18 F and 11 C labeled MCHR1 PET ligands

Abstract

The present invention provides radiolabeled 2,3,4,5-tetrahydro-1H-[1,4]diazepino[1,7-α]indol-9-yl]pyridin-2(1H)-one derivatives of general formula (I) or salts thereof, as well as precursors of the radioligands of general formulae (II) and (III), processes for their preparation, and intermediates of the processes. The radiolabeled compounds enable visualization and quantification of melanin-concentrating hormone receptor 1 (MCHR1) in various target tissues, including monitoring MCHR1 in such target tissues. The compounds exhibit high binding affinity for MCHR1 and are able to cross the blood-brain barrier, thus facilitating diagnosis of various disease states in human clinical studies.

Description

Technical Field

[0001] This invention relates to 2,3,4,5-tetrahydro-1-dimethylformyl chloride of general formula (I) for radiolabeling. H -[1,4]diaza [1,7-α]indol-9-yl]pyridine-2(1 H (II)-ketone derivatives and / or their salts, which can be used to bind and image melanin aggregation hormone receptor 1 (MCHR1) in the mammalian brain. Furthermore, this invention relates to certain precursors of formula (II) of the radioligands, methods for their preparation, and intermediates of said methods. Background Technology

[0002] Many medical diagnostic procedures use radiolabeled compounds. PET (positron emission tomography) is a highly sensitive technique that requires small amounts of radiolabeled compounds called tracers. The labeled compounds are absorbed, distributed, metabolized, and eliminated from the body in the same way as their corresponding non-radioactive counterparts. Tracers can be radiolabeled using radionuclides that can be used for PET imaging, such as… 11 C 13 N、 18 F, 64 Cu、 68 Ga、 82 Rb and 124 I. PET methods are used to detect functional impairments at the cellular level in tissue and organ studies. PET has been used in clinical oncology (imaging tumors and metastases), identifying specific brain diseases, and mapping brain and heart function.

[0003] Carbon-11 (t) 1 / 2 Carbon-11 (=20.3 min) is one of the most commonly used radioisotopes in PET because it is abundant in organic molecules and has a short half-life, allowing for multiple administrations of the tracer to the same mammal (i.e., human or animal subject) on the same day and reducing the radiation burden on patients. Radiolabeling target molecules with carbon-11 is more challenging because of its short half-life, which precludes long-duration synthesis and multi-step reactions. Some effective strategies must consider the radiochemistry of carbon-11, including designing reaction methods such that the carbon-11 radionuclide should be introduced in the final step. Purification is then performed by HPLC. 11 C-tracers significantly increase tracer loss and production time, and complicate automation.

[0004] Fluorine-18 (t) 1 / 2 =109.7 min) is another widely used PET isotope, whose radiochemistry has recently made progress with the development of kit-type reagent kits that do not require HPLC purification. By employing solid-phase extraction (SPE) columns, these completely disposable kits allow for reliable routine production.18 F-tracers include [ 18 F]FDG and other similar technologies offer shorter synthesis times, reduced human involvement, and minimal equipment maintenance. Their development will significantly improve synthesis reliability, increase radiochemical yields, and simplify the automation and preventative maintenance of production modules.

[0005] Melanin-gathering hormone (MCH) is a cyclic polypeptide composed of 19 amino acids, primarily produced by neurons in the lateral hypothalamus and the zone of indeterminate hypothalamus, with projections widely distributed throughout the entire brain (Bittencourt). Gen Comp Endocrinol , 2011, 172: 185-97). The biological effects of MCH are mediated by two G protein-coupled receptors (GPCRs) called MCH receptor 1 (MCHR1) (Saito et al., 2011, 172: 185-97). Nature , 1999, 400: 265-69; Shimomura et al., Biochem Biophys Res Commun , 1999, 261: 622-26) and MCH receptor 2 (MCHR2) (Sailer et al. ... Proc Natl Acad Sci USA (2001, 98: 7564-69). Since only MCHR1 is functional in rodents, the physiological importance of MCHR2 remains unknown due to the lack of suitable animal models.

[0006] The distribution of MCH and the expression of MCHR1 in brain regions unrelated to nutritional behavior have led to the discovery that MCH signaling is also involved in various mental illnesses, such as depression and anxiety (Smith et al.). Neuropsychopharmacology , 2006, 31: 1135-45). Given that both ependymal cells and MCH neurons are involved in glucose sensing (Guyon et al., 2006, 31: 1135-45). J Neurosci (2009, 29: 2528-33) MCH fibers can control the activity of ciliated cells to initiate an increase in cerebrospinal fluid (CSF) flow to meet metabolic demands. This strongly supports the idea that the MCH system may also be involved in non-neuronal intercellular communication, but evidence is still lacking.

[0007] Antagonism of MCHR1 is one of the feasible targets for obesity treatment (Pissios, Peptides 2009, 30: 2040-44). Among the orexin peptides in the hypothalamus, MCH is the only known peptide whose ablation leads to weight loss (Rivera et al., 2009, 30: 2040-44). Curr Med Chem, 2008 (15:1025-43). However, in order to enable quantitative in vivo assessment of MCHR1 pharmacology and facilitate preclinical-to-clinical translation, suitable PET tracers need to be developed.

[0008] SNAP-7941 is a potent MCHR1 antagonist that contains methyl ester ( Figure 1 Compound A), making its molecules suitable for introduction [ 11 C]Methyl structural portion or [ 18 F]Fluoroethyl structural moiety (Borowsky et al.) Nat Med Based on this, potential PET tracers for MCHR1 visualization were developed, such as […]. 11 C]SNAP-7941 Figure 1 (compound B) and [ 18 F]FE@SNAP Figure 1 Compound C). These tracers have some drawbacks: [ 11 [C]SNAP-7941 is a PGP substrate, therefore its uptake in the brain is low; 18 F]FE@SNAP has no selectivity for MCHR2; furthermore, both compounds exhibit low stability in rat plasma (Philippe et al., Nucl Med Biol , 2013, 40: 919-25; Philippe et al., Mol Imag Biol , 2019, 21: 257-68; Philippe et al., Sci Pharm , 2013, 81: 625-39).

[0009] Takeda reported a carbon-11 MCHR1 radioligand ( Figure 1 Compound D), which is also a substrate of efflux transport proteins in rats (Igawa et al., Curr Radiopharm (Panfen et al., 2017, 10:35-40). Limited brain uptake was increased after pretreatment with cyclosporine A (CSA) (a drug transporter inhibitor) (Panfen et al.). Drug Met. Disp. , 2019, 47(11): 1352-60; Yang et al., Int. J. Mol. Sci. , 2020, 21(19): 7023-50).

[0010] Wanyou Pharmaceuticals reported a selective and effective MCHR1 antagonist, TC-MCH7c ( Figure 1 Compound E, containing a fluorine atom linked to an aromatic ring, is an ideal compound for the synthesis and study of 18-fluoro tracers (Haga et al.). Bioorg Med Chem , 2011, 19: 883-93; Mikecz et al., Nucl Med Rev, 2017, 20(2): 111, Abstr. T1-1). The authors claim that the labeled compound showed specific uptake in the hypothalamic region of the brain of healthy rats, but no further studies have been published until now.

[0011] diaza Indole derivatives have selective antagonistic effects on MCHR1 receptors, which is known in the field (WO2016 / 166684A1), but radiolabeled MCHR1 PET ligands with similar structures have not yet been synthesized.

[0012] Based on the literature data, although MCHR1 PET radioligands have been developed, none have been proven suitable for clinical research to date.

[0013] Therefore, there is a need for MCHR1 PET radioligands that can be used in human clinical research, a need that has not been met. Summary of the Invention

[0014] This invention relates to radiolabeled compounds of general formula (I) or pharmaceutically acceptable salts thereof.

[0015] The present invention also relates to precursor compounds of general formula (II) or salts thereof.

[0016] The present invention also relates to precursor compounds of general formula (III) or salts thereof.

[0017] Furthermore, the present invention relates to a method for preparing radiolabeled compounds of general formula (I) or pharmaceutically acceptable salts thereof.

[0018] The present invention also relates to a method for preparing a precursor compound of general formula (II) or a pharmaceutically acceptable salt thereof.

[0019] This invention relates to radiolabeled compounds of general formula (I) used as PET tracers, or pharmaceutically acceptable salts thereof. Attached Figure Description

[0020] Figure 1 The structures of MCHR1 PET ligands known in the art.

[0021] Figure 2 In vivo dynamic PET imaging of the brains of healthy control Wistar rats following intravenous injection of Example 8. Representative decay-corrected dynamic PET images of the brains of healthy Wistar rats from Example 8 (…). Figure 2 A) and SUVmean time-activity curve (TAC) Figure 2 B). Black circle: Brain region.

[0022] Figure 3In vivo dynamic PET imaging of the brains of healthy control Wistar rats after intravenous injection of Example 9. Representative decay-corrected dynamic PET images of the brains of healthy Wistar rats from Example 9 (…). Figure 3 A) and SUVmean time-activity curve (TAC) Figure 3 B). Black circle: Brain region.

[0023] Figure 4 In vivo dynamic PET imaging of the brains of healthy control Wistar rats after intravenous injection of Example 10. Representative decay-corrected dynamic PET images of the brains of healthy Wistar rats from Example 10 (…). Figure 4 A) and SUVmean time-activity curve (TAC) Figure 4 B). Black circle: Brain region.

[0024] Figure 5 In vivo PET imaging of the brains of healthy control Wistar rats after intravenous injection of Reference Example 3 (alone) and CSA pretreatment. Representative decay-corrected summed PET images (0–180 min). Figure 5 A) and SUVmean time-activity curve (TAC) Figure 5 B). The black circles represent brain regions.

[0025] Figure 6 In vivo PET imaging of the brains of healthy control Wistar rats after intravenous injection of Example 11 (alone) and CSA pretreatment. Representative decay-corrected static PET images (0–40 min). Figure 6 A) and SUVmean time-activity curve (TAC) Figure 6 B). The black circles and arrows represent brain regions.

[0026] Figure 7 In vitro biodistribution of healthy control Wistar rats at 30 and 180 minutes after intravenous injection of Example 8.

[0027] Figure 8: PET / MRI summation images of the cynomolgus monkey brain 40–60 minutes after a single intravenous injection of Example 8. Figure 8A ) and PET / CT summation images between 80-100 minutes ( Figure 8B The black circle indicates a higher PET concentration.

[0028] Figure 9: Regional TAC recorded from the brain of a cynomolgus monkey. Four brain regions ( ) were observed after a single intravenous injection in Example 8. Figure 9A Images of the brain lobe and four regions ( Figure 9B (Image of ). Detailed Implementation

[0029] This invention relates to radiolabeled compounds of general formula (I) or pharmaceutically acceptable salts thereof:

[0030]

[0031] Where A is CH or N; R 1 yes 18 F and R 2 It is a CH3 or CH(CH3)2 group, or R 1 It is F and R 2 yes 11 CH3 group.

[0032] In one embodiment, the compound of general formula (I) is wherein A is CH or N, R 1 for 18 F, R 2 Compounds having a CH3 or CH(CH3)2 group, or pharmaceutically acceptable salts thereof.

[0033] In another embodiment, the compound of general formula (I) is wherein A is CH, R 1 for 18 F, R 2 Compounds having a CH3 or CH(CH3)2 group, or pharmaceutically acceptable salts thereof.

[0034] In one embodiment, the compound of general formula (I) is wherein A is CH or N, R 1 For F, R 2 for 11 A CH3 compound, or a pharmaceutically acceptable salt thereof.

[0035] In another embodiment, the compound of general formula (I) is wherein A is CH, R 1 For F, R 2 for 11 A CH3 compound, or a pharmaceutically acceptable salt thereof.

[0036] In yet another embodiment, the radiolabeled compound of general formula (I) is selected from compounds and their pharmaceutically acceptable salts:

[0037] 4-{[4-( 18 F)[fluorophenyl]methoxy}-1-[3-(propane-2-yl)-2,3,4,5-tetrahydro-1] H -[1,4]diaza [1,7-α]indol-9-yl]pyridine-2(1 H )-ketone,

[0038] 4-{[4-(18 F)[fluorophenyl]methoxy}-1-(3-methyl-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H )-ketone,

[0039] 4-[(4-fluorophenyl)methoxy]-1-[3-( 11 C) Methyl-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl]pyridine-2(1 H )-ketone,

[0040] 4-[(5-Fluoropyridin-2-yl)methoxy]-1-[3-( 11 C) Methyl-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl]pyridine-2(1 H )-ketone.

[0041] This invention also relates to fluorine-18 precursor compounds of general formula (II) or salts thereof:

[0042]

[0043] Among them, R 1 It is B(OC(R) 3 )2-C(R 3 )2-O) group; R 2 It is a CH3, CH(CH3)2, or COOC(CH3)3 group; R 3 It is a CH3 or C2H5 group.

[0044] In another embodiment, the compound of general formula (II) is selected from the following compounds:

[0045] 1-[3-(propan-2-yl)-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl]-4-{[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)phenyl]methoxy}pyridine-2(1 H )-ketone,

[0046] 1-(3-methyl-2,3,4,5-tetrahydro-1- H -[1,4]diaza [1,7-α]indol-9-yl)-4-{[4-(4,4,5,5-tetraethyl-1,3,2-dioxaborhecyclopentan-2-yl)phenyl]methoxy}pyridine-2(1 H )-ketone,

[0047] 4-{[4-(4,4,5,5-tetraethyl-1,3,2-dioxaborhecyclopentan-2-yl)phenyl]methoxy}-1-(2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H )-keto hydrochloride, and

[0048] 9-[2-oxo-4-{[4-(4,4,5,5-tetraethyl-1,3,2-dioxaborhecyclopentan-2-yl)phenyl]methoxy}pyridine-1(2 H )-[-1,2,4,5-tetrahydro-3] H -[1,4]diaza [1,7-α]indole-3-carboxylic acid tert-butyl ester.

[0049] This invention also relates to carbon-11 precursor compounds of general formula (III) or pharmaceutically acceptable salts thereof:

[0050]

[0051] Where A is CH or N.

[0052] In another embodiment, the compound of general formula (III) is selected from the following compounds:

[0053] 4-[(4-fluorophenyl)methoxy]-1-(2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H )-Ketomaleate, and

[0054] 4-[(5-Fluoropyridin-2-yl)methoxy]-1-(2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H )-Ketomaleate.

[0055] This invention relates to radiolabeled compounds of general formula (I) used as PET tracers, or pharmaceutically acceptable salts thereof.

[0056] The present invention also relates to radiolabeled compounds of general formula (I) or pharmaceutically acceptable salts thereof for visualizing the localization or distribution of MCHR1 receptors in mammals, including rodents, NHPs and humans, by PET imaging.

[0057] The term “mammal” means any vertebrate in the class Mammalia, including but not limited to guinea pigs, any rodents (such as hamsters, mice, rats), any non-human primates (such as cynomolgus monkeys, rhesus monkeys, baboons, marmosets, and green monkeys), or human subjects.

[0058] The present invention also relates to a method for preparing compounds of general formula (I).

[0059] The present invention also relates to a method for preparing compounds of general formula (II).

[0060] Therefore, compounds of general formula (I) or general formula (II) can be prepared by one of the following methods.

[0061] The following is a list of abbreviations used in common compositions and examples:

[0062]

[0063] General Method

[0064] Compounds of general formula (I) (where R) 1 for 18 F) can be synthesized by one of the following methods (reaction scheme 1):

[0065]

[0066] Boronates (compounds of general formula (II)) are used to prepare compounds of general formula (I) (where R) 1 for 18 F), in this method, copper(II)-mediated radiofluorination is carried out, preferably using Cu(OTf)2(py)4, and the fluorine-18 activity is recovered from the ion exchange column using potassium salts (preferably K2CO3 and KOTf), using a phase transfer catalyst (preferably K). 222 Under heated reaction conditions (preferably 80-90°C), in a bipolar aprotic solvent (preferably anhydrous DMA), and within a short reaction time (preferably 20 minutes) (Tredwell et al., Angew. Chem. Int. Ed. , 2014, 53(30): 7751-55).

[0067] Compounds of general formula (I) (where R) 2 for 11 CH3 can be synthesized by one of the following methods (Reaction Scheme 2):

[0068]

[0069] Compounds of general formula (I) (where R) 2 for 11 CH3) is produced via N-methylation of a secondary amine precursor (Garg et al., J. Nucl. Med. , 2017, 58(3): 473-78; Zirsberger K et al., EJNMMI Radiopharm Chem. , 2017, 2(10): 1-12). This is a nucleophilic alkylation step, using [ 11 [C]alkylating agent (preferably [ 11 A mixture of [C]MeOTf) and a bipolar aprotic solvent (preferably DMF and acetone) is reacted at low temperature (preferably -5°C) for a short reaction time (preferably 2 minutes). The secondary amine precursor (III) is known in the art (WO2016 / 166684A1).

[0070] Compounds of general formula (II) can be synthesized by one of the following methods (reaction scheme 3):

[0071]

[0072] Compound (3) is obtained by reacting compound (1) [Reference Example 1 of WO2016 / 166684] with compound (2) [Example 1 of EP1741703, step (2)] in step (i): in the presence of a catalyst (preferably cuprous iodide (I)), a base (preferably Cs2CO3), and a ligand (preferably trans-N,N'-dimethylcyclohexane-1,2-diamine), under heating conditions (preferably 110°C), in an inert solvent (preferably toluene), under an argon atmosphere. The intermediate of formula (4) is prepared from compound (3) by step (ii), i.e., a debenzylation step, preferably in the presence of hydrogen, a catalyst (preferably Pd / C), under acidic conditions (preferably AcOH / MeOH), at room temperature. Compound (6) is synthesized from compound (4) and a suitable benzyl bromide derivative (compound (5)) in the following step (iii): in the presence of a base (preferably NaH), under conventional reaction conditions (preferably room temperature), in an inert solvent (preferably DMF), and under an argon atmosphere. Compound (7) is prepared from compound (6) by step (iv), i.e., the deprotection step, preferably using HCl in EtOAc, at room temperature. The precursor compound of general formula (II) is obtained from compound (7). When R in compound (II) 2 When the group is CH3, in step (v) formaldehyde aqueous solution and NaBH(OAc)3 are used, and the solvent is a mixture of MeOH and DCM. When R in the compound of general formula (II)2 When the group is CH(CH3)2, an alkylating agent (preferably 2-iodopropane) and a base (preferably K2CO3) are used, and the reaction is carried out under heating conditions (preferably 80°C) in a bipolar aprotic solvent (preferably CH3CN) (step (vi)).

[0073] In another synthetic route, the precursor compound of general formula (II) is obtained from the compound of formula (11) according to step (iii). The compound of formula (11) is prepared from the compound of formula (10) by the debenzylation step according to step (ii). The compound of formula (10) is obtained by reacting the compound of formula (9) with the compound of formula (2) according to step (i). The compound of formula (9) is obtained from the compound of formula (8) according to step (v) or (vi). The compound of formula (8) is prepared from the compound of formula (1) by the deprotection step according to step (iv).

[0074] Reference Example 1 was originally synthesized by the Mitsunobu reaction using a 4-hydroxypyridinone intermediate (12) and 4-fluorobenzyl alcohol (13) (Haga et al., Bioorg Med Chem , 2011, 19: 883-93; Reaction scheme 4).

[0075]

[0076] In our modified method, the key intermediate is a 4-bromopyridone derivative (14). It is a common starting material for the preparation of Reference Example 1 and Reference Example 2, the borate ester precursor (Reaction Scheme 5). In the alkylation step, the compound of formula (14) is reacted with a suitable benzyl alcohol (15 or 16) in step (iii): in the presence of a base (preferably NaH), under heated reaction conditions (preferably 100°C), in a bipolar aprotic solvent (preferably anhydrous DMF), under an argon atmosphere. Reference Example 3, as a reference PET ligand, was synthesized from the precursor Reference Example 2 by a slight modification of the method described in Reaction Scheme 1 (reaction at 110°C for 13 minutes).

[0077]

[0078] Example

[0079] In this invention, TC-MCH 7c and [ 18 F]TC-MCH 7c were all synthesized via novel synthetic routes and were studied as reference examples 1 and 3.

[0080] Reference Example 1

[0081] 4-[(4-fluorophenyl)methoxy]-1-{4-[2-(pyrrolidone-1-yl)ethoxy]phenyl}pyridine-2(1 H )-ketone

[0082]

[0083] The title compound is described in Example 29 of WO2005 / 085200. It was prepared according to a different synthetic route and was isolated in the final step without chromatographic separation, as detailed below.

[0084] a.) 4-Bromo-1-{4-[(tert-butyldimethylsilyl)oxy]phenyl}-1,2-dihydropyridin-2-one

[0085]

[0086] 0.89 g (5.16 mmol) of 4-bromopyridine-2 (1 H A mixture of 4-bromo-1-{4-[(tert-butyldimethylsilyl)oxy]phenylboronic acid, 3.9 g (15.47 mmol) of 4-(tert-butyldimethylsilyl)oxy]phenylboronic acid, 1.41 g (7.77 mmol) of Cu(OAc)₂, 1.25 mL (15.47 mmol) of pyridine, and 2.0 g (4 Å) of molecular sieve in 40 mL of DCM was stirred at room temperature for 72 hours. The mixture was filtered and washed with 100 mL of brine. The organic fraction was dried over MgSO₄ and the solvent was evaporated under reduced pressure. The residue was purified by rapid silica gel chromatography (cyclohexane / EtOAc, gradient from 10:1 to 1:1) and crystallized from cyclohexane to give 0.82 g (41%) of 4-bromo-1-{4-[(tert-butyldimethylsilyl)oxy]phenyl}-1,2-dihydropyridine-2-one as a pale yellow solid.

[0087] MS (ESI): [M+H] + =380.

[0088] 1 H NMR (400MHz, DMSO) δ: 7.62 (d, J =7.3 Hz, 1H), 7.28 (d, J =8.9 Hz, 2H), 6.95 (d, J =8.9 Hz, 1H), 6.93 (s, 1H), 6.81 (d, J =2.2 Hz, 1H), 6.51 (dd, J =7.3, 2.2 Hz, 1H), 0.97-0.89 (m, 1H), 0.97 (s, 9H), 0.25-0.21 (m, 6H).

[0089] 13C NMR (101MHz, DMSO) δ: 160.0, 154.9, 139.8, 135.3, 133.4, 127.9, 121.9, 119.9, 109.2, 25.4, 17.8, -4.7.

[0090] b.) 4-Bromo-1-(4-hydroxyphenyl)pyridin-2(1H)-one

[0091]

[0092] A mixture of 2.64 g (6.94 mmol) of 4-bromo-1-{4-[(tert-butyldimethylsilyl)oxy]phenyl}-1,2-dihydropyridin-2-one [Ref. Example 1, step (a)] and 90 mL (83.3 mmol) of a 1 M tetrabutylammonium fluoride THF solution was stirred overnight at room temperature under an argon atmosphere. After solvent removal, the residue was partitioned between water and EtOAc. The organic fraction was washed with 100 mL of brine, dried over MgSO4, and the solvent was evaporated under reduced pressure. The residue was filtered through a short silica gel column using a cyclohexane / EtOAc 1:3 solution as the eluent. After solvent evaporation, the crude product was crystallized from Et2O to give 1.77 g (95.8%) of 4-bromo-1-(4-hydroxyphenyl)pyridin-2 (1 H )-ketone, a pale yellow solid.

[0093] MS (ESI): [M+H] + =266.

[0094] 1 H NMR (500MHz, DMSO) δ: 9.79 (s, 1H), 7.58 (d, J =7.3 Hz, 1H), 7.24-7.13(m, 2H), 6.95-6.82 (m, 2H), 6.79 (d, J =2.1 Hz, 1H), 6.49 (dd, J =7.3, 2.2 Hz, 1H).

[0095] 13 C NMR (126MHz, DMSO) δ: 160.1, 157.2, 139.9, 135.2, 131.3, 127.6, 121.8, 115.3, 109.0.

[0096] c.) 4-Bromo-1-{4-[2-(pyrrolidin-1-yl)ethoxy]phenyl}pyridin-2(1H)-one

[0097]

[0098] 1.22 g (4.58 mmol) of 4-bromo-1-(4-hydroxyphenyl)pyridine-2 (1 H )-ketone [Ref. Example 1, step (b)], 1.8 g (6.88 mmol) PPh3 and 0.65 mL (5.50 mmol) 1-pyrrolidineethanol in 25 mL THF were added to 1.0 mL (6.88 mmol) diethyl azodicarbonate. The mixture was stirred overnight at room temperature and then concentrated under reduced pressure. The residue was purified by rapid silica gel chromatography (DCM / MeOH, gradient from 10:1 to 1:10) and crystallized from Et2O to give 1.27 g (76%) 4-bromo-1-{4-[2-(pyrrolidine-1-yl)ethoxy]phenyl}pyridine-2(1 H )-ketone, a white solid.

[0099] MS (ESI): [M+H] + =363.

[0100] 1 H NMR (500MHz, DMSO) δ: 7.61 (d, J =7.3 Hz, 1H), 7.32-7.27 (m, 2H), 7.07-7.02 (m, 2H), 6.81 (d, J =2.0 Hz, 1H), 6.51 (dd, J =7.3, 2.2 Hz, 1H), 4.12 (t, J =5.8 Hz, 2H), 2.90-2.75 (m, 2H), 2.64-2.52 (m, 4H), 1.70 (m, 2H).

[0101] 13 C NMR (126MHz, DMSO) δ: 160.1, 158.1, 139.8, 135.3, 132.7, 127.7, 121.9, 114.6, 109.1, 66.8, 54.0, 53.9, 23.0.

[0102] d.) 4-[(4-fluorophenyl)methoxy]-1-{4-[2-(pyrrolidin-1-yl)ethoxy]phenyl}-1,2-dihydropyridine-2-one

[0103]

[0104] Under an argon atmosphere, 0.071 g (1.79 mmol) of 60% NaH dispersion was added to a 25 mL three-necked flask and suspended in 10 mL of anhydrous DMF. The suspension was cooled to 0 °C, and 0.188 mL (1.79 mmol) of 4-fluorobenzyl alcohol was added. The mixture was stirred at 0 °C for 10 minutes, and then 0.5 g (1.38 mmol) of 4-bromo-1-{4-[2-(pyrrolidone-1-yl)ethoxy]phenyl}pyridine-2(1 H The mixture was heated to 100°C for 15 minutes and reacted with a ketone [refer to Example 1, step (c)]. Complete conversion was achieved according to TLC (DCM / MeOH = 1:1) and LC-MS analysis. The mixture was cooled, quenched with 10 mL of water, and extracted with 3 × 3 mL EtOAc. The organic fraction was washed with brine, dried over MgSO4, and the solvent was evaporated under reduced pressure. The solid residue was crystallized from Et2O to give 0.178 g (32%) of 4-[(4-fluorophenyl)methoxy]-1-{4-[2-(pyrrolidone-1-yl)ethoxy]phenyl}-1,2-dihydropyridin-2-one as a white solid.

[0105] MS (ESI): [M+H] + =409.

[0106] 1 H NMR (400MHz, DMSO) δ: 7.56-7.48 (m, 3H), 7.28-7.21 (m, 4H), 7.02 (d, J =8.0 Hz, 2H), 6.06 (dd, J =7.6, 2.7 Hz, 1H), 5.96 (d, J =2.8 Hz, 1H), 5.11 (s,2H), 4.10 (t, J =5.9 Hz, 2H), 2.80 (t, J =5.8 Hz, 2H), 2.55-2.52 (m, 3H), 1.69(dt, J =6.8, 3.2 Hz, 4H).

[0107] 13 C NMR (101MHz, DMSO) δ: 166.6, 162.5, 161.9 (d, J =244.2 Hz), 157.7,139.2, 133.2, 132.0 (d, J =3.0 Hz), 130.2 (d, J=8.4 Hz), 127.9, 115.3 (d, J =21.4 Hz), 114.5, 99.9, 97.7, 68.8, 67.0, 54.1, 53.86, 23.0.

[0108] 19 F NMR (376MHz, DMSO) δ: -113.89 (tt, J =9.0, 5.5 Hz).

[0109] See Example 2

[0110] 1-{4-[2-(pyrrolidone-1-yl)ethoxy]phenyl}-4-{[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)phenyl]methoxy}pyridine-2(1 H )-ketone

[0111]

[0112] Under an argon atmosphere, 0.215 g (5.37 mmol) of 60% NaH dispersion was added to a 25 mL three-necked flask and suspended in 15 mL of anhydrous DMF. The suspension was cooled to 0 °C, and 1.17 mL (5.37 mmol) of [4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhexacyclopentan-2-yl)phenyl]methanol was added. The mixture was stirred at 0 °C for 10 minutes, and then 1.5 g (4.13 mmol) of 4-bromo-1-{4-[2-(pyrrolidone-1-yl)ethoxy]phenyl}pyridine-2(1 H The 4-ketone [Ref. Example 1, step (c)] was reacted with the mixture at 100°C for 1 hour. Complete conversion was achieved according to LC-MS analysis, but the formation of the deesterification byproduct (4-{[(2-oxo-1-{4-[2-(pyrrolidin-1-yl)ethoxy]phenyl}-1,2-dihydropyridin-4-yl)oxy]methyl}phenyl)boronic acid was also observed. The mixture was cooled, quenched with 30 mL of water, and extracted with 3 × 10 mL EtOAc. The organic fraction was washed with brine, dried over MgSO4, and the solvent was evaporated under reduced pressure. The solid residue was purified by silica gel column chromatography (CHCl3 / MeOH=10:1). The crude product was crystallized from Et2O to give 0.298 g (14%) of 1-{4-[2-(pyrrolidone-1-yl)ethoxy]phenyl}-4-{[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)phenyl]methoxy}-1,2-dihydropyridin-2-one, as a white solid.

[0113] MS (ESI): [M+H] +=517.

[0114] 1 H NMR (500MHz, DMSO) δ: 7.72 (d, J =8.0 Hz, 2H), 7.52 (d, J =7.6 Hz, 1H), 7.46 (d, J =8.0 Hz, 2H), 7.27-7.17 (m, 2H), 7.10-6.95 (m, 2H), 6.07 (dd, J =7.6, 2.7 Hz, 1H), 5.93 (d, J =2.7 Hz, 1H), 5.17 (s, 2H), 4.10 (t, J =5.9 Hz, 2H), 2.80 (t, J =5.8 Hz, 2H), 2.53 (br s, 4H), 1.69 (s, 4H), 1.30 (s, 12H).

[0115] 13 C NMR (126MHz, DMSO) δ: 166.6, 162.4, 157.7, 139.2, 139.1, 134.5, 133.2,127.9, 127.8, 126.9, 114.5, 114.4, 99.9, 97.8, 83.6, 69.3, 67.0, 54.1, 53.9,24.6, 23.0.

[0116] See Example 3

[0117] 4-{[4-( 18 F)[fluorophenyl]methoxy}-1-{4-[2-(pyrrolid-1-yl)ethoxy]phenyl}pyridine-2(1 H )-ketone

[0118]

[0119] Fluorine-18 produced in a Siemens Eclipse RD cyclotron was adsorbed onto an Oasis Max (Waters) column with 98% efficiency. The column was pre-activated with 2 mL of 0.15 M KHCO3 solution and 8 mL of Milli-Q water. The adsorption was achieved by using 805 mL of a stock solution (containing 2.3 mg / mL KHCO3). 222Fluorine-18 was recovered from the anion exchange column by elution with a mixture of a MeCN solution (prepared from 9 mg K₂CO₃ and 12 mg KOTf dissolved in 10 mL of ultrapure water) and 240 mL of an aqueous solution. The eluent was dried between 85 and 125 °C under a nitrogen stream and vacuum.

[0120] In GE TRACERlab FX FDG In the system's reactor, 5.2 mg (0.01 mol) of 1-{4-[2-(pyrrolidone-1-yl)ethoxy]phenyl}-4-{[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)phenyl]methoxy}pyridine-2(1 H )-ketone [Reference Example 2] and 6.8 mg (0.01 mol) Cu(OTf)2(py)4 were dissolved in 0.8 mL of DMA. This mixture was evaporated and transferred to a container containing 18 The solution of F was incubated at 110°C for 13 minutes. The reaction was quenched by adding 1 mL of water at 40°C. HPLC purification was performed using a Kromasil Eternity XT 10-C18 (10×150 mm) column at a flow rate of 5 mL / min; the eluent was ethanol / citrate buffer (pH=6.4) (30:70), at room temperature, with a 270 nm UV detector. R =12.3 minutes). Under these conditions, the radiochemical purity of the title compound was 98.5 ± 0.9% (n = 5), and the molar radioactivity was 1065 ± 365 GBq / mmol.

[0121] To form a radiolabeled product, 4 mL of radioactive solution was filtered through a Millex sterile filter (0.22 μm × 4 mm), diluted with 10 mL of PBS, the pH was adjusted to 6.5 with 1.6 mL of 1N NaOH solution, and 1.5 mL of 10% sodium ascorbate solution was used to ensure the formulation was resistant to radiodegradation.

[0122] non-radioactive compounds

[0123] Example 1

[0124] 4-[(4-fluorophenyl)methoxy]-1-[3-(propane-2-yl)-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl]pyridine-2(1 H )-ketone

[0125]

[0126] The title compound and its synthesis are described in Example 21 of WO2016 / 166684.

[0127] MS (ESI): 446.2 [M+H] + .

[0128] 1 H NMR (400MHz, DMSO) δ: 7.55-7.50 (m, 3H), 7.47 (d, J =8.8 Hz, 1H), 7.35 (d, J =2.1 Hz, 1H), 7.29-7.23 (m, 2H), 6.96 (dd, J =8.7, 2.1 Hz, 1H), 6.27 (s,1H), 6.05 (dd, J =7.6, 2.7 Hz, 1H), 5.96 (d, J =2.7 Hz, 1H), 5.12 (s, 2H), 4.30(d, J =7.1 Hz, 2H), 3.05-2.96 (m, 3H), 2.73-2.62 (m, 4H), 0.99 (d, J =6.6 Hz, 6H).

[0129] 13 C NMR (101MHz, DMSO) δ: 166.5, 162.8, 162.2 (d, J =244.1 Hz), 143.7,139.82 (s), 135.51 (s), 132.44 (s), 132.1 (d, J =3.1 Hz), 130.2 (d, J =8.3 Hz),127.0, 119.0, 117.5, 115.3 (d, J =21.4 Hz), 109.0, 99.5, 99.0, 97.7, 68.8,55.6, 51.9, 50.8, 45.7, 29.5, 17.9 (s).

[0130] 19 F NMR (376MHz, DMSO) δ: -113.91 (dq, J =9.1, 5.5 Hz).

[0131] Example 2

[0132] 4-[(4-fluorophenyl)methoxy]-1-(3-methyl-2,3,4,5-tetrahydro-1- H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H )-ketone

[0133]

[0134] a.) 9-Bromo-2,3,4,5-Tetrahydro-1H-[1,4]diaza [1,7-α]indole hydrochloride

[0135]

[0136] 5.35 g (14.7 mmol) of 9-bromo-1,2,4,5-tetrahydro-3- H -[1,4]diaza [1,7-α]Indole-3-carboxylic acid tert-butyl ester [Reference Example 1 of WO2016 / 166684] was added to a solution of 73 mL EtOAc containing 50 mL of 20% HCl in EtOAc. The mixture was stirred at room temperature. After 3 hours, the solid product was filtered, washed with 3 × 15 mL EtOAc, and then dried to give 4.42 g (99%) of 9-bromo-2,3,4,5-tetrahydro-1-carboxylic acid. H -[1,4]diaza [1,7-α]indole hydrochloride is a pale gray crystal.

[0137] MS (ESI): 265.0 [M+H] + .

[0138] b.) 9-Bromo-3-methyl-2,3,4,5-tetrahydro-1H-[1,4]diaza [1,7-α]indole

[0139]

[0140] 4.42 g (14.7 mmol) of 9-bromo-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indole hydrochloride [Example 1, step (a)] was suspended in a mixture of 300 mL DCM and 200 mL MeOH. 2.92 mL (39.2 mmol) of 37% formaldehyde aqueous solution was added in one go. The suspension was cooled to 0 °C, and 11.5 g (54.2 mmol) of NaBH(OAc)3 was added in one go. The cooling bath was removed, and the reaction mixture was allowed to warm to room temperature. Then 10 mL of water was added, and the organic solvent was evaporated. 100 mL of DCM was added, and the mixture was alkalized with a saturated Na2CO3 solution. The phase was separated, and the organic phase was washed with 2 × 20 mL of brine, dried over anhydrous MgSO4, filtered, and concentrated under vacuum. 3.28 g (80%) of 9-bromo-3-methyl-2,3,4,5-tetrahydro-1-methylhydrochloride was obtained. H -[1,4]diaza [1,7-α]indole is a blue crystal that requires no further purification.

[0141] MS (ESI): 279.0 [M+H] + .

[0142] c.) 4-[(4-fluorophenyl)methoxy]-1-(3-methyl-2,3,4,5-tetrahydro-1H-[1,4]diaza [1,7-α]indol-9-yl)pyridin-2(1H)-one

[0143]

[0144] 300 mg (1.1 mmol) of 9-bromo-3-methyl-2,3,4,5-tetrahydro-1 H -[1,4]diaza A mixture of [1,7-α]indole [Example 2, step (b)], 259 mg (1.2 mmol) of 4-[(4-fluorophenyl)methoxy]-1,2-dihydropyridin-2-one [WO2007 / 018248, Reference Example 2], 490 mg (1.5 mmol) of Cs₂CO₃, 168 mg (1.2 mmol) of trans-N,N'-dimethylcyclohexane-1,2-diamine, and 50 mL of toluene was stirred at room temperature for 1 hour while argon gas was bubbled into the mixture. 225 mg (1.2 mmol) of cuprous iodide (I) was added to the mixture, which was then immersed in an oil bath at 110 °C and stirred overnight at this temperature under an argon atmosphere. The mixture was evaporated to dryness, dissolved in 50 mL of DCM, and washed with 30 mL of 14% ammonia. The inorganic phase was extracted with 2 × 10 mL of DCM, and the combined organic phases were washed with 3 × 20 mL of 14% ammonia and 2 × 20 mL of brine, dried over anhydrous MgSO4, filtered, and concentrated under vacuum. The resulting 480 mg brown solid was ground with 5 mL of EtOAc, filtered, washed with 2 × 3 mL of EtOAc, and dried to give 315 mg (70%) of the title compound.

[0145] MS (ESI): 418.2 [M+H] + .

[0146] 1 H NMR (400MHz, DMSO-d6) d: 7.57-7.51 (m, 3H), 7.48 (d, J=8.8 Hz, 1H), 7.35 (d, J=2.0 Hz, 1H), 7.29-7.22 (m, 2H), 6.96 (dd, J=8.7, 2.1 Hz, 1H), 6.28(s, 1H), 6.05 (dd, J=7.6, 2.8 Hz, 1H), 5.96 (d, J=2.7 Hz, 1H), 5.12 (s, 2H), 4.38-4.29 (m, 2H), 3.06-3.01 (m, 2H), 2.66-2-56 (m, 4H), 2.35 (s, 3H).

[0147] 13C NMR (100MHz, DMSO-d6) d: 166.5, 162.8, 161.8 (d, J=244.0 Hz), 143.4,139.8, 135.5, 132.5, 132.1 (d, J=3.0 Hz), 130.2 (d, J=8.4 Hz), 127.0, 119.1,117.5, 115.3 (d, J=21.5 Hz), 109.0, 99.5, 99.1, 97.7, 68.8, 58.0, 56.8, 46.5,44.1, 28.1.

[0148] 19 F NMR (376MHz, DMSO-d6) d: -113.9 (tt, J=9.1, 5.5 Hz).

[0149] Example 3

[0150] 4-[(5-Fluoropyridin-2-yl)methoxy]-1-(3-methyl-2,3,4,5-tetrahydro-1-yl) H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H )-Ketomaleate

[0151]

[0152] The synthesis and analytical characterization of the title compound are described in Example 9 of WO2016 / 166684.

[0153] 18 Synthesis of F precursor

[0154] Example 4

[0155] 1-[3-(propan-2-yl)-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl]-4-{[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)phenyl]methoxy}pyridine-2(1 H )-ketone

[0156]

[0157] a) 9-Bromo-3-(propane-2-yl)-2,3,4,5-tetrahydro-1H-[1,4]diaza [1,7-α]indole

[0158]

[0159] 7.95 g (26.4 mmol) of 9-bromo-2,3,4,5-tetrahydro-1 H -[1,4]diaza A mixture of [1,7-α]indole hydrochloride [Example 2, step (a)], 105 mL acetonitrile, 16.40 g (119 mmol) K₂CO₃, and 14.7 mL (145 mmol) 2-iodopropane was stirred at reflux for 16 hours. After cooling to room temperature, the mixture was filtered through a diatomaceous earth saddle and the solution was evaporated to dryness. The residue was partitioned between 50 mL DCM and 50 mL water to separate the phases. The inorganic phase was extracted with 2 × 20 mL DCM, and the combined organic phases were washed with 2 × 20 mL brine, dried over anhydrous MgSO₄, filtered, and concentrated under vacuum to give 7.69 g (95%) 9-bromo-3-(propan-2-yl)-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indole is a green solid product.

[0160] MS (ESI): 307.1 [M+H] + .

[0161] b.) 4-(benzyloxy)-1-[3-(propan-2-yl)-2,3,4,5-tetrahydro-1H-[1,4]diaza [1,7-α]indol-9-yl]pyridin-2(1H)-one

[0162]

[0163] 7.69 g (25 mmol) of 9-bromo-3-(propan-2-yl)-2,3,4,5-tetrahydro-1 H -[1,4]diaza A mixture of [1,7-α]indole [Example 4, step (a)], 5.54 g (27.5 mmol) 4-(benzyloxy)-1,2-dihydropyridin-2-one [WO2005 / 085200, Example 1], 11.0 g (33.8 mmol) Cs₂CO₃, 3.92 g (27.5 mmol) trans-N,N'-dimethylcyclohexane-1,2-diamine, and 250 mL toluene was stirred at room temperature for 1 hour while argon gas was bubbled into the mixture. 5.24 g (27.5 mmol) cuprous iodide (I) was added to the mixture, which was then immersed in an oil bath at 110 °C and stirred overnight at this temperature under an argon atmosphere. The mixture was evaporated to dryness, dissolved in 150 mL of DCM, and vigorously stirred with 100 mL of 14% ammonia. The organic phase was separated and washed with 5 × 30 mL of 14% ammonia and 2 × 30 mL of brine, dried over anhydrous MgSO4, filtered, and concentrated under vacuum. The resulting 11.5 g of greenish-white solid was ground with 35 mL of EtOAc, cooled to 0 °C, filtered, washed with 10 mL of ice-cold EtOAc, and dried to give 10.0 g (94%) of 4-(benzyloxy)-1-[3-(propane-2-yl)-2,3,4,5-tetrahydro-1-yl] H -[1,4]diaza [1,7-α]indol-9-yl]pyridine-2(1 H )-ketone, is an off-white solid.

[0164] MS (ESI): 428.4 [M+H] + .

[0165] c) 4-Hydroxy-1-[3-(propane-2-yl)-2,3,4,5-tetrahydro-1H-[1,4]diaza [1,7-α]indol-9-yl]pyridin-2(1H)-one

[0166]

[0167] In a 250 mL three-necked flask, 10 g (23.4 mmol) of 4-(benzyloxy)-1-[3-(propane-2-yl)-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl]pyridine-2(1 HThe ketone [Example 4, step (b)] was suspended in 150 mL of MeOH. 6.69 mL (117 mmol) of AcOH was added to the suspension, turning it a dark green solution. After a few minutes, some precipitate formed in the solution. Argon was bubbled into the mixture for 30 minutes, followed by the addition of 4.98 g (4.7 mmol) of 10% Pd / C catalyst. After 10 minutes, the argon was replaced with hydrogen. After hydrogenation for 90 minutes, the mixture was purged with argon, and the suspension was filtered through a diatomaceous earth septum, washed with 80 mL of DCM, and evaporated to dryness to obtain a white solid. The residue was suspended in 100 mL of DCM, and 23 mL (164 mmol) of TEA was added. 100 mL of MeCN was added to the clear solution. After most of the DCM was evaporated from the solution, a slow precipitation of the product was observed. The mixture was stirred overnight at room temperature. Filtering yielded a white precipitate, providing 4.81 g (61%) of 4-hydroxy-1-[3-(propan-2-yl)-2,3,4,5-tetrahydro-1-yl] H -[1,4]diaza [1,7-α]indol-9-yl]pyridine-2(1 H )-ketone.

[0168] MS (ESI): 338.2 [M+H] + .

[0169] 1 H NMR (400MHz, DMSO-d6) d: 7.43 (d, J=8.7 Hz, 1H), 7.38 (d, J=7.5 Hz, 1H), 7.31 (d, J=2.0 Hz, 1H), 6.94 (dt, J=7.2, 3.6 Hz, 1H), 6.25 (s, 0.99 (d, J=6.6 Hz, 6H).

[0170] 13 d 50.8, 45.6, 29.5,17.9.

[0171] d.)1-[3-(propane-2-yl)-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl]-4-{[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)phenyl]methoxy}pyridine-2(1 H )-ketone

[0172]

[0173] Under an argon atmosphere, 277 mg (6.9 mmol) of 60% NaH dispersion was added to a 100 mL three-necked flask and suspended in 27 mL of anhydrous DMF. The suspension was cooled to 0 °C, and 1.80 g (5.3 mmol) of 4-hydroxy-1-[3-(propane-2-yl)-2,3,4,5-tetrahydro-1-yl] was added in a single batch. H -[1,4]diaza [1,7-α]indol-9-yl]pyridine-2(1 H )-Ketone [Example 4, step (c)]. After stirring the mixture at 0°C for 5 minutes, 2.17 g (6.9 mmol) of 2-[4-(bromomethyl)phenyl]-4,4,5,5-tetramethyl-1,3,2-dioxaborhexacyclopentane was added to the mixture. The suspension was heated to room temperature and stirred for 3 hours. Then, the suspension was cooled to 0°C and 20 mL of water was added. The temperature was raised to 25°C, and the thick suspension became difficult to stir. The solid was filtered and washed with 3 × 10 mL of water and 3 × 10 mL of Et2O. The off-white solid was dried to constant weight and then ground with 1 × 40 mL of MeCN. The solid was dried to constant weight to give 1.23 g (42%) of the title compound as an off-white solid, which was quantitatively analyzed. 1 ¹H-NMR analysis showed a purity of 91%. Due to the hydrolytic instability of the boronic ester structure, the product could not be further purified by chromatography and could be prepared for use in [the following context is missing from the original text]. 18 F-radioligand synthesis.

[0174] MS (ESI): 554.3 [M+H] + .

[0175] 1 H NMR(400MHz, CDCl3)δ: 7.85 (d, J =8.0 Hz, 2H), 7.45-7.39 (m, 3H), 7.30(d, J =7.6 Hz, 1H), 7.29 (d, J=8.6 Hz, 1H), 7.09 (dd, J =8.6, 2.0 Hz, 1H), 6.27(s, 1H), 6.07 (d, J =2.7 Hz, 1H), 6.03 (dd, J =7.5, 2.7 Hz, 1H), 5.06 (s, 2H), 4.29-4.24 (m, 2H), 3.10-2.95 (m, 3H), 2.80-2.69 (m, 4H), 1.36 (s, 12H), 1.05(d, J =6.6 Hz, 6H).

[0176] 13 C NMR (101MHz, CDCl3) δ: 167.3, 164.7, 144.0, 139.1, 138.5, 136.3, 135.3, 132.9, 129.3 (HMBC), 128.1, 127.0, 119.4, 118.1, 109.0, 100.8, 99.8,98.8, 84.0, 70.2, 56.7, 52.3, 51.4, 46.4, 30.4, 25.0, 18.4.

[0177] Example 5

[0178] 1-(3-methyl-2,3,4,5-tetrahydro-1- H -[1,4]diaza [1,7-α]indol-9-yl)-4-{[4-(4,4,5,5-tetraethyl-1,3,2-dioxaborhecyclopentan-2-yl)phenyl]methoxy}pyridine-2(1 H )-ketone

[0179]

[0180] a.) 9-(4-hydroxy-2-oxopyridin-1(2H)-yl)-1,2,4,5-tetrahydro-3H-[1,4]diaza [1,7-α]indole-3-carboxylic acid tert-butyl ester

[0181]

[0182] In a 100 mL three-necked flask, 960 mg (1.98 mmol) of 9-[4-(benzyloxy)-2-oxopyridine-1 (2... H )-[-1,2,4,5-tetrahydro-3] H-[1,4]diaza [1,7-α]indole-3-carboxylic acid tert-butyl ester [WO2016 / 166684, Reference Example 6] was suspended in 20 mL MeOH. After adding 30 mL DCM, argon gas was bubbled into the solution for 30 minutes, followed by the addition of 316 mg (0.297 mmol) of 10% Pd / C catalyst. After 10 minutes, the argon gas was replaced with hydrogen gas. After hydrogenation for 3 hours, the mixture was purged with argon gas, and the suspension was filtered through a diatomaceous earth septum, washed with 30 mL DCM, and the solution was evaporated to dryness to give a brown solid. The residue was ground with 10 mL MeCN, filtered, and washed with 2 × 5 mL MeCN. A white precipitate was filtered to give 650 mg (83%) of 9-(4-hydroxy-2-oxopyridine-1(2-oxopyridine)-1 ( ... H )-yl)-1,2,4,5-tetrahydro-3 H -[1,4]diaza [1,7-α]indole-3-carboxylic acid tert-butyl ester.

[0183] MS (ESI): 396.2 [M+H] + .

[0184] 1 H NMR (400MHz, DMSO-d6) d: 10.65 (s, 1H), 7.48 (d, J =8.7 Hz, 1H), 7.47(d, J =7.4 Hz, 1H), 7.35 (d, J =2.0 Hz, 1H), 6.97 (dd, J =8.7, 2.0 Hz, 1H), 6.32(s, 1H), 5.93 (dd, J =7.5, 2.6 Hz, 1H), 5.64 (d, J =2.6 Hz, 1H), 4.37-4.31 (m,2H), 3.65-3.62 (m, 2H), 3.58-3.54 (m, 2H), 3.05 (d, J =3.8 Hz, 2H), 1.46 (s, 9H).

[0185] 13C NMR (101MHz, DMSO-d6) d: 166.5, 162.9, 154.0, 142.3, 140.1, 136.0,132.9, 127.0, 119.5, 117.5, 109.2, 100.5, 99.6, 98.4, 79.2, 47.3(br), 46.0(br), 45.73(br), 29.4(br), 28.0.

[0186] b.) 2-[4-(bromomethyl)phenyl]-4,4,5,5-tetraethyl-1,3,2-dioxaboranecyclopentane

[0187]

[0188] In a 100 mL single-necked flask, 933 mg (5.4 mmol) of 3,4-diethylhexane-3,4-diol was dissolved in 53 mL of LCM. 1.15 g (5.35 mmol) of [4-(bromomethyl)phenyl]boric acid and 1.93 g (16.1 mmol) of anhydrous Na₂SO₄ were added to this solution. The mixture was stirred at room temperature for 48 hours. Inorganic impurities were then removed by filtration, and the filtrate was evaporated to dryness. The crude product was purified by rapid chromatography using C-hexane-EtOAc 96:4 to give 1.57 g (83%) of 2-[4-(bromomethyl)phenyl]-4,4,5,5-tetraethyl-1,3,2-dioxaborhexacyclopentane as a clear oil.

[0189] 1 H NMR (400MHz, DMSO-d6) δ: 7.68 (d, J =8.0 Hz, 2H), 7.46 (d, J =8.0 Hz,2H), 4.71 (s, 2H), 1.76-1.65 (m, 8H), 0.91 (t, J=7.4 Hz, 12H) 13 C NMR (101MHz, DMSO-d6) δ: 141.1, 134.6, 128.63, 88.4, 34.0, 25.9, 8.54.

[0190] c.) 9-[2-oxo-4-{[4-(4,4,5,5-tetraethyl-1,3,2-dioxaborane-2-yl)phenyl]methoxy}pyridin-1(2H)-yl]-1,2,4,5-tetrahydro-3H-[1,4]diaza [1,7-α]indole-3-carboxylic acid tert-butyl ester

[0191]

[0192] Under an argon atmosphere, 79 mg (2 mmol) of 60% NaH dispersion was added to a 100 mL three-necked flask and suspended in 24 mL of anhydrous DMF. The suspension was cooled to 0 °C, and 600 mg (1.5 mmol) of 9-(4-hydroxy-2-oxopyridine-1(2-)-O-dimethylamine (O-D ... H )-yl)-1,2,4,5-tetrahydro-3 H -[1,4]diaza [1,7-α]indole-3-carboxylic acid tert-butyl ester [Example 5, step (a)]. After stirring at 0°C for 10 minutes, 789 mg (2.1 mmol) of 2-[4-(bromomethyl)phenyl]-4,4,5,5-tetraethyl-1,3,2-dioxaborhexacyclopentane [Example 5, step (b)] was added in one step. The solution was heated to room temperature and stirred for 2 hours. The reaction was quenched with 500 μL of water, followed by the addition of 3 mL of 10% citric acid and 21 mL of water. The gelatinous solid residue was filtered off and washed with 10 mL of water. The solid was redissolved in 30 mL of DCM, dried over anhydrous Na2SO4, filtered, and evaporated to dryness. The crude product was purified by rapid chromatography using c-hexane-EtOAc 1:1 to give 473 mg (47%) of the title compound.

[0193] MS (ESI): 668.4 [M+H] + .

[0194] 1 H NMR (400MHz, DMSO-d6) δ: 7.74 (d, J =8.0 Hz, 2H), 7.54 (d, J =7.6 Hz, 1H), 7.50 (d, J =6.3 Hz, 1H), 7.47 (s, 2H), 7.37 (d, J =2.0 Hz, 1H), 6.99 (dd, J =8.7, 2.0 Hz, 1H), 6.33 (s, 1H), 6.07 (dd, J =7.6, 2.7 Hz, 1H), 5.93 (d, J =2.7 Hz, 1H), 5.18 (s, 2H), 4.35 (m, 2H), 3.64 (m, 2H), 3.56 (m, 2H), 3.06 (m,2H), 1.78-1.67 (m, 8H), 1.46 (s, 9H), 0.93 (t, J=7.4 Hz, 12H).

[0195] 13 C NMR(101MHz, DMSO-d6)δ: 166.5, 162.8, 154.0 (HMBC), 142.4, 139.8,139.2, 136.0, 134.5, 132.6, 127.1 (HMBC), 127.0, 126.9, 119.4, 117.5, 109.3,100.5, 99.5, 97.8, 88.3, ​​79.2, 69.2, 47.2 (HSQC), 46.0 (HSQC), 45.7 (HSQC), 29.5 (HSQC), 28.0, 26.0, 8.6.

[0196] d.)4-{[4-(4,4,5,5-tetraethyl-1,3,2-dioxaborphane-2-yl)phenyl]methoxy}-1-(2,3,4,5-tetrahydro-1H-[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1H)-one hydrochloride

[0197]

[0198] In a 100 mL single-necked flask, 430 mg (0.64 mmol) of 9-[2-oxo-4-{[4-(4,4,5,5-tetraethyl-1,3,2-dioxaborhecyclopentan-2-yl)phenyl]methoxy}pyridine-1 (2 H )-[-1,2,4,5-tetrahydro-3] H -[1,4]diaza [1,7-α]indole-3-carboxylic acid tert-butyl ester [Example 5, step (c)] was suspended in 30 mL of EtOH. Then 10 mL of DCM was added, and the suspension became a clear solution. 12 mL of EtOAc solution in 20% HCl was added. The mixture was stirred overnight at room temperature and then evaporated to dryness to give the title compound as off-white crystals. Assuming a quantitative yield, it can be used for the next step without further purification.

[0199] MS (ESI): 568.4 [M+H] + .

[0200] e.) 1-(3-methyl-2,3,4,5-tetrahydro-1H-[1,4]diaza) [1,7-α]indol-9-yl)-4-{[4-(4,4,5,5-tetraethyl-1,3,2-dioxaborphane-2-yl)phenyl]methoxy}pyridine-2(1H)-one

[0201]

[0202] In a 100 mL three-necked flask, add 389 mg (0.64 mmol) of 4-{[4-(4,4,5,5-tetraethyl-1,3,2-dioxaborphane-2-yl)phenyl]methoxy}-1-(2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H EtOAc hydrochloride [Example 5, step (d)] was dissolved in a mixture of 13 mL DCM and 8 mL MeOH. Then, 128 μL (1.72 mmol) of 37% formaldehyde aqueous solution was added, and the solution was cooled to 0 °C. 504 mg (2.38 mmol) of NaBH(OAc)3 was added, and the mixture was heated to room temperature. According to TLC (EtOAc / MeOH / TEA = 50:10:5), complete conversion was not achieved, so another batch of NaBH(OAc)3 (164 mg, 0.77 mmol) was added at 0 °C. After 5 minutes, the reaction was quenched with 3 mL of water at 0 °C. The mixture was diluted with 25 mL DCM, and the organic phase was washed with 10 mL of saturated NaHCO3 solution and 2 × 10 mL of brine, dried over anhydrous Na2SO4, filtered, and evaporated to dryness to give 360 ​​mg (96% yield in the last two steps) of the title compound as white crystals.

[0203] MS (ESI): 582.4 [M+H] + .

[0204] 1 H NMR (400MHz, DMSO) d: 7.74 (d, J =8.0 Hz, 2H), 7.55 (d, J =7.6 Hz, 1H),7.50-7.46 (m, 2H), 7.50-7.46 (m, 1H), 7.35 (d, J =2.0 Hz, 1H), 6.96 (dd, J =8.7, 2.1 Hz, 1H), 6.27 (s, 1H), 6.06 (dd, J =7.6, 2.7 Hz, 1H), 5.93 (d, J=2.7Hz, 1H), 5.18 (s, 2H), 4.33 (m, 2H), 3.06-3.00 (m, 2H), 2.59 (m, 4H), 2.35(s, 3H), 1.79-1.64 (m, 8H), 0.93 (t, J =7.4 Hz, 12H).

[0205] 13 C NMR (101MHz, DMSO) d: 166.46, 162.8, 143.4, 139.8, 139.2), 135.5,134.5, 132.5, 128.0 (HMBC), 127.0, 119.1, 117.5, 109.0, 99.5, 99.0, 97.8,88.3, ​​69.2, 58.0, 56.8, 46.5, 44.1, 28.1, 26.0, 8.6.

[0206] 11 Synthesis of C precursor

[0207] Example 6

[0208] 4-[(4-fluorophenyl)methoxy]-1-(2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H )-Ketomaleate

[0209]

[0210] In a 50 mL single-necked flask, 252 mg (0.625 mmol) of 4-[(4-fluorophenyl)methoxy]-1-(2,3,4,5-tetrahydro-1-ethylhexyl)methoxy ... H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H The ketone [Example 5, step (a) of WO2016 / 166684] was dissolved in 5 mL of DCM. Then, a solution of 72.5 mg (0.625 mmol) of maleic acid in 625 μL of MeOH was added to the mixture. A white precipitate formed immediately; the suspension was stirred at room temperature for 5 minutes. The crystals were filtered, washed successively with 2 × 2 mL DCM-MeOH 1:1 and 2 × 5 mL Et₂O, and dried to give 220 mg (68%) of the title compound.

[0211] MS (ESI): 404.2 [M+H] + .

[0212] 1 H NMR (400MHz, DMSO) δ: 8.93 (bs, 2H), 7.57-7.50 (m, 4H), 7.43 (d, J =2.0Hz, 1H), 7.29-7.23 (m, 2H), 7.05 (dd, J =5.6, 3.1 Hz, 1H), 6.43 (s, 1H), 6.07(dd, J =7.6, 2.8 Hz, 1H), 6.02 (s, 2H), 5.96 (d, J =2.7 Hz, 1H), 5.12 (s, 2H), 4.57 (d, J =5.7 Hz, 2H), 3.40-3.22 (m, 6H).

[0213] 13 C NMR (101MHz, DMSO) δ: 167.0, 166.5, 162.8, 161.8 (d, J =244.3 Hz),140.5, 139.7, 135.9, 135.7, 133.0, 132.1 (d, J =3.0 Hz), 130.2 (d, J =8.4 Hz),126.9, 119.9, 117.93, 115.3(d, J =21.4 Hz), 109.4, 100.7, 99.6, 97.7, 68.8, 46.8, 45.5, 40.9, 24.6.

[0214] 19 F NMR (376MHz, DMSO) δ: -113.85 ~ -113.93 (m).

[0215] Salt equivalent: 1:1, based on 1 ¹H NMR (400MHz, DMSO), labeled peaks: product cation 6.07 (dd, J=7.6, 2.8 Hz, 1H), maleate ion: 6.02 (s, 2H).

[0216] Example 7

[0217] 4-[(5-Fluoropyridin-2-yl)methoxy]-1-(2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H )-Ketomaleate

[0218]

[0219] In a 50 mL single-necked flask, 418 mg (1.03 mmol) of 4-[(5-fluoropyridin-2-yl)methoxy]-1-(2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H The )-ketone [Example 1, step (a) of WO2016 / 166684] was dissolved in a mixture of 15 mL DCM and 3 mL MeOH. Then, 1 mL of a MeOH solution containing 120 mg (0.625 mmol) of maleic acid was added to the mixture. After stirring for 5 minutes at room temperature, the solution became cloudy. 2 mL Et2O was added, and the suspension was stirred for 20 minutes at room temperature. The off-white crystals were filtered, washed with 2 × 4 mL Et2O, and dried to give 450 mg (84%) of the title compound.

[0220] MS (ESI): 405.2 [M+H] + .

[0221] 1 H NMR (400MHz, DMSO) δ: 8.92 (bs, 2H), 8.62 (d, J =2.9 Hz, 1H), 7.83 (td, J =8.7, 3.0 Hz, 1H), 7.66 (dd, J =8.7, 4.5 Hz, 1H), 7.54 (dd, J =8.2, 4.5 Hz, 2H), 7.43 (d, J =2.0 Hz, 1H), 7.05 (dd, J =8.7, 2.1 Hz, 1H), 6.43 (s, 1H), 6.11(dd, J =7.6, 2.8 Hz, 1H), 6.11 (dd, J =7.6, 2.8 Hz, 1H), 6.02 (s, 2H), 5.96 (d, J =2.7 Hz, 1H), 5.21 (s, 2H), 4.56 (d, J =4.9 Hz, 2H), 3.38-3.25 (m, 6H).

[0222] 13 C NMR (101MHz, DMSO) δ: 167.0, 166.4, 162.7, 158.8 (d, J =254.0 Hz), 151.7 (d, J =3.8 Hz), 140.5, 139.8, 137.3 (d, J =23.7 Hz), 136.0, 135.7, 132.9,126.9, 123.9 (d, J =18.5 Hz), 123.8 (d, J =4.7 Hz), 119.8, 117.9, 109.4, 100.7,99.5, 97.8, 69.8 (d, J =1.2 Hz), 46.8, 45.5, 40.9, 24.6.

[0223] 19 F NMR (376MHz, DMSO) δ: -128.02 (dd, J =8.8, 4.5 Hz).

[0224] Salt equivalent: 1:1, based on 1 1H NMR (400MHz, DMSO), labeled peaks: product cation 6.11 (dd, J=7.6, 2.8Hz, 1H), maleate ion: 6.02 (s, 2H).

[0225] radiosynthesis

[0226] Example 8

[0227] 4-{[4-( 18 F)[fluorophenyl]methoxy}-1-[3-(propane-2-yl)-2,3,4,5-tetrahydro-1] H -[1,4]diaza [1,7-α]indol-9-yl]pyridine-2(1 H )-ketone

[0228]

[0229] The title compound was synthesized according to the method described in Reference Example 3. 5.5 mg (0.01 mol) of 1-[3-(propan-2-yl)-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl]-4-{[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)phenyl]methoxy}pyridine-2(1 H )-ketone [Example 4] and 6.8 mg (0.01 mol) Cu(OTf)2(py)4 were dissolved in 0.8 mL of DMA. This mixture was evaporated and transferred to a container containing 18 The solution of F was incubated at 80°C for 20 minutes. The reaction was quenched by adding 1 mL of water at 40°C. HPLC purification was performed using a Waters XBridge BEHShield RP18 column (130 Å, 5 mm, 4.6 × 250 mm) at a flow rate of 1.2 mL / min; eluent 'A' = HCOOH / MeCN / H2O (1:100:900); eluent 'B' = HCOOH / MeCN / H2O (1:900:100); eluent concentration of 'B' = 22%, at room temperature, with a 270 nm UV detector (t). R =11.2 minutes). Under these conditions, the radiochemical purity of the title compound was 98.3 ± 0.4% (n = 7), and the molar radioactivity was 2371 ± 659 GBq / μmol. To form the radiolabeled product, 10 mL of the radioactive solution was filtered through a Millex sterile filter (0.22 mm × 4 mm), diluted with 6.5 mL of PBS, the pH was adjusted to 6.5 with 0.8 mL of 1N NaOH solution, and the formulation was ensured to resist radiodegradation with 0.91 mL of sodium ascorbate (1:10).

[0230] Example 9

[0231] 4-{[4-( 18 F)[fluorophenyl]methoxy}-1-(3-methyl-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H )-ketone

[0232]

[0233] The title compound consists of 1-(3-methyl-2,3,4,5-tetrahydro-1-methyl) H -[1,4]diaza [1,7-α]indol-9-yl)-4-{[4-(4,4,5,5-tetraethyl-1,3,2-dioxaborhecyclopentan-2-yl)phenyl]methoxy}pyridine-2(1 H)-Ketone [Example 5] was prepared by a copper(II)-mediated radiofluorination reaction, using a slight modification of the method described in Example 8: the reaction time was 20 minutes, and the temperature was 90°C. The HPLC flow rate was 3.2 mL / min; eluent 'A' = citrate buffer (pH=3) / EtOH (1:1); eluent 'B' = citrate buffer (pH=3) / EtOH (4:1); the concentration of eluent 'A' was 40% to >66%. Under these conditions (n=5), the radiochemical purity of the title compound was 99.8 ± 0.12% (t R =17 minutes), molar radioactivity is 1122±248 GBq / μmol.

[0234] Example 10

[0235] 4-[(4-fluorophenyl)methoxy]-1-[3-( 11 C) Methyl-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl]pyridine-2(1 H )-ketone

[0236]

[0237] [ 11 C]CO2 is produced in the Siemens Eclipse RD cyclotron accelerator, and is first reduced to [ 11 C]CH4, then further converted to [ using a commercial radiochemistry platform (ScanSys)] 11 C]CH3OTf.

[0238] Will[ 11 C]CH3OTf gas is bubbled through a gas containing 1 mg of 4-[(4-fluorophenyl)methoxy]-1-(2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H The reaction mixture was prepared in 300 mL of anhydrous DMF solution and 5 mL of 5N NaOH solution in 200 mL of acetone solution, with the temperature controlled at -5°C. After reacting for 2 minutes at room temperature, the reaction was complete. The reaction mixture was then loaded into the HPLC injection loop. The desired title compound was separated using an Akzo-Nobel Kromasyl Eternity XT-C18 column (10 mm, 10 × 150 mm), eluted with 65% citrate buffer (pH: 6.2) and 35% EtOH at a flow rate of 6 mL / min and a 254 nm UV detector (t). R=6.2 minutes). Under these conditions, the title compound had a radiochemical purity of 98.3 ± 1.3% (n = 3) and a molar radioactivity of 75 ± 9.3 GBq / μmol.

[0239] Example 11

[0240] 4-[(5-Fluoropyridin-2-yl)methoxy]-1-[3-( 11 C) Methyl-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl]pyridine-2(1 H )-ketone

[0241]

[0242] The title compound is composed of 4-[(5-fluoropyridin-2-yl)methoxy]-1-(2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H )-Ketomaleate [Example 7] was prepared according to the detailed steps in Example 10. Under these conditions, the radiochemical purity (t) was achieved. R =5.1 minutes) is 98.8±0.7% (n=5), molar radioactivity is 84.2±10.7 GBq / μmol.

[0243] use[ 125 In vitro binding assay of human MCH1 receptors using Tyr-S36057 as a radioligand

[0244] Membranes were prepared from recombinant human CHO-K1 cells (PerkinElmer, cat# ES-370-M) stably expressing the hMCHR1 receptor. Receptor binding assays were performed at 10 concentrations, with two replicates for each concentration, using incubation buffer (25 mM HEPES, pH 7.4, 10 mM MgCl2, 1 mM EDTA, 0.2% BSA), CHO-K1 cell membranes stably expressing the hMCHR1 receptor, 1% DMSO as solvent, and 0.05 nM [ 125 I]Tyr-S36057 was used as a radioligand. Nonspecific binding was determined in the presence of 1 μM MCH. The sample was incubated at 25 °C for 120 min. The binding reaction was terminated by rapid filtration and radioactivity was determined. The optimized saturated binding history K... D And History B max The concentrations were 0.1 nM and 26.0 pmole / mg protein, respectively.

[0245] The substitution of radioactive ligands by the test compound was performed via IC50. 50 Value representation, IC 50The values ​​were determined using MathIQ™ (IDBusiness Solutions Ltd., UK) via nonlinear least squares regression analysis. The suppression constant (K) is given. i ), K i Values ​​were obtained using the Cheng and Prusoff equations (Cheng, Y., Prusoff, WH, ...). Biochem. Pharmacol. (1973, 22: 3099-3108) Calculation using observation IC of the test compound. 50 The concentration of the radioligands used in the determination and the history of the radioligands K D Value (0.1 nM) (obtained experimentally in Eurofins Panlabs, Inc.). The Hill coefficient (nH) is used to define the slope of the competitive binding curve, using MathIQ. TM (Version 1.0) Calculation. A Hill coefficient that is significantly different from that in version 1.0 may indicate that the binding substitution process does not follow the law of mass action of a single binding site.

[0246] In vitro binding data of the tested compounds confirmed that Examples 1-3 exhibited high binding affinity for MCHR1. Binding data for Reference Example 1 (TC-MCH 7c) are compared with published data (Haga et al., Bioorg Med Chem (2011, 19: 883-9) shows a good correlation.

[0247] In vitro MCHR1 receptor occupancy rate in rat striatal homogenate ([ 3 H]SNAP combination)

[0248] Male Wistar rats weighing 190–210 g were used. The rats were purchased from Toxi-Coop (Budapest, Hungary). Animals arrived at the facility at least four days prior to the experiments. They were fed commercial feed and had free access to tap water. The animals were housed in a temperature-controlled room at 24±2°C and 50±10% relative humidity, with a 12-hour light / dark cycle (lights off from 6:00 PM to 6:00 AM). Throughout the experiments, the rats had unlimited access to commercially available pelleted rat feed autoclaved at 105°C and tap water. All applicable international, national, and / or institutional guidelines regarding animal care and use were followed. All procedures involving the experimental animals were reviewed and approved by the Gedeon Richter (internal) Institutional Animal Care and Use Committee and complied with European Parliament Directive 2010 / 63 / EU on the protection of animals for scientific purposes.

[0249] Animals were administered the carrier (DMA in a distilled aqueous solution of 25% HPβCD) or four study ligands (using the same formulation) via intravenous (iv) administration (2.5 mL / kg) via the tail vein.

[0250] Four rats were used in each administration group to determine MCHR1 receptor occupancy. Decapitation and brain dissection were performed 30 minutes after intravenous injection and 120 minutes after oral administration. Rats' striatal tissue preparation was completed within the subsequent 5-minute time window. Brain samples were rapidly frozen with dry ice and stored at -70°C until use.

[0251] Rat striatum was homogenized at room temperature using an Ultra-Turrax tissue homogenizer (set to 6, 10 seconds) in 9 volumes (w / v) (15 mg wet tissue / tube) of assay buffer (50 mM Tris pH=7.4, 120 mM NaCl, 5 mM KCl, 1 mM MgCl2, 2.5 mM CaCl2) and the binding assay was performed immediately.

[0252] in vitro[ 3 H]SNAP-7941 binding assays were performed in 1.2 mL deep-well plates (PS, non-sterile, 96 wells, Izinta Kft.). 40 μL of striatal homogenate (15 mg wet tissue / mL) was mixed with 210 μL of assay buffer and 50 μL of 6 nM (final assay concentration 1 nM). 3 H]SNAP-7941 (UBICHEM Research Ltd, Budapest, Hungary) was incubated together. Nonspecific binding (NSB) and total binding were measured in control animals treated with the vector. The total binding assay volume was 400 μL.

[0253] Incubate at room temperature for 45 minutes. After incubation, terminate the binding reaction by rapid filtration using a 96-well Filtermate cell collector (Perkin Elmer, Waltham, MA, USA), with the UniFilter® GF / C microporous filter plate (Perkin Elmer, Cat No.: 6055690) pre-soaked in 0.5% polyethyleneimine (PEI, dissolved in distilled water) for at least 1 hour.

[0254] The filter plate was washed three times with 0.5 mL of ice-cold washing buffer (50 mM Tris, pH 7.4). After washing, the filter plate was dried at 40 °C for 60 min, and 40 μL of Microscint-20 (Perkin Elmer, Cat#613621) scintillation solution was added to each well. Radioactivity was measured using a Microbeta 2450-0060 microplate reader (Perkin Elmer) (software: MicroBeta2 Windows Workstation, version: 2.3.0.12; serial number: 21210716).

[0255] Specific radioligand binding is defined as the difference between total binding and nonspecific binding measured in the presence of excess unlabeled ligands. Measurement is performed on drug-treated animal homogenates... 3 The in vitro receptor binding of H]SNAP-7941 and the receptor occupancy rate of the unlabeled compound administered in vivo were calculated as follows: Receptor occupancy rate (%) = 100 × [1 - (receptor marker in the drug treatment group / receptor marker in the carrier treatment group)]. MCHR1 receptor occupancy rate is expressed as [% relative to the carrier control]. 3 H]SNAP-7941 binding inhibition percentage. Data presented as average percentage of occupancy ± SD.

[0256] The in vitro occupancy data of the test compounds characterized the binding properties of Examples 1-3, indicating that not only was a high percentage of the MCHR1 receptor population occupied, but the occupancy level was also improved compared to Reference Example 1. Based on these data, there was no correlation between the binding data and the in vitro occupancy rate of Reference Example 1.

[0257] Table 1: In vitro human-human binding data and in vitro rat striatal homogenate occupancy rate

[0258]

[0259] In vivo PET imaging (positron emission tomography) in rats

[0260] laboratory animals

[0261] In vivo PET imaging and in vitro experiments were performed using 16-week-old male Wistar rats weighing 256±10g (n=16; Animalab Ltd, Budapest, Hungary). Animals were housed under standard conditions of 23±2℃ and 50±10% humidity with artificial lighting using a 12-hour light / dark cycle. All animals had free access to semi-synthetic feed (VRF1; Akronom Ltd., Budapest, Hungary) and drinking water. Animal experiments were authorized by the Animal Research Ethics Committee of the University of Debrecen, Hungary. The husbandry and handling of laboratory animals complied with all applicable provisions of Hungarian law and EU animal welfare directives and regulations. Permit No.: III / 6-KÁT-2015.

[0262] In vivo PET imaging in Example 8

[0263] Healthy control (n=16) Wistar rats were anesthetized using 3% Forane (AbbVie) and a dedicated small animal anesthesia device (Tec3 isoflurane vaporizer, Eickemeyer Veterinary Equipment, UK). After an incubation period (20 minutes), the pretreated rats were injected via the lateral caudal vein with 11.07 ± 2.9 MBq of Example 8 (dissolved in 150 μL of physiological saline). Preclinical use MiniPET-II The equipment performs dynamic (0-180 minutes) PET scans (Lajtos et al., Nuc. Int. Meth. in Phys. Res. A (2013, 707: 26-34). After 3D OSEM-LOR image reconstruction, the volume of interest (VOI) of the whole brain was manually delineated using Brain CAD image analysis software, and the quantitatively standardized uptake value (SUV) was calculated as follows: SUV = [VOI activity (Bq / mL)] / [injection activity (Bq) / animal body weight (g)], assuming a density of 1 g / mL.

[0264] To determine the effects of Example 8 in the brain, in vivo dynamic PET scans were performed under inhalation anesthesia. Qualitative analysis of the decay-corrected images was performed, and images were identified within 40 minutes after intravenous (iv) administration of Example 8. Figure 2 A). Analysis of TAC data revealed a continuous decrease in the SUVmean value in the brain ( Figure 2 B). PET images showed that Example 8 was able to cross the blood-brain barrier, and the radioactivity concentration decreased over time. During dynamic PET scans, the radioligand was very stable in rat plasma, and no radioactive metabolites were observed.

[0265] In vivo PET imaging in Examples 9 and 10

[0266] Following the PET imaging method described in Example 8, in vivo dynamic PET imaging studies were performed using the methods described in Examples 9 and 10, respectively, by injecting 11.07 ± 2.9 MBq and 12.14 ± 0.9 MBq (dissolved in 150 μL of physiological saline). In both cases, summed (0–40 minutes) decay-corrected images and TAC curves were generated. Figure 3 A and 3B, Figure 4 (A and 4B). Fluorine-18 (Example 9) and carbon-11 (Example 10) PET ligands both entered the brains of healthy Wistar rats.

[0267] In vivo PET imaging (refer to Example 3)

[0268] To determine the potential of Reference Example 3 to target MCHR receptors in the brain, in vivo dynamic PET scans were performed under inhalation anesthesia. Qualitative analysis of decay-corrected images revealed no significant radioactivity detected in the brain following intravenous (iv) administration of Reference Example 3. Figure 5 A, left). Analysis of TAC data revealed a continuous decrease in the SUVmean value in the brain (…). Figure 5 (B, upper right). PET images show that, in reference example 3, the blood-brain barrier could not be crossed.

[0269] To investigate in vivo whether Reference Example 3 is a substrate of the blood-brain barrier P-glycoprotein (Pgp), experimental animals were pre-injected intravenously with approximately 25 μg / g body weight of cyclosporine A (CSA). Ten minutes after CSA injection, approximately 10 MBq of Reference Example 3 was injected via the lateral tail vein for PET imaging. Compared to baseline, CSA administration resulted in a significantly higher brain accumulation, suggesting that Reference Example 3 may be a substrate of the Pgp pump. Figure 5 A right and Figure 5 B (bottom right).

[0270] In vivo PET imaging in Example 11

[0271] Example 11 was studied using PET imaging methods similar to Reference Example 3. In healthy Wistar rats, the compound entered the brain after CSA pretreatment. Without CSA pretreatment ( Figure 6 A, left) and after CSA pretreatment ( Figure 6 A, right) Representative decay-corrected static PET image of the brain (0–40 minutes). Circles and arrows: Brain ( Figure 6 A). Representative decay-corrected dynamic PET images and SUVmean time-activity curves (TAC) of the brain from Example 11 ( Figure 6 B).

[0272] In vitro biodistribution of rats

[0273] laboratory animals

[0274] Healthy control (n=16) Wistar rats were anesthetized using 3% Forane (AbbVie) and a dedicated small animal anesthesia device (Tec3 isoflurane vaporizer, Eickemeyer Veterinary Equipment, UK).

[0275] In vitro biodistribution of Example 8

[0276] To determine the normal biodistribution of Example 8, animals were injected with 11.26 ± 2.08 MBq of Example 8 (dissolved in 150 μL of physiological saline) via the lateral caudal vein. Rats were euthanized with 5% iodine at 30 and 180 minutes post-injection, and blood, urine, and selected organ samples were collected. The weight and radioactivity of the selected organs and tissues were measured using a calibrated gamma counter (HEWLETT PACKARD Cobra II Autogama Gamma Counter). The uptake of the radiotracer in Example 8 is expressed as %ID / g tissue. Figure 7 The biodistribution results of Example 8 are shown. Based on imaging data, the compound was primarily excreted via the liver of the tested Wistar rats.

[0277] In vivo PET imaging of non-human primates (NHP)

[0278] All experimental procedures (except for feeding and transport) were conducted at the Medicopus facility in Kaposvár, Hungary. The study was approved by the local and national animal research ethics committees and the animal health and food control department of the county government office of the Ministry of Agriculture, with license numbers BA / 73 / 0936-8 / 2022 and BA / 73 / 00023-7 / 2023. In accordance with Hungarian Government Directive 40 / 2013 (II.14.) "On Animal Experimentation" and European Parliament and Council Directive 2010 / 63 / EU "On the Protection of Animals Used for Scientific Purposes," all possible measures were taken to minimize animal pain and discomfort.

[0279] The animals were housed in a large, two-story primate enclosure, where their recommended daily metabolizable energy intake requirements and species-specific social needs were closely monitored. The animals were fed once daily, in the afternoon, after their daily training and testing sessions. The diet consisted of standard nutritionally complete laboratory feed (NHP pellets, SsnifffSpezialdi) specifically designed for non-human primates. The study included six 3-year-old cynomolgus macaques (4 males) with a daily supplement of fresh fruits and vegetables. Water was readily available. Temperature and relative humidity were maintained at 24±2℃ and 55±5%, respectively, in the enclosures and testing room.

[0280] Animals in their habitat (GTRC, K Hungary) vágósz l The animals were transported between the laboratory and the neuroimaging center (Medicopus Ltd., Kaposborg, Hungary) using a specially designed air-conditioned van (Dacia Dokker Van) equipped with a camera and specialized primate transport cages. The one-way transport took approximately one hour. Prior to the experiment, all animals underwent intensive acclimatization training and were familiarized with the transport cages through positive reinforcement training (PRT). (The transport cages were connected to the feeding cages, and the animals regularly entered the transport cages to obtain their daily food.) Therefore, the stress caused to the animals during transport to and from the measurement location was minimized.

[0281] Prior to anesthesia, animals were fasted for 6 hours and deprived of water for 2 hours, respectively. The anesthesia protocol was adjusted based on individual animal sensitivities. The primary objective was to ensure stable deep anesthesia with adequate oxygen saturation and normal body temperature throughout the experiment. For preoperative medication, animals were first lightly anesthetized with a single intramuscular injection of ketamine (0.25 mg / kg, CP Ketamin, CP-Pharma, HU) and then given a single intramuscular injection of atropine (0.04–0.05 mg / kg, Atropinum Sulphuricum, Egis, HU) to prevent salivation. Deep anesthesia was then induced by inhaling a mixture of 3.5–5% v / v isoflurane (Aerrane, Baxter, US) gas and a pure oxygen flow of 2 L / min via a Mapleson-D ventilation system. After achieving sufficient depth of anesthesia, a supraglottic airway tube (laryngeal mask airway, Fazzini, Italy) was inserted orally around the larynx of the trachea and sealed with a low-pressure cuff to ensure airway safety and stable ventilation. To maintain a stable level of deep anesthesia throughout the experiment, a mixture of 1.5V / V% isoflurane and 2L / min of pure O2 was used. After the experiment, the laryngeal mask airway was removed, and the animals were allowed to inhale pure O2 through a face mask until they regained consciousness. They were then returned to their transport cages and continuously monitored, and provided with high-sugar fruit (bananas) and water. When the animals spontaneously accepted food and exhibited normal behavior without signs of sedation, they were returned to their habitat. The total recovery (awakening) time to transportability was between 1 and 1.5 hours.

[0282] In vivo PET imaging in Example 8

[0283] Crab-eating macaques (3.2 kg, female) received a single intravenous injection (Example 8) (140 MBq / 3 mL). The PET measurement protocol was specifically optimized for cynomolgus macaques. During the measurement, simultaneous MRI (Siemens Biograph mMR MR-PET camera) and CT (Siemens Biograph TruePoint 64) scans were performed: first a 60-minute PET / MRI scan, followed by a 25-minute PET / CT scan. Figure 8A and 8B ).

[0284] Using the Level 1 VOI system in the rhesus monkey cortical hierarchy atlas (CHARM), time-activity curves (TACs) were generated for eight brain regions (prefrontal cortex, parietal lobe, temporal lobe, occipital lobe, telencephalon, diencephalon, midbrain, and hindbrain). A brief accumulation period (3-4 minutes) was detected (n=4 monkeys), followed by a 10-minute saturation period and a 45-minute slow decline period (see [link to relevant documentation]). Figure 9A and Figure 9B ).

Claims

1. A compound of general formula (I) or a pharmaceutically acceptable salt thereof, in, A is CH or N; R 1 yes 18 F and R 2 It is a CH3 or CH(CH3)2 group, or R 1 It is F and R 2 yes 11 CH3 group.

2. The compound according to claim 1 and its pharmaceutically acceptable salt, wherein, The compound is selected from the following compounds: 4-{[4-( 18 F)[fluorophenyl]methoxy}-1-[3-(propane-2-yl)-2,3,4,5-tetrahydro-1] H -[1,4]diaza [1,7-α]indol-9-yl]pyridine-2(1 H )-ketone, 4-{[4-( 18 F)[fluorophenyl]methoxy}-1-(3-methyl-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H )-ketone, 4-[(4-fluorophenyl)methoxy]-1-[3-( 11 C) Methyl-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl]pyridine-2(1 H )-ketone, 4-[(5-Fluoropyridin-2-yl)methoxy]-1-[3-( 11 C) Methyl-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl]pyridine-2(1 H )-ketone.

3. The compound according to claim 1 and its pharmaceutically acceptable salt, wherein, The compound is selected from the following compounds: 4-{[4-( 18 F)[fluorophenyl]methoxy}-1-[3-(propane-2-yl)-2,3,4,5-tetrahydro-1] H -[1,4]diaza [1,7-α]indol-9-yl]pyridine-2(1 H )-ketone, 4-{[4-( 18 F)[fluorophenyl]methoxy}-1-(3-methyl-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H )-ketone, 4-[(4-fluorophenyl)methoxy]-1-[3-( 11 C) Methyl-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl]pyridine-2(1 H )-ketone.

4. A compound of general formula (II) or a salt thereof: in, R 1 It is B(OC(R) 3 )2-C(R 3 )2-O) group, R 2 It is a CH3, CH(CH3)2 or COOC(CH3)3 group. R 3 It is a CH3 or C2H5 group.

5. The compound according to claim 4, wherein the compound is selected from the group consisting of: 1-[3-(propan-2-yl)-2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl]-4-{[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborhecyclopentan-2-yl)phenyl]methoxy}pyridine-2(1 H )-ketone, 1-(3-methyl-2,3,4,5-tetrahydro-1- H -[1,4]diaza [1,7-α]indol-9-yl)-4-{[4-(4,4,5,5-tetraethyl-1,3,2-dioxaborhecyclopentan-2-yl)phenyl]methoxy}pyridine-2(1 H )-ketone, 4-{[4-(4,4,5,5-tetraethyl-1,3,2-dioxaborhecyclopentan-2-yl)phenyl]methoxy}-1-(2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H )-keto hydrochloride, and 9-[2-oxo-4-{[4-(4,4,5,5-tetraethyl-1,3,2-dioxaborhecyclopentan-2-yl)phenyl]methoxy}pyridine-1(2 H )-[-1,2,4,5-tetrahydro-3] H -[1,4]diaza [1,7-α]indole-3-carboxylic acid tert-butyl ester.

6. A compound of general formula (III) or a pharmaceutically acceptable salt thereof: Where A is CH or N.

7. The compound according to claim 6, wherein the compound is selected from the group consisting of: 4-[(4-fluorophenyl)methoxy]-1-(2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H )-Ketomaleate, and 4-[(5-Fluoropyridin-2-yl)methoxy]-1-(2,3,4,5-tetrahydro-1 H -[1,4]diaza [1,7-α]indol-9-yl)pyridine-2(1 H )-Ketomaleate.

8. The compound according to any one of claims 1 to 3, which is used as a PET tracer.

9. The compound according to any one of claims 1 to 3, for visualizing the localization or distribution of MCHR1 receptors in mammals by PET imaging.

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