Introduction Background The present invention relates to compounds capable of binding to G-protein coupled receptors. In particular, libraries of compounds are provided for use in screening programmes against GPCR targets as well as the individual compounds for use in hit to lead and lead optimisation projects and similar stages in the drug discovery process.

The method also provides methods for making compounds and libraries.

As part of the process of discovering drugs or agrochemicals it is customary to screen libraries of compounds against biological targets to discover Hits'which are then further developed into Leads'and subsequently drugs or agrochemicals by using the techniques of medicinal chemistry. Accordingly the success or not of a drug or agrochemical discovery project is critically dependent on the quality of the hit and this in turn is dictated by the quality of the screening library.

Technological advances have enabled screening on a very large scale and the screening of hundreds of thousands of compounds at the start of a discovery program is routine. This, however, does entail a significant cost. The hits obtained from such screening efforts are not all of the best quality and often take a large amount of subsequent time and effort in order to get a good lead. It has been estimated that only about 25% of projects actually get to the lead optimisation stage and part

of the reason for this is the intractability of hits from high throughput screening.

Screening libraries are commonly collections of compounds from several sources. As a result, they typically contain compounds synthesised as a part of previous projects in the history of a company. With regard to drug discovery, these collections will be drug-like but are likely to be limited in scope and will be directed to certain areas of a particular project. It has been the common practice of many pharmaceutical companies in recent times to augment the collections by purchasing either single compounds from vendors or by contracting the synthesis of combinatorial libraries of compounds. The singly purchased compounds may have been selected to fill in areas of compound space poorly represented in the compound collections.

Combinatorial libraries are typically synthesised around well- performing chemistries with some design based on producing diversity'in compound space.

A complementary approach, and one that is increasingly preferred, is to screen focused libraries against the target of choice. Focused libraries are becoming of increasing importance in their ability to generate hits capable of rapid expansion in many areas including GPCRs. Such libraries are slightly more expensive to prepare but have attributes of reliability, reproducibility and provide a considerably higher hit rate: typically 10-100 fold and above compared with random screening. They are, however, very difficult to design and their efficiency relates directly to the amount of effort that has gone into the design. Using focused libraries, it is usually possible to get a number of hits in the low micromolar and below range. As there is a defined set of compounds there is the potential to observe indications of SAR in a chemical series and progress the chemistry efficiently.

G-protein-coupled receptors (GPCRs) are very important in the regulation of numerous body processes and a significant proportion of all drugs work by interaction with these receptors. There are several hundred known, many of which are orphans-those receptors that have no established ligands.

They fall into a class of 7-transmembrane receptors and there is only one X-ray structure known, that of the bovine rhodopsin receptor, and this is at a resolution of 2.8 Angstroms and is thus not suitable for accurate modeling work. In addition, the rhodopsin receptor is somewhat unusual in its interactions with its ligand and is not used as a drug target. Nevertheless, the overall three dimensional arrangement can be deduced from the X-ray and is in accordance with previous work based upon bacteriorhodopsin receptor which is not G-protein-coupled.

GPCRs are most often characterised by sequence homology as being comprised of several sub-families. Most attention currently is directed towards Family A receptors as being the most tractable class historically and also the one with the most potential targets.

Family A comprises about 300 receptors that are potential drug targets, approximately half of which have known ligands and the rest, the so-called orphan receptors. The group of druggable receptors is composed essentially of two types: those whose natural ligand interacts wholly within the transmembrane domain, such as the aminergic, nucleotide-like, prostaglandin receptors, etc. and those peptide liganded receptors, which have a large part of their interactions in the extracellular region and which may insert a peptide loop or tail into the transmembrane region to effect signal transduction. Examples of this class are angiotensin, cholecystokinin and opioid receptors. Irrespective of the mode of action of the natural

ligand or the GPCR family, the vast majority of drug molecules interact in the all-helical domain of the transmembrane region with exceptions being those mimics of glutamate at the metabotropic glutamate receptor and some peptide therapeutics administered parenterally. In looking for lead molecules for an unexplored or orphan GPCR it therefore makes sense to concentrate on interactions in the transmembrane domain.

The focused library provided herein is designed to interact with a range of the family A receptors. Each library is a defined set of compounds which will enhance the probability of finding a small molecule which will interact with one or more type of GPCR receptor.

For example, focused libraries can be provided having compounds which will interact with aminergic GPCRs, and peptidic GPCRs requiring an obligatory positive charge in ligands, or other types or groups of GPCRs.

Focused libraries according to this invention can provide hit rates of 1-13% or more for the requisite predicted GPCRs from both amine-and peptide-liganded classes and with agonists and antagonists.

Summary of Invention We provide herein a"focused"library of compounds which will provide"leads"for ligands which bind to Family A G-Protein coupled receptors.

In the context of the present invention, "library"means a group of compounds which are structurally related by virtue of

a core chemical structure (or"scaffold") but which differ from each other by virtue of permutation of specific substituent groups attached to the scaffold.

Generally speaking such a library will consist of or comprise a number of compounds, e. g. as many as about 100,1000, 2000,3000 or indeed 10,000 compounds. The number of compounds should be sufficient to provide an adequate diversity of related compounds without being so large as to be unduly complex/expensive to produce.

In the context of the present invention the terms"permitted substituents"and analogous terms are used to refer to defined chemical groups which may be attached to a"scaffold"to provide permutations of the chemical structure of related compounds.

Where the chemical formulae of permitted substituents are shown in this description and claims, the substituent may appear in the compound exactly as shown (i. e. simply covalently bonded to the scaffold) or may be a derivative of the shown chemical formula of the substituent by virtue of use of a reactive group to couple the substituent to the scaffold.

It will be appreciated that the total number of permutations created by the permitted substituents may be a very large number, far greater in magnitude than the actual number of compounds in an actual library. In other words, the number of possible compounds for any"virtual"library may well greatly exceed the number of synthesised compounds making up an embodiment of the"real"library. The invention is intended to encompass libraries having all, and a number, which is less than all, of the permitted substitutions represented by compounds therein.

It will be appreciated that some specific combinations of permitted substituents may be more or less difficult to synthesise and/or use in a focused library of the invention.

This does not detract from the generality of applicability of the invention as described herein. It is to be expected that real libraries will be synthesised from a selected group of permutations/combinations of permitted substituents, taking into consideration factors affecting the intended purpose of the library and its cost and complexity of synthesis.

Even if theoretically permitted, it is currently considered unlikely that any compound would be prepared for inclusion in a focused library if it had either or both of the following properties (1) molecular weight >700 (2) log p <-3 or >9 (an index of lipophilicity as calculated using commercially available"Chemenlighten 2.8" and"Biobyte"software for the log p calculation).

The present invention provides novel focused libraries of compounds. Most of the compounds defined by the permitted substitutions on the scaffolds are also novel compounds per se and the invention is intended to encompass each individual novel compound. Any known compound having a structural formula identical to any one of the compounds covered by the formulae of scaffolds and permitted substitutions described herein is hereby explicitly disclaimed per se.

Description of the Invention Library 1 (SFGO1) is a broad-spectrum library targeted largely at receptors with a requirement for a positively charged amine

in their structure activity relationships but also has features that make it a particularly good all round library. It is designed to produce both agonists and antagonists and so is expected to be especially useful in producing ligands for orphan receptors.

The central design of the library revolves around an acylurea coupled to a piperidine moiety. A combination of specific motifs R2 and R1 are appended from the central scaffold and are designed to pick up different interactions at a receptor site.

The invention provides a compound library comprising or consisting of a set of structurally related compounds of general formula (I) : wherein R1 and R2 are independently hydrogen, optionally substituted alkyl, aryl, heteroaryl or heterocyclyl.

Structural Novelty of Compounds of Library 1 Eight compounds are available for sale from Maybridge (see below). However none of these compounds are permitted in Library 1.

One other compound is mentioned in the literature. This structure could be contained in Library 1 (SFG01) but is disclaimed per se.

Methods for Synthesising Compounds of Library 1 Compounds of formula I can be made according to the following general scheme: R2 & R1 = H, Alkyl, aryl, hetero aryl or heterocyclyl A general scheme (Scheme 1) for introducing substituents to produce compounds of Library 1 (SFG01) is as follows:

Scheme 1 Methyl isonipecotate (1) can be acylated, sulphonylated or alkylated with an appropriate reagent (Rl-&num ) selected from List 1, wherein # = leaving group. Examples of leaving groups are halogen or sulphonate.

The urea compounds (3) can be formed from the reaction of potassium cyanate with an amine (R2-NH#) from List 2, wherein # = H.

The resultant intermediate compound (2) can then be reacted with the intermediate urea compound (3) to form the desired compounds of formula I.

The core chemical scaffold is formed at the final stage of this reaction scheme by the reaction of the two intermediate compounds, which have already had the permitted substituents R1 and R2 added to each intermediate compound respectively.

In the priority application (GB 0230195.0) where-OH groups are shown in place of #s, the-OH groups may be modified to form leaving groups.

The permitted substituents at positions R1 and R2 for compounds of Libray 1 (SFG01) are shown below.

List 1 List2

Specific Examples of Compounds of Library 1 Typical example of compound of Formula 2, as described in Scheme 1 ; Methyl 1- (2-phenoxyethyl) piperidine-4-carboxylate.

To a stirred solution of methyl isonipecotate (4. 00g, 28.00 mmol) and phenoxyethyl bromide (4. 00g, 20.00 mmol) in acetonitrile (30 ml) was added sodium hydrogen carbonate (7. 00g, 84.00 mmol) and the reaction stirred at 80°C for 20 hours. The reaction was cooled, filtered and concentrated under reduced pressure to give a crude brown oil. Purification by column chromatography (7% methanol: dichloromethane) gave the required compound as a light brown oil (4.8g, 65%). oH (270 MHz; CDCl3 ; Me4Si), 1.71-2. 37 (7H, m), 2.79 (2H, t, J 6. 0), 2.93-3. 0 (2H, m) 3.68 (3H, s), 4.10 (2H, t, J 6.0), 6.89-6. 97 (3H, m), 7.24-7. 32 (2H, m). HPLC 99%, m/z (APCI), 264.20 (100% [M+H] +), 232 (40, C14Hl802N), 170 (47, C9H1602N).

Typical example of compound of formula 2, as described in Scheme 1; Methyl 1-[2-(2naphthyloxy) ethyl] piperidine-4- carboxylate.

To a stirred solution of 2- (2-naphthoxy) ethanol (1.28g, 6.58 mmol), methyl isonipecotate (0.93g 6.50 mmol) and DIPEA (1.09g, 8.40 mmol) in propionitrile (13 ml) was added trialkylphosphium iodide (1.90g 7.82 mmol) and the reaction stirred at 90°C for 3 hours. The mixture was cooled to room temperature and a saturated solution of potassium carbonate (20 ml) was added.

The product was extracted with DCM (3 x 30 ml) and washed with brine, dried (MgS04) and concentrated to give a brown oil.

Purification by column chromatography (29% DCM, 29% ethyl acetate, hexane) gave the required compound as a brown solid (1. 75g, 75%) ; 6H (270 MHz; CDCl3 ; Me4Si), 1.73-1. 96 (4H, m), 2.18-2. 34 (3H, m), 2.87 (2H, t, J 6.0), 2.97-3. 03 (2H, m), 3. 68 (3H, s), 4.22 (2H, t, J 6.0), 7 13-7. 46 (3H, m), 7.70-7. 83 (4H, m). HPLC 99%, m/z (ES) 314 (100% [M+H] +), 170 (50, CgH1602N).

Typical example of compound of formula 2, as described in Scheme 1; Methyl 1-acetyl-4-carboxylate.

To a stirred solution of 1-acetyl piperidine-4-carbonyl chloride hydrochloride (l. OOg, 4.40 mmol) in methanol (20 ml) was added triethylamine (1.36g, 13.00 mmol) at 0°C. The reaction was warmed to room temperature and stirred for a further 20 hours, concentrated and diluted with water. The product was extracted with DCM (3 x 30 ml), dried (MgS04) and concentrated under reduced pressure to give a brown oil (0.80g, 98%); su (270 MHz; CDCl3 ; Me4Si), 1.64-1. 76 (2 H, m), 1.84 (1 H, s), 1.89-2. 09 (2 H, m), 2.09 (3 H, s), 2.49-2. 55 (1 H, m) 2.74- 2.85 (1 H, m), 3.08-3. 18 (1 H, m), 3.70 (3 H, s), 3.74-3. 83 (1 H, m), 4.37-4. 45 (1 H, m), m/z (ES) 186 (100%, [M+H] +), 154 (25, C8H1202N), 144 (62, C7H1202N).

Typical example of compound of formula 2, as described in Scheme 1; Methyl 1-methylpiperidine-4-carboxylate.

To a stirred solution of methyl isonipecotate (l. OOg, 7.00 mmol) in methanol (25 ml) was added formic acid (5.52g, 12.00 mmol) and a solution of formaldehyde (2.67g, 35.00 mmol, 40% aqueous) and the reaction heated at reflux. After 3 hours the reaction was cooled and concentrated under reduced pressure.

The aqueous solution was basified with NaHC03 and extracted with DCM (3 x 30 ml), dried (MgSO4) and concentrated under reduced pressure to give a brown liquid (l. OOg, 910-.) ; b (270 MHz; CDC13 ; Me4Si), 1.69-2. 03 (6 H, m), 2.23-2. 32 (4H, m), 2.78-

2.83 (2H, m), 3.68 (3 H, s); m/z (APCI) 158 (100% [M+H] +), 126 (33, C7H120N).

Typical example of compound of formula 3, as described in Scheme 1; N- (4-trifluoromethylbenzyl) urea.

To a stirred aqueous solution (60 ml) of 4- (trifluoromethyl) benzyl amine (4.28 ml, 0.03 mol) was added conc. HC1 (3 ml) and potassium cyanate (2.67g, 0.033 mol) in water (10 ml) and the solution heated at 90°C for 2 hours. The reaction was cooled to room temperature and the resultant solid filtered off to give a white crystalline solid (5. 03g, 80%); 6H (270 MHz; DMSO-d6), 4.24 (2H, d, J 6.0), 5.60 (1H, s), 6.52 (1H, m), 7.42 (2H, d, J 8.3), 7.66 (2H, d, J 8.3) ; HPLC 94. 7% ; m/z (ES) 218 (100%, [M+H] +, 158 (45, C8H6F3).

Typical example of compound of formula 3, as described in Scheme 1; N- (4-ethoxyphenyl) urea.

To a stirred solution of 4-ethoxyphenyl isocyanate (1.63g, 10.00 mmol) in anhydrous THF (12 ml) at-78°C was added dropwise ammonia (10 ml) and the reaction warmed and stirred at room temperature. After 18 hours the reaction was concentrated to give a crude product which upon recrystalisation (ethyl acetate : methanol 8: 2) gave the required compound as a white solid, (1.26g, 70%); 6H (270 MHz; DMSO-d6), 1.27 (3H, t, J 6.8), 3.90 (2H, q, J 6.8), 5.70 (2H, s), 6.78 (2H, d, J 9.0), 7.24 (2H, d, J 9.0), 8.28 (1H, s); HPLC 98%, m/z (ES) 181 (100%, [M+H] +).

Standard Synthesis of compounds of formula (I) ; To a stirred solution of the urea (3) (0.20 mmol) in DMSO (0.5 ml) was added potassium tert-butoxide (0.40 mmol) as a DMSO solution and the reactions shaken at room temperature. After 15 minutes a solution of the ester (2) (0.2 mmol, DMSO) was added and the contents shaken for a further 18 hours. The reactions were subsequently filtered over amberliteTM-IR-120 (H) resin and purified by preparative chromatography using the following conditions: Typical examples of compound of formula (I), as described in Scheme 1 ; { [ (4-butoxyphenyl) amino]carbonyl}-1-{2-(3- methylphenyl) ethyl] piperidine-4-carboxamide

Yield 26.6mg, 30%; 6H (270 MHz; DMSO-d6) 1.01 (3H, t, J 7.3), 1.50 (2H, q of t, J 7.3, 6.8), 1.76 (2H, t of t, J 6.8, 6.3), 1.89-1. 99 (2H, m), 2.14-2. 19 (2H, m), 2.38 (3H, s), 2.76-2. 78 (1H, m), 3.00-3. 06 (2H, m), 3.33-3. 41 (2H, m), 3.50-3. 53 (2H, m), 3.70-3. 76 (2H, m), 4.01 (2H, t, J 6.3), 6.97 (2H, d, J 9.03), 7.13-7. 17 (3H, m), 7.29-7. 32 (1H, m), 7.50 (2H, d, J 9.03) 10.35 (1H, s), 10.88 (1H, s); HPLC 100%, m/z (ES) 438 (100%, [M+H] +).

Typical example of compound of formula (I), as described in Scheme 1; N-{[(4-isopropylphenyl) amino] carbonyl}-1-(3- phenoxypropyl) piperidine-4-carboxamide.

Yield 25. 1mg, 30%; Sn (270 MHz; DMSO-d6) 1.26 (6H, d, J 6.8), 1.92-1. 93 (2H, m), 2.11-2. 24 (4H, m), 2.60-2. 62 (1H, m), 2.93 (1H, q, J 6.8), 3.01-3. 08 (2H, m), 3.32-3. 38 (H, m), 3.00-3. 06 (2H, m), 3.33-3. 41 (2H, m), 3.50-3. 53 (2H, m), 3.68-3. 74 (2H, m), 4.13 (2H, t, J 5.9), 7.03 (2H, d, J 8.5), 7.04-7. 06 (1H, m), 7.26-7. 30 (2H, m), 7.36-7. 39 (2H, m), 7.50 (2H, d, J 8.5) 10.45 (1H, s), 10.91 (1H, s); HPLC 96%, m/z (ES) 424 (96%, [M+H] +).

Typical example of compound of formula (I), as described in Scheme 1; 1-Acetyl-N-{[(2-ethoxyphenyl) amino] carbonyl} piperidine-4-carboxamide.

Yield 8.6mg, 13%; 6H (270 MHz; DMSO-d6) 1.30-1. 33 (1H, m), 1.33 (3H, t, J 6.8), 1.76-1. 78 (1H, m), 1.79-1. 81 (2H, m), 1.98 (3H, s), 2.47-2. 49 (1H, m), 2.50-2. 52 (1H, m), 2.97-2. 99 (1H, m), 3.31-3. 32 (1H, m), 4.08 (2H, q, J 6.8), 4.38-4. 39 (1H, m), 6.88-7. 02 (3H, m), 8.13-8. 17 (1H, m), 10.70 (1H, s), 10.96 (1H, s); HPLC 98%, m/z (ES) 334 (96%, [M+H] +).

Typical example of compound of formula (I), as described in Scheme 1; 1-Acetyl-N-L (4-£luorophenyl) amino] carbonyl} piperidine-4-carboxamide.

Yield 8.3mg, 14%; dH (270 MHz; DMSO-d6) 1.73-1. 75 (1H, m), 1.75- 7.77 (1H, m), 1.78-1. 80 (2H, m), 1.98 (3H, s), 2.47-2. 49 (1H, m), 2.50-2. 52 (1H, m), 2.97-2. 99 (1H, m), 3.81-3. 86 (1H, m), 4.34-4. 39 (1H, m), 7.11-7. 19 (2H, m), 7.51-7. 56 (2H, m), 10.48 (1H, s), 10.74 (1H, s); HPLC 98%, m/z (ES) 308 (90%, [M+H] +).

Analytical HPLC conditions Mobile phase.

0. 2% TFA/water, ACN Flow rate 25 ml/min.

Gradient: 85/15 H20 + 0. 2% for 1.5 min.

TFA/ACN 5/95 in 9.5 min. for 1.5 min.

85/15 in 0.5 min.

Detector: ELS. (approx. 1. 5ml/min flow split to Sedex 55 ELSD) Gas (Nitrogen) 2.0 bar Nebulizer 40°C Column: Waters SymmetryPrep 19mm x 150m x 7mm C18