WO2009001021A1 - Imidazopyridine derivatives useful as enzyme inhibitors for the treatment of cell proliferative and autoimmune diseases - Google Patents

Imidazopyridine derivatives useful as enzyme inhibitors for the treatment of cell proliferative and autoimmune diseases Download PDF

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Publication number
WO2009001021A1
WO2009001021A1 PCT/GB2007/002393 GB2007002393W WO2009001021A1 WO 2009001021 A1 WO2009001021 A1 WO 2009001021A1 GB 2007002393 W GB2007002393 W GB 2007002393W WO 2009001021 A1 WO2009001021 A1 WO 2009001021A1
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piperazin
imidazo
pyridin
bromo
phenyl
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PCT/GB2007/002393
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French (fr)
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Vassilios Bavetsias
Edward Mcdonald
Spyridon Linardopoulos
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Chroma Therapeutics Ltd.
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Publication of WO2009001021A1 publication Critical patent/WO2009001021A1/en

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D471/00—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00
    • C07D471/02—Heterocyclic compounds containing nitrogen atoms as the only ring hetero atoms in the condensed system, at least one ring being a six-membered ring with one nitrogen atom, not provided for by groups C07D451/00 - C07D463/00 in which the condensed system contains two hetero rings
    • C07D471/04—Ortho-condensed systems
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents

Definitions

  • This invention relates to certain specific imidazopyridine compounds which inhibit members of the aurora kinase family of enzymes and to their use in the treatment of cell proliferative diseases, including cancer, and inflammation.
  • DNA is packaged with histones, to form chromatin.
  • the aurora kinases are a family of serine/threonine kinases which have been identified as key regulators of the mitotic cell division process (Bischoff and Plowman, 1999 Trends Cell Biol 9, 454-459) which may become deregulated in cancer and other hyperproliferative diseases (Warner et al, 2003, MoI Can Ther 2, 589-595).
  • the three members of this family identified so far are referred to as Aurora- A, Aurora-B and Aurora-C.
  • Higher eukaryotic cells typically express two or more Aurora kinases. It has been shown that inhibition of Aurora B affects several facets of mitosis including histone H3 phosphorylation, chromosome segregation and cytokinesis.
  • aurora kinase inhibitors Several structural classes of aurora kinase inhibitors are known, see for example WO
  • X is -N-, -CH 2 -N-, -CH 2 -CH-, or -CH-;
  • R 1 is a radical of formula (IA)
  • Z is -CH 2 -, -NH-, -O-, -S(O)- -S-, -S(O) 2 or a divalent monocyclic carbocyclic or heterocyclic radical having 3-7 ring atoms;
  • AIk is an optionally substituted divalent C 1 -C 6 alkylene radical
  • A is hydrogen or an optionally substituted monocyclic carbocyclic or heterocyclic ring having 5-7 ring atoms; r, s and t are independently 0 or 1 , provided that when A is hydrogen then at least one of r and s is 1 ;
  • R 2 is halogen, -CN, -CF 3 , -OCH 3 , or cyclopropyl
  • R 3 is a radical of formula (IB)
  • Q is hydrogen or an optionally substituted phenyl or monocyclic heterocyclic ring with 5 or 6 ring atoms;
  • AIk 1 and AIk 2 are, independently, optionally substituted divalent C r C 3 alkylene radicals
  • the present invention includes all tautomers of the compounds of the Examples herein, and their salts, hydrates, solvates, and N-oxides, as well as mixtures thereof in any proportions.
  • 'solvate' is used herein to describe a molecular complex comprising the compound of the invention and a stoichiometric amount of one or more pharmaceutically acceptable solvent molecules, for example, ethanol.
  • solvent molecules for example, ethanol.
  • 'hydrate' is employed when said solvent is water.
  • the compounds with which the invention is concerned are inhibitors of the Aurora kinase family, namely Aurora kinases A and/or B and/or C, and are therefore of use in the treatment of cell proliferative disease, such as cancer, and in treatment of aitoimmune diseases and inflammation, in humans and other mammals.
  • the compounds of the invention may be used in the preparation of a composition for the treatment of cell-proliferation disease, for example cancer cell proliferation and inflammatory and autoimmune diseases, for example rheumatoid arthritis.
  • cell-proliferation disease for example cancer cell proliferation and inflammatory and autoimmune diseases, for example rheumatoid arthritis.
  • the invention provides a method for the treatment of the foregoing disease types, which comprises administering to a subject suffering such disease an effective amount of a compound of the invention.
  • the specific dose level for any particular patient will depend upon a variety of factors including the activity of the specific compound employed, the age, body weight, general health, sex, diet, time of administration, route of administration, rate of excretion, drug combination and the severity of the particular disease undergoing treatment. Optimum dose levels and frequency of dosing will be determined by clinical trial as is required in the art.
  • Tablets and capsules for oral administration may be in unit dose presentation form, and may contain conventional excipients such as binding agents, for example syrup, acacia, gelatin, sorbitol, tragacanth, or polyvinyl-pyrrolidone; fillers for example lactose, sugar, maize-starch, calcium phosphate, sorbitol or glycine; tabletting lubricant, for example magnesium stearate, talc, polyethylene glycol or silica; disintegrants for example potato starch, or acceptable wetting agents such as sodium lauryl sulphate.
  • the tablets may be coated according to methods well known in normal pharmaceutical practice.
  • Oral liquid preparations may be in the form of, for example, aqueous or oily suspensions, solutions, emulsions, syrups or elixirs, or may be presented as a dry product for reconstitution with water or other suitable vehicle before use.
  • Such liquid preparations may contain conventional additives such as suspending agents, for example sorbitol, syrup, methyl cellulose, glucose syrup, gelatin hydrogenated edible fats; emulsifying agents, for example lecithin, sorbitan monooleate, or acacia; nonaqueous vehicles (which may include edible oils), for example almond oil, fractionated coconut oil, oily esters such as glycerine, propylene glycol, or ethyl alcohol; preservatives, for example methyl or propyl p-hydroxybenzoate or sorbic acid, and if desired conventional flavouring or colouring agents.
  • suspending agents for example sorbitol, syrup, methyl cellulose, glucose syrup, gelatin hydrogenated edible fats
  • emulsifying agents for example lecithin, sorbitan monooleate, or acacia
  • nonaqueous vehicles which may include edible oils
  • almond oil fractionated coconut oil
  • oily esters such as glycerine, propylene glycol
  • the drug may be made up into a cream, lotion or ointment.
  • Cream or ointment formulations which may be used for the drug are conventional formulations well known in the art, for example as described in standard textbooks of pharmaceutics such as the British Pharmacopoeia.
  • the active ingredient may also be administered parenterally in a sterile medium.
  • the drug can either be suspended or dissolved in the vehicle.
  • adjuvants such as a local anaesthetic, preservative and buffering agents can be dissolved in the vehicle.
  • the compounds of the invention are those of the Examples below, and may be prepared by the methods described therein:
  • Flash column chromatography was performed using Merck silica gel 60 (0.025 - 0.04 mm). Column chromatography was also performed on a FlashMaster personal unit using isolute Flash silica columns or a Biotage SP1 purification system using Biotage Flash silica cartridges. Ion exchange chromatography was performed using acidic lsolute Flash SCX-II cartridges. 1 H NMR spectra were recorded on a Bruker Avance dpx250 or a Bruker Avance-500. Samples were prepared as solutions in a deuterated solvent and referenced to the appropriate internal non-deuterated solvent peak or tetramethylsilane.
  • Method A (10 mins) - nominal mass, LC injection with a 10 minute gradient (MeOH and 0.1% formic acid), positive ionisation and an injection volume of 3 ⁇ L.
  • Method B (6 mins) - nominal mass, LC injection with a 6 minute gradient (MeOH and 0.1% formic acid), positive ionisation and an injection volume of 2 ⁇ L.
  • High resolution mass spectra were also obtained using the Agilent 6210 time-of- Flight LCMS (with 1200 series LC) instrumental set-up and the following conditions: LC injection with a 10 minute gradient (MeOH and 0.1 % formic acid), +ve ionisation and an injection volume of 1 ⁇ L.
  • 3-Fluoro-5-formyl pyridine (0.50 g, 4.0 mmol, 1 eq) was dissolved in a mixture of EtOH (9.9 mL) and AcOH (1.1 mL).
  • N-Boc piperazine (1.86 g, 10.0 mmol, 2.5 eq) was added in one portion and the mixture was stirred and cooled in a water-ice bath for 10 minutes before the portionwise addition of NaBH 3 CN (0.24 g, 3.8 mmol, 0.95 eq). The mixture was allowed to warm up to rt and stirred for 17h.
  • reaction mixture was heated at 85 ° C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH 3 until complete dissolution was observed.
  • This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v; 94:6).
  • reaction mixture was heated at 85 ° C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH 3 until complete dissolution was observed.
  • This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v; 94:6).
  • reaction mixture was heated at 85 ° C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH 3 until complete dissolution was observed.
  • This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v; 94:6).
  • reaction mixture was heated at 85 ° C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH 3 until complete dissolution was observed.
  • This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v; 94:6).
  • reaction mixture was heated at 85 ° C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH 3 until complete dissolution was observed.
  • This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v; 94:6).
  • reaction mixture was heated at 85 ° C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH 3 until complete dissolution was observed.
  • This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v; 95:5).
  • reaction mixture was heated at 85 ° C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH 3 until complete dissolution was observed.
  • This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v; 95:5).
  • reaction mixture was heated at 85 ° C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH 3 until complete dissolution was observed.
  • This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v; 94:6).
  • reaction mixture was stirred at 80 0 C for 20 h, then allowed to cool to room temperature and concentrated in vacuo.
  • the residue was absorbed on silica gel, the free-running powder was placed on a 10 g isolute silica column, and elution with a gradient of methanol (0 to 6%) in ethyl acetate / dichloromethane (v:v; 1 :1 ) afforded the title compound as a pale yellow solid (0.031 g, 48%).
  • kinase buffer 50 mM Tris pH 7.5, 10 mM NaCI, 2.5 mM MgCL 2 , 1 mM DTT, 20 ⁇ M ATP, 0.025 ⁇ Ci/ ⁇ l 33 P-ATP, and 100 ⁇ g/ml MBP was added to a 96- well plate followed by addition of 250 ng of Aurora-A enzyme per well. The plate was shaken for approximately 2 min on a flat-bed plate shaker (Wellmix, Denley, UK) and incubated for 2 hours at room temperature. The reaction was stopped by the addition of 30 ⁇ l of 2% orthophosphoric acid.
  • the reaction mixture was then transferred and filtered on the 96-well Multi-Screening Filter Plate (MATAH0P00, MILLIPORE), pre- treated with 50 ⁇ l per well of 0.5% orthophosphoric acid.
  • the plate was washed twice with 200 ⁇ l of 0.5% orthophosphoric acid and vacuum-dried. 25 ⁇ l of scintillant (MicroscintTM20, PerkinElmer) per well was added and the plate was shaken for 10 min. Finally, the plate was re-sealed with TopSealA and the signal from the filter- bound component was read on TopCount-NXTTM (PerkinElmer Life Sciences UK Ltd., Hounslow, UK).
  • TopCount-NXTTM PerkinElmer Life Sciences UK Ltd., Hounslow, UK.
  • Cell viability assay Determination of IC 50 of Aurora inhibitors in HCT116 cell line using MTT assay.
  • the effects of compounds of the Examples on cellular proliferation were determined using the MTT assay according to manufacturer's instructions (Sigma). Briefly, the human colon tumour cells HCT116, were seeded in triplicate into 96-well plates at 2500 cells/well 24 hours before treatment with a range of concentrations of Aurora inhibitors (0 - 50 ⁇ M). After 72 hrs, 15 ⁇ l/well of 0.5% 3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT) was added to the cells for 4 hrs at 37 0 C, the dye- stained viable cells were extracted by adding 150 ⁇ l/well of dimethyl sulphoxide (DMSO). The optical density was measured at 570 nm using the Wallac VICTOR 2 T M 1420 Multilabel Counter (PerkinElmer). The IC 50 was calculated using the Prism software.
  • DMSO dimethyl sulphoxide
  • Range A IC 50 ⁇ 200 nM for Aurora A inhibition, and IC 50 ⁇ 2000 nM for HCT116 cell growth inhibition.
  • Range B IC 50 >200 nM for Aurora A inhibition, and IC 50 >2000 nM for HCT116 cell growth inhibition.
  • the compounds of examples 7, 24, 25, 26, 28, 33, 34 had IC 50 S ⁇ 60 nM in the enzyme assay, and ⁇ 600 nM in the cellular assay.

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Abstract

The 62 specific compounds disclosed in the description, which fall within the general structural formula (I) below, are inhibitors of aurora kinase enzymes: wherein X is -N-, -CH2-N-, -CH2-CH-, or -CH-; R1 is a radical of formula (IA) wherein Z is -CH2-, -NH-, -O-, -S(O)- -S-, -S(O)2 or a divalent monocyclic carbocyclic or heterocyclic radical having 3-7 ring atoms; AIk is an optionally substituted divalent C1-C6 alkylene radical; A is hydrogen or an optionally substituted monocyclic carbocyclic or heterocyclic ring having 5-7 ring atoms; r, s and t are independently 0 or 1, provided that when A is hydrogen then at least one of r and s is 1; R2 is halogen, -CN, -CF3, -OCH3, or cyclopropyl; and R3 is a radical of formula (IB) wherein Q is hydrogen or an optionally substituted phenyl or monocyclic heterocyclic ring with 5 or 6 ring atoms; Z1 is -S-, -S(O)-, -S(O)2-, -O-, -SO2NH-, -NHSO2-, NHC(=O)NH, -NH(C=S)NH-, Or -N(R4)- wherein R4 is hydrogen, C1-C3 alkyl, cycloalkyl, or benzyl; and AIk1 and AIk2 are, independently, optionally substituted divalent C1-C3 alkylene radicals; and m, n and p are independently O or 1.

Description

IMIDAZOPYRIDINE DERIVATIVES USEFUL AS ENZYME INHIBITORS FOR THE TREATMENT OF CELL PROLIFERATIVE AND AUTOIMMUNE DISEASES
This invention relates to certain specific imidazopyridine compounds which inhibit members of the aurora kinase family of enzymes and to their use in the treatment of cell proliferative diseases, including cancer, and inflammation.
Background to the Invention
In eukaryotic cells DNA is packaged with histones, to form chromatin. Approximately
150 base pairs of DNA are wrapped twice around an octamer of histones (two each of histones 2A, 2B, 3 and 4) to form a nucleosome, the basic unit of chromatin. The ordered structure of chromatin needs to be modified in order to allow transcription of the associated genes. Transcriptional regulation is key to differentiation, proliferation and apoptosis, and is, therefore, tightly controlled. Control of the changes in chromatin structure (and hence of transcription) is mediated by covalent modifications to histones, most notably of the N-terminal tails. Covalent modifications (for example methylation, acetylation, phosphorylation and ubiquitination) of the side chains of amino acids are enzymatically mediated (A review of the covalent modifications of histones and their role in transcriptional regulation can be found in Berger SL 2001 Oncogene 20, 3007-3013; See Grunstein, M 1997 Nature 389, 349-352; Wolffe AP 1996 Science 272, 371-372; and Wade PA et al 1997 Trends Biochem Sci 22, 128-132 for reviews of histone acetylation and transcription).
The aurora kinases are a family of serine/threonine kinases which have been identified as key regulators of the mitotic cell division process (Bischoff and Plowman, 1999 Trends Cell Biol 9, 454-459) which may become deregulated in cancer and other hyperproliferative diseases (Warner et al, 2003, MoI Can Ther 2, 589-595). The three members of this family identified so far are referred to as Aurora- A, Aurora-B and Aurora-C. Higher eukaryotic cells typically express two or more Aurora kinases. It has been shown that inhibition of Aurora B affects several facets of mitosis including histone H3 phosphorylation, chromosome segregation and cytokinesis. Aurora A and C localise to spindle poles with Aurora A being required for bipolar spindle formation in a number of systems (Giet and Prigent, 1999, J.Cell.Sci 11 , 3591-3601 ). Aurora A and B have been shown to be overexpressed in a number of human cancers and their overexpression in cells in vitro leads to transformation, centrosome abnormalities and aneuploidy (Bischoff et al, 1998, EMBO J. 17, 3052). Cells which overexpress Aurora A have been shown to form tumours in aythymic ce. The observations contained in these manuscripts suggest that increase in jrora kinase activity may serve to promote tumour development by providing rowth advantage or by inducing genetic instability and that Aurora Kinase inhibition hould have therapeutic benefit in cancer.
Aurora Kinase Inhibitors.
Several structural classes of aurora kinase inhibitors are known, see for example WO
02/00649, WO 2004/000833, WO 03/055491 , WO 2004/058752, WO 2004/058781 , WO 04105765, WO 05004872, WO 04113324, US 6143764 and US 2004/0049032, but the foregoing is by no means an exhaustive list.
Co-pending international patent application no. PCT/GB2006/004854 describes and claims compound of formula (I) and salts, hydrates, solvates, and N-oxides thereof, having aurora kinase inhibitory activity:
Figure imgf000003_0001
wherein
X is -N-, -CH2-N-, -CH2-CH-, or -CH-;
R1 is a radical of formula (IA)
-|-[Z]r-[Alk]s-A (IA)
wherein
Z is -CH2-, -NH-, -O-, -S(O)- -S-, -S(O)2 or a divalent monocyclic carbocyclic or heterocyclic radical having 3-7 ring atoms;
AIk is an optionally substituted divalent C1-C6 alkylene radical;
A is hydrogen or an optionally substituted monocyclic carbocyclic or heterocyclic ring having 5-7 ring atoms; r, s and t are independently 0 or 1 , provided that when A is hydrogen then at least one of r and s is 1 ;
R2 is halogen, -CN, -CF3, -OCH3, or cyclopropyl; and
R3 is a radical of formula (IB)
-|-[Alki]m-[Zi]p-[Alk2]n-Q (IB)
wherein
Q is hydrogen or an optionally substituted phenyl or monocyclic heterocyclic ring with 5 or 6 ring atoms;
Z1 is -S-, -S(O)-, -S(O)2-, -O-, -SO2NH-, -NHSO2-, NHC(=O)NH, -NH(C=S)NH-, or -N(R4)- wherein R4 is hydrogen, C1-C3 alkyl, cycloalkyl, or benzyl; and
AIk1 and AIk2 are, independently, optionally substituted divalent CrC3 alkylene radicals; and
m, n and p are independently O or 1.
Deacription of the Invention
The present invention relates to compounds having aurora kinase inhibitory activity, falling within the general class of compounds described and claimed in PCT/GB2006/004854, but not specifically disclosed therein. More specifically, a first aspect of the invention is a compound selected from the group consisting of those of the Examples herein, and their salts, hydrates, solvates, and N-oxides.
Since the compounds of the invention fall within the general formula (I) above of PCT/GB2006/004854, they, like the compounds of PCT/GB2006/004854, may exist in two tautomeric forms one of which is shown in formula (I), the other being shown in formula (II):
Figure imgf000005_0001
A third possible tautomeric form is shown in formula (III):
Figure imgf000005_0002
The present invention includes all tautomers of the compounds of the Examples herein, and their salts, hydrates, solvates, and N-oxides, as well as mixtures thereof in any proportions.
The term 'solvate' is used herein to describe a molecular complex comprising the compound of the invention and a stoichiometric amount of one or more pharmaceutically acceptable solvent molecules, for example, ethanol. The term 'hydrate' is employed when said solvent is water.
Utilities
As mentioned above, the compounds with which the invention is concerned are inhibitors of the Aurora kinase family, namely Aurora kinases A and/or B and/or C, and are therefore of use in the treatment of cell proliferative disease, such as cancer, and in treatment of aitoimmune diseases and inflammation, in humans and other mammals.
Therefore, in another aspect the invention provides the use of a compound of the present invention in the preparation of a composition for inhibiting the activity of an aurora kinase enzyme, f f/o-»rr α evxaommrpJle The compounds with which the invention is concerned may be used for the inhibition of aurora kinase activity, such as aurora-A, ex vivo or in vivo.
In one aspect of the invention, the compounds of the invention may be used in the preparation of a composition for the treatment of cell-proliferation disease, for example cancer cell proliferation and inflammatory and autoimmune diseases, for example rheumatoid arthritis.
In another aspect, the invention provides a method for the treatment of the foregoing disease types, which comprises administering to a subject suffering such disease an effective amount of a compound of the invention.
It will be understood that the specific dose level for any particular patient will depend upon a variety of factors including the activity of the specific compound employed, the age, body weight, general health, sex, diet, time of administration, route of administration, rate of excretion, drug combination and the severity of the particular disease undergoing treatment. Optimum dose levels and frequency of dosing will be determined by clinical trial as is required in the art.
The invention also includes pharmaceutical composition comprising a compound of the invention, together with a pharmaceutically acceptable carrier.
The compounds with which the invention is concerned may be prepared for administration by any route consistent with their pharmacokinetic properties. The orally administrable compositions may be in the form of tablets, capsules, powders, granules, lozenges, liquid or gel preparations, such as oral, topical, or sterile parenteral solutions or suspensions. Tablets and capsules for oral administration may be in unit dose presentation form, and may contain conventional excipients such as binding agents, for example syrup, acacia, gelatin, sorbitol, tragacanth, or polyvinyl-pyrrolidone; fillers for example lactose, sugar, maize-starch, calcium phosphate, sorbitol or glycine; tabletting lubricant, for example magnesium stearate, talc, polyethylene glycol or silica; disintegrants for example potato starch, or acceptable wetting agents such as sodium lauryl sulphate. The tablets may be coated according to methods well known in normal pharmaceutical practice. Oral liquid preparations may be in the form of, for example, aqueous or oily suspensions, solutions, emulsions, syrups or elixirs, or may be presented as a dry product for reconstitution with water or other suitable vehicle before use. Such liquid preparations may contain conventional additives such as suspending agents, for example sorbitol, syrup, methyl cellulose, glucose syrup, gelatin hydrogenated edible fats; emulsifying agents, for example lecithin, sorbitan monooleate, or acacia; nonaqueous vehicles (which may include edible oils), for example almond oil, fractionated coconut oil, oily esters such as glycerine, propylene glycol, or ethyl alcohol; preservatives, for example methyl or propyl p-hydroxybenzoate or sorbic acid, and if desired conventional flavouring or colouring agents.
For topical application to the skin, the drug may be made up into a cream, lotion or ointment. Cream or ointment formulations which may be used for the drug are conventional formulations well known in the art, for example as described in standard textbooks of pharmaceutics such as the British Pharmacopoeia.
For topical application to the eye, the drug may be made up into a solution or suspension in a suitable sterile aqueous or non aqueous vehicle. Additives, for instance buffers such as sodium metabisulphite or disodium edeate; preservatives including bactericidal and fungicidal agents such as phenyl mercuric acetate or nitrate, benzalkonium chloride or chlorhexidine, and thickening agents such as hypromellose may also be included.
The active ingredient may also be administered parenterally in a sterile medium. Depending on the vehicle and concentration used, the drug can either be suspended or dissolved in the vehicle. Advantageously, adjuvants such as a local anaesthetic, preservative and buffering agents can be dissolved in the vehicle.
As stated, the compounds of the invention are those of the Examples below, and may be prepared by the methods described therein:
EXAMPLES General experimental comments
Commercially available starting materials, reagents and dry solvents were used as supplied. Flash column chromatography was performed using Merck silica gel 60 (0.025 - 0.04 mm). Column chromatography was also performed on a FlashMaster personal unit using isolute Flash silica columns or a Biotage SP1 purification system using Biotage Flash silica cartridges. Ion exchange chromatography was performed using acidic lsolute Flash SCX-II cartridges. 1H NMR spectra were recorded on a Bruker Avance dpx250 or a Bruker Avance-500. Samples were prepared as solutions in a deuterated solvent and referenced to the appropriate internal non-deuterated solvent peak or tetramethylsilane. Chemical shifts were recorded in ppm (δ) downfield of tetramethylsilane. LC-MS spectra were recorded on a Waters LCT with a Waters Alliance 2795 sepearations module, using a Phenomenex Gemini C18 column and either of the following conditions:
Method A (10 mins) - nominal mass, LC injection with a 10 minute gradient (MeOH and 0.1% formic acid), positive ionisation and an injection volume of 3 μL. Column: Phenomenex Gemini Ci8 column (5 micron, 50 x 4.6 mm) Method B (6 mins) - nominal mass, LC injection with a 6 minute gradient (MeOH and 0.1% formic acid), positive ionisation and an injection volume of 2 μL. Column: Phenomenex Gemini Ci8 column (3 micron, 30 x 4.6 mm). If method not stated, then Method A was followed.
High resolution mass spectra were obtained using the above instrumental set-up and the following conditions:
Accurate mass, LC injection with a 10 minute gradient (MeOH and 0.1% formic acid), +ve ionisation and an injection volume of 4 μL. Column: Phenomenex Gemini Ci8 column (5 micron, 50 x 4.6 mm)
High resolution mass spectra were also obtained using the Agilent 6210 time-of- Flight LCMS (with 1200 series LC) instrumental set-up and the following conditions: LC injection with a 10 minute gradient (MeOH and 0.1 % formic acid), +ve ionisation and an injection volume of 1 μL. Column: Phenomenex Gemini C18 column (3 micron, 30 x 4.6 mm)
Synthesis of key intermediates:
1. 4,5-dichloro-3-nitropyridin-2-amine
Figure imgf000008_0001
It was synthesised by the method described in example 1 of PCT/GB2006/004854.
2. 5-bromo-4-chloro-3-nitropyridin-2-amine
Figure imgf000008_0002
It was synthesised by the method described in example 2 of PCT/GB2006/004854.
Example 1 te/t-Butyl-4-(4-(6-bromo-7-(4-(cyclobutylmethyl)piperazin-1-yl)-3H-imidazo[4,5- b]pyridin-2-yl)phenyl)piperazine-1-carboxylate
Figure imgf000009_0001
To a mixture of 5-bromo-4-(4-(cyclobutylmethyl)piperazin-1-yl)-3-nitropyridin-2-amine (prepared as described in example 149 of PCT/GB2006/004854; 0.12 g, 0.32 mmol, 1eq) in EtOH (5.6 ml_) and DMF (0.74 ml_), tert-butyl 4-(4-formylphenyl)piperazine-1- carboxylate (0.102 g, 0.35 mmol, 1.1 eq) was added followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.96 mL, 0.96 mmol). The reaction mixture was heated at 85 0C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH3 until complete dissolution was observed. This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v 94:6) to give the title compound as an off-white solid (0.043 g, 22%); 1H-NMR (500Mz, DMSO-c/6): δ 1.43 (s, 9H, C(CH3)3), 1.60-1.75 (m, 2H), 1.76- 1.94 (m, 2H), 2.00-2.11 (m, 2H), 2.35-2.48 (brs, 2H), 2.50-2.65 (m, 4H), 3.43-3.52 (m, 4H), 3.55-3.69 (brs, 4H), 7.07 (d, J = 9.0 Hz, 2H, ArH, C6H4), 8.04 (d, J = 8.5 Hz, 2H, ArH, C6H4), 8.17 (s, 1H, imidazo[4,5-b]pyridine 5-H)1 13.22 (br s, 1 H1 imidazo[4,5-b]pyridine N-H), two signals (one from the cyclobutyl ring and that from one of the two piperazinyl ring were hidden under solvents' signals; LC (Method B) - MS (ESI, m/z) 3.86 min - 610/612 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 610.2504, calculated for C30H4iN7O2Br (M+H)+: 610.2499.
6-Bromo-7-(4-(cyclobutylmethyl)piperazin-1-yl)-2-(4-(piperazin-1-yl)phenyl)-3H- imidazo[4,5-/b]pyridine
Figure imgf000010_0001
te/t-Butyl-4-(4-(6-bromo-7-(4-(cyclobutylmethyl)piperazin-1-yl)-3H-imidazo[4,5- t)]pyridin-2-yl)phenyl)piperazine-1-carboxylate (0.040 g, 0.066 mmol) was suspended in DCM (4.0 ml_) and the mixture cooled in an ice bath. TFA (1.0 ml_) was added and the resulting solution was allowed to warm up to room temperature and stirred for 2 h. The mixture was passed through an SCX column (2 g), the filtrate collected and the solvent removed in vacuo to give the title compound as a solid (0.032 g, 96%); 1H-NMR (500Mz, DMSO-c/6): δ 1.60-1.74 (m, 2H), 1.77-1.94 (m, 2H), 2.00-2.10 (m, 2H), 2.42 (d, J = 7.0 Hz, 2H, NCH2), 2.52-2.60 (m, 5H), 2.85 (t, J = 4.8 Hz, 4H), 3.16- 3.21 (m, 4H), 3.55-3.65 (m, 4H), 7.03 (d, J = 8.5 Hz, 2H, ArH, C6H4), 8.02 (d, J = 9.0 Hz, 2H, ArH, C6H4), 8.16 (s, 1 H1 imidazo[4,5-Jb]pyridine 5-H); LC (Method B) - MS (ESI, m/z) 2.16 min - 510/512 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 510.1974, calculated for C25H33N7Br (M+H)+: 510.1975.
Example 2
5-Chloro-4-(4-(cyclobutylmethyl)piperazin-1-yl)-3-nitropyridin-2-amine
Figure imgf000010_0002
te/f-Butyl 4-(cyclobutylmethyl)piperazine-1-carboxylate prepared as described in Example 149 of PCT/GB2006/004854 (0.095 g, 0.37 mmol, 1.1 eq) was dissolved in DCM (1.1 mL) and the mixture cooled in a ice-water bath before the dropwise addition of TFA (0.55 mL). Stirring was continued at this temperature for 1 h, and the solvents were removed in vacuo. The resulting crude material was azeotroped with toluene and dried.
The resulting 1-(cyclobutylmethyl)piperazine (supposedly 0.057 g, 0.37 mmol, 1.1 eq) was suspended in 'PrOH (0.37 mL) and DIPEA (0.19 mL). To this solution, 4,5- dichloro-3-nitropyridin-2-amine (0.07 g, 0.34 mmol, 1 eq) was added and the reaction mixture was heated and stirred for 17 h at 650C. The mixture was filtered, washed with 1PrOH (3 x 3 mL), Et2O (2 x 3 mL), and dried to give the title compound as a bright yellow powder (0.045 g, 37%); 1H-NMR (500 MHz, DMSO-Cf6): δ 1.58-1.71 (m, 2H), 1.74-1.92 (m, 2H), 1.95-2.07 (m, 2H), 2.37 (d, J = 7.0 Hz, 2H, NCH2), 2.41-2.54 (m, 5H), 2.97-3.10 (m, 4H), 6.93 (s, 2H, NH2), 8.05 (s, 1 H, pyridine 6-H); LC (Method B) - MS (ESI, m/z): Rt = 1.90 min - 326/328 [(M+H+), Br isotopic pattern, 100%].
terf-Butyl 4-(4-(6-chloro-7-(4-(cyclobutylmethyl)piperazin-1-yl)-3H-imidazo[4,5- jb]pyridin-2-yl)benzyl)piperazine-1-carboxylate
Figure imgf000011_0001
4-chloro-4-(cyclobutylmethyl)piperazin-1-yl)-3-nitropyridin-2-amine (0.045 g, 0.14 mmol, 1eq) in EtOH (2.4 mL) and DMF (0.33 mL), te/t-butyl 4-(4- formylphenyl)piperazine-1-carboxylate (0.046 g, 0.15 mmol, 1.1 eq) was added followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.41 mL, 0.41 mmol). The reaction mixture was heated at 85°C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH3 until complete dissolution was observed. This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v 94:6) to give the title compound as a off-white solid (0.010 g, 12%); 1H-NMR (500Mz, CDCI3): δ 1.48 (s, 9H, C(CH3)3), 1.71-2.02 (m, 4H), 2.10-2.20 (m, 2H), 2.42-2.50 (m, 4H), 2.57 (d, 2H, J = 6.5 Hz, NCH2), 2.62-2.80 (m, 5H), 3.43-3.52 (m, 4H), 3.63 (s, 2H, NCH2), 3.87-3.99 (m, 4H, piperazine N(CH2)2), 7.53 (d, J = 7.5 Hz, 2H, ArH, C6H4), 8.11 (d, J = 8.0 Hz, 2H, ArH, C6H4), 8.16 (s, 1H, imidazo[4,5-b]pyridine 5-H), 13.78 (br s, 1 H, imidazo[4,5- -b]pyridine N-H); LC (Method B) - MS (ESI, m/z) 2.57 min - 580/582 [(M+H)\ Cl isotopic pattern]. ESI-HRMS: Found: 580.3163, calculated for C31H43CIN7O2 (M+H)+: 580.3161.
6-Chloro-7-(4-(cyclobutylmethyl)piperazin-1-yl)-2-(4-(piperazin-1-ylmethyl)phenyl)- 3H-imidazo[4,5-ib]pyridine
Figure imgf000012_0001
tert-Butyl-4-(4-(6-chloro-7-(4-(cyclobutylmethyl)piperazin-1-yl)-3/-/-imidazo[4,5- /?]pyridin-2-yl)benzyl)piperazine-1-carboxylate (0.008 g, 0.013 mmol) was suspended in DCM (0.6 mL) and the mixture cooled in an ice bath. TFA (0.15 mL) was added and the resulting solution was allowed to warm up to room temperature and stirred for 2 h. The mixture was passed through an SCX column (2 g), the filtrate collected and the solvent removed in vacuo to give the title compound as a solid (0.004 g, 64%); 1H-NMR (500Mz, DMSO-c/6): δ 1.77-1.92 (m, 5H), 2.00-2.10 (m, 2H), 2.28-2.37 (m, 4H), 2.41 (d, 2H, J = 7.0 Hz, NCH2), 2.52-2.60 (m, 4H), 2.73 (t, 4H, J = 4.8Hz, piperazine N(CH2)2), 3.50 (s, 2H), 3.63-3.71 (m, 4H, piperazine N(CH2J2), 7.45 (d, J = 8.0 Hz, 2H, ArH, C6H4), 8.10 (s, 1H, imidazo[4,5-b]pyridine 5-H)1 8.12 (d, J = 8.5 Hz, 2H, ArH, C6H4); LC (Method B) - MS (ESI, m/z) 2.05 min - 480/482 [(M+H)+, Cl isotopic pattern]. ESI-HRMS: Found: 480.2632, calculated for C26H35CIN7 (M+H)+: 480.2637.
Example 3 te/t-Butyl
Figure imgf000012_0002
3-Fluoro-5-formyl pyridine (0.50 g, 4.0 mmol, 1 eq) was dissolved in a mixture of EtOH (9.9 mL) and AcOH (1.1 mL). N-Boc piperazine (1.86 g, 10.0 mmol, 2.5 eq) was added in one portion and the mixture was stirred and cooled in a water-ice bath for 10 minutes before the portionwise addition of NaBH3CN (0.24 g, 3.8 mmol, 0.95 eq). The mixture was allowed to warm up to rt and stirred for 17h. The solvents were removed in vacuo and the crude material was purified purified by column chromatography on a Biotage SP1 system (DCM/EtOAc; v/v 1 :1 ) to give the title compound (1.12 g, 95%); 1H-NMR (500 MHz, CDCI3): δ 1.46 (s, 9H, C(CH3J3), 2.40 (t, 4H, J = 4.8 Hz, piperazine N(CH2)2), 3.44 (t, 4H, J = 5.0 Hz, piperazine N(CH2)2). 3.54 (s, 2H, NCH2), 7.42-7.47 (m, 1 H, py 4-H), 8.34-8.37 (m, 1H, py 2-H), 8.38 (d, 1 H1 J = 2.5 Hz, py 6-H); 13C-NMR (125 MHz, CDCI3): δ 28.4, 52.9, 59.4, 79.7, 123.0 (d, 2J0F = 18 Hz), 135.6 (d, 4JCF = 3 Hz), 137.1 (d, 2JCF = 23 Hz), 145.9 (d, 2JCF = 3 Hz), 159.7 (d, 1JcF = 255 Hz); LC(Method B) - MS (ESI, m/z): Rt = 2.30 min - 196 [(M- Boc+H)+, 100%].
5-Bromo-4-(4-((5-fluoropyridin-3-yl)nnethyl)piperazin-1-yl)-3-nitropyridin-2-amine
Figure imgf000013_0001
tert-Butyl 4-((5-fluoropyridin-3-yl)methyl)piperazine-1-carboxylate (0.30 g, 1.07 mmol, 1.1 eq) was dissolved in DCM (3.2 mL) and the mixture cooled in a ice-water bath before the dropwise addition of TFA (1 mL). Stirring was continued at this temperature for 1 h and the solvents were removed in vacuo. The resulting crude material was azeotroped with toluene and dried. The resulting 1-((5-fluoropyridin-3- yl)methyl)piperazine (supposedly 0.21 g, 0.42 mmol, 1 eq) was suspended in 1PrOH (3.2 mL) and DIPEA (0.8 mL). To this solution, the 5-bromo-4-chloro-3-nitropyridin-2- amine (0.25 g, 0.97 mmol, 1.0 eq) was added, and the mixture heated and stirred for 17 h at 65°C. The mixture was filtered, washed with 1PrOH (3 x 3 mL), cold H2O (3 x 3 mL), Et2O (2 x 3 mL) and dried to give the title compound as a bright yellow solid (0.29 g, 66%); 1H-NMR (500 MHz, DMSO-c/6): δ 2.50-2.59 (br s, 4H, piperazine N(CH2)2), 3.02-3.12 (m, 4H, piperazine N(CH2)2), 3.62 (s, 2H, NCH2), 6.97 (s, 2H, NH2), 7.64-7.70 (m, 1 H, Fpy 4-H), 8.16 (s, 1H, pyridine 6-H), 8.39-8.43 (m, 1 H, Fpy 2-H), 8.47 (d, 1 H, J = 2.5 Hz, Fpy 6-H); LC (Method B) - MS (ESI, m/z): Rt = 2.39 min - 411/413 [(M+H)+, Br isotopic pattern, 100%].
te/t-Butyl-4-(4-(6-bromo-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-b]pyridin-2-yl)benzyl)piperazine-1-carboxylate
Figure imgf000014_0001
To a mixture of 5-bromo-4-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.080 g, 0.19 mmol) in EtOH (3.2 mL) and DMF (0.45 mL), tert- butyl 4-(4-formylbenzyl)piperazine-1-carboxylate (0.064 g, 0.21 mmol, 1.1 eq) was added followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.58 mL, 0.58 mmol). The reaction mixture was heated at 85 0C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH3 until complete dissolution was observed. This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v 90:10) to give the title compound as an off-white solid (0.015 g, 12%); 1H-NMR (500Mz, DMSO-c/6): δ 1.39 (s, 9H, C(CHs)3), 2.34 (t, 4H, J = 4.5 Hz, piperazine N(CH2)2), 2.60-2.67 (m, 4H, piperazine N(CH2)2), 3.29-3.36 (m, 4H, piperazine N(CH2)2), 3.55 (s, 2H, NCH2), 3.63-3.73 (m, 6H), 7.46 (d, J = 8.0 Hz, 2H, ArH, C6H4), 7.68-7.74 (m, 1 H, py 4-H), 8.14 (d, J = 8.0 Hz, 2H, ArH, C6H4), 8.23 (s, 1H, imidazo[4,5-ib]pyridine 5-H), 8.44-8.47 (m, py 2-H ), 8.49 (d, 1H, J = 3.0 Hz, py 6-H), 13.48 (br s, 1 H, imidazo[4,5-/3]pyridine N-H); LC (Method B) - MS (ESI, m/z) 2.92 min - 665/667 [(M+H)+, Br isotopic pattern], ESI- HRMS: Found: 665.2356, calculated for C32H39FBrN8O2 (M+H)+: 665.2358.
6-Bromo-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-2-(4-(piperazin-1- ylmethyl)phenyl)-3H-imidazo[4,5-ιfc>]pyridine
Figure imgf000014_0002
te/Y-Butyl-4-(4-(6-bromo-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3/-/- imidazo[4,5-ιb]pyridin-2-yl)benzyl)piperazine-1-carboxylate (0.013 g, 0.019 mmol) was suspended in DCM (1.0 mL) and the mixture cooled in an ice bath. TFA (0.25 mL) was added and the resulting solution was allowed to warm up to room temperature and stirred for 2 h. The mixture was passed through an SCX column (2 g), the filtrate collected and the solvent removed in vacuo to give the title compound as a solid (0.005 g, 45%); 1H-NMR (500Mz, DMSO-Cf6): δ 2.26-2.36 (m, 4H1 piperazine N(CH2)2), 2.60-2.67 (m, 4H, piperazine N(CH2)2), 2.71 (t, 4H, J = 4.5 Hz1 piperazine N(CH2)2), 3.49 (s, 2H1 NCH2), 3.63-3.72 (m, 6H)1 7.45 (d, J = 8.0 Hz1 2H, ArH, C6H4), 7.68-7.75 (m, 1 H1 py 4-H), 8.13 (d, J = 8.0 Hz, 2H, ArH1 C6H4), 8.23 (s, 1 H, imidazo[4,5-b]pyridine 5-H), 8.44-8.47 (m, py 2-H ), 8.49 (d, 1 H, J = 2.5 Hz, py 6- H); LC (Method B) - MS (ESI, m/z) 2.19 min - 565/567 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 565.1828, calculated for C27H3iFBrN8 (M+H)+: 565.1834.
Example 4
2-(4-(6-Bromo-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3/-/-imidazo[4,5- /b]pyridin-2-yl)phenoxy)ethanol
Figure imgf000015_0001
To a mixture of 5-bromo-4-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.060 g, 0.14 mmol) in EtOH (2.4 mL) and DMF (0.33 mL), 4-(2- 2-hydroxyethoxy)benzaldehyde (0.027 g, 0.16 mmol, 1.1 eq) was added followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.44 mL, 0.44 mmol). The reaction mixture was heated at 85 0C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH3 until complete dissolution was observed. This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v 90:10) to give the title compound as an off- white solid (0.015 g, 20%); 1H-NMR (500Mz, DMSO-Cf6): δ 2.59-2.67 (m, 4H, piperazine N(CH2)2), 3.63-3.71 (m, 6H), 3.75 (td, 2H, J = 5.5, 5.5 Hz, CH2OH), 4.08 (t, 2H1 J = 4.8 Hz1 OCH2), 4.87 (t, 1 H1 J = 5.5 Hz1 OH)1 7.10 (d, J = 9.0 Hz1 2H, ArH1 C6H4), 7.68-7.75 (m, 1 H, py 4-H)1 8.12 (d, J = 8.5 Hz, 2H, ArH, C6H4), 8.20 (s, 1 H, imidazo[4,5-<b]pyridine 5-H)1 8.44-8.48 (m, py 2-H ), 8.49 (d, 1 H, J = 3.0 Hz, py 6-H)1 13.34 (br s, 1 H, imidazo[4,5-b]pyridine N-H); LC (Method B) - MS (ESI, m/z) 3.37 min - 527/529 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 527.1194, calculated for C24H25FBrN6O2 (M+H)+: 527.1201.
Example 5
2-(4-(6-bromo-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- ib]pyridin-2-yl)phenoxy)-Λ/,Λ/-dimethylethanamine
Figure imgf000016_0001
To a mixture of 5-bromo-4-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.060 g, 0.14 mmol) in EtOH (2.4 mL) and DMF (0.33 ml_), 4-(2- dimethylaminoethoxy)benzaldehyde (0.031 g, 0.16 mmol, 1.1 eq) was added followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.44 mL, 0.44 mmol). The reaction mixture was heated at 85 0C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH3 until complete dissolution was observed. This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v 90:10) to give the title compound as a off-white solid (0.005 g, 6%); 1H-NMR (500Mz, DMSO-d6): δ (NMe2 and CH2N masked under DMSO peak), 2.60-2.69 (m, 4H, piperazine N(CH2)2), 3.64-3.70 (m, 6H), 4.13-4.22 (m, 2H, OCH2), 7.11 (d, J = 9.0 Hz, 2H, ArH, C6H4), 7.69-7.75 (m, 1 H, py 4-H), 8.13 (d, J = 9.0 Hz, 2H1 ArH, C6H4), 8.21 (s, 1 H, imidazo[4,5Hb]pyridine 5-H)1 8.45-8.47 (m, py 2-H ), 8.49 (d, 1H, J = 2.5 Hz, py 6-H)1 13.35 (br s, 1H, imidazo[4,5- jb]pyridine N-H); LC (Method B) - MS (ESI, m/z) 2.32 min - 554/556 [(M+H)\ Br isotopic pattern]. ESI-HRMS: Found: 554.1679, calculated for C26H30FBrN7O (M+H)+: 554.1674.
Example 6 tø/t-Butyl-4-(6-bromo-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-b]pyridin-2-yl)benzyl carbamate
Figure imgf000017_0001
To a mixture of 5-bromo-4-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.110 g, 0.27 mmol, 1 eq), EtOH (4.7 mL) and DMF (0.63 ml_), te/t-butyl /V-(4-formylbenzyl)carbamate (0.069 g, 0.29 mmol, 1.1 eq) was added followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.80 mL, 0.80 mmol). The reaction mixture was heated at 85 °C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH3 until complete dissolution was observed. This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v; 94:6). The title compound was obtained after trituration with diethyl ether as an off-white solid (0.025 g, 15%); 1H- NMR (500Mz, DMSO-c/e): 1.40 (s, 9H, C(CH3)3), 2.58-2.68 (m, 4H, piperazine N(CHz)2), 3.63-3.70 (m, 6H, NCH2 and piperazine N(CH2)2), 4.19 (d, J = 6.0 Hz, 2H, NCH2), 7.38 (d, J = 8.0 Hz, 2H, ArH, C6H4), 7.70-7.75 (m, 1H, py 4-H), 8.12 (d, J = 8.0 Hz, 2H, ArH, C6H4), 8.24 (s, 1H, imidazo[4,5-ιb]pyridine 5-H), 8.46 (m, 1 H, py 2-H ), 8.50 (d, 1 H, J = 2.5 Hz, py 6-H), 13.51 (br s, 1 H, imidazo[4,5-ib]pyridine N-H); LC (Method B) - MS (ESI, m/z) 4.06 min - 596/598 [(M+H)+, Br isotopic pattern]; ESI- HRMS: Found: 596.1776, calculated for C28H32BrFN7O2 (M+H)+: 596.1780.
(4-(6-Bromo-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- £>]pyridin-2-yl)phenyl)methanamine
Figure imgf000017_0002
te/t-Butyl-4-(4-(6-bromo-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-<b]pyridin-2-yl)benzyl)piperazine-1-carboxylate (0.022 g, 0.037 mmol) was suspended in DCM (1.7 ml_) and the mixture cooled in an ice bath. TFA (0.40 ml_) was added and the resulting solution was allowed to warm up to room temperature and stirred for 2 h. The mixture was passed through an SCX column (2 g), the filtrate collected and the solvent removed in vacuo to give the title compound as a solid (0.016 g, 87%); 1H-NMR (500Mz, DMSO-c/6): 2.62-2.67 (m, 4H, piperazine N(CH2)2), 3.65-3.71 (m, 6H1 NCH2 and piperazine N(CHz)2). 3.94 (s, 2H, NCH2), 7.55 (d, J = 8.0 Hz, 2H, ArH, C6H4), 7.69-7.74 (m, 1H, py 4-H), 8.17 (d, J = 8.5 Hz, 2H, ArH, C6H4), 8.25 (s, 1H1 imidazo[4,5-b]pyridine 5-H), 8.45-8.48 (m, 1 H, py 2-H ), 8.50 (d, 1H, J = 3.0 Hz, py 6-H); LC (Method B) - MS (ESI, m/z) 2.07 min - 496/498 [(M+H)+, Br isotopic pattern]; ESI-HRMS: Found: 496.1249, calculated for C23H24BrFN7 (M+H)+: 496.1255.
Example 7 tert-Butyl 4-(4-(6-bromo-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-jb]pyridin-2-yl)phenyl)piperazine-1-carboxylate
Figure imgf000018_0001
To a mixture of 5-bromo-4-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.90 g, 0.22 mmol, 1 eq), EtOH (3.8 mL) and DMF (0.52 mL), tert-butyl-4-(4-formylphenyl)tetrahydro-1-(2H)pyrazine carboxylate (0.069 g, 0.24 mmol, 1.1 eq) was added followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.66 mL, 0.66 mmol). The reaction mixture was heated at 85 °C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH3 until complete dissolution was observed. This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v; 94:6). The title compound was obtained after trituration with diethyl ether as an off-white solid (0.025 g, 15%); 1H-NMR (500Mz, DMSO-Cf6): δ 1.42 (s, 9H, C(CH3J3), 2.59-2.67 (m, 4H, piperazine N(CH2)2), 3.23-3.29 (m, 4H, piperazine N(CH2)2), 3.43-3.52 (m, 4H, piperazine N(CH2)2), 3.60-3.70 (m, 6H), 7.07 (d, J = 9.0 Hz, 2H, ArH, C6H4), 7.70- 7.75 (m, 1 H, py 4-H), 8.13 (d, J = 9.0 Hz, 2H, ArH, C6H4), 8.18 (s, 1 H, imidazo[4,5- jb]pyridine 5-H), 8.45-8.48 (m, 1 H, py 2-H ), 8.50 (d, 1 H, J = 3.0 Hz, py 6-H)1 13.28 (brs, 1 H, imidazo[4,5-ιb]pyridine N-H); LC (Method B) - MS (ESI, m/z) 4.59 min - 651/653 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 651.2200, calculated for C3IH37FBrN8O2 (M+H)+: 651.2201.
6-Bromo-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-2-(4-(piperazin-1- yl)phenyl)-3/-/-imidazo[4,5-ό]pyridine
Figure imgf000019_0001
te/f-Butyl 4-(4-(6-bromo-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-jb]pyridin-2-yl)phenyl)piperazine-1-carboxylate (0.014 g, 0.021 mmol) was suspended in DCM (1.5 ml_) and the mixture cooled in an ice bath. TFA (0.30 mL) was added and the resulting solution was allowed to warm up to room temperature and stirred for 2 h. The mixture was passed through an SCX column (2 g), the filtrate collected and the solvent removed in vacuo to give the title compound as a solid (0.008 g, 69%); 1H-NMR (500Mz, DMSO-c/6): 2.61-2.66 (m, 4H, piperazine N(CH2J2), 3.11-3.16 (m, 4H, piperazine N(CH2J2), 3.38-3.43 (m, 4H, piperazine N(CH2)2), 3.63-3.70 (m, 6H), 7.10 (d, J = 9.0 Hz, 2H, ArH, C6H4), 7.69-7.74 (m, 1 H, py 4-H), 8.06 (d, J = 8.5 Hz, 2H, ArH, C6H4), 8.18 (s, 1 H, imidazo[4,5-ιb]pyridine 5-H), 8.45-8.47 (m, 1 H, py 2-H ), 8.50 (d, 1H, J = 3.0 Hz, py 6-H); LC (Method B) - MS (ESI, m/z) 2.19 min - 551/553 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 551.1668, calculated for C26H29FBrN8 (M+H)+: 551.1677.
Example 8
5-Chloro-4-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3-nitropyridin-2-amine
Figure imgf000020_0001
te/t-Butyl 4-((5-fluoropyridin-3-yl)methyl)piperazine-1-carboxylate (0.12 g, 0.42 mmol, 1.1 eq) was dissolved in DCM (1 mL) and the mixture cooled in a ice-water bath before the dropwise addition of TFA (0.5 mL). Stirring was continued at this temperature for 1 h and the solvents were removed in vacuo. The resulting crude material was azeotroped with toluene and dried. The resulting 1-((5-fluoropyridin-3- yl)methyl)piperazine (supposedly 0.082 g, 0.42 mmol, 1 eq) was suspended in 1PrOH (0.4 mL) and DIPEA (0.15 mL). To this solution, the 4,5-dichloro-3-nitropyridin- 2-amine (0.08 g, 0.38 mmol, 1.0 eq) was added, and the mixture heated and stirred for 17 h at 65°C. The mixture was filtered, washed with 1PrOH (3 x 3 mL), cold H2O (3 x 3 mL), Et2O (2 x 3 mL) and dried to give the title compound as a bright yellow solid (0.122 g, 44%); LC (Method B) - MS (ESI, m/z): Rt = 2.30 min - 367/369 [(M+H)+, Cl isotopic pattern, 100%].
tert-Butyl4-(4-(6-chloro-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-£>]pyridin-2-yl)phenyl)piperazine-1-carboxylate
Figure imgf000020_0002
To a mixture of 5-chloro-4-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.100 g, 0.27 mmol, 1 eq), EtOH (4.7 mL) and DMF (0.63 mL), terf-butyl-4-(4-formylphenyl)tetrahydro-1-(2H)pyrazine carboxylate (0.087 g, 0.3 mmol, 1.1 eq) was added followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.82 mL, 0.82 mmol). The reaction mixture was heated at 85°C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH3 until complete dissolution was observed. This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v; 94:6). The title compound was obtained after trituration with diethyl ether as an off-white solid (0.050 g, 30%); 1H-NMR (500Mz, DMSO-c/6): δ 1.43 (s, 9H, C(CH3J3), 2.59-2.67 (m, 4H, piperazine N(CH2)2), 3.45-3.52 (m, 4H, piperazine N(CH2J2), 3.66 (s, 2H, NCH2), 3.65-3.74 (m, 4H, piperazine N(CH2)2), missing 4H piperazine N(CH2)2 under solvents peaks, 7.07 (d, J = 9.5 Hz, 2H, ArH, C6H4), 7.64-7.74 (m, 1 H, py 4-H)1 8.01- 8.07 (m, 3H, ArH and imidazo [4,5-ό]pyridine 5-H), 8.44-8.47 (m, 1 H), 8.49 (d, J = 2.5 Hz, 1H, py 6-H), 13.19 (br s, 1H, imidazo[4,5-b]pyridine N-H); LC (Method B) - MS (ESI, m/z) 4.51 min - 607/609 [(M+H)+, Cl isotopic pattern]; ESI-HRMS: Found: 607.2743, calculated for C31H36CIFN8O2 (M+H)+: 607.2707.
6-Chloro-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-2-(4-(piperazin-1- yl)phenyl)-3/-/-imidazo[4,5-ύ]pyridine
Figure imgf000021_0001
terf-Butyl4-(4-(6-chloro-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3/-/- imidazo[4,5-ιb]pyridin-2-yl)phenyl)piperazine-1-carboxylate (0.050 g, 0.082 mmol) was suspended in DCM (5.9 mL) and the mixture cooled in an ice bath. TFA (1.32 mL) was added and the resulting solution was allowed to warm up to room temperature and stirred for 2 h. The mixture was passed through an SCX column (2 g), the filtrate collected and the solvent removed in vacuo to give the title compound as a solid (0.028 g, 67%); 1H-NMR (500Mz, DMSO-d6): 2.59-2.67 (m, 4H, piperazine N(CH2J2), 2.82-2.90 (m, 4H, piperazine N(CH2)2), 3.15-3.24 (m, 4H, piperazine N(CH2J2), 3.64-3.75 (m, 6H, piperazine N(CH2)2 and NCH2), 7.03 (d, J = 9.0 Hz, 2H, ArH, C6H4), 7.68-7.75 (m, 1 H, py 4-H), 7.99-8.07 (m, 3H), 8.44-8.47 (m, 1H), 8.49 (d, J = 2.5 Hz, 1 H, py 6-H); LC (Method B) - MS (ESI, m/z) 2.14 min - 507/509 [(M+H)\ Cl isotopic pattern]; ESI-HRMS: Found: 507.2183, calculated for C26H29CIFN8 (M+H)+: 507.2182.
Example 9 te/t-Butyl 4-(6-chloro-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-£>]pyridin-2-yl)benzylcarbamate
Figure imgf000022_0001
To a mixture of 5-chloro-4-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.100 g, 0.27 mmol, 1 eq), EtOH (4.7 ml.) and DMF (0.63 ml_), terf-butyl Λ/-(4-formylbenzyl)carbamate (0.071 g, 0.3 mmol, 1.1 eq) was added followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.82 mL, 0.82 mmol). The reaction mixture was heated at 85°C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH3 until complete dissolution was observed. This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v; 94:6). The title compound was obtained after trituration with diethyl ether as an off-white solid (0.040 g, 27%); 1H- NMR (500Mz, DMSO-CZ6): 1.41 (s, 9H, C(CHa)3), 2.60-2.67 (m, 4H1 piperazine N(CH2)2), 3.66 (s, 2H, NCH2), 3.68-3.75 (m, 4H, piperazine N(CH2J2), 4.19 (d, J = 6.0 Hz, 2H, CH2NH), 7.39 (d, J = 8.0 Hz, 2H, ArH, C6H4), 7.43 (t, 1 H, J = 5.0 Hz, NHBoc), 7.71 (ddd, 1 H, J = 10, 2.5, 1.5 Hz, py 4-H), 8.08-8.15 (m, 3H, ArH C6H4 and imidazo[4,5-/?]pyridine 5-H), 8.44-8.47 (m, 1H, py 2-H ), 8.49 (d, 1H1 J = 2.5 Hz, py 6- H), 13.43 (br s, 1 H, imidazo[4,5-ιb]pyridine N-H); LC (Method B) - MS (ESI, m/z) 3.97 min - 552/554 [(M+H)+, Cl isotopic pattern]; ESI-HRMS: Found: 552.2283, calculated for C28H32CIFN7O2 (M+H)+: 552.2285.
(4-(6-Chloro-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- /)]pyridin-2-yl)phenyl)methanamine
Figure imgf000023_0001
terf-Butyl 4-(6-chloro-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-/3]pyndin-2-yl)benzylcarbamate (0.040 g, 0.072 mmol) was suspended in DCM (5.1 ml_) and the mixture cooled in an ice bath. TFA (1.1 mL) was added and the resulting solution was allowed to warm up to room temperature and stirred for 2 h. The mixture was passed through an SCX column (2 g), the filtrate collected and the solvent removed in vacuo to give the title compound as a solid (0.017 g, 52%); 1H-NMR (500Mz, DMSO-d6): 2.60-2.67 (m, 4H, piperazine N(CHZ)2), 3.67 (s, 2H), 3.69-3.75 (m, 4H, piperazine N(CH2)2), 3.81 (s, 2H), 7.50 (d, J = 8.0 Hz, 2H, ArH, C6H4), 7.69-7.75 (m, 1 H, py 4-H), 8.10 (s, 1H, imidazo[4,5-/3]pyridine 5-H), 8.12 (d, 2H, J = 8.5 Hz, ArH C6H4), 8.44-8.47 (m, 1 H, py 2-H ), 8.49 (d, 1 H, J = 3.0 Hz, py 6- H); LC (Method B) - MS (ESI, m/z) 2.00 min - 452/454 [(M+H)+, Cl isotopic pattern]; ESI-HRMS: Found: 452.1761 , calculated for C23H24CIFN7 (M+H)+: 452.1760.
Example 10
5-Chloro-3-nitro-4-(4-((6-(trifluoromethyl)pyridin-3-yl)methyl)piperazin-1-yl)pyridin-2- amine
Figure imgf000023_0002
terf-Butyl 4-((6-(trifluoromethyl)pyridin-3-yl)methyl)piperazine-1 -carboxylate (prepared as described in example 124 of PCT/GB2006/004854; 0.37 g, 1.06 mmol, 1.1 eq) was dissolved in DCM (3.2 mL) and the mixture cooled in a ice-water bath before the dropwise addition of TFA (1 ml_). Stirring was continued at this temperature for 1 h and the solvents were removed in vacuo. The resulting crude material was azeotroped with toluene and dried.
1-((6-(Trifluoromethyl)pyridin-3-yl)methyl)piperazine (supposedly 0.261 g, 1.06 mmol, 1.1 eq) was suspended in 1PrOH (3.2 ml_) and DIPEA (0.8 ml_). To this solution, 4,5- dichloro-3-nitropyridin-2-amine (0.20 g, 0.96 mmol, 1 eq) was added and the mixture heated and stirred for 17 h at 65°C. The mixture was filtered, washed with 1PrOH (3 x 3 mL), cold H2O (3 x 3 ml_), Et2O (2 x 3 mL), and dried to give the title compound (0.308 g, 70%); 1H-NMR (500 MHz, DMSO-d6): δ 2.50-2.62 (m, 4H, piperazine N(CH2)2), 3.02-3.14 (m, 4H, piperazine N(CH2J2), 3.70 (s, 2H, NCH2), 6.96 (brs, 2H, NH2), 7.88 (d, J = 8.0 Hz, 1 H, CF3py-H), 8.04 (d, J = 8.0 Hz, 1 H. CF3py-H), 8.06 (s, 1 H, chloropyridine 6-H), 8.72 (s, 1 H, CF3py-H) ; LC (Method B) - MS (ESI, m/z): Rt = 3.22 min - 417/419 [(M+H+), Cl isotopic pattern, 100%].
terf-Butyl 4-(6-chloro-7-(4-((6-(trifluoromethyl)pyridin-3-yl)methyl)piperazin-1-yl)-3/-/- imidazo[4,5-fb]pyridin-2-yl)benzyl carbamate
Figure imgf000024_0001
To a mixture of 5-chloro-3-nitro-4-(4-((6-(trifluoromethyl)pyridin-3- yl)methyl)piperazin-1-yl)pyridin-2-amine (0.100 g, 0.24 mmol, 1 eq), EtOH (4.3 mL) and DMF (0.57 mL), fe/f-butyl Λ/-(4-formylbenzyl)carbamate (0.062 g, 0.26 mmol, 1.1 eq) was added followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.72 mL, 0.72 mmol). The reaction mixture was heated at 85°C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH3 until complete dissolution was observed. This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v; 94:6). The title compound was obtained after trituration with diethyl ether as an off-white solid (0.029 g, 20%); 1H-NMR (500Mz, DMSO-c/6): δ 1.41 (s, 9H, C(CHg)3), 2.60-2.70 (m, 4H, piperazine N(CH2J2), 3.67-3.77 (m, 6H, NCH2 and piperazine N(CH2)2), 4.19 (d, J = 6.0 Hz1 2H, CH2NH), 7.38 (d, J = 8.0 Hz, 2H, ArH, C6H4), 7.43 (t, 1H, J = 5.5 Hz, NHBoc), 7.90 (d, 1 H, J = 8.0 Hz, pyridine H), 8.05-8.15 (m, 4H1 ArH C6H4, pyridine H, imidazo[4,5-ιb]pyridine 5-H), 8.75-8.79 (m, 1 H, pyridine H), 13.43 (br s, 1 H, imidazo[4,5-k]pyridine N-H); LC (Method B) - MS (ESI, m/z) 4.47 min - 602/604 [(M+H)+, Cl isotopic pattern]; ESI-HRMS: Found; 602.2252, calculated for C29H32CIF3N7O2 (M+H)+: 602.2253.
4-(6-Chloro-7-(4-((6-(trifluoromethyl)pyridin-3-yl)methyl)piperazin-1-yl)-3/-/- imidazo[4,5-(b]pyridin-2-yl)phenyl)methanamine
Figure imgf000025_0001
te/f-butyl 4-(4-(6-chloro-7-(4-((6-(trifluoromethyl)pyridin-3-yl)methyl)piperazin-1-yl)- 3H-imidazo[4,5-/?]pyridin-2-yl)phenyl)piperazine-1-carboxylate (0.029 g, 0.048 mmol) was suspended in DCM (3.4 ml_) and the mixture cooled in an ice bath. TFA (0.8 ml_) was added and the resulting solution was allowed to warm up to room temperature and stirred for 2 h. The mixture was passed through an SCX column (2 g), the filtrate collected and the solvent removed in vacuo to give the title compound as a solid (0.011 g, 46%); 1H-NMR (500Mz, DMSO-c/6): 2.60-2.69 (m, 4H, piperazine N(CH2)2), 3.68-3.76 (m, 6H, NCH2 and piperazine N(CH2J2), 3.84 (s, 2H), 7.51 (d, J = 8.0 Hz, 2H, ArH, C6H4), 7.90 (d, 1 H, J = 8.0 Hz, pyridine H), 8.05-8.16 (m, 4H), 8.77 (s, 1H, pyridine H ); LC (Method B) - MS (ESI, m/z) 2.43 min - 502/504 [(M+H)+, Cl isotopic pattern]; ESI-HRMS: Found: 502.1729, calculated for C24H23CIF3N7 (M+H)+: 502.1728.
Example 11 terf-Butyl 4-(4-(6-chloro-7-(4-((6-(trifluoromethyl)pyridin-3-yl)methyl)piperazin-1-yl)- 3H-imidazo[4,5-/3]pyridin-2-yl)phenyl)piperazine-1-carboxylate
Figure imgf000026_0001
To a mixture of 5-chloro-3-nitro-4-(4-((6-(trifluoromethyl)pyridin-3- yl)methyl)piperazin-1-yl)pyridin-2-amine (0.100 g, 0.24 mmol, 1 eq), EtOH (4.3 mL) and DMF (0.57 mL), te/t-butyl-4-(4-formylphenyl)tetrahydro-1-(2H)pyrazine carboxylate (0.077 g, 0.26 mmol, 1.1 eq) was added followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.72 mL, 0.72 mmol). The reaction mixture was heated at 85°C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH3 until complete dissolution was observed. This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v; 94:6). The title compound was obtained after trituration with diethyl ether as an off-white solid (0.042 g, 27%); 1H-NMR (500Mz, DMSO-c/6): 1.43 (s, 9H, C(CH3J3), 2.60-2.67 (m, 4H, piperazine N(CH2J2), 3.27 (t, 4H, J = 5.2 Hz, piperazine N(CH2)2), 3.45-3.51 (m, 4H, piperazine N(CH2J2), 3.66-3.72 (m, 4H, piperazine N(CH2)2), 3.73 (s, 2H, NCH2), 7.06 (d, J = 9.0 Hz, 2H, ArH, C6H4), 7.90 (dd, 1H, pyridine H), 8.03 (d, J = 9.0 Hz, 2H, ArH1 C6H4), 8.04 (s, 1 H, imidazo[4,5-ό]pyridine 5- H), 8.06-8.11 (m, 1H, pyridine H ), 8.75-8.78 (m, 1H, pyridine H), 13.19 (br s, 1H, imidazo[4,5-ib]pyridine N-H); LC (Method B) - MS (ESI, m/z) 4.93 min - 657/659 [(M+H)+, Cl isotopic pattern]; ESI-HRMS: Found: 657.2675, calculated for C32H37CIF3N8O2 (M+H)+: 657.2671.
6-Chloro-2-(4-(piperazin-1-yl)phenyl)-7-(4-((6-(trifluoromethyl)pyridin-3- yl)methyl)piperazin-1-yl)-3H-imidazo[4,5-b]pyridine
Figure imgf000026_0002
tert-butyl 4-(4-(6-chloro-7-(4-((6-(trifluoromethyl)pyridin-3-yl)methyl)piperazin-1-yl)- 3H-imidazo[4,5-b]pyridin-2-yl)phenyl)piperazine-1-carboxylate (0.042 g, 0.064 mmol) was suspended in DCM (4.6 ml_) and the mixture cooled in an ice bath. TFA (1.0 mL) was added and the resulting solution was allowed to warm up to room temperature and stirred for 2 h. The mixture was passed through an SCX column (2 g), the filtrate collected and the solvent removed in vacuo to give the title compound as a solid (0.027 g, 76%); 1H-NMR (500Mz1 DMSO-c/6): δ 2.60-2.67 (m, 4H, piperazine N(CH2J2), 2.82-2.90 (m, 4H, piperazine N(CH2J2), 3.16-3.22 (m, 4H, piperazine N(CH2)2), 3.65-3.71 (m, 4H, piperazine N(CH2J2), 3.73 (s, 2H, NCH2), 7.03 (d, J = 9.0 Hz, 2H, ArH, C6H4), 7.90 (d, J = 8.0 Hz, 1 H, pyridine H), 8.01 (d, J = 9.0 Hz, 2H, ArH, C6H4), 8.04 (s, 1H, imidazo[4,5-*b]pyridine 5-H), 8.06-8.11 (m, 1H, pyridine H ), 8.75- 8.78 (m, 1 H, pyridine H); LC (Method B) - MS (ESI, m/z) 2.55 min - 557/559 [(M+H)+, Cl isotopic pattern]; ESI-HRMS: Found: 557.2147, calculated for C27H29CIF3N8 (M+H)+: 557.2150.
Example 12
2-(4-(6-Bromo-7-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- /j/pyridin-2-yl)phenoxy)ethanol
Figure imgf000027_0001
To a mixture of 5-bromo-4-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (prepared as described in example 144 of PCT/GB2006/004854 0.062 g, 0.14 mmol) in EtOH (2.6 mL) and DMF (0.35 mL), 4-(2-2- hydroxyethoxy)benzaldehyde (0.027 g, 0.16 mmol, 1.1 eq) was added followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.44 mL, 0.44 mmol). The reaction mixture was heated at 85 0C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH3 until complete dissolution was observed. This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v 90:10) to give the title compound as an off- white solid (0.018 g, 24%); 1H-NMR (500Mz, DMSO-c/6): δ 2.59-2.67 (m, 7H, piperazine N(CH2)2 and CH3-thiazole), 3.63-3.71 (m, 6H), 3.75 (td, 2H, J = 5.5, 5.5 Hz, CH2OH), 4.04-4.11 (m, 2H, CH2CH2OH), 4.90 (t, 1H, J = 5.5 Hz, OH), 7.11 (d, J = 9.0 Hz, 2H, ArH, C6H4), 7.33 (s, 1 H, 5H-thiazole), 8.13 (d, J = 9.0 Hz, 2H, ArH, C6H4), 8.21 (s, 1H, imidazo[4,5-ib]pyridine 5-H)1 13.35 (br s, 1H, imidazo[4,5- ώ]pyridine N-H); LC (Method B) - MS (ESI, m/z) 2.96 min - 529/531 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 529.1014, calculated for C23H26BrN6O2S (M+H)+: 529.1016.
Example 13
2-(4-(6-Bromo-7-(4-((2-methylthiazol-4-yl)nnethyl)piperazin-1-yl)-3H-imidazo[4,5- b]pyridin-2-yl)phenoxy)-Λ/,Λ/-dimethylethanamine
Figure imgf000028_0001
To a mixture of 5-bromo-4-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.062 g, 0.14 mmol) in EtOH (2.6 ml_) and DMF (0.35 ml_), 4-(2- dimethylaminoethoxy)benzaldehyde (0.032 g, 0.16 mmol, 1.1 eq) was added, followed by a freshly prepared aqueous solution of Na2S2O4 (1M; 0.44 ml_, 0.44 mmol). The reaction mixture was heated at 85 0C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH3 until complete dissolution was observed. This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v 90:10) to give the title compound as an off-white solid (0.007 g, 8.9%); 1H-NMR (500Mz, DMSO-c/6): δ 2.27 (s, 6H, NMe2), 2.63-2.75 (m, 9H, piperazine N(CH2)2, CH3-thiazole, CH2N), 3.64-3.70 (m, 6H), 3.61-3.69 (m, 6H, piperazine N(CH2)2, CH2N), 4.11-4.17 (m, 2H, OCH2), 7.10 (d, J = 9.0 Hz, 2H, ArH, C6H4), 7.33 (s, 1 H, 5H-thiazole), 8.12 (d, J = 9.0 Hz, 2H, ArH, C6H4), 8.20 (s, 1 H, imidazo[4,5-/b]pyridine 5-H)1 13.39 (br s, 1 H, imidazo[4,5- 6]pyridine N-H); LC (Method B) - MS (ESI, m/z) 2.06 min - 556/558 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 556.1488, calculated for C25H31BrN7O (M+H)+: 556.1489. Example 14
1-(4-(6-Bromo-7-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3/-/-imidazo[4,5- b]pyridin-2-yl)phenyl)piperidin-4-ol
Figure imgf000029_0001
5-bromo-4-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3-nitropyridin-2-amine (0.062 g, 0.14 mmol) in EtOH (2.6 mL) and DMF (0.35 mL), 4-(4-hydroxypiperidin-1- yl)benzaldehyde (0.034 g, 0.16 mmol, 1.1 eq) was added, followed by a freshly prepared aqueous solution of Na2S2O4 (1M; 0.44 mL, 0.44 mmol). The reaction mixture was heated at 85 0C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH3 until complete dissolution was observed. This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v 90:10) to give the title compound as an off-white solid (0.031 g, 39%); 1H-NMR (500Mz, DMSO-d6): δ 1.41-1.51 (m, 2H), 1.79-1.87 (m, 2H), 2.60-2.92 (m, 6H, piperazine N(CH2)2, CH2), 2.96-3.05 (m, 2H), 3.60-3.75 (m, 9H), 4.70 (d, J = 4.5 Hz, 1 H, CHOH), 7.05 (d, J = 9.0 Hz, 2H, ArH, C6H4), 7.33 (s, 1H, 5H- thiazole), 8.01 (d, J = 9.0 Hz, 2H, ArH, C6H4), 8.17 (s, 1 H, imidazo[4,5-<b]pyridine 5- H), 13.39 (br s, 1 H, imidazo[4,5-<b]pyridine N-H); LC (Method B) - MS (ESI, m/z) 3.07 min - 568/570 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 568.1489, calculated for C26H31BrN7OS (M+H)+: 568.1489.
Example 15
4-(5-(6-Bromo-7-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- ifc>]pyridin-2-yl)pyridin-2-yl)morpholine
Figure imgf000029_0002
To a mixture of 5-bromo-4-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.062 g, 0.15 mmol, 1 eq), EtOH (2.68 ml_) and DMF (0.35 mL), 6-morpholin-4-yl-pyridine-3-carbaldehyde (0.032 g, 0.17 mmol, 1.1 eq) was added followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.45 mL, 0.45 mmol). The reaction mixture was heated at 85 °C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH3 until complete dissolution was observed. This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v; 95:5). The title compound was obtained after trituration with diethyl ether as a off-white solid (0.020 g, 24%); 1H- NMR (500Mz, DMSO-de): δ 2.63-2.71 (m, 7H), 3.56-3.62 (s, 4H), 3.62-3.68 (m, 6H), 3.70-3.75 (m, 4H), 7.33 (s, 1 H, thiazole 5-H), 8.20 (s, 1 H, imidazo[4,5-6]pyridine 5- H), 8.27 (dd, J = 9.0, 2.5 Hz, 1 H, py H ), 8.92 (d, J = 2.0 Hz, 1 H, py H), 13.31 (brs, 1H, imidazo[4,5-ib]pyridine N-H); LC (Method B) - MS (ESI, m/z) 3.05 min - 555/557 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 555.1280, calculated for C24H28BrN8OS (M+H)+: 555.1285.
Example 16 tert-Butyl 4-(5-(6-bromo-7-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-(b]pyridin-2-yl)pyridin-2-yl)piperazine-1-carboxylate
Figure imgf000030_0001
To a mixture of 5-bromo-4-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.079 g, 0.19 mmol, 1 eq), EtOH (3.3 mL) and DMF (0.44 mL), te/t-butyl-4-(5-formylpyrid-2-yl)piperazine-1-carboxylate (0.061 g, 0.21 mmol, 1.1 eq) was added followed by a freshly prepared aqueous solution of Na2S2O4 (1M; 0.57 mL, 0.57 mmol). The reaction mixture was heated at 85 °C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH3 until complete dissolution was observed. This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v; 95:5). The title compound was obtained after trituration with diethyl ether as an off-white solid (0.033 g, 27%); 1H- NMR (500Mz, DMSO-c/e): δ 1.44 (s, 9H, C(CH3J3), 2.63-2.71 (m, 7H), 3.44-3.49 (m, 4H), 3.60-3.69 (m, 10H), 6.98 (d, J = 9.0 Hz, 1 H, py 3-H), 7.33 (s, 1 H, thiazole 5-H), 8.18 (S, 1H, imidazo[4,5-/fc>]pyridine 5-H), 8.26 (dd, J = 8.8, 2.2 Hz, 1 H, py 4-H ), 8.91 (d, J = 2.5 Hz, 1H, py 6-H); LC (Method B) - MS (ESI, m/z) 3.84 min - 654/656 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 654.1961 , calculated for C29H37BrN9O2S (M+H)+: 654.1969.
4-((4-(6-Bromo-2-(6-(piperazin-1-yl)pyridin-3-yl)-3/-/-imidazo[4,5-b]pyridin-7- yl)piperazin-1-yl)methyl)-2-methylthiazole
Figure imgf000031_0001
te/if-Butyl 4-(5-(6-bromo-7-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-b]pyridin-2-yl)pyridin-2-yl)piperazine-1-carboxylate (0.028 g, 0.043 mmol) was suspended in DCM (3 mL) and the mixture cooled in an ice bath. TFA (0.60 mL) was added and the resulting solution was allowed to warm up to room temperature and stirred for 2 h. The mixture was passed through an SCX column (2 g), the filtrate collected and the solvent removed in vacuo to give the title compound as a solid (0.022 g, 93%); 1H-NMR (500Mz, DMSO-d6): δ 2.63-2.71 (m, 7H, piperazine N(CH2)2 and CH3-thiazole), 2.78-2.86 (m, 4H, piperazine N(CH2)2), 3.53-3.60 (m, 4H, piperazine N(CH2J2), 3.61-3.70 (m, 6H), 6.95 (d, J = 9.0 Hz, 1H, py 3-H), 7.33 (s, 1H, thiazole 5-H), 8.19 (s, 1H, imidazo[4,5-/b]pyridine 5-H), 8.22 (dd, J = 9.0, 2.0 Hz, 1 H, py 4-H ), 8.89 (d, J = 2.5 Hz, 1H, py 6-H); LC (Method B) - MS (ESI, m/z) 2.04 min - 554/556 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 554.1448, calculated for C24H29BrN9S (M+H)+: 554.1444.
Example 17
5-Chloro-4-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3-nitropyridin-2-amine
Figure imgf000032_0001
te/t-Butyl 4-((2-methylthiazol-4-yl)methyl)piperazine-1-carboxylate (prepared as described in example 144 of PCT/GB2006/004854, 0.24 g, 0.79 mmol) was suspended in DCM (2.4 mL) and the mixture cooled in an ice bath. TFA (0.75 mL) was added and the resulting solution was allowed to warm up to room temperature and stirred for 2 h. The resulting 2-methyl-4-(piperazin-1-ylmethyl)thiazole ( supposedly 0.20 g, 0.79 mmol, 1.1 eq) was suspended in 1PrOH (2.4 mL) and DIPEA (0.6 mL). To this solution, 4,5-dichloro-3-nitropyridin-2-amine (0.150 g, 0.72 mmol, 1 eq) was added and the mixture heated and stirred for 17 h at 65°C. The mixture was filtered, washed with 1PrOH (3 x 3 mL), cold H2O (3 x 3 mL), Et2O (2 x 3 mL), and dried to give the title compound (0.17 g, 60%); 1H-NMR (500 MHz, DMSO-Cf6): δ 2.51-2.59 (m, 4H, piperazine N(CH2)2), 3.06 (t, 4H, J = 4.3, piperazine N(CH2J2), 3.59 (s, 2H, NCH2), 6.94 (brs, 2H, NH2), 7.28 (s, 1H. thiazole 5-H), 8.05 (s, 1H, pyridine 6- H); LC (Method B) - MS (ESI, m/z): Rt = 2.00 min - 369/371 [(M+H+), Cl isotopic pattern, 100%].
tert-Butyl 4-(4-(6-chloro-7-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-/fc>]pyridin-2-yl)phenyl)piperazine-1-carboxylate
Figure imgf000032_0002
To a mixture of 5-chloro-4-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.100 g, 0.27 mmol, 1 eq), EtOH (4.73 mL) and DMF (0.63 mL), te/Y-butyl-4-(4-formylphenyl)tetrahydro-1-(2H)pyrazine carboxylate (0.087 g, 0.3 mmol, 1.1 eq) was added followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.82 ml_, 0.82 mmol). The reaction mixture was heated at 85°C for 24 h, then allowed to cool to room temperature and diluted with DCM and a few drops of aqueous NH3 until complete dissolution was observed. This solution was deposited on two preparative silica TLC plates and eluted with DCM/MeOH (v/v; 94:6). The title compound was obtained after trituration with diethyl ether as a off-white solid (0.025 g, 15%); 1H-NMR (500Mz, DMSO-c/6): δ 1.43 (s, 9H, C(CH3)3), 2.63-2.69 (m, 7H1 piperazine N(CH2)2 and thiazole Me), (piperazine N(CH2)2 masked under water or DMSO peak), 3.45-3.51 (m, 4H, piperazine N(CH2)2), 3.63 (s, 2H, NCH2), 3.65-3.71 (m, 4H, piperazine N(CH2J2), 7.07 (d, J = 9.0 Hz, 2H1 ArH, C6H4), 7.31 (s, 1 H, thiazole 5-H)1 8.00-8.06 (m, 3H, ArH, C6H4 and imidazo[4,5-ιb]pyridine 5-H)1 13.18 (br s, 1 H, imidazo[4,5-ιb]pyridine N-H); LC (Method B) - MS (ESI, m/z) 3.93 min - 609/611 [(M+H)+, Cl isotopic pattern]; ESI-HRMS: Found: 609.2648, calculated for C30H38CIN8O2S (M+H)+: 609.2521.
4-((4-(6-Chloro-2-(4-(piperazin-1-yl)phenyl)-3H-imidazo[4,5-d]pyridin-7-yl)piperazin-1- yl)methyl)-2-methylthiazole
Figure imgf000033_0001
tert-Butyl-4-(4-(6-bromo-7-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-<fc>]pyridin-2-yl)phenyl)piperazine-1-carboxylate (0.025 g, 0.041 mmol) was suspended in DCM (2.9 mL) and the mixture cooled in an ice bath. TFA (0.66 mL) was added and the resulting solution was allowed to warm up to room temperature and stirred for 2 h. The mixture was passed through an SCX column (2 g), the filtrate collected and the solvent removed in vacuo to give the title compound as a solid (0.009 g, 43%); 1H-NMR (500Mz, DMSO-d6): δ 2.63-2.69 (m, 7H, piperazine N(CH2)2 and thiazole Me), 2.96-3.03 (m, 4H, piperazine N(CH2)2), 3.64 (s, 2H1 NCH2), 3.65-3.71 (m, 4H, piperazine N(CH2J2), 7.07 (d, J = 9.0 Hz, 2H, ArH, C6H4), 7.32 (s, 1 H, thiazole 5-H), 8.00-8.06 (m, 3H, ArH, C6H4 and imidazo[4,5- b]pyridine 5-H), 13.18 (br s, 1H, imidazo[4,5-b]pyridine N-H); LC (Method B) - MS (ESI, m/z) 2.09 min - 509/511 [(M+H)+, Cl isotopic pattern]; ESI-HRMS: Found: 509.1997, calculated for C25H30CIN8S (M+H)+: 509.1997.
Example 18
4-(3-(6-Bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-3/-/-imidazo[4,5-b]pyridin-2- yl)benzyl)morpholine
Figure imgf000034_0001
To a mixture of 5-bromo-4-[4-(4-chloro-benzyl)-piperazin-1-yl]-3-nitro-pyridin-2- ylamine (prepared as described in example 26 of PCT/GB2006/004854; 0.043 g, 0.10 mmol) and EtOH (6.0 ml_) was added 3-(morpholinomethyl)benzaldehyde (0.027 g, 0.13 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.40 mL, 0.40 mmol). The reaction mixture was stirred at 80 0C for 20 h, then allowed to cool to room temperature and concentrated in vacuo. The resulting residue was absorbed on silica, and the free-running powder was placed on a 10 g isolute silica column. Elution with a gradient of methanol (0 to 8%) in ethyl acetate/dichloromethane (v:v; 1:1) afforded a pale yellow solid. The title compound was obtained as a white solid after trituration with diethyl ether (0.017 g, 29%); 1H- NMR (500 MHz, DMSO-d6) 2.40 (br s, 4H) and 2.61 (br s, 4H) (morpholine N(CH2)2 and piperazine N(CH2)2), 3.60 (br t, J = 4.39 Hz, 4H) and 3.67 (br s, 4H) (morpholine O(CH2)2 and piperazine N(CH2J2), 3.56 (s, 2H) and 3.57 (s, 2H) (NCH2-C6H4CI and C6H4CH2), 7.41 (m, 4 H, C6H4CI), 7.44 (d, J = 7.0 Hz, 1H, 4-ArH or 6-ArH), 7.49 (t, J = 7.8 Hz, 1 H, 5-ArH), 8.06 (d, J = 7.4 Hz, 1 H, 4-ArH or 6-ArH), 8.14 (s, 1 H, 2-ArH), 8.24 (s, 1H, imidazo[4,5-b]pyridine 5-H), 13.51 (br s, 1 H, imidazo[4,5-jb]pyridine N-
H);
LC (Method B) - MS (ESI, m/z): Rt = 2.49 min - 581 , 583, 585 [(M+H)+, BrCI isotopic pattern]; ESI-HRMS: Found: 581.1421 , calculated for C28H3iBrCIN6O (M+H)+:
581.1426.
Example 19 terf-Butyl 4-(4-(6-bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-3H-imidazo[4,5- b]pyridin-2-yl)benzyl)piperazine-1-carboxylate
Figure imgf000035_0001
To a mixture of 5-bromo-4-[4-(4-chloro-benzyl)-piperazin-1-yl]-3-nitro-pyridin-2- ylamine (0.043 g, 0.10 mmol) and EtOH (6.0 mL) was added tert-butyl 4-(4- formylbenzyl)piperazine-1-carboxylate (0.040 g, 0.13 mmol) followed by a freshly prepared aqueous solution of Na2S2θ4 (1 M; 0.40 mL, 0.40 mmol). The reaction mixture was stirred at 80 0C for 24 h, then allowed to cool to room temperature and concentrated in vacuo. The residue was absorbed on silica gel, the free-running powder was placed on a 10 g isolute silica column, and elution with a gradient of methanol (0 to 7%) in ethyl acetate / dichloromethane (v:v; 1 :1 ) afforded the title compound as a pale yellow solid (0.036 g, 53%). 1H-NMR (500 MHz, DMSO-d6) 1.39 (s, 9H, OC(CH3)3), 2.33 (br t, J = 4.7 Hz, 4H), 2.61 (br s, 4H), 3.33 (br s, 4H), 3.67 (br s, 4H) (4 x piperazine N(CH2)2), 3.56 (s, 2H) and 3.57 (s, 2H) (NCH2-C6H4CI and C6H4CH2, 7.41 (m, 4 H, C6H4CI), 7.47 (d, J = 7.6 Hz, 2H) and 8.14 (d, J = 7.6 Hz, 2H) (3,5-C6H4 and 2,6-C6H4), 8.23 (s, 1 H, imidazo[4,5-<b]pyridine 5-H), 13.48 (br s, 1 H, imidazo[4,5-b]pyridine N-H); LC (Method B) - MS (ESI, m/z): Rt = 3.19 min - 680, 682, 684 [(M+H)+, BrCI isotopic pattern].
6-Bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-2-(4-(piperazin-1-ylmethyl)phenyl)-3H- imidazo[4,5-6]pyridine
Figure imgf000035_0002
A solution of tert-butyl 4-(4-(6-bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-3H- imidazo[4,5-£»]pyridin-2-yl)benzyl)piperazine-1-carboxylate (0.030 g, 0.045 mmol) in dichloromethane (4.0 mL) and trifluoroacetic acid (0.6 mL) was stirred at room temperature for 1 h, then it was concentrated in vacuo. The residue was dissolved in methanol and the solution was placed on a 5g isolute SCX Il column. Elution with methanol, and then 0.1M, and 0.4M ammonia in methanol afforded a yellow solid. The title compound was obtained as a pale yellow solid after trituration with diethyl ether (0.02Og, 80%). 1H-NMR (500 MHz, DMSO-d6) 2.33 (br s, 4H), 2.61 (br s, 4H), 2.72 (br t, J = 4.5 Hz, 4H), 3.66 (br s, 4H) (piperazine N(CH2)2), 3.50 (s, 2H) and 3.57 (s, 2H) (NCH2-C6H4CI and C6H4CH2, 7.41 (m, 4 H, C6H4CI), 7.45 (d, J = 8.1 Hz, 2H) and 8.13 (d, J = 8.1 Hz, 2H) (3,5-C6H4 and 2,6-C6H4), 8.23 (s, 1H, imidazo[4,5- jb]pyridine 5-H);
LC (Method B) - MS (ESI, m/z): Rt = 2.54 min - 580, 582, 584 [(M+H)+, BrCI isotopic pattern]. ESI-HRMS: Found: 580.1575, calculated for C28H32BrCIN7 (M+H)+: 580.1585.
Example 20 terf-Butyl 4-(4-(6-bromo-7-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-b]pyridin-2-yl)benzyl)piperazine-1-carboxylate
Figure imgf000036_0001
To a mixture of 5-bromo-4-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (prepared as described in example 74 of PCT/GB2006/004854; 0.039 g, 0.10 mmol) and EtOH (4.0 ml_) was added tert-butyl 4-(4- formylbenzyl)piperazine-1-carboxylate (0.040 g, 0.13 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1M; 0.40 mL, 0.40 mmol). The reaction mixture was stirred at 80 0C for 20 h, then allowed to cool to room temperature and concentrated in vacuo. The residue was absorbed on silica gel, the free-running powder was placed on a 10 g isolute silica column, and elution with a gradient of methanol (0 to 6%) in ethyl acetate / dichloromethane (v:v; 1 :1 ) afforded the title compound as a pale yellow solid (0.031 g, 48%). 1H-NMR (500 MHz, DMSO-d6) 1.39 (s, 9H, OC(CH3)3), 2.40 (s, 3H, isoxazole-5-CH3), 2.34 (br t, J = 4.8 Hz, 4H), 2.64 (br s, 4H), 3.33 (br s, 4H), and 3.67 (br s, 4H) (4 x piperazine N(CH2)2), 3.56 (s, 2H) and 3.60 (s, 2H) (NCHs-isoxazole and C6H4CH2), 6.25 (s, 1H, isoxazole 4-H), 7.47 (d, J = 8.1 Hz, 2H) and 8.12 (d, J = 8.1 Hz, 2H) (3,5-C6H4 and 2,6-C6H4), 8.24 (s, 1 H, imidazo[4,5-/3]pyridine 5-H), 13.48 (br s, 1H, imidazo[4,5-fo]pyridine N-H);
LC (Method B) - MS (ESI, m/z): Rt = 3.00 min - 651, 653 [(M+H)\ Br isotopic pattern].
3-((4-(6-Bromo-2-(4-(piperazin-1-ylmethyl)phenyl)-3/-/-imidazo[4,5-ύ]pyridin-7- yl)piperazin-1-yl)methyl)-5-methylisoxazole
Figure imgf000037_0001
A solution of tert-butyl 4-(4-(6-bromo-7-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1- yl)-3/-/-imidazo[4,5-ib]pyridin-2-yl)benzyl)piperazine-1-carboxylate (0.024 g, 0.037 mmol) in dichloromethane (4.0 ml_) and trifluoroacetic acid (0.5 mL) was stirred at room temperature for 1 h, then it was concentrated in vacuo. The residue was dissolved in methanol and the solution was placed on a 5g isolute SCX Il column. Elution with methanol and then 0.1 M, and 0.4M ammonia in methanol afforded a yellow solid. The title compound was obtained as an off-white solid after trituration with diethyl ether (0.01Og, 50%). 1H-NMR (500 MHz, DMSOd6) 2.40 (s, 3H, isoxazole-5-CH3), 2.34 (br s, 4H), 2.64 (br s, 4H), 2.74 (br t, J = 4.5 Hz, 4H) and 3.66 (br s, 4H) (4 x piperazine N(CH2J2), 3.51 (s, 2H) and 3.60 (s, 2H) (NCH2-isoxazole and C6H4CH2), 6.25 (s, 1 H, isoxazole 4-H), 7.46 (d, J = 8.1 Hz, 2H) and 8.13 (d, J = 8.1 Hz, 2H) (3,5-C6H4 and 2,6-C6H4), 8.23 (s, 1 H, imidazo[4,5-*b]pyridine 5-H); LC (Method B) - MS (ESI, m/z): Rt = 2.28 min - 551 , 553 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 551.1870, calculated for C26H32BrN8O (M+H)+: 551.1877.
Example 21 ferf-Butyl 4-((2-methylpyridin-4-yl)methyl)piperazine-1-carboxylate
Figure imgf000038_0001
To a mixture of 1-BOC-piperazine (0.615 g, 3.30 mmol) and 1 ,2 DCE (12 ml_) was added 2-methylpyridine-4-carboxaldehyde (0.363 g, 3.0 mmol) followed by the slow addition of sodium triacetoxyborohydride (0.885 g, 4.19 mmol). The reaction mixture was stirred at room temperature for 24 h, then partitioned between ethyl acetate (70 ml_) and saturated aqueous NaHCO3 (50 mL). The aqueous layer was extracted with ethyl acetate (2 x 50 mL), and the combined extracts were dried (Na2SO4) and concentrated in vacuo. The residue was then absorbed on silica gel and placed on a 20 g isolute column. Elution of the column with ethyl acetate / dichloromethane (v/v; 1 :1 ), ethyl acetate, and finally 5% methanol in ethyl acetate afforded the title compound as a colourless viscous oil (0.626 g, 72 %). 1H-NMR (500 MHz, DMSO- d6) 1.39 (s, 9H, C(CH3)3), 2.31 (t, J = 5.0 Hz, 4H) and 3.32 (br t, J = 4.6 Hz, 4H) (2 x piperazine N(CH2J2), 2.44 (s, 3H, pyridine 2-CH3), 3.46 (s, 2H, NCH2-pyridyl), 7.11 (d, J = 5.5 Hz, 1 H, pyridine 5-H), 7.17 (s, 1 H, pyridine 3-H), 8.37 (d, J = 5.0 Hz, 1 H, pyridine 6-H);
LC (Method B) - MS (ESI, m/z): Rt = 1.96 min - 292 [(M+H)+, 70%], 236 [(M-1Bu)+, 100%].
5-Bromo-4-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yI)-3-nitropyridin-2-amine
Figure imgf000038_0002
A solution of te/t-butyl 4-((2-methylpyήdin-4-yl)methyl)piperazine-1-carboxylate (0.350 g, 1.20 mmol) in dichloromethane (10 mL) and TFA (7 mL) was stirred at room temperature for 1.5 h. The solvents were then removed in vacuo, and the residue was suspended in isopropanol (18 mL). To this mixture 2-amino-5-bromo-4-chloro-3- nitropyridine (0.304 g, 1.2 mmol) was added followed by diisopropylethylamine (1.3, ml_, 7.45 mmol). The reaction mixture was stirred at 45 0C for 20 h, then the solvent was removed in vacuo and the resulting residue was absorbed on silica gel and placed on a 20 g isolute column. Elution of the column with ethyl acetate/ dichloromethane (v/v; 1 :1 ) and 2% methanol in ethyl acetate / dichloromethane (v/v; 1 :1) afforded the title compound as yellow solid (0.200 g, 41 %) after trituration with isopropanol. 1H-NMR (500 MHz, DMSO-d6) 2.45 (s, 3H, pyridine 2-CH3), 2.52 (br s- obscured by solvent peak, 4H) and 3.07 (br s, 4H) (2 x piperazine N(CH2)2), 3.52 (s, 2H, NCH2-pyridyl), 6.97 (s, 2H, NH2), 7.13 (d, J = 5.0 Hz, 1 H, 2-methylpyridine 5-H), 7.19 (S, 1H, 2-methylpyridine 3-H), 8.16 (s, 1 H, pyridine 6-H)1 8.37 (d, J = 5.0 Hz, 1 H, 2-methylpyridine 6-H);
LC (Method B) - MS (ESI, m/z): Rt = 1.91 min - 407, 409 [(M+H)+, Br isotopic pattern].
2-(4-(6-Bromo-7-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- 6]pyridin-2-yl)phenoxy)ethanol
Figure imgf000039_0001
To a mixture of 5-bromo-4-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.041 g, 0.1 mmol) and EtOH (4.0 ml_) was added 4-(2- hydroxyethoxy)benzaldehyde (0.022 g, 0.13 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.40 mL, 0.40 mmol). The reaction mixture was stirred at 80 0C for 20 h, then allowed to cool to room temperature and concentrated in vacuo. The resulting residue was absorbed on silica, and the free-running powder was placed on a 10 g isolute silica column. Elution with a gradient of methanol (0 to 8%) in ethyl acetate/dichloromethane (v:v; 3:2) afforded a pale yellow solid. This solid was triturated with diethyl ether; the precipitate was collected by filtration, washed with water (2 x 5ml_) and diethyl ether (2 x 6 mL) to obtain the title compound as a white solid (0.005 g, 10%); 1H-NMR (500 MHz, CD3OD) 2.58 (s, 3H, pyridine 2-CH3), 2.83 (br s, 4H) and 3.83 (br s, 4H) (2 x piperazine N(CHz)2). 3.76 (s, 2H, NCH2- pyridyl), 3.92 (t, J = 4. 8 Hz, 2H) and 4.15 (t, J = 4.6 Hz, 2H) (OCH2CH2OH), 7.12 (d, J = 8.8 Hz, 2H) and 8.10 (d, J= 8.8 Hz, 2H) (C6H4O), 7.38 (d, J = 4.9 Hz, 1H, 2- methylpyridine 5-H), 7.44 (S, 1 H, 2-methylpyridine 3-H), 8.21 (s, 1 H, imidazo[4,5- bjpyridine 5-H), 8.42 (d, J = 5.2 Hz, 1H, 2-methylpyridine 6-H);
LC (Method B) - MS (ESI, m/z): Rt = 2.97 min - 523, 525 [(M+H)+, Br isotopic pattern];
ESI-HRMS: Found: 523.1448, calculated for C25H28BrN6O2(Mn-H)+: 523.1452.
Example 22
2-(4-(6-Bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-3H-imidazo[4,5-/b]pyridin-2- yl)phenoxy)ethanol
Figure imgf000040_0001
To a mixture of 5-bromo-4-[4-(4-chloro-benzyl)-piperazin-1-yl]-3-nitro-pyridin-2- ylamine (0.036 g, 0.08 mmol) and EtOH (6.0 ml_) was added 4-(2- hydroxyethoxy)benzaldehyde (0.019 g, 0.11 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.40 mL, 0.40 mmol). The reaction mixture was stirred at 80 0C for 20 h, then allowed to cool to room temperature and concentrated in vacuo. The resulting residue was absorbed on silica, and the free-running powder was placed on a 10 g isolute silica column. Elution with ethyl acetate/dichloromethane (v:v; 3:7) and then a gradient of methanol (0 to 3%) in ethyl acetate/dichloromethane (v:v; 1 :1 ) afforded the title compound as a pale yellow solid after trituration with diethyl ether (0.011 g, 26%); 1H-NMR (500 MHz, DMSO-d6) 2.60 (br s, 4H) and 3.65 (br s, 4H) (2 x piperazine N(CH2J2), 3.57 (s, 2H, NCH2-C6H4CI), 3.74 (q, J = 5.2 Hz, 2H, CH2OH) and 4.07 (t, J = 4.9 Hz, 2H, OCH2CH2OH), 4.88 (t, J = 5.2 Hz, 1 H, CH2OH), 7.41 (m, 4H, C6H4CI), 7.10 (d, J = 8.8 Hz, 2H) and 8.12 (d, J = 8.7 Hz, 2H) (3,5-C6H4 and 2,6-C6H4), 8.20 (s, 1H, imidazo[4,5-b]pyridine 5-H), 13.37 (s, 1 H, imidazo[4,5-£>]pyridine N-H);
LC (Method B) - MS (ESI1 m/z): Rt = 3.63 min - 542, 544, 546 [(M+H)+, BrCI isotopic pattern]; ESI-HRMS: Found: 542.0947, calculated for C25H26BrCIN5O2 (M+H)+: 542.0953. Example 23 te/t-Butyl 4-(6-bromo-7-(4-(4-chlorobenzy|)piperazin-1-yl)-3H-imidazo[4,5-ib]pyridin-2- yl)benzyl carbamate
Figure imgf000041_0001
To a mixture of 5-bromo-4-[4-(4-chloro-benzyl)-piperazin-1-yl]-3-nitro-pyridin-2- ylamine (0.043 g, 0.10 mmol) and EtOH (6.0 ml_) was added te/t-butyl N-(4- formylbenzyl)carbamate (0.031 g, 0.13 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.40 ml_, 0.40 mmol). The reaction mixture was stirred at 80 0C for 20 h, then allowed to cool to room temperature and concentrated in vacuo. The residue was absorbed on silica gel, the free-running powder was placed on a 10 g isolute silica column, and elution with a gradient of methanol (0 to 2%) in ethyl acetate / dichloromethane (v:v; 1 :1) afforded the title compound as a pale yellow solid (0.027 g, 44%). 1H-NMR (500 MHz, DMSO-d6) 1.41 (s, 9H, OC(CHs)3), 2.60 (br s, 4H) and 3.65 (br s, 4H) (2 x piperazine N(CH2J2), 3.56 (s, 2H, NCAV2-C6H4CI), 4.18 (d, J = 5.5 Hz, 2H, C6H4CH2), 7.41 (m, 7H, C6H4CI, 2,6-C6H4, and CONH), 8.14 (d, J = 8.2 Hz, 2H) (3,5-C6H4), 8.23 (s, 1 H, imidazo[4,5-6]pyridine 5-H), 13.45 (br s, 1 H, imidazo[4,5-<b]pyridine N-H);
LC (Method B) - MS (ESI, m/z): Rt = 4.17 min - 611 , 613, 615 [(M+H)+, BrCI isotopic pattern].
(4-(6-Bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-3H-imidazo[4,5-Jb]pyridin-2- yl)phenyl)methanamine
Figure imgf000041_0002
A solution of terf-butyl 4-(6-bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-3H- imidazo[4,5-<b]pyridin-2-yl)benzylcarbamate (0.026 g, 0.043 mmol), dichloromethane (4 ml_) and TFA (0.7 ml_) was stirred at room temperature for 1 h. The solvents were then removed in vacuo, and the resulting residue was dissolved in methanol/dichloromethane and the solution was placed on a 5g isolute SCX Il column. Elution with methanol and then 0.1 M, and 0.2 M ammonia in methanol afforded the title compound as a pale yellow solid (0.02Og, 91%). 1H-NMR (500 MHz, DMSOd6) 2.61 (br s, 4H) and 3.65 (br s, 4H) (2 x piperazine N(CH2J2), 3.57 (s, 2H) and 3.79 (s, 2H) (NCH2-C6H4CI and C6H4CH2), 7.41 (m, 4H, C6H4CI), 7.49 (d, J = 8.2 Hz, 2H) and 8.12 (d, J = 8.2 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 8.22 (s, 1 H, imidazo[4,5-b]pyridine 5-H);
LC (Method B) - MS (ESI, m/z): Rt = 2.42 min - 511 , 513, 515 [(M+H)+, BrCI isotopic pattern]. ESI-HRMS: Found: 511.1007, calculated for C24H25BrCIN6 (M+H)+: 511.1007.
Example 24 te/t-Butyl 4-(4-(6-bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-3H-imidazo[4,5- <b]pyridin-2-yl)phenyl)piperazine-1-carboxylate
Figure imgf000042_0001
To a mixture of 5-bromo-4-[4-(4-chloro-benzyl)-piperazin-1-yl]-3-nitro-pyridin-2- ylamine (0.043 g, 0.10 mmol) and EtOH (6.5 ml_) was added te/t-butyl 4-(4- formylphenyl)piperazine-1-carboxylate (0.038 g, 0.13 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1M; 0.40 ml_, 0.40 mmol). The reaction mixture was stirred at 80 0C for 20 h, then allowed to cool to room temperature and concentrated in vacuo. The residue was absorbed on silica gel, the free-running powder was placed on a 10 g isolute silica column, and elution with a gradient of methanol (0 to 3%) in ethyl acetate / dichloromethane (v:v; 1 :1) afforded the title compound as a pale yellow solid (0.038 g, 57%). 1H-NMR (500 MHz, DMSO-d6) 1.43 (s, 9H, OC(CHa)3), 2.60 (br s, 4H), 3.26 (br s, obscured by water peak), 3.48 (br t, J = 5.3 Hz, 4H), and 3.64 (br s, 4H) (4 x piperazine N(CHz)2), 3.57 (s, 2H, NCH2- C6H4CI), 7.41 (m, 4H, C6H4CI), 7.07 (d, J = 8.9 Hz1 2H) and 8.04 (d, J = 8.9 Hz, 2H)
(2,6-C6H4 and 3,5-C6H4), 8.17 (s, 1H, imidazo[4,5-/b]pyridine 5-H), 13.23 (br s, 1H, imidazo[4,5-b]pyridine N-H);
LC (Method B) - MS (ESI, m/z): Rt = 4.55 min - 666, 668, 670 [(M+H)+, BrCI isotopic pattern].
6-Bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-2-(4-(piperazin-1-yl)phenyl)-3H- imidazo[4,5-£>]pyridine
Figure imgf000043_0001
A solution of te/t-butyl 4-(4-(6-bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-3/-/- imidazo[4,5-έ>]pyridin-2-yl)phenyl)piperazine-1-carboxylate (0.032 g, 0.048 mmol), dichloromethane (5 ml_) and TFA (0.7 mL) was stirred at room temperature for 1 h. The solvents were then removed in vacuo, and the resulting residue was dissolved in methanol/dichloromethane and placed on a2 g isolute SCX Il column. Elution with methanol and then 0.1 M, and 0.2M ammonia in methanol afforded the title compound as a pale yellow solid (O.OOδg, 30%) after a trituration with diethyl ether. 1H-NMR (500 MHz, DMSOd6) 2.60 (br s, 4H), 2.88 (br t, J = 4.9 Hz, 4H), 3.20 (br t, J = 5.3 Hz, 4H) and 3.64 (br s, 4H) (4 x piperazine N(CH2)2), 3.57 (s, 2H, NCH2-C6H4CI), 7.41 (m, 4H, C6H4CI), 7.04 (d, J = 8.9 Hz, 2H) and 8.02 (d, J = 8.7 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 8.16 (s, 1 H, imidazo[4,5-<b]pyridine 5-H), 13.15 (br s, imidazo[4,5- jb]pyridine N-H);
LC (Method B) - MS (ESI, m/z): Rt = 2.50 min - 566, 568, 570 [(M+H)+, BrCI isotopic pattern]. ESI-HRMS: Found: 566.1432, calculated for C27H30BrCIN7 (M+H)+: 566.1429.
Example 25
2-(4-(6-Bromo-7-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3/-/-imidazo[4,5- b]pyridin-2-yl)phenoxy)ethanol
Figure imgf000044_0001
To a mixture of 5-bromo-4-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.040 g, 0.1 mmol) and EtOH (4.0 ml_) was added 4-(2- hydroxyethoxy)benzaldehyde (0.022 g, 0.13 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.40 mL, 0.40 mmol). The reaction mixture was stirred at 80 0C for 20 h, then allowed to cool to room temperature and concentrated in vacuo. The resulting residue was absorbed on silica, and the free-running powder was placed on a 10 g isolute silica column. Elution with a gradient of methanol (0 to 5%) in ethyl acetate/dichloromethane (v:v; 1 :1) afforded the title compound as a yellow solid after trituration with diethyl ether (0.008 g, 16%); 1H-NMR (500 MHz, DMSO-d6) 2.40 (s, 3H, isoxazole 5-CH3), 2.60 (br s, 4H) and 3.64 (br s, 4H) (2 x piperazine N(CH2)2), 3.59 (s, 2H, NCH2-isoxazole), 3.74 (q, J = 5.1 Hz, 2H) and 4.07 (t, J = 5.0 Hz, 2H) (OCH2CH2OH), 4.93 (t, J = 5.5 Hz, 1 H, CH2OH), 6.26 (s, 1 H, isoxazole 4-H), 7.10 (d, J = 8.9 Hz, 2H) and 8.12 (d, J = 8.9 Hz, 2H) (3,5-C6H4 and 2,6-C6H4), 8.20 (s, 1 H, imidazo[4,5-t>]pyridine 5-H)1 13.39 (s, 1 H, imidazo[4,5- /b]pyridine N-H);
LC (Method B) - MS (ESI, m/z): Rt = 3.39 min - 513, 515 [(M+H)+, Br isotopic pattern]; ESI-HRMS: Found: 513.1242, calculated for C23H26BrN6O3 (M+H)+: 513.1244.
Example 26 te/t-butyl 4-(4-(6-Bromo-7-(4-(1-phenylethyl)piperazin-1-yl)-3H-imidazo[4,5-jb]pyridin- 2-yl)phenyl)piperazine-1-carboxylate
Figure imgf000045_0001
To a mixture of 5-bromo-3-nitro-4-[4-(1-phenyl-ethyl)-piperazin-1-yl]-pyridin-2-ylamine (prepared as described in example 70 of PCT/GB2006/004854; 0.042 g, 0.1 mmol) and EtOH (6.5 ml_) was added ferf-butyl 4-(4-formylphenyl)piperazine-1-carboxylate (0.038 g, 0.13 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.40 mL, 0.40 mmol). The reaction mixture was stirred at 80 0C for 20 h, then allowed to cool to room temperature and concentrated in vacuo. The residue was absorbed on silica gel, the free-running powder was placed on a 10 g isolute silica column, and elution with a gradient of methanol (0 to 3%) in ethyl acetate / dichloromethane (v:v; 1 :1 ) afforded the title compound as a yellow solid (0.031 g, 48%). 1H-NMR (500 MHz, DMSO-d6) 1.36 (d, J = 6.7 Hz, 3H, CHCH3), 1.43 (s, 9H, OC(CHs)3), 2.53 (m, 2H), 2.60 (m, 2H), 3.26 (br t, 4H), 3.48 (m, 5H), and 3.61 (br s, 4H) (piperazine NCH2 and CHCH3), 7.07 (d, J = 9.0 Hz, 2H) and 8.03 (d, J = 8.8 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 7.25 (m, 1 H) and 7.36 (m, 4H) (PhH), 8.15 (s, 1 H, imidazo[4,5-<fc>]pyridine 5-H), 13.23 (br s, 1 H, imidazo[4,5-ό]pyridine N-H); LC (Method B) - MS (ESI, m/z): Rt = 4.14 min - 646, 648, [(M+H)+, Br isotopic pattern],
6-Bromo-7-(4-(1-phenylethyl)piperazin-1-yl)-2-(4-(piperazin-1-yl)phenyl)-3H- imidazo[4,5-ιb]pyridine
Figure imgf000045_0002
A solution of fe/f-butyl 4-(4-(6-bromo-7-(4-(1-phenylethyl)piperazin-1-yl)-3H- imidazo[4,5-ιb]pyridin-2-yl)phenyl)piperazine-1-carboxylate (0.062 g, 0.095 mmol), dichloromethane (9 mL) and TFA (2 mL) was stirred at room temperature for 1 h and 10 min. The solvents were then removed in vacuo, and the resulting residue was dissolved in methanol/dichloromethane and placed on a 5g isolute SCX Il column. Elution with methanol and then 0.1 M, and 0.2M ammonia in methanol afforded the title compound as a pale yellow solid (0.05Og, 96%). 1.36 (d, J = 6.7 Hz, 3H, CHCW3), 2.54 (m, 2H), 2.63 (m, 2H), 2.85 (br t, J = 5.0 Hz, 4H), 3.19 (br t, J = 4.7 Hz, 4H), and 3.61 (br s, 4H) (piperazine N(CH2)), 3.50 (q, J = 6.7 Hz, 1H1 CHCH3), 7.03 (d, J = 9.0 Hz, 2H) and 8.01 (d, J = 8.8 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 7.25 (m, 1H) and 7.36 (m, 4H) (PhH), 8.15 (s, 1H, imidazo[4,5-6]pyridine 5-H); LC (Method B) - MS (ESI, m/z): Rt = 2.39 min - 546, 548 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 546.1980, calculated for C28H33BrN7 (M+H)+: 546.1975.
Example 27
2-(4-(6-bromo-7-(4-(1-phenylethyl)piperazin-1-yl)-3f/-imidazo[4,5-/?]pyridin-2- yl)phenoxy)ethanol
Figure imgf000046_0001
To a mixture of 5-bromo-3-nitro-4-[4-(1-phenyl-ethyl)-piperazin-1-yl]-pyridin-2-ylamine (0.041 g, 0.1 mmol) and EtOH (5.0 mL) was added 4-(2- hydroxyethoxy)benzaldehyde (0.022 g, 0.13 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.40 mL, 0.40 mmol). The reaction mixture was stirred at 80 0C for 20 h, then allowed to cool to room temperature and concentrated in vacuo. The resulting residue was absorbed on silica, and the free-running powder was placed on a 10 g isolute silica column. Elution with a gradient of methanol (0 to 3.5%) in ethyl acetate/dichloromethane (v:v; 1 :1) afforded the title compound as a white solid after trituration with diethyl ether (0.016 g, 30%); 1H-NMR (500 MHz, DMSO-de) 1.36 (d, J = 6.7 Hz, 3H, CHCH3), 2.55 (m, 2H), 2.65 (br s, 2H) and 3.63 (br s, 4H) (piperazine NCH2), 3.50 (q, J = 6.8 Hz, 1H, CHCH3), 3.75 (q, J = 5.2 Hz, 2H, CH2OH), 4.08 (t, J = 4.9 Hz, 2H, OCH2CH2OH), 4.88 (t, J = 5.5 Hz, 1H, CH2OH), 7.10 (d, J = 8.9 Hz, 2H) and 8.12 (d, J = 8.7 Hz, 2H) (3,5-C6H4 and 2,6-C6H4), 7.26 (m, 1H) and 7.36 (m, 4H) (PhH), 8.18 (s, 1H, imidazo[4,5-b]pyridine 5-H), 13.30 (s, 1H, imidazo[4,5-ib]pyridine N-H);
LC (Method B) - MS (ESI, m/z): Rt = 3.26 min - 522, 524 [(M+H)+, Br isotopic pattern]; ESI-HRMS: Found: 522.1496, calculated for C26H29BrN5O2 (M+H)+:
522.1499.
Example 28 tert-butyl 4-(4-(6-bromo-7-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-b]pyridin-2-yl)phenyl)piperazine-1-carboxylate
Figure imgf000047_0001
To a mixture of 5-bromo-4-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.040 g, 0.10 mmol) and EtOH (4 ml_) was added terf-butyl 4- (4-formylphenyl)piperazine-1-carboxylate (0.038 g, 0.13 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.40 ml_, 0.40 mmol). The reaction mixture was stirred at 80 0C for 20 h, then allowed to cool to room temperature and concentrated in vacuo. The residue was absorbed on silica gel, the free-running powder was placed on a 10 g isolute silica column, and elution with ethyl acetate / dichloromethane (v:v; 2:3), and 3% methanol in ethyl acetate / dichloromethane (v:v; 1 :1 ) afforded the title compound as a pale yellow solid (0.045 g, 70%). 1H-NMR (500 MHz, DMSO-d6) 1.43 (s, 9H, OC(CH3)3), 2.40 (s, 3H, isoxazole 5-CH3), 2.62 (br s, 4H), 3.26 (br t, 4H), 3.47 (br s, 4H), and 3.62 (br s, 4H) (4 x piperazine N(CH2)2), 3.59 (s, 2H, N-CH2-isoxazole), 6.26 (s, 1H, 4-H isoxazole), 7.07 (d, J = 9.0 Hz, 2H) and 8.03 (d, J = 9.3 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 8.17 (s, 1 H, imidazo[4,5-b]pyridine 5-H), 13.30 (br s, 1 H, imidazo[4,5-b]pyridine N-H);
LC (Method B) - MS (ESI, m/z): Rt = 4.68 min - 637, 639, [(M+H)+, Br isotopic pattern].
3-((4-(6-Bromo-2-(4-(piperazin-1-yl)phenyl)-3H-imidazo[4,5-ό]pyridin-7-yl)piperazin-1- yl)methyl)-5-methy!isoxazole
Figure imgf000048_0001
To a mixture of te/t-butyl 4-(4-(6-bromo-7-(4-((5-methylisoxazol-3- yl)methyl)piperazin-1-yl)-3/-/-imidazo[4,5-ib]pyridin-2-yl)phenyl)piperazine-1- carboxylate (0.032 g, 0.05 mmol) and dichloromethane (4.5 ml_) was added trifluoroacetic acid (0.6 ml_). The reaction mixture was stirred at room temperature for 1 h and 10 min, then the solvents were removed in vacuo. The resulting residue was dissolved in methanol and the solution was placed on a 5g isolute SCX Il column. Elution with methanol and then 0.1 M, and 0.3M ammonia in methanol afforded the title compound as an off-white solid (0.015g, 56%) after a trituration with diethyl ether. 1H-NMR (500 MHz, DMSOd6) 2.40 (s, 3H, isoxazole 5-CH3), 2.63 (br s, 4H), 2.85 (t, J = 4.9 Hz, 4H), 3.17 (t, J = 4.9 Hz, 4H), and 3.63 (br s, 4H) (piperazine N(CH2)2), 3.60 (s, 2H, N-CH2-isoxazole), 6.25 (s, 1 H1 4-H isoxazole), 7.04 (d, J = 9.0 Hz, 2H) and 8.02 (d, J = 8.9 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 8.17 (s, 1 H, imidazo[4,5-/?]pyridine 5-H);
LC (Method B) - MS (ESI1 m/z): Rt = 2.30 min - 537, 539 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 537.1719, calculated for C25H30BrN8O (M+H)+: 537.1720.
Example 29
6-bromo-2-(4-methoxyphenyl)-7-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-ϋ]pyridine
Figure imgf000048_0002
To a mixture of 5-bromo-4-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.041 g, 0.1 mmol) and EtOH (4.0 mL) was added 4- methoxybenzaldehyde (0.021 g, 0.16 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.40 mL, 0.40 mmol). The reaction mixture was stirred at 80 0C for 18 h, then allowed to cool to room temperature and concentrated in vacuo. The resulting residue was absorbed on silica, and the free-running powder was placed on a 10 g isolute silica column. Elution with a gradient of methanol (0 to 5%) in ethyl acetate/dichloromethane (v:v; 1 :1) afforded a white solid after trituration with diethyl ether (0.017 g, 35%); 1H-NMR (500 MHz, DMSO-d6) 2.47 (s, 3H, pyridine 2- CH3), 2.61 (br s, 4H) and 3.66 (br s, 4H) (2 x piperazine N(CH2)2), 3.56 (s, 2H, NCH2- pyridyl), 3.84 (s, 3H, OCH3), 7.10 (d, J = 8.0 Hz, 2H) and 8.11 (d, J= 8.8 Hz, 2H) (2,6- C6H4O and 3,5-C6H4O), 7.19 (d, J = 5.0 Hz, 1 H, 2-methylpyridine 5-H), 7.25 (s, 1H, 2- methylpyridine 3-H), 8.20 (s, 1 H, imidazo[4,5-6]pyridine 5-H), 8.40 (d, J = 5.0 Hz, 1H, 2-methylpyridine 6-H), 13.34 (br s, 1H, imidazo[4,5-b]pyridine N-H); LC (Method B) - MS (ESI, m/z): Rt = 3.20 min - 493, 495 [(M+H)+, Br isotopic pattern]; ESI-HRMS: Found: 493.1348, calculated for C24H26BrN6O (M+H)+: 493.1346.
Example 30 tert-Butyl 4-(4-(6-bromo-7-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-t>]pyridin-2-yl)phenyl)piperazine-1-carboxylate
Figure imgf000049_0001
To a mixture of 5-bromo-4-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.041 g, 0.1 mmol) and EtOH (4 mL) was added te/t-butyl 4-(4- formylphenyl)piperazine-1-carboxylate (0.038 g, 0.13 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1M; 0.40 mL, 0.40 mmol). The reaction mixture was stirred at 80 0C for 20 h, then allowed to cool to room temperature and concentrated in vacuo. The residue was absorbed on silica gel, the free-running powder was placed on a 10 g isolute silica column, and elution with a gradient of methanol (0 to 5%) in ethyl acetate / dichloromethane (v:v; 1 :1) afforded the title compound as a pale yellow solid (0.032 g, 49%). 1H-NMR (500 MHz, DMSOd6) 1.43 (s, 9H, C(CHs)3), 2.48 (s, 3H, pyridine 2-CH3), 2.61 (br s, 4H), 3.26 (brs, 4H), 3.48 (br t, J = 4.8 Hz, 4H), and 3.66 (br t, J = 4.4 Hz, 4H), (piperazine N(CH2J2), 3.56 (s, 2H, NCtfa-pyridyl), 7.07 (d, J = 9.0 Hz, 2H) and 8.04 (d, J= 8.9 Hz, 2H) (2,6-C6H4 and 3,5- C6H4), 7.19 (d, J = 5.2Hz, 1 H, 2-methylpyridine 5-H), 7.25 (s, 1 H, 2-methylpyridine 3- H), 8.17 (s, 1 H1 imidazo[4,5-b]pyridine 5-H), 8.40 (d, J = 5.2 Hz, 1H, 2-methylpyridine 6-H), 13.23 (br s, 1 H, imidazo[4,5-/?]pyridine N-H);
LC (Method B) - MS (ESI, m/z): Rt = 4.21 min - 647, 649 [(M+H)+, Br isotopic pattern].
6-Bromo-7-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yl)-2-(4-(piperazin-1- yl)phenyl)-3/-/-imidazo[4,5-]fc>]pyridine
Figure imgf000050_0001
To a mixture of tert-butyl 4-(4-(6-bromo-7-(4-((2-methylpyridin-4-yl)methyl)piperazin- 1-yl)-3H-imidazo[4,5-b]pyridin-2-yl)phenyl)piperazine-1-carboxylate (0.027 g, 0.04 mmol) and dichloromethane (4.5 ml_) was added trifluoroacetic acid (0.8 ml_). The reaction mixture was stirred at room temperature for 1 h and 15 min, then the solvents were removed in vacuo, the resulting residue was dissolved in methanol/dichloromethane and the solution was placed on a 5g isolute SCX Il column. Elution with methanol and then 0.1 M, and 0.3M ammonia in methanol afforded the title compound as a white solid (0.011 g, 52%) after a trituration with diethyl ether. 1H-NMR (500 MHz, DMSOd6) 1H-NMR (500 MHz, DMSO-d6) 2.43 (s, 3H, pyridine 2-CH3), 2.61 (br s, 4H), 2.85 (br t, J = 4.8 Hz, 4H), 3.19 (br t, J = 4.8 Hz, 4H), and 3.60 (br s, 4H), (piperazine N(CH2)2), 3.56 (s, 2H, NCH2-pyridyl), 7.04 (d, J = 8.9 Hz, 2H) and 8.02 (d, J= 8.7 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 7.19 (d, J = 4.7 Hz, 1 H, 2-methylpyridine 5-H)1 7.25 (s, 1 H, 2-methylpyridine 3-H), 8.17 (s, 1H, imidazo[4,5-jb]pyridine 5-H), 8.40 (d, J = 5.0 Hz, 1 H, 2-methylpyridine 6-H), 13.23 (br s, imidazo[4,5-jb]pyridine N-H);
LC (Method B) - MS (ESI, m/z): Rt = 1.99 min - 547, 549 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 547.1919, calculated for C27H32BrN8 (M+H)+: 547.1928.
Example 31 terf-Butyl 4-(6-bromo-7-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-6]pyridin-2-yl)benzylcarbamate
Figure imgf000051_0001
To a mixture of 5-bromo-4-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.060 g, 0.15 mmol) and EtOH (6.0 ml_) was added terf-butyl N- (4-formylbenzyl)carbamate (0.046 g, 0.19 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.60 ml_, 0.60 mmol). The reaction mixture was stirred at 80 0C for 19 h, then allowed to cool to room temperature and concentrated in vacuo. The residue was absorbed on silica gel, and the free-running powder was placed on a 10 g isolute silica column. Elution with a gradient of methanol (0 to 3%) in ethyl acetate / dichloromethane (v:v; 1 :1) afforded the title compound as a pale yellow solid (0.057 g, 66%). 1H-NMR (500 MHz, DMSO-d6) 1.41 (s, 9H, OC(CH3)3), 2.40 (s, 3H, isoxazole 5-CH3), 2.64 (br s, 4H) and 3.66 (br s, 4H) (piperazine N(CHz)2), 3.60 (s, 2H, N-CH2-isoxazole), 4.20 (d, J = 6.1 Hz, 2H, CH2NH), 6.25 (s, 1H, 4-H isoxazole), 7.40 (d, J = 8.2 Hz, 2H) and 8.14 (d, J = 8.3 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 7.43 (br t, J = 7.10 Hz, 1 H, CH2NH), 8.23 (s, 1H, imidazo[4,5- ύ]pyridine 5-H), 13.48 (br s, 1 H, imidazo[4,5-b]pyridine N-H); LC (Method B) - MS (ESI, m/z): Rt = 4.10 min - 582, 584 [(M+H)+, Br isotopic pattern].
(4-(6-Bromo-7-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- ύ]pyridin-2-yl)phenyl)methanamine
Figure imgf000052_0001
To a mixture of te/t-butyl 4-(6-bromo-7-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1- yl)-3H-imidazo[4,5-ύ]pyridin-2-yl)benzylcarbamate (0.053 g, 0.09 mmol) and dichloromethane (9 mL) was added trifluoroacetic acid (1.7 ml_). The reaction mixture was stirred at room temperature for 1 h and 15 min, then the solvents were removed in vacuo, the resulting residue was dissolved in methanol, and the solution was placed on a 5g isolute SCX Il column. Elution with methanol and then 0.1 M, and 0.2M ammonia in methanol afforded the title compound as a white solid (0.040g, 91%). 1H-NMR (500 MHz, DMSOd6) 2.40 (s, 3H, isoxazole 5-CH3), 2.64 (br s, 4H) and 3.66 (br t, 4H) (piperazine N(CH2J2), 3.60 (s, 2H) and 3.79 (s, 2H) (N-CH2- isoxazole and CH2NH2), 6.25 (s, 1 H, 4-H isoxazole), 7.49 (d, J = 8.3 Hz, 2H) and 8.12 (d, J = 8.2 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 8.22 (s, 1 H, imidazo[4,5-b]pyridine 5-H);
LC (Method B) - MS (ESI, m/z): Rt = 2.15 min - 482, 484 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 482.1299, calculated for C22H25BrN7O (M+H)+: 482.1298.
Example 32
1-(4-(6-bromo-7-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- b]pyridin-2-yl)phenyl)-/V,Λ/-dimethylmethanamine
Figure imgf000052_0002
To a solution of (4-(6-bromo-7-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-jb]pyridin-2-yl)phenyl)methanamine (0.035 g, 0.07 mmol) in tetrahydrofuran (2 ml_) and methanol (2 mL) was added 37% aqueous formaldehyde (17 mg, 0.21 mmol). The reaction mixture was stirred at room temperature for 5 min, then sodium cyanoborohydride (11.5 mg) was added and stirring was continued at room temperature for 4.5 h. The solvents were removed in vacuo, and the resulting residue was absorbed on silica gel and placed on a 10 g isolute column. Elution with 2% methanol in ethyl acetate / dichloromethane (v/v; 1 :1) and a gradient of methanol (2 to 10 %) in chloroform afforded a white solid. This solid was dissolved in methanol, and placed on a 5 g SCX Il column. Elution with methanol and 0.1 M ammonia in methanol afforded the title compound as a white solid (0.002 g, 6%) after trituration with diethyl ether. 1H-NMR (500 MHz, DMSOd6) 2.18 (s, 6H, NMe2), 2.42 (s, 3H, isoxazole 5-CH3), 2.65 (br s, 4H) and 3.67 (br s, 4H) (piperazine N(CH2J2), 3.46 (s, 2H) and 3.61 (s, 2H) (N-CH2-isoxazole and CH2NMe2), 6.25 (s, 1 H, 4-H isoxazole), 7.46 (d, J = 8.0 Hz, 2H) and 8.15 (d, J = 7.9 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 8.24 (s, 1 H, imidazo[4,5-fc>]pyridine 5-H), 13.50 (br s, 1H, imidazo[4,5-/b]pyridine N-H); LC (Method B) - MS (ESI, m/z): Rt = 2.16 min - 510, 512 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 510.1608, calculated for C24H29BrN7O (M+H)+: 510.1612.
Example 33
1-(4-(6-Bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-3/-/-imidazo[4,5-jb]pyridin-2- yl)phenyl)-Λ/,/V-dimethylmethanamine
Figure imgf000053_0001
To a solution of (4-(6-bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-3H-imidazo[4,5- b]pyridin-2-yl)phenyl)methanamine (0.030 g, 0.06 mmo) in tetrahydrofuran (1.7 mL) and methanol (1.7 mL) was added 37% aqueous formaldehyde (14 mg, 0.17 mmol). The reaction mixture was stirred at room temperature for 5 min, then sodium cyanoborohydride (10 mg) was added and stirring was continued at room temperature for 3 h. The solvents were removed in vacuo, and the resulting residue was dissolved in methanol / dichloromethane and placed on a 5 g SCX Il column. Elution with methanol, 0.1 M ammonia in methanol, and finally 0.2M ammonia in methanol afforded the title compound as a pale yellow solid (0.010 g, 32%) after trituration with diethyl ether. 1H-NMR (500 MHz, DMSOd6) 2.17 (s, 6H, NMe2), 2.61 (br s, 4H) and 3.67 (br s, 4H) (2 x piperazine N(CH2)2), 3.46 (s, 2H) and 3.57 (s, 2H) (NCH2-C6H4CI and C6H4CH2), 7.41 (m, 4H, C6H4CI), 7.45 (d, J = 8.1 Hz, 2H) and 8.14 (d, J = 8.1 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 8.23 (s, 1H, imidazo[4,5-<b]pyridine 5-H), 13.45 (br s, 1 H, imidazo[4,5-<b]pyridine N-H);
LC (Method B) - MS (ESI, m/z): Rt = 2.36 min - 539, 541 , 543 [(M+H)+, BrCI isotopic pattern]. ESI-HRMS: Found: 539.1320, calculated for C26H29BrCIN6 (M+H)+: 539.1320.
Example 34
2-(4-(2-Amino-5-chloro-3-nitropyridin-4-yl)piperazin-1-yl)-/V-(thiazol-2-yl)acetamide
Figure imgf000054_0001
2-(Piperazin-1-yl)-Λ/-(thiazol-2-yl)acetamide X 2HCI salt (0.209 g, 0.70 mmol) was suspended in iPrOH (12 ml_) and diisopropylethylamine (0.295 g, 2.30 mmol). To this solution, 4,5-dichloro-3-nitropyridin-2-amine (0.130 g, 0.63 mmol) was added and the mixture heated and stirred for 17 h at 45°C. The mixture was then allowed to cool to room temperature, diluted with isopropanol (10 ml_), filtered, washed with 'PrOH (3 x 3 ml_), and Et2O (2 x 5 ml_), and dried to give the title compound (0.200 g, 80%); 1H-NMR (500 MHz, DMSO-c/6): δ 2.60-2.71 (m, 4H, piperazine N(CH2J2), 3.03-3.14 (m, 4H, piperazine N(CH2J2), 3.38 (s, 2H, NCH2CO), 6.95 (brs, 2H, NH2), 7.22 (d, 1H, J = 3.5 Hz), 7.47 (d, 1 H, J = 3.5 Hz) (thiazole 4-H and thiazole 5-H), 8.07 (s, 1H, pyridine 6-H)1 11.90 (br s, 1 H, CONH); LC (Method B) - MS (ESI, m/z): Rt = 3.01 min - 398/400 [(M+H+), Cl isotopic pattern, 100%].
te/f-Butyl 4-(4-(6-chloro-7-(4-(2-oxo-2-(thiazol-2-ylamino)ethyl)piperazin-1-yl)-3H- imidazo[4,5-ιb]pyridin-2-yl)phenyl)piperazine-1-carboxylate
Figure imgf000055_0001
To a mixture of 2-[4-(2-amino-5-chloro-3-nitro-pyridin-4-yl)-piperazin-1-yl]-Λ/-thiazol-2- yl-acetamide (0.048 g, 0.12 mmol) and EtOH (5 ml_) was added tert-butyl 4-(4- formylphenyl)piperazine-1-carboxylate (0.045 g, 0.16 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.48 ml_, 0.48 mmol). The reaction mixture was stirred at 80 0C for 18 h, then allowed to cool to room temperature and concentrated in vacuo. The residue was absorbed on silica gel, the free-running powder was placed on a 10 g isolute silica column, and elution with a gradient of methanol (0 to 3%) in ethyl acetate / dichloromethane (v:v; 1 :1 ) afforded the title compound as a pale yellow solid (0.040 g, 53%).1 H-NMR (500 MHz, DMSO-d6) 1.43 (s, 9H, C(CHs)3), 2.77 (br t, 4H), 3.26 (br t obscured by water peak), 3.48 (br t, 4H), and 3.71 (br s, 4H) (piperazine N(CH2)2), 3.40 (s, 2H, NCH2CONH), 7.06 (d, J = 9.3 Hz, 2H) and 8.04 (d, J= 8.2Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 7.23 (d, J = 3.5 Hz, 1 H) and 7.49 (d, J = 3.5 Hz, 1H), (thiazole 4-H, 5-H), 8.05 (s, 1 H, imidazo[4,5-ό]pyridine 5-H)1 11.87 (br s, 1H, CONH), 13.18 (br s, 1 H, imidazo[4,5-<b]pyridine N-H); LC (Method B) - MS (ESI, m/z): Rt = 4.93 min - 638, 640 [(M+H)+, Cl isotopic pattern].
2-(4-(6-Chloro-2-(4-(piperazin-1-yl)phenyl)-3H-imidazo[4,5-b]pyridin-7-yl)piperazin-1- yl)-Λ/-(thiazol-2-yl)acetamide
Figure imgf000055_0002
To a mixture of terf-butyl 4-(4-(6-chloro-7-(4-(2-oxo-2-(thiazol-2- ylamino)ethyl)piperazin-1-yl)-3H-imidazo[4,5-ib]pyridin-2-yl)phenyl)piperazine-1- carboxylate (0.036 g, 0.06 mmol) and dichloromethane (5.0 ml_) was added trifluoroacetic acid (0.8 mL). The reaction mixture was stirred at room temperature for 1 h and 10 min, then the solvents were removed in vacuo. The resulting residue was dissolved in methanol / dichloromethane, and the solution was placed on a 5g isolute SCX Il column. Elution with methanol and then 0.1 M, and 0.3M ammonia in methanol afforded the title compound as an off-white solid (0.018g, 56%) after a trituration with diethyl ether. 1H-NMR (500 MHz, DMSO-d6) 2.77 (br t, J = 4.2 Hz, 4H), 2.84 (t, J = 5.0 Hz, 4H), 3.19 (t, J = 5.2 Hz, 4H), and 3.71 (br s, 4H) (piperazine N(CH2)2), 3.40 (s, 2H, NCH2CONH), 7.03 (d, J = 9.0 Hz, 2H) and 8.02 (d, J= 8.8 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 7.23 (d, J = 3.5 Hz, 1H) and 7.49 (d, J = 3.6 Hz, 1 H), (thiazole 4-H, 5- H), 8.05 (s, 1 H, imidazo[4,5-(b]pyridine 5-H)1 13.10 (br s, imidazo[4,5-b]pyridine N-H); LC (Method B) - MS (ESI1 m/z): Rt = 2.52 min - 538, 540 [(M+H)+, Cl isotopic pattern]. ESI-HRMS: Found: 538.1901 , calculated for C25H29CIN9OS (M+H)+: 538.1899.
Example 35 terf-Butyl 4-(6-bromo-7-(4-(1-phenylethyl)piperazin-1-yl)-3H-imidazo[4,5-d]pyridin-2- yl)benzylcarbamate
Figure imgf000056_0001
To a mixture of 5-bromo-3-nitro-4-[4-(1-phenyl-ethyl)-piperazin-1-yl]-pyridin-2-ylamine (0.057 g, 0.14 mmol) and EtOH (9.0 mL) was added fe/t-butyl /V-(4- formylbenzyl)carbamate (0.040 g, 0.17 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.56 mL, 0.56 mmol). The reaction mixture was stirred at 80 0C for 18 h, then allowed to cool to room temperature and concentrated in vacuo. The residue was absorbed on silica gel, and the free-running powder was placed on a 10 g isolute silica column. Elution with a gradient of methanol (0 to 2.5%) in ethyl acetate / dichloromethane (v:v; 1 :1 ) afforded the title compound as a yellow solid (0.036 g, 44%). 1H-NMR (500 MHz, DMSO-d6) 1.36 (d, J = 6.7 Hz, 3H CHCH3), 1.41 (s, 9H, OC(CH3)3), 2.53 (m, 2H) and 2.64 (m, 2H), 3.64 (br s, 4H) (piperazine
N(CH2)), 3.50 (q, 1 H, CHCH3), 4.20 (d, J = 5.9 Hz, 2H, CH2NH), 7.28 (m, 1H) and
7.35 (m, 4H) (PhH), 7.39 (d, J = 8.3 Hz, 2H) and 8.12 (d, J = 8.1 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 7.43 (br t, 1 H, CH2NH), 8.21 (s, 1 H, imidazo[4,5-/?]pyridine 5-H), 13.45
(br s, 1H1 imidazo[4,5-jb]pyridine N-H);
LC (Method B) - MS (ESI, m/z): Rt = 3.80 min - 591 , 593 [(M+H)+, Br isotopic pattern].
(4-(6-bromo-7-(4-(1-phenylethyl)piperazin-1-yl)-3H-imidazo[4,5-fo]pyridin-2- yl)phenyl)methanamine
Figure imgf000057_0001
To a mixture of te/t-butyl 4-(6-bromo-7-(4-(1-phenylethyl)piperazin-1-yl)-3H- imidazo[4,5-t)]pyridin-2-yl)benzylcarbamate (0.030 g, 0.05 mmol) and dichloromethane (5.0 ml_) was added trifluoroacetic acid (1.0 mL). The reaction mixture was stirred at room temperature for 1 h and 15 min, then the solvents were removed in vacuo. The resulting residue was dissolved in methanol / dichloromethane, and the solution was placed on a 2 g isolute SCX Il column. Elution with methanol and then 0.1 M ammonia in methanol afforded the title compound as an off-white solid (0.003g, 12%) after a trituration with diethyl ether. 1H-NMR (500 MHz, DMSOd6) 1.36 (d, J = 6.7 Hz, 3H CHCH5), 2.55 (m, 2H) and 2.63 (m, 2H), 3.64 (br s, 4H) (piperazine N(CH2)), 3.50 (q, J = 6.8 Hz, 1 H, CHCH3), 3.81 (s, 2H, CH2NH), 7.26 (m, 1 H) and 7.36 (m, 4H) (PhH), 7.50 (d, J = 8.3 Hz, 2H) and 8.12 (d, J = 8.3 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 8.21 (s, 1 H, imidazo[4,5-d]pyridine 5-H); LC (Method B) - MS (ESI, m/z): Rt = 2.29 min - 491 , 493 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 491.1568, calculated for C25H28BrN6 (Mn-H)+: 491.1553.
Example 36 terf-Butyl 4-(6-chloro-7-(4-(2-oxo-2-(thiazol-2-ylamino)ethyl)piperazin-1-yl)-3H- imidazo[4,5-ιb]pyridin-2-yl)benzylcarbamate
Figure imgf000058_0001
To a mixture of 2-[4-(2-amino-5-chloro-3-nitro-pyridin-4-yl)-piperazin-1-yl]-/V-thiazol-2- yl-acetamide (0.066 g, 0.17 mmol) and EtOH (7.0 ml_) was added te/t-butyl /V-(4- formylbenzyl)carbamate (0.047 g, 0.20 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.67 ml_, 0.67 mmol). The reaction mixture was stirred at 80 0C for 18 h, then allowed to cool to room temperature and concentrated in vacuo. The residue was absorbed on silica gel, and the free-running powder was placed on a 10 g isolute silica column. Elution, first with ethyl acetate / dichloromethane (v:v; 1 :1 ), and then 2% methanol in ethyl acetate / dichloromethane (v:v; 1:1) afforded the title compound as a pale yellow solid (0.053 g, 55%). 1H-NMR (500 MHz, DMSO-de) 1.41 (s, 9H, C(CH3J3), 2.78 (t, J = 4.2 Hz, 4H) and 3.75 (br s, 4H) (piperazine N(CH2J2), 3.40 (s, 2H, NCH2CONH)1 4.20 (d, J = 5.9 Hz, 2H, CH2NH), 7.23 (d, J = 3.5 Hz, 1H) and 7.49 (d, J = 3.5 Hz, 1H) (thiazole 4-H, 5-H)1 7.39 (d, J = 8.2 Hz, 2H) and 8.12 (d, J= 8.4 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 7.43 (br t, 1H, CH2NH), 8.11 (s, 1 H, imidazo[4,5-<b]pyridine 5-H), 11.92 (br s, 1 H, CONH), 13.45 (br s, 1H, imidazo[4,5-ib]pyridine N-H);
LC (Method B) - MS (ESI, m/z): Rt = 4.45 min - 583, 585 [(M+H)+, Cl isotopic pattern].
2-(4-(2-(4-(aminomethyl)phenyl)-6-chloro-3H-imidazo[4,5-t>]pyridin-7-yl)piperazin-1- yl)-Λ/-(thiazol-2-yl)acetamide
Figure imgf000058_0002
To a mixture of te/t-butyl 4-(6-chloro-7-(4-(2-oxo-2-(thiazol-2-ylamino)ethyl)piperazin- 1-yl)-3H-imidazo[4,5-b]pyridin-2-yl)benzylcarbamate (0.048 g, 0.08 mmol) and dichloromethane (8.0 mL) was added trifluoroacetic acid (1.6 mL). The reaction mixture was stirred at room temperature for 1 h, then the solvents were removed in vacuo, the resulting residue was dissolved in methanol / dichloromethane, and the solution was placed on a 5 g isolute SCX Il column. Elution with methanol and then 0.1 M, and 0.2M ammonia in methanol afforded the title compound as a pale solid (0.036g, 91 %). 1H-NMR (500 MHz, DMSOd6) 2.78 (br s, 4H) and 3.74 (br s, 4H) (piperazine N(CH2)2), 3.40 (s, 2H, NCH2CONH), 3.79 (s, 2H, CH2NH2), 7.23 (d, J = 3.5 Hz, 1 H, thiazole 4-H or 5-H), and 7.49 (m, 3H) and 8.12 (d, J = 8.3 Hz, 2H) (thiazole 4-H or 5-H, 2,6-C6H4 and 3,5-C6H4), 8.11 (s, 1H, imidazo[4,5-ιb]pyridine 5-
H);
LC (Method B) - MS (ESI, m/z): Rt = 2.42 min - 483, 485 [(M+H)+, Cl isotopic pattern]. ESI-HRMS: Found: 483.1477, calculated for C22H24CIN8OS (M+H)+:
483.1477.
Example 37
Te/t-butyl 4-(4-(6-chloro-7-(4-(4-chlorobenzyl)piperazin-1-yl)-3H-imidazo[4,5- £>]pyridin-2-yl)phenyl)piperazine-1-carboxylate
Figure imgf000059_0001
To a mixture of 5-chloro-4-(4-(4-chlorobenzyl)piperazin-1-yl)-3-nitropyridin-2-amine (prepared as described in example 37 of PCT/GB2006/004854; 0.042 g, 0.11 mmol) and EtOH (7 mL) was added te/t-butyl 4-(4-formylphenyl)piperazine-1-carboxylate (0.039 g, 0.14 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.44 mL, 0.44 mmol). The reaction mixture was stirred at 80 0C for 20 h, then allowed to cool to room temperature and concentrated in vacuo. The residue was absorbed on silica gel, and the free-running powder was placed on a 10 g isolute silica column. Elution with ethyl acetate / dichloromethane (v:v; 1 :1), and then 2.5% methanol in ethyl acetate / dichloromethane (v:v; 1 :1 ) afforded the title compound as an off-white solid (0.023 g, 34%). 1H-NMR (500 MHz, DMSO-d6) 1.43 (s, 9H, OC(CHa)3), 2.58 (br t, 4H), 3.48 (br t, 4H), and 3.67 (br s, 4H) (piperazine N(CH2)2), 3.56 (s, 2H, NCH2-C6H4CI), 7.41 (m, 4H, C6H4CI), 7.06 (d, J = 8.8 Hz1 2H) and 8.03 (d, J = 7.7 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 8.04 (s, 1 H, imidazo[4,5-ιb]pyridine 5-H), 13.18 (br s, 1H, imidazo[4,5-/b]pyridine N-H);
LC (Method B) - MS (ESI1 m/z): Rt = 4.45 min - 622, 624, 626 [(M+H)+, Cl2 isotopic pattern].
6-Chloro-7-(4-(4-chlorobenzyl)piperazin-1-yl)-2-(4-(piperazin-1-yl)phenyl)-3/-/- imidazo[4,5-ib]pyridine
Figure imgf000060_0001
A solution of tert-butyl 4-(4-(6-chloro-7-(4-(4-chlorobenzyl)piperazin-1-yl)-3H- imidazo[4,5-jb]pyhdin-2-yl)phenyl)piperazine-1-carboxylate (0.021 g, 0.03 mmol), dichloromethane (3 ml_) and TFA (0.8 ml_) was stirred at room temperature for 1 h. The solvents were then removed in vacuo, and the resulting residue was dissolved in methanol/dichloromethane and placed on a 2g isolute SCX Il column. Elution with methanol and then 0.1 M, and 0.2M ammonia in methanol afforded the title compound as a white solid (0.011g, 70%) after a trituration with diethyl ether. 1H-NMR (500 MHz, DMSOd6) 2.59 (br s, 4H), 2.84 (br t, J = 4.7 Hz, 4H), 3.19 (br t, J = 5.0 Hz, 4H) and 3.67 (br s, 4H) (4 x piperazine N(CH2)2), 3.56 (s, 2H, NCH2-C6H4CI), 7.41 (m, 4H, C6H4CI), 7.03 (d, J = 8.9 Hz, 2H) and 8.01 (d, J = 8.7 Hz, 2H) (2,6-C6H4 and 3,5- C6H4), 8.04 (s, 1 H, imidazo[4,5-b]pyridine 5-H);
LC (Method B) - MS (ESI, m/z): Rt = 2.52 min - 522, 524, 526 [(M+H)+, Cl2 isotopic pattern]. ESI-HRMS: Found: 522.1940, calculated for C27H30CI2N7 (M+H)+: 522.1934.
Example 38 tert-Butyl 4-(4-(6-chloro-7-(4-(1-phenylethyl)piperazin-1-yl)-3H-imidazo[4,5-fc>]pyridin- 2-yl)phenyl)piperazine-1-carboxylate
Figure imgf000061_0001
To a mixture of 5-chloro-3-nitro-4-(4-(1-phenylethyl)piperazin-1-yl)pyridin-2-amine (prepared as described in example 17 of PCT/GB2006/004854; 0.040 g, 0.11 mmol) and EtOH (7 mL) was added te/f-butyl 4-(4-formylphenyl)piperazine-1-carboxylate (0.039 g, 0.14 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.44 mL, 0.44 mmol). The reaction mixture was stirred at 80 0C for 20 h, then allowed to cool to room temperature and concentrated in vacuo. The residue was absorbed on silica gel, the free-running powder was placed on a 10 g isolute silica column, and elution with a gradient of methanol (0 to 2.5%) in ethyl acetate / dichloromethane (v:v; 1:1) afforded the title compound as a yellow solid (0.026 g, 39%). 1H-NMR (500 MHz, DMSO-d6) 1.35 (d, J = 6.7 Hz, 3H, CHCH3), 1.43 (s, 9H, OC(CHa)3), 2.54 (m, 2H), 2.63 (m, 2H), 3.48 (m, 5H), and 3.65 (br s, 4H) (piperazine NCH2 and CHCH3), 7.06 (d, J = 9.0 Hz, 2H) and 8.03 (m, 3H) (imidazo[4,5-ιb]pyridine 5-H1 2,6-C6H4 and 3,5-C6H4), 7.26 (m, 1H) and 7.36 (m, 4H) (PhH), 13.17 (br s, 1 H, imidazo[4,5-£>]pyridine N-H);
LC (Method B) - MS (ESI, m/z): Rt = 4.07 min - 602, 604, [(M+H)\ Cl isotopic pattern].
6-Chloro-7-(4-(1-phenylethyl)piperazin-1-yl)-2-(4-(piperazin-1-yl)phenyl)-3H- imidazo[4,5-6]pyridine
Figure imgf000061_0002
A solution of tert-butyl 4-(4-(6-chloro-7-(4-(1-phenylethyl)piperazin-1-yl)-3H- imidazo[4,5-(b]pyridin-2-yl)phenyl)piperazine-1-carboxylate (0.021 g, 0.035 mmol) in dichloromethane (3 mL) and TFA (0.7 mL) was stirred at room temperature for 1 h. The solvents were then removed in vacuo, and the resulting residue was dissolved in methanol/dichloromethane and placed on a 2g isolute SCX Il column. Elution with methanol and then 0.1 M, and 0.2M ammonia in methanol afforded the title compound as an off-white solid (0.009g, 51 %) after a trituration with diethyl ether. 1H-NMR (500 MHz1 DMSOd6) 1.36 (d, J = 6.6 Hz, 3H, CHCH3), 2.53 (m, 2H), 2.61 (m, 2H), 2.85 (t, J = 4.5 Hz, 4H), 3.19 (t, J = 4.8 Hz, 4H), and 3.65 (br s, 4H) (piperazine NCH2), 3.51 (q, J = 6.4 Hz, 1H, CHCH3), 7.03 (d, J = 8.9 Hz, 2H) and 8.00 (d, J = 8.8 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 7.25 (m, 1H) and 7.35 (m, 4H) (PhH), 8.02 (s, 1 H, imidazo[4,5-£>]pyridine 5-H);
LC (Method B) - MS (ESI, m/z): Rt = 2.33 min - 502, 504, [(M+H)+, Cl isotopic pattern]. ESI-HRMS: Found: 502.2480, calculated for C28H33CIN7 (M+H)+: 502.2480.
Example 39
3-((4-(6-Bromo-2-(4-(4-methylpiperazin-1-yl)phenyl)-3H-imidazo[4,5-b]pyridin-7- yl)piperazin-1-yl)methyl)-5-methylisoxazole
Figure imgf000062_0001
To a mixture of 5-bromo-4-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.052 g, 0.13 mmol) and EtOH (5 mL) was added 4-(4- methylpiperazino)benzaldehyde (0.032 g, 1.16 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.52 mL, 0.52 mmol). The reaction mixture was stirred at 80 0C for 18 h, then allowed to cool to room temperature and concentrated in vacuo. The residue was absorbed on silica gel, and the free-running powder was placed on a 10 g isolute silica column. Elution with ethyl acetate / chloroform (v:v; 1 :1), and then a gradient of methanol (5 to 12%) in chloroform afforded a yellow solid. This solid was triturated with ether, the yellow precipitate was collected by filtration, and washed with diethyl ether, and water. The title compound was obtained as a yellow solid (0.022 g, 31%). 1H-NMR (500 MHz, DMSO-d6) 2.23 (s, 3H, piperazine N-Me), 2.40 (s, 3H1 isoxazole 5-CH3), 2.46 (br t, J =4.8 Hz, 4H), 2.63 (br s, 4H), and 3.63 (br t, J = 4.7 Hz, 4H) (piperazine N(CH2)2), 3.60 (s, 2H, N-CH2- isoxazole), 6.25 (s, 1H, 4-H isoxazole), 7.06 (d, J = 9.0 Hz, 2H) and 8.02 (d, J = 8.9
Hz1 2H) (2,6-C6H4 and 3,5-C6H4), 8.17 (s, 1H, imidazo[4,5-ό]pyridine 5-H), 13.20 (br s,
1H, imidazo[4,5-jb]pyridine N-H);
LC (Method B) - MS (ESI, m/z): Rt = 2.33 min - 551 , 553 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 551.1870, calculated for C26H32BrN8O (M+H)+:
551.1877.
Example 40
3-((4-(6-Chloro-2-(4-methoxyphenyl)-3/-/-imidazo[4,5-/t)]pyridin-7-yl)piperazin-1- yl)methyl)-5-methylisoxazole
Figure imgf000063_0001
To a mixture of 5-chloro-4-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (prepared as described in example 80 of PCT/GB2006/004854;0.036 g, 0.1 mmol) and EtOH (5 ml) was added 4- methoxybenzaldehyde (0.020 g, 0.15 mmol), followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.40 ml, 0.40 mmol). The reaction mixture was stirred at 80 0C for 2Oh, then allowed to cool to room temperature and concentrated in vacuo. The resulting residue was absorbed on silica, and the free-running powder was placed on a 10 g isolute silica column. Elution with ethyl acetate/dichloromethane (v:v; 1 :1 ) and then 2.5% methanol in ethyl acetate/dichloromethane (v:v; 1 :1 ) afforded a yellow solid. The title compound was obtained as a pale yellow solid after trituration with diethyl ether (0.021 g, 48%); 1H-NMR (500 MHz, DMSO-d6) 2.40 (s, 3H, isoxazole 5- CH3), 2.64 (br t, 4H) and 3.68 (br s, 4H) (piperazine N(CH2)2), 3.60 (s, 2H, NCH2- isoxazole), 3.84 (s, 3H, OCH3), 6.25 (s, 1 H, isoxazole 4-H), 7.10 (d, J = 8.9 Hz, 2H) and 8.12 (d, J = 8.8 Hz, 2H) (3,5-C6H4 OMe and 2,6-C6H4OMe), 8.07 (s, 1H, imidazo[4,5-£>]pyridine 5-H), 13.30 (br s, 1 H, imidazo[4,5-jb]pyridine N-H); LC (Method B) - MS (ESI, m/z): Rt = 3.60 min - 439, 441 [(M+H)+, Cl isotopic pattern]. ESI-HRMS: Found: 439.1657, calculated for C22H24CIN6O2 (M+H)+: 439.1644.
Example 41
1-(4-(7-(4-Benzylpiperidin-1-yl)-6-bromo-3H-imidazo[4,5-ό]pyridin-2-yl)phenyl)-Λ/,Λ/- dimethylmethanamine
Figure imgf000064_0001
This was prepared using the same procedure as for 1-(4-(6-bromo-7-(4-(pyridin-3- ylmethyl)piperazin-1-yl)-3H-imidazo[4,5-b]pyridin-2-yl)phenyl)-Λ/,Λ/- dimethylmethanamine (example 61 of PCT/GB2006/004854), but here using (4-(7-(4- benzylpiperidin-1-yl)-6-bromo-3H-imidazo[4,5-ib]pyridin-2-yl)phenyl)methanamine (example 133 of PCT/GB2006/004854) (16 mg, 0.034 mmol), THF (1 ml_), MeOH (1 ml_), formaldehyde (3.0 eq, 0.10 mmol, 10.0 μl_) and NaBH3CN (3.0 eq, 0.10 mmol, 6 mg). The same purification procedure gave the product (9 mg, 53%) as a colourless solid; δH (500 MHz, DMSO-d6) 1.40-1.48 (m, 2H, 2 x piperidine CHAHB), 1.71-1.75 (m, 2H, 2 x piperidine CHAHB), 1.79-1.85 (m, 1 H, piperidine CH), 2.17 (s, 6H, N(CHa)2), 2.62 (d, J = 7.0 Hz, 2H, PhCH2), 3.30-3.32 (m, 2H, 2 x piperidine NCHAHB), 3.45 (s, 2H, CH2NMe2), 3.86 (d, br, J = 13.0 Hz, 2H, 2 x piperidine NCHAHB), 7.20 (t, J = 7.3 Hz, 1 H, phenyl H-4), 7.23 (d, J = 7.0 Hz, 2H, phenyl H-2 & H-6), 7.31 (t, J = 7.5 Hz, 2H, phenyl H-3 & H-5), 7.45 (d, J = 7.3 Hz, 2H, phenyl H-3' & H-5'), 8.12 (d, J = 8.0 Hz, 2H, phenyl H-2' & H-6'), 8.22 (s, 1H, imidazo[4,5-<b]pyridine H-5), 13.42 (s, br, 1 H, imidazo[4,5-b]pyridine NH);
LC (Method B) - MS (ESI, m/z): Rt = 4.37 min - 504, 506 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 504.1759, calculated for C27H3IBrN5 (M+H)+: 504.1757.
Example 42 1-(4-(6-Bromo-7-(4-phenoxypiperidin-1-yl)-3H-imidazo[4,5-/3]pyridin-2-yl)phenyl)-Λ/,A/- dimethylmethanamine
Figure imgf000065_0001
This was prepared using the same procedure as for 1-(4-(6-bromo-7-(4-(pyridin-3- ylmethyl)piperazin-1-yl)-3H-imidazo[4,5-ιb]pyridin-2-yl)phenyl)-Λ/,/V- dimethylmethanamine (example 61 of PCT/GB2006/004854), but here using (4-(6- bromo-7-(4-phenoxypiperidin-1-yl)-3H-imidazo[4,5-ib]pyridin-2- yl)phenyl)methanamine (example 135 of PCT/GB2006/004854) (8 mg, 0.017 mmol), THF (0.5 mL), MeOH (0.5 mL), formaldehyde (3.0 eq, 0.051 mmol, 5.0 μl_) and NaBH3CN (3.0 eq, 0.051 mmol, 3 mg). The same purification procedure gave the product (6 mg, 70%) as a colourless solid; δH (500 MHz, DMSO-d6) 1.85-1.90 (m, 2H, 2 x piperidine CHAHB), 2.18 (s, 6H, N(CH3J2), 2.50 (hidden by DMSO peak, 2H, 2 x piperidine CHA/-/B), 3.47 (s, 2H, CH2NMe2), 3.59-3.63 (m, 2H, 2 x piperidine NCHAHB), 3.88-3.91 (m, 2H, 2 x piperidine NCHAHB), 4.68-4.73 (m, 1H, piperidine CH), 6.94 (t, J = 7.3 Hz, 1H1 phenyl H-A), 7.05 (t, d = 7.6 Hz, 2H, phenyl H-2 & H-6), 7.31 (t, J = 7.3 Hz, 2H, phenyl H-3 & H-5), 7.46 (d, J = 8.0 Hz, 2H, phenyl H-3' & H- 5'), 8.14 (d, J = 8.1 Hz, 2H, phenyl H-2' & H-6'), 8.24 (s, 1H, imidazo[4,5-/b]pyridine H-5);
LC (Method B) - MS (ESI, m/z): Rt = 4.11 min - 506, 508 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 506.1553, calculated for C26H29BrN5O (M+H)+: 506.1549.
Example 43
5-Bromo-3-nitro-4-(4-(pyridin-3-yloxy)pipehdin-1-yl)pyridin-2-amine
Figure imgf000066_0001
This was prepared using the same procedure as for 2-(4-(2-amino-5-bromo-3- nitropyridin-4-yl)piperazin-1-yl)-Λ/-(thiazol-2-yl)acetamide, but here using 5-bromo-4- chloro-3-nitropyridin-2-amine (200 mg, 0.79 mmol), DIPEA (3.5 eq, 2.77 mmol, 0.48 ml_), isopropanol (4 mL) and 3-(piperidin-4-yloxy)pyridine (1.1 eq, 0.87 mmol, 154 mg). Concentration in vacuo to half volume after 6 h gave a bright yellow solid, which was filtered and washed with cold water (2 x 2 mL) to give the product (256 mg, 82%) as a yellow solid; δH (500 MHz, DMSOd6) 1.78-1.83 (m, 2H, 2 x piperidine CHAHB), 2.06-2.10 (m, 2H, 2 x piperidine CHAHB), 3.00-3.04 (m, 2H, 2 x piperidine NCHAHB), 3.25-3.29 (m, 2H, 2 x piperidine NCHAHB), 4.72 (quintet, J = 4.4 Hz, 1H, piperidine CH), 6.98 (s, br, 2H, NH2), 7.32 (ddd, J = 8.5, 4.7, 0.6 Hz, 1 H, pyridine H-5), 7.48 (ddd, J = 8.5, 3.0, 1.3 Hz, 1 H, pyridine H-4), 8.16 (dd, J = 4.6, 1.2 Hz, 1 H, pyridine H- 6), 8.17 (s, 1H, bromopyridine H-6), 8.32 (d, J = 2.8 Hz, 1H, pyridine H-2); LC (Method B) - MS (ESI, m/z): Rt = 3.90 min - 394, 396 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 394.0509, calculated for Ci5H17BrN5O3 (M+H)+: 394.0509.
ferf-Butyl 4-(6-bromo-7-(4-(pyridin-3-yloxy)piperidin-1-yl)-3H-imidazo[4,5-jb]pyridin-2- yl)benzylcarbamate
Figure imgf000066_0002
This was prepared using the same procedure as for 4-(6-bromo-2-(4- (dimethylamino)phenyl)-3H-imidazo[4,5-/?]pyridin-7-yl)-Λ/-phenylpiperazine-1- carboxamide (example 39 of PCT/GB2006/004854), but here using 5-bromo-3-nitro- 4-(4-(pyridin-3-yloxy)piperidin-1-yl)pyridin-2-amine (100 mg, 0.25 mmol), DMF (0.20 ml_), ethanol (1.20 ml_), 1 M Na2S2O4 (3 eq, 0.75 mmol, 0.75 ml_) and tert-butyl Λ/-(4- formylbenzyl)carbamate (1.1 eq, 0.28 mmol, 66 mg). After 6 h, concentration in vacuo and purification by preparative tic (CH2CI2-MeOH, 9:1 ) gave the product (31 mg, 21 %) as a colourless solid; δH (500 MHz, DMSO-d6) 1.42 (s, 9H, C(CH3J3), 1.87- 1.92 (m, 2H, 2 x piperidine CHAHB), 2.20-2.22 (m, 2H, 2 x piperidine CHAHB), 3.63 (app t, J = 9.5 Hz, 2H, 2 x piperidine NCHAHB), 3.89-3.92 (m, 2H, 2 x piperidine NCHAHB), 4.21 (d, J = 5.7 Hz, 2H, CH2-NHBOC), 4.81 (app septet, J = 4.0 Hz, 1 H, piperidine CH), 7.36 (dd, J = 8.4, 4.6 Hz, 1 H, pyridine H-5), 7.41 (d, J = 7.9 Hz, 2H, phenyl H-Z & H-5), 7.45 (t, J = 5.4 Hz, 1 H, CH2-NHBOC), 7.52-7.54 (m, 1 H, pyridine H-A), 8.14 (d, J = 7.9 Hz, 2H, phenyl H-2 & H-6), 8.19 (d, J = 4.5 Hz, 1 H, pyridine H- 6), 8.26 (s, 1H, imidazo[4,5-<b]pyridine H-5), 8.38 (d, J = 2.7 Hz, 1 H, pyridine H-2), 13.48 (s, br, 1 H, imidazo[4,5-ό]pyridine NH);
LC (Method B) - MS (ESI, m/z): Rt = 5.44 min - 579, 581 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 579.1713, calculated for C28H32BrN6O3 (M+H)+: 579.1713.
(4-(6-Bromo-7-(4-(pyridin-3-yloxy)piperidin-1-yl)-3H-imidazo[4,5-ib]pyridin-2- yl)phenyl)methanamine
Figure imgf000067_0001
This was prepared using the same procedure as for (4-(6-bromo-7-(4-(pyridin-3- ylmethyl)piperazin-1-yl)-3H-imidazo[4,5-t)]pyridin-2-yl)phenyl)methanamine (example 60 of PCT/GB2006/004854), but here using terf-butyl 4-(6-bromo-7-(4-(pyridin-3- yloxy)piperidin-1-yl)-3H-imidazo[4,5-ιb]pyridin-2-yl)benzylcarbamate (11 mg, 0.019 mmol), TFA (0.2 ml_) and CH2CI2 (1 ml_). The same purification procedure gave the desired product (8 mg, 89%) as a colourless solid; δH (500 MHz, DMSOd6) 1 -87-1.93 (m, 2H, 2 x piperidine CHAHB), 2.20-2.22 (m, 2H, 2 x piperidine CHAHB), 3.60-3.65 (m, 2H, 2 x piperidine NCHAHB), 3.82 (s, 2H, CH2NH2), 3.88-3.91 (m, 2H, 2 x piperidine NCHAHB), 4.81 (quintet, J = 3.9 Hz, 1 H, piperidine CH), 7.36 (dd, J = 8.4, 4.6 Hz, 1H, pyridine H-5), 7.51 (d, J = 8.2 Hz, 2H, phenyl H-Z & H-5), 7.52-7.54 (m, 1 H, pyridine HA), 8.14 (d, J = 8.0 Hz, 2H, phenyl H-2 & H-6), 8.19 (d, br, J = 4.6 Hz, 1 H, pyridine H-6), 8.25 (s, 1 H, imidazo[4,5-6]pyhdine H-5), 8.38 (d, J = 3.9 Hz, 1 H, pyridine H-2);
LC (Method B) - MS (ESI, m/z): Rt = 3.05 min - 479, 481 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 479.1182, calculated for C23H24BrN6O (M+H)+: 479.1189.
Example 44
1-(4-(6-Bromo-7-(4-(pyridin-3-yloxy)piperidin-1-yl)-3H-imidazo[4,5-b]pyridin-2- yl)phenyl)-Λ/,Λ/-dimethylmethanamine
Figure imgf000068_0001
This was prepared using the same procedure as for 1-(4-(6-bromo-7-(4-(pyridin-3- ylmethyl)piperazin-1-yl)-3H-imidazo[4,5-jb]pyridin-2-yl)phenyl)-/V,/V- dimethylmethanamine (example 61 of PCT/GB2006/004854), but here using (4-(6- bromo-7-(4-(pyridin-3-yloxy)piperidin-1-yl)-3H-imidazo[4,5-<b]pyridin-2- yl)phenyl)methanamine (7 mg, 0.015 mmol), THF (0.5 ml_), MeOH (0.5 ml_), formaldehyde (3.0 eq, 0.044 mmol, 4.5 μl_) and NaBH3CN (3.0 eq, 0.044 mmol, 3 mg). The same purification procedure gave the product (6 mg, 81%) as a pale yellow solid; δH (500 MHz, DMSO-d6) 1.88-1.93 (m, 2H, 2 x piperidine CHAHB), 2.21 (s, br, 6H, N(CHa)2), 2.50 (hidden by DMSO peak, 2H, 2 x piperidine CHAHB), 3.28 (s, 2H, NCH2), 3.61-3.65 (m, 2H, 2 x piperidine NCHAHB), 3.89-3.92 (m, 2H, 2 x piperidine NCHAHB), 4.79-4.82 (m, 1 H, piperidine CH), 7.36 (dd, J = 8.4, 4.6 Hz, 1 H, pyridine H- 5), 7.49-7.52 (m, 1H1 pyridine H-4), 7.53 (d, J = 8.5 Hz, 2H, phenyl H-3 & H-5), 8.17- 8.20 (m, 3H, phenyl H-2 & H-6, pyridine H-6), 8.26 (s, 1H1 imidazo[4,5-b]pyridine H-
5), 8.38 (d, J = 2.6 Hz, 1 H, pyridine H-2), 13.51 (s, br, 1 H, imidazo[4,5-Jb]pyridine
NH);
LC (Method B) - MS (ESI, m/z): Rt = 3.05 min - 507, 509 [(M+H)+, Br isotopic pattern]. ESI-HRMS: Found: 507.1497, calculated for C25H28BrN6O (M+H)+:
507.1502.
Example 45 tert-Butyl 4-(2-amino-5-cyclopropyl-3-nitropyridin-4-yl)piperazine-1-carboxylate
Figure imgf000069_0001
A solution of ferf-butyl 4-(2-amino-5-bromo-3-nitropyridin-4-yl)piperazine-1- carboxylate (prepared as rescribed in example 14 of PCT/GB2006/004854; 1 g, 2.50 mmol) in DME (2OmL) was treated with Pd(OAc)2 (5 mol %, 0.12 mmol, 30 mg), PCy3 (10 mol %, 0.25 mmol, 70 mg), cyclopropylboronic acid (3 eq, 7.51 mmol, 642 mg) and K3PO4 (3.5 eq, 8.71 mmol, 1.84 g) and stirred with microwave heating at 150 0C for 45 minutes. After this time the reaction mixture was filtered through a bed of celite, concentrated in vacuo and purified by column chromatography (hexane- EtOAc, 2:1 ) to give the product (221 mg, 25%); δH (500 MHz, DMSO-d6) 0.65-0.68 (m, 2H, 2 x cyclopropyl CHAHB), 0.88-0.92 (m, 2H, 2 x cyclopropyl CHAHB), 1.42 (s, 9H, C(CH3)3), 1.77-1.82 (m, 1 H, cyclopropyl CH), 3.01 (t, J = 4.5 Hz, 4H, piperazine N(CH2)2), 3.44 (s, br, 4H, piperazine N(CH2)2), 6.53 (s, br, 2H, NH2), 7.79 (s, 1H, pyridine H-6);
LC (Method B) - MS (ESI, m/z): Rt = 4.16 min - 364 [(M+H)+]. ESI-HRMS: Found: 364.1982, calculated for C17H26N5O4 (M+H)+: 364.1979.
terf-Butyl 4-(6-cyclopropyl-2-(4-methoxyphenyl)-3H-imidazo[4,5-ό]pyridin-7- yl)piperazine-1 -carboxylate
Figure imgf000070_0001
This was prepared using the same procedure as for 4-(6-bromo-2-(4- (dimethylamino)phenyl)-3H-imidazo[4,5-b]pyridin-7-yl)-Λ/-phenylpiperazine-1- carboxamide (example 39 of PCT/GB2006/004854), but here using terf-butyl 4-(2- amino-5-cyclopropyl-3-nitropyridin-4-yl)piperazine-1-carboxylate (82 mg, 0.22 mmol), DMF (0.15 mL), ethanol (0.85 mL), 1 M Na2S2O4 (3 eq, 0.67 mmol, 0.67 mL) and 4- methoxybenzaldehyde (1.1 eq, 0.25 mmol, 0.03 mL). After 6 h, concentration in vacuo and purification by preparative tic (CH2CI2-MeOH, 95:5) gave the product (31 mg, 31%) as a pale yellow solid; δH (500 MHz, DMSO-d6) 0.72 (app d, J = 4.4 Hz, 2 x cyclopropyl CHAHB), 0.96-1.00 (m, 2H, 2 x cyclopropyl CHAHB), 1.44 (s, 9H, C(CHa)3), 2.03-2.07 (m, 1 H, cyclopropyl CH), 3.55 (s, br, 4H, piperazine N(CH2)2), 3.61 (s, br, 4H, piperazine N(CH2J2), 3.83 (s, 3H, OCH3), 7.08 (d, J = 8.8 Hz, 2H, phenyl H-3 & H-5), 7.80 (s, 1H, imidazo[4,5-<b]pyridine H-5), 8.11 (d, J = 8.7 Hz, 2H, phenyl H-2 & H-6), 13.06 (s, br, 1H1 imidazo[4,5-£>]pyridine NH); LC (Method B) - MS (ESI, m/z): Rt = 4.69 min - 450 [(M+H)+]. ESI-HRMS: Found: 450.2500, calculated for C25H32N5O3 (M+H)+: 450.2499.
6-Cyclopropyl-2-(4-methoxyphenyl)-7-(piperazin-1-yl)-3H-imidazo[4,5-)b]pyridine
Figure imgf000070_0002
A solution of te/t-butyl 4-(6-cyclopropyl-2-(4-methoxyphenyl)-3H-imidazo[4,5- jb]pyridin-7-yl)piperazine-1-carboxylate (20 mg, 0.055 mmol) in TFA (0.3 mL) and CH2CI2 (1.5 mL) was stirred for 3h at room temperature . Concentration in vacuo and purification on a 5g SCX cartridge (eluting with MeOH, then 0.5M NH3 in MeOH) gave the product (15 mg, 97%) as a pale yellow solid; δH (500 MHz, DMSO-d6) 0.73 (app q, J = 4.4 Hz, 2H, 2 x cyclopropyl CHAHB), 0.96-1.00 (m, 2H, 2 x cyclopropyl CHAHB), 2.03-2.08 (m, 1 H, cyclopropyl CH), 2.98 (s, br, 4H1 piperazine N(CH2J2), 3.64 (s, br, 4H, piperazine N(CH2J2), 3.84 (s, 3H, OCH3), 7.09 (d, J = 8.6 Hz, 2H, methoxyphenyl H-3 & H-5), 7.78 (s, 1H, imidazo[4,5-b]pyridine H-S), 8.12 (d, J = 8.7 Hz, 2H, methoxyphenyl H-2 & H-6), 13.02 (s, br, 1 H, imidazo[4,5-ό]pyridine NH); LC (Method B) - MS (ESI1 m/z): Rt = 2.02 min - 350 [(M+H)+]. ESI-HRMS: Found: 350.1979, calculated for C20H24N5O (M+H)+: 350.1975.
6-Cyclopropyl-2-(4-methoxyphenyl)-7-(4-(pyridin-3-ylmethyl)piperazin-1-yl)-3H- imidazo[4,5-ib]pyridine
Figure imgf000071_0001
This was prepared using the same procedure as for 1-(4-(6-bromo-7-(4-(pyridin-3- ylmethy!)piperazin-1-yl)-3H-imidazo[4,5-jb]pyridin-2-yl)phenyl)-Λ/,Λ/- dimethylmethanamine (example 61 of PCT/GB2006/004854), but here using 6- cyclopropyl-2-(4-methoxyphenyl)-7-(piperazin-1-yl)-3H-imidazo[4,5-o]pyridine (15 mg, 0.043 mmol), THF (0.5 ml_), MeOH (0.5 ml_), 3-pyridinecarboxaldehyde (3.0 eq, 0.13 mmol, 14 mg) and NaBH3CN (3.0 eq, 0.13 mmol, 8 mg). Purification by preparative tic (DCM-MeOH, 95:5) gave the product (8 mg, 42%) as a colourless solid; δH (500 MHz, DMSO-d6) 0.69-0.73 (m, 2H, 2 x cyclopropyl CHAHB), 0.94-0.98 (m, 2H, 2 x cyclopropyl CHAHB), 2.01-2.06 (m, 1 H, cyclopropyl CH), 2.61 (s, br, 4H, piperazine N(CH2)2), 3.61 (s, 2H, NCH2), 3.69 (s, br, 4H, piperazine N(CH2J2), 3.83 (s, 3H, OCH3), 7.07 (d, J = 8.8 Hz, 2H, methoxyphenyl H-3 & H-5), 7.39 (dd, J = 7.7, 4.8 Hz, 1 H, pyridine H-5), 7.76 (s, 1 H, imidazo[4,5-6]pyridine H-5), 7.79 (d, br, J = 7.8 Hz, 1H, pyridine H-A), 8.10 (d, J = 8.8 Hz, 2H, methoxyphenyl H-2 & H-6), 8.49 (dd, J = 4.7, 1.5 Hz, 1 H, pyridine H-6), 8.56 (d, br, J = 1.7 Hz, 1 H, pyridine H-2), 13.00 (s, br, 1 H, imidazo[4,5-ό]pyridine NH);
LC (Method B) - MS (ESI, m/z): Rt = 2.40 min - 441 [(M+H)+]. ESI-HRMS: Found: 441.2398, calculated for C26H29N6O (M+H)+: 441.2397; Example 46
2-Chloro-Λ/-(thiazol-2-yl)acetamide
Figure imgf000072_0001
This is a known compound (P. Vicini et al., Bioorg. Med. Chem. 2006, 14, 3859- 3864), and here was prepared as follows: A solution of 2-aminothiazole (500 mg, 4.99 mmol) in CH2CI2 (20 mL) was treated with Et3N (1.1 eq, 5.49 mmol, 0.77 mL) and dropwise with chloroacetyl chloride (1.1 eq, 5.49 mmol, 0.44 mL). After 18 h, the reaction mixture was washed with water (20 mL), brine (20 mL), dried (MgSO4) and concentrated in vacuo. Filtration through a short pad of silica (hexane-EtOAc, 1 :1) gave the product (491 mg, 56%) as a pale yellow solid; δH (500 MHz, DMSO-d6) 4.38 (s, 2H, CH2), 7.27 (d, J = 3.5 Hz, 1 H, thiazole H-A or H-5), 7.50 (d, J = 3.5 Hz, 1H, thiazole H-4 or H-5), 12.41 (s, br, 1 H, NH).
5-Cyclopropyl-3-nitro-4-(piperazin-1-yl)pyridin-2-amine
Figure imgf000072_0002
A solution of te/f-butyl 4-(2-amino-5-cyclopropyl-3-nitropyridin-4-yl)piperazine-1- carboxylate (55 mg, 0.15 mmol) in TFA (0.2 mL) and CH2CI2 (1 mL) was stirred for 3h at room temperature. Concentration in vacuo and purification on a 5g SCX cartridge (eluting with MeOH, then 0.5M NH3 in MeOH) gave the product (39 mg, 97%) as a pale yellow solid; δH (500 MHz, DMSO-d6) 0.66-0.68 (m, 2H1 2 x cyclopropyl CHAHB), 0.88-0.91 (m, 1 H, 2 x cyclopropyl CHAHB), 2.58 (s, br, 1 H, cyclopropyl CH), 2.77-2.79 (m, 4H, piperazine N(CH2J2), 2.98-3.00 (m, 4H, piperazine N(CH2J2), 6.44 (s, br, 2H, NH2), 7.74 (s, 1 H, pyridine H-6); LC (Method B) - MS (ESI, m/z): Rt = 2.40 min - 264 [(M+H)*]. 2-(4-(2-Amino-5-cyclopropyl-3-nitropyridin-4-yl)piperazin-1-yl)-Λ/-(thiazol-2- yl)acetamide
Figure imgf000073_0001
A solution of 5-cyclopropyl-3-nitro-4-(piperazin-1-yl)pyridin-2-annine (36 mg, 0.14 mmol) in CH2CI2 (1 ml_) was treated with DIPEA (2.5 eq, 0.34 mmol, 0.06 mL) and 2- chloro-Λ/-(thiazol-2-yl)acetamide (1.1 eq, 0.15 mmol, 26 mg) and stirred at 40 0C for 18 h. After this time, concentration in vacuo and preparative tic purification (CH2CI2- MeOH, 95:5) gave the product (43 mg, 78%) as an orange solid; 6H (500 MHz, DMSO-de) 0.67-0.70 (m, 2H, 2 x cyclopropyl CHAHB), 0.89-0.93 (m, 2H, 2 x cyclopropyl CHAHB), 1.75-1.81 (m, 1 H, cyclopropyl CH), 2.67 (t, J = 4.2 Hz, 4H, piperazine N(CH2J2), 3.14 (t, J = 4.4 Hz, 4H, piperazine N(CH2)2), 3.36 (s, 2H, NCH2), 6.49 (s, br, 2H, NH2), 7.23 (d, J = 3.5 Hz, thiazole H-4 or H-5), 7.49 (d, J = 3.6 Hz, 1H, thiazole H-4 or H-5), 7.77 (s, 1 H, pyridine H-G), 11.87 (s, br, 1 H, NH); LC (Method B) - MS (ESI, m/z): Rt = 2.39 min - 404 [(M+H)+]. ESI-HRMS: Found: 404.1501 , calculated for C17H22N7O3S (M+H)+: 404.1499.
2-(4-(6-Cyclopropyl-2-(4-methoxyphenyl)-3H-imidazo[4,5-ib]pyridin-7-yl)piperazin-1- yl)-Λ/-(thiazol-2-yl)acetamide
Figure imgf000073_0002
This was prepared using the same procedure as for 4-(6-bromo-2-(4- (dimethylamino)phenyl)-3H-imidazo[4,5-b]pyridin-7-yl)-/V-phenylpiperazine-1- carboxamide (example 39 of PCT/GB2006/004854), but here using 2-(4-(2-amino-5- cyclopropyl-3-nitropyridin-4-yl)piperazin-1-yl)-Λ/-(thiazol-2-yl)acetamide (37 mg, 0.092 mmol), DMF (0.15 ml_), ethanol (0.85 mL), 1M Na2S2O4 (3 eq, 0.28 mmol, 0.28 mL) and 4-methoxybenzaldehyde (1.1 eq, 0.10 mmol, 14 mg). After 6 h, concentration in vacuo and purification by preparative tic (CH2CI2-MeOH, 95:5) gave the product (14 mg, 31 %) as a colourless solid; δH (500 MHz, DMSO-d6) 0.73 (app d, J = 5.3 Hz, 2H, 2 x cyclopropyl CHAHB), 0.97-1.00 (m, 2H, 2 x cyclopropyl CHAHB), 2.04-2.09 (m, 1 H, cyclopropyl CH), 2.78 (s, br, 4H, piperazine N(CH2J2), 3.41 (s, 2H, NCH2), 3.74 (s, br, 4H, piperazine N(CH2)2), 3.84 (s, 3H, OCH3), 7.09 (d, J = 8.7 Hz, 2H, methoxyphenyl H-3 & H-5), 7.25 (d, J = 3.2 Hz, 1 H, thiazole H-4 or H-5), 7.50 (d, J = 3.6 Hz, 1 H, thiazole H-4 or H-5), 7.78 (s, 1 H, imidazo[4,5-<b]pyridine H-5), 8.13 (d, J = 8.6 Hz, 2H, methoxyphenyl H-2 & H-6), 13.04 (s, br, 1H, imidazo[4,5-jb]pyridine NH); LC (Method B) - MS (ESI, m/z): Rt = 3.29 min - 490 [(M+H)+]. ESI-HRMS: Found: 490.2022, calculated for C25H28N7O2S (M+H)+: 490.2022.
Example 47 terf-Butyl 4-(2-amino-5-cyano-3-nitropyridin-4-yl)piperazine-1-carboxylate
Figure imgf000074_0001
A solution of fe/t-butyl 4-(2-amino-5-bromo-3-nitropyridin-4-y!)piperazine-1- carboxylate (1 g, 2.50 mmol) in DMF (12.5 mL) was treated with Pd2(dba)3 (0.05 eq, 0.125 mmol, 114 mg), dppf (0.10 eq, 0.25 mmol, 138 mg) and Zn(CN)2 (0.8 eq, 1.98 mmol, 234 mg) and stirred under microwave heating at 180 0C for 30 minutes. After this time, additional equal amounts of Pd2(dba)3, dppf and Zn(CN)2 were added and microwave heating was applied for a further 30 minutes at 180 0C. The solvent was then removed in vacuo and the residue partitioned between water and EtOAc. Further extraction of the aqueous layer with EtOAc (x 2) and CH2CI2 (x 2), drying of the combined extracts with Na2SO4, concentration in vacuo and column chromatography (hexane-EtOAc, 1 :1) gave recovered starting material (212 mg), and the product (121 mg, 14%) as an orange solid;
LC (Method B) - MS (ESI, m/z): Rt = 4.55 min - 249 [(M+H)+].
fert-Butyl 4-(6-cyano-2-(4-methoxyphenyl)-3H-imidazo[4,5-ιb]pyridin-7-yl)piperazine- 1-carboxylate
Figure imgf000075_0001
This was prepared using the same procedure as for 4-(6-bromo-2-(4- (dimethylamino)phenyl)-3H-imidazo[4,5-ιb]pyridin-7-yl)-Λ/-phenylpiperazine-1- carboxamide (example 39 of PCT/GB2006/004854), but here using te/t-butyl 4-(2- amino-5-cyano-3-nitropyridin-4-yl)piperazine-1-carboxylate (94 mg, 0.27 mmol), DMF (0.3 mL), ethanol (2.1 ml_), 1 M Na2S2O4 (3 eq, 0.80 mmol, 0.80 ml_) and 4- methoxybenzaldehyde (1.2 eq, 0.32 mmol, 44 mg). After 6 h, concentration in vacuo and purification by preparative tic (CH2CI2-MeOH, 98:2) gave the product (19 mg, 16%) as a colourless solid; δH (500 MHz, DMSO-d6) 1.46 (s, 9H, C(CH3J3), 3.58 (s, br, 4H, piperazine N(CH2)2), 3.85 (s, 3H, OCH3), 4.06 (t, J = 4.4 Hz, 4H, piperazine N(CH2)2), 7.11 (d, J = 8.9 Hz, 2H, methoxyphenyl H-3 & H-5), 8.13 (d, J = 8.8 Hz, 2H, methoxyphenyl H-2 & H-6), 8.30 (s, 1 H, imidazo[4,5-b]pyridine H-5), 13.52 (s, br, 1 H, imidazo[4,5-jb]pyridine NH);
LC (Method B) - MS (ESI, m/z): Rt = 5.55 min - 435 [(M+H)+]. ESI-HRMS: Found: 435.2141 , calculated for C23H27N6O3 (M+H)+: 435.2139.
2-(4-Methoxyphenyl)-7-(4-(pyridin-3~ylmethyl)piperazin-1-yl)-3H-imidazo[4,5- b]pyridine-6-carbonitrile
Figure imgf000076_0001
A solution of terf-butyl 4-(6-cyano-2-(4-methoxyphenyl)-3H-imidazo[4,5-ιb]pyridin-7- yl)piperazine-1-carboxylate (15 mg, 0.034 mmol) in CH2CI2 (1 ml_) at 0 0C was treated dropwise with TFA (0.3 ml_), warmed to room temperature and stirred for 3 h. Concentration in vacuo and rapid purification by sex cartridge (eluting with MeOH, then 0.2M NH3 in MeOH) gave the intermediate secondary amine. This was immediately re-dissolved in THF (0.5 mL) and MeOH (0.5 ml_) and treated with pyridine 3-carboxaldehyde (3 eq, 0.10 mmol, 11 mg) and, after 5 minutes, sodium cyanoborohydride (3 eq, 0.10 mmol, 6 mg). After 18 h at room temperature, concentration in vacuo and preparative tic (CH2CI2-MeOH, 9:1 ) gave the product (9 mg, 62% for two steps) as a colourless solid; δH (500 MHz, DMSO-d6) 2.63 (s, br, 4H, piperazine N(CH2J2), 3.61 (s, 2H, NCH2), 3.83 (s, 3H, OCH3), 4.10 (t, J = 4.7 Hz, 4H, piperazine N(CH2J2), 7.09 (d, J = 8.9 Hz, 2H, methoxyphenyl H-3 & H-S), 7.39 (dd, J = 7.1 , 4.4 Hz, 1 H, pyridine H-5), 7.77 (dt, J = 7.8, 2.0 Hz, 1 H, pyridine H-A), 8.10 (d, J = 8.9 Hz, 2H, methoxyphenyl H-2 & H-6), 8.26 (s, 1H, imidazo[4,5-/b]pyridine H-5), 8.50 (dd, J = 4.7, 1.5 Hz, 1H, pyridine H-6), 8.56 (s, br, 1H, pyridine H-2), 13.57 (s, br, 1 H1 imidazo[4,5-jfc>]pyridine NH);
LC (Method B) - MS (ESI, m/z): Rt = 3.06 min - 426 [(M+H)+]. ESI-HRMS: Found: 426.2042, calculated for C24H24N7O (M+H)+: 426.2037.
Example 48
1-(4-(6-Bromo-7-(4-((6-(trifluoromethyl)pyridin-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-b]pyridin-2-yl)phenyl)-Λ/,Λ/-dimethylmethanamine
Figure imgf000077_0001
This was prepared using the same procedure as for 4-(6-bromo-2-(4- (dimethylamino)phenyl)-3f/-imidazo[4,5-ib]pyridin-7-yl)-Λ/-phenylpiperazine-1- carboxamide (example 39 of PCT/GB2006/004854), but here using 5-bromo-3-nitro- 4-(4-((6-(trifluoromethyl)pyridin-3-yl)methyl)piperazin-1-yl)pyridin-2-amine (prepared as described in example 124 of PCT/GB2006/004854; 10 mg, 0.022 mmol), DMF (0.05 ml_), ethanol (0.45 mL), 1M Na2S2O4 (3 eq, 0.066 mmol, 0.066 mL) and 4- dimethylaminomethylbenzaldehyde (1.1 eq, 0.026 mmol, 4 mg). After 6 h, concentration in vacuo and purification by preparative tic (CH2CI2-MeOH, 95:5) gave the product (4 mg, 33%) as a colourless solid; δH (500 MHz, DMSO-d6) 2.17 (s, 6H, N(CHa)2), 2.64 (t, J = 4.5 Hz, 4H, piperazine N(CH2J2), 3.46 (s, 2H, CH2NMe2), 3.68 (t, J = 4.5 Hz, 4H, piperazine N(CH2J2), 3.74 (s, 2H, NCH2), 7.45 (d, J = 8.1 Hz, 2H, phenyl H-Z & H-5), 7.90 (d, J = 8.2 Hz, 1H1 pyridine H-S), 8.09 (d, br, J = 8.0 Hz, 1H, pyridine H-4), 8.13 (d, J = 8.1 Hz, 2H, phenyl H-I & H-6), 8.24 (s, 1 H, imidazo[4,5- jb]pyridine H-5), 8.77 (s, br, 1 H, pyridine H-2), 13.47 (s, br, 1 H, imidazo[4,5-jb]pyridine NH);
LC (Method B) - MS (ESI, m/z): Rt = 2.50 min - 574, 576 [(M+H)+, Br isotopic pattern. ESI-HRMS: Found: 574.1535, calculated for C26H28BrF3N7 (M+H)+: 574.1536.
Example 49 terf-Butyl 4-(4-(6-bromo-7-(4-((6-(trifluoromethyl)pyridin-3-yl)methyl)piperazin-1-yl)- 3H-imidazo[4,5-b]pyridin-2-yl)benzyl)piperazine-1-carboxylate
Figure imgf000078_0001
This was prepared using the same procedure as for 4-(6-bromo-2-(4- (dimethylamino)phenyl)-3/-/-imida2o[4,5-jb]pyridin-7-yl)-Λ/-phenylpiperazine-1- carboxamide (example 39 of PCT/GB2006/004854), but here using 5-bromo-3-nitro- 4-(4-((6-(trifluoromethyl)pyridin-3-yl)methyl)piperazin-1 -yl)pyridin-2-amine (75 mg, 0.16 nnmol), DMF (0.2 ml_), ethanol (1.3 ml_), 1 M Na2S2O4 (3 eq, 0.48 mmoi, 0.48 mL) and fe/t-butyl 4-(4-formylbenzyl)piperazine-1-carboxylate (1.2 eq, 0.20 mmol, 62 mg). After 6 h, concentration in vacuo and purification by preparative tic (CH2CI2- MeOH, 95:5) gave the product (28 mg, 24%) as a colourless solid; δH (500 MHz, DMSO-dβ) 1.40 (s, 9H, C(CH3)3), 2.35 (t, J = 5.0 Hz, 4H, piperazine N(CH2J2), 2.66 (s, br, 4H, piperazine N(CH2)2), 3.36 (s, br, 4H, piperazine N(CH2J2), 3.56 (s, 2H, NCH2), 3.69 (s, br, 4H, piperazine N(CH2J2), 3.75 (s, 2H, NCH2), 7.48 (d, J = 8.2 Hz, 2H, phenyl H-Z & H-5), 7.92 (d, br, J = 8.0 Hz, 1H, pyridine H-5), 8.10 (dd, J = 8.0, 1.3 Hz, 1H, pyridine H-A), 8.15 (d, J = 8.2 Hz, 2H, phenyl H-2 & H-Q), 8.25 (s, 1 H, imidazo[4,5-<b]pyridine H-5), 8.78 (s, br, 1H, pyridine H-2);
LC (Method B) - MS (ESI, m/z): Rt = 3.48 min - 715, 717 [(M+H)+, Br isotopic pattern. ESI-HRMS: Found: 715.2321 , calculated for C33H39BrF3N8O2 (M+H)+: 715.2326.
6-Bromo-2-(4-(piperazin-1-ylmethyl)phenyl)-7-(4-((6-(trifluoromethyl)pyridin-3- yl)methyl)piperazin-1-yl)-3H-imidazo[4,5-ib]pyridine
Figure imgf000079_0001
This was prepared using the same procedure as for (4-(6-bromo-7-(4-(pyridin-3- ylmethyl)piperazin-1-yl)-3H-imidazo[4,5-b]pyridin-2-yl)phenyl)methanamine (example 60 of PCT/GB2006/004854), but here using tert-butyl 4-(4-(6-bromo-7-(4-((6- (trifluoromethyl)pyridin-3-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5-t)]pyridin-2- yl)benzyl)piperazine-1-carboxylate (15 mg, 0.021 mmol), TFA (0.25 ml_) and CH2CI2 (1 mL). The same purification procedure gave the desired product (10 mg, 77%) as a pale yellow solid; δH (500 MHz, DMSO-d6) 2.30 (s, br, 4H, piperazine N(CH2J2), 2.64 (s, br, 4H, piperazine N(CH2J2), 2.70 (t, J = 4.6 Hz, 4H, piperazine N(CH2J2), 3.49 (s, 2H, NCH2), 3.67 (s, br, 4H, piperazine N(CH2J2), 3.73 (s, 2H, NCH2), 7.45 (d, J = 8.2 Hz, 2H, phenyl H-Z & HS), 7.91 (d, J = 8.0 Hz, 1 H, pyridine H-5), 8.08 (d, br, J = 8.2 Hz, 1 H, pyridine H-A), 8.12 (d, J = 8.2 Hz, 2H, phenyl H-2 & H-Q), 8.24 (s, 1 H, imidazo[4,5-/b]pyridine H-5), 8.77 (s, br, 1 H, pyridine H-2);
LC (Method B) - MS (ESI, m/z): Rt = 2.67 min - 615, 617 [(M+H)+, Br isotopic pattern. ESI-HRMS: Found: 615.1796, calculated for C28H31BrF3N8 (M+H)+: 615.1802.
Example 50 te/t-Butyl 4-(4-(6-bromo-7-(4-(1-(pyridin-4-yl)ethyl)piperazin-1-yl)-3H-imidazo[4,5- ib]pyridin-2-yl)benzyl)piperazine-1-carboxylate
Figure imgf000079_0002
This was prepared using the same procedure as for 4-(6-bromo-2-(4- (dimethylamino)phenyl)-3/-/-imidazo[4,5-t)]pyridin-7-yl)-A/-phenylpiperazine-1- carboxamide (example 39 of PCT/GB2006/004854), but here using 5-bromo-3-nitro- 4-(4-(1-(pyridin-4-yl)ethyl)piperazin-1-yl)pyridin-2-amine (prepared as described in example 53 of PCT/GB2006/004854 50 mg, 0.12 mmol), DMF (0.2 mL), ethanol (1.4 ml_), 1 M Na2S2O4 (3 eq, 0.37 mmol, 0.37 mL) and tert-butyl 4-(4- formylbenzyl)piperazine-1-carboxylate (1.2 eq, 0.15 mmol, 47 mg). After 6 h, concentration in vacuo and purification by preparative tic (CH2CI2-MeOH, 95:5) gave the product (32 mg, 40%) as a colourless solid; δH (500 MHz, DMSO-d6) 1.35 (d, J = 6.7 Hz, 3H, CHCH3), 1.39 (s, 9H, C(CH3J3), 2.34 (t, J = 4.8 Hz, 4H, piperazine N(CHa)2), 2.53-2.56 (m, 2H, piperazine NCH2), 2.63-2.66 (m, 2H, piperazine NCH2), 3.33 (s, br, 4H, piperazine N(CH2J2), 3.56 (s, 2H, NCH2), 3.58 (q, J = 6.8 Hz, 1H, CHCH3), 3.66 (s, br, 4H, piperazine N(CH2)2), 7.40 (d, J = 5.9 Hz, 2H, pyridine H-3 & H-5), 7.47 (d, J = 8.2 Hz, 2H, phenyl H-3 & H-5), 8.13 (d, J = 8.4 Hz, 2H, phenyl H-2 & H-6), 8.22 (s, 1 H, imidazo[4,5-/?]pyridine H-5), 8.54 (d, J = 5.8 Hz, 2H, pyridine H-2 & H-6), 13.47 (s, br, 1 H, imidazo[4,5-b]pyridine NH);
LC (Method B) - MS (ESI, m/z): Rt = 2.74 min - 661 , 663 [(M+H)+, Br isotopic pattern.
6-Bromo-2-(4-(piperazin-1 -ylmethyl)phenyl)-7-(4-(1 -(pyridin-4-yl)ethyl)piperazin-1 -yl)- 3H-imidazo[4,5-£>]pyridine
Figure imgf000080_0001
This was prepared using the same procedure as for (4-(6-bromo-7-(4-(pyridin-3- ylmethyl)piperazin-1-yl)-3H-imidazo[4,5-b]pyridin-2-yl)phenyl)methanamine (example 60 of PCT/GB2006/004854), but here using ferf-butyl 4-(4-(6-bromo-7-(4-(1-(pyridin- 4-yl)ethyl)piperazin-1-yl)-3H-imidazo[4,5-t)]pyridin-2-yl)benzyl)piperazine-1- carboxylate (18 mg, 0.027 mmol), TFA (0.2 mL) and CH2CI2 (1 mL). The same purification procedure gave the desired product (12 mg, 80%) as a yellow solid; 6H (500 MHz1 DMSO-d6) 1.35 (d, J = 6.8 Hz, 3H, CHCH3), 2.33 (s, br, 4H, piperazine N(CHz)2), 2.74 (t, J = 4.4 Hz, 4H, piperazine N(CHJO2), 3.31 (hidden by the water peak, 4H, piperazine N(CH2J2), 3.50 (s, 2H, NCH2), 3.57 (q, J = 6.7 Hz, 1 H, CHCH3), 3.64 (s, br, 4H, piperazine N(CH2)2), 7.40 (d, J = 6.0 Hz, 2H1 pyridine H-3 & H-S), 7.46 (d, J = 8.3 Hz, 2H, phenyl H-3 & H-5), 8.12 (d, J = 8.2 Hz, 2H, phenyl H-2 & H- 6), 8.22 (s, 1 H, imidazo[4,5-ό]pyridine H-5), 8.54 (d, J = 5.9 Hz, 2H, pyridine H-2 & H-
6);
LC (Method B) - MS (ESI, m/z): Rt = 2.14 min - 561 , 563 [(M+H)+, Br isotopic pattern. ESI-HRMS: Found: 561.2087, calculated for C28H34BrN8 (M+H)+: 561.2084.
Example 51 terf-Butyl 4-(4-(6-bromo-7-(4-(1-(pyridin-3-yl)ethyl)piperazin-1-yl)-3H-imidazo[4,5- ib]pyridin-2-yl)benzyl)piperazine-1-carboxylate
Figure imgf000081_0001
This was prepared using the same procedure as for 4-(6-bromo-2-(4- (dimethylamino)phenyl)-3H-imidazo[4,5-jb]pyridin-7-yl)-Λ/-phenylpiperazine-1- carboxamide (example 39 of PCT/GB2006/004854), but here using 5-bromo-3-nitro- 4-(4-(1-(pyridin-3-yl)ethyl)piperazin-1-yl)pyridin-2-amine (prepared as described in example 52 of PCT/GB2006/00485475 mg, 0.18 mmol), DMF (0.2 ml_), ethanol (1.4 ml_), 1 M Na2S2O4 (3 eq, 0.55 mmol, 0.55 ml_) and tert-butyl 4-(4- formylbenzyl)piperazine-1-carboxylate (1.2 eq, 0.22 mmol, 71 mg). After 6 h, concentration in vacuo and purification by preparative tic (CH2CI2-MeOH, 9:1 ) gave the product (17 mg, 14%) as a colourless solid; δH (500 MHz, DMSO-d6) 1.26 (d, J = 6.8 Hz, 3H, CHCH3), 1.39 (s, 9H, C(CH3)3), 2.33 (s, br, 4H, piperazine N(CH2)2), 2.58 (s, br, 2H, piperazine NCH2), 2.63 (s, br, 2H, piperazine NCH2), 3.30 (hidden by water peak, 4H, piperazine N(CH2J2), 3.55 (s, 2H, NCH2), 3.61-3.67 (m, 4H, CHCH3, piperazine N(CH2)2), 7.39 (dd, J = 7.8, 5.0 Hz, 1 H, pyridine H-5), 7.47 (d, J = 8.1 Hz, 2H, phenyl H-3 & H-5), 7.79 (d, br, J = 7.8 Hz, 1 H, pyridine H-4), 8.13 (d, J = 8.2 Hz1 2H, phenyl H-2 & H-6), 8.22 (s, 1H1 imidazo[4,5-<b]pyridine H-5), 8.48 (dd, J = 4.6, 1.5
Hz, 1 H, pyridine H-6), 8.57 (d, J = 1.7 Hz, 1 H, pyridine H-2), 13.52 (s, br, 1 H, imidazo[4,5-ib]pyridine NH);
LC (Method B) - MS (ESI, m/z): Rt = 2.63 min - 661 , 663 [(M+H)+, Br isotopic pattern. ESI-HRMS: Found: 661.2609, calculated for C33H42BrN8O2 (M+H)+:
661.2608.
6-Bromo-2-(4-(piperazin-1 -ylmethyl)phenyl)-7-(4-(1 -(pyridin-3-yl)ethyl)piperazin-1 -yl)- 3H-imidazo[4,5-/?]pyridine
Figure imgf000082_0001
This was prepared using the same procedure as for (4-(6-bromo-7-(4-(pyridin-3- ylmethyl)piperazin-1-yl)-3/-/-imidazo[4,5-fo]pyridin-2-yl)phenyl)methanamine (example 60 of PCT/GB2006/004854), but here using te/f-butyl 4-(4-(6-bromo-7-(4-(1-(pyridin- 3-yl)ethyl)piperazin-1-yl)-3H-imidazo[4,5-b]pyridin-2-yl)benzyl)piperazine-1- carboxylate (10 mg, 0.015 mmol), TFA (0.2 mL) and CH2CI2 (1 mL). The same purification procedure gave the desired product (7 mg, 83%) as a pale yellow solid; δH (500 MHz, DMSOd6) 1.40 (d, J = 6.8 Hz, 3H, CHCH3), 2.33 (s, br, 4H, piperazine N(CH2)2), 2.54-2.56 (m, 2H, piperazine NCH2), 2.62-2.66 (m, 2H, piperazine NCH2), 2.73 (t, J = 4.5 Hz, 4H, piperazine N(CH2)2), 3.50 (s, 2H, NCH2), 3.64 (q, J = 6.8 Hz, 1 H, CHCH3), 3.65 (s, br, 4H, piperazine N(CH2)2), 7.39 (dd, J = 7.7, 4.7 Hz, 1 H, pyridine H-5), 7.46 (d, J = 8.3 Hz, 2H, phenyl H-3 & H-5), 7.78 (dt, J = 7.8, 1.9 Hz, 1 H, pyridine H-4), 8.12 (d, J = 8.2 Hz, 2H, phenyl H-2 & H-6), 8.22 (s, 1H, imidazo[4,5-<b]pyridine H-5), 8.48 (dd, J = 4.8, 1.8 Hz, 1 H, pyridine H-6), 8.58 (d, J = 1.7 Hz, 1H, pyridine H-2);
LC (Method B) - MS (ESI, m/z): Rt = 1.94 min - 561 , 563 [(M+H)+, Br isotopic pattern. ESI-HRMS: Found: 561.2080, calculated for C28H34BrN8 (M+H)+: 561.2084.
Example 52 2-(4-(6-Bromo-7-(4-(pyridin-3-ylmethyl)piperazin-1-yl)-3/-/-imidazo[4,5-t»]pyridin-2- yl)phenoxy)ethanol
Figure imgf000083_0001
This was prepared using the same procedure as for 4-(6-bromo-2-(4- (dimethylamino)phenyl)-3/-/-imidazo[4,5-ιb]pyridin-7-yl)-Λ/-phenylpiperazine-1- carboxamide (example 39 of PCT/GB2006/004854), but here using 5-bromo-3-nitro- 4-(4-(pyridin-3-ylmethyl)piperazin-1-yl)pyridin-2-amine (prepared as described in example 27 of PCT/GB2006/004854; 75 mg, 0.19 mmol), DMF (0.2 ml_), ethanol (1.3 ml_), 1 M Na2S2O4 (3 eq, 0.57 mmol, 0.57 ml_) and 4-(2-hydroxyethoxy)benzaldehyde (1.1 eq, 0.21 mmol, 35 mg). After 6 h, concentration in vacuo and purification by preparative tic (CH2CI2-MeOH, 9:1 ) gave the product (27 mg, 29%) as a pale yellow solid; δH (500 MHz, DMSO-d6) 2.63 (s, br, 4H, piperazine N(CH2J2), 3.62 (s, 2H, NCH2), 3.67 (t, J = 4.4 Hz, 4H, piperazine N(CH2J2), 3.76 (q, J = 5.0 Hz, 2H, CH2OH), 4.08 (t, J = 4.9 Hz, 2H, CH2CH2OH), 4.90 (t, J = 4.5 Hz, 1 H, OH), 7.11 (d, J = 8.6 Hz, 2H, phenyl H-Z & H-5), 7.40 (dd, J = 7.7, 4.7 Hz, 1 H, pyridine H-S), 7.79 (d, br, J = 7.7 Hz, 1H, pyridine H-A), 8.13 (d, J = 8.6 Hz, 2H, phenyl H-2 & H-6), 8.21 (s, 1 H, imidazo[4,5-b]pyridine H-5), 8.50 (d, br, J = 4.5 Hz, 1 H, pyridine H-6), 8.58 (s, br, 1 H, pyridine H-2), 13.36 (s, br, 1 H, imidazo[4,5-ιb]pyridine NH);
LC (Method B) - MS (ESI, m/z): Rt = 2.78 min - 509, 511 [(M+H)+, Br isotopic pattern. ESI-HRMS: Found: 509.1293, calculated for C24H26BrN6O2 (M+H)+: 509.1295.
Example 53 tert-Butyl 4-(4-(6-bromo-7-(4-((2~methylpyridin-4-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-jb]pyridin-2-yl)benzyl)piperazine-1-carboxylate
Figure imgf000084_0001
This was prepared using the same procedure as for 4-(6-bromo-2-(4- (dimethylamino)phenyl)-3H-imidazo[4,5-t»]pyridin-7-yl)-A/-phenylpiperazine-1- carboxamide (example 39 of PCT/GB2006/004854), but here using 5-bromo-4-(4-((2- methylpyridin-4-yl)methyl)piperazin-1-yl)-3-nitropyridin-2-amine (50 mg, 0.12 mmol), DMF (0.15 mL), ethanol (0.85 mL), 1 M Na2S2O4 (3 eq, 0.37 mmol, 0.37 mL) and tert- butyl 4-(4-formylbenzyl)piperazine-1-carboxylate (1.1 eq, 0.14 mmol, 41 mg). After 6 h, concentration in vacuo and purification by preparative tic (CH2CI2-MeOH, 95:5) gave the product (18 mg, 22%) as a pale yellow solid; δH (500 MHz, DMSO-d6) 1.39 (s, 9H, C(CHa)3), 2.34 (t, J = 5.0 Hz, 4H, piperazine N(CH2)2), 2.48 (s, 3H, CH3), 2.62 (s, br, 4H, piperazine N(C/-/2)2), 3.30 (hidden by water peak, 4H, piperazine N(CH2)2), 3.55 (s, 2H, NCH2), 3.57 (s, 2H, NCH2), 3.68 (s, br, 4H, piperazine N(CH2)2), 7.19 (d, br, J = 5.0 Hz, 1 H, methyl pyridine H-5), 7.25 (s, br, 1 H, methylpyridine H-3), 7.46 (d, J = 8.1 Hz, 2H, phenyl H-3 & H-5), 8.14 (d, J = 8.1 Hz, 2H, phenyl H-2 & H-6), 8.22 (s, 1H, imidazo[4,5-b]pyridine H-5), 8.39 (d, J = 5.0 Hz, 1 H, methylpyridine H-6); LC (Method B) - MS (ESl1 m/z): Rt = 2.75 min - 661 , 663 [(M+H)+, Br isotopic pattern. ESI-HRMS: Found: 661.2608, calculated for C33H42BrN8O2 (M+H)+: 661.2609.
6-Bromo-7-(4-((2-methylpyhdin-4-yl)methyl)piperazin-1-yl)-2-(4-(piperazin-1- ylmethyl)phenyl)-3H-imidazo[4,5-ιb]pyridine
Figure imgf000084_0002
This was prepared using the same procedure as for (4-(6-bromo-7-(4-(pyridin-3- ylmethyl)piperazin-1-yl)-3H-imidazo[4,5-ib]pyridin-2-yl)phenyl)methanamine (example 60 of PCT/GB2006/004854), but here using te/t-butyl 4-(4-(6-bromo-7-(4-((2- methylpyridin-4-yl)methyl)piperazin-1-yl)-3/-/-imidazo[4,5-/b]pyridin-2- yl)benzyl)piperazine-1-carboxylate (10 mg, 0.015 mmol), TFA (0.25 ml_) and CH2CI2 (1 ml_). The same purification procedure gave the desired product (8 mg, 94%) as a pale yellow solid; δH (500 MHz, DMSOd6) 2.48 (s, 3H, CH3), 2.63 (s, br, 4H, piperazine N(CH2)2), 3.58 (s, 2H, NCH2), 3.69 (t, J = 4.4 Hz, 4H, piperazine N(CH2J2), 3.82 (s, 2H, NCH2), 7.21 (d, J = 5.0 Hz, 1 H, methyl pyridine H-S), 7.26 (s, br, 1 H, methylpyridine H-3), 7.51 (d, J = 8.2 Hz, 2H, phenyl H-Z & H-S), 8.14 (d, J = 8.2 Hz, 2H, phenyl H-I & H-6), 8.24 (s, 1 H, imidazo[4,5-<b]pyridine HS), 8.41 (d, J = 5.0 Hz, 1 H, methylpyridine H-6);
LC (Method B) - MS (ESI, m/z): Rt = 2.00 min - 561 , 563 [(M+H)+, Br isotopic pattern. ESI-HRMS: Found: 561.2077, calculated for C28H34BrN8 (M+H)+: 561.2084;
Example 54 fe/t-Butyl 4-(6-bromo-7-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-<fc>]pyridin-2-yl)benzyl carbamate JML 1217-14-1
Figure imgf000085_0001
This was prepared using the same procedure as for 4-(6-bromo-2-(4- (dimethylamino)phenyl)-3H-imidazo[4,5-t)]pyridin-7-yl)-Λ/-phenylpiperazine-1- carboxamide (example 39 of PCT/GB2006/004854), but here using 5-bromo-4-(4-((2- methylpyridin-4-yl)methyl)piperazin-1-yl)-3-nitropyridin-2-amine (100 mg, 0.24 mmol), DMF (0.3 mL), ethanol (1.7 mL), 1 M Na2S2O4 (3 eq, 0.74 mmol, 0.74 mL) and tert- butyl Λ/-(4-formylbenzyl)carbamate (1.1 eq, 0.27 mmol, 64 mg). After 6 h, concentration in vacuo and purification by preparative tic (CH2CI2-MeOH, 95:5) gave the product (25 mg, 17%) as a pale yellow solid; δH (500 MHz, DMSO-d6) 1.41 (s, 9H, C(CHs)3), 2.63 (s, br, 4H, piperazine N(CH2)2), 3.57 (s, 2H, NCH2), 3.68 (s, br, 4H, piperazine N(CH2)2), 4.19 (d, J = 4.8 Hz, 2H, CH2NHBOC), 7.19 (d, J = 4.8 Hz, 1 H, methylpyridine H-5), 7.25 (s, br, 1 H, methylpyridine H-3), 7.39 (d, J = 8.1 Hz, 2H, phenyl H-3 & H-5), 7.43 (t, br, J = 5.0 Hz, 1 H, CH2NHBOC), 8.12 (d, J = 7.9 Hz, 2H, phenyl H-2 & H-6), 8.23 (s, 1H, imidazo[4,5-jb]pyridine H-5), 8.40 (d, J = 5.0 Hz, 1 H, methylpyridine H-6), 13.45 (s, br, 1 H, imidazo[4,5-ϋ]pyridine NH); LC (Method B) - MS (ESI, m/z): Rt = 3.82 min - 592, 594 [(M+H)+, Br isotopic pattern. ESI-HRMS: Found: 592.2033, calculated for C29H35BrN7O2 (M+H)+: 592.2030.
Example 55
(4-(6-Bromo-7-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- /b]pyridin-2-yl)phenyl)methanamine
Figure imgf000086_0001
This was prepared using the same procedure as for (4-(6-bromo-7-(4-(pyridin-3- ylmethyl)piperazin-1-yl)-3H-imidazo[4,5-ifc)]pyridin-2-yl)phenyl)methanamine (example 60 of PCT/GB2006/004854), but here using ferf-butyl 4-(6-bromo-7-(4-((2- methylpyridin-4-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5-£>]pyridin-2- yl)benzylcarbamate (15 mg, 0.025 mmol), TFA (0.25 mL) and CH2CI2 (1 ml_). The same purification procedure gave the desired product (11 mg, 88%) as a pale yellow solid; LC (Method B) - MS (ESI, m/z): Rt = 1.88 min - 492, 494 [(M+H)+, Br isotopic pattern. ESI-HRMS: Found: 492.1509, calculated for C24H27BrN7 (M+H)+: 492.1506.
Example 56
1-(4-(6-Bromo-7-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- fo]pyridin-2-yl)phenyl)-Λ/,Λ/-dimethylmethanamine
Figure imgf000087_0001
This was prepared using the same procedure as for 1-(4-(6-bromo-7-(4-(pyridin-3- ylmethyl)piperazin-1-yl)-3H-imidazo[4,5-b]pyridin-2-yl)phenyl)-Λ/,/\/- dimethylmethanamine (example 61 of PCT/GB2006/004854), but here using (4-(6- bromo-7-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yl)-3/-/-imidazo[4,5-jb]pyridin-2- yl)phenyl)methanamine (10 mg, 0.024 mmol), THF (0.5 mL), MeOH (0.5 ml_), formaldehyde (6.0 eq, 0.12 mmol, 10.0 μl_) and NaBH3CN (6.0 eq, 0.12 mmol, 8 mg). The same purification procedure gave the product (6 mg, 57%) as a colourless solid; δH (500 MHz, DMSO-d6) 2.17 (s, 6H, N(CH3J2), 2.48 (s, 3H, CH3), 2.63-2.68 (m, 4H, piperazine N(CH2)2), 3.45 (s, 2H, NCH2), 3.57 (s, 2H, NCH2), 3.69 (s, br, 4H, piperazine N(CH2)2), 7.19 (d, br, J = 5.4 Hz, 1 H, methylpyridine H-5), 7.25 (s, br, 1 H, methylpyridine H-3), 7.45 (d, J = 8.2 Hz, 2H, phenyl H-3 & H-5), 8.14 (d, J = 8.3 Hz, 2H, phenyl H-2 & H-6), 8.23 (s, 1 H, imidazo[4,5-b]pyridine H-5), 8.40 (d, J = 5.2 Hz, 1 H, methylpyridine H-6);
LC (Method B) - MS (ESI, m/z): Rt = 2.13 min - 520, 522 [(M+H)+, Br isotopic pattern. ESI-HRMS: Found: 520.1816, calculated for C26H31BrN7 (M+H)+: 520.1819.
Example 57
2-(4-(6-Bromo-7-(4-((6-(trifluoromethyl)pyridin-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-b]pyridin-2-yl)phenoxy)ethanol
Figure imgf000087_0002
This was prepared using the same procedure as for 4-(6-bromo-2-(4- (dimethylamino)phenyl)-3H-imidazo[4,5-(b]pyridin-7-yl)-/V-phenylpiperazine-1- carboxamide (example 39 of PCT/GB2006/004854), but here using 5-bromo-3-nitro- 4-(4-((6-(trif!uoromethyl)pyridin-3-yl)methyl)piperazin-1 -yl)pyridin-2-amine (50 mg, 0.11 mmol), DMF (0.15 ml_), ethanol (0.85 ml_), 1 M Na2S2O4 (3 eq, 0.32 mmol, 0.32 ml_) and 4-(2-hydroxyethoxy)benzaldehyde (1.1 eq, 0.12 mmol, 20 mg). After 6 h, concentration in vacuo and purification by preparative tic (CH2CI2-MeOH, 95:5) gave the product (21 mg, 34%) as a colourless solid; δH (500 MHz, DMSO-d6) 2.63 (s, br, 4H, piperazine N(CH2)2), 3.66 (s, br, 4H, piperazine N(CH2)2), 3.73 (s, 2H, NCH2), 3.74 (q, br, J = 5.3 Hz, 2H, CH2OH), 4.06 (t, J = 5.0 Hz, 2H, CH2CH2OH), 4.93 (t, br, J = 5.5 Hz, 1 H, OH), 7.09 (d, J = 8.9 Hz, 2H, phenyl H-3 & H-5), 7.92 (d, J = 8.0 Hz, 1 H, pyridine H-5), 8.09 (dd, J = 8.1 , 1.2 Hz, 1 H, pyridine H-4), 8.12 (d, J = 8.8 Hz, 2H, phenyl H-2 & H-6), 8.21 (s, 1 H, imidazo[4,5-5]pyridine H-5), 8.77 (s, br, 1 H1 pyridine H-2), 13.39 (s, br, 1 H, imidazo[4,5-jb]pyridine NH);
LC (Method B) - MS (ESI, m/z): Rt = 3.85 min - 577, 579 [(M+H)+, Br isotopic pattern. ESI-HRMS: Found: 577.1164, calculated for C25H25BrF3N6O2 (M+H)+: 577.1169.
Example 58 te/f-Butyl 4-(4-(6-bromo-7-(4-((6-(trifluoromethyl)pyridin-3-yl)methyl)piperazin-1-yl)- 3H-imidazo[4,5-ib]pyridin-2-yl)phenyl)piperazine-1-carboxylate
Figure imgf000088_0001
This was prepared using the same procedure as for 4-(6-bromo-2-(4- (dimethylamino)phenyl)-3H-imidazo[4,5-/?]pyridin-7-yl)-Λ/-phenylpiperazine-1- carboxamide (example 39 of PCT/GB2006/004854), but here using 5-bromo-3-nitro- 4-(4-((6-(trifluoromethyl)pyridin-3-yl)methyl)piperazin-1-yl)pyridin-2-amine (75 mg, 0.16 mmol), DMF (0.2 mL), ethanol (1.3 ml_), 1 M Na2S2O4 (3 eq, 0.48 mmol, 0.48 ml_) and 4-(4-formylphenyl)piperazine-1-carboxylic acid tert-butyl ester (1.1 eq, 0.18 mmol, 52 mg). After 6 h, concentration in vacuo and purification by preparative tic (CH2CI2-MeOH, 95:5) gave the product (25 mg, 22%) as a colourless solid; δH (500 MHz, DMSO-d6) 1.42 (s, 9H, C(CH3J3), 2.63 (s, br, 4H, piperazine N(CH2J2), 3.26 (t, J = 4.4 Hz, 4H, piperazine N(CH2J2), 3.47 (s, br, 4H, piperazine N(CH2)2), 3.65 (s, br, 4H, piperazine N(CH2J2), 3.73 (s, 2H, NCH2), 7.07 (d, J = 9.0 Hz, 2H, phenyl H-3 & H- 5), 7.91 (d, J = 8.0 Hz, 1H, pyridine H-5), 8.04 (d, J = 8.8 Hz, 2H, phenyl H-2 & H-Q), 8.08 (dd, J = 8.1 , 1.4 Hz, 1 H, pyridine H-A), 8.18 (s, 1 H, imidazo[4,5-b]pyridine H-5), 8.77 (s, br, 1H, pyridine H-2), 13.28 (s, br, 1H, imidazo[4,5-6]pyridine NH); LC (Method B) - MS (ESI, m/z): Rt = 5.05 min - 701 , 703 [(M+H)+, Br isotopic pattern. ESI-HRMS: Found: 701.2167, calculated for C32H37BrF3N8O2 (M+H)+: 701.2170.
6-Bromo-2-(4-(piperazin-1-yl)phenyl)-7-(4-((6-(trifluoromethyl)pyridin-3- yl)methyl)piperazin-1-yl)-3H-imidazo[4,5-jb]pyridine
Figure imgf000089_0001
This was prepared using the same procedure as for (4-(6-bromo-7-(4-(pyridin-3- ylmethyl)piperazin-1-yl)-3H-imidazo[4,5-ib]pyridin-2-yl)phenyl)methanamine (example 60 of PCT/GB2006/004854), but here using tert-butyl 4-(4-(6-bromo-7-(4-((6- (trifluoromethyl)pyridin-3-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5-d]pyridin-2- yl)phenyl)piperazine-1-carboxylate (18 mg, 0.026 mmol), TFA (0.25 ml.) and CH2CI2 (1 mL). The same purification procedure gave the desired product (11 mg, 71 %) as a yellow solid; δH (500 MHz, DMSO-d6) 2.64 (s, br, 4H, piperazine N(CH2J2), 2.85 (s, br, 4H, piperazine N(CH2)2), 3.19 (s, br, 4H, piperazine N(CH2J2), 3.67 (s, br, 4H, piperazine N(CH2J2), 3.74 (s, 2H, NCH2), 7.04 (d, J = 8.4 Hz, 2H, phenyl H-3 & H-5), 7.92 (d, J = 8.0 Hz, 1H, pyridine H-5), 8.03 (d, J = 8.3 Hz, 2H, phenyl H-2 & H-Q), 8.09 (d, br, J = 7.8 Hz, 1 H, pyridine H-A), 8.18 (s, 1H, imidazo[4,5-b]pyridine H-5), 8.78 (s, br, 1 H, pyridine H-2), 13.23 (s, br, 1 H, imidazo[4,5-/b]pyridine NH); LC (Method B) - MS (ESI1 m/z): Rt = 2.65 min - 601 , 603 [(M+H)+, Br isotopic pattern. ESI-HRMS: Found: 601.1644, calculated for C27H29BrF3N8 (M+H)+: 601.1645.
Example 59 te/t-Butyl 4-(6-chloro-7-(4-(pyridin-3-ylmethyl)piperazin-1-yl)-3H-imidazo[4,5- ib]pyridin-2-yl)benzylcarbamate
Figure imgf000090_0001
This was prepared using the same procedure as for 4-(6-bromo-2-(4- (dimethylamino)phenyl)-3/-/-imidazo[4,5-b]pyridin-7-yl)-/V-phenylpiperazine-1- carboxamide (example 39 of PCT/GB2006/004854), but here using 5-chloro-3-nitro- 4-(4-(pyridin-3-ylmethyl)piperazin-1-yl)pyridin-2-amine (prepared as described in example 79 of PCT/GB2006/004854; 37 mg, 0.106 mmol), DMF (0.2 mL), ethanol (1.0 mL), 1 M Na2S2O4 (3 eq, 0.32 mmol, 0.32 mL) and te/t-butyl Λ/-(4- formylbenzyl)carbamate (1.1 eq, 0.1 17 mmol, 27 mg). After 6 h, concentration in vacuo and purification by preparative tic (CH2CI2-MeOH, 9:1 ) gave the product (18 mg, 32%) as a colourless solid; LC (Method B) - MS (ESI1 m/z): Rt = 3.53 min - 534, 536 [(M+H)+, Cl isotopic pattern. ESI-HRMS: Found: 534.2382, calculated for C28H33CIN7O2 (M+H)+: 534.2384.
(4-(6-Chloro-7-(4-(pyridin-3-ylmethyl)piperazin-1-yl)-3H-imidazo[4,5-ib]pyhdin-2- yl)phenyl)methanamine
Figure imgf000091_0001
This was prepared using the same procedure as for (4-(6-bromo-7-(4-(pyridin-3- ylmethyl)piperazin-1-yl)-3/-/-imidazo[4,5-i5]pyridin-2-yl)phenyi)methanamine (example 60 of PCT/GB2006/004854, but here using terf-butyl 4-(6-chloro-7-(4-(pyridin-3- ylmethyl)piperazin-1-yl)-3H-imidazo[4,5-b]pyridin-2-yl)benzylcarbamate (13 mg, 0.024 mmol), TFA (0.2 mL) and CH2CI2 (1 ml_). The same purification procedure gave the desired product (9 mg, 85%) as a pale yellow solid; LC (Method B) - MS (ESI, /77/z): Rt = 1.75 min - 434, 436 [(M+H)+, Cl isotopic pattern. ESI-HRMS: Found: 434.1870, calculated for C23H25CIN7 (M+H)+: 434.1860.
Example 60
1-(4-(6-Chloro-7-(4-(pyridin-3-ylmethyl)piperazin-1-yl)-3/-/-imidazo[4,5-jfcι]pyridin-2- yl)phenyl)-Λ/,Λ/-dimethylmethanamine
Figure imgf000091_0002
This was prepared using the same procedure as for 1-(4-(6-bromo-7-(4-(pyridin-3- ylmethyl)piperazin-1-yl)-3H-imidazo[4,5-6]pyridin-2-yl)phenyl)-Λ/,Λ/- dimethylmethanamine (example 61 of PCT/GB2006/004854), but here using (4-(6- chloro-7-(4-(pyridin-3-ylmethyl)piperazin-1-yl)-3/-/-imidazo[4,5-ib]pyridin-2- yl)phenyl)methanamine (8 mg, 0.018mmol), THF (0.3 mL), MeOH (0.3 mL), formaldehyde (3.0 eq, 0.054 mmol, 4.5 μL) and NaBH3CN (3 eq, 0.054 mmol, 3.5 mg). The same purification procedure gave the product (5 mg, 59%) as a colourless solid; δH (500 MHz, DMSO-Cl6) 2.17 (s, 6H, N(CHs)2), 2.62 (t, J = 4.4 Hz, 4H, piperazine N(CH2)2), 3.45 (s, 2H, NCH2), 3.61 (s, 2H, NCH2), 3.70 (s, br, 4H, piperazine N(CH2J2), 7.39 (dd, J = 7.7, 4.8 Hz, 1H, pyridine H-S), 7.44 (d, J = 8.2 Hz, 2H, phenyl H-Z & /7-5), 7.78 (dt, J = 7.8, 1.8 Hz, 1 H, pyridine H-A), 8.10 (s, 1 H, imidazo[4,5-/b]pyridine H-5), 8.13 (d, J = 8.2 Hz1 2H, phenyl H-2 & H-6), 8.48 (dd, J = 4.8, 1.6 Hz, 1 H, pyridine H-6), 8.56 (d, J = 1.5 Hz, 1 H, pyridine H-2), 13.43 (s, br, 1 H, imidazo[4,5-jfc>]pyridine NH);
LC (Method B) - MS (ESI, m/z): Rt = 1.75 min - 462, 464 [(M+H)+, Cl isotopic pattern. ESI-HRMS: Found: 462.2171 , calculated for C25H29CIN7 (M+H)+: 462.2168.
Example 61 ferf-butyl 4-(4-(6-chloro-7-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-ifc>]pyridin-2-yl)phenyl)piperazine-1-carboxylate
Figure imgf000092_0001
To a mixture of 5-chloro-4-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.06 g, 0.17 mmol) and EtOH (5 mL) was added te/t-butyl 4-(4- formylphenyl)piperazine-1-carboxylate (0.064 g, 0.22 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.7 mL, 0.7 mmol). The reaction mixture was stirred at 8O0C for 18 h, then allowed to cool to room temperature and concentrated in vacuo. The residue was purified by flash column chromatography (elution with ethyl acetate / hexane (v:v; 7 : 3) followed by 100 % ethyl acetate and then 5% methanol in ethyl acetate to provide 62 mg of the title compound as a yellow powder ( 62 %). 1H-NMR (500 MHz, DMSO-d6) 1.43 (s, 9H, piperazine N-Boc), 2.40 (s, 3H, isoxazole 5-CH3), 2.63 (br s, 4H), 3.26 (br t, J = 4.8 Hz, 4H) , 3.48 (br, t, J = 4.7 Hz, 4 H), 3.59 (s, 2H, N-CH2-isoxazole), 3.67 (br, s, 4 H), 6.25 (s, 1H, 4-H isoxazole), 7.06 (d, J = 9.5 Hz, 2H) and 8.02 (d, J = 8.9 Hz, 2H) (2,6-C6H4 and 3,5- C6H4), 8.05 (s, 1 H, imidazo[4,5-6]pyridine 5-H)1 13.19 (br s, 1 H, imidazo[4,5- £>]pyridine N-H); LC (method B) - MS (ESI1 m/z): Rt = 3.55 min, - 537, 539 [(M- 1Bu)+, Cl isotopic pattern].
3-((4-(6-Chloro-2-(4-piperazin-1-yl)phenyl)-3H-imidazo[4,5-fc>]pyridin-7-yl)piperazin-1- yl)methyl)-5-methylisoxazole
Figure imgf000093_0001
A solution of terf-butyl 4-(4-(6-chloro-7-(4-((5-methylisoxazol-3-yl)piperazin-1-yl)-3H- imidazo[4,5-b]pyridine-2-yl)phenyl)piperazine-1-carboxylate (0.042 g, 0.07mmol), dichloromethane (3 ml_) and TFA (3 ml_) was stirred at room temperature for 0.5 h. Volatiles were then removed in vacuo, and the resulting residue was dissolved in methanol and placed on a 2g isolute SCX Il column. Elution with methanol and then 0.1 M, and 0.2M ammonia in methanol afforded the title compound as a yellow powder (0.028g, 81 %) . 1H-NMR (500 MHz, DMSO-d6), 2.40 (s, 3H, isoxazole 5- CH3), 2.63 (br t, J 4.2 Hz1 4H), 2.85 (br t, J = 4.7 Hz, 4H), 3.20 (br, t, J - 5.1 Hz, 4H), 3.59 (s, 2H, N-CH2-isoxazole), 3.67 (br, s, 4 H), 6.24 (s, 1 H, 4-H isoxazole), 7.04 (d, J = 9.0 Hz, 2H) and 8.02 (d, J = 9.0 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 8.04 (s, 1 H, imidazo[4,5-ιbjpyridine 5-H), 13.19 (br s, 1H, imidazo[4,5-b]pyridine N-H); LC (Method B) - MS (ESI, m/z): Rt = 2.10 min, - 493, 495 [(M + H)+, Cl isotopic pattern], ESI-HRMS: Found 493.2224, calculated for C25H30CIN8O (M + H)+ : 493.2225
Example 62 te/t-Butyl 4-(6-chloro-7-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-/b]pyridine-2-yl)benzylcarbamate
Figure imgf000094_0001
To a mixture of 5-chloro-4-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3- nitropyridin-2-amine (0.06 g, 0.17 mmol) and EtOH (5 mL) was added terf-butyl Λ/-(4- formylbenzyl)carbamate (0.064 g, 0.22 mmol) followed by a freshly prepared aqueous solution of Na2S2O4 (1 M; 0.7 mL, 0.7 mmol). The reaction mixture was stirred at 8O0C for 18 h, then allowed to cool to room temperature and concentrated in vacuo. The residue was purified by flash column chromatography (elution with ethyl acetate / hexane (v:v; 7 : 3) followed by 100 % ethyl acetate and then 5% methanol in ethyl acetate to provide 60 mg of the title compound as a yellow powder (66 %). 1H-NMR (500 MHz, DMSO-d6) 1.41 (s, 9H, piperazine N-Boc), 2.40 (s, 3H, isoxazole 5-CH3), 2.64 (br, s, 4 H), 3.62 (s, 2 H, CH2isoxazole), 3.71 (br, s, 4 H), 4.20 (d, J = 5.9 Hz, 2 H, CH2NHBoC), 6.25 (s, 1 H, 4-H isoxazole), 7.38 (d, J = 8.2 Hz, 2H) 7.43 (br, t, J = 5.9 Hz, 1 H, NHBoc), 8.10 (d, J = 8.2 Hz, 2H) (2,6-C6H4 and 3,5-C6H4), 8.12 (s, 1 H, imidazo[4,5-<b]pyridine 5-H), 13.43 (br s, 1 H, imidazo[4,5- ib]pyridine N-H);
LC (Method B) - MS (ESI, m/z): Rt = 3.27 min, - 538, 540 [(M + H)+, Cl isotopic pattern].
(4-(6-Chloro-7-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- b]pyridin-2-yl)phenyl)methanamine
Figure imgf000095_0001
A solution of te/t-butyl 4-(6-chloro-7-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)- 3H-imidazo[4,5-ιt>]pyridine-2-yl)benzylcarbamate (0.04 g, 0.07 mmol), dichloromethane (3 ml_) and TFA (3 mL) was stirred at room temperature for 0.5 h. Volatiles were then removed in vacuo, and the resulting residue was dissolved in methanol and placed on a 2g isolute SCX Il column. Elution with methanol and then 0.1 M, and 0.2M ammonia in methanol afforded the title compound as a yellow powder (0.03g, 92.5 %) . 1H-NMR (500 MHz, DMSO-d6), 2.40 (s, 3H, isoxazole 5- CH3), 2.64 (br t, J = 5 Hz, 4H), 3.70 (br, t, J = 5 Hz, 4H), 3.60 (s, 2 H) and 3.82 (s, 2H) (CH2NH2 and CH2-isoxazole), 6.25 (s, 1H, 4-H isoxazole), 7.50 (d, J = 8.3 Hz, 2H, 2,6-C6H4 Or S1S-C6H4), 8.10 (s, 1 H, imidazo[4,5-/b]pyridine 5-H), 8.12 (d, J = 8.3 Hz, 2H, 2,6-C6H4 or 3,5-C6H4). LC (Method B) - MS (ESI, m/z): Rt = 2.10 min, - 438, 440 [(M + H)+, Cl isotopic pattern]. ESI-HRMS: Found 438.1802, calculated for C22H25CIN7O (M + H)+ : 438.1803.
Biological Results
The compounds of the above examples were tested in the assay described below and shown to have Aurora-A inhibitory activity.
Filterplate assay for identification and evaluation of Aurora-A inhibitors
40 μl master mix of kinase buffer (50 mM Tris pH 7.5, 10 mM NaCI, 2.5 mM MgCL2, 1 mM DTT, 20 μM ATP, 0.025 μCi/μl 33P-ATP, and 100 μg/ml MBP was added to a 96- well plate followed by addition of 250 ng of Aurora-A enzyme per well. The plate was shaken for approximately 2 min on a flat-bed plate shaker (Wellmix, Denley, UK) and incubated for 2 hours at room temperature. The reaction was stopped by the addition of 30 μl of 2% orthophosphoric acid. The reaction mixture was then transferred and filtered on the 96-well Multi-Screening Filter Plate (MATAH0P00, MILLIPORE), pre- treated with 50 μl per well of 0.5% orthophosphoric acid. The plate was washed twice with 200 μl of 0.5% orthophosphoric acid and vacuum-dried. 25μl of scintillant (Microscint™20, PerkinElmer) per well was added and the plate was shaken for 10 min. Finally, the plate was re-sealed with TopSealA and the signal from the filter- bound component was read on TopCount-NXT™ (PerkinElmer Life Sciences UK Ltd., Hounslow, UK).
Cell viability assay: Determination of IC50 of Aurora inhibitors in HCT116 cell line using MTT assay.
The effects of compounds of the Examples on cellular proliferation were determined using the MTT assay according to manufacturer's instructions (Sigma). Briefly, the human colon tumour cells HCT116, were seeded in triplicate into 96-well plates at 2500 cells/well 24 hours before treatment with a range of concentrations of Aurora inhibitors (0 - 50 μM). After 72 hrs, 15 μl/well of 0.5% 3-(4,5-dimethylthiazol-2-yl)-2,5- diphenyltetrazolium bromide (MTT) was added to the cells for 4 hrs at 370C, the dye- stained viable cells were extracted by adding 150 μl/well of dimethyl sulphoxide (DMSO). The optical density was measured at 570 nm using the Wallac VICTOR2 TM 1420 Multilabel Counter (PerkinElmer). The IC50 was calculated using the Prism software.
IC50 results were allocated to one of two ranges as follows:
Range A: IC50 < 200 nM for Aurora A inhibition, and IC50 < 2000 nM for HCT116 cell growth inhibition. Range B: IC50 >200 nM for Aurora A inhibition, and IC50 >2000 nM for HCT116 cell growth inhibition.
The Table below summarises the results of testing compounds of the invention in the Filterplate Aurora A enzyme inhibiton assay, and in the cell viability assay described above:
Results Table
Figure imgf000096_0001
Figure imgf000097_0001
Figure imgf000098_0001
For example, the compounds of examples 7, 24, 25, 26, 28, 33, 34 had IC50S < 60 nM in the enzyme assay, and < 600 nM in the cellular assay.
The legend "Not Tested" in the above Table means the compound in question had not yet been tested in the relevant assay at the date of filing the present application. However, such compounds are confidently expected to have activity in the relevant assays in view of their close structural similarity to other active compounds of the invention.

Claims

Claims:
A compound selected from the group consisting of:
e-Bromo-y^^cyclobutylmethyOpiperazin-i-yl^^^piperazin-i-yOphenyO-S/V- imidazo[4,5-ό]pyridine,
6-Chloro-7-(4-(cyclobutylmethyl)piperazin-1-yl)-2-(4-(piperazin-1-ylmethyl)phenyl)- 3H-imidazo[4,5-£>]pyridine,
6-Bromo-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-2-(4-(piperazin-1- ylmethyl)phenyl)-3H-imidazo[4,5-b]pyridine,
2-(4-(6-Bromo-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- ib]pyridin-2-yl)phenoxy)ethanol,
2-(4-(6-bromo-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- /b]pyridin-2-yl)phenoxy)-Λ/,/V-dimethylethanamine,
tenf-Butyl-4-(6-bromo-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-b]pyridin-2-yl)benzylcarbamate,
6-Bromo-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-2-(4-(piperazin-1- yl)phenyl)-3H-imidazo[4,5-<b]pyridine,
6-Chloro-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-2-(4-(piperazin-1- yl)phenyl)-3H-imidazo[4,5-jb]pyridine,
(4-(6-Chloro-7-(4-((5-fluoropyridin-3-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- b]pyridin-2-yl)phenyl)methanamine,
4-(6-Chloro-7-(4-((6-(trifluoromethyl)pyridin-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-/3]pyridin-2-yl)phenyl)methanamine,
6-Chloro-2-(4-(piperazin-1-yl)phenyl)-7-(4-((6-(trifluoromethyl)pyridin-3- yl)methyl)piperazin-1-yl)-3AV-imidazo[4,5-ifc)]pyridine, 2-(4-(6-Bromo-7-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3/-/-imidazo[4,5- b/pyridin-2-yl)phenoxy)ethano),
2-(4-(6-Bromo-7-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- b]pyridin-2-yl)phenoxy)-/V,/V-dimethylethanamine,
1-(4-(6-Bromo-7-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3/-/-imidazo[4,5- b]pyridin-2-yl)phenyl)pipetϊdin-4-ol,
4-(5-(6-Bromo-7-(4-((2-methylthiazol-4-yl)methyl)piperazin-1-yl)-3f/-imidazo[4,5- jb]pyridin-2-yl)pyridin-2-yl)morpholine,
4-((4-(6-Bromo-2-(6-(piperazin-1-yl)pyridin-3-yl)-3H-imidazo[4,5-fe]pyridin-7- yl)piperazin-1-yl)methyl)-2-methylthiazole,
4-((4-(6-Chloro-2-(4-(piperazin-1-yl)phenyl)-3/V-imidazo[4,5-ib]pyridin-7-yl)piperazin-1- yl)methyl)-2-methylthiazole,
4-(3-(6-Bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-3H-imidazo[4,5-/3]pyridin-2- yl)benzyl)morpholine,
6-Bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-2-(4-(piperazin-1-ylmethyl)phenyl)-3/-/- imidazo[4,5-ib]pyridine,
3-((4-(6-Bromo-2-(4-(piperazin-1-ylmethyl)phenyl)-3H-imidazo[4,5-^]pyridin-7- yl)piperazin-1-yl)methyl)-5-methylisoxazole,
2-(4-(6-Bromo-7-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- b]pyridin-2-yl)phenoxy)ethanol,
2-(4-(6-Bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-3H-imidazo[4,5-/b]pyridin-2- yl )phenoxy)ethanol ,
(4-(6-Bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-3H-imidazo[4,5-ib]pyridin-2- yl)phenyl)methanamine, 6-Bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-2-(4-(piperazin-1-yl)phenyl)-3f/- imidazo[4,5-ifc>]pyιϊdine,
2-(4-(6-Bromo-7-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- jfc>]pyridin-2-yl)phenoxy)ethanol,
6-Bromo-7-(4-(1-phenylethyl)piperazin-1-yl)-2-(4-(piperazin-1-yl)phenyl)-3H- imidazo[4,5-<b]pyridine,
2-(4-(6-bromo-7-(4-(1-phenylethyl)piperazin-1-yl)-3H-imidazo[4,5-it)]pyridin-2- yl)phenoxy)ethanol,
3-((4-(6-Bromo-2-(4-(piperazin-1-yl)phenyl)-3/-/-imidazo[4,5-/?]pyridin-7-yl)piperazin-1- yl)methyl)-5-methylisoxazole,
6-bromo-2-(4-methoxyphenyl)-7-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yl)-3/-/- imidazo[4,5-ib]pyridine,
6-Bromo-7-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yl)-2-(4-(piperazin-1- yl)phenyl)-3H-imidazo[4,5-b]pyridine,
(4-(6-Bromo-7-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3/-/-imidazo[4,5- o]pyridin-2-yl)phenyl)methanamine,
1-(4-(6-bromo-7-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- b]pyridin-2-yl)phenyl)-Λ/,/V-dimethylnnethanamine,
1-(4-(6-Bromo-7-(4-(4-chlorobenzyl)piperazin-1-yl)-3H-imidazo[4,5-ό]pyridin-2- yl)phenyl)-/V,/V-dimethylmethanamine,
2-(4-(6-Chloro-2-(4-(piperazin-1-yl)phenyl)-3H-imidazo[4,5-jb]pyridin-7-yl)piperazin-1- yl)-Λ/-(thiazol-2-yl)acetamide,
(4-(6-bromo-7-(4-(1-phenylethyl)piperazin-1-yl)-3/-/-imidazo[4,5-/?]pyridin-2- yl)phenyl)methanamine, 2-(4-(2-(4-(aminomethyl)phenyl)-6-chloro-3H-imidazo[4,5-b]pyridin-7-yl)piperazin-1- yl)-Λ/-(thiazol-2-yl)acetamide,
6-Chloro-7-(4-(4-chlorobenzyl)piperazin-1-yl)-2-(4-(piperazin-1-yl)phenyl)-3/-/- imidazo[4,5-ib]pyridine,
θ-Chloro-Z^^I-phenylethyOpiperazin-i-yl^^^piperazin-i-yOphenyO-S/V- imidazo[4,5-ib]pyridine,
3-((4-(6-Bromo-2-(4-(4-methylpiperazin-1-yl)phenyl)-3H-imidazo[4,5-/3]pyridin-7- yl)piperazin-1-yl)methyl)-5-methylisoxazole,
3-((4-(6-Chloro-2-(4-methoxyphenyl)-3H-imidazo[4,5-jb]pyridin-7-yl)piperazin-1- yl)methyl)-5-methylisoxazole,
1-(4-(7-(4-Benzylpiperidin-1-yl)-6-bromo-3H-imidazo[4,5-/3]pyridin-2-yI)phenyl)-Λ/,Λ/- dimethylmethanamine,
1-(4-(6-Bronno-7-(4-phenoxypiperidin-1-yl)-3/-/-imidazo[4,5-ib]pyridin-2-yl)phenyl)-Λ/>/V- dimethylmethanamine,
(4-(6-Bromo-7-(4-(pyridin-3-yloxy)piperidin-1-yl)-3H-imidazo[4,5-d]pyridin-2- yi)phenyl)methanamine,
1-(4-(6-Bromo-7-(4-(pyridin-3-yloxy)piperidin-1-yl)-3/-/-imidazo[4,5-ifc>]pyridin-2- yl)phenyl)-/V,Λ/-dimethylmethanamine,
6-Cyclopropyl-2-(4-methoxyphenyl)-7-(4-(pyridin-3-ylmethyl)piperazin-1-yl)-3/-/- imidazo[4,5-ib]pyridine,
2-(4-(6-Cyclopropyl-2-(4-methoxyphenyl)-3H-imidazo[4,5-ib]pyridin-7-yl)piperazin-1- yl)-Λ/-(thiazol-2-yl)acetamide,
2-(4-Methoxyphenyl)-7-(4-(pyridin-3-ylmethyl)piperazin-1-yl)-3H-imidazo[4,5- ό]pyridine-6-carbonithle, 1-(4-(6-Bromo-7-(4-((6-(trifluoromethyl)pyridin-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-t)]pyridin-2-yl)phenyl)-Λ/,/V-climethylnnethanamine,
6-Bromo-2-(4-(piperazin-1-ylmethyl)phenyl)-7-(4-((6-(trifluoromethyl)pyridin-3- yl)methyl)piperazin-1-yl)-3H-imidazo[4,5-jfc>]pyridine,
6-Bromo-2-(4-(piperazin-1 -ylmethyl)phenyl)-7-(4-(1 -(pyridin-4-yl)ethyl)piperazin-1 -yl)- 3H-imidazo[4,5-ifc>]pyridine,
6-Bromo-2-(4-(piperazin-1-ylmethyl)phenyl)-7-(4-(1-(pyridin-3-yl)ethyl)piperazin-1-yl)- 3H-imidazo[4,5-jb]pyridine,
2-(4-(6-Bromo-7-(4-(pyridin-3-ylmethyl)piperazin-1-yl)-3/-/-imidazo[4,5-d]pyridin-2- yl)phenoxy)ethanol,
6-Bromo-7-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yl)-2-(4-(piperazin-1- ylmethyl)phenyl)-3/-/-imidazo[4,5-b]pyridine,
(4-(6-Bromo-7-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- b]pyridin-2-yl)phenyl)methanamine,
1-(4-(6-Bromo-7-(4-((2-methylpyridin-4-yl)methyl)piperazin-1-yl)-3/-/-imidazo[4,5- b]pyridin-2-yl)phenyl)-Λ/,/V-dimethylmethanamine,
2-(4-(6-Bromo-7-(4-((6-(trifluoromethyl)pyridin-3-yl)methyl)piperazin-1-yl)-3H- imidazo[4,5-ib]pyridin-2-yl)phenoxy)ethanol,
6-Bromo-2-(4-(piperazin-1-yl)phenyl)-7-(4-((6-(trifluoromethyl)pyridin-3- yl)methyl)piperazin-1-yl)-3H-imidazo[4,5-ib]pyridine,
(4-(6-Chloro-7-(4-(pyridin-3-ylmethyl)piperazin-1-yl)-3f/-imidazo[4,5-ib]pyridin-2- yl)phenyl)methanamine,
1-(4-(6-Chloro-7-(4-(pyridin-3-ylmethyl)piperazin-1-yl)-3/-/-imidazo[4,5-/3]pyridin-2- yl)phenyl)-Λ/,Λ/-dimethylmethanamine, 3-((4-(6-Chloro-2-(4-piperazin-1-yl)phenyl)-3H-imidazo[4,5-ib]pyriciin-7-yl)piperazin-1- yl)methyl)-5-methylisoxazole,
(4-(6-Chloro-7-(4-((5-methylisoxazol-3-yl)methyl)piperazin-1-yl)-3H-imidazo[4,5- b]pyridin-2-yl)phenyl)methanamine,
and salts, hydrates, solvates, and N-oxides thereof.
2. A pharmaceutical composition comprising a compound as claimed in claim 1 , together with a pharmaceutically acceptable carrier.
3. The use of a compound as claimed in claim 1 in the preparation of a composition for inhibiting the activity of an aurora kinase enzyme.
4. The use as claimed in claim 3 for the inhibition of aurora-A activity, ex vivo or in vivo.
5. A method of inhibiting the activity of an aurora kinase enzyme comprising contacting the enzyme with an amount of a compound as claimed in claim 1 effective for such inhibition.
6. A method as claimed in claim 5 for the inhibition of aurora-A activity, ex vivo or in vivo.
7. The use of a compound as claimed in claim 1 in the preparation of a composition for the treatment of cell-proliferation disease or autoimmune disease,
8. A method for the treatment of cell-proliferation disease or autoimmune disease, which comprises administering to a subject suffering such disease an effective amount of a compound as claimed in claim 1.
9. The use as claimed in claim 7 or a method as claimed in claim 8 for the treatment of cancer cell proliferation.
10. The use as claimed in claim 7 or a method as claimed in claim 8 for the treatment of rheumatoid arthritis.
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