (3 H, s), 3,93 (3 H, s), 5.15 (1 H, q), 7.64 (1 H, s), 8.05 (1 H, s), 8.54 (1 H, d, br); HPLC (column: DAICEL Chiralpack AS-H, 5 mm, 250 mm x 4.6 mm; mobile phase: 80:20:0.1 Hexane: IPA: DEA; flow rate: 1.0 ml/min; temperature: 25 ⁰C; injection volume: 20 μl; detection: UV absorbance at 225 nm; Sample concentration: 1.0 mg per ml prepared in mobile phase; run time 30 minutes). Retention Time: major enantiomer 14.3 min.

Preparation 4 (S)-2-(4-Methoxy-6-methylcarbamoyl-pyridin-3-yloxy)-propionic acid hydrochloride

(S)-2-(4-Methoxy-6-methylcarbamoyl-pyridin-3-yloxy)-propionic acid methyl ester (crude product from Preparation 3A) (assumed to be 72 mmol), was dissolved in methanol (150 ml_), lithium hydroxide (3.93 g, 94 mmol) added portionwise and the mixture stirred at room temperature for 18 hours. The reaction mixture was evaporated to dryness and the residue partitioned between water (300 mL) and a 1:1 v/v mixture of ethyl acetate and diethyl ether (300 mL). The aqueous layer was separated and washed sequentially with ethyl acetate (2 X 200 mL), diethyl ether (2 X 200 mL) and finally a 1 :1 v/v mixture of ethyl acetate and diethyl ether (300 mL). The aqueous layer was acidified to pH 1 by addition of concentrated hydrochloric acid (~30 mL) and then evaporated to dryness. The residue was re- dissolved in acetonitrile (200 mL) and water (200 mL) and then evaporated under reduced pressure until the volume of solvent had reduced to -100 mL. On standing at room temperature a white solid started to precipitate. Acetonitrile (100 mL) was added and the mixture stirred and sonicated to complete the precipitation. The resulting white solid was filtered off, washed with acetonitrile and dried under vacuum to afford the title compound as a white solid, 25.87 g. LRMS (APCI⁺): m/z [M+H]⁺ 255 Preparation 5 (S)-2-(4-Methoxy-6-methylcarbamoyl-pyridin-3-yloxy)-propionic acid

To a slurry of (S)-2-(4-methoxy-6-methylcarbamoyl-pyridin-3-yloxy)-propionic acid methyl ester (50 g, 186.4 mmol) in water (150 ml) was added aqueous sodium hydroxide solution (25% w/w, 31.3 g, 195.7 mmol) maintaining the temperature between 22-30 ⁰C. The reaction was then stirred for 2 hours at room temperature and then aqueous hydrochloric acid (6 M, 18 ml) was added to give a pH of to 2-3 by paper. The water was then removed under vacuum and then dichloromethane (2 x 100 ml) was added and removed under vacuum to give a solid foam which was dried under vacuum at 40 ⁰C overnight to give the title compound (56.2 g crude, quantitative yield). ¹H-NMR (d₆-DMSO, 300 MHz) δ: 1.46 (3 H, d), 2.78 (3 H, d), 3.90 (3 H, s), 4.80 (1 H, q), 7.59 (1 H, s), 7.98 (1 H, s), 8.52 (1 H, q); LRMS (ES): m/z 255 [M+H]⁺.

Preparation 6

(2R)-1 -r(2f?)-4-Benzoyl-2-methyl-piperazin-1 -yll-2-benzyloxy-propan-1 -one

1-Hydroxybenzotriazole hydrate (825mg, 5.3mmol), N-methylmorpholine (808μl_, 7.3mmol), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (1.13g, 5.8mmol) and (3R)-(3-methyl-piperazin-1-yl)-phenyl-methanone [(1.Og, 4.9mmol), J. Med. Chem. 43(23), 4499; 2000] were added to a solution of (R)-(+)-2- benzyloxypropionic acid (972mg, 5.3mmol) in dichloromethane (1OmL) and the mixture was stirred for 18 hours. Additional (R)-(+)-2-benzyloxypropionic acid (486mg, 2.65mmol), 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (5.65mg, 2.9mmol), 1-hydroxybenzotriazole hydrate (413mg, 2.65mmol) and N- methylmorpholine (808μL, 7.3mmol) were added and the mixture was heated under reflux for 3 hours. Water was then added to the reaction mixture and the aqueous solution was extracted with dichloromethane (2x1 OmL). The combined organic extracts were washed with 2M hydrochloric acid, 1M sodium hydroxide solution and brine. The organic phase was dried over magnesium sulfate and concentrated in vacuo. Purification by column chromatography on silica gel, eluting with pentane:ethyl acetate, 50:50, followed by ethyl acetate:methanol, 90:10, afforded the title compound as a white foam in 81% yield, 1.45g LRMS (APCO: m/z [M-H]- 367

Preparation 7 (2R)-1-[(2/^:?)-4-Benzoyl-2-methyl-piperazin-1-yl)-2-hvdroxy-propan-1-one

Pd(OH)₂ (300mg, 2.2mmol) and ammonium formate (1.37g, 22mmol) were added to a solution of (2R)-1-[(2R)-4-benzoyl-2-methyl-piperazin-1-yl]-2-benzyloxy- propan-1-one (1.45g, 3.9mmol) in ethanol (3OmL) and the mixture was heated at 6O⁰C for 2 hours. TIc analysis then showed that the reaction had not reached completion so additional Pd(OH)2 (300mg, 2.2mmol) was added, followed by ammonium formate (1.37g, 22mmol) added at 45 minute intervals until all of the starting material was consumed. The reaction mixture was then cooled and filtered through Arbocel^®, washing through with ethanol (1OmL). The filtrate was concentrated in vacuo and the residue was purified by column chromatography on silica gel, eluting with dichloromethane:methanol, 95:5, to afford the title compound as a white gum in 91 % yield. 1q. LRMS (APCI⁺): m/z [M+H]⁺ 277

Preparation 8

5-((1S)-2-r(2R)-4-Benzoyl-2-methyl-piperazin-1-vn-1-methyl-2-oxo-ethoxy)-4- methoxy-pyridine-2-carboxylic acid methyl ester

Di-tert-butyl azodicarboxylate (0.14 g, 0.6 mmol) and polymer supported triphenylphosphine (0.25 g, 0.75 mmol) were added to a stirred solution of (2R)-1- [(2/?)-4-benzoyl-2-methyl-piperazin-1-yl)-2-hydroxy-propan-1-one (0.083 g, 0.3 mmol) and 5-hydroxy-4-methoxy-pyridine-2-carboxylic acid methyl ester (0.083 g, 0.45 mmol) (Tetrahedron Letters, 38, 1297 (1997)) in dichloromethane (3 ml_) at O⁰C. After one hour the reaction mixture was allowed to warm to room temperature and additional di-tert-butyl azodicarboxylate (0.14 g, 0.6 mmol) and polymer supported triphenylphosphine (0.25 g, 0.75 mmol) were added. After 18 hours at room temperature the reaction mixture was filtered through Arbocel^®, washing through with dichloromethane (10 mL). The filtrate was washed with saturated aqueous sodium hydrogencarbonate solution (20 mL), then the organic phase separated, dried (MgSO₄) and evaporated under reduced pressure. Purification of the residue by column chromatography on silica gel, eluting with ethyl acetate:pentane, 80:20, then ethyl acetate and finally ethyl acetate:methanol, 95:5, to afford the title compound as a white solid, 0.1 g. LRMS (APCI⁺): m/z [M+H]⁺ 442

Preparation 9

(3fi)-(3-methyl-piperazin-1 -yl)-phenyl-methanone hydrochloride

To a slurry of (R)-2-methylpiperazine dihydrochloride (300 g, 1.73 mol) in acetonitrile (2.6 L) was added potassium carbonate (479 g, 3.46 mol) with stirring under nitrogen. The reaction was then heated to 70 ⁰C for 18 hours. The reaction was then cooled to room temperature and the inorganic slurry was filtered and washed with acetonitrile (0.9 L). The filtrate was then cooled to 3 ⁰C with stirring under nitrogen. To this solution was added benzoyl chloride (221 ml, 1.90 mol) over 2.5 hours maintaining the temperature below 10 ⁰C. The reaction was allowed to warm to room temperature and stirred for 18 hours. The resultant slurry was then cooled to 3 ⁰C and then filtered under vacuum. The filter cake was washed with acetonitrile (300 ml) and was then dried under vacuum at 60 ⁰C overnight to give the title compound as a white crystalline solid (336 g, 1.40 mol, 81% yield). ¹H-NMR (d₄-methanol, 300 MHz), δ: 1.36 (3 H, s, br), 3.19-3.32 (2 H, m), 3.40- 3.50 (4 H, m), 4.79 (2 H, s), 7.51 (5 H, s). Preparation 10 (3fi)-(3-methyl-piperazin-1-yl)-phenyl-methanone

To a suspension of (3R)-(3-methyl-piperazin-1-yl)-phenyl-methanone hydrochloride in ethyl acetate was added water and then aqueous potassium carbonate solution was added until a pH of 9-10 was observed in the aqueous phase. The layers were separated and the organic phase was distilled to dryness to give the title compound as an oil.

LRMS (El): m/z 203 [M]⁺, 160, 105; ¹H-NMR (d₄-methanol, 300 MHz), δ: 1.05, (3 H, s br), 2.47-3.17 (m, 5 H), 3.58 (m, br, 1 H), 4.49 (d, br, 1 H), 7.37-7.49 (m, 5 H)

Solid Form data

DSC measurements were made using a TA Instruments Q1000 Differential Scanning Calorimeter. A sample of the crystalline form of Example 1 (2.822 mg) was heated at 20°C/minute, from ambient to 25O⁰C, in a non-crimped aluminium pan. Flow gas was nitrogen at 20 cm³/min. The crystalline form of 5-{(1S)-2-[(2R)- 4-benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2- carboxylic acid methylamide shows a sharp endothermic melting peak around 127°C (Figure 1)

TGA measurements were made using TA Instruments TGA2950 Hi-Res Thermogravimetric Analyser using nitrogen purge gas at a rate of 50 cm³/min from ambient to 250⁰C at a heating rate of 20°C/min. The sample size was 6.077mg. The TGA plot for the crystalline form of 5-{(1S)-2-[(2R)-4-benzoyl-2- methyl-piperazin-1-yl]-1-methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide is shown in Figure 1.

PXRD patterns for the crystalline form of 5-{(1S)-2-[(2R)-4-benzoyl-2-methyl- piperazin-1-yl]-1-methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide were determined by mounting 5mg of solid onto a silicon wafer. Radiation used was CuK_αi(λ=1.5406A) scanned through an angular range of 3° to 35° 20 with a step size of 0.014° 2Θ on a Bruker D8 powder X-ray diffractometer and Positional Sensitive Detector. Variable temperature and humidity were provided by use of an ANSYCO Sycos H-Hot Humidifier system.

PXRD patterns for the crystalline form of 5-{(1S)-2-[(2R)-4-benzoyl-2-methyl- piperazin-1-yl]-1-methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide at variable temperature and relative humidity are provided, respectively, in Figures 2 and 3.

The DSC/TGA results in Figure 1 demonstrate that the dehydration event occurs over a temperature range of 20-100 ⁰C. There is little change in the PXRD pattern over a similar temperature range (Figure 2), demonstrating that the crystalline structure is independent of water content.

The moisture sorption profile was determined by Dynamic Vapour Sorption (DVS) using a Surface Measurement Systems Ltd. DVS 1 by exposing 19.267mg of solid to a controlled relative humidity (%RH) environment and the weight change recorded over time. The humidity was changed from 45 - 0 - 90 - 0 %RH in 15% intervals. The flow gas used was nitrogen. The DVS isotherm for crystalline 5- {(1S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2-oxo-ethoxy}-4- methoxy-pyridine-2-carboxylic acid methylamide, as shown in Figure 4, shows that there is a direct correlation between % Relative Humidity (RH) and water uptake (0% RH anhydrous; 90% RH monohydrate). There is insignificant change in the PXRD pattern over a similar RH range (Figure 3), again demonstrating that the crystalline structure is independent of water content.

2Θ Angles, d-spacings and relative intensities (Table 1) were calculated from the single crystal structure of 5-{(1S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1-yl]-1- methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide using the "Reflex Powder Diffraction" module of Accelrys MS Modelling™ [version 3.0]. Pertinent simulation parameters were:

Wavelength = 1.5406 A (Cu Ka) Polarisation Factor = 0.5

Pseudo-Voigt Profile (U = 0.01 , V = -0.001 , W = 0.002)

The main characteristic PXRD peaks for crystalline 5-{(1S)-2-[(2R)-4-benzoyl-2~ methyl-piperazin-1-yl]-1-methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide are given in Table 2 below. Table 1 - Calculated PXRD peaks for the crystalline form of 5-{(1S)-2-[(2R)-4- benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2- carboxylic acid methylamide.

Table 2 - Main characteristic PXRD peaks for the crystalline form of 5-{(1S)-2- [(2R)-4-benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2-oxo-ethoxy}-4-methoxy- pyridine-2-carboxylic acid methylamide.

The calculated pattern represents that of a pure phase of the crystalline form of 5-{(1 S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1 -yl]-1 -methyl-2-oxo-ethoxy}-4- methoxy-pyridine-2-carboxylic acid methylamide since it is derived from a single crystal structure. A comparison of the measured and calculated patterns is shown in Figure 5 and demonstrates that the bulk is represented by the single crystal structure. Slight discrepancies between peak intensities can be attributed to preferred orientation effects in the measured pattern.

Biological Data The ability of 5-{(1S)-2-[(2R)-4-benzoyl-2~methyl-piperazin-1-yl]-1-methyl-2-oxo- ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide to modulate gp120 activity, in particular inhibit the interaction of gp120 with CD4, is demonstrated using a gp160 induced cell-cell fusion assay to determine the IC₅₀ values of compounds against HIV-1 fusion. The gp160 induced cell-cell fusion assay uses a HeLa P4 cell line and a CHO-Tat10 cell line.

The HeLa P4 cell line expresses CCR5 and CD4 and has been transfected with HIV-1 LTR-β-Galactosidase. The media for this cell line is Dulbecco modified eagle's medium (D-MEM) (without L-glutamine) containing 10% foetal calf serum (FCS), 2mM L-glutamine penicillin/streptomycin (Pen/Strep; 100U/mL penicillin + 10mg/mL streptomycin), and 1μg/ml puromycin.

The CHO cell line is a Tat (transcriptional trans activator)-expressing clone from a CHO JRR17.1 cell line that has been transfected with pTat puro plasmid. The media for this cell line is rich medium for mammalian cell culture originally developed at Roswell Park Memorial Institute RPMH 640 (without L-glutamine) containing 10% FCS, 2mM L-glutamine, 0.5 mg/ml Hygromycin B and 12μg/ml puromycin. The CHO JRR17.1 line expresses gp160 (JRFL) and is a clone that has been selected for its ability to fuse with a CCR5/CD4 expressing cell line.

Upon cell fusion, Tat present in the CHO cell is able to transactivate the HIV-1 long terminal repeat (LTR) present in the HeLa cell leading to the expression of the β-Galactosidase enzyme. This expression is then measured using a Fluor Ace™ β-Galactosidase reporter assay kit (Bio-Rad cat no. 170-3150). This kit is a quantitative fluorescent assay that determines the level of expression of β- galactosidase using 4-methylumbelliferul-galactopyranoside (MUG) as substrate. β-Galactosidase hydrolyses the fluorogenic substrate resulting in release of the fluorescent molecule 4-methylumbelliferone (4MU). Fluorescence of 4- methylumbelliferone is then measured on a fluorometer using an excitation wavelength of 360nm and emission wavelength of 460nm.

Compounds that inhibit fusion will give rise to a reduced signal and, following solubilisation in an appropriate solvent and dilution in culture medium, a dose- response curve for each compound can be used to calculate IC₅₀ values.

The IC₅₀ value for 5-{(1S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2- oxo-ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide is 14nM.

The ability of 5-{(1S)-2-[(2R)-4-benzoyi-2-methyl-piperazin-1-yl]-1-methyl-2-oxo- ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide to inhibit the interaction of purified recombinant HIV-1 gp120 with soluble CD4 (sCD4) can be demonstrated using Alphascreen™ technology (Perkin Elmer Life Sciences).

5-{(1S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2-oxo-ethoxy}-4- methoxy-pyridine-2-carboxylic acid methylamide is solubilised (100% DMSO (dimethylsulphoxide)) and diluted in alphascreen buffer (25mM HEPES (N-2- hydroxyethylpiperazine-N-2-ethanesulphonic acid) pH7.4, 10OmM NaCI, 0.03% TWEEN® (polyethylene glycol sorbitan monolaurate), 0.3% BSA (bovine serum albumin)) to the required concentration. 4μl of solution is added to an opaque proxiplate (Perkin Elmer Life Sciences). 4μl of purified, recombinant, histidine- tagged gp120 (BaL strain, expressed in CHO cells) is added followed by 4μl biotinylated soluble CD4 (Autogen Bioclear). Alphascreen beads (8μl) are added to each well (Perkin Elmer) and the reaction incubated for 2hrs at RT. The concentration of reagents used is batch dependent. The alphascreen beads consist of streptavidin coated donor beads, that bind to biotinylated soluble CD4, and nickel chelate acceptor beads which bind to histidine-tagged gp120. If gp120 is bound to sCD4, laser excitation of donor beads at 680nm leads to excitation of acceptor beads and emission at 520-620nm. This is measured using a Fusion α FT HP plate reader (Perkin Elmer Life Sciences). Inhibition of gp120 binding to sCD4 is measured by the decrease in emission from the acceptor beads.

5-{(1S)-2-[(2R)-4-Benzoyl-2-methyl-piperazin-1-yl]-1-methyl-2-oxo-ethoxy}-4- methoxy-pyridine-2-carboxylic acid methylamide has been found to inhibit this interaction with an IC₅₀ of 1.5μM.

Claims

Claims:

1. A crystalline form of 5-{(1 S)-2-[(2R)-4-benzoyl-2-methyl-piperazin-1-yl]-1- methyl-2-oxo-ethoxy}-4-methoxy-pyridine-2-carboxylic acid methylamide which exhibits the following characteristic powder X ray diffraction peaks when measured using Cu Ka radiation (Wavelength = 1.5406 A):

2. A crystalline form as claimed in claim 1 which is further characterised by the following characteristic powder X ray diffraction peaks when measured using Cu Ka radiation. (Wavelength = 1.5406 A):

3. A crystalline form as claimed in claim 1 or 2 which is further characterised by the following characteristic powder X ray diffraction peaks when measured using Cu Ka radiation (Wavelength = 1.5406 A):

12.1

12.4

12.9

13.9

15.1

16.2

17.9

20.2

21.0

21.7

4. A crystalline form as claimed in any one of claims 1 to 3 which is further characterised by the following characteristic powder X ray diffraction peaks when measured using Cu Ka radiation (Wavelength = 1.5406 A):

5. A crystalline form as claimed in any preceding claim which is further characterised by a DSC trace which shows a sharp endotherm at 127⁰C.

6. A pharmaceutical composition including the crystalline form as claimed in any one of claims 1 to 5 together with one or more pharmaceutically acceptable excipients, diluents or carriers.

7. The crystalline form as claimed in any one of claims 1 to 5, for use as a medicament.

8. The crystalline form as claimed in any one of claims 1 to 5 for use in the treatment of HIV, a retroviral infection genetically related to HIV, or AIDS.

9. The use of the crystalline form as claimed in any one of claims 1 to 5 for the manufacture of a medicament for the treatment of HIV, a retroviral infection genetically related to HIV, or AIDS.

10. A method of treatment of a mammal suffering from HIV, a retroviral infection genetically related to HIV, or AIDS which comprises treating said mammal with an effective amount of the crystalline form as claimed in any one of claims 1 to 5.

11. A pharmaceutical composition comprising the crystalline form as claimed in any one of claims 1 to 5 and one or more additional therapeutic agents.

12. A process for preparing the crystalline form as claimed in any one of claims 1 to 5 comprising crystallising the compound of formula (I)

from a solvent system comprising at least one ester and water.

13. The process as claimed in claim 12, wherein the ester is n-butyl acetate.

14. The process as claimed in claim 13, wherein the solvent system comprises 0.2-5% w/w water.