Novel compounds for tau imaging

Abstract

The present disclosure provides novel compounds of formula (I), wherein R 1 , R 2 , R 3 , R 4 , and R 5 as described herein], methods for their preparation, pharmaceutical compositions thereof, tau imaging formulations, and methods of using the compounds for tau imaging. [Formula 1] TIFF2026504445000092.tif20128

Description

[Technical Field]

[0001] The present invention relates to novel compounds, intermediates for the preparation of these compounds, methods of using these compounds for tau imaging, compositions and formulations of these compounds for diagnostic imaging, and imaging methods using these compounds, compositions, and formulations. [Background technology]

[0002] Alzheimer's disease (AD), the leading cause of dementia, affects 1% of the population between the ages of 65 and 69, increasing to 40% to 50% in those aged 95 and older. Patients with AD exhibit clear clinical symptoms, including cognitive impairment and memory deficits. In these patients, the presence of AD is confirmed by the presence of severe senile plaque burden and neurofibrillary tangles (NFTs) in the cerebral cortex during postmortem histopathological examination. Mature senile plaques are composed of extracellular β-amyloid peptides derived from enzymatic processing of amyloid precursor protein and intracellular neurofibrillary tangles (NFTs) derived from filaments of hyperphosphorylated tau protein. Hyperphosphorylated tau aggregates, such as neurofibrillary tangles, are associated with the degree of cognitive impairment in Alzheimer's disease. In AD and various other tauopathies, tau aggregates appear in specific brain regions and patterns associated with disease risk, onset, and / or progression, and these regions and patterns are known to those skilled in the art. In AD patients, tau-containing tangles first appear in brain regions closely associated with memory, and pathological studies have shown that tangles may correlate more strongly with cognitive performance than plaques. Signals generated by tau imaging agents in these regions and patterns can be used by those skilled in the art to better monitor and diagnose the risk, onset, and progression of specific disease states (see Correlation of Alzheimer's disease neuropathologic changes with cognitive status: a review of the literature. Nelson PT, et al., J Neuropathol Exp Neurol. 2012 May;71(5):362-81).Therefore, there is a need for simple, non-invasive methods to detect and / or quantitate tau deposits in patients (see M. Maruyama et al., "Imaging of tau pathology in a tauopathy mouse model and in Alzheimer patients compared to normal controls," Neuron, 79:1094-1108, 2013; C. Mathis and W. Klunk, "Imaging Tau Deposits In Vivo: Progress in Viewing More of The Proteopathy Picture," Neuron, 79:1035-10-37, 2013).

[0003] Existing drugs capable of imaging tau using positron emission tomography (PET) are known in the art, for example, such drugs are described in WO2009 / 102498 and WO2011 / 119565. Furthermore, compounds approved by the US Food and Drug Administration (FDA) [ 18 [(18)F]T807 (also known as AV-1451, the structure of which is shown below) is described in WO 2013 / 176698 (see also [(18)F]T807, a novel tau positron emission tomography imaging agent for Alzheimer's disease. Xia CF, et al., Alzheimer's Dement. 2013 Nov;9(6):666-76).

[0004] [ka]

[0005] However, existing tau imaging compounds have technical attributes that can be improved by the design of innovative agents that can provide improved tau images with improved tau signal and minimal non-tau signal or improved tau selectivity. Thus, there is a great need for improved methods for the detection and / or quantification of tau in patients.

[0006] Current clinically validated tau tracers are limited to detecting neurofibrillary tangles seen in late-stage Alzheimer's disease (see An Autoradiographic Evaluation of AV-1451 Tau PET in Dementia; Lowe et al. Acta Neuropathologica Communications (2016) 4:58 and Tau PET Imaging in Neurodegenerative Tauopathies—Still a Challenge; Leuzy et al. Molecular Psychiatry (2019) 24:1112–1134). There remains a need for detection of early-stage AD tau deposits and tau isoforms in non-AD (3R or 4R) tauopathies, such as progressive supranuclear palsy (PSP), corticobasal degeneration (CBD), and Pick's disease (PiD), as well as atypical Alzheimer's disease, chronic traumatic encephalopathy (CTE), and frontotemporal dementia (FTD). Novel PET tracers may be useful in advancing our understanding of early AD tau and non-AD tau-related neurodegenerative processes and in providing more effective tools for improving patient stratification and early inclusion in clinical trials evaluating novel therapeutics for these diseases. Detection of early tau accumulation can provide an endpoint for evaluating treatment efficacy in early AD patients, while identification of PET ligands in non-AD tauopathies (e.g., PSP, CBD, or PiD) may be useful for disease differentiation and staging, as well as for quantifying treatment efficacy in patients. Currently, no selective imaging agents for non-AD tauopathies are available to aid in better understanding the mechanisms underlying these neurodegenerative diseases and their progression. Summary of the Invention

[0007] Imaging tau in the brain using improved imaging agents has several potential advantages. Improved tau imaging could improve diagnosis by identifying potential patients with high levels of tau in the brain who are believed to be at a high probability of developing AD. Imaging with improved tau imaging agents could also be useful for monitoring tau accumulation and localization and / or the progression of AD and / or other tauopathies by PET. Once anti-tau drug treatments become available, tau imaging could provide an essential tool for monitoring treatment. The present disclosure provides novel compounds, compositions, formulations, and methods for tau imaging. Therefore, improved techniques that improve the ability to image tau in patients are also needed to expand the clinical benefit and impact of diagnostic tau imaging. Improved imaging agents could provide clearer and better PET images due to greater tau selectivity. Improved tau imaging agents could also improve our understanding of the development and progression of dementia in general, and AD and non-AD tauopathies in particular, leading to more effective treatments. [Brief explanation of the drawings]

[0008] [Figure 1] 1 shows autoradiography of AD brain sections for Kd measurement in Example 3 according to the protocol described in Assay Example 32. [Figure 2] 1 shows autoradiography of AD brain sections for selectivity measurements of Example 3 according to the protocol described in Assay Example 33. [Figure 3] 1 shows autoradiography of PSP brain sections from Example 4 for measurement of binding according to the protocol described in Assay Example 34. [Figure 4] 1 shows autoradiography of CBD brain sections from Example 4 for measurement of binding according to the protocol described in Assay Example 34. DETAILED DESCRIPTION OF THE INVENTION

[0009] The present invention provides a compound of formula 1:

[0010] [ka] During the ceremony, n is 0, 1, or 2; R 1 is H, halo, C1-C3 alkyl, or C3-C6 cycloalkyl; R 2 is H, halo, C1-C3 alkyl, C2-C3 alkenyl, C2-C3 alkynyl, -O-(C1-C4 alkyl), C1-C4 alkylhalo, C2-C4 alkenylhalo, or

[0011] [ka] and R 3 is H or F, R 4 is F, 18 F, C1-C4 alkylF, C1-C4 alkyl 18 F, -O-(C1-C4 alkyl)F, or -O-(C1-C4 alkyl) 18 F, R 5 is H, halo, or C1-C4 alkyl; where n is 0 and R 1 is methyl and R 2 , R 3 , and R 5 If each is H, then R 4 is F or 18 Not F.

[0012] In Formula I, n can be 0, 1, or 2. When n is 0, the ring is a 4-membered azetidin-1-yl ring. When n is 1, the ring is a 5-membered pyrrolidin-1-yl ring. When n is 2, the ring is piperidin-1-yl. In each of these ring systems, conventional nomenclature identifies the nitrogen atom at the 1-position of the ring. In selected embodiments, R 4 The substituent is attached to the ring at position 3. In another embodiment, when n is 2 in the piperidin-1-yl ring system, R 4 can be located at the 3- or 4-position of the ring.

[0013] Compounds of formula I do not include the two compounds exemplified below:

[0014] [ka]

[0015] In another aspect, the disclosure provides a compound of formula I above, or a pharmaceutically acceptable salt thereof, wherein n is 0, 1, or 2; and R 1 is H, halo, C1-C2 alkyl, or cyclopropane; R 2 is H, halo, -CH3, -CH2CH3, -OCH3, -CH=CH2, -CF=CH2,

[0016] [ka] -OCH2CH2F, or

[0017] [ka] and R 3 is H or F, and R 4 is F, 18 F, -CH2F, -CH2 18 F, -CH2CH2F, -CH2CH2 18 F, -CH2CH2CH2F, -CH2CH2CH2 18 F, -OCH2CH2F, or -OCH2CH2 18 F and R5 is H, halo, or C1-C4 alkyl.

[0018] In another aspect, the disclosure provides a compound of formula I above, or a pharmaceutically acceptable salt thereof, wherein n is 0 or 1 and R 1 is H, halo, C1-C2 alkyl, or cyclopropane; R 2 is H, halo, -CH3, -OCH3, -CH=CH2, -CF=CH2,

[0019] [ka] -OCH2CH2F, or

[0020] [ka] and R 3 is H or F, and R 4 are F or 3-position of the cycloalkyl ring, respectively. 18 F and R 5 is H, halo, or -CH3.

[0021] In another aspect, the present disclosure provides a compound of formula I above, or a pharmaceutically acceptable salt thereof, wherein n is 0 and R 1 H, halo, -CH3 or -CH2CH3, and R 2 is H, halo, -CH3, -OCH3, -CH=CH2, -CF=CH2,

[0022] [ka] or

[0023] [ka] and R 3 is H or F, and R 4 are F or 3-position of the cycloalkyl ring, respectively.18 F and R 5 is H, halo, or -CH3.

[0024] In another aspect, the present disclosure provides a compound of formula I above, or a pharmaceutically acceptable salt thereof, wherein R 1 is -CH3 and R 2 is H and R 3 is H and R 4 is F or 18 F and R5 is Cl.

[0025] In another aspect, the present disclosure provides a compound of formula I above, or a pharmaceutically acceptable salt thereof, wherein n is 0 and R 1 , R 2 , R 3 , and R 5 are H and R, respectively. 4 is F or 18 Provide something that is F.

[0026] In another aspect, the present disclosure provides a compound of formula I above, or a pharmaceutically acceptable salt thereof, wherein n is 0 and R 1 is H and R 2 is Cl, F or I, and R 3 is H and R 4 is F or 18 F and R 5 provides what is H.

[0027] In another aspect, the present disclosure provides a compound of formula I above, or a pharmaceutically acceptable salt thereof, wherein n is 0 and R 1 , R 3 , and R 5 are H and R, respectively. 2 are -CH3, -CH2CH3, -CF=CH2,

[0028] [ka] F, Cl, I, or R4 is F or 18 Provide something that is F.

[0029] In another aspect, the present disclosure provides a compound of formula I above, or a pharmaceutically acceptable salt thereof, wherein n is 0 and R 1 is -CH3 or -CH2CH3, and R 2 , R 3 and R 5 are H and R, respectively. 4 is F or 18 Provide something that is F.

[0030] In another aspect, the present disclosure provides a compound of formula I above, or a pharmaceutically acceptable salt thereof, wherein n is 2 and R 1 is H and R 2 is H or -CH3, R3 is H, R4 is F, 18 F, -C1-C3 alkylF, or -C1-C3 alkyl 18 F and R 5 provides what is H.

[0031] In some embodiments, the disclosure provides a compound of Formula I, or a pharmaceutically acceptable salt thereof, wherein the compound is:

[0032] [Table 1-1]

[0033] [Table 1-2]

[0034] [Table 1-3]

[0035] In yet another aspect, the present disclosure provides a pharmaceutical composition comprising a compound according to Formula I, as described in various aspects above, or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers, diluents, or stabilizers. Examples of pharmaceutical compositions and processes for their preparation can be found in "Remington: The Science and Practice of Pharmacy", Loyd, V., et al. Eds., 22nd Ed., Mack Publishing Co., 2012.

[0036] In another aspect, the present disclosure provides a pharmaceutical composition comprising a compound according to Formula I, as described in its various aspects above, or a pharmaceutically acceptable salt thereof, wherein the carrier comprises ethanol, water, and a buffer suitable for injection into a patient. Preferably, the composition includes sodium chloride in an amount that renders the formulation suitable for injection into a patient.

[0037] In this form, the buffering agent may include sodium chloride, sodium phosphate, or sodium ascorbate.

[0038] Examples of diluents include water for injection and saline. The diluent may be included in the pharmaceutical composition in an amount sufficient to provide a concentration of the radiolabeled embodiment of the compound of Formula I, or a pharmaceutically acceptable salt thereof, suitable for facilitating the diagnosis of patients at risk for or suffering from dementia or AD.

[0039] Examples of stabilizers, particularly radiolytic stabilizers, include ethanol, ascorbic acid, monothioglycerol, vitamin E, and cysteine.

[0040] The compounds of the present disclosure are preferably formulated as pharmaceutical compositions for intravenous administration to patients, preferably humans. Such pharmaceutical compositions and processes for preparing such compositions are known in the art (see, e.g., Remington: The Science and Practice of Pharmacy (PPG Erbino, 21st ed., Lippincott Williams & Wilkins, 2006)). Methods of using tau imaging agents for PET imaging of tau are known to those skilled in the art (see, e.g., [(18)F]T807, a novel tau positron emission tomography imaging agent for Alzheimer's disease. Xia CF, et al., Alzheimer's Dement. 2013 Nov;9(6):666-76). [(18)F]T807 is also known as [18F]AV-1451.

[0041] In another aspect, the present invention provides a pharmaceutical composition for imaging tau, comprising a compound of formula (I) or a pharmaceutically acceptable salt thereof. The tau imaging formulation is preferably formulated for use in humans. In one embodiment, the tau imaging formulation comprises a compound according to formula I or a pharmaceutically acceptable salt thereof formulated in 10% EtOH (v / v), 0.45% (w / v) sodium ascorbate in 0.9% sodium chloride.

[0042] The present disclosure also provides a method of imaging tau, comprising introducing into a patient a detectable amount of a compound of Formula I, or a pharmaceutically acceptable salt thereof. In another aspect, the present disclosure provides a method of imaging tau, comprising introducing into a patient a pharmaceutical composition comprising a detectable amount of a compound of Formula I, or a pharmaceutically acceptable salt thereof.

[0043] The present disclosure provides a method for imaging tau, comprising introducing into a mammal a detectable amount of a pharmaceutical composition described according to embodiments herein, allowing sufficient time for the pharmaceutical composition to bind to tau, and detecting the radiolabeled compound. A preferred method for detecting the radiolabeled compound uses PET.

[0044] The present invention provides the use of a compound of Formula 1 or a pharmaceutically acceptable salt thereof. Additionally, the present disclosure also provides that a compound of Formula 1 (or a pharmaceutically acceptable salt) can be used in the manufacture of a radiopharmaceutical for imaging tau in a patient, preferably a human.

[0045] The present disclosure provides processes for producing compounds according to Formula 1 having an F radiolabel. In certain embodiments, the present disclosure provides methods for preparing compounds of Formula 1, or pharmaceutically acceptable salts thereof, from precursor compounds outlined herein.

[0046] As used herein, "alkyl," "C1, C2, C3, C4, C5, or C6 alkyl," or "C1-C6 alkyl" is intended to include C1, C2, C3, C4, C5, or C6 straight-chain (straight-chain) saturated aliphatic hydrocarbon groups, and C3, C4, C5, or C6 branched-chain saturated aliphatic hydrocarbon groups. For example, C1-C6 alkyl is intended to include C1, C2, C3, C4, C5, and C6 alkyl groups. Examples of alkyl include moieties having 1 to 6 carbon atoms, such as, but not limited to, methyl, ethyl, n-propyl, i-propyl, n-butyl, s-butyl, t-butyl, n-pentyl, i-pentyl, or n-hexyl. In some embodiments, a straight chain or branched chain alkyl has 6 or fewer carbon atoms (e.g., C1-C6 for straight chain, C3-C6 for branched chain), and in other embodiments, a straight chain or branched chain alkyl has 4 or fewer carbon atoms. When this term is used in conjunction with the term halogen or, particularly, F atom or 18F atom, it refers to a halogen or F( 18F) atoms replace any one of the hydrogen atoms on the carbon chain. In some embodiments, F or 18 The F atom is attached to the terminal carbon atom of the chain.

[0047] The term "-O-(C1-C4 alkyl)" refers to an alkoxyl group containing 1 to 4 carbon atoms. The alkyl group may be a straight or branched alkyl chain. This term does not include halogen or, in particular, F atoms or 18 When used with the term F atom, it refers to a halogen or F( 18 F) can be attached to replace one of the hydrogen atoms attached to the carbon backbone of the chain. In some embodiments, the F atom or 18 The F atom is attached to the terminal carbon in the straight-chain alkyl group.

[0048] As used herein, the term "alkylhalo" includes saturated aliphatic groups similar in length and possible substitution to the alkyls described above, but in which at least a halogen atom replaces any one of the hydrogen atoms on the carbon chain.

[0049] As used herein, the term "alkenyl" includes unsaturated aliphatic groups similar in length and possible substitution to the alkyls described above, but containing at least one double bond. For example, the term "alkenyl" includes straight-chain alkenyl groups (e.g., ethenyl, propenyl, butenyl, pentenyl, hexenyl, heptenyl, octenyl, nonenyl, decenyl) and branched-chain alkenyl groups. In some embodiments, a straight-chain or branched-chain alkenyl group has six or fewer carbon atoms in its backbone (e.g., C2-C6 for straight chain, C3-C6 for branched chain). The term "C2-C4" includes alkenyl groups having two to four carbon atoms. The term "C2-C6" includes alkenyl groups having two to six carbon atoms. The term "C3-C6" includes alkenyl groups having three to six carbon atoms. Depending on the substituents attached to the two carbons of the double bond, the configuration around the double bond can be described as either a cis or trans double bond. The term refers to a halogen, or specifically to an F atom or 18When used with F atoms, halogen or F( 18 The F) atom can be bonded to replace one of the hydrogen atoms attached to the carbon backbone of the alkenyl chain.

[0050] As used herein, a "C2-C6 alkenylene linker" is intended to include a C2, C3, C4, C5, or C6 chain (straight or branched) divalent unsaturated aliphatic hydrocarbon group.

[0051] As used herein, the term "alkenylhalo" includes unsaturated aliphatic groups similar in length and possible substitution to the alkyls described above, but in which at least a halogen atom replaces any one of the hydrogen atoms on the carbon chain.

[0052] As used herein, the term "alkynyl" includes unsaturated aliphatic groups similar in length and possible substitution to the alkyl groups described above, but which contain at least one triple bond. For example, "alkynyl" includes straight-chain alkynyl groups (e.g., ethynyl, propynyl, butynyl, pentynyl, hexynyl, heptynyl, octynyl, nonynyl, decynyl) and branched-chain alkynyl groups. In some embodiments, a straight-chain or branched-chain alkynyl group has 6 or fewer carbon atoms in its backbone (e.g., C2-C6 for straight chain, C3-C6 for branched chain). The term "C2-C6" includes alkynyl groups having 2-6 carbon atoms. The term "C2-C4" includes alkynyl groups having 2-4 carbon atoms. The term "C3-C6" includes alkynyl groups having 3-6 carbon atoms. As used herein, a "C2-C6 alkynylene linker" is intended to include a C2, C3, C4, C5, or C6 chain (straight or branched) divalent unsaturated aliphatic hydrocarbon group. For example, a C2-C6 alkenylene linker is intended to include C2, C3, C4, C5, and C6 alkenylene linker groups.

[0053] As used herein, the term "cycloalkyl" refers to a group having 3 to 30 carbon atoms (e.g., C3 to C4 12 , C3~C 10, or C3-C8), a saturated or partially unsaturated hydrocarbon mono- or polycyclic (e.g., fused, bridged, or spiro) system. Examples of cycloalkyl include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclopentenyl, cyclohexenyl, cycloheptenyl, 1,2,3,4-tetrahydronaphthalenyl, and adamantyl. For polycyclic cycloalkyls, only one of the rings in the cycloalkyl need be non-aromatic.

[0054] The term "radiolabelled compound" refers to one of the compounds described below that contains an 18F atom.

[0055] The term "leaving group" (LG) refers to -NO2, trialkylamine, alkylsulfonate, or arylsulfonate. Alkylsulfonates of the present disclosure include C1-C4 alkylsulfonates. Arylsulfonates of the present disclosure include phenylsulfonates in which the phenyl group is optionally substituted once with C1-C4 alkyl, halogen, or nitro. Methanesulfonate (mesylate) and ethanesulfonate are preferred alkylsulfonates. Benzenesulfonate, 4-methylbenzenesulfonate (tosylate), 4-bromobenzenesulfonate, and 4-nitrobenzenesulfonate are preferred arylsulfonates.

[0056] [ka]

[0057] The bond designated as indicates the point of attachment of the 4-methylpyrazolyl ring to the rest of the molecule.

[0058] As used herein, the term " pharmaceutically acceptable salt " refers to the salt of the compound of the present invention that is deemed acceptable for clinical and / or veterinary use. Examples of pharmaceutically acceptable salts and the general methodology for their preparation can be found in "Handbook of Pharmaceutical Salts: Properties, Selection and Use" P.Stahl, et al., 2nd Revised Edition, Wiley-VCH, 2011 and S.M.Berge, et al., "Pharmaceutical Salts", Journal of Pharmaceutical Sciences, 1977, 66(1), 1-19.

[0059] As used herein, the term "effective amount" refers to a dosage that is effective in imaging tau. As a skilled artisan, the attending physician can easily determine an effective amount by using conventional techniques and observing results obtained under similar circumstances. When determining the effective amount or dose of a compound, many factors are taken into consideration, including (but not limited to) whether the compound or its salt is administered; if used, the co-administration of other drugs; the type of mammal; its size, age, and general health; the degree of involvement or severity of the disorder; the response of the individual patient; the mode of administration; the bioavailability characteristics of the administered preparation; the selected dosage regimen; the use of other concomitant drugs, and other relevant circumstances.

[0060] As used herein, the term "patient" refers to a mammal. Preferably, the patient is a human or companion mammal, such as a dog or cat; or other domesticated mammal, such as a cow, pig, horse, sheep, rabbit, mouse, rat, and goat.

[0061] The treating physician, veterinarian, or other medical professional will be able to determine the effective amount of the compound for treatment of the patient in need thereof. Preferred pharmaceutical compositions can be formulated as injectable solutions. The solutions can include the compounds of the present disclosure in an amount effective to treat the patient in need thereof.

[0062] Novel compounds of Formula I have been discovered to be advantageous for tau imaging, preferably including human clinical imaging. Some preferred compounds of Formula I possess a combination of properties that are particularly useful for tau imaging, including high affinity for tau. Some preferred compounds exhibit favorable tissue distribution and pharmacokinetics in vivo. Some compounds exhibit high affinity binding to tau ex vivo and / or in vitro and label tau-containing tissue samples from AD brains with high selectivity over Aβ and / or non-tau binding.

[0063] Abbreviations used herein are defined according to Daub GH, et al., "The Use of Acronyms in Organic Chemistry," Aldrichimica Acta, 1984, 17(1), 6-23. Other terms used herein include the following: "AD" refers to Alzheimer's disease; "Boc" or "BOC" refers to tert-butoxycarbonyl; "cat" refers to methyl methyl acrylate; "amt" refers to catalytic amount, "CT" or "CAT" refers to computed tomography, "DMAP" refers to 4-(dimethylamino)pyridine, "DMF" refers to dimethylformamide, "DMPAO" refers to (2,6-dimethylanilino)(oxo)acetic acid, "DMSO" refers to dimethylsulfoxide, "EOS" refers to end of synthesis, "ESI" refers to electrospray ionization, "EtOH" refers to ethanol, "HPLC" refers to high performance liquid chromatography, "hr" or "h" refers to time, "HRMS" refers to high resolution mass spectrometry, "LCMS" refers to liquid chromatography mass spectrometry, and "mCT" refers to microcomputerized tomography. "MeOH" refers to methanol, "minute, min" refers to minute, "mPET" refers to micropositron emission tomography, "MS" refers to mass spectrometry, "OMs" refers to O-mesyl, "OTs" refers to O-tosyl, "PBS" refers to phosphate buffered saline, "PET" refers to positron emission tomography, "PHF" refers to paired helical filaments, "Prec" refers to precursor, "Prep" refers to compound preparation, "RCP" refers to radiochemical purity, "RT" refers to room temperature, "SM" refers to starting material, "TAC" refers to time activity curve, "OTMS" refers to O-trimethylsilyl, and "WFI" refers to water for injection.

[0064] General Chemical Synthesis The following schemes, preparations, precursors, and examples are presented to facilitate the practice of the present invention. Suitable reaction conditions for each step in these schemes, preparations, precursors, and examples are known in the art, and appropriate modifications of the reaction conditions, including substitutions of solvents and co-reagents, are within the skill of one of ordinary skill in the art.

[0065] Furthermore, those skilled in the art will understand that, under some circumstances, the order in which moieties are introduced is not critical. The specific order of steps required to produce compounds of Formula I will depend on the particular compound being synthesized, the starting compound, and the relative lability of the substituted moieties, as is well recognized by skilled chemists. Those skilled in the art will understand that not all substituents are compatible with all reaction conditions. These compounds can be protected or modified at convenient points in the synthesis by methods well known in the art. The intermediates and final products of the present disclosure can be further purified, if desired, by common techniques such as recrystallization or chromatography on solid supports such as silica gel or alumina.

[0066] The compounds of the present disclosure or salts thereof can be prepared by various procedures known in the art, some of which are illustrated in the following schemes, preparations, precursors, and examples. Specific synthetic steps in each of the described routes can be combined in different ways, or steps from different schemes can be combined to prepare the compounds or salts of the present disclosure. The products of each step in the following schemes can be recovered by conventional methods well known in the art, including extraction, evaporation, precipitation, chromatography, filtration, trituration, and crystallization. In the following schemes, all substituents are as previously defined unless otherwise indicated. Reagents and starting materials are readily available to those skilled in the art.

[0067] All reactions are carried out under a nitrogen atmosphere unless otherwise noted. Reagents, solvents, and supplies are purchased from commercial sources unless otherwise indicated. Compounds are purified using an automated Teledyne Isco flash chromatography system. HRMS data are acquired on a Waters QT mass spectrometer using electrospray ionization positive scan mode. Nominal resolution MS data are acquired on a Waters Micromass ZQ mass spectrometer using ESI positive ionization scan mode. [Example]

[0068] Scheme 1 (5H-pyrido[1,2-a]benzimidazole ring formation)

[0069] [ka]

[0070] Method A: Step 1 Copper-catalyzed benzo[4,5]imidazo[1,2-a]pyridine formation 5H-Pyrido[1,2-a]benzimidazole (IV) can be prepared by reacting appropriately substituted bromoaniline (II), bromopyridine (III), a base such as cesium carbonate, a catalytic amount of copper(I) iodide, and 1,10-phenanthroline in a nonpolar solvent such as a mixture of xylenes. Upon completion of the reaction, the mixture is cooled to room temperature and filtered to remove solids. The desired material is isolated using standard procedures. In some of the following preparations, X is a halogen such as bromine or chlorine. In other preparations, X is a cyclic amine. The cyclic amine may be substituted with a linker "L," which may be a bond or an alkyl or O-alkyl. A leaving group "LG," e.g., -NO, trialkylamine, alkylsulfonate, or arylsulfonate, is attached to the cyclic amine via L to form the precursor. Alternatively, R4 may be attached to the cyclic amine via L.

[0071] Method B: Step 2 Synthesis of N-phenylpyridin-2-amine An appropriately substituted aniline (V), a substituted pyridine 1-oxide (VI), bromotripyrrolidinophosphonium hexafluorophosphate, and a base (e.g., diisopropylethylamine) are combined in methylene chloride and stirred at room temperature until the reaction is complete. The reaction mixture is concentrated, and the residue is subjected to silica gel chromatography to isolate the desired phenylpyridine compound (VII).

[0072] Method B: Step 3 Oxidative ring closure to form benzo[4,5]imidazo[1,2-a]pyridine: A mixture of appropriately substituted pyridin-2-amine (VII) and (diacetoxyiodo)benzene in a solvent is stirred for 1-5 hours while monitoring the reaction for completion. The mixture is concentrated, and the desired benzo[4,5]imidazo[1,2-a]pyridine (VIII) is isolated by chromatography.

[0073] Scheme 2 Amine Coupling

[0074] [ka]

[0075] Method C RuPhos-catalyzed Buchwald amine coupling: Halobenzo[4,5]imidazo[1,2-a]pyridine (VIII), cyclic amine (IX), base, and chloro(2-dicyclohexylphosphino-2',6'-di-i-propoxy-1,1'-biphenyl)[2-(2-aminoethylphenyl)]palladium(II), methyl t-butyl ether adduct (Ru Phos-Pd) (catalytic amount) are combined in dioxane and the mixture is heated. The reaction can be monitored by LCMS. Upon completion, the reaction mixture is cooled and the desired compound (XI) is isolated using standard chromatography or other purification techniques.

[0076] Method D XPhos-catalyzed Buchwald amine coupling: A mixture of bromobenzo[4,5]imidazo[1,2-a]pyridine (1 equivalent), cyclic amine (1.3 to 2 equivalents), potassium phosphate monohydrate (7 equivalents), and chloro(2-dicyclohexylphosphino-2',4',6'-tri-i-propyl-1,1'-biphenyl)[2-(2-aminoethyl)phenyl]palladium(II) methyl t-butyl ether adduct (XPhos-Pd) (0.2 equivalents) in dioxane (0.07-0.15 M) is heated at 100 °C. The reaction is monitored by LCMS. Upon completion, the reaction mixture is cooled to room temperature and concentrated. The desired material is isolated by column chromatography (silica gel) eluting with a methylene chloride:methanol gradient.

[0077] Method E Copper-catalyzed amine coupling: A mixture of bromobenzo[4,5]imidazo[1,2-a]pyridine (1 equivalent), cyclic amine (1.5 equivalents), L-proline (0.5 equivalents), copper(I) iodide (0.3 equivalents), and anhydrous potassium phosphate (3 equivalents) in dimethyl sulfoxide (0.25 M) is heated to 100-110 °C in a 20 ml reaction vial. The reaction is monitored by LCMS. Upon completion, the reaction mixture is cooled to room temperature and poured into deionized water. The aqueous mixture is extracted with 90:10 methylene chloride:methanol (5x). The combined organic layers are washed with 50% ammonium hydroxide, dried over magnesium sulfate, filtered, and concentrated. The crude material is purified by column chromatography on silica gel using a methylene chloride:methanol gradient.

[0078] Method F Copper-catalyzed amine coupling: A mixture of 2,4-dibromo-5-fluoropyridine (1 equivalent), the appropriate amine (1-1.4 equivalents), copper(I) iodide (0.1-0.15 equivalents), 2-((2,6-dimethylphenyl)amino)-2-oxoacetic acid (DMPAO) (0.15-0.2 equivalents), potassium phosphate (3 equivalents), and dimethyl sulfoxide (0.2-0.6 M) is purged under nitrogen for 10 minutes. The reaction is heated at 100°C for 2-24 hours. Water is added, and the mixture is extracted with ethyl acetate. The combined organic layers are dried over magnesium sulfate, filtered, and washed with dichloromethane. The solvent is removed under vacuum. Optionally, the reaction is cooled to room temperature and added dropwise to water. The solid is isolated by filtration and washed with water. The solid is redissolved in 10% methanol in dichloromethane and adsorbed onto silica gel. The crude product is purified by column chromatography on silica gel using a gradient of 5-65% ethyl acetate in hexanes.

[0079] Scheme 3 Precursor formation

[0080] [ka]

[0081] Method G A mixture of the appropriate 1-(benzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol (XII) (where L is as defined above), methanesulfonyl chloride or p-toluenesulfonyl chloride is stirred to give LG (as defined above), and base (6-25 mM) in methylene chloride. (Optionally, catalytic DMAP can be added.) Upon completion, the reaction is worked up using conventional neutralization and / or extraction techniques. The desired precursor XIII can be isolated by chromatographic techniques.

[0082] Scheme 4 Fluorine-18 radiolabeling procedure

[0083] [ka]

[0084] General radiosynthesis method A solution of the appropriately substituted precursor (XIII) (where L' and LG are as defined above) was added to the anhydrous cryptand 2.2.2-K2CO3 [ 18 The resulting mixture is maintained at 140 °C for 10 min, then hydrolyzed with 1 mL of 1 N NaOH at 65 °C for 3 min. After cooling to 60 °C, the crude reaction mixture is neutralized with 2 mL of 0.5 N HCl (1 mL of 1 N HCl + 1 mL of WFI). The crude reaction product is then loaded onto a semi-preparative HPLC column for purification using isocratic elution.

[0085] Preparation of starting materials Preparation 1 3-Bromo-8-methylbenzo[4,5]imidazo[1,2-a]pyridine

[0086] [ka] 2,4-Dibromopyridine (20.0 g, 84.6 mmol), copper(I) iodide (3.22 g, 16.9 mmol), 1,10-phenanthroline (6.10 g, 33.8 mmol), cesium carbonate (110 g, 338 mmol), diatomaceous earth (16 g), and p-xylene (170 mL) were added together. 2-Bromo-4-methylaniline (10.6 mL, 84.6 mmol) was added to the resulting slurry, and nitrogen gas was bubbled through the vigorously stirred mixture for 10 minutes. The reaction mixture was heated to 135 °C for 24 hours, cooled to room temperature, and filtered. The filter cake was washed with methylene chloride and ethyl acetate, and the combined organic filtrate was concentrated under reduced pressure onto silica gel. The crude reaction product was purified by chromatography on silica gel using a gradient of 0 to 10% ethyl acetate in methylene chloride. The resulting brown solid was slurried in methylene chloride, triturated with hexane, and isolated by filtration to give the title compound (6.52 g, 25.0 mmol, 30% yield) as a shiny yellow solid. ES / MS m / z ( 79Br / 81 Br)260.92 / 262.92[M+H]

[0087] The compounds in Table 1 below were prepared as generally described in Scheme 1, Method A (Step 1), essentially following the procedures described above for Preparation 1.

[0088] [Table 2-1]

[0089] [Table 2-2]

[0090] Preparation 24 4-chloro-N-(4-cyclopropylphenyl)pyridin-2-amine

[0091] [ka] A solution of 4-cyclopropylaniline (0.213 g, 1.6 mmol), 4-chloropyridine N-oxide (0.25 g, 1.9 mmol), and diisopropylethylamine (0.98 mL, 5.6 mmol) in dichloromethane (8 mL) was treated with bromotripyrrolidinophosphonium hexafluorophosphate (PyBrop) (1.19 g, 2.6 mmol) and stirred at room temperature overnight. The reaction mixture was purified by column chromatography on silica gel (0-70% ethyl acetate in hexanes) to give the pure product as a light brown solid (0.266 g, 68%). ES / MS m / z ( 35 Cl / 37 Cl)245.1 / 246.9(M+H)

[0092] Preparation 27 3-chloro-8-cyclopropylbenzo[4,5]imidazo[1,2-a]

[0093] [ka] A solution of 4-chloro-N-(4-cyclopropylphenyl)pyridin-2-amine (0.266 g, 1.09 mmol) in hexafluoroisopropanol (11 mL) was treated with bis(tert-butylcarbonyloxy)iodobenzene (0.46 g, 1.14 mmol) and stirred at room temperature overnight. The reaction mixture was concentrated and purified by column chromatography on silica gel (0-60% ethyl acetate in hexanes) to give the title compound as a light brown solid (0.22 g, 84%). ES / MS m / z ( 35 Cl / 37 Cl)243.3 / 245.0(M+H)

[0094] Preparation 29 3-chloro-8-iodobenzo[4,5]imidazo[1,2-a]pyridine

[0095] [ka] This compound was prepared in 54% yield from 4-chloro-N-(4-iodophenyl)pyridin-2-amine essentially following the procedure of Preparation 27. ES / MS m / z ( 35 Cl / 37 Cl)328.8 / 330.7

[0096] Preparation 30 3-chloro-8-ethylbenzo[4,5]imidazo[1,2-a]pyridine

[0097] [ka] Compound 30 was prepared essentially according to Scheme 1. Method B, Steps 2 and 3. To a mixture of 4-chloropyridine N-oxide (1.3 g, 10 mmol) and 4-ethylaniline (1.2 g, 10 mmol) in dichloromethane (50 mL) was added diisopropylethylamine (5.2 mL, 30 mmol) and bromotripyrrolidinophosphonium hexafluorophosphate (PyBrop) (5.1 g, 11 mmol) at room temperature. The resulting solution was stirred overnight at room temperature, then concentrated and purified by column chromatography on silica gel (0 to 50% ethyl acetate in hexanes) to give a red solid (1.4 g, 60%). This material was dissolved in hexafluoroisopropanol (40 mL) and bis(tert-butylcarbonyloxy)iodobenzene (2.56 g, 6.3 mmol) was added. The reaction was stirred at room temperature for 3-4 hours, then concentrated and purified by column chromatography on silica gel (0-70% ethyl acetate in hexanes) to give the title compound as a yellow solid (1.2 g, 5.2 mmol, 52% over two steps). ES / MS m / z ( 35 Cl / 37 Cl)231.1 / 232.9(M+H)

[0098] Preparation 31 1-(7-bromo-8-fluorobenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol

[0099] [ka] 3,7-Dibromo-8-fluorobenzo[4,5]imidazo[1,2-a]pyridine (0.120 g, 0.349 mmol), 3-hydroxyazetidine hydrochloride (0.076 g, 0.698 mmol), sodium tert-butoxide (0.134 g, 1.40 mmol), chloro(2-dicyclohexylphosphino-2',6'-diisopropoxy-1,1'-biphenyl)[2-(2'-amino-1,1'-biphenyl)]palladium II) (Ruphos Pd G2) (0.027 g, 0.035 mmol), and dioxane (3 mL) were added and purged with nitrogen gas for 10 minutes. The mixture was heated in a microwave oven at 110 °C for 45 minutes and then cooled to room temperature. The solvent was removed in vacuo. The residue was purified by column chromatography on silica gel (2% to 30% methanol in methylene chloride). Selected fractions were combined and concentrated to give the title compound as a yellow solid (28 mg, 24%). ES / MS m / z 336.1 (M+H).

[0100] The compounds in Table 2 below were prepared essentially according to Preparation 31 above and as generally described in Scheme 2, Method C.

[0101] [Table 3]

[0102] Preparation 39 1-(benzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol

[0103] [ka] Dimethyl sulfoxide (110 mL) was added to a solid mixture of 3-bromo-8-methylbenzo[4,5]imidazo[1,2-a]pyridine (6.57 g, 25.2 mmol), azetidin-3-ol hydrochloride (5.52 g, 50.4 mmol), copper(I) iodide (480 mg, 2.52 mmol), DMPAO (972 mg, 5.04 mmol), and tribasic potassium phosphate (21.4 g, 101 mmol). Nitrogen gas was bubbled through the stirred slurry for 15 minutes. The mixture was heated at 90°C for 24 hours. The reaction mixture was cooled to room temperature, and water (1000 mL) was added slowly with vigorous stirring. The precipitated solid was isolated by vacuum filtration and dissolved in 10% methanol in methylene chloride (500 mL). The aqueous filtrate was extracted with 10% methanol in methylene chloride (3 x 250 mL). The organic extracts were combined with the solution of the solid isolated from the first aqueous filtration, dried over sodium sulfate, filtered, and concentrated under reduced pressure. The resulting solid was dissolved in methylene chloride, adsorbed onto silica gel, and purified by chromatography on silica gel using a gradient of 0-30% methanol in methylene chloride to afford the title compound as a green-gray solid (3.18 g, 12.6 mmol, 50% yield). HRMS m / z): 254.1296 (M+H).

[0104] The compounds in Table 3 below were prepared essentially according to Preparation 39 above and as generally described in Scheme 2, Method E.

[0105] [Table 4-1]

[0106] [Table 4-2]

[0107] Preparation 49 1-(7-Bromobenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol

[0108] [ka] Dimethyl sulfoxide (6.7 mL) was added to a mixture of 3,7-dibromobenzo[4,5]imidazo[1,2-a]pyridine (500 mg, 1.53 mmol, 1.0 equiv.), azetidin-3-ol hydrochloride (336 mg, 3.06 mmol, 2.0 equiv.), copper(I) iodide (29.0 mg, 0.153 mmol, 0.1 equiv.), DMPAO (59.0 mg, 0.306 mmol, 0.2 equiv.), and potassium phosphate (977 mg, 4.60 mmol, 3.0 equiv.) and sparged with nitrogen for 10 minutes. The vial was sealed and heated at 90 °C for 20 hours, then cooled and diluted with water (40 mL). The precipitate was collected by filtration and dissolved in 10% methanol in methylene chloride. This solution was combined with the organic layer obtained by extracting the aqueous filtrate with 10% methanol in methylene chloride (3 x 20 mL). The combined organic phases were dried over sodium sulfate, filtered, and concentrated onto silica gel. The crude product was purified by column chromatography on silica gel to give the title compound as a brown solid (170 mg, 0.535 mmol, 35%). ES / MS m / z 319.94 (M+H).

[0109] The compounds in Table 4 below were prepared essentially according to Preparation 49 above and as generally described in Scheme 2, Method F.

[0110] [Table 5]

[0111] Preparation 53 3-chloro-7-(1-fluorovinyl)benzo[4,5]imidazo[1,2-a]pyridine

[0112] [ka] A mixture of 3-chloro-7-iodobenzo[4,5]imidazo[1,2-a]pyridine (160 mg, 0.49 mmol), (1-fluorovinyl)(methyl)diphenylsilane (242 mg, 1.0 mmol), cesium fluoride (228 mg, 1.5 mmol), copper(I) iodide (10 mg, 0.05 mmol), and tetrakis(triphenylphosphine)palladium(0) (58 mg, 0.05 mmol) in dimethylformamide (4 mL) was stirred at room temperature for 2 days. The reaction was purified by column chromatography on silica gel to give the title compound (25 mg, 21%).

[0113] Preparation 54 2-((1-(7-vinylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-yl)oxy)ethanol

[0114] [ka]

[0115] Step 1: Preparation of 1-(7-vinylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol

[0116] [ka] A mixture of dimethoxyethane:ethanol:water (7:2:1 v / v, 5.0 mL) was added to 1-(7-bromobenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol (165 mg, 0.516 mmol, 1.0 equiv.) and potassium carbonate (213 mg, 1.55 mmol, 3.0 equiv.) in a screw-cap vial. Vinylboronic acid pinacol ester (130 μL, 0.773 mmol, 1.5 equiv.), [1,1'-bis(di-tert-butylphosphino)ferrocene]-dichloropalladium(II) (16.8 mg, 0.026 mmol, 0.05 equiv.), and dimethyl sulfoxide (5.0 mL) were combined and sparged with nitrogen for 10 minutes. The vial was capped and heated at 100 °C for 16 hours. The reaction mixture was cooled to room temperature, diluted with water (40 mL), and filtered. The filter cake was dissolved in 10% methanol in methylene chloride, and the organic layer obtained by extracting the aqueous filtrate with methylene chloride (3 x 20 mL) was combined. The combined organic phases were dried over sodium sulfate, filtered, and concentrated onto silica gel. The crude product was purified by column chromatography on silica gel (0-10-20% methanol in methylene chloride) to give the title compound as a brown solid (75.0 mg, 0.283 mmol, 55%). ES / MS m / z 266.03 (M+H).

[0117] Step 2: Preparation of 3-(3-(2-((tetrahydro-2H-pyran-2-yl)oxy)ethoxy)azetidin-1-yl)-7-vinylbenzo[4,5]imidazo[1,2-a]pyridine

[0118] [ka] To a solution of 1-(7-vinylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol (31 mg, 0.12 mmol) and 2-(2-bromoethoxy)tetrahydro-2H-pyran (50 mg, 0.24 mmol, 2 equiv.) in dimethylformamide (1 mL) was added sodium hydride (60% in mineral oil, 10 mg, 0.24 mmol, 2 equiv.). The reaction was stirred at room temperature for 3 hours. Additional sodium hydride (60% in mineral oil, 10 mg, 0.24 mmol, 2 equiv.) was added and stirred at room temperature for 16 hours. The reaction was diluted with water (5 mL). The aqueous phase was extracted with dichloromethane (3 × 4 mL). The organic phase was concentrated in vacuo and then placed under high vacuum to give the crude product as a brown residue (47 mg, quantitative yield). ES / MS m / z 394.4(M+H)

[0119] Step 3: Preparation of 2-((1-(7-vinylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-yl)oxy)ethanol

[0120] [ka] p-Toluenesulfonic acid monohydrate (23 mg, 0.12 mmol, 1 equiv.) was added to a solution of 3-(3-(2-((tetrahydro-2H-pyran-2-yl)oxy)ethoxy)azetidin-1-yl)-7-vinylbenzo[4,5]imidazo[1,2-a]pyridine (47 mg, 0.12 mmol) in methanol (1 mL). The reaction was stirred at room temperature for 2 hours and then concentrated in vacuo. The residue was dissolved in a 9:1 dichloromethane:methanol mixture (6 mL) and washed with saturated aqueous sodium bicarbonate solution (5 mL). The aqueous phase was extracted with a 9:1 dichloromethane:methanol mixture (2 x 6 mL). The organic layer was dried over sodium sulfate and concentrated in vacuo to give the title compound as a brown residue (45 mg, quantitative yield). ES / MS m / z 310.2 (M+H)

[0121] Preparation 55 1-(7-ethylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol

[0122] [ka] 2-Nitrobenzenesulfonyl chloride (175 mg, 0.792 mmol, 3.0 equiv.) was added to a slurry of 1-(7-vinylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol (75.0 mg, 0.264 mmol, 1.0 equiv.) and hydrazine hydrate (65%, 101 μL, 1.32 mmol, 5.0 equiv.) in a mixture of methylene chloride (1.7 mL) and dimethylformamide (0.85 mL) at room temperature. The reaction mixture was stirred for 16 hours, concentrated onto silica gel, and then purified by column chromatography on silica gel to give the title compound as a beige solid (28.0 mg, 0.104 mmol, 40%). ES / MS m / z 268.16 (M+H).

[0123] Preparation 56 3-(3-fluoroazetidin-1-yl)-7-vinylbenzo[4,5]imidazo[1,2-a]pyridine

[0124] [ka] 7-Bromo-3-(3-fluoroazetidin-1-yl)benzo[4,5]imidazo[1,2-a]pyridine (115 mg, 0.36 mmol, 1.0 equiv.), potassium vinyltrifluoroborate (72 mg, 0.54 mmol, 1.5 equiv.), and potassium carbonate (149 mg, 1.08 mmol, 3.0 equiv.) were added to a scintillation vial, followed by a 7:2:1 mixture of dimethoxyethane:ethanol:water (3.5 mL). Catalyst [1,1'-bis(di-tert-butylphosphino)ferrocene]dichloropalladium(II) (23 mg, 0.036 mmol, 0.1 equiv.) was added, and the reaction vial was sealed and heated to 80 °C for 16 h. The reaction mixture was cooled to room temperature, concentrated onto silica gel, and purified by column chromatography on silica gel (0-5-10% methanol in methylene chloride) to give the title compound as a light brown solid (33.0 mg, 0.124 mmol, 34%). ES / MS m / z 261.1 (M+H).

[0125] Preparation 57 1-(8-fluoro-7-vinylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol

[0126] [ka] 1-(7-Bromo-8-fluorobenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol (22 mg, 0.066 mmol), vinyl pinacolborane (20 mg, 0.13 mmol), potassium carbonate (27 mg, 0.196 mmol), and [1,1'-bis(di-tert-butylphosphino)ferrocene]dichloropalladium(II) (4.0 mg, 0.0066 mmol) were added and purged with nitrogen gas for 10 minutes. Dimethoxyethane / ethanol / water (7 / 2 / 1.3 mL) and dimethyl sulfoxide (5 mL) were added. The reaction mixture was heated at 80°C overnight. The reaction mixture was partitioned between ethyl acetate and water. The organic layer was separated, washed with brine, and dried over sodium sulfate. The solid was filtered and washed with dichloromethane. The solvent was removed in vacuo. Purification was carried out by column chromatography on silica gel (0-30% methanol in methylene chloride) to give the title compound as a grey solid (17 mg, 91%). ES / MS m / z 284.2 (M+H).

[0127] Preparation 58 1-(2-fluoro-7-methylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol

[0128] [ka] A vial was charged with 3-(3-((tert-butyldimethylsilyl)oxy)azetidin-1-yl)-2-fluoro-7-methylbenzo[4,5]imidazo[1,2-a]pyridine (0.022 g, 0.0571 mmol) and 1 M tetrabutylammonium fluoride in tetrahydrofuran (1.0 mL, 1 mmol). The reaction mixture was heated to 45° C. for 4 h. The solvent was removed in vacuo. The residue was purified by column chromatography on silica gel (0-20% methanol in dichloromethane). Selected fractions were concentrated to give the title compound as a white solid (11 mg, 71%). ES / MS m / z 272.1 (M+H).

[0129] Preparation 59 3-(1-(benzo[4,5]imidazo[1,2-a]pyridin-3-yl)piperidin-4-yl)propan-1-ol

[0130] [ka] A slurry of 3-bromobenzo[4,5]imidazo[1,2-a]pyridine (0.3 g, 1.21 mmol), 3-(piperidin-4-yl)propan-1-ol (260.8 mg, 1.82 mmol), (2-biphenyl)dicyclohexylphosphine (51.1 mg, 0.146 mmol), lithium bis(trimethylsilyl)amide (1 M in tetrahydrofuran, 3 mL, 2.67 mmol), and tris(dibenzylideneacetone)dipalladium(0) (55.5 mg, 60.7 mmol) was sparged with nitrogen and heated at 100° C. overnight. Purification was carried out by column chromatography on silica gel to give the title compound as a light brown solid (53.3 mg, 14%).

[0131] Precursor preparation Preparation of P1 1-(8-methylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-yl 4-methylbenzenesulfonate

[0132] [ka] A mixture of 1-(8-methylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol (0.010 g, 0.036 mmol), p-toluenesulfonyl chloride (13.6 mg, 0.071 mmol), and triethylamine (6 equiv.) in methylene chloride (6-25 mM) was stirred at room temperature. Upon completion, the reaction was treated with saturated aqueous sodium bicarbonate solution with vigorous stirring. The phases were separated, and the aqueous layer was extracted with methylene chloride (4 times). The combined organic phases were dried over sodium sulfate, filtered, and concentrated. The crude material was purified by column chromatography on silica gel using a methylene chloride:methanol gradient to afford the title compound as an off-white solid (8 mg, 52%). LCMS (ESI+) 435.89 (M+H).

[0133] Alternative preparation of P1 A suspension of 1-8-methylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl]azetidin-3-ol (3.18 g, 12.6 mmol) in dichloromethane (135 mL) was treated with triethylamine (17.5 mL, 126 mmol) and stirred for 10 minutes, after which p-toluenesulfonic anhydride (12.31 g, 37.7 mmol) was added. The reaction mixture was stirred at room temperature for 22 hours. Additional p-toluenesulfonic anhydride (1.84 g, 5.6 mmol) was added and stirred for 6 hours. The reaction mixture was concentrated, resuspended in methylene chloride (175 mL), and treated with 1 N aqueous sodium hydroxide (150 mL). The biphasic mixture was stirred vigorously for 90 minutes, transferred to a separatory funnel, and the layers were separated. The organic layer was vigorously shaken with 1N aqueous sodium hydroxide (2 × 100 mL, 1 × 150 mL), dried over magnesium sulfate, filtered, concentrated, and placed under high vacuum. A solution of the isolated solid in 10% methanol in methylene chloride was concentrated under reduced pressure onto silica gel (24 g). Purification of the title compound was carried out by chromatography on silica gel using a gradient of 0 to 10% methanol in methylene chloride. The resulting solid was suspended in methylene chloride (approximately 50 mL), sonicated, and triturated with diethyl ether (750 mL). The precipitated solid was collected by filtration, rinsed with diethyl ether, and dried under vacuum to give the title compound as a beige solid (3.21 g, 7.89 mmol, 63% yield).

[0134] The precursors in Table 5 below were prepared essentially according to Preparation P1 or alternative preparations according to Scheme 3 of Method G.

[0135] [Table 6-1]

[0136] [Table 6-2]

[0137] [Table 6-3]

[0138] Preparation of precursor 21 3-(3-(tert-butyldimethylsilyl)oxy)azetidin-1-yl)-7-iodobenzo[4,5]imidazo[1,2-a]pyridine

[0139] [ka]

[0140] Step 1: Preparation of 3-(3-(tert-butyldimethylsilyl)oxy)azetidin-1-yl)-7-(tributylstannyl)benzo[4,5]imidazo[1,2-a]pyridine

[0141] [ka] A solution of 7-bromo-3-(3-((tert-butyldimethylsilyl)oxy)azetidin-1-yl)benzo[4,5]imidazo[1,2-a]pyridine (432 mg, 1.0 mmol), lithium chloride (260 mg, 6.0 mmol), bis(tributyltin) (1.74 g, 3.0 mmol), and tetrakis(triphenylphosphine)palladium(0) (120 mg, 0.1 mmol) in dioxane (20 mL) was sparged with nitrogen for 5 minutes and then stirred at 100 °C for 4–5 hours. The reaction mixture was diluted with dichloromethane, filtered, and concentrated. The residue was purified by column chromatography on silica gel (0–15% methanol in dichloromethane) to afford the title compound (574 mg, 89%) as a yellow solid.

[0142] Step 2: Preparation of 3-(3-(tert-butyldimethylsilyl)oxy)azetidin-1-yl)-7-iodobenzo[4,5]imidazo[1,2-a]pyridine

[0143] [ka] Iodine (76 mg, 0.3 mmol) in dichloromethane was added dropwise to a stirred solution of 3-(3-((tert-butyldimethylsilyl)oxy)azetidin-1-yl)-7-(tributylstannyl)benzo[4,5]imidazo[1,2-a]pyridine (190 mg, 0.3 mmol) in dichloromethane (6.0 mL) at 0° C. After 5 min, the reaction was quenched with 5% aqueous sodium bisulfite, extracted with dichloromethane, dried over sodium sulfate, filtered, and concentrated. The residue was purified by column chromatography on silica gel (0-20% methanol in dichloromethane) to give the title compound as a solid (42 mg, 29%). ES / MS m / z 480.3 (M+H)

[0144] Alternative preparation of precursor 21 1-(7-Iodobenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-yl 4-methylbenzenesulfonate

[0145] [ka] Six to seven drops of acetyl chloride were added to a solution of 3-(3-((tert-butyldimethylsilyl)oxy)azetidin-1-yl)-7-iodobenzo[4,5]imidazo[1,2-a]pyridine (42 mg, 0.088 mmol) in methanol (6 mL). The reaction was stirred overnight at room temperature, concentrated, and dried under high vacuum for several hours. The residue was redissolved in dichloromethane (40 mL), and triethylamine (0.25 mL, 1.76 mmol), p-toluenesulfonyl chloride (100 mg, 0.53 mmol), and a catalytic amount of 4-(N,N-dimethylamino)pyridine were added. The mixture was stirred overnight at 40 °C, extracted with 1 N sodium hydroxide, dried over sodium sulfate, filtered, and concentrated. The residue was purified by column chromatography on silica gel (0-10% methanol in ethyl acetate) followed by a second silica gel column (0-15% methanol in dichloromethane) to give the title compound as a yellow solid (24 mg, 52%). ES / MS m / z 520.2 (M+H).

[0146] Preparation of precursor 22 1-(7-(1-fluorovinyl)benzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-yl 4-methylbenzenesulfonate

[0147] [ka] p-Toluenesulfonic anhydride (273 mg, 0.84 mmol), 4-(N,N-dimethylamino)pyridine (1 mg, 0.008 mmol), and triethylamine (0.1 mL) were added to a solution of 1-(7-(1-fluorovinyl)benzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol (20 mg, 0.07 mmol) in dichloromethane (3 mL). The mixture was stirred at room temperature and monitored by LCMS until all of the alcohol had been converted to the ditosylate product. The mixture was washed with aqueous sodium bicarbonate and extracted with 10% methanol in dichloromethane. The organic layer was dried over magnesium sulfate, and the solvent was removed under reduced pressure. Potassium hydroxide (1 N aqueous solution, 1 mL) and methanol (2 mL) were added. The mixture was stirred at room temperature for 1 hour until all of the ditosylate had been converted to the desired compound. The pH of the mixture was adjusted by slow addition of hydrochloric acid (1 N) and washed with aqueous sodium bicarbonate. The aqueous mixture was extracted with 10% methanol in methylene chloride. The organic solvent was removed under reduced pressure. The crude product was purified by column chromatography on silica (0-10% methanol in dichloromethane) to give the title compound as an off-white solid (14.4 mg, 46%). ES / MS m / z 438.14 (M+H).

[0148] Preparation of precursor 23 1-(7-ethynylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-yl 4-methylbenzenesulfonate

[0149] [ka]

[0150] Step 1: Preparation of 3-(3-(tert-butyldimethylsilyl)oxy)azetidin-1-yl)-7-((trimethylsilyl)ethynyl)benzo[4,5]imidazo[1,2-a]pyridine

[0151] [ka] A mixture of 7-bromo-3-(3-((tert-butyldimethylsilyl)oxy)azetidin-1-yl)benzo[4,5]imidazo[1,2-a]pyridine (216 mg, 0.5 mmol), ethynyltrimethylsilane (0.25 mL, 1.8 mmol), tetrakis(triphenylphosphine)palladium (58 mg, 0.05 mmol), and copper(I) iodide (12 mg, 0.03 mmol) in trimethylamine and dimethylformamide (2 mL, 1:1) was sparged with nitrogen and then stirred at 90 °C overnight. The mixture was filtered through a pad of diatomaceous earth and washed with 10% methanol in dichloromethane. The filtrate was evaporated under reduced pressure and purified by column chromatography on silica gel (0–10% methanol in dichloromethane) to give the title compound (140 mg, 62%) as a light brown solid. ES / MS m / z 450.57(M+H)

[0152] Step 2: Preparation of 1-(7-((trimethylsilyl)ethynyl)benzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol

[0153] [ka] Acetyl chloride (25 μL) was added to a solution of 3-(3-((tert-butyldimethylsilyl)oxy)azetidin-1-yl)-7-((trimethylsilyl)ethynyl)benzo[4,5]imidazo[1,2-a]pyridine (140 mg, 0.31 mmol) in methanol (1 mL) at 0° C. The reaction mixture was stirred at room temperature overnight. The mixture was diluted with aqueous sodium bicarbonate and extracted with 10% methanol in dichloromethane. The organic layer was dried over magnesium sulfate and the solvent was removed under reduced pressure. The crude product was purified by column chromatography on silica (0-6% methanol in dichloromethane) to give the title compound as a yellow solid (30 mg, 29%). ES / MS m / z 336.27 (M+H)

[0154] Step 3: Preparation of 1-(7-ethynylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-yl 4-methylbenzenesulfonate

[0155] [ka] p-Toluenesulfonic anhydride (535 mg, 1.6 mmol), 4-(N,N-dimethylamino)pyridine (1 mg, 0.008 mmol), and triethylamine (0.1 mL) were added to a solution of 1-(7-((trimethylsilyl)ethynyl)benzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol (55 mg, 0.16 mmol) in dichloromethane (4 mL). The mixture was stirred at room temperature and monitored by LCMS until all the alcohol had been converted to the ditosylate product. The mixture was diluted with aqueous sodium bicarbonate and extracted with 10% methanol in dichloromethane. The organic layer was dried over magnesium sulfate, and the solvent was removed under reduced pressure. The crude product was purified by column chromatography on silica (0–6% methanol in dichloromethane). To the ditosylate intermediate was added potassium hydroxide (1N aqueous solution, 0.5 mL) and methanol:tetrahydrofuran (1:1, 2 mL). The mixture was stirred at room temperature for 1 h until all the ditosylate was converted to the desired compound. The pH of the mixture was adjusted by slow addition of aqueous hydrochloric acid (1N) and washed with aqueous sodium bicarbonate. The aqueous mixture was extracted with 10% methanol in methylene chloride. The organic solvent was removed under reduced pressure. The crude product was purified by column chromatography on silica (0-10% methanol in dichloromethane) to give the title compound as a pale yellow solid (16.9 mg, 25%). ES / MS m / z 418.15 (M+H).

[0156] Preparation of precursor 24 1-(7-(1-methyl-1H-pyrazol-4-yl)benzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-yl methanesulfonate

[0157] [ka] A stirred solution of 1-(7-bromobenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol (85 mg, 0.27 mmol), 1-methylpyrazole-4-boronic acid pinacol ester (112 mg, 0.54 mmol), and [1,1'-bis(di-tert-butylphosphino)ferrocene]dichloropalladium(II) (18 mg, 0.027 mmol) in dioxane (5 mL) and 2 M aqueous carbonate (1 mL) was sparged with nitrogen gas for 5 minutes and then heated at 100 °C for 3–4 hours. The reaction was diluted with 10% methanol in dichloromethane, filtered, and concentrated. The residue was dried under high vacuum for several hours and then dissolved in dichloromethane (50 mL). Triethylamine (0.4 mL) and methanesulfonyl chloride (154 mg, 1.35 mmol) were added at room temperature. After 1-2 hours at room temperature, the dichloromethane layer was washed with aqueous sodium bicarbonate, dried over sodium sulfate, filtered, and concentrated. The residue was purified by column chromatography on silica gel (0-20% methanol in dichloromethane) to give the title compound as a brown solid (24 mg, 22% yield over two steps). ES / MS m / z 397.8 (M+H).

[0158] Preparation of precursor 25 (S)-1-(4-chloro-8-methylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl)pyrrolidin-3-yl 4-methylbenzenesulfonate (P25):

[0159] [ka] To a solution of (S)-1-(8-methylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl)pyrrolidin-3-yl 4-methylbenzenesulfonate (20 mg, 0.048 mmol, 1.0 equiv.) in 10% methanol in methylene chloride (2 mL) was added N-chlorosuccinimide (8 mg, 0.062 mmol, 1.3 equiv.). The reaction was stirred at room temperature for 10 minutes and then washed with 1N sodium hydroxide (2 x 1 mL) and water (1 mL). The organic layer was concentrated. This material was purified on a 12 g silica gel column using a gradient of 0% to 40% ethyl acetate in methylene chloride. The product-containing fractions were concentrated to give a pale yellow solid (18 mg, 82% yield). ES / MS m / z (35 Cl / 37 Cl)456.3 / 458.1

[0160] Preparation of precursor 26 1-(8-methylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-yl 4-methylbenzenesulfonate

[0161] [ka] Precursor 26 was prepared in 81% yield from 1-(4-chloro-8-methylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-yl 4-methylbenzenesulfonate essentially following the procedure for precursor 25. ES / MS m / z (35 Cl / 37 Cl)442.2 / 444.1

[0162] Preparation of each example Radiolabeling procedure Radiolabeled syntheses were performed using a GE TRACERlab FXF-N automated radiosynthesizer with starting activities ranging from 0.1 Ci to 2.4 Ci. Average synthesis times ranged from 60 ± 10 min, and average decay-corrected yields ranged from 12 to 54%.

[0163] The [18F]fluorinated active material was retained on a Sep-Pak Accell Plus QMA Carbonate Plus Light Cartridge (Waters, 40 mg of sorbent per cartridge, 40 μm particle size) and eluted into the reaction vessel using 0.8 mL of cryptand 2.2.2-K2CO3 solution [cryptand 2.2.2 (7 mg) and potassium carbonate (0.75 mg) in HO (0.4 mL) and acetonitrile (0.4 mL)].

[0164] The eluted active material was dried by heating at 70 °C under a stream of inert gas and vacuum for approximately 5 min. The temperature was then increased to 100 °C under vacuum for 5 min to yield anhydrous cryptand 2.2.2-K2CO3[18F]Fluoride. A solution of the precursor [0.5 mg–2.0 mg in DMSO (1–2 mL)] was added to the reaction vessel containing anhydrous cryptand 2.2.2-K2CO3[18F]Fluoride, and the resulting mixture was maintained at elevated temperature (100–150 °C) for 10–20 min. The crude reaction mixture was cooled (50–65 °C) and diluted by the addition of HO (2.5–3 mL). The diluted crude reaction mixture was loaded onto a semi-preparative HPLC column for purification using isocratic elution (Agilent ZORBAX Eclipse XDB-C18 9.4 × 250 mm, flow rate 4 mL / min; see Table 6 for details). The fractions isolated from the HPLC column contained the radiolabeled examples as shown in Table 6.

[0165] Each example listed in Table 6 below was prepared essentially according to Example 1 and as generally described in Scheme 4.

[0166] [Table 7-1]

[0167] [Table 7-2]

[0168] [Table 7-3]

[0169] [Table 7-4] * Precursor 20 is 18 It rearranged during the F-labeling reaction.

[0170] Preparation of extemporaneous injection formulation HPLC fractions containing radiolabeled examples, prepared essentially according to the procedure of Example 1, were diluted with 30-40 mL of 0.5% aqueous sodium ascorbate (w / v). The diluted solution was passed through either a Sep-Pak® C18 Plus Light Cartridge (Waters, 130 mg of sorbent per cartridge, particle size 55-105 μm) or a Sep-Pak® Vac C18 Cartridge (Waters, 50 mg of sorbent per cartridge, particle size 55-105 μm), and the retained material was washed with 5-10 mL of water. The material was eluted from the cartridge using dehydrated alcohol USP (0.4-1 mL) and combined with a solution of 0.5% sodium ascorbate (w / v) in 0.9% sodium chloride injection USP to give the product (material in 10% ethanol (v / v), 0.45% (w / v) sodium ascorbate in 0.9% sodium chloride injection USP, total volume 4-10 mL).

[0171] Product samples were removed and analyzed by HPLC to determine radiochemical purity, radiochemical character, chemical purity, and specific activity. Peak(s) from the UV chromatogram and radiochromatogram were integrated to determine radiochemical purity (%RCP), radiochemical character, chemical purity, and specific activity. RCP ranged from 92-100%. Stability data for selected examples is shown in Table 7 below.

[0172] [Table 8]

[0173] The examples in Table 8 below were prepared essentially according to Scheme 1, Method A (Step 1).

[0174] [Table 9]

[0175] Each example listed below in Table 9 was prepared essentially according to Scheme 2, Method C.

[0176] [Table 10]

[0177] Each example in Table 10 below was prepared essentially according to Method D in Scheme 2.

[0178] [Table 11]

[0179] Each example in Table 11 below was prepared essentially according to Method E in Scheme 2.

[0180] [Table 12]

[0181] Each example in Table 12 below was prepared essentially according to the procedure for Precursor 25.

[0182] [Table 13]

[0183] Example 5 3-(3-fluoroazetidin-1-yl)-7-iodobenzo[4,5]imidazo[1,2-a]pyridine

[0184] [ka]

[0185] Step 1: Preparation of 3-(3-fluoroazetidin-1-yl)-7-(tributylstannyl)benzo[4,5]imidazo[1,2-a]pyridine

[0186] [ka] The title compound was prepared in 46% yield from 7-bromo-3-(3-fluoroazetidin-1-yl)benzo[4,5]imidazo[1,2-a]pyridine using essentially the same procedure as in Step 1 of Precursor 21. ES / MS m / z 531.0 (M+H)

[0187] Step 2: Preparation of 3-(3-fluoroazetidin-1-yl)-7-iodobenzo[4,5]imidazo[1,2-a]pyridine

[0188] [ka] The title compound was prepared in 64% yield from 3-(3-fluoroazetidin-1-yl)-7-(tributylstannyl)benzo[4,5]imidazo[1,2-a]pyridine using essentially the same procedure as in Step 2 of Precursor 21. ES / MS m / z 367.9 (M+H)

[0189] Example 19 7-Ethyl-3-(3-fluoroazetidin-1-yl)benzo[4,5]imidazo[1,2-a]pyridine

[0190] [ka] 2-Nitrobenzenesulfonyl chloride (75.0 mg, 0.337 mmol, 3.0 equiv.) was added to a slurry of 3-(3-fluoroazetidin-1-yl)-7-vinylbenzo[4,5]imidazo[1,2-a]pyridine (30.0 mg, 0.112 mmol, 1.0 equiv.) and hydrazine hydrate (43.0 μL, 0.56 mmol, 5.0 equiv.) in methylene chloride (3 mL) and dimethylformamide (1 mL) at room temperature. The reaction mixture gradually cleared to an orange solution, and the solution was stirred for 16 h. The reaction mixture was concentrated onto silica gel and purified by column chromatography on silica gel (0–10% methanol in methylene chloride) to give the product, contaminated with an aromatic by-product. The yellow solid was dissolved in methylene chloride and washed with saturated aqueous sodium bicarbonate and water. The organic layer was dried over magnesium sulfate and concentrated. The crude solid was dissolved in methylene chloride and purified by column chromatography on silica gel (0-10% methanol in methylene chloride) to give the title compound as a beige solid (14.0 mg, 0.052 mmol, 46%). ES / MS m / z 270.1 (M+H).

[0191] Example 21 8-Fluoro-3-(3-fluoroazetidin-1-yl)-7-vinylbenzo[4,5]imidazo[1,2-a]pyridine

[0192] [ka] 7-Bromo-8-fluoro-3-(3-fluoroazetidin-1-yl)benzo[4,5]imidazo[1,2-a]pyridine (0.100 g, 0.30 mmol), vinyl pinacolborane (0.091 g, 0.59 mmol), potassium carbonate (0.122 g, 0.89 mmol), and dichloro[1,1'-bis(di-tert-butylphosphino)ferrocene]palladium(II) (Pd-118) (0.019 g, 0.030 mmol) were added. Dimethoxyethane / ethanol / water (7 / 2 / 1, 6.0 mL) and dimethyl sulfoxide (0.4 mL) were added. The reaction mixture was heated at 80 °C overnight. The reaction mixture was partitioned between ethyl acetate and water. The organic layer was separated, washed with brine, and dried over sodium sulfate. The solid was filtered and washed with dichloromethane. The solvent was removed in vacuo. Purification was carried out by column chromatography on silica gel (0-95% ethyl acetate in methylene chloride) to give the title compound as a brown solid (46 mg, 54%). ES / MS m / z 286.2 (M+H).

[0193] Example 22 3-(3-(2-fluoroethoxy)azetidin-1-yl)-7-vinylbenzo[4,5]imidazo[1,2-a]pyridine

[0194] [ka] To a solution of 1-(7-vinylbenzo[4,5]imidazo[1,2-a]pyridin-3-yl)azetidin-3-ol (27 mg, 0.1 mmol) in dimethylformamide (3 mL) was added 1-bromo-2-fluoroethane (38 mg, 0.3 mmol, 3 equiv.), followed by potassium tert-butoxide (34 mg, 0.3 mmol, 3 equiv.). The reaction was stirred at room temperature for 72 hours. Additional 1-bromo-2-fluoroethane (63 mg, 0.5 mmol, 5 equiv.) was added, followed by potassium tert-butoxide (67 mg, 0.6 mmol, 6 equiv.). The reaction was stirred at room temperature for 15 minutes, then cooled to 0 °C (ice bath), and water (3 mL) was added. The aqueous phase was extracted with dichloromethane (4 x 5 mL), concentrated in vacuo, and then purified by column chromatography on silica gel (12 g column, 0% to 30% methanol in dichloromethane) to give the title compound as a brown solid (17 mg, 55%). ES / MS m / z 312.2 (M+H).

[0195] Example 30 7-ethynyl-3-(3-fluoroazetidin-1-yl)benzo[4,5]imidazo[1,2-a]pyridine

[0196] [ka] Potassium hydroxide solution (1N, 1 mL) was added dropwise to a solution of 3-(3-fluoroazetidin-1-yl)-7-((trimethylsilyl)ethynyl)benzo[4,5]imidazo[1,2-a]pyridine (20 mg, 0.07 mmol) in methanol:tetrahydrofuran (2 mL 1:1) at 0°C. The reaction was stirred at room temperature for 1.5 hours. The reaction mixture was neutralized with hydrochloric acid (1N) followed by aqueous sodium bicarbonate. The aqueous layer was extracted with dichloromethane (3x). The combined organic layers were dried over magnesium sulfate and the solvent removed under reduced pressure. The crude product was purified by column chromatography on silica (0-10% methanol in dichloromethane) to give the title compound as a yellow solid (41 mg, 31% yield over two steps). ES / MS m / z 266.14 (M+H).

[0197] Example 17 3-(3-fluoroazetidin-1-yl)-7-(1-methyl-1H-pyrazol-4-yl)benzo[4,5]imidazo[1,2-a]pyridine

[0198] [ka] A stirred solution of 7-bromo-3-(3-fluoroazetidin-1-yl)benzo[4,5]imidazo[1,2-a]pyridine (48 mg, 0.15 mmol), 1-methylpyrazole-4-boronic acid pinacol ester (62 mg, 0.3 mmol), and [1,1'-bis(di-tert-butylphosphino)ferrocene]dichloropalladium(II) (10 mg, 0.015 mmol) in dioxane (5 mL) and 2 M aqueous carbonate solution was sparged with nitrogen gas for 5 minutes and then heated at 100 °C for 3–4 hours. The reaction was cooled, diluted with 10% methanol in dichloromethane, filtered, and concentrated. The residue was purified by column chromatography on silica gel (0–20% methanol in dichloromethane) to give the title compound as a solid (30 mg, 62%). ES / MS m / z 322.0 (M+H).

[0199] biological analysis Assay Example 31: K of the compound of Example 3 using tau from a human donor with Alzheimer's disease i and K. d decision.

[0200] Preparation of Paired Helical Fiber (PHF) Tau: Purified soluble PHF tau was isolated from AD brain tissue using a protocol modified from that described by Jicha, et al. (G.A. Jicha, A. O'Donnell, C. Weaver (1999) "Hierarchical phosphorylation of recombinant tau by the paired-helical filament-associated protein kinase is dependent on cyclic AMP-dependent protein kinase," J Neurochem. 72(1):214). Briefly, AD cortices were homogenized using a handheld Kinematica Polytron followed by high-pressure batch gas expansion using a Parr cell disruption bomb. The crude homogenate was centrifuged at 28 kg to pellet cellular debris. Soluble PHF tau was isolated from the supernatant by affinity chromatography on an Affigel-10 column immobilized with the tau antibody MC1, which recognizes the pathological conformation of tau (GA Jicha, R. Bowser, IG Kazam (1997), "Alz-50 and MC-1, a new monoclonal antibody raised to paired helical filaments, recognizes conformational epitope on recombinant tau," J Neurosci Res. 48(2):12.).

[0201] Example 3: Determination of Ki for 3-(3-fluoroazetidin-1-yl)-7-methylbenzo[4,5]imidazo[1,2-a]pyridine

[0202] I C 50The molar concentration of competing ligand that reduces specific binding of the radioligand by 50% is defined as the molar concentration of competing ligand that reduces specific binding of the radioligand by 50%. In this competition assay, the competing ligand was the non-radiolabeled compound of Example 3, and the radiolabeled compound, i.e., radioligand, was 7-[6-(F)fluoropyridin-3-yl]-5H-pyrido[4,3-B]indole (also known as [F]AV-1451 or T807). Binding of [F]AV-1451 to PHF tau was determined against various concentrations of the non-radiolabeled compound of Example 3. The reaction mixture (200 μl) contained PHF tau (0.12 μg), 0.1 to 0.5 nM [F]AV-1451, and serial dilutions of the non-radiolabeled compound of Example 3 from 316 nM to 0.01 nM. Assays were performed in 96-well polypropylene microplates in PBS, pH 7.4, containing 0.01% bovine serum albumin. Nonspecific binding was defined as radioligand binding in the presence of 2-[4-(2-fluoranylethyl)-1-piperidyl]pyrimido[1,2-a]benzimidazole T808 / AV-680 (5 μM), a known ligand for PHF tau (Zhang, J. (2012), "A highly selective and specific PET tracer for imaging of tau pathologies," J Alzheimers Dis., 31(3):A,C.601). After 1.5 hours of incubation at 37°C, bound radioactivity was analyzed using a Millipore MultiScreen™ PET filter. HTS Millipore MultiScreen using a Vacuum Manifold HTS The cells were harvested onto 96-well glass fiber FB filter plates, followed by five washes with PBS, pH 7.4. Filters containing bound [18F]AV-1451 were assayed for radioactivity in a Wizard 2480 automated gamma counter (Perkin Elmer). Using these assay conditions, the total bound fraction is typically less than 10% of the added radioligand [18F]AV-1451. IC 50 is determined using ActivityBase or XLfit model 205 (or equivalent), where: y=A+(BA) / (1+((C / x)^D) Y=inhibition rate (%) X = concentration of cold competing ligand (nM) A = Minimum Y (0%) B=Max Y(100%) C=IC50 D = slope coefficient

[0203] Ki (i.e., the equilibrium dissociation constant for binding of non-radiolabeled compounds) was calculated using the Cheng-Prusoff equation as the IC 50 Calculate from the value (Cheng Y., Prusoff WH (1973), "Relationship between the inhibition constant (KI) and the concentration of inhibitor which causes 50 percent inhibition (I 50 ) of an enzymatic reaction”Biochem Pharmacol 22(23):3099-3108). Ki = IC50 / (1+[L] / Kd) [L] = concentration of [18F]AV-1451 (typically about 0.5 nM) K d = Dissociation constant of [18F]AV-1451 (0.57 nM).

[0204] The K of the compound of Example 3 (3-(3-fluoroazetidin-1-yl)-7-methylbenzo[4,5]imidazo[1,2-a]pyridine) against [F]AV-1451 was 0.23 nM for PHF tau obtained from a donor with Alzheimer's disease, indicating that the compound of Example 3 binds to PHF tau. Therefore, PET imaging using the compound of Example 3 and examination of the imaging pattern are useful for detecting the presence of PHF tau in patients and may be helpful in confirming the diagnosis of AD or non-AD tauopathy.

[0205] The Example compounds disclosed herein exhibited a K of less than 3 nM as assessed according to this assay. Thus, the Example compounds bind to PHF tau, and radiolabeled Example compounds are useful for detecting PHF tau.

[0206] Determination of Kd for the compound of Example 3 (3-(3-[18F]-fluoroazetidin-1-yl)-7-methylbenzo[4,5]imidazo[1,2-a]pyridine)

[0207] The dissociation constant [Kd] of radiolabeled Compound of Example 3 (18F) was determined by saturation binding, and the total and nonspecific binding of the radioligand were measured at various radioligand concentrations. The reaction mixture (250 μl) contained PHF tau (0.15 μg) serially diluted in PBS from 25 nM to 0.3 nM, and the compound of Example 3. The assay was performed in 96-well polypropylene microplates in PBS containing 0.01% bovine serum albumin. Nonspecific binding was defined as the binding of the radioligand in the presence of 2-[4-(2-fluoranylethyl)-1-piperidyl]pyrimido[1,2-a]benzimidazole (also known as T808 / AV-680) (10 μM). The radioligand in this assay was the compound of Example 3. After 1.5 hours of incubation at 37°C, bound radioactivity was collected by vacuum filtration onto Millipore MultiScreenHTS 96-well glass fiber FB filter plates using a Millipore MultiScreenHTS Vacuum Manifold, followed by five washes with PBS. Filters containing bound Example 3 compound were assayed for radioactivity in a Wizard 2480 automated gamma counter (Perkin Elmer). Using these assay conditions, the percent total binding is typically less than 10% of the added radioligand. Total and nonspecific binding data were analyzed by nonlinear regression analysis using Graphpad Prism to determine the Kd of the radioligand.

[0208] The Kd of the compound of Example 3 was 1.5±0.2 nM for PHF tau obtained from Alzheimer's disease donors, indicating that this compound binds to PHF tau with high affinity. Therefore, PET imaging and examination of the imaging pattern using this compound are useful for detecting the presence of tau in patients and can confirm the diagnosis of AD or non-AD tauopathy.

[0209] Assay Example 32 The compound of Example 3 ( 18 F) Measurement of binding Kd The compound of Example 3 ( 18Autoradiography was used to measure the Kd of [(18)F]T807, a novel tau positron emission tomography imaging agent for Alzheimer's disease. Xia CF, et al., Alzheimer's Dement. 2013 Nov;9(6):666-76 (see Zhang, J. (2012), "A highly selective and specific PET tracer for imaging of tau pathologies," J Alzheimers Dis., 31(3):601). In this experiment, 15 adjacent frontal lobe sections from each of two AD brains, one with tau-rich and amyloid-rich (tau+Aβ+) and one with tau-poor and amyloid-rich (tau-Aβ+), were used to define nonspecific binding. Each section was covered with 0.5 mL of 3-(3-[18F]-fluoroazetidin-1-yl)-7-methylbenzo[4,5]imidazo[1,2-a]pyridine serially diluted from approximately 250 nM in binding buffer (2.5% dimethyl sulfoxide + 2.5% ethanol in 1x PBS, pH 7.4). After 60 min of incubation at room temperature, unbound ligand was removed by successive wash cycles (2 min in 1x PBS, 2 min in 30% ethanol in 1x PBS, 2 min in 70% ethanol in 1x PBS, and 2 min in 1x PBS). After drying under a hood, each section was exposed overnight to a phosphorimaging screen. The autoradiographic signals recorded on the phosphorimaging screen were read using a GE Healthcare Life Sciences Typhoon FLA 7000 Phosphorimager. Signal intensity over the gray matter was measured using Fujifilm Multi Gauge software. The Kd of the compounds was determined by nonlinear regression analysis of the bound concentration of the compound of Example 3 versus the concentration of the free compound.

[0210] Compound of Example 3 on AD brain slices for Kd determination ( 18An autoradiography from F) is shown in Figure 1. The Kd of the compound of Example 3 for native tau aggregates in AD brain tissue, as determined by nonlinear regression analysis, was 1.7 nM, indicating that this compound binds to tau. 18 F) PET imaging and imaging patterns can be useful in detecting the presence of tau in patients and can confirm the diagnosis of AD or non-AD tauopathy.

[0211] Assay Example 33 Selectivity of 3-(3-[18F]-fluoroazetidin-1-yl)-7-methylbenzo[4,5]imidazo[1,2-a]pyridine for tau and beta-amyloid in AD human brain tissue. method Based on the results of anti-tau and anti-amyloid immunostaining of the brain sections, three groups of human brain sections were selected for autoradiography experiments to determine the binding selectivity of the compound of Example 3 to pathological tau. Figure 2 shows the autoradiography of the compound of Example 3 on the three groups of human brain sections. Group A is AD brain sections rich in pathological tau (labeled as tau+Aβ+), Group B is AD brain sections with little pathological tau (labeled as tau-Aβ+), and Group C1 is normal brain sections with tau-Aβ-. As shown in Figure 2, the human AD brain sections used in Group A are #0185, #28770, #30121, #30311, and #30461. The human AD brain sections used in Group B are #33562, #32656, #33998, #35682, and #33563. The normal human brain sections in group C are #29092 and #32566. Selectivity was calculated using tissue sections from the same donor. Beta-amyloid burden was quantified in each of these three groups of brains by autoradiography on adjacent 10 μm sections using the amyloid tracer [18F]W372 (2-(6-fluoro-3-pyridyl)-6-methoxy-imidazo[1,2]thiazolo[2,3-c]pyridine). [18F]W372 is a selective amyloid binding tracer discovered by Siemens and evaluated in IND105173 (see also US 9,023,316). Each section was covered with 0.5 ml of binding buffer (2.5% dimethyl sulfoxide + 2.5% ethanol in 1×PBS, pH 7.4, approximately 20 μCi / slide) containing the compound of Example 3 and incubated for 60 minutes. Unbound tracer was then removed using successive washing cycles (2 min PBS, 2 min 30% EtOH / PBS, 2 min 70% EtOH / PBS, 2 min PBS). Sections were air-dried, placed on phosphor imaging plates (Fuji IP plates), and exposed overnight. IP plates were read using a GE Healthcare Life Sciences Typhoon FLA 7000 Phosphorimager. Signal intensity on the gray matter was measured using Fujifilm Multi Gauge software.After subtracting the background signal (the signal in the cortical region of Group C), the signals of each section of Group A and Group B were normalized with the corresponding signal from the autoradiography of each adjacent section using [F]W372. Calculations were performed based on Group B brain sections #32656 and #33998, in which pathological tau was not detectable by immunohistochemistry. The normalized signal of the brain section of Group B is the relative signal level of the compound of Example 3 relative to [F]W372 resulting from binding to native β-amyloid aggregates. The binding level to native tau aggregates in Group A sections was estimated by subtracting the amount of total signal resulting from binding to β-amyloid (calculated by multiplying the total signal from [F]W372 binding to β-amyloid in adjacent sections by the relative signal of the compound of Example 3 relative to [F]W372 determined from the section of Group B). The resulting difference was then divided by the signal due to binding to β-amyloid to estimate selectivity.

[0212] A strong signal was observed in the gray matter (cortical region) of the sections from group A (tau+Aβ+), whereas a weak or no signal was detected in the cortical region of the sections from group B (tau-Aβ+). No autoradiographic signal was observed in the normal brain sections from group C (tau-Aβ-). These results indicate that the compound of Example 3 specifically binds to native tau aggregates in human AD brains and has weak or no interaction with native β-amyloid aggregates.

[0213] Because the IHC results showed that brain sections #32656 and #33998 from Group B were free of tau protein aggregates, the normalized autoradiography signals in the cortical regions of these AD brain sections result from the binding of the compound of Example 3 to native β-amyloid aggregates. The selectivity of the binding of the compound of Example 3 to native tau aggregates relative to native β-amyloid aggregates is reflected in the ratio of the signal from Group A (tau+Aβ+) to the average signal from brain sections #32656 and #33998.

[0214] The compound of Example 3 contains five tau + Aβ + Based on the brain specimen and two tau-Aβ+ brain specimens, the tau:Aβ selectivity ratio is approximately 31.4, and the gray matter to white matter (GM / WM) signal ratio is approximately 18.9. The autoradiography signal of the compound of Example 3 on normal brain sections is weak and uniform, showing little difference between gray matter and white matter, indicating low non-specific binding. The selectivity ratio of the two native tau aggregates compared to the binding of native β-amyloid aggregates in the gray matter region of human AD brains was observed to be approximately 31-fold.

[0215] The results provided in the above biological assays support the use of the compound of Example 3 as a radiolabeled example that can be used in conjunction with a PET imaging probe to detect levels of aggregated tau protein in patients with AD and / or other neurodegenerative diseases, such as CTE. These results also suggest the use of compounds of other examples as useful PET imaging probes for aggregated tau protein to aid in diagnosing and monitoring patients with AD and other neurodegenerative disorders associated with aggregated tau protein.

[0216] Assay Example 34 The compound of Example 4 ( 18 Autoradiographic determination of F) binding The compound of Example 4 ( 18Autoradiography was used to measure tau positron emission tomography (PET) ([(18)F]T807, a novel tau positron emission tomography imaging agent for Alzheimer's disease. Xia CF, et al., Alzheimer's Dement. 2013 Nov;9(6):666-76) (see Zhang, J. (2012), "A highly selective and specific PET tracer for imaging of tau pathologies," J Alzheimers Dis., 31(3):601). Experiments used 10 μm adjacent sections from clinically diagnosed cases of AD, PSP, or CBD confirmed as tau-positive by IHC using AT8 or AT100 antibodies using standard techniques, or from control tissues defined as amyloid- and tau-negative by IHC. Each section was analyzed using the method described in Example 4 [ 18 The sections were coated with [F] (40 μCi / ml in binding buffer (2.5% dimethyl sulfoxide + 2.5% ethanol in 1x PBS, pH 7.4)). After 60 min of incubation at room temperature, unbound ligand was removed by successive washing cycles (2 min in 1x PBS, 2 min in 30% ethanol in 1x PBS, 2 min in 70% ethanol in 1x PBS, and 2 min in 1x PBS). After drying under a hood, each section was exposed to a phosphorimaging screen overnight. The autoradiographic signal recorded on the phosphorimaging screen was read using an Amersham Typhoon Bio-Imaging System. Individual tissue samples were compared with adjacent slices using either AT8 or AT100. A positive correlation with the tau antibody indicates binding to the non-AD tau (PSP or CBD) being tested.

[0217] The compound of Example 4 ( 18 F) Autoradiography is shown in Figure 3. Sections from PSP patients were 18The autoradiographic signals were comparable to those seen by IHC with F-labeled AT100 antibody or AT8. ARG positivity was observed in areas rich in AT8-positive tufted astrocytes, a classic neuropathological feature observed in PSP.

[0218] The compound of Example 4 ( 18 An autoradiography of Example 3 compound (F) is shown in Figure 4. A strong signal was observed in tau-positive tissue from CBD patients. Note the binding in white matter regions of CBD patients, which have been reported to have abundant tau deposits and therefore tau aggregate-rich regions. The presence of an ARG signal for Example 3 compound correlates with tau-positive regions in PSP and CBD human tissue, indicating that this compound binds to non-AD tau. Both Figures 3 and 4 show the binding of Example 4 compound (F) to human AD tissue. 18 F), showing strong binding to AD tau. 18 PET imaging and imaging patterns using F) are useful in detecting the presence of AD and non-AD tau in patients and can confirm the diagnosis of AD or non-AD tauopathy.

Claims

1. A compound of the following formula: or a pharmaceutically acceptable salt thereof: 【Chemistry 1】 [In the formula, n is 0, 1, or 2; R １ is H, halo, C １ ~C ３ Alkyl, or C ３ ~C ６ is cycloalkyl, R ２ is H, halo, C １ ~C ３ Alkyl, C ２ ~C ３ Alkenyl, C ２ ~C ３ Alkynyl, —O—(C １ ~C ４ alkyl), C １ ~C ４ Alkyl halo, C ２ ~C ４ alkenylhalo, or 【Chemistry 2】 and R ３ is H or F, R ４ , F, １８ F, C １ ~C ４ -Alkyl F, C １ ~C ４ -Alkyl １８ F, —O—(C １ ~C ４ -alkyl)F, or -O-(C １ ~C ４ -alkyl) １８ F, R ５ is H, halo, or C １ ~C ４ is alkyl, where n is 0 and R １ is methyl, and R ２ is H and R ３ is H and R ５ is H, then R ４ is F or １８ Not F].

2. n is 0, 1, or 2; R １ is H, halo, C １ ~C ２ alkyl, or cyclopropyl; R ２ は、H、ハロ、-CH ３ 、-CH ２ CH ３ ,-OCH ３ 、-EH=EH ２ 、-CF=CH ２ 、 【Transformation 3】 -OCH ２ CH ２ F, or 【Chemistry 4】 and R ３ is H or F, R ４ is F, １８ F, -CH ２ F, -CH ２ １８ F, -CH ２ CH ２ F, -CH ２ CH ２ １８ F, -CH ２ CH ２ CH ２ F, -CH ２ CH ２ CH ２ １８ F, -OCH ２ CH ２ F, or -OCH ２ CH ２ １８ F, and R ５ is H, halo, or C １ ~C ４ 2. The compound of claim 1, or a pharmaceutically acceptable salt thereof, wherein: R is alkyl;

3. n is 0 or 1; R １ is H, halo, C １ ~C ２ alkyl, or cycloalkyl; R ２ は、H、ハロ、-CH ３ ,-OCH ３ 、-EH=EH ２ 、-CF=CH ２ 、 【Transformation 5】 -OCH ２ CH ２ F, or 【Transformation 6】 and R ３ is H or F, R ４ is F or １８ F, R ５ is H, halo, or —CH ３ 3. The compound of claim 1 or 2, or a pharmaceutically acceptable salt thereof:

4. n is 0, R １ is H, halo, -CH ３ , or -CH ２ CH ３ and R ２ は、H、ハロ、-CH ３ ,-OCH ３ 、-EH=EH ２ 、-CF=CH ２ 、 【Transformation 7】 or 【Transformation 8】 and R ３ is H or F, R ４ is F or １８ F, R ５ is H, halo, or —CH ３ 3. The compound of claim 1 or 2, or a pharmaceutically acceptable salt thereof:

5. R １ is -CH ３ and R ２ is H, R ３ is H, R ５ 5. The compound or pharmaceutically acceptable salt of claim 4, wherein:

6. n is 0, R １ , R ２ , R ３ , and R ５ are H, respectively, R ４ is F or １８ 5. The compound of claim 4, wherein R is H or a pharmaceutically acceptable salt thereof.

7. n=0, R １ is H, R ２ is Cl, F or I, R ３ is H, R ４ is F or １８ F, R ５ or a pharmaceutically acceptable salt thereof.

8. n is 0, R １ , R ３ , and R ５ are H, respectively, R ２ は、-CH ３ 、-CH ２ CH ３ 、-CF=CH ２ 、 【Chemistry 9】 F, Cl, or I; R ４ is F or １８ 2. The compound of claim 1, or a pharmaceutically acceptable salt thereof, wherein:

9. n is 0, R １ is -CH ３ or -CH ２ CH ３ and R ２ , R ３ and R ５ are H, respectively, R ４ is F or １８ 2. The compound of claim 1, or a pharmaceutically acceptable salt thereof, wherein:

10. n is 2, R １ is H, R ２ is H or -CH ３ and R ３ is H, R ４ , F, １８ F, C １～ C ３ Alkyl F, C １～ C ３ Alkyl １８ F, R ５ or a pharmaceutically acceptable salt thereof.

11. The compound is Table 1-1 Table 1-2 2. The compound of claim 1, or a pharmaceutically acceptable salt thereof:

12. A pharmaceutical composition comprising a compound according to any one of claims 1 to 11 or a pharmaceutically acceptable salt thereof, and one or more pharmaceutically acceptable carriers or diluents.

13. 13. The pharmaceutical composition of claim 12, wherein the carrier comprises ethanol, water, and a buffer suitable for injection into a patient.

14. 1. A method for imaging aggregated tau in a mammal, comprising: １８ introducing into a mammal a detectable amount of a compound of any one of claims 1 to 11, or a pharmaceutically acceptable salt thereof, having an F substituent; allowing sufficient time for the compound to bind to aggregated tau; and detecting said compound.

15. 15. The method of claim 14, wherein the mammal is a human.

16. 15. The method of claim 14, wherein the mammal is a human and is suspected of having a neurological disease or disorder.

17. 17. The method of claim 16, wherein the human is suspected of having Alzheimer's disease.

18. 17. The method of claim 16, wherein the human is suspected of having progressive supranuclear palsy (PSP), corticobasal degeneration (CBD), Pick's disease (PiD), atypical Alzheimer's disease, chronic traumatic encephalopathy (CTE), and frontotemporal dementia (FTD).

19. 17. The method of claim 16, wherein the human is suspected of having early Alzheimer's disease tau.

20. 17. The method of claim 16, wherein the human is suspected of having non-Alzheimer's disease tau.

21. A compound of the following formula: or a pharmaceutically acceptable salt thereof: 【Chemistry 10】 [In the formula, n is 0, 1, or 2; R １ is H, halo, C １ ~C ３ Alkyl, or C ３ ~C ６ is cycloalkyl, R ２ is H, halo, C １ ~C ３ Alkyl, C ２ ~C ３ Alkenyl, C ２ ~C ３ Alkynyl, —O—(C １ ~C ４ alkyl), C １ ~C ４ Alkyl halo, C ２ ~C ４ alkenylhalo, or 【Chemistry 11】 and R ３ is H or F, L is a bond, C １ ~C ４ -Alkyl F, C １ ~C ４ -Alkyl １８ F, —O—(C １ ~C ４ -alkyl)F, or -O-(C １ ~C ４ -alkyl) １８ F, LG is a leaving group, R ５ is H, halo, or C １ ~C ４ is alkyl, where n is 0 and R １ is methyl, and R ２ is H and R ３ is H and R ５ is H, then L is not a bond.

22. The LG is —NO ２ , trialkylamine, alkylsulfonate, arylsulfonate, mesylate group, tosylate group, triflate group, Cl, Br, I, or nosylate group, or a pharmaceutically acceptable salt thereof.

23. 19. The compound or pharmaceutically acceptable salt of claim 18, wherein the leaving group is mesylate or tosylate.

Images