WO2007028445A1 - 6-indolyl-4-yl-amino-5-halogeno-2-pyrimidinyl-amino derivatives - Google Patents

Abstract

Bis Anilinopyrimidine compounds of formula (I) are useful as inhibitors of Syk kinase and thus useful in treating diseases resulting from inappropriate mast cell activation, for instance allergic and inflammatory diseases.

Description

6-INDOLYL-4-YL-AMINO-5-HALOGENO-2-PYRIMIDINYL-AMINO DERIVATIVES

The present invention relates to novel chemical compounds which have activity against the spleen tyrosine kinase (Syk kinase), processes for their preparation, pharmaceutically acceptable formulations containing them and their use in therapy.

Allergic rhinitis and asthma are diseases associated with hypersensitivity reactions and inflammatory events involving a multitude of cell types including mast cells, eosinophils, T cells and dendritic cells. Following exposure to allergen, high affinity immunoglobulin receptors for IgE (FcεRI) and IgG (FcγRI) become cross-linked and activate downstream processes in mast cells and other cell types leading to the release of pro-inflammatory mediators and airway spasmogens. In the mast cell, for example, IgE receptor cross-linking by allergen leads to release of mediators including histamine from pre-formed granules, as well as the synthesis and release of newly synthesised lipid mediators including prostaglandins and leukotrienes.

Syk kinase is a non-receptor linked tyrosine kinase which is important in transducing the downstream cellular signals associated with cross-linking FcεRI and or FcγRI receptors, and is positioned early in the signalling cascade. In mast cells, for example, the early sequence of FcεRI signalling following allergen cross-linking of receptor-lgE complexes involves first Lyn (a Src family tyrosine kinase) and then Syk kinase. Inhibitors of Syk kinase activity would therefore be expected to inhibit all downstream signalling cascades thereby alleviating the immediate allergic response and adverse events initiated by the release of pro-inflammatory mediators and spasmogens (Wong, B., Grossbard, E. B. Payan, D. G & Masuda, E. S. Expert Opin. Investig. Drugs (2004) 13 (7) 743-762).

Recently, it has been shown that the Syk kinase inhibitor R112 (Rigel), dosed intranasally in a phase I/I I study for the treatment of allergic rhinitis, gave a statistically significant decrease in PGD₂, a key immune mediator that is highly correlated with improvements in allergic rhinorrhea, as well as being safe across a range of indicators, thus providing the first evidence for the clinical safety and efficacy of a topical Syk kinase inhibitor. Meltzer, EIi O.; Berkowitz, Robert B.; Grossbard, Elliott B. An intranasal Syk -kinase inhibitor ( R112 ) improves the symptoms of seasonal allergic rhinitis in a park environment. Journal of Allergy and

Clinical Immunology (2005), 115(4), 791-796. In a more recent phase Il clinical trial for allergic rhinitis (Clinical Trials.gov Identifier NCT0015089), R112 was however shown as having a lack of efficacy versus placebo.

Rheumatoid Arthritis (RA) is an auto-immune disease affecting approximately 1% of the population. It is characterised by inflammation of articular joints leading to debilitating destruction of bone and cartilage. Recent clinical studies with Rituximab, which causes a reversible B cell depletion, (J. CW. Edwards et al 2004, New Eng. J. Med. 350: 2572-2581 ) have shown that targeting B cell function is an appropriate therapeutic strategy in auto-immune diseases such as RA. Clinical benefit correlates with a reduction in auto-reactive antibodies (or Rheumatoid Factor) and these studies suggest that B cell function and indeed auto-antibody production are central to the ongoing pathology in the disease.

Studies using cells from mice deficient in the Syk kinase have demonstrated a non- redundant role of this kinase in B cell function. The deficiency in Syk kinase is characterised by a block in B cell development (M. Turner et al 1995 Nature 379: 298-302 and Cheng et al 1995, Nature 378: 303-306). These studies, along with studies on mature B cells deficient in Syk kinase (Kurasaki et al 2000, Immunol. Rev. 176:19-29), demonstrate that Syk kinase is required for the differentiation and activation of B cells. Hence, inhibition of Syk kinase in RA patients is likely to block B cell function and hence to reduce Rheumatoid Factor production. In addition to the role of Syk kinase in B cell function, of relevance to the treatment of RA, is the requirement for Syk kinase activity in Fc receptor (FcR) signalling. FcR activation by immune complexes in RA has been suggested to contribute to the release of multiple pro-inflammatory mediators.

The contribution of Syk kinase dependent processes to the pathology of RA has been reviewed in Wong et al (2004, ibid).

WO 03/057695 (Boehringer lngelheim Pharmaceuticals, Inc.) describes substituted [1 ,6]-naphthyridines that inhibit Syk kinase.

WO2003/063794, WO2004/014382, WO2005/012294 and WO2005/16893 (Rigel Pharmaceuticals, Inc) describes a series of 2,4-pyrimidinediamine compounds which inhibit Syk kinase, for use in treating autoimmune diseases.

WO2005/026158 (Novartis AG) describes 2,4-di (hetero)-arylamino-pyrimidine derivatives which have ZAP-70 and/or Syk inhibitory activities.

WO 04/035604 discloses the structural co-ordinates of the human Syk protein.

There remains however the need to identify further compounds which are inhibitors of Syk kinase.

Thus, the present invention provides a compound of formula (I):

in which:

R¹, R² and R³ is each independently selected from hydrogen, halogen, -Co.₆hydroxy, -C_1-6 alkyl, -NR⁵R⁶, -CN, -Co-aalkylene-COzH, OC(O)C_1-6 alkyl, C(O)C_1-6 alkoxy, -C_1-2 alkyl substituted by 1 or more fluorine atoms, -C_1-6 alkoxy, -C(O)NR⁵R⁶, -OCH₂C(O)NR⁵R⁶, -NR⁵C(O)R⁶, -SC_1-6alkyl, -S(O)C_1-6alkyl, -S(O)₂C_1-6alkyl,- NHS(O)₂R⁷, -S(O)₂NR⁵R⁶ or -S(O)₂NR⁵R⁸, such that at least one of R¹, R² and R³ is hydrogen; or

R¹ is hydrogen and one of R² and R³ is hydrogen, C_1-6alkyl, or C_1-6alkoxy and the other is a 5- or 6-membered heteroaryl ring comprising from 1 to 3 heteroatoms selected from O, N and S bonded to the phenyl ring through a ring carbon atom and which heteroaryl ring is unsubstituted or substituted on one or more ring carbon atoms by C_1-6alkyl or oxo (=0), or on a ring nitrogen atom by C_1-6alkyl; or

R¹ is hydrogen, halogen or C_1-6 alkyl and R² and R³ are joined and together form, in combination with the carbon atoms on the phenyl group to which they are attached, a 5- or 6-membered ring wherein: the ring is heterocyclic and comprises one heteroatom selected from O, N or S, the ring is heteroaryl or heterocyclic and comprises two heteroatoms selected from O, N or S, which maybe the same or different, and excluding two S atoms, the ring is heteroaryl and comprises three heteroatoms of which two are N atoms and the third is nitrogen or oxygen, which ring may be unsubstituted or substituted on: one or more ring carbon atoms which each may be independently substituted by oxo (=0) or by 1 or 2 substituents which may be the same or different selected from C_1-6 alkyl, hydroxyl, or halo, one or more ring nitrogen atoms by C_1-6 alkyl, CH₂C_3-7cycloalkyl, hydroxy C_1-6 alkyl, C_Lβalkylcarbonyl, or C_1-6haloalkylcarbonyl, and a ring sulphur atom by (O)₂; and

R⁴ is C_1-3 alkyl;

R⁵ and R⁶ are is each independently hydrogen or C_halky!, or R⁵ and R⁶ together with the nitrogen atom to which they are attached form a 5- or 6-membered heterocyclic ring which may comprise a second heteroatom selected from O, S or N and which may be substituted on the second N, if present, by C_1-6alkyl, and which may be substituted on a ring carbon by oxo (=0),

(which may be the same or different), or halogen;

R⁷ is C_1-6alkyl or phenyl optionally substituted by C_1-6alkyl;

R⁸ is C_3-7cycloalkyl, C^alkyl (optionally terminally substituted with hydroxyl or tetrahydrofuranyl); and

X is halogen; or

a salt or solvate, preferably a pharmaceutically acceptable salt or solvate, thereof.

Compounds of the present invention are useful as inhibitors of Syk kinase and thus useful in treating diseases resulting from inappropriate mast cell activation, for instance allergic and inflammatory diseases.

In a further aspect, the present invention provides for a compound of formula (I) as hereinbefore defined, and excluding a compound of the formula (IA):

in which:

R^9a and R^1Oa is each independently hydrogen, C_1-6alkyl or CH₂C_3-7cycloalkyl;

R^4a is C_1-3 alkyl; and

X^a is halogen.

In a further embodiment, two of R¹, R² and R³ are hydrogen.

In a further embodiment, representative values of R¹, R² and R³ include: hydroxy, - NH₂, methyl, -OCOCH₃, -OCH₃, -C₂H₄OH, -CONH₂, -CONHCH₃, -CONH(C₂H₅), - SO₂NH₂, -SO₂NHR⁸ and -SO₂CH₃

In a further embodiment, R¹ and R³ is each hydrogen and R² is a 5- or 6-membered heteroaryl ring or a heterocyclic ring comprising from 1 to 3 heteroatoms selected from O, N and S bonded to the phenyl ring through a ring carbon or nitrogen (if present) atom and which heterocyclic ring is unsubstituted or substituted on one or more ring carbon atoms by d_-6alkyl or oxo (=0), or on a ring nitrogen atom by C_1-6alkyl. In a further embodiment, R¹ and R³ is each hydrogen and representative examples of R² include:

In a further embodiment, R¹ and R² is each hydrogen and R³ is a 5- or 6-membered heteroaryl ring or a heterocyclic ring comprising from 1 to 3 heteroatoms selected from O, N and S bonded to the phenyl ring through a ring carbon or nitrogen (if present) atom and which heterocyclic ring is unsubstituted or substituted on one or more ring carbon atoms by C_1-6alkyl or oxo (=0), or on a ring nitrogen atom by C^alkyL

In a further embodiment, R¹ and R² is each hydrogen and representative examples of

R include:

In a further embodiment, representative examples of the 5- or 6- membered saturated or unsaturated ring formed by R² and R³, fused with the phenyl ring, include:

In a further embodiment, representative examples of the 5- or 6- membered saturated or unsaturated ring formed by R² and R³, fused with the phenyl ring, and including any substituents which may be present, include:

In a further embodiment, R⁴ is ethyl or propyl.

In a further embodiment, representative examples of R⁵ and R⁶ include: H, methyl, and ethyl.

In a further embodiment, representative examples of R⁸ include cyclopropyl, hydroxyethyl and tetrahydro-2-furanylmethyl

In a further embodiment, representative examples of X include fluoro, chloro and bromo.

In a further embodiment, X is fluoro. Representative examples of compounds of formula (I) include: /^-(i .i-dioxido^.S-dihydro-i-benzothien-e-yO-Λ^-ethyl-S-fluoro-Λ^-IH-indazol^-yl- 2,4-pyrimidinediamine; 3-({4-[ethyl(1/-/-indazol-4-yl)amino]-5-fluoro-2-pyrimidinyl}amino)benzene sulfonamide;

3-[6-({4-[ethyl(1 /-/-indazol-4-yl)amino]-5-fluoro-2-pyrimidinyl}amino)-1 H-indazol-1 -yl]- 1-propanol;

Λ/²-1 H-1 ,2,3-Benzotriazol-5-yl-/V⁴-ethyl-5-fluoro-/V⁴-1 H-indazol-4-yl-2,4- pyrimidinediamine; or salt or solvate thereof, in particular, a pharmaceutically acceptable salt or solvate thereof.

It will be understood that compounds of formula (I) may exist in alternative tautomeric forms, for example when R² represents a non-hydrogen substituent on the nitrogen atom in the 2-position.

When used herein, and unless otherwise indicated, the term "heteroaryl" includes single or fused rings comprising up to four hetero-atoms in the ring selected from oxygen, nitrogen and sulphur and optionally substituted with up to three substituents. Representative heteroaryl rings comprise from 4 to 7, preferably 5 or 6, ring atoms. A fused heteroaryl ring system may include carbocyclic rings and need only include one heterocyclic ring.

When used herein, and unless otherwise indicated, the term "heterocyclic" includes non-aromatic single or fused rings comprising up to four hetero-atoms in the ring selected from oxygen, nitrogen and sulphur and optionally substituted with up to three substituents. Representative heterocyclic rings comprise from 4 to 7, preferably 5 to 6, ring atoms. A fused heterocyclic ring system may include carbocyclic rings and need only include one heterocyclic ring.

Examples of substituents for such heteroaryl and heterocyclic rings include C_1-6 alkyl, hydroxyC_1-6alkyl, C_1-6alkylcarbonyl, halogen, C₁^aIkOXy, cyano, hydroxy, nitro, amino, -N(CHa)₂, -NHC(O)C^alkyl, -CF₃, -C(O)OC_1-6 alkyl, -C(O)NR¹⁹R²⁰ (wherein R¹⁹ and R²⁰ independently represent hydrogen or C_1-6 alkyl, e.g., methyl), -S(O)₂N(CH₃)₂, or - S(O)₂CH₃, and, for heterocyclic, oxo (=0).

Examples of "-C_1-2 alkyl substituted by 1 or more fluorine atoms" include, but is not restricted to, -CF₃. References to alkyl include references to both straight chain and branched chain aliphatic isomers of the corresponding alkyl. It will be appreciated that references to alkylene and alkoxy shall be interpreted similarly.

References to C_3-7 cycloalkyl include references to all alicyclic (including branched) isomers of the corresponding alkyl.

When used herein, the term "pharmaceutically acceptable" refers to those compounds, materials, compositions, and dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, or other problem or complication, commensurate with a reasonable benefit/risk ratio. The skilled artisan will appreciate that pharmaceutically acceptable salts of the compound of the present invention may be prepared. As used herein, the term "pharmaceutically acceptable salts" refers to salts that retain the desired biological activity of the subject compound and exhibit minimal undesired toxicological effects. These pharmaceutically acceptable salts may be prepared in situ during the final isolation and purification of the compound, or by separately reacting the purified compound in its free acid or free base form with a suitable base or acid, respectively. Indeed, in certain embodiments of the invention, pharmaceutically acceptable salts may be preferred over the respective free base or free acid because such salts impart greater stability or solubility to the molecule thereby facilitating formulation into a dosage form.

In certain embodiments, the compound of the present invention may contain one or more acidic functional groups. Suitable pharmaceutically acceptable salts include salts of such acidic functional groups. Representative salts include pharmaceutically acceptable metal salts such as sodium, potassium, lithium, calcium, magnesium, aluminum, and zinc salts; carbonates and bicarbonates of a pharmaceutically- acceptable metal cation such as sodium, potassium, lithium, calcium, magnesium, aluminum, and zinc; pharmaceutically acceptable organic primary, secondary, and tertiary amines including aliphatic amines, aromatic amines, aliphatic diamines, and hydroxy alkylamines such as methylamine, ethylamine, 2-hydroxyethylamine, diethylamine, triethylamine, ethylenediamine, ethanolamine, diethanolamine, and cyclohexylamine.

Compounds of the present invention are basic and accordingly generally capable of forming pharmaceutically acceptable acid addition salts by treatment with a suitable acid. Suitable acids include pharmaceutically acceptable inorganic acids and pharmaceutically acceptable organic acids. Representative pharmaceutically acceptable acid addition salts include hydrochloride, hydrobromide, nitrate, methylnitrate, sulfate, bisulfate, sulfamate, phosphate_A acetate, hydroxyacetate, phenylacetate, propionate, butyrate, isobutyrate, valerate, maleate, hydroxymaleate, acrylate, fumarate, malate, tartrate, citrate, salicylate, p-aminosalicyclate, glycollate, lactate, heptanoate, phthalate, oxalate, succinate, benzoate, o-acetoxybenzoate, chlorobenzoate, methylbenzoate, dinitrobenzoate, hydroxybenzoate, methoxybenzoate, mandelate, tannate, formate, stearate, ascorbate, palmitate, oleate, pyruvate, pamoate, malonate, laurate, glutarate, glutamate, estolate, methanesulfonate (mesylate), ethanesulfonate (esylate), 2-hydroxyethanesulfonate, benzenesulfonate (besylate), p-aminobenzenesulfonate, p-toluenesulfonate (tosylate), and napthalene-2-sulfonate.

As used herein, the term "compound" refers to one or more compounds. Similarly, the term "a compound of the present invention" refers to one or more compounds of the present invention.

The compound of the present invention may exist in solid or liquid form. In the solid state, the compound of the present invention may exist in crystalline or non- crystalline form, or as a mixture thereof. For a compound of the present invention that is in crystalline form, the skilled artisan will appreciate that pharmaceutically acceptable solvates may be formed wherein solvent molecules are incorporated into the crystalline lattice during crystallization. Solvates may involve non-aqueous solvents such as, but not limited to, ethanol, isopropanol, n-butanol, i-butanol, acetone, tetrahydrofuran, dioxane, DMSO, acetic acid, ethanolamine, and ethyl acetate, or they may involve water as the solvent that is incorporated into the crystalline lattice. Solvates wherein water is the solvent incorporated into the crystalline lattice are typically referred to as "hydrates." Hydrates include stoichiometric hydrates as well as compositions containing variable amounts of water. The invention includes all such solvates.

The skilled artisan will further appreciate that certain compounds of the invention that exist in crystalline form, including the various solvates thereof, may exhibit polymorphism (i.e. the capacity to occur in different crystalline structures). These different crystalline forms are typically known as "polymorphs." The invention includes all such polymorphs. Polymorphs have the same chemical composition but differ in packing, geometrical arrangement, and other descriptive properties of the crystalline solid state. Polymorphs, therefore, may have different physical properties such as shape, density, hardness, deformability, stability, and dissolution properties. Polymorphs typically exhibit different melting points, IR spectra, and X-ray powder diffraction patterns, which may be used for identification. The skilled artisan will appreciate that different polymorphs may be produced, for example, by changing or adjusting the reaction conditions or reagents, used in making the compound. For example, changes in temperature, pressure, or solvent may result in polymophs. In addition, one polymorph may spontaneously convert to another polymorph under certain conditions. The compound of the present invention may contain one or more asymmetric centers (also referred to as a chiral centre) and may, therefore, exist as individual enantiomers, diastereoisomers, or other stereoisomeric forms, or as mixtures thereof. Chiral centers, such as chiral carbon atoms, may also be present in a substituent such as an alkyl group. Where the stereochemistry of a chiral centre is not specified the structure is intended to encompass any stereoisomer and all mixtures thereof. Thus, the compound of the present invention containing one or more chiral centres may be used as racemic mixtures, enantiomerically enriched mixtures, or as enantiomerically pure individual stereoisomers. Generally it is preferred to use a compound of formula (I) in the form of a purified single enantiomer.

Individual stereoisomers of a compound according to Formula (I) which contain one or more asymmetric centre may be resolved by methods known to those skilled in the art. For example, such resolution may be carried out (1) by formation of diastereoisomeric salts, complexes or other derivatives; (2) by selective reaction with a stereoisomer-specific reagent, for example by enzymatic oxidation or reduction; or (3) by gas-liquid or liquid chromatography in a chiral enviomment, for example, on a chiral support such as silica with a bound chiral ligand or in the presence of a chiral solvent. The skilled artisan will appreciate that where the desired stereoisomer is converted into another chemical entity by one of the separation procedures described above, a further step is required to liberate the desired form. Alternatively, specific stereoisomers may be synthesized by asymmetric synthesis using optically active reagents, substrates, catalysts or solvents, or by converting one enantiomer to the other by asymmetric transformation.

The compound of the present invention may also contain double bonds or other centres of geometric asymmetry. Where the stereochemistry of a centre of geometric asymmetry is not specified, the structure is intended to encompass the trans (E) geometric isomer, the cis (Z) geometric isomer, and all mixtures thereof. Likewise, all tautomeric forms are also included the compound of the present invention whether such tautomers exist in equilibrium or predominately in one form.

Compounds of the present invention, as well as salts and solvates thereof, may be prepared by one or more of the general synthetic schemes described hereinafter. Scheme 1A

Scheme 1B

Scheme 1C

t-butyl nitrite

in which schemes R¹, R², R³ R⁴ and X are as hereinbefore defined and Pr is a protecting group such as 2-(trimethylsilyl)ethoxymethyl or 1 ,1- dimethylethylcarboxylate.

Accordingly, in a further aspect, the present invention provides a process for preparing a compound of formula (I), or a salt or solvate thereof, which process comprises:

reacting a compound of formula (II):

or a protected derivative thereof wherein L¹ represents a suitable leaving group, with a compound of formula (III)

or a protected derivative thereof, wherein X, R¹, R², R³ and R⁴ are as defined above.

The process may be performed in the presence of a solvent (for example, aqueous acetone or propan-2-ol), in the presence of concentrated hydrochloric acid, at a suitable temperature, preferably in the range of 0-170° C.

Alternatively, with a 2-(trimethylsilyl)ethoxymethyl protecting group, the process may be performed in the presence of a solvent such as 2-propanol, and hydrogen chloride in ether, at a suitable temperature, preferably in the range of 0-100° C, preferably under a nitrogen atmosphere.

Alternatively, with a 1 ,1-dimethylethylcarboxylate protecting group, the process may be performed in the presence of a solvent such as 2-propanol, and a dilute acid such as dilute hydrochloric acid, at a suitable temperature, preferably in the range of 0- 100° C.

Examples of suitable leaving groups (L¹) include a halide such as chloride, bromide or iodide. Other leaving groups include, but are not restricted to, an alkyl and aryl sulfide, an alkyl and aryl sulfinyl, an alkyl and aryl sulfonyl, and an alcohol derived leaving group (such as triflate, mesylate, methylsulfonate or tosylate).

Alternatively the process may be performed in the presence of a non-nucleophilic base such as triethylamine or di-isopropylethylamine, as a melt, at a suitable temperature, preferably in the range of 0-180° C.

Examples of protecting groups and the means for their removal can be found in T. W. Greene 'Protective Groups in Organic Synthesis' (J. Wiley and Sons, 1991). Suitable amine protecting groups include, but are not restricted to, sulphonyl (such as tosyl), acyl (such as benzyloxycarbonyl or t-butoxycarbonyl) and arylalkyl (such as benzyl), which may be removed by hydrolysis or hydrogenolysis as appropriate. Other suitable amine protecting groups include trifluoroacetyl (-C(O)CF₃), which may be removed by base catalysed hydrolysis, or a solid phase resin bound benzyl group, such as a Merrifield resin bound 2,6-dimethoxybenzyl group (Ellman linker) which may be removed by acid catalysed hydrolysis (using, for example, trifluoroacetic acid). Compounds of formula (II) may be prepared from the corresponding compound of formula (IV):

wherein L¹, and X are as hereinbefore defined, or a protected derivative thereof, by reacting with a sodium hydride in DMF, followed by adding alkyl iodide R⁴I, at a temperature of about 0° C, under a nitrogen atmosphere.

Compounds of formula (IV) may be prepared by reacting a corresponding (protected) indazole-4-amine with a pyrimidine compound of formula (V):

in which L² is a leaving group such as chloro and L¹ and X are as hereinbefore defined; in a solvent such as propan-2-ol or acetone in the presence or a base such as di-isopropylethylamine, and at a temperature in the range 120-180° C.

Compounds of formula (II) may also be prepared by reacting a compound of formula (Vl):

in which R⁴, L¹ and X are as hereinbefore defined, or a protected derivative thereof, with an organic nitrite derivative, such as, but not limited to, tert-butyl nitrite. This reaction is preferably performed in the presence of an acid, such as acetic acid or hydrochloric acid. The aforementioned reaction may be performed in the presence of acetic anhydride to yield a product in which an indazolyl nitrogen atom is protected.

Compounds of formula (Vl) may be prepared from the corresponding compound of formula (VII):

in which L¹ and X are as hereinbefore defined, or a protected derivative thereof, with an alkyl iodide R⁴I in the presence of a base such as cesium carbonate, in an aprotic solvent such as DMF.

Compounds of formula (VII) may be prepared by reacting the corresponding optionally protected aniline of formula (VIII):

with a pyrimidine compound of formula (IX):

in which L¹, L² and X are as hereinbefore defined, in aqueous methanol, at a temperature of about 70° C.

In a further embodiment, the compound of formula (II) is the following N-acetyl protected derivative:

wherein L¹, X₁ and R⁴ are as defined above. This may be prepared from the corresponding N-acetyl compounds of formula (Vl), (VII) and (VIII), as hereinbefore described.

Compounds of formula (III) are either obtainable from commercial sources, known in the art or may be prepared from the corresponding nitro compound using conventional reducing conditions for such a transformation, for example sodium dithionite, hydrogen gas in the presence of palladium, or iron metal with hydrochloric acid.

Compounds of formula (II) in which the leaving group represents other than halogen, may be obtained via the corresponding halogen by conventional methods.

Certain compounds of formula (II) and are new and form a further aspect of the invention.

Compounds of the present invention are useful as inhibitors of Syk and thus useful in treating diseases resulting from inappropriate mast cell activation, for instance allergic and inflammatory diseases.

Thus, in a further aspect, the present invention provides for a compound of formula (I), or a pharmaceutically acceptable salt or solvate thereof, for use in therapy.

In a further embodiment, the present invention provides for a method of treating of treating inappropriate mast cell activation which method comprises administering to a patient in need thereof an effective compound of formula I, or a or a pharmaceutically acceptable salt or solvate thereof.

In a further aspect, the present invention provides a method comprising administering to a patient in need thereof an effective compound of formula (I), or a pharmaceutically acceptable salt or solvate thereof, to inhibit a Syk kinase. In a further aspect, the present invention provides a method of treating an inflammatory disease which comprises administering to a patient in need thereof an effective compound of formula I, or a pharmaceutically acceptable salt or solvate thereof.

In a further aspect, the present invention provides a method of treating an allergic disorder which comprises administering to a patient in need thereof an effective compound of formula (I), or a pharmaceutically acceptable salt or solvate thereof.

Diseases and pathological conditions thought to be mediated by Syk kinase include inflammatory and allergic disorders involving mast cell activation, such as chronic obstructive pulmonary disease (COPD), adult respiratory distress syndrome (ARDS), asthma, ulcerative colitis, Crohn's Disease, bronchitis, conjunctivitis, psoriasis, sclerodoma, urticaria, dermatitis, and allergic rhinitis. They also include inflammatory conditions which involve B cells, for instance lupus and rheumatoid arthritis.

Likely disease targets for compounds of the present invention include chronic obstructive pulmonary disease (COPD), asthma and allergic rhinitis.

In a further aspect, the present invention provides a method of treating chronic obstructive pulmonary disease which comprises administering to a patient in need thereof an effective compound of formula (I), or a pharmaceutically acceptable salt or solvate thereof.

In a further aspect, the present invention provides a method of treating asthma which comprises administering to a patient in need thereof an effective compound of formula (I), or a pharmaceutically acceptable salt or solvate thereof.

In a further aspect, the present invention provides a method of treating allergic rhinitis which comprises administering to a patient in need thereof an effective compound of formula (I), or a pharmaceutically acceptable salt or solvate thereof.

Compounds of the present invention may also be used in combination with other classes of therapeutic agents which are known in the art. Representative clases of agents for use in such combinations include, for treating asthma, anti-inflammatory steroids (in particular corticosteroids), topical glucocorticoid agonists, PDE4 inhibitors, IKK2 inhibitors, A2a agonists, β₂-adrenoreceptor agonists (including both slow acting and long acting β₂-adrenoreceptor agonists), alpha 4 integrin inhibitors, and anti-muscarinics, and, for treating allergies, the foregoing agents, as well as H1 and H1/H3 antagonists. Representative agents for use in combination therapy for treating severe asthma include topically acting p38 inhibitors, and IKK2 inhibitors. Anti-inflammatory corticosteroids are well known in the art. Representative examples include fluticasone propionate (e.g. see US patent 4,335,121 ), beclomethasone 17- propionate ester, beclomethasone 17,21-dipropionate ester, dexamethasone or an ester thereof, mometasone or an ester thereof (e.g. mometasone furoate), ciclesonide, budesonide, and flunisolide. Further examples of anti-inflammatory corticosteroids are described in WO 02/12266 A1 (Glaxo Group Ltd), in particular, the compounds of Example 1 ( 6α,9α-difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy- 16α-methyl-3-oxo-androsta-1 ,4-diene-17β-carbothioic acid S-fluoromethyl ester) and Example 41 (6α,9α-difluoro-11 β-hydroxy-16α-methyl-17α-[(4-methyl-1 ,3-thiazole-5- carbonyOoxyJ-S-oxo-androsta-i ^-diene-^β-carbothioic acid S-fluoromethyl ester), or a pharmaceutically acceptable salt thereof.

Examples of β₂-adrenoreceptor agonists include salmeterol (e.g. as racemate or a single enantiomer such as the R-enantiomer), salbutamol, formoterol, salmefamol, fenoterol or terbutaline and salts thereof, for example the xinafoate salt of salmeterol, the sulphate salt or free base of salbutamol or the fumarate salt of formoterol. Long- acting β₂-adrenoreceptor agonists are preferred, especially those having a therapeutic effect over a 24 hour period such as salmeterol or formoterol.

Examples of anti-histamines include azelastine, levocabastine, olopatidine, methapyrilene, loratadine, cetirizine, desloratadine or fexofenadine.

Examples of anticholinergic compounds include muscarinic (M) receptor antagonists, in particular M-| , M2, M-1/M2, or M3 receptor antagonists, in particular a (selective) M3 receptor antagonist. Examples of anticholinergic compounds are described in

WO 03/011274 A2 and WO 02/069945 A2 / US 2002/0193393 A1 and US 2002/052312 A1. Examples of muscarinic M3 antagonists include ipratropium bromide, oxitropium bromide or tiotropium bromide.

Representative PDE4 or mixed PDE3/4 inhibitors that may be used in combination with compounds of the invention include AWD-12-281 (Elbion), PD-168787 (Pfizer), roflumilast, and cilomilast (GlaxoSmithKline). Further examples of PDE4 inhibitors are described in WO 2004/103998 (Glaxo Group Ltd).

The present invention also provides for so-called "triple combination" therapy, comprising a compound of formula (I) or a pharmaceutically acceptable salt thereof together with β₂-adrenoreceptor agonist and an anti-inflammatory corticosteroid.

Preferably this combination is for treatment and/or prophylaxis of asthma, COPD or allergic rhinitis. The β₂-adrenoreceptor agonist and/or the anti-inflammatory corticosteroid can be as described above and/or as described in WO 03/030939 A1. A representative example of such a "triple" combination comprises a compound of formula (I) or a pharmaceutically acceptable salt thereof, salmeterol or a pharmaceutically acceptable salt thereof (e.g. salmeterol xinafoate) and fluticasone propionate. Preferred compounds of formula (I) for use as inhibitors of Syk kinase are those which exhibit selectivity for the Syk kinase against other key kinases such as Aurora A, Aurora B, JNK3, for instance at least 1Ox (based on either pKi or plC₅₀ values for the enzymes)

The compound of the present invention will normally, but not necessarily, be formulated into pharmaceutical compositions prior to administration to a patient. Accordingly, in another aspect the invention is directed to pharmaceutical compositions comprising a compound of the invention and one or more pharmaceutically acceptable excipient.

The pharmaceutical compositions of the invention may be prepared and packaged in bulk form wherein a safe and effective amount of a compound of the invention can be extracted and then given to the patient, such as with powders or syrups. Alternatively, the pharmaceutical compositions of the invention may be prepared and packaged in unit dosage form wherein each physically discrete unit contains a safe and effective amount of a compound of the invention. When prepared in unit dosage form, the pharmaceutical compositions of the invention typically contain from about 0.1 to 99.9 wt.%, depending on the nature of the formulation.

The pharmaceutical compositions of the invention typically contain one compound of the invention. However, in certain embodiments, the pharmaceutical compositions of the invention contain more than one compound of the invention. For example, in certain embodiments the pharmaceutical compositions of the invention contain two compounds of the invention. In addition, the pharmaceutical compositions of the invention may optionally further comprise one or more additional pharmaceutically active compounds.

As used herein, "pharmaceutically acceptable excipient" means a pharmaceutically acceptable material, composition or vehicle involved in giving form or consistency to the pharmaceutical composition. Each excipient must be compatible with the other ingredients of the pharmaceutical composition when commingled, such that interactions which would substantially reduce the efficacy of the compound of the invention when administered to a patient and would result in pharmaceutically unacceptable compositions are avoided. In addition, each excipient must of course be of sufficiently high purity to render it pharmaceutically acceptable.

The compound of the invention and the pharmaceutically acceptable excipient or excipients will typically be formulated into a dosage form adapted for administration to the patient by the desired route of administration. For example, dosage forms include those adapted for (1 ) inhalation, such as aerosols and solutions; and (2) intranasal administration, such as solutions or sprays. It will be appreciated by the skilled person that a preferred route of administration for treating asthma and COPD is inhalation and that a preferred route of administration for treating allergic rhinitis is intranasal administration.

Suitable pharmaceutically acceptable excipients will vary depending upon the particular dosage form chosen. In addition, suitable pharmaceutically acceptable excipients may be chosen for a particular function that they may serve in the composition. For example, certain pharmaceutically acceptable excipients may be chosen for their ability to facilitate the production of uniform dosage forms. Certain pharmaceutically acceptable excipients may be chosen for their ability to facilitate the production of stable dosage forms. Certain pharmaceutically acceptable excipients may be chosen for their ability to facilitate the carrying or transporting the compound of the present invention once administered to the patient from one organ, or portion of the body, to another organ, or portion of the body. Certain pharmaceutically acceptable excipients may be chosen for their ability to enhance patient compliance.

Suitable pharmaceutically acceptable excipients include the following types of excipients: Diluents, fillers, lubricants, glidants, granulating agents, wetting agents, solvents, co-solvents, suspending agents, emulsifiers, anticaking agents, chelating agents, viscosity increasing agents, antioxidants, preservatives, stabilizers, surfactants, and buffering agents. The skilled artisan will appreciate that certain pharmaceutically acceptable excipients may serve more than one function and may serve alternative functions depending on how much of the excipient is present in the formulation and what other ingredients are present in the formulation.

Skilled artisans possess the knowledge and skill in the art to enable them to select suitable pharmaceutically acceptable excipients in appropriate amounts for use in the invention. In addition, there are a number of resources that are available to the skilled artisan which describe pharmaceutically acceptable excipients and may be useful in selecting suitable pharmaceutically acceptable excipients. Examples include Remington's Pharmaceutical Sciences (Mack Publishing Company), Remington: The Science and Practice of Pharmacy, (Lippincott Williams & Wilkins), The Handbook of Pharmaceutical Additives (Gower Publishing Limited), and The Handbook of Pharmaceutical Excipients (the American Pharmaceutical Association and the Pharmaceutical Press).

The pharmaceutical compositions of the invention are prepared using techniques and methods known to those skilled in the art. Some of the methods commonly used in the art are described in Remington's Pharmaceutical Sciences (Mack Publishing Company).

Dosage forms for nasal or inhaled administration may conveniently be formulated as aerosols, solutions, drops, gels or dry powders. Dosage forms for topical administration to the nasal cavity (nasal administration) include pressurised aerosol formulations and aqueous formulations administered to the nose by pressurised pump. Formulations which are non-pressurised and adapted for nasal administration are of particular interest. Suitable formulations contain water as the diluent or carrier for this purpose. Aqueous formulations for administration to the nose may be provided with conventional excipients such as buffering agents, tonicity modifying agents and the like. Aqueous formulations may also be administered to the nose by nebulisation.

In a further embodiment, dosage forms for nasal administration are provided in a metered dose device. The dosage form may be provided as a fluid formulation for delivery from a fluid dispenser having a dispensing nozzle or dispensing orifice through which a metered dose of the fluid formulation is dispensed upon the application of a user-applied force to a pump mechanism of the fluid dispenser. Such fluid dispensers are generally provided with a reservoir of multiple metered doses of the fluid formulation, the doses being dispensable upon sequential pump actuations. The dispensing nozzle or orifice may be configured for insertion into the nostrils of the user for spray dispensing of the fluid formulation into the nasal cavity. In one embodiment, the fluid dispenser is of the general type described and illustrated in WO-A-2005/044354. The dispenser has a housing which houses a fluid discharge device having a compression pump mounted on a container for containing a fluid formulation. The housing has at least one finger-operable side lever which is movable inwardly with respect to the housing to cam the container upwardly in the housing to cause the pump to compress and pump a metered dose of the formulation out of a pump stem through a nasal nozzle of the housing. A particularly preferred fluid dispenser is of the general type illustrated in Figures 30-40 of WO-A- 2005/044354.

For compositions suitable and/or adapted for inhaled administration, it is preferred that the compound or salt of formula (I) is in a particle-size-reduced form, and more preferably the size-reduced form is obtained or obtainable by micronisation. The preferable particle size of the size-reduced (e.g. micronised) compound or salt or solvate is defined by a D50 value of about 0.5 to about 10 microns (for example as measured using laser diffraction).

Aerosol formulations, e.g. for inhaled administration, can comprise a solution or fine suspension of the active substance in a pharmaceutically acceptable aqueous or non-aqueous solvent. Aerosol formulations can be presented in single or multidose quantities in sterile form in a sealed container, which can take the form of a cartridge or refill for use with an atomising device or inhaler. Alternatively the sealed container may be a unitary dispensing device such as a single dose nasal inhaler or an aerosol dispenser fitted with a metering valve (metered dose inhaler) which is intended for disposal once the contents of the container have been exhausted. Where the dosage form comprises an aerosol dispenser, it preferably contains a suitable propellant under pressure such as compressed air, carbon dioxide or an organic propellant such as a hydrofluorocarbon (HFC). Suitable HFC propellants include 1 ,1 ,1 , 2,3, 3,3-heptafluoropropane and 1 ,1 ,1 ,2-tetrafluoroethane. The aerosol dosage forms can also take the form of a pump-atomiser. The pressurised aerosol may contain a solution or a suspension of the active compound. This may require the incorporation of additional excipients e.g. co-solvents and/or surfactants to improve the dispersion characteristics and homogeneity of suspension formulations. Solution formulations may also require the addition of co-solvents such as ethanol. Other excipient modifiers may also be incorporated to improve, for example, the stability and/or taste and/or fine particle mass characteristics (amount and/or profile) of the formulation.

For pharmaceutical compositions suitable and/or adapted for inhaled administration, it is preferred that the pharmaceutical composition is a dry powder inhalable composition. Such a composition can comprise a powder base such as lactose, glucose, trehalose, mannitol or starch, the compound of formula (I) or salt or solvate thereof (preferably in particle-size-reduced form, e.g. in micronised form), and optionally a performance modifier such as L-leucine or another amino acid, and/or metals salts of stearic acid such as magnesium or calcium stearate. Preferably, the dry powder inhalable composition comprises a dry powder blend of lactose and the compound of formula (I) or salt thereof. The lactose is preferably lactose hydrate e.g. lactose monohydrate and/or is preferably inhalation-grade and/or fine-grade lactose. Preferably, the particle size of the lactose is defined by 90% or more (by weight or by volume) of the lactose particles being less than 1000 microns (micrometres) (e.g. 10-1000 microns e.g. 30-1000 microns) in diameter, and/or 50% or more of the lactose particles being less than 500 microns (e.g. 10-500 microns) in diameter. More preferably, the particle size of the lactose is defined by 90% or more of the lactose particles being less than 300 microns (e.g. 10-300 microns e.g. 50-300 microns) in diameter, and/or 50% or more of the lactose particles being less than 100 microns in diameter. Optionally, the particle size of the lactose is defined by 90% or more of the lactose particles being less than 100-200 microns in diameter, and/or 50% or more of the lactose particles being less than 40-70 microns in diameter. Most importantly, it is preferable that about 3 to about 30% (e.g. about 10%) (by weight or by volume) of the particles are less than 50 microns or less than 20 microns in diameter. For example, without limitation, a suitable inhalation-grade lactose is E9334 lactose (10% fines) (Borculo Domo Ingredients, Hanzeplein 25, 8017 JD Zwolle, Netherlands).

Optionally, in particular for dry powder inhalable compositions, a pharmaceutical composition for inhaled administration can be incorporated into a plurality of sealed dose containers (e.g. containing the dry powder composition) mounted longitudinally in a strip or ribbon inside a suitable inhalation device. The container is rupturable or peel-openable on demand and the dose of e.g. the dry powder composition can be administered by inhalation via the device such as the DISKUS ™ device, marketed by GlaxoSmithKline. The DISKUS ™ inhalation device is for example described in GB 2242134 A, and in such a device at least one container for the pharmaceutical composition in powder form (the container or containers preferably being a plurality of sealed dose containers mounted longitudinally in a strip or ribbon) is defined between two members peelably secured to one another; the device comprises: a means of defining an opening station for the said container or containers; a means for peeling the members apart at the opening station to open the container; and an outlet, communicating with the opened container, through which a user can inhale the pharmaceutical composition in powder form from the opened container.

A composition of the present invention, for intranasal administration, may also be adapted for dosing by insufflation, as a dry powder formulation.

It will be appreciated that when the compound of the present invention is administered in combination with other therapeutic agents normally administered by the inhaled, intravenous, oral or intranasal route, that the resultant pharmaceutical composition may be administered by the same routes.

The compound of the present invention may conveniently be administered in amounts of, for example, 1 μg to 100 mg. The precise dose will of course depend on the age and condition of the patient and the particular route of administration chosen.

Biological test methods

Compounds of the invention may be tested for in vitro activity in accordance with the following assays:

1. Enzyme Assay - Time-resolved fluorescence resonance energy transfer kinase assay

Recombinant human Syk was expressed as a His-tagged protein^*. The activity of Syk was assessed using a time-resolved fluorescence resonance energy transfer (TR-FRET) assay.

Version A - 3μl of substrate reagent containing biotinylated peptide, Biotin- AAAEEIYGEI (0.5μM final), ATP (30μM final) and MgCI₂ (1OmM final) in HEPES pH 7.4, (4OmM final), were added to wells containing 0.2μl of various concentrations of compound or DMSO vehicle (3.3% final) in Greiner low volume 384 well black plate. The reaction was initiated by the addition of 3μl of Syk (2OnM final) in HEPES pH 7.4 (4OmM final). The reaction was incubated for 40min at room temperature, then terminated by the addition of 3μl of read reagent containing 60 mM EDTA, 15OmM NaCI, 5OnM Streptavidin APC (Prozyme, San Leandro, California, USA), 0.5nM antiphosphotyrosine antibody labelled with W-1024 europium chelate (Wallac OY, Turku, Finland) in 4OmM HEPES pH 7.4, 0.03% BSA. The reaction was further incubated for 60min at room temperature. The degree of phosphorylation of Biotin- AAAEEIYGEI was measured using a BMG Rubystar plate reader (BMG LabTechnologies Ltd, Aylesbury, UK) as a ratio of specific 665 nm energy transfer signal to reference europium 620 nm signal.

Version B - Syk was pre-activated at room temperature for 30 mins in the presence of 16.6mM MgCI₂, 8.3mM ATP and then diluted to 4nM in 4OmM Hepes pH 7.4, 0.01% BSA. 3μl of substrate reagent containing biotinylated peptide, Biotin- AAAEEIYGEI (0.5μM final), ATP (30μM final) and MgCI₂ (1OmM final) in 4OmM HEPES pH 7.4, 0.01% BSA, were added to wells containing 0.1 μl of various concentrations of compound or DMSO vehicle (1.7% final) in Greiner low volume 384 well black plate. The reaction was initiated by the addition of 3μl of diluted Syk (2nM final). The reaction was incubated for 60min at room temperature, then terminated by the addition of 3μl of read reagent containing 60 mM EDTA, 15OmM NaCI, 5OnM Streptavidin APC (Prozyme, San Leandro, California, USA), 0.5nM antiphosphotyrosine antibody labelled with W-1024 europium chelate (Wallac OY, Turku, Finland) in 4OmM HEPES pH 7.4, 0.03% BSA. The reaction was further incubated for 45min at room temperature. The degree of phosphorylation of Biotin- AAAEEIYGEI was measured using a BMG Rubystar plate reader (BMG LabTechnologies Ltd, Aylesbury, UK) as a ratio of specific 665 nm energy transfer signal to reference europium 620 nm signal. Compounds according to the present invention were assayed in this, or a similar Time-resolved fluorescence resonance energy transfer kinase assay, and gave IC₅₀ values less than 10μM.

* Preparation of Recombinant Human Full Length Spleen Tyrosine Kinase (Svk) Full length human Syk was expressed with a 6His tag on the N-terminal using the baculovirus system (Invitrogen, Paisley, Scotland). The cells were disrupted by dounce homogenisation, the debris removed by centrifugation and the lysate contacted with NiNTA Superflow (Qiagen, Crawley, UK). The NiNTA was packed into a column and eluted using 10 column volumes each of buffer (2OmM Tris pHδ.O, 30OmM NaCI, 1OmM βMcEtOH, 10% glycerol), buffer + 1 M NaCI, buffer + 2OmM Imidazole and buffer + 30OmM imidazole. The 30OmM Imidazole fractions were pooled buffer exchanged using G25M (Amersham Biosciences, Buckinghamshire, UK) into 2OmM MES pH 6.0, 2OmM NaCI, 1OmM βMcEtOH,10% glycerol. The buffer exchanged 6His-Syk was loaded onto a Source15S column (Amersham Biosciences, Buckinghamshire, UK) and the column eluted using a NaCI gradient 0-50OmM over 50 column volumes. The 6His-Syk containing fractions were pooled and concentrated by ultra-filtration. The identity of 6His-Syk was confirmed by peptide mass finger printing and intact LC-MS.

2. Whole Cell Assay - cFms assay

Principle of the assay

Cells of the mouse fibroblast cell line NIH-3T3 are stably transfected with a cFms- SYK chimera. Addition of the ligand (MCSF) produces dimerisation of the chimera resulting in autophosphorylation of the SYK kinase domain. Following cell lysis phosphorylated SYK is detected by ELISA.

Stimulation of cFms-SYK cells with MCSF Version A

Cells are plated at a density of 1x10⁵/well in a volume of 200μl growth medium (DMEM containing 10% heat inactivated foetal calf serum, 1% L-glutamine, 400μg/ml geneticin and 400μg/ml zeocin) in 96 well Collagen 1 coated tissue culture plates. Following incubation at 37°C, 10% CO_2, for 2Oh, the cell supernatant is removed and replaced with 200μl DMEM containing 1 % penicillin/streptomycin (serum free DMEM). The cells are incubated for an hour under the conditions described above. The medium is removed, 50μl appropriately diluted compound solution added and the plate incubated for a further hour. Cells are stimulated with 25μl MCSF (0.66μg/ml final) for 20min at 37°C. After removal of the supernatant, the cells are washed with cold PBS and lysed with 100μl lysis buffer for 4h at 4°C.

Stimulation of cFms-SYK cells with MCSF Version B Cells are plated at a density of 1x105/well in a volume of 200μl growth medium (DMEM containing 10% heat inactivated foetal calf serum, 1% L-glutamine, 400μg/ml geneticin and 400μg/ml zeocin) in 96 well Collagen 1 coated tissue culture plates. Following incubation at 37°C, 10% CO2, for 2Oh the cell supernatant is removed and 50μl appropriately diluted compound solution added and the plate incubated for an hour. Cells are stimulated with 25μl MCSF (0.66μg/ml final) for 20min at 37°C. After removal of the supernatant, the cells are washed with cold PBS and lysed with 10Oμl lysis buffer for 4h at 4⁰C.

cFms ELISA

85μL cell lysate is transferred to a 96 well ELISA plate coated with goat anti human M-CSF R capture antibody and incubated for 16 hours at 4°C. The plate is washed and a biotinylated anti-phosphotyrosine detection antibody added (100μl/well) for 2 hours at room temperature. This is removed and replaced with 100μl Streptavidin- HRP for 30min. Captured phosphorylated SYK is visualised using 10Oμl TMB substrate. The reaction is terminated with 50μl 1 M sulphuric acid and the absorbance measured at 450nm.

Compound Preparation Compound is prepared as a 1 OmM stock in DMSO and a dilution series prepared in DMSO using 9 successive 5-fold dilutions. This dilution series is diluted a further 1 :333 with serum free DMEM to give the concentration range to be tested of 1x10⁵ to 1.54x10^"11M. Compound dilutions are prepared using the Biomek 2000 or Biomek Nx automated robotic pipetting systems.

3. B Cell Proliferation Assay

Background The population of B cells observed in this assay are the naive mature IgM/lgD expressing population. These form at least 70% of the purified B cell population (the rest being isotype switched memory B cells) and are the only cells that proliferate as the cells are stimulated with anti-lgM.

Anti-lgM drives signalling through the B cell receptor which is Syk dependant. Proliferation is a functional measure of B cell signalling that can be measured by observing the incorporation of tritiated methyl thymidine into the cells.

Protocol Purified human tonsillar B cells are resuspended in Buckleys* medium at a concentration of 1.25 x 10⁶ ml.

160μl of cells re-suspended in Buckley's medium is added to the compound and control wells of a 96 well plate. The control wells are located on column 11 and 12 of the 96 well plate. The background wells are located in column 12 and 20μl of 10μM control is added to provide an appropriate background control. 20μl of 1% DMSO is added to the wells in column 11 for the stimulated control.

The compound titrations are located between columns 1 and 10. Three compounds are run in duplicate on each plate and row A and B are used for the control compound titration.

The final concentration of DMSO is 0.1 % in the assay. The cells are left for 45min, after 4 min the proliferative stimulus is added to the first 1 1 wells of the 96 well plate and 20μl of medium is added to column 12. F(ab')2 fragments of a polyclonal goat anti-sera raised to human IgM is used at a final concentration of 15μg/ ml to stimulate the cells. (Biosource. Cat no: AMI 4601 ).

Tritiated methyl thymidine is added to the cells at a concentration of 1μCi per well. (Amersham, TRK 758). The radioactivity is added 65 hours after the initial stimulus and is left on the cells for 6 to 8 hours. After pulsing with methyl thymidine the cells are harvested on a Skatron 96 well cell harvester onto glass fibre mats. Once these have dried these are counted on a Wallac 1450 Microbeta scintillation counter.

Data is downloaded as an XL file and IC50's determined using Activity base.

* Buckleys Medium: 450 ml Iscoves (Sigma I 3390), 50ml FCS, 2.5 g BSA, 5ml Pen/ strep, 5ml Glutamine (20OmM), 500μl Apo transferrin (50mg/ml) Sigma (T 1147), 100μl Bovine Insulin (10mg/ml) Sigma (I 1882).

Compound Preparation

Compound is prepared as a 1OmM stock in DMSO and a dilution series prepared in DMSO using 9 successive 3-fold dilutions. This dilution series is diluted a further 1 :100 with Buckleys medium to give the concentration range to be tested of 100μM to 5nM. This is added as 20μl to 96 well plates in duplicate to generate two IC50's for each compound tested. Each plate is run in the presence of a control compound, which acts as an internal standard. .

4. LAD2 Assay

Principle of the assay

LAD2 is a stem cell factor (SCF)-dependent human mast cell line that was established by the NIH from bone marrow aspirates from a patient with mast cell sarcoma/leukaemia. LAD2 cells resemble CD34+-derived human mast cells and express functional FcεRI. The FcεRI is up-regulated in the presence of IL-4, SCF and IgE, subsequent cross linking of cell-bound IgE results in degranulation which can be measured as hexosaminidase release.

Priming LAD2 cells to up-regulate FcεRI LAD2 cells are re-suspended at 1x10⁵/ml in complete stem pro-34SFM (Gibco Cat 10640-019 media containing Stem Pro-34 nutrient supplement (1 :40), glutamine (2mM), penicillin (100μg/ml), streptomycin (100μg/ml)) with additional supplements of human recombinant SCF (100ng/ml; R&D systems), human recombinant Interleukin- 4 (6ng/ml; R&D Systems) and IgE (100μg/ml; Calbiochem). Cells are then maintained for 5 days at 37°C, 5% CO2 in a humidified atmosphere.

Compound Preparation

Compounds are titrated from a 2mM stock in 100% DMSO to give 9 successive 1 :3 dilutions (V 96-well Nunc; Biomek 2000). From this master plate 3μl is dispensed into a daughter plate (flat 96-well NuncBiomek Fx) which is then diluted 1 :40 in RPMI with 2mM glutamine, and 20μl of the diluted compound transferred into the Greiner cell plate. Therefore the final compound concentration range is 1x10^'5M to 5x10^"10M in a constant 0.5% DMSO. Control wells are treated with 0.5% DMSO.

Activation of LAD2 cells with anti-lgE: Version A

Primed LAD2 cells are centrifuged (30Og, 5min), the supernatant discarded and the cell pellet re-suspended at 1x10⁴ cells/ml in RPMI supplemented with glutamine (2mM). Following a further centrifugation (30Og, 5min) the cells are re-suspended in fresh RPMI with glutamine (2mM), adjusted to a density of 2.85x10⁵/ml, and pipetted into sterile V-well plates (70μl/well; Greiner) containing 20μl diluted compound (prepared as detailed above). Cells are then incubated for 1h (37°C, 5% CO₂ in a humidified atmosphere) before activating with a sub-maximal concentration of anti- lgE (10μl volume to give a final assay dilution of 1 :2700; Sigma). Following a 40min incubation (37°C, 5% CO₂ in a humidified atmosphere), plates are centrifuged (120Og, 10min, 4°C) and the supernatant removed for hexosaminidase assay. The cell pellet is lysed in 100μl/well triton-X (0.5% in RPMI 2mM glutamine) at 37°C for 30 min.

Activation of LAD2 cells with anti-lgE: Version B Primed LAD2 cells are centrifuged (400g, 5min), the supernatant discarded and the cell pellet re-suspended at 1x10⁴ cells/ml in RPMI supplemented with glutamine (2mM). Following a further centrifugation (40Og, 5min) the cells are re-suspended in fresh RPMI with glutamine (2mM), adjusted to a density of 5.7 x10⁵/ml, and pipetted into sterile V-well plates (70μl/well; Greiner) containing 20μl diluted compound (prepared as detailed above). Cells are then incubated for 1 h (37°C, 5% CO₂ in a humidified atmosphere) before activating with a sub-maximal concentration of anti- lgE (10μl volume to give a final assay dilution of 1 :2700; Sigma). Following a 40min incubation (37°C, 5% CO₂ in a humidified atmosphere), plates are centrifuged (120Og, 10min, 4^CC) and the supernatant removed for hexosaminidase assay. The cell pellet is lysed in 100μl/well triton-X (0.5% in RPMI 2mM glutamine) at 37°C for 30min.

Beta-hexosaminidase assay Beta-hexosaminidase activity is measured by the conversion of 4-methylumbelliferyl N-acetyl-ε-D glucosaminide (Sigma) to a fluorescent product.

Supernatant or lysate (25μl) is incubated with an equal volume of 4- methylumbelliferyl N-acetyl-ε-D glucosaminide (500μM in 0.2M sodium citrate buffer, pH 4.5) in black 96-well plate (Nunc) for 1 hour at 37⁰C. The reaction is then terminated by addition of Trizma pH9 (90μl) and the fluorescent product measured using excitation 356nm and emission 450nm (Tecan Safire)

A useful screening strategy comprises assay 1 (enzyme assay (pKi), assay 2 and then assay 3 (B Cell Proliferation) or assay 4 (LAD2).

Intermediates and Examples

General

All temperatures are in ⁰C.

DCM refers to dichloromethane

DMSO refers to dimethylsulfoxide.

DMF refers to Λ/,Λ/-dimethylformamide

IPA refers to propan-2-ol TFA refers to trifluoroacetic acid

THF refers to tetrahydrofuran.

HPLC refers to high performance liquid chromatography.

ADDP refers to 1 ,1'(azodicarbonyl)dipiperidine

SPE refers to solid phase extraction cartridges marketed by lsolute TBTU refers to O-benzotriazol-1-yl-Λ/,Λ/,Λ/',Λ/',-bis(tetramethylene)uronium tetrafluoroborate

HOBt refers to N-hydroxybenzotriazole hydrate

¹H NMR spectra were recorded using a Bruker DPX 400MHz, referenced to tetramethylsilane.

LC/MS was conducted on a Supelcosil LCABZ+PLUS column (3.3 cm x 4.6 mm ID) eluting with 0.1% HCO2H and 0.01 M ammonium acetate in water (solvent A) and 0.05% HCO2H 5% water in acetonitrile (solvent B), using the following elution gradient 0.0-7min 0%B, 0.7-4.2min 100%B, 4.2-5.3min 0%B, 5.3-5.5min 0%B at a flow rate of 3ml/min. The mass spectra were recorded on a Fisons VG Platform spectrometer using electrospray positive and negative mode (ES+ve and ES-ve).

"Mass directed autoprep" / "preparative mass directed HPLC" was conducted on a system such as; a Waters FractionLynx system comprising of a Waters 600 pump with extended pump heads, Waters 2700 autosampler, Waters 996 diode array and Gilson 202 fraction collector on a 10 cm 2.54 cm ID ABZ+ column, eluting with either 0.1% formic acid or trifluoroacetic acid in water (solvent A) and 0.1 % formic or trifluoroacetic acid in acetonitrile (solvent B) using the appropriate elution gradient. Mass spectra were recored on Micromass ZMD mass spectrometer using electrospray positive and negative mode, alternate scans. The software used was MassLynx 3.5 with OpenLynx and FractionLynx optio or using equivalent alternative systems.

"Hydrophobic frits" refers to filtration tubes sold by Whatman. SPE (solid phase extraction) refers to the use of cartridges sold by International Sorbent Technology Ltd.

The Flashmaster Il is an automated multi-user flash chromatography system, available from Argonaut Technologies Ltd, which utilises disposable, normal phase, SPE cartridges (2 g to 100 g). It provides quaternary on-line solvent mixing to enable gradient methods to be run. Samples are queued using the multi-functional open access software, which manages solvents, flow-rates, gradient profile and collection conditions. The system is equipped with a Knauer variable wavelength uv-detector and two Gilson FC204 fraction-collectors enabling automated peak cutting, collection and tracking.

Silica chromatography techniques include either automated (Flashmaster) techniques or manual chromatography on pre-packed cartridges (SPE) or manually- packed flash columns.

Microwave chemistry was typically performed in sealed vessels, irradiating with a suitable microwave reactor system, such as a Biotage Initiator™ Microwave Synthesiser.

When the name of a commercial supplier is given after the name of a compound or a reagent, for instance "compound X (Aldrich)" or "compound X / Aldrich", this means that compound X is obtainable from a commercial supplier, such as the commercial supplier named.

Similarly, when a literature or a patent reference is given after the name of a compound, for instance compound Y (EP 0 123 456), this means that the preparation of the compound is described in the named reference.

The names of the above mentioned Examples have been obtained using the compound naming programme "ACD Name Pro 6.02".

Intermediate 1 - Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1-(([2- (trimethylsilyl)ethyl1oxy)methyl)-1 /-/-indazol-4-amine

To a flask charged with Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-1-({[2- (trimethylsilyl)ethyl]oxy}methyl)-1H-indazol-4-amine (10.84g) in DMF (400ml) was added potassium carbonate (4.98g) portionwise. The mixture was stirred for 30min prior to the addition of ethyl iodide (2.63ml). The reaction mixture was stirred overnight. The solvent was removed in vacuo and the residue partitioned with water (3x 300ml) and DCM (3x 300ml). The dried organic extracts (MgSO₄) were filtered and concentrated. The crude residue was purified by chromatography on silica cartridges (2x 7Og) eluting with 20% ethylacetate in cyclohexane to afford, after evaporation of the solvents, Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1-({[2- (trimethylsilyl)ethyl]oxy}methyl)-1H-indazol-4-amine as a yellow gum which solidified on cooling (6.25g). LC/MS Rt 3.88min, MH⁺ 422, 424.

Intermediate 2 _: Λ/-(5-bromo-2-chloro-4-pyrimidinyl)-Λ/-ethyl-1-({r2-

(trimethylsilyl)ethvπoxy}methvD-1/-/-indazol-4-amine

To a flask charged with Λ/-(2-chloro-5-bromo-4-pyrimidinyl)-1-({[2-

(trimethylsilyl)ethyl]oxy}methyl)-1H-indazol-4-amine (2.Og) in DMF (100ml) was added potassium carbonate (0.79g) portionwise. The mixture was stirred for 30min prior to the addition of ethyl iodide (0.42ml). The reaction mixture was stirred at 23⁰C for 16h after which time a further aliquot of ethyl iodide (0.42ml) was added and the reaction stirred for 6h. At this time an additional aliquot of ethyl iodide (0.42ml) was added and the reaction stirred for 16h. The reaction was concentrated in vacuo and the residue partitioned with water (50ml) and DCM (3x 50ml). The dried organic extracts (hydrophobic frit) were concentrated and the crude residue purified by chromatography (2Og cartridge) eluting with an ethyl acetate / cyclohexane gradient. Evaporation of the solvents gave Λ/-(5-bromo-2-chloro-4-pyrimidinyl)-Λ/-ethyl-1-({[2- (trimethylsilyl)ethyl]oxy}methyl)-1H-indazol-4-amine as a pale yellow oil. (1.0g). LC/MS; Rt 4.17min, MH⁺ 482, 484, 486.

Intermediate 3 - Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-1-((f2- (trimethylsilyl)ethylloxy)methyl)-1/-/-indazol-4-amine

To a stirred solution of a mixture of 2-({[2-(trimethylsilyl)ethyl]oxy}methyl)-2H-indazol- 4-amine and 1-({[2-(trimethylsilyl)ethyl]oxy}methyl)-1 H-indazol-4-amine (20.4g) in N,N-diisopropylethylamine (155ml) was added 2,4-dichloro-5-fluoropyrimidine (13.6g, Aldrich). The mixture was heated under nitrogen at reflux for 3h. Further 2,4- dichloro-5-fluoropyrimidine (0.97g) was added over 1 h before the mixture was allowed to cool. The volatile components were evaporated in vacuo and the residue was re-dissolved in DCM and divided into 2 portions of approximately 1 :2 ratio. The first portion was absorbed onto a 330g silica cartridge before being purified using a CombiFlash^® Companion™ system eluting with a gradient of ethyl acetate in cyclohexane (0-100%). The required pure fractions were combined and the solvent was evaporated in vacuo to give the Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-1-({[2- (trimethylsilyl)ethyl]oxy}methyl)-1H-indazol-4-amine as an orange/brown solid (4.99g). LC/MS; Rt 3.57min, MH⁺ 394,396.

The second portion was absorbed onto a 75Og silica cartridge before being purified using a CombiFlash^® Companion™ system eluting with a gradient of ethyl acetate in cyclohexane (0-100%). The required pure fractions were combined and the solvent was evaporated in vacuo to give a further quantity of the Λ/-(2-chloro-5-fluoro-4- pyrimidinyl)-1 -({[2-(trimethylsilyl)ethyl]oxy}methyl)-1 H-indazol-4-amine as an orange/brown solid (10.96g). LC/MS; Rt 3.57min, MH⁺ 394,396.

Intermediate 4 - 1-({r2-(trimethylsilyl)ethvπoxy)methyl)-1 H-indazol-4-amine mixture with 2-({r2-(trimethylsilyl)ethylloxy)methyl)-2H-indazol-4-amine

Two vigorously stirred solutions, each containing 4-nitro-1-({[2- (trimethylsilyl)ethyl]oxy}methyl)-1 H-indazole mixture with 4-nitro-2-({[2- (trimethylsilyl)ethyl]oxy}methyl)-2H-indazole (21.17g) in ethanol (150ml), were hydrogenated at room temperature and 1atm. of pressure using 10% palladium on carbon catalyst (2.65g) for 1.5h. The mixtures were both filtered through Celite and the filter cakes were washed with portions of ethanol. The combined ethanol filtrates and washings from both reactions were evaporated in vacuo to give 1-({[2- (trimethylsilyl)ethyl]oxy}methyl)-1 H-indazol-4-amine mixture with 2-({[2- (trimethylsilyl)ethyl]oxy}methyl)-2H-indazol-4-amine as a brown oil (36.83g). LC/MS; Rt 3.14 and 3.35min, MH⁺ 264.

Intermediate 5 - 4-nitro-1-({[2-(trimethylsilyl)ethylloxy)methyl)-1 H-indazole mixture with 4-nitro-2-((f2-(trimethylsilyl)ethylloxy)methyl)-2H-indazole

To a stirred solution of 4-nitroindazole (24.Og, Bionet Research Intermediates Catalogue) in dry tetrahydrofuran (380ml), stirred under nitrogen and cooled using an ice bath was added portion wise sodium-tert-butoxide (14.85g) followed after 5min by 2-(trimethylsilyl)ethoxymethylchloride (28.2ml). After removal of the cooling bath, the mixture was stirred at room temperature for 3h. Methanol (240ml) was added to the mixture and the solvent was evaporated in vacuo. The residue was partitioned between chloroform (300ml) and water (200ml), the phases were separated and the aqueous phase was extracted with chloroform (3x100ml). The combined organic phases were dried (MgSO₄), filtered and the solvent evaporated in vacuo to give 4- nitro-1-({[2-(trimethylsilyl)ethyl]oxy}methyl)-1H-indazole mixture with 4-nitro-2-({[2- (trimethylsilyl)ethyl]oxy}methyl)-2H-indazole as a orange/brown oil (42.34g). LC/MS; Rt 3.68 and 3.77min, MH⁺ 294.

Intermediate 6 - Λ/-(5-bromo-2-chloro-4-pyrimidinvQ-1-((r2- (trimethylsilyl)ethylloxy)methyl)-1/-/-indazol-4-amine

To a flask charged with a mixture of 1-({[2-(trimethylsilyl)ethyl]oxy}methyl)-1H- indazol-4-amine and 2-({[2-(trimethylsilyl)ethyl]oxy}methyl)-2H-indazol-4-amine (5.3g) in methanol (30ml) was added a solution of 5-bromo-2,4-dichloropyrimidine (4.6g, Aldrich), in methanol (30ml). Water (180ml) was added to the stirred solution and the mixture heated at 7O⁰C for 5h. The mixture was allowed to cool to room temperature and then diluted with water (200ml). The mixture was decanted to leave an organic residue. This residue was purified by chromatography on an silica cartridge (100g) eluting with an ethyl acetate / cyclohexane gradient to afford Λ/-(5-bromo-2-chloro-4- pyrimidinyl)-1-({[2-(trimethylsilyl)ethyl]oxy}methyl)-1/-/-indazol-4-amine as a white solid (4.22g). LC/MS; Rt 4.75min, MH⁺ 453 / 455 / 457.

Method 1

A microwave vessel was charged with Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1- ({[2-(trimethylsilyl)ethyl]oxy}methyl)-1H-indazol-4-amine (0.05g) and aniline (0.2mmol), followed by and a solution of acetone / water / cone, hydrochloric acid (150:100:1 , 3ml) and N-methyl pyrrolidinone (0.1ml) The mixture was heated by microwave irradiation in a sealed vessel at 15O⁰C for 60min. The crude material was added to an SCX cartridge (10g), primed with methanol. The cartridge was eluted with 3x column volumes of methanol and then flushed with 2x column volumes ammonia in methanol solution (2M). The basic fractions were combined and concentrated and the residue purified by preparative mass directed HPLC to afford the title compound.

ff-λ H- 1 ,2,3-benzotriazol-5- 1 /-/-1 ,2,3-benzotriazol-5-amine 390 2.9 yl-Λ/*-ethyl-5-fluoro-Λ/⁴-1 H- Lancaster Synthesis indazol-4-yl-2,4- pyrimidinediamine trifluoroacetate

ΛΛethyl-5-fluoro-/V⁴-1H- 1 /-/-indazol-6-amine / Aldrich 398 3.0 indazoM-yl-Λ/²-! H-indazol-6- yl-2,4-pyrimidinediamine trifluoroacetate

Λr-ethyl-5-fluoro-ΛT-1H- 3-(1 ,3-oxazol-5-yl)aniline / Ryan Scientific 416 3.24 indazoM-yl-Λf-β-αS- Product List oxazol-5-yl)phenyl]-2,4- pyrimidinediamine

Example 9 - 3-((4-rethyl(1H-indazol-4-yl)aminol-5-fluoro-2- pyrimidinyl)amino)benzenesulfonannide

A mixture of Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1H-indazol-4-amine (30.9mg) and 3-aminobenzenesulphonamide (36.5mg, Apollo Scientific) in acetone (1.5ml) and water (1 ml) had cone, hydrochloric acid (40μl) added before heating in a sealed vessel at 130⁰C in a microwave reactor for 15min. The mixture was further heated at 150⁰C in a microwave reactor for 15min and then for a further 15min at 160°C. The solvent was evaporated in vacuo, and the residue was purified by mass-directed auto prep. The required fractions were combined and the solvent evaporated in vacuo to give the title compound as a cream solid (29.3mg). LC/MS; Rt 3.01 min, MH⁺ 428.

Intermediate 7 - N-(2-chloro-5-fluoro-4-pyrimidinyl)-N-ethyl-1 H-indazol-4-amine

A solution of Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1-({[2-

(trimethylsilyl)ethyl]oxy}methyl)-1H-indazol-4-amine (4.2g) in IPA (20ml) and hydrochloric acid (5M, 60ml) was stirred with heating at 55°C for 4 hours. After allowing to cool, the solvents were evaporated in vacuo and the residue was basified with saturated aqueous sodium carbonate solution and extracted with ethyl acetate (3x 100ml). The combined organic phases were washed with water (3x 100ml), dried (MgSO₄), filtered, and the solvent evaporated in vacuo. The residue was absorbed onto a silica SPE cartridge (100g) which was then eluted with an ethyl acetate / cyclohexane gradient (0-100%) over 40min followed by a gradient of methanol in ethyl acetate (0-20%). The required fractions were combined and the solvent was evaporated in vacuo to give the N-(2-chloro-5-fluoro-4-pyrimidinyl)-N-ethyl-1 H- indazol-4-amine as a yellow oil (1.55g). LC/MS; Rt 3.05min, MH⁺ 292, 294.

Intermediate 8 - /V-(2-chloro-5-fluoro-4-pyrimidinvD-1H-indazol-4-amine

A solution of 2,4-dichloro-5-fluoropyhmidine (0.2g), 4-aminoindazole (0.16g, Ryan Scientific Product List) and Sodium hydrogen carbonate (0.3g) in THF (1.2ml) and ethanol (4.8ml) was stirred at room temperature overnight. The solvents were evaporated in vacuo and the residue partitioned between DCM and water, leaving an insoluble precipitate which was isolated by filtration and dried in vacuo to give Λ/-(2- chloro-5-fluoro-4-pyrimidinyl)-1H-indazol-4-amine (0.093g). LC/MS; Rt 2.60, MH⁺ 264.

Example 10 - 6-((4-rEthyl(1H-indazol-4-yl)amino1-5-fluoro-2-pyrimidinyl)amino)-1- methyl-1.3-dihvdro-2H-benzimidazol-2-one

The mixture of 6-({4-[ethyl(1H-indazol-4-yl)amino]-5-fluoro-2-pyrimidinyl}amino)-1- methyl-1 ,3-dihydro-2H-benzimidazol-2-one and 5-({4-[ethyl(1 H-indazol-4-yl)amino]-5- fluoro-2-pyrimidinyl}amino)-1-methyl-1 ,3-dihydro-2H-benzimidazol-2-one (3:7) Formic acid (3:7, 12mg) was separated on a Phenomenz Luna C18 column (100mm x 21.2mm ID 5uM) using 0.1% TFA / H₂O and (0.05% TFA / methanol) / acetonitrile (1 :1 ) gradient of 25 to 35% over 25min.

The more polar material was evaporated in vacuo, and recolumned using the above conditions. The product fractions were evaporated, the residue dissolved in methanol, applied to an SCX-2 SPE cartridge (5g) and eluted with 2N methanolic ammonia. The filtrate was evaporated in vacuo to give the title compound (2.7mg). LC/MS; Rt 2.66min, MH⁺ 419.

Example 11 - 5-({4-rEthyl(1H-indazol-4-yl)aminol-5-fluoro-2-pyrimidinyl}amino)-1- methyl-1.3-dihvdro-2H-benzimidazol-2-one

The mixture of 6-({4-[ethyl(1H-indazol-4-yl)amino]-5-fluoro-2-pyrimidinyl}amino)-1- methyl-1 ,3-dihydro-2/-/-benzimidazol-2-one and 5-({4-[ethyl(1H-indazol-4-yl)amino]-5- fluoro-2-pyrimidinyl}amino)-1-methyl-1 ,3-dihydro-2H-benzimidazol-2-one (3:7) Formic acid (3:7, 12mg) was separated on a Phenomenz Luna C18 column (100mm x 21.2mm ID 5uM) using 0.1 % TFA / H₂O and (0.05% TFA / methanol) / acetonitrile (1 :1) gradient of 25 to 35% over 25min.

The less polar material was evaporated in vacuo, the residue dissolved in methanol, applied to an SCX-2 SPE cartridge (5g) and eluted with 2N methanolic ammonia. The filtrate was evaporated in vacuo to give the title compound (8mg). LC/MS; Rt 2.67min, MH⁺ 419.

Intermediate 9 - 6-((4-rEthyl(1H-indazol-4-yl)amino1-5-fluoro-2-pyrimidinyl)amino)-1- methyl-1 ,3-dihvdro-2/-/-benzimidazol-2-one and 5-({4-Fethyl(1 H-indazol-4-yl)aminol-5- fluoro-2-pyrimidinyl)amino)-1-methyl-1 ,3-dihydro-2/-/-benzimidazol-2-one (3:7) Formic acid(1 :1 )

A mixture of 6-amino-1-methyl-1 ,3-dihydro-2H-benzimidazol-2-one and 5-amino-1- methyl-1 ,3-dihydro-2H-benzimidazol-2-one (3:7, 33mg) and Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1/-/-indazol-4-amine (50mg) were dissolved in a mixture of acetone / water / cone, hydrochloric acid (1.2ml, 150:100:1). The solution was heated at 160⁰C by microwave irradiation in a sealed vessel for 30min. The reaction was then evaporated to dryness and the crude residue was purified twice by mass directed autoprep to give the title compound (20mg) (3:7 mixture). LC/MS; Rt 2.5min, MH⁺ 419.

Intermediate 10 - 6-Amino-1-methyl-1.3-dihvdro-2H-benzimidazol-2-one and 5- amino-1 -methyl-1 ,3-dihydro-2/-/-benzimidazol-2-one (3:7)

A mixture of 1-methyl-6-nitro-1,3-dihydro-2H-benzimidazol-2-one and 1-methyl-5- nitro-1,3-dihydro-2H-benzimidazol-2-one (3:7, 110mg) in dry ethanol (30ml) was added to 10% Palladium on carbon catalyst (20mg) and stirred under a hydrogen atmosphere (1Atm) for 3h. The mixture was filtered through celite and evaporated in vacuo to give the title compounds (3:7 mixture, 85mg). LC/MS; Rt 0.34min. NMR; [CD₃CN] δH Major component:- 6.65-6.63,(1H,d), 6.33-6.23.,(2H₁ m), 3.83,(2H, s, NH₂), 3.14,(3H, s). Minor component:^.67-6.65,(1 H, d), 6.33-6.23, (2H, m), 3.86,(2H₁ S, NH₂), 3.13,(3H, s).

Intermediate 11 - 1-Methyl-6-nitro-1,3-dihvdro-2/-/-benzimidazol-2-one - 1-methyl-5- nitro-1.3-dihydro-2/-/-benzimidazol-2-one (3:7)

To a solution of 5-nitro-1,3-dihydro-2H-benzimidazol-2-one (LOg, Aldrich) in dry acetone (20ml) was added portionwise, powdered potassium hydroxide (0.32g). A solution of methyl iodide (1.58g) in dry acetone (5ml) was added dropwise over 10- 15min. The mixture was stirred at room temperature under nitrogen for 18h. Acetone (20ml) was added and the mixture filtered. The filtrate was evaporated in vacuo and the residue stirred with water and filtered. The solid was purified further by chromatography, pre-absorbing onto silica gel (10g), adding to a silica cartridge (100g) and eluting with an ethyl acetate / cyclohexane gradient (0 to 100%) over 60min. to afford the title compounds (3:7 mixture, 110mg). LC/MS; Rt 2.28min. NMR; [CD₃CN] δH Major component :- 8.09-8.06,(1H, dd), 7.89- 7.88.,(1H, d), 7.15-7.13,(1H, d), 3.38,(3H, s). Minor component :- 8.04-8.01,(1H, dd), 7.93-7.92, (1 H, d), 7.17-7.15,(1 H, d), 3.39,(3H, s).

Example 12 - formic acid - 6-((4-rethyl(1/-/-indazol-4-yl)aminol-5-fluoro-2- pyrimidinyl)amino)-2,3-dihvdro-1/-/-isoindol-1-one (1:1)

Λ/-(2-Chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1H-indazol-4-amine (40mg) and 6-amino- 2,3-dihydro-1H-isoindol-1-one (20mg) were dissolved in a mixture of acetone / water / cone, hydrochloric acid (1.2ml, 150:100:1). The solution heated at 160⁰C by microwave irradiation in a sealed vessel for 30min. The reaction mixture was then evaporated to dryness and the crude residue was purified by mass directed autoprep to give 6-({4-[ethyl(1 H-indazol-4-yl)amino]-5-fluoro-2-pyrimidinyl}amino)-2,3-dihydro- 1H-isoindol-1-one (32mg). LC/MS; Rt 2.77min, MH⁺ 404.

Intermediate 12 - 6-Amino-2_l3-dihvdro-1/-/-isoindol-1-one

6-Nitro-2,3-dihydro-1H-isoindol-1-one (33mg) was dissolved in glacial acetic acid (2ml) and added to 10% palladium on carbon (6mg). The mixture was stirred under a hydrogen atmosphere (1Atm.) for 72h. The mixture was filtered through a celite cartridge (10g), and the filtrate diluted with ethanol and loaded onto a SPE cartridge (SCX-2, 5g). The product was eluted with ammonia in methanol (2N). The appropriate ammonical fractions were combined and evaporated in vacuo to give 6- amino-2,3-dihydro-1/-/-isoindol-1-one (20mg). LC/MS; Rt O.74min.

Intermediate 13 - 6-Nitro-2,3-dihvdro-1H-isoindol-1-one

Methyl 2-methyl-5-nitrobenzoate (1.0g, Salor) was dissolved in dry chloroform (40ml). N-Bromosuccinimide (1.09g) was added, followed by benzoyl peroxide (200mg) and the mixture was heated at reflux for 18h. The reaction mixture was cooled to room temperature and evaporated in vacuo. The residual solid was purified by chromatography on a silica cartridge (10Og) and eluting with an ethyl acetate / cyclohexane gradient (0 to 50%) over 60min. Fractions containing methyl 2- (bromomethyl)-5-nitrobenzoate were combined and evaporated to give a mixture with methyl 2-methyl-5-nitrobenzoate (1.16g) (72:28). This mixture was used without further purification. A portion of the solid (200mg) was dissolved in methanol (4.5ml) and 0.880 ammonia (0.14ml) was added. The solution was heated in a sealed vessel by microwave irradiation at 100⁰C for 5min. The solvent was removed in vacuo and the residue partitioned between ethyl acetate and water. The organic phase was separated and the aqueous extracted with ethyl acetate (x2). The organic phases were combined and evaporated in vacuo. The residue was purified first by flash chromatography (silica cartridge, 2Og, eluting with an ethyl acetate / cyclohexane gradient (0 to 100%) over 20min, then 0 to 20% ethyl acetate / (methanol-DCM, 1 :1) over 10min. Product containing fractions were combined and evaporated in vacuo and purified further by mass directed autoprep to give the title compound (33mg). LC/MS; Rt 2.04min.

Example 13 - 6-((4-fEthyl(1 H-indazol-4-yl)amino1-5-fluoro-2-pyrimidinyl}amino)-2- methyl-2,3-dihvdro-1 H-isoindol-1 -one

Λ/-(2-Chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1H-indazol-4-amine (29mg) and 6-amino- 2-methyl-2,3-dihydro-1 H-isoindol-1 -one (19mg) were dissolved in a mixture of acetone / water / cone, hydrochloric acid (1.0ml, 150:100:1 ). The solution heated at

160^cC by microwave irradiation in a sealed vessel for 30min. The reaction mixture was evaporated to dryness and the crude residue was purified by mass directed autoprep to give 6-({4-[ethyl(1H-indazol-4-yl)amino]-5-fluoro-2-pyrimidinyl}amino)-2- methyl-2,3-dihydro-1 H-isoindol-1 -one (14mg). LC/MS; Rt 2.87min, MH⁺ 418.

Intermediate 14 - 6-Amino-2-methyl-2,3-dihvdro-1 H-isoindol-1 -one

2-Methyl-6-nitro-2,3-dihydro-1 H-isoindol-1 -one (58mg) was dissolved in glacial acetic acid (4ml) and added to 10% palladium on carbon (10mg). The mixture was stirred under a hydrogen atmosphere (1Atm) for 48h. The mixture was filtered through a celite cartridge (10g), and the filtrate diluted with ethanol and loaded onto a SPE cartridge (SCX-2, 5g). The product was eluted with ammonia in methanol (2N). The appropriate ammonical fractions were combined and evaporated in vacuo to give the title compound (38mg). LC/MS; Rt 1.57min.

Intermediate 15 - 2-Methyl-6-nitro-2,3-dihvdro-1 H-isoindol-1 -one

A mixture of methyl 2-(bromomethyl)-5-nitrobenzoate with methyl 2-methyl-5- nitrobenzoate (200mg, 72:28, EP357510) was dissolved in methanol (3ml) and methylamine in methanol (2N, 1.1 ml) was added. The solution was heated in a sealed vessel by microwave irradiation at 100⁰C for 5min. The solvent was removed in vacuo and the residue triturated with water and filtered. The solid was dissolved in minimum volume of DCM and applied to a SPE cartridge (silica, 5g). The cartridge was eluted with a stepped gradient of ethyl acetate / cyclohexane (10 to 100%). The product containing fractions were combined and evaporated in vacuo to give the title compound (58mg). LC/MS; Rt 2.15min.

Method 2

Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1/-/-indazol-4-amine (30mg) and the appropriate aniline (1.1 eq) were dissolved in a mixture of acetone / water / cone, hydrochloric acid (1.0ml, 150:100:1). The solution was heated by microwave irradiation in a sealed vessel at 160°C for 30min. The reaction mixture was evaporated to dryness and the crude residue purified by mass directed autoprep to give the title compound.

The following examples were prepared using Method 2;

Method 3

A mixture of Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-N-methyl-1H-indazol-4-amine (28.0mg) and aniline (0.15mmol) in acetone / water / cone, hydrochloric acid (150:100:1 , 1.5 ml) was heated at 8O⁰C for 2 days. The reaction was evaporated to dryness and the crude residue was dissolved in methanol and was loaded on to an SCX-2 SPE cartridge, which was eluted with methanol followed by 10% ammonia/methanol. The methanolic ammonia fraction was evaporated, and the residue was further purified by mass directed autoprep to give the desired product.

The following examples were prepared using Method 3:

Intermediate 16 - /V-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-methyl-1H-indazol-4-amine

Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-methyl-1-({[2-(trimethylsilyl)ethyl]oxy}methyl)- 1/-/-indazol-4-amine (2.12g) in 5M hydrochloric acid / IPA (2:1 , 60ml) was heated at 55°C for 3.5h. The cooled reaction was then evaporated to dryness. The resulting residue was basified with saturated sodium carbonate solution (70ml) and extracted with ethyl acetate (3x 70ml). The combined organic extracts were washed with water (3x 70ml), dried (MgSO₄) and the solvent evaporated. The residue was purified by chromatography on a silica cartridge (100g) eluting with an ethyl acetate / cyclohexane gradient (0-50%) over 40min to give, after evaporation of the solvents Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-methyl-1 H-indazol-4-amine (285mg). LC/MS; Rt 2.97min. MH⁺ 278.

Intermediate 17 - Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-methyl-1-((r2- (trimethylsilyl)ethvHoxy}methyl)-1 H-indazol-4-amine

Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-1-({[2-(trimethylsilyl)ethyl]oxy}methyl)-1 H-indazol- 4-amine (1.938g) was dissolved in DMF (20ml) and cooled to 0⁰C in an ice-bath under nitrogen. Sodium hydride (60% in mineral oil, 236mg) was added and the mixture stirred for 15min. Methyl iodide (459μl) was added and the mixture was allowed to warm at room temperature and stirred for 3h. Saturated ammonium chloride (5ml) was added and the mixture reduced to dryness. The crude residue was partitioned between ethyl acetate and water. The organic extract was washed with water (3x 15ml), dried (MgSO₄) and the solvent evaporated in vacuo to give Λ/- (2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-methyl-1-({[2-(trimethylsilyl)ethyl]oxy}methyl)-1 H- indazol-4-amine. LC/MS; Rt 3.82min. MH⁺ 408.

Intermediate 18 - 3-(4-methyl-1.3-oxazol-5-yl)aniline

A solution of 4-methyl-5-(3-nitrophenyl)-1,3-oxazole (5Og) in methanol (600ml) was prepared in parr-shaker bottle; Pd/C (5g) and hydrogenated at 3.5kg pressure for 2Oh. The reaction mixture was filtered through celite and the filtrate was concentrated. The residue was purified by chromatography on silica eluting with 40% ethyl acetate in pet. ether to get 3-(4-methyl-1 ,3-oxazol-5-yl)aniline as a yellow solid. (18g).

Intermediate 19 - 4-methyl-5-(3-nitrophenyl)-1 ,3-oxazole

To a cooled (0⁰C) solution of 1-isocyanatoethyl 4-methylphenyl sulfone (6Og, 0.287mol) in dry methanol (800ml), dry powdered potassium carbonate was added and stirred at room temperature for 1 h. The reaction was recooled to O⁰C and 3- nitrobenzaldehyde added. The reaction mixture was warmed to room temperature, followed by heating at 55°C overnight with stirring. The excess solvent was removed under reduced pressure and the residue was dissolved in water (100ml). The mixture was extracted with ethyl acetate (3x 350ml). The combined organic phases were washed with brine, water, dried over anhydrous sodium sulfate and concentrated. The crude product was purified by chromatography on silica eluting with 15% ethyl acetate in pet. ether, to give, after evaporation of the solvents, 4-methyl-5-(3- nitrophenyl)-1 ,3-oxazole as yellow solid. (5Og).

Intermediate 20 - 1-isocvanatoethyl 4-methylphenyl sulfone

To cooled (O⁰C) solution of {1-[(4-methylphenyl)sulfonyl]ethyl}formamide (11Og, 0.484mol) in dry THF (1000ml), phosphorus oxychloride (148.4g, 0.968mol) was added slowly and stirred for 30min. Triethylamine (293.8g, 2.9mol) was added dropwise over 30min and the reaction mixture stirred for 2hr at 0⁰C. The reaction was quenched with water, extracted with ethyl acetate (3x 350ml). The combined organic extracts were washed with brine, water, dried over anhydrous sodium sulfate and concentrated. The crude 1-isocyanatoethyl 4-methylphenyl sulfone was taken to next step without further purification. Yield (crude): 6Og.

Intermediate 21 - (1-f(4-methylphenyl)sulfonyllethyl)formamide

A stirred mixture of dry acetonitrile (500ml) and dry toluene (500ml) was cooled to - 30°C, followed by the addition of formamide (50.7g), acetaldehyde (37g) and trimethylsilylchloride (106.4g) at -30⁰C under a nitrogen atmosphere. The reaction mixture was stirred at room temperature for 1 h and then warmed to 55°C. p-Toluene sulphonic acid was added and the reaction mixture was stirred at this temperature overnight. The reaction mixture was cooled to 0⁰C, tert-butyl ether (300ml) added and the mixture stirred for 1 h. The white precipitate formed was isolated by filtration and dried. The crude {1-[(4-methylphenyl)sulfonyl]ethyl}formamide was taken into next step without further purification. Yield (crude): 110g

Intermediate 22 - 6-amino-1.4-dihvdro-2/-/-3.1-benzoxazin-2-one

A mixture of 6-nitro-1 ,4-dihydro-2H-3,1-benzoxazin-2-one (23.0 g, WO9940094), methanol (~60ml) and glacial acetic acid (450ml) was heated to 85°C and the iron powder (19.7g, 3eq.) added in portions. After 5 min, when ~1 equivalent of the iron had been added, an exotherm set in and the reaction mixture temperature rose to 95⁰C. The rest of the iron was slowly added, the temperature kept at 95⁰C for a further 15 min and then the mixture was allowed to cool. A precipitate formed when the temperature fell to ~60°C. When cold, the mixture was rendered alkaline with .880 ammonia and the product extracted with THF (7x 300 ml). A 3-phase system formed during this process, and the whole was filtered (slowly) through Celite held in the funnel with cloth instead of filter paper. The combined extracts were dried with sodium sulphate and evaporated to yield 6-amino-1 ,4-dihydro-2H-3,1-benzoxazin-2- one as a dark mustard-coloured solid (13g). LC/MS; Rt 0.79min.

Method 4

A mixture of Λ/-(2,5-dichloro-4-pyrimidinyl)-N-ethyl-1H-indazol-4-amine (30.0mg) and the appropriate aniline (0.146mmol) in acetone / water / cone, hydrochloric acid (150: 100: 1 , 1.5ml) was heated at 80°C for 16-24h. The reaction was then evaporated to dryness and the crude residue was purified by mass directed autoprep to give the desired product.

The following examples were prepared using Method 4

PB61506-1

Intermediate 23 - /V-(2,5-dichloro-4-pyrimidinyl)-Λ/-ethyl-1H-indazol-4-amine

N-(2,5-dichloro-4-pyrimidinyl)-N-ethyl-1-({[2-(trimethylsilyl)ethyl]oxy}methyl)-1 H- indazol-4-amine (520mg) in 5M hydrochloric acid / IPA (2:1 ; 15ml) was heated at 55⁰C for 6.5h. The cooled mixture was evaporated to dryness in vacuo and the crude residue was left at room temperature overnight. The residue was basified with saturated sodium carbonate solution (20ml) and extracted with ethyl acetate (3x 30ml). The combined organic extracts were washed with water (2x 20ml), dried (MgSO₄) and the solvent evaporated in vacuo. The residue was purified by chromatography on a silica cartridge (5Og) eluting with an ethyl acetate / cyclohexane gradient (0-50%) over 40min. Then product fractions were combined and reduced to dryness in vacuo. The residue was purified by mass directed autoprep to give, after evaporation of the solvents, Λ/-(2,5-dichloro-4-pyrimidinyl)-Λ/-ethyl-1H-indazol-4- amine (79mg). LC/MS; Rt 3.33min. MH⁺ 308, 310.

Intermediate 24 - N-(2.5-dichloro-4-pyrimidinyl)-N-ethyl-1-((r2- (trimethylsilyl)ethylloxy}methyl)-1 H-indazol-4-amine

N-(2,5-dichloro-4-pyrimidinyl)-1-({[2-(trimethylsilyl)ethyl]oxy}methyl)-1 H-indazol-4- amine (870mg) was dissolved in DMF (20ml) at room temperature under nitrogen. Cesium carbonate (760mg) was added and the mixture stirred for 30min. Ethyl iodide (255μl) was added and the mixture was left to stir at 60⁰C for 1.5h. The solvent was evaporated in vacuo and the residue partitioned between ethyl acetate and water. The organic extract was washed with water (3x 15ml), dried (MgSO₄) and reduced to dryness. The residue was purified by chromatography on a silica cartridge (5Og) eluting with an ethyl acetate / cyclohexane gradient (0-50%) over 40min to give N- (2,5-dichloro-4-pyrimidinyl)-N-ethyl-1-({[2-(trimethylsilyl)ethyl]oxy}methyl)-1 H-indazol- 4-amine (520mg). LC/MS; Rt 4.08min. MH⁺ 438, 440.

Intermediate 25 - N-(2.5-dichloro-4-pyrimidinyl)-1-((f2- (trimethylsilyl)ethvnoxy)methyl)-1 H-indazol-4-amine

A mixture of 1-({[2-(trimethylsilyl)ethyl]oxy}methyl)-1 H-indazol-4-amine and 2-({[2- (trimethylsilyl)ethyl]oxy}methyl)-2H-indazol-4-amine (5g) was mixed with 2,4,5- trichloropyrimidine (2.17ml, Aldrich) in N,N-diisopropylethylamine (40ml) and heated at 15O⁰C for 5h. The reaction was then evaporated and the residue was purified by chromatography on silica cartridges (2x 100g) eluting with ethyl acetate / cyclohexane gradients (0-100%) over 40min to give, after evaporation of the solvents, N-(2,5-dichloro-4-pyrimidinyl)-1 -({[2-(trimethylsilyl)ethyl]oxy}methyl)-1 H- indazol-4-amine (945mg). LC/MS; Rt 3.80min. MH⁺ 410.

Example 36 - Λ/⁴-ethyl-5-fluoro-/v⁴-1H-indazol-4-yl-Λ/²-r3-(methylsulfonvnphenyll-2,4- pyrimidinediamine trifluoroacetate

Λ/-(2-Chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1/-/-indazol-4-amine (29.1 mg) was placed in a microwaveable vessel, suspended in water (1.5ml) acetone (1ml) and 2N HCI (20μl) and treated with [3-(methylsulfonyl)phenyl]amine hydrochloride (31.4mg, Acros) This was then irradiated in the sealed vessel using a Biotage microwave at 160⁰C for 1h. The reaction mixture was allowed to cool, evaporated to dryness, dissolved in methanol and applied to a SCX-2 column (500mg) that had been preconditioned with methanol. The column was washed with methanol (2ml) and the crude product was eluted with methanolic ammonia solution (2N, 2ml). The solution was evaporated to dryness, dissolved in DMSO and purified by Mass Directed HPLC. The fractions containing product were evaporated to dryness to give the title compound (0.02Og) LC/MS; Rt 3.04min, MH⁺ 427.

Similarly prepared were the following:

Example 54 - 5-((4-rethyl(1 H-indazol-4-yl)amino1-5-fluoro-2-pyrimidinyl)amino)-1 ,3- dihydro-2H-benzimidazol-2-one trifluoroacetate

Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1/-/-indazol-4-amine (16.3mg) was placed in a microwaveable vessel, suspended in water (1.5ml), acetone (1ml) and hydrochloric acid (2N, 20μl) and treated with 5-amino-1 ,3-dihydro-2H-benzimidazol- 2-one (12.5mg, ASINEX-REAG). The sealed vessel was irradiated in a biotage microwave at 70⁰C for 1h. The reaction was treated with 5-amino-1 ,3-dihydro-2H- benzimidazol-2-one (8.3mg, ASINEX-REAG), and irradiated at 100°C for 1h. The reaction was transferred to a greenhouse tube and heated at 70⁰C under reflux conditions over the weekend. The reaction mixture was allowed to cool, then evaporated to dryness, dissolved in methanol and applied to a SCX-2 column (500mg) that had been preconditioned with methanol. The column was washed with methanol (2ml) and the crude product was eluted with methanolic ammonia solution (2N, 2ml). The solution was evaporated to dryness, dissolved in DMSO and purified by Mass Directed HPLC. The fractions containing product were evaporated to dryness to give the title compound (0.0057g) LC/MS; Rt 2.63min, MH⁺ 404.95.

Intermediate 26 - 1-acetyl-Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1H-indazol-4- amine

To a mixture of Λ/-{3-[(2-chloro-5-fluoro-4-pyrimidinyl)(ethyl)amino]-2- methylphenyljacetamide (7.2g) in chloroform (45ml) was added potassium acetate (2.3g), acetic anhydride (2.11ml), acetic acid (1.25ml), 18-crown-6 (1.2g), and t-butyl nitrite (5.28ml). The mixture was stirred at reflux overnight and then extracted with saturated aqueous sodium bicarbonate solution. The organic layer was separated using a hydrophobic frit and the solvent evaporated in vacuo. The residue was purified by chromatography, on silica cartridges (2x 10Og) and eluting with a cyclohexane / ethyl acetate gradient. The title compound was isolated by evaporation in vacuo of the desired fractions (5.97g). LC/MS; Rt 3.3min, MH⁺ 333 / 335.

Intermediate 27 - 1-acetyl-Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1 /-/-indazol-4- amine

A suspension of Λ/-{3-[(2-chloro-5-fluoro-4-pyrimidinyl)(ethyl)amino]-2- methylphenyljacetamide (37.Og) and potassium acetate (11.25g) in chlorobenzene (185ml) was treated sequentially with glacial acetic acid (9.8ml), acetic anhydride (16.25ml) and t-butyl nitrite (30.2ml) at ambient temperature. The mixture was stirred for 10min, then warmed to 70⁰C and held at this temperature for 7h before slowly cooling to ambient temperature. The orange mixture was treated cautiously with 1 M aqueous potassium carbonate (259ml) over 20min, stirred for a further 30min and the layers were separated. The organic layer was washed with water (150ml) and concentrated to ca 110ml under reduced pressure. The residue was diluted by addition of propan-1-ol (296ml) and the organic layer was concentrated to ca 110ml under reduced pressure. Further propan-1-ol (222ml) was added and the organic layer was concentrated to ca 110ml under reduced pressure. The residue was diluted with further propan-1-ol (1 10ml), warmed to 40-50⁰C, and treated with water (222ml) over 20min. The slurry was aged at ca 50⁰C for 15min, then cooled to 5°C over 3h. The product was isolated by filtration, washed with water / propan-1-ol (1 :1 , 2x 75ml), and dried in vacuo to give the title product as a yellow solid (34.91 g).

Intermediate 28 - Λ/-{3-f(2-chloro-5-fluoro-4-pyrimidinyl)(ethyl)aminol-2- methylphenvDacetamide

A mixture of Λ/-{3-[(2-chloro-5-fluoro-4-pyrimidinyl)amino]-2-methylphenyl}acetamide (6.9g) in anhydrous DMF (50ml) was stirred with ethyl iodide (2.06ml) and potassium carbonate (3.6g). The reaction mixture was stirred for approximately 5.5h and the solvent removed in vacuo. The residue was partitioned between ethyl acetate (50ml) and water (25ml). The organic layer was separated and washed with water (25ml x2) and dried with brine (25ml) and over magnesium sulphate. The solvent was evaporated in vacuo to give the title compound (7.2g). LC/MS; Rt 2.8min, MH⁺ 323, 325.

Intermediate 29 - Λ/-(3-r(2-chloro-5-fluoro-4-pyrimidinyl)aminol-2- methylphenyl}acetamide

A mixture of 3-acetylamino-2-methylaniline (EP425345A1 , 7.72g) and 2,4-dichloro-5- fluoropyrimidine (7.84g) in methanol / water (1 :3, 188ml) was stirred at 70⁰C for 1 h. The reaction mixture was cooled and then filtered. The residue was washed with ice cold methanol / water (1 :3) and the solid dried in vacuo at 50⁰C and then at 65°C to give the title compound as a white solid (6.9g). LC/MS; Rt 2.2min, MH⁺ 295 / 297.

Intermediate 30 Λ/-(3-[(2-chloro-5-fluoro-4-pyrimidinyl)amino1-2- methylphenvDacetamide

2,4-Dichloro-5-fluoropyrimidine (23.Og), 3-acetylamino-2-methylaniline (EP425345A1 , 23.71 g) and sodium acetate (22.52g) were slurried in IPA (115ml) and water (345ml), and warmed to 70⁰C. After 10min at 70⁰C The mixture was seeded with Λ/-{3-[(2- chloro-5-fluoro-4-pyrimidinyl)amino]-2-methylphenyl}acetamide, the mixture was stirred at 7O⁰C for 5h, then allowed to cool to ambient temperature overnight. The mixture was further cooled to 5-10°C, aged for 2h and the solid isolated by filtration. The solid was washed with water / IPA (3:1 , 100ml), then water (2x 100ml), and dried in vacuo to give the title product as a white solid (35.15g).

Example 55 - 3-r6-({4-rethyl(1/-/-indazol-4-vπamino1-5-fluoro-2-pyrimidinyl)amino)-1/-/- indazol-1 -yli-1 -propanol

To a flask charged with 1-acetyl-Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1/-/- indazol-4-amine (5.Og) and 3-(6-amino-1H-indazol-1-yl)-1-propanol (3.71 g, 80% purity) in IPA (40ml) was added hydrochloric acid (2M, 10ml). The mixture was heated at reflux for 16h. The reaction was allowed to cool and then water added (200ml). To this solution was added aqueous sodium hydroxide (2M) to adjust the pH to 10. The mixture was stirred for 1 h then filtered to afford an off-white solid. This material was chromatographed on a silica cartridge (100g) eluting with a gradient of 0-30% methanol, 1% triethylamine, in DCM over an 80min period. Concentration of the appropriate fractions afforded material which still required further purification. This material was split into 2 portions. The first portion was chromatographed on a silica cartridge (100g) eluting with DCM / MeOH / triethylamine (94:5:1). The pure fractions were reduced to dryness to give the title compound as a pale yellow solid (1.25g). LC/MS; Rt 2.92min, MH⁺ 445.

The impure fractions were reduced to dryness, combined with the second portion of material and chromatographed on a silica cartridge (100g) eluting with DCM / MeOH / triethylamine (94:5:1). The resulting impure product fractions from the second column were combined, concentrated and dried in vacuo. This material was dissolved in 10ml of DMF to give approx 14ml of solution. 9 chromatographic separations were performed by taking 1.6ml of sample solution and adding 3.4ml of DMF and 0.7ml of TFA for each separation. A column of 7micron Kromasil C8 (25 x 5cm) was used with a gradient of 0 to 50%B in 40min (where A is water +0.25% TFA and B is acetonitrile +0.25% TFA). The peaks at retention time 35.5 to 37mins were bulked from all the chromatographic separations. The bulked fraction was evaporated to remove acetonitrile and divided into 2 portions. Each portion was applied to an Amberchrom CG161 column (25cm x 2 cm) to adsorb the compound. The column was washed in turn with water, 0.2M ammonium hydroxide, water, and IPA and the compound eluted with acetone. The acetone elution samples were bulked and dried. This material was dissolved in methanol and passed through an aminopropyl cartridge (2Og) then concentrated in vacuo and dried at 40⁰C under vacuum to afford a further batch of the title compound (1.3g). NMR; [D₆-DMSO] δH 8.27,(1 H, s), 7.98,(1 H, d), 7.92,(1 H, d), 7.90,(1 H, d), 7.58,(1 H, d), 7.51 ,(1 H, d), 7.38,(1 H, m), 7.26,(1 H₁ dd), 7.05,(1 H, d), 4.33,(2H, t), 4.20,(2H, q), 3.38,(2H, t), 1.97,(2H, m), 1.26,(3H, t).

Intermediate 31 - 3-(6-amino-1 H-indazol-1-yl)-1-propanol

6-Aminoindazole (5g, Aldrich) was dissolved in methanol (100ml) and benzaldehyde (4.22ml) was added. The reaction mixture was stirred at room temperature under nitrogen for 1 h. The solvent was removed under vacuum and the resulting white solid was dissolved in dry DMF (50ml). Sodium hydride (1.77g, 60% in mineral oil) was added and the reaction mixture was stirred in a cold water bath under a nitrogen atmosphere for 10min. [(3-Bromopropyl)oxy](1 ,1-dimethylethyl)dimethylsilane (10.17ml, Aldrich) was added and the reaction mixture was stirred at room temperature under nitrogen atmosphere for 1h. Saturated aqueous ammonium chloride (50ml) was added to quench the reaction. The mixture was partitioned between ethyl acetate (100ml) and water (50ml), the two phases were separated, the organic layer dried (hydrophobic frit) and concentrated under vacuum. The residue was dissolved in methanol (15ml) and hydrochloric acid (5ml, 1 M) was added. The reaction mixture was left in solution at room temperature for 16h. The solvent was removed under vacuum, the residue partitioned between ethyl acetate (100ml), water (50ml) and saturated aqueous sodium hydrogencarbonate (50ml). The phases were separated and the organic layer was dried (hydrophobic frit) and concentrated under vacuum. The residue was absorbed onto silica and purified by chromatography eluting with a DCM / (methanol + 1%triethylamine) gradient (0-25%). The fractions containing the desired product were combined and concentrated under vacuum to give the title compound as a yellow solid (3.35g). LC/MS; Rt 1.68min, MH⁺ 192. Purity: 75%. Intermediate 32 - /V-f3-r(2-chloro-5-fluoro-4-pyrimidinyl)(ethyl)amino1-2-

methylphenvDacetamide

A suspension of potassium carbonate (11.25g) in DMF (100ml) was treated with N- {3-[(2-chloro-5-fluoro-4-pyrimidinyl)amino]-2-methylphenyl}acetamide (20.Og) over ca 1min. The resulting yellow suspension was warmed to 50°C and treated with ethyl iodide (6.0ml) over 15min, rinsing in with further DMF (5ml). The mixture was stirred at this temperature for 14h, warmed to 60⁰C and treated with water (100ml) dropwise over 10min. The mixture was seeded with Λ/-{3-r(2-chloro-5-fluoro-4- pyrimidinyl)(ethyl)aminol-2-methylphenyl)acetamide. aged for 30min at 6O⁰C and treated with further water (100ml) over 30min. The yellow suspension was aged for 1h, cooled to 10⁰C and aged for 2h. The product was isolated by filtration, washed with water / DMF (2:1 , 60ml) and then water (2x 60ml). The product was dried in vacuo at 55-60°C to give the title product as a yellow solid (20.1g).

Example 56 - Λ/²-1H-1.2,3-Benzotriazol-5-yl-/V⁴-ethyl-5-fluoro-Λ/⁴-1 H-indazol-4-yl-2,4- pyrimidinediamine

1 -Acetyl-Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1 H-indazol-4-amine (5.Og), 1 H- 1 ,2,3-benzotriazol-5-amine (2.4g, Lancaster), was stirred with IPA (30ml) and hydrochloric acid (2N, 6ml,), at reflux for 18h. The reaction mixture was allowed to cool to room temperature, diluted with water (50ml) and stirred in an ice/water bath for 1 h. The solid was isolated by filtration, triturated with ether, refiltered and dried under vacuum. The dry solid was dissolved in methanol (minimium volume) and ether added until solid precipitated. The solid was isolated by filtration, dissolved in methanol and passed through an aminopropyl SPE cartridge and the methanol evaporated in vacuo to give the impure product (0.352g).

The aqueous filtrate was basified with 2N sodium hydroxide and extracted with ethyl acetate (x3). The ethyl acetate extracts were combined, dried (MgSO4), filtered and the solvent evaporated in vacuo. The residue was recrystalised from ethyl acetate and the crystals dried at 4O⁰C under high vacuum to give the title compound (0.893g). LC/MS; Rt 2.90min, MH⁺ 390.

NMR; [D₆-DMSO] δH 9.63,(NH, s), 8.45,(1 H, s), 7.98,(1 H, d), 7.92,(1 H, s), 7.80- 7.78,(1 H₁ d), 7.52-7.48,(2H, m), 7.40-7.36,(1 H, m), 7.05-7.03,(1 H, d), 4.17-4.12,(2H, m), 1.27-1.24,(3H, t).

A second crop of the title compound was obtained from the filtrate and dried at 40⁰C under high vacuum (0.25g). LC/MS; Rt 2.98min, MH⁺ 390.

All filtrates and impure products were combined and reduced to dryness to give crude product (3.2g). This material was dissolved in 17ml of DMF to produce a solution of 190mg/ml. A trial run of 1ml of this solution in DMF (4ml) and TFA (0.5ml) was injected onto a column of 7micron Kromasil C8 (25 x 5cm) and eluted with a gradient of 0 to 100%B in 40min (where A is water + 0.25% TFA and B is acetonitrile + 0.25% TFA). The peak with MH⁺ = 390 was collected. This method was then used on 5 x 3ml portions of the solution in DMF (2ml) and TFA (0.5ml). The peaks from all the chromatographic separations were combined and evaporated to remove acetonitrile and applied to an Amberchrom CG 161 column (25cm x 2 cm) to adsorb the compound. The column was washed with 0.2M ammonium hydroxide (200ml), then water (400ml) until neutral pH. The product was eluted with methanol and dried

(1 -4g).

This material was dissolved in methanol and passed through an aminopropyl cartridge (2Og) then concentrated in vacuo and dried at 40⁰C under vacuum to afford a further batch of the title compound (1.04g, -80% purity).

Example 57 - N⁴-ethyl-5-fluoro-N⁴-1 H-indazol-4-yl-N²-r4-(4-methyl-1- piperazinyl)phenyll-2,4-pyrimidinediamine

A mixture containing 1-acetyl-Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1/-/-indazol- 4-amine (75mg) and 4-(4-methyl-1-piperazinyl)aniline (56mg, Acros Organics) in IPA (2ml) and hydrochloric acid (2N, 0.2ml) was irradiated in a sealed microwave vessel at 145⁰C for 1.25h. The resulting solution was allowed to cool and was then applied to an SCX cartridge which had been pre-treated with methanol. The cartridge was eluted with methanol followed by 10% ammonia in methanol. Appropriate fractions were combined and evaporated in vacuo and the residual black gum purified by mass directed autoprep to give a gum. This was dissolved in methanol and applied to an aminopropyl cartridge. Elution with methanol gave N⁴-ethyl-5-fluoro-N⁴-1 H- indazol-4-yl-N²-[4-(4-methyl-1-piperazinyl)phenyl]-2,4-pyhmidinediamine as an off white solid (36mg). LC/MS; MH⁺ 447, Rt 2.34min.

Example 58 - Λ/⁴-Ethyl-5-fluoro-Λ/⁴-1H-indazol-4-yl-Λ/²-[3-(3-pyridinyl)phenyll-2,4- pyrimidinediamine

Λ^-Ethyl-S-fluoro-Λ/^tS-CS-pyridinyOphenyll-Λ^-ti-Ct^-^rimethylsilyOethylloxyϊmethyl)- 1/-/-indazol-4-yl]-2,4-pyrimidinediamine (43mg) dissolved in dry THF (1-2ml) was treated with TBAF in THF (1 N, 2-3eq). The mixture was heated at reflux for 16h, diluted with water and extracted with ethyl acetate (x3). The organic phases were combined, dried (MgSO₄) and evaporated to dryness. The crude residue was purified twice by mass directed autoprep and the residue taken up in methanol and passed through an aminopropyl SPE cartridge (1g). The solvent was evaporated in vacuo to leave the title compound (9mg). LC/MS; MH⁺ 426, Rt 3.07min.

Intermediate 33 Λ/⁴-Ethyl-5-fluoro-Λ/²-r3-(3-pyridinyl)phenyll-/V⁴-π-αr2-

(trimethylsilyl)ethylloxy)methyl)-1 /-/-indazol-4-yll-2.4-Pyrimidinediamine

Λ/-(2-Chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1-({[2-(trimethylsilyl)ethyl]oxy}methyl)-1/-/- indazol-4-amine (50mg) and 3-(3-pyridinyl)aniline (Maybridge CombiChem, 2eq) , chloro[2'-(dimethylamino)-2-biphenylyl]palladium - (1R,4S)-bicyclo[2.2.1]hept-2- yl[(1 S,4R)-bicyclo[2.2.1]hept-2-yl]phosphane (1 :1 , 0.2eq), cesium carbonate (3eq), were dissolved in dioxane / water (2.5ml, 4:1 ). The solution was heated in a sealed vessel at 150⁰C for 1 h by microwave irradiation. The reaction mixture was partitioned between ethyl acetate and water. The aqueous extracted with ethyl acetate (x2). The organic phases were combined, dried (MgSO₄) and evaporated to dryness .The crude residue was purified on a silica cartridge (5g), eluting with an ethyl acetate / cyclohexane gradient (0 to 100%) over 30min. The appropriate fractions were combined and evaporated in vacuo to give the title compound (43mgJ. LC/MS; MH⁺ 556, Rt 4.0min.

Example 59 - 3-[6-((4-FEtIwI(I H-indazol-4-vDaminol-5-fluoro-2-pyrimidinyl)amino)-3- methyl-2H-indazol-2-vπ-1-propanol

1 -Acetyl-Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1 H-indazol-4-amine (75mg) and 2-(3-{[(1 , 1 -dimethylethyl)(dimethyl)silyl]oxy}propyl)-3-methyl-2H-indazol-6-amine (86mg), were dissolved in IPA (4ml). Concentrated hydrochloric acid (100μl) was added and the mixture heated at 80⁰C for 18h. The reaction mixture was diluted with methanol and filtered through an aminopropyl SPE cartridge (10g), the cartridge was washed with methanol and the combined filtrate and washings reduced to dryness. The residue was applied to a silica cartridge and eluted with an (1% Et3N / 50%MeOH / 50%EtOAc) / cyclohexane gradient (0 to 50%). The eluted fractions were combined and reduced to dryness. The residue was purified by mass directed autoprep to give the title compound (41 mg). LC/MS; MH⁺ 461 , Rt 2.77min.

Intermediate 34 - 2-(3-{r(1 ,1-Dimethylethyl)(dimethyl)silylloxy)propyl)-3-methyl-2/-/- indazol-6-amine

2-(3-{[(1 J-dimethylethyl)(dimethyl)silyl]oxy}propyl)-3-methyl-6-nitro-2/-/-indazole (2.08g) in dry ethanol (50ml) and ethyl acetate (50ml) was added to palladium on carbon (10%, 300mg) and stirred under 1Atm. of hydrogen for 3h. The mixture was filtered through celite and the filtrate evaporated to dryness. The residue was purified by chromatography on a silica cartridge (100g), eluting with an ethyl acetate / cyclohexane gradient (0 -100%) over 40min, to give, after evaporation of the solvents, the title compound (1.64g). LC/MS; MH⁺ 320, Rt 3.24min.

Intermediates 35 & 36 - 2-(3-(r(1.1-dimethylethvn(dimethyl)silylloxy)propyl)-3-methyl- 6-nitro-2/-/-indazole and 1-(3-(F(1.1-dimethylethyl)(dimethyl)silylloxy)propyl)-3-methyl- 6-nitro-1H-indazole

3-Methyl-6-nitro-1/-/-indazole (5g, WO 2002059110) in dry DMF (40ml) was cooled in an ice / water bath to ~5°C, stirring under nitrogen. Sodium hydride (60% in mineral oil 1.13g) was added portionwise. The reaction mixture was stirred for a further 30min. in the ice / water bath and [(3-bromopropyl)oxy](1 ,1- dimethylethyl)dimethylsilane (6.72ml) in dry DMF (5ml) was added dropwise. The mixture was allowed to warm to room temperature and stirred for 18h. The reaction mixture was quenched with water and extracted with ethyl acetate (x3). The organic phases were combined, dried (MgSO₄), and evaporated to dryness. The residue was purified on a silica cartridge (100g), eluting with an ethyl acetate / cyclohexane gradient (0 - 50%) over 40min to give a mixture of the two products (9.07g). The mixture was dissolved in cyclohexane and applied to a silica cartridge (100g), the cartridge was eluted with cyclohexane (200ml), 10% ethyl acetate / cyclohexane (200ml) and then 30% ethyl acetate / cyclohexane. Appropriate fractions were combined and reduced to dryness to give the two title compounds:- The eluting product first, 1-(3-{[(1,1-dimethylethyl)(dimethyl)silyl]oxy}propyl)-3-methyl- 6-nitro-1H-indazole (6.4g). NMR; [CDCI₃] δH 8.39,(1 H, s), 7.98-7.96., (1 H, d), 7.74-7.72, (1 H, d), 4.52-4.51 ,(2H₁ t), 3.58-3.55, (2H, t), 2.62,(3H₁ s), 2.16-2.10,(2H, m), 0.92,(9H, s), 0.04,(6H, s). LC/MS; MH⁺ 350, Rt 4.24min. The eluting product second was purified further by trituation with cyclohexane and filtration to give 2-(3-{[(1 ,1-dimethylethyl)(dimethyl)silyl]oxy}propyl)-3-methyl-6-nitro- 2H-indazole (2.08g). NMR; [CDCI₃] δH 8.63,(1 H₁, s), 7.85-7.83, (1 H, d), 7.67-7.65., (1 H₁ d), 4.55-4.52,(2H₁ t), 3.62-3.60, (2H, t), 2.69,(3H, s), 2.23-2.17, (2H, m), 0.91 ,(9H, s), 0.06,(6H, s). LC/MS; MH⁺ 350, Rt 4.24min.

Example 60 - 3-[6-({4-rEthyl(1H-indazol-4-yl)amino1-5-fluoro-2-pyrimidinyl)amino)-3- methyl-1 H-indazol-1 -yll-1 -propanol

1 -Acetyl-Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1 H-indazol-4-amine (75mg) and 1 -(3-{[(1 , 1 -dimethylethyl)(dimethyl)silyl]oxy}propyl)-3-methyl-1 /-/-indazol-6-amine (86mg), were dissolved in IPA (4ml). Concentrated hydrochloric acid (100μl) was added and the mixture heated at 80°C for 18h. The reaction mixture was diluted with methanol and filtered through an aminopropyl SPE cartridge (10g), the cartridge washed with methanol and the combined filtrate and washings reduced to dryness. The residue was applied to a silica cartridge and eluted with an (1% Et3N / 50%MeOH / 50%EtOAc) / cyclohexane gradient (0 to 50%). The eluted fractions were combined and reduced to dryness. The residue was purified by mass directed autoprep to give the title compound (24mg). LC/MS; MH⁺ 461 , Rt 2.98min.

Intermediate 37 - 1-(3-(r(1.1-Dimethylethyl)(dimethyl)silvnoxy>propyl)-3-methyl-1/-/- indazol-6-amine

1 -(3-{[(1 , 1 -dimethylethyl)(dimethyl)silyl]oxy}propyl)-3-methyl-6-nitro-1 H-indazole (6.4g) in dry ethanol (150ml) was added to palladium on carbon (10%, 650mg) and stirred under 1Atm. of hydrogen for 4h. The mixture was filtered through celite and the filtrate evaporated to dryness. The residue was purified on a silica cartridge (100g), eluting with an ethyl acetate / cyclohexane gradient (0 -100%) over 40min, to give, after evaporation of the solvents, the title compound (5.17g). LC/MS; MH⁺ 320, Rt 3.53min.

Example 61 - 4-r6-((4-rEthyl(1 H-indazol-4-yl)aminol-5-fluoro-2-pyrimidinyl}amino)-1 H- indazol-1 -yll-1 -butanol

1 -Acetyl-Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1 H-indazol-4-amine (75mg) and 4-(6-amino-1H-indazol-1-yl)-1 -butanol (55mg), were dissolved in IPA (4ml). Concentrated hydrochloric acid (100μl) was added and the mixture heated at 150°C in a sealed vessel in a microwave for 1h. The reaction mixture was reduced to dryness, dissolved in methanol and loaded onto an SCX SPE cartridge (5g). The cartridge was eluted with a methanolic ammonia (2N) / DCM gradient (0 to 10%). Appropriate fractions were combined and reduced to dryness. The residue was purified further on a silica cartridge (2Og), eluting with a gradient of (1% triethylamine / methanol) / DCM (0-15%) over 30min, to give, after evaporation of the solvents, the title compound (35mg). LC/MS; MH⁺ 461 , Rt 3.02min.

Intermediate 38 - 4-(6-Amino-1H-indazol-1-yl)-1 -butanol

6-Aminoindazole (1g) was dissolved in anhydrous methanol (30ml) and treated with benzaldehyde (0.92ml). The reaction mixture was stirred under nitrogen at room temperature for 90min. The solvent was evaporated in vacuo, the residue dissolved in anhydrous DMF (15ml) and stirred in an ice/water bath under nitrogen. Sodium hydride (60% in mineral oil, 360mg) was added and stirring continued for 15min before addition of the tert-butyl(4-iodobutoxy)-dimethylsilane (1.94ml). The reaction mixture was stirred at room temperature for 45min before quenching with hydrochloric acid (5M, 5ml) and stirring at room temperature overnight. The reaction mixture was partitioned between ethyl acetate and saturated sodium bicarbonate solution. The aqueous phase was extracted with ethyl acetate and the combined organic extracts dried (MgSO₄), filtered and reduced to dryness. The residue was dissolved in DCM, loaded on to a silica cartridge (7Og) and the cartridge eluted with an ethyl acetate / cyclohexane gradient (0 - 100%), then methanol / DCM (0 - 25%) over 30min. Appropriate fractions were combined, reduced to dryness and the residue dried in vacuo to give the title compound (520 mg). LC/MS; Rt 1.79min, MH⁺ 206.

Example 62 - Λ/²-(1.1-dioxido-2,3-dihvdro-1-benzothien-6-yl)-5-fluoro-Λ/⁴-1 /-/-indazol- 4-yl-Λ/⁴-propyl-2,4-Pyrimidinediamine

{4-[(2-Chloro-5-fluoro-4-pyrimidinyl)(propyl)amino]-1 H-indazol-1 -yl}methanol / Λ/-(2- chloro-5-fluoro-4-pyrimidinyl)-Λ/-propyl-1/-/-indazol-4-amine (1 :1 , 48mg) and 2,3- dihydro-1-benzothiophen-6-amine 1 ,1-dioxide, (34mg, Butt Park Screening Library) were dissolved in a mixture of acetone / water / cone, hydrochloric acid (1ml, 150:100:1 ). The solution was heated at 16O⁰C for 30min by microwave in a sealed vessel. The reaction mixture was evaporated to dryness and the crude residue was purified by mass directed autoprep to give the title compound (19mg). LC/MS; MH⁺ 453, Rt 3.14min.

Intermediate 39 - {4-r(2-Chloro-5-fluoro-4-pyrimidinyl)(propyl)amino1-1 H-indazol-1 - vDmethanol / Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-propyl-1 /-/-indazol-4-amine

Λ/-(2-Chloro-5-fluoro-4-pyrimidinyl)-Λ/-propyl-1-({[2-(trimethylsilyl)ethyl]oxy}methyl)- 1/-/-indazol-4-amine (1.69g) was stirred in a mixture of hydrochloric acid (5N) / IPA (45ml, 2:1) at 55°C for 4h. The solvents were evaporated in vacuo and the residue partitioned between ethyl acetate and saturated sodium bicarbonate. The aqueous was extracted with ethyl acetate (x2). The organic phases were combined and dried (MgSO₄) and reduced to dryness in vacuo. The residue was purified on a silica cartridge (10Og), using an ethyl acetate / cyclohexane gradient (0 to 50%) over 60min. The appropriate fractions were combined and the solvents evaporated in vacuo to give the title compound (96mg). LC/MS; MH⁺ 336, Rt 3.16min (CH₂OH), and MH⁺ 306, Rt 3.30min (H).

Intermediate 40 Λ/-(2-Chloro-5-fluoro-4-pyrimidinyl)-Λ/-propyl-1-({[2-

(trimethylsilv0ethvπoxy)methvD-1 H-indazol-4-amine

Λ/-(2-Chloro-5-fluoro-4-pyrimidinyl)-1-({[2-(trimethylsilyl)ethyl]oxy}methyl)-1H-indazol- 4-amine (1.768g) and potassium carbonate (0.8g) were stirred in dry DMF (100ml) at room temperature under nitrogen for 30min. lodopropane (0.92g) was added and stirring continued for 18h. The solvent was evaporated in vacuo and the residue partitioned between ethyl acetate and water. The aqueous was extracted with ethyl acetate (x2). The combined organic phases were dried (MgSO₄) and reduced to dryness in vacuo. The residue was purified on a silica cartridge (10Og), using an ethyl acetate / cyclohexane gradient (0 to 50%) over 60min. The appropriate fractions were combined and evaporated in vacuo to give the title compound (1.69g). LC/MS; MH⁺ 436, Rt 4.02min. Example 63 - Λ^-ethyl-S-fluoro-Λ^-d H-indazol^-vD-Λ^-ri-αrifluoroacetvn^.S-dihvdro- 1/-/-indol-5-yll-2.4-pyrimidinediamine

A mixture of 1-acetyl-Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1H-indazol-4-amine (1.36g), 1-(trifluoroacetyl)-5-indolinamine (1.125g), hydrochloric acid (2N, 1.58ml) in IPA (15ml) was heated in a sealed vial by microwave irradiation for 90min at 120⁰C. The reaction mixture was poured into saturated sodium bicarbonate (50ml), extracted with ethyl acetate (2x 150ml). The combined organic extracts were washed with water (100ml), dried (hydrophobic frit) and concentrated in vacuo. The residue was adsorbed onto a silica cartridge (7Og) and eluted with an ethyl acetate / cyclohexane gradient (20-50%). The required fractions were combined and the solvent was removed under reduced pressure to yield the title compound as a white solid (0.19Og). LC/MS; Rt 3.46min, MH⁺ 486.

Intermediate 41 - 1-(trifluoroacetyl)-5-indolinamine

A solution of 5-nitro-1-(trifluoroacetyl)indoline (2g) in ethanol (50ml) was added to platinum on carbon (5%, 0.2g) and stirred under 1atm. of hydrogen for 1 h. The mixture was filtered through celite and the filtrate concentrated under reduced pressure to yield the title compound as a green solid (1.8g). LC/MS; Rt 2.23min, MH⁺ 231.

Intermediate 42 - 5-nitro-1-(trifluoroacetyl)indoline

N,N-Diisopropylethylamine (2.55ml) was added to a solution of 5-nitroindoline (2g, Aldrich) in DCM (20ml). The solution was cooled to 0°C (ice/salt water bath), trifluoroacetic anhydride (2.60ml) added and the mixture left to stir at 0⁰C for 10min. The solution was poured into saturated sodium bicarbonate (100ml) and extracted with DCM (2x 100ml). The combined organic extracts were washed with water (100ml), dried (hydrophobic frit) and concentrated under reduced pressure. The residue was dissolved in cyclohexane (60ml) and ethyl acetate (40ml), the volume reduced to 50ml and the mixture left to stand overnight. The resulting crystals were filtered off, washed with cyclohexane and dried under vacuum at 50⁰C for 2h to yield the title compound as a solid (2g). NMR; [D₆-DMSO] δH 8.26-8.24,(1 H, m), 8.23- 8.21 , (2H, m), 4.39,(2H, t), 3.37-3.31 ,(2H, m).

Example 64 - /V⁴-ethyl-5-fluoro-Λ/⁴-1H-indazol-4-yl-Λ/²-{4-[(4-methyl-1- piperazinyl)carbonyl]phenyl}-2,4-pyrimidinediamine

A mixture containing 1-acetyl-Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1 /-/-indazol- 4-amine (33mg) and 4-[(4-methyl-1-piperazinyl)carbonyl]aniline (33mg, lnterchim S.A.) in IPA (1 ml) and hydrochloric acid (2N, 0.25ml) was stirred in a sealed microwave vessel and irradiated at 140°C for 1.25h. The reaction mixture was applied to an SCX cartridge which had been pre-treated with methanol. The cartridge was eluted with methanol followed by methanolic ammonia (2M). Appropriate fractions were blown down to dryness and the residue purified by mass directed autoprep. Appropriate fractions were blown down to dryness and the residue dissolved in methanol and applied to an aminopropyl cartridge which had been pre-treated with methanol. The cartridge was eluted with methanol and appropriate fractions reduced to dryness under a stream of nitrogen to give the title compound as a white solid (8mg). LC/MS; MH⁺ 475, Rt 2.45min.

Example 65 - Λ/⁴-ethyl-5-fluoro-N⁴-1 H-indazol-4-yl-N²-[3-(4-methyl-1- piperazinyl)phenyl]-2,4-pyrimidinediannine

A mixture containing 1-acetyl-Λ/-(2-chloro-5-fluoro-4-pyrimidinyl)-Λ/-ethyl-1H-indazol- 4-amine (33mg) and [3-(4-methyl-1-piperazinyl)phenyl]amine (29mg, EP 533267) in IPA (1ml) and hydrochloric acid (2N, 0.25ml) was stirred in a sealed microwave vessel and irradiated at 140⁰C for 1.25h. The reaction mixture was then applied to an SCX cartridge which had been pre-treated with methanol. The cartridge was eluted with methanol followed by methanolic ammonia (2M). Appropriate fractions reduced to dryness under a stream of nitrogen and the residue purified by mass directed autoprep. Appropriate fractions were reduced to dryness under a stream of nitrogen and the residue dissolved in methanol and applied to an aminopropyl cartridge which had been pre-treated with methanol. Elution with methanol followed by reduction to dryness under a stream of nitrogen of the appropriate fractions gave the title compound as a foam (22mg). LC/MS; MH⁺ 447, Rt 2.53min.

Claims

Claims

1. A compound of formula (I):

in which:

R¹, R² and R³ is each independently selected from hydrogen, halogen, -C_0-6hydroxy, -C_1-6 alkyl, -NR⁵R⁶, -CN, -Co-aalkylene-CC^H, OC(O)C_1-6 alkyl, C(O)C_1-6 alkoxy, -C_1-2 alkyl substituted by 1 or more fluorine atoms, -C_1-6 alkoxy, -C(O)NR⁵R⁶,

-OCH₂C(O)NR^sR^b, -NR^bC(O)R^b, -SC_1-6alkyl, -S(O)C_1-6alkyl, -S(O)₂C^alkyl,- NHS(O)₂R⁷, -S(O)₂NR⁵R⁶ or -S(O)₂NR⁵R⁸, such that at least one of R¹, R^2a and R^3a is hydrogen; or

R¹ is hydrogen and one of R² and R³ is hydrogen, C_1-6alkyl, or C_1-6alkoxy and the other is a 5- or 6-membered heteroaryl ring comprising from 1 to 3 heteroatoms selected from O, N and S bonded to the phenyl ring through a ring carbon atom and which heteroaryl ring is unsubstituted or substituted on one or more ring carbon atoms by C_1-6alkyl or oxo (=0), or on a ring nitrogen atom by C_1-6alkyl; or

R¹ is hydrogen, halogen or C_1-6 alkyl and R² and R³ are joined and together form, in combination with the carbon atoms on the phenyl group to which they are attached, a 5- or 6-membered ring wherein: the ring is heterocyclic and comprises one heteroatom selected from O, N or S, the ring is heteroaryl or heterocyclic and comprises two heteroatoms selected from O, N or S, which maybe the same or different, and excluding two S atoms, the ring is heteroaryl and comprises three heteroatoms of which two are N atoms and the third is nitrogen or oxygen, which ring may be unsubstituted or substituted on: one or more ring carbon atoms which each may be independently substituted by oxo (=O) or by 1 or 2 substituents which may be the same or different selected from C_1-6 alkyl, hydroxyl, or halo, one or more ring nitrogen atoms by C_1-6 alkyl, CH₂C_3-7cycloalkyl, hydroxyC_1-6 alkyl, C_1-6 alkylcarbonyl, or C_1-6 haloalkylcarbonyl, and a ring sulphur atom by (O)₂; and

R⁴ is C_1-3 alkyl; R⁵ and R⁶ are is each independently hydrogen or C_1-6alkyl, or R⁵ and R⁶ together with the nitrogen atom to which they are attached form a 5- or 6-membered heterocyclic ring which may comprise a second heteroatom selected from O, S or N and which may be substituted on the second N, if present, by C_1-6alkyl, and which may be substituted on a ring carbon by oxo (=0), Ci_-6alkyl, di-C^alkyl (which may be the same or different), or halogen;

R⁷ is C_1-6 alkyl or phenyl optionally substituted by C_1-6alkyl;

R⁸ is C_3-7cycloalkyl, C_1-6alkyl (optionally terminally substituted with hydroxyl or tetrahydrofuranyl); and

X is halogen; or a salt or solvate, preferably a pharmaceutically acceptable salt or solvate, thereof.

2. A compound of formula (I) as claimed in claim 1 excluding a compound of the formula (IA):

in which:

R^9a and R^1Oa is each independently hydrogen, C_1-6alkyl or CH₂C_3-7cycloalkyl;

R^4a is C_1-3alkyl; and

X^a is halogen; or a salt or solvate, preferably a pharmaceutically acceptable salt or solvate, thereof.

3. A compound of formula (I) as claimed in claim 1 or 2 in which two of R¹, R² and R³ are hydrogen.

4. A compound of formula (I) as claimed any one of claims 1 to 3 in which R¹ and R³ is each hydrogen and R² is a 5- or 6-membered heteroaryl ring or a heterocyclic ring comprising from 1 to 3 heteroatoms selected from O, N and S bonded to the phenyl ring through a ring carbon or nitrogen (if present) atom and which heterocyclic ring is unsubstituted or substituted on one or more ring carbon atoms by C_1-6alkyl or oxo (=0), or on a ring nitrogen atom by C^alkyl.

5. A compound of formula (I) as claimed any one of claims 1 to 3 in which R¹ and R² is each hydrogen and R³ is a 5- or 6-membered heteroaryl ring or a heterocyclic ring comprising from 1 to 3 heteroatoms selected from O, N and S bonded to the phenyl ring through a ring carbon or nitrogen (if present) atom and which heterocyclic ring is unsubstituted or substituted on one or more ring carbon atoms by C_1-6alkyl or oxo (=0), or on a ring nitrogen atom by C_1-6alkyl.

6. A compound of formula (I) as claimed in any one of claims 1 to 4 in which the 5- or 6- membered saturated or unsaturated ring formed by R² and R³, fused with the phenyl ring, include:

7. A compound of formula (I) as claimed in any one of claims 1 to 6 in which R⁴ is ethyl or propyl.

8. A compound of formula (I) as claimed in any one of claims 1 to 7 in which X is fluoro.

9. A compound of formula (I) as claimed in claim 1 selected from any one of Examples 1 to 65.

10. A compound of formula (I) as claimed in claim 1 selected from: ^-(i .i-dioxido^.S-dihydro-i-benzothien-θ-yO-Λ^-ethyl-δ-fluoro-Λ^-IH-indazol^-yl- 2,4-pyrimidinediamine; 3-({4-[ethyl(1/-/-indazol-4-yl)amino]-5-fluoro-2-pyrimidinyl}amino)benzene sulfonamide;

3-[6-({4-[ethyl(1 H-indazol-4-yl)amino]-5-fluoro-2-pyrimidinyl}amino)-1 H-indazol-1 -yl]-

1-propanol;

/V²-1 H-1 ^.S-Benzotriazol-δ-yl-ΛΛethyl-S-fluoro-ΛΛi H-indazol-4-yl-2,4- pyrimidinediamine; or salt or solvate thereof, in particular, a pharmaceutically acceptable salt or solvate thereof.

11. A process for preparing a compound of formula (I), or a salt or solvate thereof, as defined in any one of claims 1 to 10, which process comprises reacting a compound of formula (II):

or a protected derivative thereof wherein L¹ represents a suitable leaving group, with a compound of formula (III)

or a protected derivative thereof, wherein X, R¹, R², R³ and R⁴ are as defined for formula (I) in claim 1.

12. A pharmaceutical formulation comprising a compound of formula (I), or a salt or solvate thereof, as defined in any one of claims 1 to 10 and pharmaceutically acceptable excipients.

13. A compound of formula (I) or a salt or solvate thereof as defined in any one of claims 1 to 10 for use in therapy.

14. The use of a compound of formula (I) or a salt or solvate thereof as defined in any one of claims 1 to 10 in the manufacture of a medicament for treating a disease associated with inappropriate mast cell activation

15. The use of a compound of formula (I) or a salt or solvate thereof as defined in any one of claims 1 to 10 in the manufacture of a medicament to inhibit a Syk kinase.

16. The use of a compound of formula (I) or a salt or solvate thereof as defined in any one of claims 1 to 10 in the manufacture of a medicament for of treating an inflammatory disease

17. The use of a compound of formula (I) or a salt or solvate thereof as defined in any one of claims 1 to 10 in the manufacture of a medicament for treating an allergic disorder.

18. The use of a compound of formula (I) or a salt or solvate thereof as defined in any one of claims 1 to 10 in the manufacture of a medicament for treating rhinitis.