Slack-activating compounds and their medical uses

Abstract

This invention relates to novel compounds structurally derived from 2-chloro-11-(4-methylpiperazin-1-yl)dibenzo[b,f][1,4]oxazepine and their synthesis. Comparative studies have shown that the compounds of this invention bind to and activate potassium channels, particularly Slack. Based on their activating activity, the compounds of this invention and pharmaceutical compositions containing them can be used in a variety of therapeutic applications for the treatment or prevention of disease conditions.

Description

[Technical Field]

[0001] This invention relates to novel compounds structurally derived from 2-chloro-11-(4-methylpiperazin-1-yl)dibenzo[b,f][1,4]oxazepine and their synthesis. Comparative studies have shown that the compounds of this invention bind to and activate potassium channels, particularly Slack. Based on their activating activity, the compounds of this invention and pharmaceutical compositions containing them can be used in a variety of therapeutic applications for the treatment or prevention of disease conditions. The compounds of this invention are particularly useful in the treatment of systemic pain / inflammatory pain and / or pruritus / local itching. [Background technology]

[0002] Itching (also known as pruritus) is a debilitating symptom associated with various skin disorders, systemic diseases such as chronic kidney disease or cholestatic liver disease, mental illnesses, and neurological disorders, or it is an idiopathic symptom of unknown cause. Itching is defined as an unpleasant sensation that triggers the urge to scratch, and approximately one-third of dermatology patients and nearly 15% of the general population report symptoms of itching. 1、2 Like pain, acute itching serves as an important defense mechanism for detecting potentially harmful stimuli. However, chronic itching (i.e., lasting longer than six weeks in humans) does not serve a useful function and is rather distressing, often impairing quality of life to the same extent as chronic pain.

[0003] Itching can be classified into histamine-dependent (histaminergic) and histamine-independent (non-histaminergic) types. Since most types of chronic itching are histamine-independent and resistant to antihistamines, there is an urgent need to develop new treatment strategies.

[0004] Most of the known mechanisms of pruriception (itching sensation) begin with the activation of pruriceptive sensory neurons. Several major receptors, including members of the Mrgpr (Mas-related G protein-coupled receptor) family, have been found to be important for detecting non-histaminergic chemical itch signals. 3 Recent single-cell RNA sequencing (scRNA-seq) studies have revealed that itch-related genes are highly expressed in distinct subsets of sensory neurons. For example, in a pioneering scRNA-seq study by Usoskin et al., 11 major types of sensory neurons were identified, and three non-peptidergic populations (termed NP1, NP2, and NP3) were proposed to be pruriceptive. 4 After activation, these sensory neurons send signals to the posterior horn of the spinal cord, where the continuous information is further processed and transmitted to the ascending itch signaling pathway to the brain. 5 .

[0005] The excitability of sensory neurons is driven by various types of ion channels, among which K + channels are the most numerous and diverse class, being governed by over 75 genes in humans and characterized by tissue-dependent expression patterns. 6 Notably, Slack (K + 1.1, also known as the gene Kcnt1), a K + channel activated by intracellular Na Na and inhibited by divalent cations, is highly enriched in the pruriceptive NP1, NP2, and NP3 subsets of mouse sensory neurons 7、8 (Figure 1A). This suggests that Slack has a role in itching. Indeed, the significant functional role of Slack in pruriception is reflected by the increased scratching behavior of Slack knockout mice after exposure to pruritogens such as chloroquine and histamine. 4、9 (Figure 1C) and human sensory neurons 10 . 11 (Figure 1C) and human sensory neurons12 The observation that Slack expression (Figure 1B) is enriched in itch-associated cell populations supports the hypothesis that activators of the Slack channel have therapeutic potential for treating itch.

[0006] Previous studies using library screening of pharmacologically active compounds have reported that the first-generation antipsychotic roxapine activates Slack. 13 In preliminary experiments, the inventors observed that systemic administration of low-dose roxapine significantly alleviated chloroquine-induced scratching behavior in wild-type mice, but did not affect scratching behavior in Slack knockout mice (data not shown). This indicates that the roxapine-induced antipruritic effect is dependent on Slack activation.

[0007] Despite the need for medications to treat histamine-independent itching, unfortunately, the clinical use of roxapine is limited by typical adverse events associated with first-generation antipsychotics, such as blockade activity at dopamine receptors and other receptors. 14、15 .

[0008] The inventors, starting with the structure of roxapine, developed novel structural roxapine derivatives that have an activating effect on Slack. Notably, the roxapine derivatives of the present invention are characterized by a more favorable pharmacological profile distinct from that of roxapine. Furthermore, the compounds of the present invention acting as Slack activators have equivalent or improved binding properties to Slack, reduced BBB permeability leading to fewer undesirable side effects, and improved off-target activity compared to the parent molecule roxapine. [Overview of the project]

[0009] In general, the main aspects of the present invention can be described as follows with a brief explanation.

[0010] In a first embodiment, the present invention relates to formula I: [ka] (In the formula, A is one or two substituents independently selected from -H, -F, -Cl, -Br, -I, -CF3, -OCF3, -CF2H, and -OCF2H. B is one or two substituents independently selected from -F, -Cl, -Br, -I, -CF3, -OCF3, -CF2H, and -OCF2H. X is selected from -S- and -O-, m is between 0 and 3. Y is -H; Nitrile group; A 5- to 6-membered unsaturated heterocycle containing three heteroatoms selected from O and N, preferably two or three of which are nitrogen, and the heterocycle is monosubstituted or disubstituted with =S or =O, preferably with =O; or, Linear-C 1~3 Alkyl, linear-C 1~3 The alkyl group -(CH2)-, preferably one -(CH2)-, is optionally monosubstituted with -OH, and the linear-chain -C 1~3 The alkyl group's terminal -CH3 is -OR 1 Alternatively, -SR 1 And preferably -OR 1 It is monosubstituted with R 1 However, it is a straight chain -C which is -H, -(CH2)-COOH or -(CH2)-(CH2)-OH. 1~3 Alkyl; or, -(CH2) i -CX 1 R 2 (Here, X 1 is, =NR 3 Or = O, R 3 is -H or -OH, i is either 0 or 1. R 2 は-OR 4 or -NR 5 R 6 And R4 is -H or -C 1~2 It is alkyl, R 5 and R 6 (The element is selected independently of -H or -methyl); Selected from, Here, it is referred to as "Z", [ka] is an aliphatic group comprising at least one, preferably one or two heterocycles, the heterocycles collectively comprising two nitrogen atoms. Of the two nitrogen atoms, the first nitrogen atom is Z in formula I [ka] Linked to, the second of the two nitrogen atoms is Z in formula I [ka] Connected to, preferably, [ka] It is connected to the Z group, Each of the two nitrogen atoms is a heteroatom in one of the two linked four-membered heterocycles (each heterocycle has only one heteroatom), and the two linked four-membered heterocycles form a spirocycle, preferably the four-membered heterocycle having the first nitrogen atom is linked to the four-membered heterocycle having the second nitrogen atom via a shared carbon atom, thereby forming a spirocycle having two heteroatoms which are two nitrogen atoms, or The first of two nitrogen atoms is linked to a four-membered heterocycle having one heteroatom, the second of the two nitrogen atoms is the heteroatom in the four-membered heterocycle to which the first nitrogen atom is linked, and the first nitrogen atom is further substituted with methyl, or The present invention relates to compounds having two nitrogen atoms that are heteroatoms in a 6- to 7-membered heterocycle having two heteroatoms, and the heterocycle is optionally substituted with methyl, preferably monosubstituted, or to salts, complexes, diastereomers, enantiomers and / or tautomers of compounds having formula (I).

[0011] In a second embodiment, the present invention relates to a pharmaceutical composition comprising any one of the compounds described herein, or a salt, solvate, or ester thereof, and a pharmaceutically acceptable carrier or excipient.

[0012] In a third embodiment, the present invention relates to a compound or composition used to treat a disease in a subject, wherein the compound or composition contains an effective amount of a compound according to the first embodiment of the present invention, or a salt, solvate, or ester thereof, the disease is treatable by activating potassium channels in cells associated with the pathology of the disease, and the treatment comprises administering the compound or composition to a subject in need of treatment.

[0013] In a fourth embodiment, the present invention relates to a method for synthesizing a compound according to the first embodiment of the present invention. [Modes for carrying out the invention]

[0014] The elements of the present invention are described in more detail here. These elements are listed with specific embodiments, but it should be understood that they can be combined in any way and in any number to create additional embodiments. The various examples and preferred embodiments described are not to be construed as limiting the invention to only the explicitly described embodiments. The specification of this application should be understood to support and encompass embodiments that combine the explicitly described embodiments with any number of disclosed and / or preferred elements. Furthermore, unless otherwise specified in the context, any permutations and combinations of all elements described herein should be considered disclosed by the specification of this application.

[0015] In a first embodiment, the present invention relates to formula I: [ka] (In the formula, A is one or two substituents independently selected from -H, -F, -Cl, -Br, -I, -CF3, -OCF3, -CF2H, and -OCF2H. B is one or two substituents independently selected from -F, -Cl, -Br, -I, -CF3, -OCF3, -CF2H, and -OCF2H. X is selected from -S- and -O-, m is between 0 and 3. Y is -H; Nitrile group; A 5- to 6-membered unsaturated heterocycle containing three heteroatoms selected from O and N, preferably two or three of which are nitrogen, and the heterocycle is monosubstituted or disubstituted with =S or =O, preferably with =O; or, Linear-C 1~3 Alkyl, linear-C 1~3 The alkyl group -(CH2)-, preferably one -(CH2)-, is optionally monosubstituted with -OH, and the linear-chain -C 1~3 The alkyl group's terminal -CH3 is -OR 1 Alternatively, -SR 1 And preferably -OR 1 It is monosubstituted with R 1 However, it is a straight chain -C which is -H, -(CH2)-COOH or -(CH2)-(CH2)-OH. 1~3 Alkyl; or, -(CH2) i -CX 1 R 2 (Here, X 1 is, =NR 3 Or = O, R 3 is -H or -OH, i is either 0 or 1. R 2 は-OR 4 or -NR5 R 6 And R 4 is -H or -C 1~2 It is alkyl, R 5 and R 6 (The element is selected independently of -H or -methyl); Selected from, Here, it is referred to as "Z", [ka] is an aliphatic group comprising at least one, preferably one or two heterocycles, the heterocycles collectively comprising two nitrogen atoms. Of the two nitrogen atoms, the first nitrogen atom is Z in formula I [ka] Linked to, the second of the two nitrogen atoms is Z in formula I [ka] Connected to, preferably, [ka] It is connected to the Z group, Each of the two nitrogen atoms is a heteroatom in one of the two linked four-membered heterocycles (each heterocycle has only one heteroatom), and the two linked four-membered heterocycles form a spirocycle, preferably the four-membered heterocycle having the first nitrogen atom is linked to the four-membered heterocycle having the second nitrogen atom via a shared carbon atom, thereby forming a spirocycle having two heteroatoms which are two nitrogen atoms, or The first of two nitrogen atoms is linked to a four-membered heterocycle having one heteroatom, the second of the two nitrogen atoms is the heteroatom in the four-membered heterocycle to which the first nitrogen atom is linked, and the first nitrogen atom is further substituted with methyl, or The present invention relates to compounds having two nitrogen atoms that are heteroatoms in a 6- to 7-membered heterocycle having two heteroatoms, and the heterocycle is optionally substituted with methyl, preferably monosubstituted, or to salts, complexes, diastereomers, enantiomers and / or tautomers of compounds having formula (I). In a preferred embodiment of the present invention, the compound is [ka] No.

[0016] In preferred embodiments of the present invention, the compound is a salt, complex, diastereomer, enantiomer and / or tautomer of a compound having formula (I).

[0017] In the context of the present invention, when a first chemical group is “substituted” by a second chemical group, this indicates that the second chemical group is bonded to the first chemical group, preferably by replacing a hydrogen atom of the first chemical group at the position where each bond between the first and second chemical groups is formed. In this context, the term “substituted” does not indicate the nature of a chemical reaction. Similarly, when a first chemical group is a “substituent” of a second chemical group, this indicates that the chemical groups are bonded to each other.

[0018] The terms “carbocyclic,” “carbocyclic,” or “carbocykryl,” unless otherwise indicated, generally encompass monocyclic rings of 3 to 9 members, preferably 4 to 8 members, 3 to 6 members, or 5 to 7 members, more preferably 5 or 6 members, containing 3 to 9, preferably 4 to 8, 3 to 6, or 5 to 7, more preferably 5 or 6 ring carbon atoms. In preferred embodiments, the carbocyclic according to the present invention contains 4, 5, 6, or 7 ring carbon atoms. The carbocyclic may be saturated, partially or completely unsaturated, or aromatic, where saturation means that only single bonds are present, partially or completely unsaturated means that Hückel's rule for aromaticity is not satisfied, while one or more double bonds may be present in appropriate positions, and aromatic means that Hückel's (4n+2) rule is satisfied. Thus, the terms “carbocyclic,” “carbocyclic,” or “carbocykryl” may encompass, in particular, cycloalkyl, cycloalkenyl, and phenyl. Preferably, the terms “carbocyclic,” “carbocyclyl,” or “carbocyclic” include cycloalkyl groups and cycloalkenyl groups, such as cyclopropane rings, cyclobutane rings, cyclopentane rings, and cyclohexane rings. In preferred embodiments, “carbocyclic,” “carbocyclic,” or “carbocyclyl” is a non-aromatic “carbocyclic,” “carbocyclic,” or “carbocyclyl.”

[0019] As used herein, the term "cycloalkyl" typically refers to a monocyclic or alicyclic group having 3 to 9 or 4 to 7 ring carbon atoms and no ring heteroatoms, unless otherwise specified. Examples of such cycloalkyls include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, cyclononyl, and cyclodecyl, or cyclopropyl, cyclobutyl, cyclopentyl, and cyclohexyl.

[0020] The terms "aryl" or "aromatic carbocyclic" preferably encompass six-membered aromatic carbocyclic rings based on carbon atoms as ring members. A preferred example is phenyl.

[0021] The terms “heterocyclic,” “heterocyclic,” or “heterocyclyl” generally encompass monocyclic rings of 3 to 9 members, preferably 4 to 7 members, preferably 5 to 7 members, 5 to 6 members, 6 to 7 members, or 3 to 5 members, particularly 4, 5, 6, or 7 members, unless otherwise indicated. A heterocyclic ring typically contains one or more heteroatoms selected from N, O, and S as ring members, for example, 1, 2, 3, or 4, preferably 1, 2, or 3, preferably 1 or 2, where the S atom as a ring member can exist as S, SO, or SO2, the N atom as a ring member can exist as N or NH, and the O atom as a ring member can exist as O. Preferably, the heteroatom is a N atom. The remaining ring members are carbon atoms.

[0022] In the context of the present invention, when referring to heteroatoms in a heterocycle, heteroatom "N" preferably refers to a ring member existing in the form of N or NH, heteroatom "O" preferably refers to a ring member existing in the form of O, and heteroatom "S" preferably refers to a ring member existing in the form of S, SO or SO2.

[0023] A heterocyclyl may be saturated, partially or completely unsaturated, or aromatic. Saturation means that only single bonds are present; partially or completely unsaturated means that one or more double bonds may be present in appropriate positions, but Hückel's rule for aromaticity is not satisfied; on the other hand, aromatic means that Hückel's (4n+2) rule is satisfied.

[0024] As used herein, the term “heterocycloalkyl” refers to the heterocyclil as defined above unless otherwise indicated, where the monocyclic ring is a monocyclic alicyclic group having typically 3 to 9 or 4 to 7 ring atoms, in each case 5 to 7 ring atoms, 5 to 6 ring atoms, 6 to 7 ring atoms, 3 to 5 ring atoms, and particularly 4, 5, 6, or 7 ring atoms, and the ring members include one or more heteroatoms selected from N, O, and S, for example 1, 2, 3, or 4, preferably 1, 2, or 3, preferably 1 or 2, where the S atom as a ring member may exist as S, SO, or SO2, the N atom as a ring member may exist as N or NH, and the O atom as a ring member may exist as O. The remaining ring members are carbon atoms. Preferably, unless otherwise indicated, the term “heterocycloalkyl” includes monocyclic rings with 3 to 9 members, preferably 4 to 7 members, preferably 5 to 7 members, 5 to 6 members, 6 to 7 members, or 3 to 5 members, particularly 4, 5, 6, or 7 members, respectively.

[0025] The terms “hetaryl,” “hetaryl,” “aromatic heterocycle,” or “aromatic heterocyclic ring” encompass monocyclic five- or six-membered aromatic heterocycles containing one, two, three, or four heteroatoms as ring members, unless otherwise indicated. In preferred embodiments, the heterocyclyl is a non-aromatic heterocyclyl.

[0026] As used herein, the term “spiro ring” means, unless otherwise indicated, a group having at least two, preferably two monocyclic rings, two of which are linked by a covalent atom that is a member of both linked monocyclic rings.

[0027] As used herein, the term "carbospiro ring" means, unless otherwise indicated, a group having at least two, preferably two, carbon rings, two of which are linked by a covalent atom that is a member of both linked carbon rings.

[0028] As used herein, the term “heterospiro ring” means, unless otherwise indicated, a group having at least two, preferably two monocyclic rings, wherein the monocyclic rings are linked by a covalent atom that is a member of both linked monocyclic rings, and at least one of the monocyclic rings is a heterocyclic ring.

[0029] In the context of the present invention, the spiro ring preferably comprises at least one, preferably one or two heteroatoms, the heteroatoms of which are preferably nitrogen. Preferably, the spiro ring in the context of the present invention comprises two monocyclic rings, each having four ring atoms.

[0030] In the context of the present invention, the term "alkyl" refers to a straight-chain saturated hydrocarbon or a branched-chain saturated hydrocarbon. For example, in certain embodiments, alkyl may refer to methyl, ethyl, propyl, isopropyl (also called 2-propyl or 1-methylethyl), butyl, isobutyl, tert-butyl, n-pentyl, iso-pentyl, sec-pentyl, neo-pentyl, 1,2-dimethyl-propyl, iso-amyl, n-hexyl, isohexyl, sec-hexyl, n-heptyl, iso-heptyl, n-octyl, 2-ethyl-hexyl, n-nonyl, n-decyl, n-undecyl, or n-dodecyl.

[0031] In a preferred embodiment, m is 0 to 2, more preferably 0 or 1, and most preferably m is 1. In a preferred embodiment, m is 0.

[0032] In a preferred embodiment, Y is not -H. In a preferred embodiment, Y is Nitrile group; A 5- to 6-membered unsaturated heterocycle containing three heteroatoms selected from O and N, preferably two or three of which are nitrogen, and the heterocycle being monosubstituted or disubstituted with =S or =O, preferably with =O; Linear-C 1~3 Alkyl, linear-C 1~3The alkyl group -(CH2)-, preferably one -(CH2)-, is optionally monosubstituted with -OH, and the linear-chain -C 1~3 The alkyl group's terminal -CH3 is -OR 1 Alternatively, -SR 1 And preferably -OR 1 It is monosubstituted with R 1 However, it is a straight chain -C which is H, -(CH2)-COOH or -(CH2)-(CH2)-OH. 1~3 Alkyl; or, -(CH2) i -CX 1 R 2 (Here, X 1 is, =NR 3 Or = O, R 3 is -H or -OH, i is either 0 or 1. R 2 は-OR 4 or -NR 5 R 6 And R 4 is -H or -C 1~2 It is alkyl, R 5 and R 6 (The element is selected independently of -H or -methyl); Selected from.

[0033] In a preferred embodiment, Y is not -H. In a preferred embodiment, Y is not a nitrile group. In a preferred embodiment, Y is A 5- to 6-membered unsaturated heterocycle containing three heteroatoms selected from O and N, preferably two or three of which are nitrogen, and the heterocycle being monosubstituted or disubstituted with =S or =O, preferably with =O; Linear-C 1~3 Alkyl, linear-C 1~3 The alkyl group -(CH2)-, preferably one -(CH2)-, is optionally monosubstituted with -OH, and the linear-chain -C 1~3 The alkyl group's terminal -CH3 is -OR 1 Alternatively, -SR 1 And preferably -OR1 is monosubstituted, and R 1 is H, -(CH2)-COOH or -(CH2)-(CH2)-OH, a linear -C 1~3 alkyl; or, -(CH2) i -CX 1 R 2 (where X 1 is =NR 3 or =O, and R 3 is -H or -OH, i is 0 or 1, R 2 is -OR 4 or -NR 5 R 6 and R 4 is -H or -C 1~2 [[ID=[]37]]alkyl, and R 5 and R 6 are independently selected from -H or -methyl); selected from.

[0034] In a preferred embodiment, Y is not -H. In a preferred embodiment, Y is a 5- to 6-membered unsaturated heterocyclic ring containing three heteroatoms selected from O and N, preferably two or three heteroatoms are nitrogen, the heterocyclic ring is =S or =O, preferably monosubstituted or disubstituted with =O, a 5- to 6-membered unsaturated heterocyclic ring; or, a linear -C 1~3 alkyl, a linear -C 1~3 alkyl's -(CH2)-, preferably one of the -(CH2)- is optionally monosubstituted with -OH, a linear -C 1~3 alkyl's terminal -CH3 is -OR 1 or -SR 1 and preferably monosubstituted with -OR 1 and R 1 is H, -(CH2)-COOH or -(CH2)-(CH2)-OH, a linear -C 1~3 alkyl; selected from.

[0035] In a preferred embodiment, Y is -H.

[0036] In a preferred embodiment, Y is a nitrile group.

[0037] In a preferred embodiment, Y is a 5- to 6-membered unsaturated heterocycle containing three heteroatoms selected from O and N, preferably two or three of which are nitrogen atoms, and the heterocycle is monosubstituted or disubstituted with =S or =O, preferably with =O.

[0038] In a preferred embodiment, Y is a linear-C 1~3 Selected from alkyl groups, linear-C 1~3 The alkyl group's -(CH2)- is arbitrarily monosubstituted with -OH, and the linear-C 1~3 The alkyl group's terminal -CH3 is -OR 1 or -SR 1 And preferably -OR 1 It is monosubstituted with R 1 is -H, -(CH2)-COOH, or -(CH2)-(CH2)-OH. More preferably, Y is a linear-chain -C 1~2 Selected from alkyl groups, linear-C 1~2 The alkyl group's -(CH2)- is arbitrarily monosubstituted with -OH, and the linear-C 1~2 The alkyl group's terminal -CH3 is -OR 1 or -SR 1 And preferably -OR 1 It is monosubstituted with R 1 is H, -(CH2)-COOH, or -(CH2)-(CH2)-OH. Most preferably, Y is -OR 1 or -SR 1 And preferably -OR 1 It is a monosubstituted methyl group, R 1 These are H, -(CH2)-COOH, or -(CH2)-(CH2)-OH.

[0039] In a preferred embodiment, Y is a linear-C 1~3 Selected from alkyl groups, linear-C 1~3The alkyl group's -(CH2)- is arbitrarily monosubstituted with -OH, and the linear-C 1~3 The alkyl group's terminal -CH3 is -OR 1 or -SR 1 And preferably -OR 1 It is monosubstituted with R 1 is -H or -(CH2)-COOH. More preferably, Y is a linear -C 1~2 Selected from alkyl groups, linear-C 1~2 The alkyl group's -(CH2)- is arbitrarily monosubstituted with -OH, and the linear-C 1~2 The alkyl group's terminal -CH3 is -OR 1 or -SR 1 And preferably -OR 1 It is monosubstituted with R 1 is H or -(CH2)-COOH. Most preferably, Y is -OR 1 or -SR 1 And preferably -OR 1 It is a monosubstituted methyl group, R 1 This is either H or -(CH2)-COOH.

[0040] In a preferred embodiment, Y is a linear-C 1~3 Selected from alkyl groups, linear-C 1~3 The alkyl group's -(CH2)- is arbitrarily monosubstituted with -OH, and the linear-C 1~3 The alkyl group's terminal -CH3 is -OR 1 or -SR 1 And preferably -OR 1 It is monosubstituted with R 1 is -(CH2)-COOH. More preferably, Y is a linear-C 1~2 Selected from alkyl groups, linear-C 1~2 The alkyl group's -(CH2)- is arbitrarily monosubstituted with -OH, and the linear-C 1~2 The alkyl group's terminal -CH3 is -OR 1 or -SR 1 And preferably -OR 1 It is monosubstituted with R 1 is -(CH2)-COOH. Most preferably, Y is -OR 1or -SR 1 And preferably -OR 1 It is a monosubstituted methyl group, R 1 It is -(CH2)-COOH.

[0041] In a preferred embodiment, Y is -(CH2) i -CX 1 R 2 (Here, X 1 is, =NR 3 Or = O, R 3 is -H or -OH, and i is 0 or 1. R 2 is -OR 4 or -NR 5 R 6 And R 4 is -H or -C 1~2 It is alkyl, R 5 and R 6 (is independently selected from -H or -methyl).

[0042] In a preferred embodiment, i is 0.

[0043] In a preferred embodiment, R 3 It is -OH.

[0044] In a preferred embodiment, R 5 and R 6 Both are -H.

[0045] In a preferred embodiment, i is 0, and X 1 is=NR 3 And R 3 is -OH, and R 2 -NR 5 R 6 And R 5 and R 6 Both are -H.

[0046] In a preferred embodiment, i is 0 or 1, and X 1 is = O, and R 2-NR 5 R 6 And R 5 and R 6 Both are -H.

[0047] In a preferred embodiment, X 1 is = O, and R 2 は-OR 4 And R 4 is -H or -C 1~2 It is alkyl, preferably H.

[0048] In a preferred embodiment, when referring to Y, the 5- to 6-membered unsaturated heterocycle containing three heteroatoms, which are nitrogen, A five-membered unsaturated heterocycle comprising three heteroatoms selected from O and N, preferably two or three of which are nitrogen, and the five-membered unsaturated heterocycle being monosubstituted with =S or =O, preferably =O; A six-membered heterocycle containing three nitrogen heteroatoms, which are disubstituted with =O or =S, preferably with =O; That is the case.

[0049] In a preferred embodiment, when referring to Y, the 5- to 6-membered unsaturated heterocycle containing three heteroatoms that are nitrogen is a 5-membered heterocycle containing three heteroatoms that are nitrogen, and the 5-membered heterocycle is monosubstituted with =O.

[0050] In a preferred embodiment, Y is Selected from the group consisting of TIFF2026509430000012.tif176170, more preferably Y is Selected from the group consisting of TIFF2026509430000013.tif165170, and more preferably Y is Selected from the group consisting of TIFF2026509430000014.tif124170.

[0051] In a more preferred embodiment, Y is Selected from the group consisting of TIFF2026509430000015.tif84170, more preferably Y is Selected from the group consisting of TIFF2026509430000016.tif112170.

[0052] In a more preferred embodiment, Y is Selected from the group consisting of TIFF2026509430000017.tif115170.

[0053] In a preferred embodiment, Z is an aliphatic group comprising two nitrogen atoms and one heterocycle, wherein the first of the two nitrogen atoms constitutes Z. [ka] Linked to, the second nitrogen atom of the two nitrogen atoms is Z, [ka] Preferably, [ka] Linked to the other, the two nitrogen atoms are heteroatoms in a 6- to 7-membered heterocycle having two heteroatoms, and the heterocycle is optionally substituted, preferably monosubstituted, with methyl atoms. In a preferred embodiment, the 6- to 7-membered heterocycle having two heteroatoms is a 6-membered heterocycle having two heteroatoms. In another preferred embodiment, the 6- to 7-membered heterocycle having two heteroatoms is a 7-membered heterocycle having two heteroatoms.

[0054] In a preferred embodiment, Z is Selected from the group consisting of TIFF2026509430000021.tif115170.

[0055] In a more preferred embodiment, Z is [ka] Selected from.

[0056] In the most preferred embodiment, Z is [ka] That is the case.

[0057] In a preferred embodiment, the compound is of formula II: [ka] (wherein B, X, A, Y, m, and Z are defined in any embodiment herein)

[0058] In the most preferred embodiment, the compound is of formula III: [ka] (wherein B, X, A, Y, and Z are defined in any embodiment herein)

[0059] In a preferred embodiment, B is selected from -Cl or -CF3. In a particularly preferred embodiment, B is -Cl. In a particularly preferred embodiment, B is -CF3. In a preferred embodiment, B is defined in any other embodiment of this specification except that B is not -CF3. In a preferred embodiment, B is defined in any other embodiment of this specification except that B is not -Cl.

[0060] In a preferred embodiment, A is selected from -H or -F. In the most preferred embodiment, A is H.

[0061] In the most preferred embodiment, X is -O-. In a preferred embodiment of the present invention, A is one or two substituents independently selected from -H, -F, -Cl, -Br, -I, -CF3, -OCF3, -CF2H, and -OCF2H. B is one or two substituents independently selected from -F, -Cl, -Br, -I, -CF3, -OCF3, -CF2H, and -OCF2H, X is selected from -S- and -O-, m is 1, Y is selected from TIFF2026509430000026.tif122170, Z is selected from TIFF2026509430000027.tif111170.

[0062] In a preferred embodiment, the compound has the formula II:

Chemical formula

[0063] In the most preferred embodiment, the compound has the formula III:

Chemical formula

[0064] In a preferred embodiment of the present invention, the compound has the formula (III), A is selected from -H or -F, and B is selected from -Cl or -CF3. In a more preferred embodiment of the present invention, the compound has the formula (III), A is selected from -H or -F, B is selected from -Cl or -CF3, and X is -O-. In a more preferred embodiment of the present invention, the compound has the formula (III), A is selected from -H or -F, B is selected from -Cl or -CF3, X is -O-, and Z is

Chemical formula

[0065] In a more preferred embodiment of the present invention, the compound has formula (III), where A is selected from -H or -F, B is selected from -Cl or -CF3, X is -O-, and Z is [ka] And Y is, Selected from TIFF2026509430000033.tif112170.

[0066] In a preferred embodiment, the compound has formula (III), where A is -H and B is -Cl. In a preferred embodiment, the compound has formula (III), where A is -H, B is -Cl, and X is -O-. In a more preferred embodiment, the compound has formula (III), where A is -H, B is -Cl, X is -O-, and Z is [ka] In a preferred embodiment, the compound has formula III, where A is -H, B is -Cl, X is -O-, and Y is Selected from TIFF2026509430000035.tif112170.

[0067] In a more preferred embodiment, the compound has formula III, where A is -H, B is -Cl, X is -O-, and Z is [ka] And Y is, Selected from TIFF2026509430000037.tif112170.

[0068] In a preferred embodiment of the present invention, the compound has the formula (II), (i) A is selected from -H or -F, (ii) B is selected from -Cl or -CF3, (iii) m is 1, (iv) Y is selected from the group consisting of TIFF2026509430000038.tif157170, Z is selected from the group consisting of TIFF2026509430000039.tif123170, Preferably, X is -O-.

[0069] In the context of the present invention, the compounds are referred to by the following numbering.

[0070] TIFF2026509430000040.tif181170TIFF2026509430000041.tif213170TIFF2026509430000042.tif254170TIFF2026509430000043.tif235170TIFF2026509430000044.tif222170TIFF2026509430000045.tif91170

[0071] In a particularly preferred embodiment, the compound is selected from the group consisting of Compound 1 to Compound 21. Preferably, the compound is selected from the group consisting of Compound 2 to Compound 21. Preferably, the compound is selected from the group consisting of Compound 2, Compound 3, Compound 4, Compound 5, Compound 6, Compound 7, Compound 8, Compound 9, Compound 10, Compound 11, Compound 12, Compound 18, Compound 19, Compound 20, Compound 21. More preferably, the compound is selected from the group consisting of Compound 2, Compound 3, Compound 4, Compound 5, Compound 6, Compound 7, Compound 8, Compound 9, and Compound 10.

[0072] In a particularly preferred embodiment, the compound is selected from the group consisting of compound 6, compound 8, compound 9, compound 10, compound 11, compound 12, compound 15, compound 16, compound 17, compound 18, compound 19, compound 20, and compound 21; more preferably, the compound is selected from the group consisting of compound 6, compound 8, compound 9, compound 10, compound 11, compound 12, compound 18, compound 19, compound 20, and compound 21; and more preferably, the compound is selected from the group consisting of compound 6, compound 8, compound 9, and compound 10.

[0073] In a particularly preferred embodiment, the compound is selected from the group consisting of compound 6, compound 7, compound 8, compound 9, compound 10, compound 11, compound 15, compound 16, compound 17, compound 18, compound 19, compound 20, and compound 21; more preferably, the compound is selected from the group consisting of compound 6, compound 7, compound 8, compound 9, compound 10, compound 11, compound 18, compound 19, compound 20, and compound 21; and more preferably, the compound is selected from the group consisting of compound 6, compound 7, compound 8, compound 9, and compound 10.

[0074] In a particularly preferred embodiment, the compound is selected from the group consisting of compound 10, compound 11, compound 15, compound 18, compound 19, and compound 20. In a particularly preferred embodiment, Y is [ka] And m is arbitrarily 1.

[0075] In preferred embodiments of the present invention, the compound is selected from the group consisting of compound 3 and compounds 5 to 21, preferably from the group consisting of compound 3 and compounds 5 to 14, preferably from the group consisting of compound 3 and compounds 5 to 11, and more preferably from the group consisting of compound 3 and compounds 5 to 10. In preferred embodiments, the compound is selected from the group consisting of compound 3, compounds 5 to 10, and compounds 18 to 21. In preferred embodiments, the compound is one of compounds 2 to 21.

[0076] In a preferred embodiment of the present invention, if X is -S-, then Y is not -(CH)2-O-CH2-CH2-OH. In a preferred embodiment of the present invention, X is -S-, B is -Cl, A is -H, m is 1, and Z is [ka] If this is the case, then Y is not -(CH)2-O-CH2-CH2-OH

[0077] In a preferred embodiment of the present invention, when m is 0, Y is not -H. In a preferred embodiment of the present invention, when m is 1, Y is not -H. In a preferred embodiment of the present invention, when m is 0 and B is optionally further Cl, Y is not -(CH2)-(CH2)-OH. In a preferred embodiment of the present invention, when m is 1 and B is optionally further Cl, Y is not -(CH2)-OH.

[0078] In a preferred embodiment of the present invention, the compound is [ka] It is not a compound selected from the group. In a preferred embodiment, the compound is [ka] isn't it.

[0079] In a preferred embodiment, the compound is a potassium channel in cells, preferably a potassium channel Slack(K) Na It has activity as an activator of 1.1). In the context of the present invention, the terms “Slack” or “Slack channel” refer to a class of sodium-activated potassium channels. These potassium channels are primarily expressed in nerve tissue, where they are involved in neuronal excitability, burst firing, and firing rate adaptation. Preferably, Slack channels are encoded by the gene Kcnt1. Preferably, Slack channels refer to any isoform of potassium channel subfamily T member 1 disclosed in UniProt entries Q5JUK3 and Q6ZPR4 (as of February 2, 2023).

[0080] In a preferred embodiment, the compound has an EC of less than 100 μM, preferably 0.5 μM to 80 μM. 50 This activates potassium channels. In this context, EC 50 Preferably, 2 mM Ca 2+ , 2 mM Mg 2+ and Na containing 0.03% DMSO + EC determined in a buffer that does not contain EC 50 It refers to.

[0081] In preferred embodiments, the compound does not cross the blood-brain barrier (BBB) ​​when administered to animals, or crosses the BBB less effectively than a reference compound such as roxapine.

[0082] In preferred embodiments, the compound does not bind to human dopamine receptors, and more preferably, the compound is less likely to bind to human dopamine receptors compared to a reference compound such as roxapine.

[0083] In a preferred embodiment of the present invention, the compound is an α-1A adrenergic receptor (α 1A ), dopamine receptor D1 (D1), short form of dopamine receptor D2 (D 2S or D 2Sh), histamine H1 receptor (H1), serotonin receptor (preferably 5-HT 2A and 5-HT 2B ), binds to an off-target selected from the group consisting of norepinephrine transporter (NET), serotonin transporter (SET), and cannabinoid receptor (CB1). In a preferred embodiment of the present invention, the compound binds to the α-1A adrenergic receptor (α 1A ), dopamine receptor D1 (D1), short form of dopamine receptor D2 (D 2S or D 2Sh ), histamine H1 receptor (H1), serotonin receptor (preferably 5-HT 2A and 5-HT 2B ), binds to a target selected from the group consisting of norepinephrine transporters (NETs) and serotonin transporters (SETs). In preferred embodiments of the present invention, the compound binds to dopamine receptor D1 (D1), histamine H1 receptor (H1), and serotonin receptor (preferably 5-HT1). 2A and 5-HT 2B The compound binds to a target selected from the group consisting of ). By binding to such off-targets, the antipruritic effect of the compound of the present invention can be further enhanced. In a preferred embodiment, the compound binds to an H1 receptor (preferably, the compound of the present invention is an antagonist of the H1 receptor). In a further preferred embodiment, the compound of the present invention binds to a 5-HT receptor, more preferably a 5-HT2 receptor, and especially a 5-HT 2A The compound binds to receptors, preferably 5-HT receptors, 5-HT2 receptors and / or 5-HT2 receptors. 2A It is a receptor agonist.

[0084] In a preferred embodiment of the present invention, the compound is compound 6, compound 8, compound 10, or compound 11. In a preferred embodiment of the present invention, the compound is compound 6, compound 8, or compound 10. In a more preferred embodiment of the present invention, the compound is compound 6 or compound 10.

[0085] In another preferred embodiment of the present invention, the compound is compound 6 or compound 21. In a more preferred embodiment of the present invention, the compound is compound 21.

[0086] In preferred embodiments of the present invention, the compound is isolated. In preferred embodiments, the compound is dissolved in a solvent or dispersion. In preferred embodiments, the compound has a purity of at least 75%, optionally at least 90%, and optionally at least 95%. In this context, purity refers to the absence of any foreign matter (especially chemical substances) that may be present in a composition containing the compound of the present invention. Impurities may be naturally occurring or may be added or generated during the synthesis and / or purification of the compound of the present invention. For example, impurities may include starting materials, solvents, intermediates or reactants, decomposition products of any of the above or the desired compound, the remainder of protecting groups after deprotection, and combinations thereof.

[0087] The compounds of the present invention may exist in tautomer forms. Preferably, a description of a single tautomer, such as a functional group or a single tautomer in a compound, is understood to represent the compound or group in all of its tautomer forms. For example, unless otherwise indicated, compound 10, compound 11, compound 15, compound 17, compound 18, compound 19, and / or compound 20 preferably refer to both of the following tautomer forms, respectively.

[0088] TIFF2026509430000050.tif228170TIFF2026509430000051.tif175170

[0089] Preferably, compound 10 is compound 10 tautomer 1 or compound 10 tautomer 2. Preferably, compound 11 is compound 11 tautomer 1 or compound 11 tautomer 2. Preferably, compound 15 is compound 15 tautomer 1 or compound 15 tautomer 2. Preferably, compound 17 is compound 17 tautomer 1 or compound 17 tautomer 2. Preferably, compound 18 is compound 18 tautomer 1 or compound 18 tautomer 2. Preferably, compound 19 is compound 19 tautomer 1 or compound 19 tautomer 2. Preferably, compound 20 is compound 20 tautomer 1 or compound 20 tautomer 2.

[0090] In a second embodiment, the present invention relates to a pharmaceutical composition comprising any one of the compounds described herein, or a salt, solvate, or ester thereof, and a pharmaceutically acceptable carrier or excipient.

[0091] Preferably, a pharmaceutically acceptable composition comprises a compound according to any embodiment of the present invention, the compound in the form of a prodrug, salt, racemic mixture, crystalline form, polymorph, solvate, or combination thereof.

[0092] The term “pharmaceutically acceptable carrier or excipient” means a carrier or excipient useful in the preparation of a pharmaceutical composition, including excipients that are generally safe, non-toxic, not biologically or otherwise undesirable, and acceptable for both veterinary and human medicinal use. This includes solvents, emulsifiers, suspending agents, disintegrants, binders, stabilizers, diluents, gelling agents, preservatives, lubricants, surfactants, and other similar carriers. Such a pharmaceutically acceptable carrier or excipient must be compatible with the other components of the composition and “acceptable” in the sense that it is not harmful to the patient.

[0093] In preferred embodiments of the pharmaceutical composition, the compound, or its salt, solvate, or ester, is present in an amount of 0.1% w / w to 10% w / w, optionally 1% w / w to 10% w / w. In preferred embodiments, the salt of the compound is the hydrochloride salt of the compound.

[0094] In preferred embodiments of the pharmaceutical composition, the composition is an oral composition, a topical composition, or an injectable. In preferred embodiments of the pharmaceutical composition, the composition is an oral composition, a topical composition, or a parenteral composition. In preferred embodiments, the composition is in the form of a patch, suppository, syringe, injector pen, spray bottle, mask, pastille and / or implant inhaler, nebulizer, cream and / or vaporizer. A preferred embodiment of the pharmaceutical composition is a unit dosage form. In a particular preferred embodiment, the composition is in the form of a cream.

[0095] In a preferred embodiment of the composition, the dosage form of the composition is a sustained-release dosage form. Preferably, the sustained-release dosage form releases the compound over 5 hours, 10 hours, 18 hours, 24 hours, 36 hours, 48 ​​hours, 72 hours, 1 week, 2 weeks, 1 month, 6 months, 12 months and / or 2 years. In this context, "over ~" includes releases in which the compound is continuously, periodically, or irregularly released into the body of the subject. Preferably, the sustained-release dosage form is an implant. Preferably, the implant contains the compound of the first embodiment of the present invention.

[0096] The present invention further relates to the following special preferred embodiments. These special preferred embodiments can be combined with any aspect, embodiment, and claim herein.

[0097] In a particularly preferred embodiment, Y is a linear-C 1~3 Selected from alkyl groups, linear-C 1~3 The alkyl group's terminal -CH3 is -OR 1 It is monosubstituted with R 1 is -(CH2)-COOH. More preferably, Y is a linear-C 1~2 Selected from alkyl groups, linear-C 1~2 The alkyl group's terminal -CH3 is -OR 1 It is monosubstituted with R 1is -(CH2)-COOH. Most preferably, Y is -OR 1 It is a monosubstituted methyl group, R 1 It is -(CH2)-COOH.

[0098] In a particularly preferred embodiment, m=1, and Y is a linear-C 1~3 Selected from alkyl groups, linear-C 1~3 The alkyl group's terminal -CH3 is -OR 1 It is monosubstituted with R 1 is -(CH2)-COOH. More preferably, m=1 and Y is a linear-C 1~2 Selected from alkyl groups, linear-C 1~2 The alkyl group's terminal -CH3 is -OR 1 It is monosubstituted with R 1 It is -(CH2)-COOH. Most preferably, m=1 and Y is -OR 1 It is a monosubstituted methyl group, R 1 It is -(CH2)-COOH.

[0099] In a particularly preferred embodiment, when referring to Y, a 5- to 6-membered unsaturated heterocycle containing three heteroatoms selected from O and N is, A five-membered unsaturated heterocycle containing three heteroatoms that are N, which are monosubstituted with =S or =O, preferably =O; or, A six-membered heterocycle containing three nitrogen heteroatoms, which are disubstituted with =O or =S, preferably with =O; That is the case.

[0100] In a preferred embodiment, when referring to Y, the 5- to 6-membered unsaturated heterocycle containing three heteroatoms selected from O and N is a 5-membered heterocycle containing three heteroatoms that are nitrogen, and the 5-membered heterocycle is monosubstituted with =O.

[0101] In a particular embodiment, the present invention relates to a compound or composition for the treatment of a disease in a subject, wherein the composition comprises an effective amount of a compound according to the first embodiment of the present invention, or a salt, solvate, or ester thereof.

[0102] In a third embodiment, the present invention relates to a compound or composition for the treatment of a disease in a subject, wherein the composition comprises an effective amount of a compound according to the first embodiment of the present invention, or a salt, solvate, or ester thereof, the disease being treatable by activating potassium channels in cells associated with the pathology of the disease, and the treatment comprising administering the compound or composition to a subject in need of treatment.

[0103] Preferably, the subject of the present invention is an animal. In this context, the term “animal” includes humans. Preferably, the animal is a mammal or a bird, and most preferably a mammal. Preferably, the mammal is a human. Preferably, the mammal is a non-human primate, dog, cat, sheep, cattle, goat, pig, mouse, rat, rabbit, or guinea pig.

[0104] In this context, the term “treatment” includes the application or administration of a therapeutic agent (such as the compounds of the present invention) or procedure to a patient in need of treatment, the purpose of which is to cure, restore, alleviate, eliminate, alter, correct, mitigate, improve, act upon, or prevent a disease, symptoms of a disease, or predisposition to a disease. Therefore, the term “treatment” may encompass preventive treatment of a disease or symptoms of a disease.

[0105] Preferably, the application or administration includes intravenous, intraperitoneal, subcutaneous, intramuscular, intrathecal, epidural, topical, oral, gastric, and / or rectal administration. In preferred embodiments, the application or administration includes intrathecal and / or epidural administration. In preferred embodiments, the application or administration includes intravenous, intraperitoneal, subcutaneous, intramuscular, topical, oral, gastric, and / or rectal administration. Administration to the CNS, particularly intrathecal or epidural administration, is likely to produce a stronger analgesic effect in cases of neuropathic pain compared to other types of administration such as oral or intraperitoneal administration.

[0106] In a preferred embodiment, the disease is a pruritic disorder, such as acute or chronic itching. In a preferred embodiment, the pruritic disorder is related to a secondary condition, such as those selected from the following: Dermatological disorders such as xerosis or xerosis (dry skin), dermatitis or eczema (e.g., atopic dermatitis), psoriasis (e.g., psoriasis vulgaris), prurigo (e.g., nodular prurigo), urticaria (e.g., chronic idiopathic urticaria), connective tissue disorders (e.g., dermatomyositis), and post-burn pruritus; Renal impairment (e.g., chronic kidney disease, chronic renal failure, or end-stage renal disease), dialysis (e.g., hemodialysis), uremic pruritus; Hepatobiliary disorders (e.g., cholestasis, primary biliary cholangitis, primary sclerosing cholangitis, secondary sclerosing cholangitis, hepatitis, toxic liver disease, chronic liver disease, or cirrhosis), cholestatic pruritus; Endocrine disorders (e.g., hyperthyroidism or diabetes mellitus); Metabolic disorders (e.g., iron deficiency or iron overload); Benign or malignant neoplasms (e.g., solid tumors, carcinomas, or hematological malignancies (e.g., Hodgkin lymphoma, non-Hodgkin lymphoma, myeloproliferative disorders, or polycythemia vera)); Infectious diseases (e.g., viral infections such as herpes simplex, herpes zoster, chickenpox, human immunodeficiency virus (HIV), and infections associated with hepatitis; bacterial infections or parasitic diseases); Neurological disorders (e.g., neurodegenerative diseases, multiple sclerosis, brain tumors, postherpetic neuralgia, small fiber neuropathy, brachioradialis pruritus, or dorsal paresthesia), neurogenic itching, neurogenic pruritus; Mental illness (e.g., depression, obsessive-compulsive disorder, delusional disorder, eating disorder, or anxiety disorder); Drug-induced pruritus (e.g., caused by opioids, antibiotics, antimalarial drugs, ACE inhibitors, angiotensin receptor blockers, antiarrhythmics, antidepressants, antidiabetic drugs, antihypertensive drugs, anticonvulsants, anti-inflammatory drugs, beta-blockers, bronchodilators, calcium channel blockers, diuretics, hormones, immunosuppressants, antilipids, nerve blockers, plasma expanders, tranquilizers, or uric acid-lowering drugs); Age-related pruritus; Pregnancy pruritus; and / or, Chronic idiopathic pruritus.

[0107] In a preferred embodiment of the present invention, pruritic disorders are associated with secondary conditions such as those selected from the following: Dermatological disorders such as xerosis or xerosis (dry skin), dermatitis or eczema (e.g., atopic dermatitis), psoriasis (e.g., psoriasis vulgaris), prurigo (e.g., nodular prurigo), urticaria (e.g., chronic idiopathic urticaria), connective tissue disorders (e.g., dermatomyositis), and post-burn pruritus; Renal impairment (e.g., chronic kidney disease, chronic renal failure, or end-stage renal disease), dialysis (e.g., hemodialysis), uremic pruritus; Hepatobiliary disorders (e.g., cholestasis, primary biliary cholangitis, hepatitis, chronic liver disease, or cirrhosis), cholestatic pruritus; Benign or malignant neoplasms (e.g., solid tumors, carcinomas, or hematological malignancies (e.g., Hodgkin lymphoma, non-Hodgkin lymphoma, or polycythemia vera)); Neurogenic itching (e.g., brachioradialis pruritus or dorsal paresthesia), neurogenic itching; Drug-induced pruritus (e.g., caused by opioids); Age-related pruritus; and / or, Chronic idiopathic pruritus.

[0108] In a preferred embodiment, the myeloproliferative disorder is a myeloproliferative neoplasm.

[0109] In preferred embodiments, the composition used is a pharmaceutical composition according to a second aspect of the present invention. Furthermore, treatment may include the application or administration of a further antipruritic agent before, simultaneously with, or after the application or administration of a compound or composition of the present invention. Preferably, the further antipruritic agent is selected from systemic agents, oral agents and / or topical agents. Preferably, the further antipruritic agent is selected from the group of corticosteroids, antihistamines, anesthetics, phosphodiesterase-4 inhibitors, capsaicin, μ-opioid receptor antagonists, antidepressants, immunosuppressants, anticonvulsants, Janus kinase inhibitors, κ-opioid receptor agonists and thalidomide-based and / or butorphanol-based drugs.

[0110] Preferably, the treatment according to the present invention is performed before, simultaneously with, or after further antipruritic treatment, which preferably is selected from the group of applications or administrations of aluminum triacetate solution, olive oil, jewelweed, calamine lotion, sodium bicarbonate paste, ammonium hydroxide and / or papain-based topical creams, cannabis, purslane (portulaca oleracaea), ashoka (sarco asoca or saraca asoca), fig (fificus carica) and / or cannabinoids.

[0111] In a preferred embodiment, the disease is pain or a pain-related adverse sensation in the subject.

[0112] In a preferred embodiment, pain or pain-related adverse sensations are Neuropathic pain induced by traumatic nerve injury, cancer and cancer treatment (e.g., chemotherapy), neurological disorders (e.g., multiple sclerosis), neurodegenerative diseases (e.g., Parkinson's disease), trigeminal neuralgia, diabetic peripheral neuropathy, stroke, herpes zoster, HIV, leprosy, Guillain-Barré syndrome, vascular diseases, vascular malformations, and autoimmune diseases. acute postoperative pain, Inflammatory pain, rheumatoid arthritis, osteoarthritis, and / or Pain dysregulation, Selected from.

[0113] In a preferred embodiment, the composition comprises at least one additional therapeutic agent and / or pain reliever. Preferably, the at least one additional therapeutic agent is an agent for the treatment of a secondary pathological condition as described herein.

[0114] In a fourth embodiment, the present invention relates to a method for synthesizing a compound according to the first embodiment of the present invention.

[0115] In this context, a method for synthesizing a compound may include one or more synthesis steps. Preferably, the synthesis steps are a chemical reaction step and / or a purification step.

[0116] In particular, the method of supplying the compound may include any of the synthetic steps and / or similar synthetic steps disclosed herein, which are readily known and available to those skilled in the art of organic synthesis.

[0117] In particular, the method for supplying the compound may include any chemical reaction steps and / or similar chemical reaction steps disclosed herein, which are readily known and available to those skilled in the art of organic synthesis.

[0118] In particular, the method for supplying the compound may include any purification steps and / or similar purification steps disclosed herein, which are readily known and available to those skilled in the art of organic synthesis.

[0119] Preferably, the chemical reaction step is carried out before or after the purification step.

[0120] In this context, purification refers to any preferred method for removing impurity fractions. Such purification methods are known and include, for example, column chromatography, selective precipitation, tritulation, and elution of impurities with a preferred solvent in which the desired compound is insoluble. The impurity fraction removed may be such that the compound is prepared in a substantially pure form, i.e., with the percentage purity described above. In other embodiments, the compound of the present invention is supplied in an impure form by the method.

[0121] The method for synthesizing the compounds according to the present invention can be started, interrupted, and continued at any point. Preferably, the method for synthesizing the compounds according to the present invention is interrupted after a chemical reaction step or a purification step. If possible, the method includes the isolation of by-products or intermediates of the chemical reaction step of the present invention.

[0122] The synthesis steps, such as the chemical reaction step or the purification step according to the present invention, can be carried out using any suitable solvent. For example, the choice of solvent may depend on the stability of the product, the polarity of the solvent and / or the compound, the boiling point of the solvent, the acidity of the solvent, whether the solvent is protic / aprotic, and / or the biocompatibility of the solvent. Preferred solvents include aliphatic alcohols (methanol, ethanol, n-propanol, isopropanol, etc.), acetone, acetonitrile, ether, N,N-dimethylformamide (DMF), tetrahydrofuran (THF), dichloromethane, water, or mixtures of two or more of these liquids. Preferably, the synthesis steps according to the present invention can be carried out under an inert gas such as nitrogen, helium, neon, argon, krypton, xenon, radon, and / or sulfur hexafluoride. Furthermore, the solvent used in the synthesis steps, such as the chemical reaction step or the purification step, can be purified using such gases.

[0123] In a preferred embodiment, a method for synthesizing the compound according to the present invention includes a synthesis step, preferably a chemical reaction step, for synthesizing a lactam core. Preferably, the lactam core has a structure [ka] (wherein A, X, and B are defined in any embodiment herein)

[0124] In a preferred embodiment, the lactam core of the present invention is synthesized according to synthesis route A or synthesis route B, the corresponding chemical reaction steps shown in Table 1. The individual chemical reaction steps in synthesis routes A and B are preferably performed sequentially, and therefore, Chemical reaction step 1a → Chemical reaction step 2a → Chemical reaction step 3a, or Chemical reaction step 1b → Chemical reaction step 2b, It will be done in this order.

[0125] TIFF2026509430000053.tif230170TIFF2026509430000054.tif56170

[0126] In a further preferred embodiment, a method for synthesizing the compounds according to the present invention includes a synthesis step, preferably a chemical reaction step, of converting a lactam core to its imidoyl chloride derivative and / or converting the lactam core and / or its imidoyl chloride derivative to the compounds of the present invention. The above conversion of the lactam core to its imidoyl chloride derivative and / or converting the lactam core and / or its imidoyl chloride derivative to the compounds of the present invention is preferably carried out using synthesis route C, the corresponding chemical reaction steps shown in Table 2. The individual chemical reaction steps in synthesis route C are preferably carried out sequentially, and therefore in the order of chemical reaction step 1c → chemical reaction step 2c.

[0127] TIFF2026509430000055.tif176170

[0128] In a preferred embodiment, a method for synthesizing the compound according to the present invention includes carrying out synthesis route A, followed by synthesis route C. In a preferred embodiment, a method for synthesizing the compound according to the present invention includes carrying out synthesis route B, followed by synthesis route C.

[0129] In a preferred embodiment, synthesis route B includes an additional chemical reaction step 0b that is carried out before chemical reaction step 1b. [ka] (Chemical reaction step 0b)

[0130] In a preferred embodiment of the present invention, [ka] but, [ka] In that case, preferably, [ka] If is piperazine or homopiperazine, synthesis route C includes chemical reaction step 3c after chemical reaction step 2c: [ka] (Chemical reaction step 3c) (In the formula, Y 1 Y 1 (Defined as Y having the additional characteristic that it is not H).

[0131] In a preferred embodiment, a method for synthesizing a compound according to the present invention includes initiating the synthesis from any synthesis step, preferably any chemical reaction step, or a purification step performed before or after any chemical reaction step disclosed herein. In a preferred embodiment, a method for synthesizing a compound according to the present invention includes carrying out a reaction using a reaction intermediate or reaction product of a synthesis step, preferably a chemical reaction step, as disclosed herein, preferably in synthesis route A, synthesis route B and / or synthesis route C.

[0132] In a preferred embodiment, a method for synthesizing a compound according to the present invention comprises carrying out at least one synthesis step, preferably at least one chemical reaction step, which is disclosed herein, preferably in synthesis route A, synthesis route B and / or synthesis route C.

[0133] In a preferred embodiment of the present invention, chemical reaction step 1a is carried out in a solvent containing potassium carbonate. In a preferred embodiment, chemical reaction step 1a is carried out in N,N-dimethylformamide (DMF). In a preferred embodiment, chemical reaction step 1a is carried out at 100°C to 140°C, most preferably at 120°C.

[0134] In a preferred embodiment, chemical reaction step 2a is carried out using a reducing agent, preferably SnCl2·2H2O, and optionally chemical reaction step 2a is carried out in a 1:1 mixture of ethanol / concentrated HCl.

[0135] In a preferred embodiment, the chemical reaction step 3a is: [ka] The process includes treating with an acid, more preferably sulfuric acid, and most preferably concentrated sulfuric acid. In a preferred embodiment, the reaction is carried out in DMF. In a preferred embodiment, chemical reaction step 3a is carried out in DMF at 100°C to 140°C, most preferably 120°C.

[0136] In a preferred embodiment, chemical reaction step 0b involves thionyl chloride, [ka] This includes the reaction of adding to [the substance].

[0137] In a preferred embodiment of the present invention, chemical reaction step 1b is: [ka] of, [ka] This includes adding dropwise to the solution. In a preferred embodiment, the chemical reaction step 1b is carried out at -20°C to 50°C, more preferably -10°C to 30°C, and most preferably 0°C.

[0138] In a preferred embodiment of the present invention, chemical reaction step 2b is carried out in a basic solution, preferably the basic solution containing NaOH. In a preferred embodiment, chemical reaction step 2b is carried out in DMF. In a preferred embodiment of the present invention, chemical reaction step 2b is carried out at 120°C to 170°C, more preferably 140°C to 160°C, and most preferably 150°C.

[0139] In a preferred embodiment, chemical reaction step 1c is carried out in phosphorus oxychloride, more preferably under reflux, and optionally the phosphorus oxychloride contains N,N-dimethylaniline.

[0140] In a preferred embodiment of the present invention, chemical reaction step 2c is carried out in p-xylene, and the reaction is optionally carried out at 110°C to 150°C, preferably 120°C to 140°C.

[0141] In a preferred embodiment of the present invention, chemical reaction step 3c is carried out using an additional base in the reaction mixture, preferably triethylamine.

[0142] In preferred embodiments, particularly when referring to the first and fourth embodiments of the present invention, A is one substituent. In preferred embodiments, A is two substituents. In preferred embodiments, B is one substituent. In preferred embodiments, B is two substituents. Preferably, when B is two substituents, B is substituents at carbon positions 2 and 3, and more preferably, B is substituent-Cl at carbon positions 2 and 3. In preferred embodiments, A and B are each one substituent.

[0143] In preferred embodiments, and particularly when referring to the first and fourth aspects of the present invention, A is one substituent. In preferred embodiments, A is two substituents. In preferred embodiments, B is one substituent. In preferred embodiments, B is two substituents. Preferably, when B is two substituents, B is substituents at carbon positions 3 and 4, and more preferably, B is substituent-Cl at carbon positions 3 and 4. In preferred embodiments, A and B are each one substituent.

[0144] In the context of the present invention, when referring to the carbon positions of A and / or B with respect to structure (I), the numbering preferably refers to the following: [ka] Preferably, in synthesis pathways A to C, the positions of substituents A and B in each reactant are selected to coincide with the positions of substituents A and B in structure (I).

[0145] In preferred embodiments of the present invention, when referring to the chemical reaction steps of the present invention, each of them is: [ka] teeth, [ka] And, [ka] teeth, [ka] And, [ka] teeth, [ka] And, [ka] teeth, [ka] And, [ka] teeth, [ka] And, [ka] teeth, [ka] And, [ka] teeth, [ka] And, [ka] teeth, [ka] And, [ka] teeth, [ka] And, [ka] teeth, [ka] And, [ka] teeth, [ka] And, [ka] teeth, [ka] And, [ka] teeth, [ka] That is the case.

[0146] In a further special embodiment, the present invention relates to formula I: [ka] (In the formula, A is one or two substituents independently selected from -H, -F, -Cl, -Br, -I, -CF3, -OCF3, -CF2H, and -OCF2H. B is one or two substituents independently selected from -F, -Cl, -Br, -I, -CF3, -OCF3, -CF2H, and -OCF2H. X is selected from -S- and -O-, m is between 0 and 3. Y is -H; Nitrile group; A 5- to 6-membered unsaturated heterocycle containing three heteroatoms selected from O and N, preferably two or three of which are nitrogen, and the heterocycle is monosubstituted or disubstituted with =S or =O, preferably with =O; or, Linear-C 1~3 Alkyl, linear-C1~3 The alkyl group -(CH2)-, preferably one -(CH2)-, is optionally monosubstituted with -OH, and the linear-chain -C 1~3 The alkyl group's terminal -CH3 is -OR 1 Alternatively, -SR 1 And preferably -OR 1 It is monosubstituted with R 1 However, it is a straight chain -C which is -H, -(CH2)-COOH or -(CH2)-(CH2)-OH. 1~3 Alkyl; or, -(CH2) i -CX 1 R 2 (Here, X 1 is, =NR 3 Or = O, R 3 is -H or -OH, i is either 0 or 1. R 2 は-OR 4 or -NR 5 R 6 And R 4 is -H or -C 1~2 It is alkyl, R 5 and R 6 (The element is selected independently of -H or -methyl); Selected from, Here, it is referred to as "Z", [ka] is an aliphatic group comprising at least one, preferably one or two heterocycles, wherein the heterocycles together comprise at least two nitrogen atoms. Of the two nitrogen atoms, the first nitrogen atom is Z in formula I [ka] Linked to, the second of the two nitrogen atoms is Z in formula I [ka] Preferably, [ka] It is connected to the Z group, Each of the two nitrogen atoms is a heteroatom in one of the two linked four-membered heterocycles (each heterocycle has only one heteroatom), and the two linked four-membered heterocycles form a spirocycle, preferably the four-membered heterocycle having the first nitrogen atom is linked to the four-membered heterocycle having the second nitrogen atom via a shared carbon atom, thereby forming a spirocycle having two heteroatoms which are two nitrogen atoms, or The first of two nitrogen atoms is linked to a four-membered heterocycle having one heteroatom, the second of the two nitrogen atoms is the heteroatom in the four-membered heterocycle to which the first nitrogen atom is linked, and the first nitrogen atom may be further substituted with methyl, or The present invention relates to compounds having two nitrogen atoms that are heteroatoms in a 6- to 7-membered heterocycle having two heteroatoms, and the heterocycle is optionally substituted with methyl atoms, preferably monosubstituted, or to salts, complexes, diastereomers, enantiomers and / or tautomers of compounds having formula (I).

[0147] The present invention further relates to the following listed embodiments. These listed embodiments can be combined with any aspect, embodiment, and claim herein.

[0148] Item 1: A compound, or a salt, complex, diastereomer, enantiomer and / or tautomer thereof, wherein the compound is of formula: [ka] (In the formula, A is -H, B is one substituent selected from -F, -Cl, -Br, -I, -CF3, -OCF3, -CF2H, and -OCF2H. X is selected from -S- and -O-, m is between 0 and 3. Y is Nitrile group; A 5- to 6-membered unsaturated heterocycle containing three heteroatoms selected from O and N, preferably two or three of which are nitrogen, and the heterocycle is monosubstituted or disubstituted with =S or =O, preferably with =O; or, Linear-C 1~3 Alkyl, linear-C 1~3 The alkyl group -(CH2)-, preferably one -(CH2)-, is optionally monosubstituted with -OH, and the linear-chain -C 1~3 The alkyl group's terminal -CH3 is -OR 1 Alternatively, -SR 1 And preferably -OR 1 It is monosubstituted with R 1 However, it is a straight chain -C which is H, -(CH2)-COOH or -(CH2)-(CH2)-OH. 1~3 Alkyl; or, -(CH2) i -CX 1 R 2 (Here, X 1 is, =NR 3 Or = O, R 3 is -H or -OH, i is either 0 or 1. R 2 は-OR 4 or -NR 5 R 6 And R 4 is -H or -C 1~2 It is alkyl, R 5 and R 6 (The element is selected independently of -H or -methyl); Selected from, [ka] teeth, [ka] (is) has, The compound, [ka] A compound, or a salt, complex, diastereomer, enantiomer, and / or tautomer thereof, that is not any of the compounds mentioned above.

[0149] Item 2: (v) A is selected from -H, (vi) B is selected from -Cl or -CF3, (vii) m is 0 to 2, more preferably m is 0 or 1, and most preferably m is 1. (viii) Y is A compound of item 1, selected from the group consisting of TIFF2026509430000101.tif151170.

[0150] Item 3: The compound is Selected from the group TIFF2026509430000102.tif232170TIFF2026509430000103.tif237170TIFF2026509430000104.tif183170, Preferably, the compound is selected from the group consisting of compound 6, compound 8, compound 9, compound 10, compound 11, compound 12, compound 18, compound 19, compound 20, and compound 21, and more preferably, the compound is selected from the group consisting of compound 6, compound 8, compound 9, and compound 10, as described in item 1 or 2.

[0151] Item 4: The compound is [ka] A compound described in any one of items 1 to 3.

[0152] Item 5: The compound is a potassium channel in cells, preferably a potassium channel Slack (K Na A compound according to any one of items 1 to 4, having the activity of an activator as described in 1.1), preferably the compound activates potassium channels at an EC50 of less than 100 μM, preferably 0.5 μM to 80 μM.

[0153] Item 6: The compound is When administered to animals, it does not cross the blood-brain barrier (BBB), or crosses the BBB less effectively than reference compounds such as roxapine. A compound according to any one of items 1 to 5, which does not bind to human dopamine receptors, or more preferably has less binding to human dopamine receptors compared to a reference compound such as roxapine.

[0154] Item 7: A compound according to any one of items 1 to 6, wherein the compound has been isolated and / or the compound has a purity of at least 75%, optionally at least 90%, and optionally at least 95%.

[0155] Item 8: A pharmaceutical composition comprising a compound described in any one of items 1 to 7, or a salt, solvate, or ester thereof, and a pharmaceutically acceptable carrier or excipient.

[0156] Item 9: A pharmaceutical composition according to Item 8, wherein the compound, or its salt, solvate, or ester, is present in an amount of 0.1% w / w to 10% w / w, optionally 1% w / w to 10% w / w.

[0157] Item 10: A compound or composition for the treatment of a disease in a subject, wherein the compound or composition has the formula: [ka] (In the formula, A is -H, B is one substituent selected from -F, -Cl, -Br, -I, -CF3, -OCF3, -CF2H, and -OCF2H. X is selected from -S- and -O-, m is between 0 and 3. Y is Nitrile group; A 5- to 6-membered unsaturated heterocycle containing three heteroatoms selected from O and N, preferably two or three of which are nitrogen, and the heterocycle is monosubstituted or disubstituted with =S or =O, preferably with =O; or, Linear-C 1~3 Alkyl, linear-C 1~3 The alkyl group -(CH2)-, preferably one -(CH2)-, is optionally monosubstituted with -OH, and the linear-chain -C 1~3 The alkyl group's terminal -CH3 is -OR 1 Alternatively, -SR 1 And preferably -OR 1 It is monosubstituted with R 1 However, it is a straight chain -C which is H, -(CH2)-COOH or -(CH2)-(CH2)-OH. 1~3 Alkyl; or, -(CH2) i -CX 1 R 2 (Here, X 1 is, =NR 3 Or = O, R 3 is -H or -OH, i is either 0 or 1. R 2 は-OR 4 or -NR 5 R 6 And R 4 is -H or -C 1~2 It is alkyl, R 5 and R 6 (The element is selected independently of -H or -methyl); Selected from, [ka] teeth, [ka] A compound having ( ), or a salt, complex, diastereomer, enantiomer and / or tautomer thereof, preferably comprising an effective amount of a compound, or a salt, solvate or ester thereof as described in any one of items 1 to 6, preferably the composition is a pharmaceutical composition as described in item 7 or 8, wherein the disease is treatable by activating potassium channels in cells associated with the pathology of the disease, and the treatment comprises administering the compound or composition to a subject in need of treatment.

[0158] Item 11: The disease is a pruritic disorder such as acute or chronic itching, and optionally, the pruritic disorder is related to a secondary pathological condition such as one selected from the following: Dermatological disorders such as xerosis or xerosis (dry skin), dermatitis or eczema (e.g., atopic dermatitis), psoriasis (e.g., psoriasis vulgaris), prurigo (e.g., nodular prurigo), urticaria (e.g., chronic idiopathic urticaria), connective tissue disorders (e.g., dermatomyositis), and post-burn pruritus; Renal impairment (e.g., chronic kidney disease, chronic renal failure, or end-stage renal disease), dialysis (e.g., hemodialysis), uremic pruritus; Hepatobiliary disorders (e.g., cholestasis, primary biliary cholangitis, primary sclerosing cholangitis, secondary sclerosing cholangitis, hepatitis, toxic liver disease, chronic liver disease, or cirrhosis), cholestatic pruritus; Endocrine disorders (e.g., hyperthyroidism or diabetes mellitus); Metabolic disorders (e.g., iron deficiency or iron overload); Benign or malignant neoplasms (e.g., solid tumors, carcinomas, or hematological malignancies (e.g., Hodgkin lymphoma, non-Hodgkin lymphoma, myeloproliferative disorders, or polycythemia vera)); Infectious diseases (e.g., viral infections such as herpes simplex, herpes zoster, chickenpox, human immunodeficiency virus (HIV), and infections associated with hepatitis; bacterial infections or parasitic diseases); Neurological disorders (e.g., neurodegenerative diseases, multiple sclerosis, brain tumors, postherpetic neuralgia, small fiber neuropathy, brachioradialis pruritus, or dorsal paresthesia), neurogenic itching, neurogenic pruritus; Mental illness (e.g., depression, obsessive-compulsive disorder, delusional disorder, eating disorder, or anxiety disorder); Drug-induced pruritus (e.g., caused by opioids, antibiotics, antimalarial drugs, ACE inhibitors, angiotensin receptor blockers, antiarrhythmics, antidepressants, antidiabetic drugs, antihypertensive drugs, anticonvulsants, anti-inflammatory drugs, beta-blockers, bronchodilators, calcium channel blockers, diuretics, hormones, immunosuppressants, antilipids, nerve blockers, plasma expanders, tranquilizers, or uric acid-lowering drugs); Age-related pruritus; Pregnancy pruritus; and / or, Chronic idiopathic pruritus.

[0159] Item 12: The disease is pain or pain-related adverse sensation in the subject, preferably the pain is Neuropathic pain induced by traumatic nerve injury, cancer and cancer treatment (e.g., chemotherapy), neurological disorders (e.g., multiple sclerosis), neurodegenerative diseases (e.g., Parkinson's disease), trigeminal neuralgia, diabetic peripheral neuropathy, stroke, herpes zoster, HIV, leprosy, Guillain-Barré syndrome, vascular diseases, vascular malformations, and autoimmune diseases. acute postoperative pain, Inflammatory pain, rheumatoid arthritis, osteoarthritis, and / or Pain dysregulation, A compound or composition selected from item 10 or 11.

[0160] Item 13: A compound or composition according to any one of items 10 to 12, wherein the administration includes intravenous, intraperitoneal, subcutaneous, intramuscular, intrathecal, epidural, topical, oral, gastric and / or rectal administration.

[0161] Item 14: A compound or composition described in any one of items 10 to 13, wherein the compound is compound 6.

[0162] Item 15: A method for synthesizing any one of items 1 to 7, preferably the method comprising a synthesis step of synthesizing a lactam core, preferably the lactam core having the following structure: [ka] A method comprising (wherein A, X, and B are defined in any one of items 1 to 7).

[0163] As used herein, terms such as “(of this) invention,” “according to the present invention,” and “according to the present invention” are intended to refer to all aspects and embodiments of the invention described and / or claimed herein.

[0164] Where used herein and in embodiments individually disclosed in accordance with the present invention, the term “comprising” should be interpreted as encompassing both “including” and “consisting of,” as both meanings are specifically intended. Where used herein, “and / or” should be interpreted as each of two designated features or components, with or without other features or components. For example, “A and / or B” should be interpreted as each of (i) A, (ii) B, and (iii) A and B, each in exactly the same way as they are individually described herein. In the context of the present invention, the terms “about” and “approximately” indicate a range of accuracy that a person skilled in the art would understand to be guaranteed that the technical effect of the feature in question is still assured. The terms typically indicate deviations of ±20%, ±15%, ±10%, and, for example, ±5% from the given numerical value. As a person skilled in the art will understand, a particular such deviation from the numerical value for a given technical effect depends on the nature of the technical effect. For example, natural or biological technical effects may generally have larger deviations than artificial or engineered technical effects. As those skilled in the art will understand, a particular deviation of a numerical value for a given technical effect depends on the nature of the technical effect. For example, natural or biological technical effects may generally have larger deviations than artificial or engineered technical effects. When an indefinite or definite article is used when referring to a singular noun, for example, the singular form ("a", "an", or "the"), this includes the plural form of that noun unless otherwise specifically stated.

[0165] Applying the teachings of the present invention to specific problems or environments, and including variations or additional features of the present invention (such as further aspects and embodiments), should be understood to be within the capabilities of those skilled in the art in light of the teachings contained herein.

[0166] Unless otherwise indicated by the context, the above descriptions and definitions of features are not limited to any particular aspect or embodiment of the present invention, but apply equally to all aspects and embodiments described herein.

[0167] All references, patents, and publications cited herein constitute part of this specification by direct reference. [Brief explanation of the drawing]

[0168] [Figure 1] This figure shows the expression of Kcnt1 and important pruritus receptors across a subset of sensory neurons from publicly available single-cell RNA-seq data. (A) Expression pattern 4 in mouse DRG neurons (6-8 weeks old). (B) Expression pattern 16 in human DRG neurons (24-65 years old). Downloaded from https: / / sensoryomics.shinyapps.io / RNA-Data / . (C) Expression pattern 17 in non-human primate DRG neurons (5-14 years old). Downloaded from https: / / ernforsgroup.shinyapps.io / macaquedr. [Figure 2]This figure shows the establishment of a modified version of the FluxOR assay. Cultured HEK293 cells (HEK-Slack cells) stably expressing human Slack were incubated with the compounds of the present invention in different buffers. (A) In the buffer provided with the FluxOR assay kit, Slack activation (shown as an increase in the F / F (baseline) ratio) was detected after incubation with both roxapine and vehicle (FluxOR assay buffer containing 0.03% DMSO). Since Slack is activated by Na+, vehicle-induced Slack activation was most likely mediated by Na+ present in the FluxOR assay buffer and vehicle. (B) In a Na+-free buffer (NaCl replaced with choline chloride), Slack activation was observed after incubation with 30 μM roxapine and 140 mM NaCl, but not after incubation with vehicle (Na+-free buffer containing 2 mM Ca2+, 2 mM Mg2+, and 0.03% DMSO). (C) DMSO concentrations of 0.03% to 3% did not activate Slack in a Na+-free buffer containing 2 mM Ca2+ and 2 mM Mg2+. (D) A dose-response experiment using roxapine in a Na+-free buffer containing 2 mM Ca2+, 2 mM Mg2+, and 0.03% DMSO yielded an EC50 value of 23.45 μM in the initial preliminary experiment. (E) A dose-response experiment using Slack inhibitor compound 31 (Reference 18) pre-stimulated with 25 μM roxapine in a Na+-free buffer containing 2 mM Ca2+, 2 mM Mg2+, and 0.03% DMSO showed that compound 31 inhibited the F / F (baseline) ratio in a dose-dependent manner (IC50 = 2.1 μM), confirming that the readings were Slack-dependent. All further experiments using FluxOR, shown in Figures 3A and 3B, were performed using a Na+-free buffer containing 2 mM Ca2+, 2 mM Mg2+, and 0.03% DMSO. All conditions were measured at least three times, and data are shown as mean ± SD. [Figure 3]The figure shows that the novel compound is an activator of the Slack channel. (A) and (B) Dose-response experiments with the novel compound in the FluxOR potassium ion channel assay. HEK293 cells (HEK-Slack cells) that stably express human Slack were assayed for the thallium response via potassium channels using FluxOR. Each compound was incubated at six concentrations. For each sample, the fluorescence value was calculated and then normalized to the maximum fluorescence value of roxapine. For ease of comparison, data from roxapine measurements are shown in all five graphs. Each data point is the average of three replicates. Data represent mean ± sd. (C) Patch-clamp recordings confirm that the novel compound induces a potassium current via Slack. Whole-cell voltage recordings in HEK-Slack cells were performed at baseline and after incubation with the novel compound (50 μM), roxapine (lox; 50 μM), or vehicle (external solution containing 0.03% DMSO). Roxapine was incubated as a positive control in each series of experiments. The current densities (pA / pF) (fold increase) of the vehicle, roxapine, and novel compound relative to baseline at a voltage of +80mV are shown. Corresponding current-voltage curves and representative outward potassium traces from these records are shown in Figure 4. **P<0.01, ***P<0.001 (Kruskal-Wallis test with Dunn correction) for the vehicle; box plots represent maximum and minimum values, and boxes represent the first, second (median), and third quartiles. (D) Correlation between the relative EMAX values ​​of the novel compound obtained by the FluxOR assay (shown in (A) and (B)) and the relative current densities obtained by the patch-clamp experiment (shown in (C)). Pearson correlation. (E) Time-dependent fluorescence / fluorescence baseline ratios of the dose-response experiment shown in (B), compared to roxapine and vehicle. [Figure 4]This figure shows that the novel compound induces a potassium current via Slack. Whole-cell voltage recordings were performed in HEK-Slack cells at baseline and after incubation with the novel compound (50 μM), roxapine (50 μM), or vehicle (external solution containing 0.03% DMSO). (A) Current-voltage (IV) curves from patch-clamp experiments shown in Figure 3C. Roxapine was incubated as a positive control in each series of experiments. Cell size in each group n=8–20. Data are shown as mean ± SD. (B) Representative outward K+ (IK) trace of roxapine at +80 mV. (C) Current-voltage (IV) curves from patch-clamp experiments shown in Figure 3B. Roxapine was incubated as a positive control in each series of experiments. Cell size in each group n=8–20. [Figure 5] This figure shows the in vitro pharmacokinetic characterization of a novel compound. The blood-brain barrier (BBB) ​​permeability of the compound of the present invention was estimated using a BBB-specific parallel artificial membrane permeability assay. The predicted degree of BBB permeability is reflected by the log Pe value, and the fraction of solute lost to the membrane in this assay is reflected by the membrane retention coefficient (MR). Note that a high MR value limits the predictive validity of the corresponding log Pe value. [Figure 6] This figure shows the in vivo pharmacokinetic profiles of the novel compounds. (A) Time course of brain and plasma concentrations of compounds 1-10 in mice. Animals were administered 1 mg / kg of each compound intraperitoneally using a cassette administration procedure (simultaneous administration of 3-4 compounds to one animal), and plasma and brain levels were measured by LC-MS analysis at different time points. Note that the y-axis scale is different in all figures. Data are shown as mean ± sem for 3 mice in each group. (B) In vivo brain / plasma ratio of the novel compounds in mice. Note that compounds 6 and 10 have particularly low brain / plasma ratios, suggesting limited brain permeability. Data represent the mean. Additional pharmacokinetic parameters are shown in Table 6. [Figure 7]This figure shows the in vitro pharmacological screening of compound 6, compound 10, and roxapine. The compounds were tested at a concentration of 10 μM in binding, enzyme, and uptake assays of 44 targets (mostly human, with the exception of rat-derived BZD, NMDA, MAO-A, Ca2+ channels, KV channels, and Na+ channels). Compound binding was calculated as the percentage inhibition of binding to each target-specific radiolabeled ligand (agonist or antagonist, shown in parentheses). Compound enzyme inhibitory effect was calculated as the percentage inhibition of control enzyme activity. Results showing inhibition greater than 50% (or stimulation in assays performed under baseline conditions) are considered to represent a significant effect of the test compound and are shown in black. Results showing inhibition or stimulation between 25% and 50% (showing weak to moderate effects) and results less than 25% (mostly considered to be due to signal fluctuations around the control level) are shown in gray. Measurements were performed twice. [Figure 8]This figure shows that compounds 6 and 10 inhibit histamine-independent acute pruritic behavior and neuropathic pain behavior. (A), (B) Motor function. Compounds 6, 10, roxapine, or a vehicle (0.9% NaCl containing 10% 2-hydroxypropyl-β-cyclodextrin) were administered intraperitoneally, and (A) an accelerated rotarod test was performed, followed by (B) a vertical pole test 15 minutes later. Note that compounds 6 and 10 did not affect the time spent on the rotarod or vertical pole, i.e., they did not inhibit motor coordination, but roxapine inhibited motor coordination in a dose-dependent manner in both models. Each group n=8. Box plots show maximum and minimum values, and boxes show the first quartile, second quartile (median), and third quartile. Dotted lines show cutoff times. *p<0.05, ***p<0.001, Kruskal-Wallis test. (C)~(G) Acute pruritic behavior. Compound 6, compound 10, or vehicle was administered intraperitoneally, and 15 minutes later, a different pruritic substance was administered subcutaneously to the nape of the neck. The number of scratching behaviors over a 30-minute period was counted. In each panel, the time course of scratching behavior is shown on the left, and the total number of scratching behaviors over 30 minutes is shown on the right. (C) Compounds 6 and 10 dose-dependently inhibited chloroquine-induced scratching behavior. (D) Compound 6 inhibited chloroquine-induced scratching behavior in wild-type mice, but not in Slack- / - littermates. (E) Compound 6 alleviated SLIGRL-induced scratching behavior, but (F) histamine-induced scratching behavior. Each group n=7~8. Data represent mean ± sem. *p<0.05, **p<0.01, ***p<0.001; One-way ANOVA with Dunnett correction (C), Kruskal-Wallis test with Dunn correction (D), or unpaired t-test (E, F). (G) Neuropathic pain behavior. Neuropathic pain was surgically induced in a partial nerve injury (SNI) model. Twenty-eight days later, mechanical hypersensitivity of the affected hind paw (determined using a dynamic plantar aesthesiometer and indicated by a decrease in paw retraction latency) was detected in all mice.Next, compound 10 or vehicle was administered intraperitoneally, and mechanical sensitivity was evaluated over 3 hours. Note that compound 10 inhibited neuropathic pain behavior. (H) Effect of compound 6 in wild-type mice pretreated with a Slack inhibitor. Compound 31 (30 mg / kg) or vehicle was administered intraperitoneally 5 minutes before compound 6 (30 mg / kg intraperitoneally), and chloroquine was administered subcutaneously 15 minutes later. The data show that the antipruritic effect of compound 6 was partially antagonized by the Slack inhibitor. Each group n=7-8. Data are shown as mean ± SEM. *p<0.05, unpaired t-test. [Figure 9] This figure shows that the effects of compound 6 in motor function and acute pruritus models did not differ between sexes. (A) Figure 8A, (B) Figure 8B, (C) Figure 8C, and (D) Figure 8F show the breakdown of results for compound 6 in male and female mice. Statistical significance in a and b was assessed by the Kruskal-Wallis test. Box plots represent maximum and minimum values, and boxes represent the first quartile, second quartile (median), and third quartile. The dotted line indicates the cutoff time. Statistical significance in c and f was assessed by one-way ANOVA with Dunnett correction. Data represent mean ± sem. [Figure 10]This figure shows that compound 6 inhibits persistent itching behavior. (A)-(C) Efficacy of compound 6 in a DNFB model of persistent itching. (A) Experimental figure showing induction of spontaneous itching behavior by topical application of 2,4-dinitrofluorobenzene (DNFB) to the nape of the neck (twice, 14 days apart), intraperitoneal drug delivery 105 minutes after the second DNFB application, and the start of video recording 15 minutes thereafter. (B) Compound 6 significantly reduced the number of scratching behaviors compared to the vehicle, and (C) Compound 6 significantly reduced the number of head shaking behaviors. Each group n=6-7. (D)-(F) Efficacy of compound 6 in an MC903 model of persistent itching. (D) Experimental figure showing induction of spontaneous scratching by topical application of MC903 to the nape of the neck (once daily for 7 days), intraperitoneal drug delivery on day 8, and the start of video recording 15 minutes thereafter. Compared to the vehicle, compound 6 significantly reduced (E) the number of scratching behaviors and (F) the number of head shaking behaviors. Each group n=8. Data represent mean ± sem. *p<0.05, **p<0.01, unpaired t-test. [Figure 11] This figure shows the sex-related effects of compound 6 and the correlation of behavioral outcomes in chronic pruritus models. (A), (B) The effect of compound 6 in chronic pruritus models does not show a difference based on sex. (A) Breakdown of results in male and female mice from Figures 10B and 10C (DNFB model) and (B) Figures 10E and 10F (MC903 model). Data represent mean ± sem. (C), (D) Correlation between the number of head shakes and the number of scratching behaviors from (C) Figures 10B and 10C, (D) Figures 10E and 10F, and (E) combinations thereof. Statistical significance was assessed by Pearson correlation. [Figure 12] This figure shows the potential side effects of compound 6. Pulse oximetry in unanesthetized mice using the MouseOX Plus instrument revealed that intraperitoneal delivery of compound 6 or vehicle did not alter (A) heart rate or (B) respiratory rate, while morphine significantly reduced both parameters (n=6 in each group). *p<0.05, **p<0.01, ***p<0.001 for vehicle, two-way MC ANOVA and Dunnett test. [Figure 13] This figure shows that compound 6 reduces the neuronal excitability of itch-sensitive sensory neurons. Cultured DRG neurons from mice were incubated overnight with inflammatory soup, followed by whole-cell current-clamp recordings of IB4-connected neurons. (A) Recordings from DRG neurons showing action potential (AP) firing in response to current injection (200 pA to 950 pA at 150 pA intervals for a duration of 1000 milliseconds) at baseline, in the presence of compound 6 (50 μM), and after washing. (B) Group data show that compound 6 significantly blocked AP firing compared to baseline. n=6 neurons. (C) Recordings from DRG neurons showing single APs induced by injection of small amounts of current (0 pA to 220 pA at 20 pA intervals for a duration of 10 milliseconds) at baseline and in the presence of compound 6 (50 μM). (D), (E) Group data show that compound 6 significantly increased (D) the rheobase (the amount of current required to generate an AP) and significantly decreased (E) the AP amplitude. n=7 neurons. Data represent mean ± sem. *p<0.05, **p<0.01, ***p<0.001, paired t-test. [Figure 14] This figure shows the antinociceptive effect of compound 6 in a CFA-induced inflammatory pain model in mice. Mechanical sensitivity of the hind paw was determined using a dynamic planter estesiometer. 24 hours after CFA injection into the hind paw, mice were intraperitoneally treated with either compound 6 (10 mg / kg) or a vehicle (10% cyclodextrin in 0.9% NaCl). Note that the paw retraction latency significantly increased 0.5 and 1 hour after administration of compound 6 (n=11-12 mice in each group). Results are shown as mean ± SEM. Data were analyzed by two-way repeated measures ANOVA with Sidac's post-hoc test. *P<0.05, **P<0.01. [Figure 15]This figure shows that topically administered Slack activators inhibit histamine-independent pruritic behavior. Compound 6, compound 21, or vehicle was topically applied to the nape of the neck. Thirty minutes later, chloroquine was subcutaneously administered to the nape of the neck, and the number of scratching behaviors over the 30 minutes was counted. The time course of scratching behavior is shown on the left, and the total number of scratching behaviors over the 30 minutes is shown on the right. Note that both compound 6 and compound 21 significantly inhibited chloroquine-induced scratching behavior. Data represent mean ± SEM (n=6). *P<0.05, **P<0.01. [Examples]

[0169] Certain aspects and embodiments of the present invention are described herein by reference with reference to the description, drawings, and tables provided herein. Such examples of methods, uses, and other aspects of the present invention are representative examples and should not be construed as limiting the scope of the present invention to such representative examples only.

[0170] Examples are shown below.

[0171] Example 1: Synthesis of Slack-activating compound Based on the antipsychotic drug roxapine, we developed a series of novel Slack-activating compounds. These novel compounds were synthesized under a linear synthetic pathway A to C in an argon atmosphere, as described below.

[0172] Synthetic Route A: The roxapine skeleton contains a tricyclic core and a piperazine ring substituted with alkyl residues. Starting from methyl esters of salicylic acid and 2-fluoronitrobenzene derivatives, diaryl ethers were obtained by nucleophilic aromatic substitution reactions. Subsequent reduction of the nitro group with tin(II) chloride enabled the formation of an intramolecular amide bond.

[0173] Synthesis Path A [ka]

[0174] Synthetic route B: Alternatively, the same lactam core was obtained by first forming an amide bond between 2-fluorobenzoic acid and a 2-aminophenol derivative, followed by intramolecular nucleophilic aromatic substitution.

[0175] Synthesis Path B [ka]

[0176] Synthetic pathway C: The lactam was then converted to imidoyl chloride by POCl3. In the final step, a substituted piperazine substituted the chloride to obtain the desired roxapine derivative.

[0177] Synthesis route C [ka] p-xylene

[0178] Synthetic Route A - General synthesis of lactams starting from methyl salicylate and 2-fluoronitrobenzene derivatives GP1 - Diaryl ether formation: Potassium carbonate (1.5 equivalents) was added to a solution of the corresponding methyl salicylate (1.5 equivalents) and the corresponding 2-fluoronitrobenzene (1.0 equivalent) in DMF (1.5 M with respect to 2-fluoronitrobenzene). The resulting solution was heated overnight in an oil bath at 120°C. The solvent was evaporated under reduced pressure, and the residue was transferred to water and extracted three times with ethyl acetate. The combined organic phase was dried over magnesium sulfate, filtered, and evaporated. The resulting crude product was purified by flash chromatography.

[0179] GP2 - Nitro group reduction: A solution of the corresponding methyl 2-(2-nitrophenoxy)benzoate derivative (1.0 equivalent) in a 1:1 (0.5 M) mixture of ethanol / concentrated HCl was mixed with a solution of SnCl2·2H2O (4.0 equivalents) in concentrated HCl (3 M). The resulting solution was stirred overnight at room temperature. After that time, the temperature was set to 0°C and the pH of the reaction solution was made slightly basic by the addition of sodium carbonate. The resulting solution was then extracted three times with ethyl acetate. The combined organic phase was dried over magnesium sulfate, filtered, and evaporated. The resulting crude product was purified by flash chromatography.

[0180] GP3 - Formation of lactam rings by intramolecular condensation: A solution of the corresponding methyl 2-(2-aminophenoxy)benzoate derivative (1.0 equivalent) in GP3I:DMF (0.2 M) was treated with concentrated sulfuric acid (1.3 equivalents) and heated overnight in an oil bath at 120°C. After that time, the reaction mixture was cooled to 0°C and several milliliters of water were added. The precipitated product was then filtered and dried under vacuum to obtain the crude product, which was used in the next step without further purification.

[0181] Synthetic Route B: General synthesis of lactams starting from 2-fluorobenzoic acid and 2-aminophenol derivatives Formation of GP4 amide: Freshly distilled thionyl chloride (2.0 equivalents) was added to a solution of the corresponding 2-fluorobenzoic acid (1.0 equivalent) in THF (1M), and the mixture was heated under reflux in an oil bath for 2 hours. Then, excess thionyl chloride and THF were removed under reduced pressure, and the resulting residue was dissolved again in THF (2.5M). This solution was added dropwise to a solution of the corresponding 2-aminophenol derivative (1.0 equivalent) and triethylamine (2 equivalents) in THF (2.5M for the 2-aminophenol derivative) at 0°C, and the reaction mixture was stirred overnight at room temperature. After that time, the reaction mixture was concentrated under reduced pressure, and the residue was transferred to ethyl acetate. The organic phase was first washed with aqueous HCl (2M), water, and saturated aqueous NaCl, then dried over magnesium sulfate, filtered, and evaporated. The resulting crude product was purified by flash chromatography.

[0182] GP5 - Formation of a lactam ring by intramolecular nucleophilic aromatic substitution: A solution of the corresponding 2-fluoro-N-(2-hydroxyphenyl)benzamide derivative (1.0 equivalent) in DMF (0.25 M) was treated with freshly powdered NaOH (1.0 equivalent) and heated in an oil bath at 150°C for 5 hours. After that time, the reaction mixture was cooled to 0°C and several milliliters of water were added. The precipitated product was then filtered and dried under vacuum to obtain the crude product, which was used in the next step without further purification.

[0183] Synthetic Pathway C - General Synthesis of Roxapine Derivatives Starting from Lactams GP6 - Reaction with phosphorus oxychloride: The corresponding lactam (1.0 equivalent) was dissolved in freshly distilled phosphorus oxychloride (0.5 M), and N,N-dimethylaniline (0.6 equivalents) was added. The resulting mixture was heated under reflux in an oil bath for 5 hours. Then, excess phosphorus oxychloride was removed under reduced pressure, and the resulting residue was placed in toluene and washed once with cold water. The organic phase was dried over magnesium sulfate, filtered, and evaporated to obtain the crude product, which was immediately used in the next step without further purification.

[0184] GP7 - Formation of roxapine derivatives: To a solution of a suitable imidoyl chloride (1.0 equivalent) in p-xylene (0.15 M with respect to imidoyl chloride), the corresponding amine (2.0 equivalents) was added. The resulting mixture was heated in an oil bath at 140°C for 5 hours. The solvent was evaporated, and the crude product was purified by preparative HPLC or flash chromatography. When unsubstituted piperazine or homopiperazine was used, the alkylation of these derivatives was obtained by GP8.

[0185] GP8 - Alkylation of roxapine derivatives: A mixture of roxapine derivative (1 equivalent), corresponding alkyl chloride (2 equivalents), and triethylamine (10 equivalents) in acetonitrile (0.15 M) was heated under reflux in an oil bath for 16 hours. After completion, the reaction mixture was evaporated, and the residue was purified by preparative HPLC or flash chromatography.

[0186] Example 2: Characterization of the synthesized compound The compounds were synthesized using the above synthetic route. To characterize these compounds, NMR spectra were recorded using a Bruker Avance DPX250, Bruker Avance 300, Bruker Avance 400, or Bruker Avance 500, all operated at ambient temperature by Bruker (Karlsruhe, Germany). Proton spectra were recorded using CDCl3 or DMSO-d6. 1 The 1H NMR chemical shifts were used as a reference for the residual signals of CHCl3 (δ=7.26 ppm) and DMSO-d5 (δ=2.50 ppm). 13The ¹³C NMR chemical shift was referenced to the centerline of the solvent signal (δ=77.16 ppm for CDCl3, δ=39.52 ppm for DMSO-d6). Chemical shifts are given on a δ scale (ppm). Coupling constants (J) are given in Hz. Multiplicity is indicated as follows: br (broad signal), s (singular), d (double), t (tripular), q (quadular), quint (quinular), or m (multiple). ESI-MS was measured on a Shimadzu LCMS-2020, and HRMS was measured on a Thermo Scientific MALDI Orbitrap XL. TLC was performed on Marcherey-Nagel (ALUGRAM®) silica gel plates and visualized with a UV lamp (254 nm and / or 366 nm). All product purification was performed by flash chromatography using a puriFlash XS420 from Interchim (Montluçon, France) and a Silica HP 30 μm column as the stationary phase. Analysis and half-section HPLC were performed using a Shimadzu prominence system equipped with a Shimadzu SPD20A UV / Vis detector from Shimadzu (Duisburg, Germany). The stationary phases were Luna 10 μm 100 Å, C18(2) (250 mm × 4.6 mm) and Luna 10 μm 100 Å, C18(2) (250 mm × 21.20 mm) from Phenomenex (Aschaffenburg, Germany), and the eluent was a mixture of ACN and formic acid aqueous solution (0.1%). Flow rates were set to 1 mL / min and 21 mL / min. All tested compounds showed a purity of over 95% (254 nm).

[0187] 11-(4-methylpiperazin-1-yl)-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine (compound 2) [ka] The compound was synthesized from 11-chloro-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine (63 mg, 0.210 mmol) and 1-methylpiperazine (46.6 μL, 0.420 mmol) according to GP7. Purification of the crude product by preparative HPLC yielded the title compound as a brown oily substance and formate (22 mg, 26%). 1 H NMR (300 MHz, CDCl3) δ 8.35 (s, 1H), 7.95 (br s, 1H), 7.70 (ddd, J = 8.5, 2.3, 0.5 Hz, 1H), 7.62 (d, J = 2.3 Hz, 1H), 7.36 (d, J = 8.5 Hz, 1H), 7.18-7.07 (m, 3H), 7.01 (td, J = 7.4, 2.0 Hz, 1H), 3.62 (br s, 4H), 2.69 (br s, 4H), 2.44 (s, 3H); 13C[1H] NMR (75 MHz, CDCl3) δ 166.6, 163.2, 158.7, 151.4, 139.9, 129.7, 127.5, 127.2, 127.0, 126.0, 124.9, 124.1, 123.5, 122.1, 120.3, 54.0, 46.7, 45.3;HPLC purity (254nm):96%;MALDI-HRMS:m / z C 19 H 19 The calculated value of F3N3O[M+H]+ is 362.1474, and the measured value is 362.1488.

[0188] 11-(4-methyl-1,4-diazepan-1-yl)-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine (compound 3) [ka] The compound was synthesized from 11-chloro-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine (63 mg, 0.212 mmol) and 1-methylhomopiperazine (49 mg, 0.423 mmol) according to GP7. Purification of the crude product by preparative HPLC yielded the title compound as a brown oily substance and formate (73 mg, 92%).1 H NMR (300 MHz, CDCl3) δ 9.97 (br s, 1H), 8.51 (s, 1H), 7.68 (dd, J = 8.5, 1.9 Hz, 1H), 7.58 (d, J = 2.1 Hz, 1H), 7.35 (d, J = 8.5 Hz, 1H), 7.12-7.06 (m, 3H), 6.98-6.95 (m, 1H), 4.06-3.45 (m, 4H), 3.08-2.84 (m, 4H), 2.56 (s, 3H), 2.29 (br s, 1H), 2.01 (br s, 1H); 13C[1H] NMR (75 MHz, CDCl3) δ 168.2, 162,8, 158.3, 151.2, 140.4, 129.4, 127,4, 127.0, 126.9, 126.2, 124,2, 124.0, 123.6, 122,2, 120,2, 56.9, 56.3, 49.1, 46.9, 45.4, 26.3;HPLC purity (254nm):97%;MALDI-HRMS:m / z C 20 H 21 The calculated value of F3N3O[M+H]+ is 376.1631, and the measured value is 376.1639.

[0189] 8-Fluoro-11-(4-methylpiperazin-1-yl)-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine (compound 4) [ka] The compound was synthesized from 11-chloro-8-fluoro-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine (50 mg, 0.158 mmol) and 1-methylpiperazine (32 mg, 0.317 mmol) according to GP7. Purification of the crude product by preparative HPLC yielded the title compound as a yellow solid (25 mg, 42%). 1H NMR (300 MHz, CDCl3) δ 7.68 (dd, J = 8.5, 1.9 Hz, 1H), 7.58 (d, J = 2.1 Hz, 1H), 7.35 (d, J = 8.5 Hz, 1H), 7.12-7.06 (m, 3H), 6.98-6.95 13C[1H] NMR (75 MHz, CDCl3) δ 163.2, 161.5, 159.6, 159.4, 147.7, 141.6, 128.4, 127.7, 123.7, 124.1, 122.2, 120.1, 113.4, 110.9, 54.7, 47.3, 46.04; HPLC purity (254 nm): >99%; MALDI-HRMS: m / z C 19 H 18 The calculated value of F4N3O[M+H]+ is 380.1381, and the measured value is 380.1377.

[0190] 2-(4-(2-chlorodibenzo[b,f][1,4]oxazepine-11-yl)piperazine-1-yl)ethane-1-ol (compound 5) [ka] The compound was synthesized from 2,11-dichlorodibenzo[b,f][1,4]oxazepane (198 mg, 0.750 mmol) and hydroxyethylpiperazine (195 mg, 1.50 mmol) according to GP7. Purification of the crude product by preparative HPLC yielded the title compound as a brown oily substance and formate (216 mg, 72%). 1H NMR (300 MHz, CDCl3) δ 8.40 (s, 1H), 7.41 (dd, J = 8.6, 2.6 Hz, 1H), 7.31 (d, J = 2.6 Hz, 1H), 7.19 (d, J = 8.6 Hz, 1H), 7.16-7.07 (m, 13C[1H] NMR (75 MHz, CDCl3) δ 166.8, 159.3, 158.3, 151.7, 139.6, 133.0, 130.5, 128.8, 127.1, 125.9, 125.1, 124.5, 122.9, 120.2, 59.7, 56.9, 52.2, 45.8;HPLC purity (254nm):99%;MALDI-HRMS:m / z C 19 H 21 Calculated value of ClN3O2[M+H]+: 358.1317, measured value: 358.1324.

[0191] 2-(2-(4-(2-chlorodibenzo[b,f][1,4]oxazepine-11-yl)piperazine-1-yl)ethoxy)acetic acid (compound 6) [ka] The compound was synthesized according to GP8 from commercially available amoxapine (75 mg, 0.239 mmol), 2-(2-chloroethoxy)acetic acid (66 mg, 0.478 mmol), and triethylamine (0.34 mL, 2.39 mmol). Purification of the crude product by preparative HPLC yielded the title compound as a brown oily substance and formate (47 mg, 42%). 1H NMR (300 MHz, CDCl3) δ 8.24 (br s, 1H), 7.41 (dd, J = 8.7, 2.4 Hz, 1H), 7.34 (d, J = 2.4 Hz, 1H), 7.18 (d, J = 8.7 Hz, 1H), 7.15-6.98 (m, 13C[1H] NMR (75 MHz, CDCl3) δ 168.8, 164.7, 159.3, 159.0, 151.6, 139.1, 133.4, 130.8, 128.7, 127.2, 126.0, 125.7, 123.9, 123.0, 120.3, 61.4, 59.5, 55.8, 41.4;HPLC purity (254nm):99%;MALDI-HRMS:m / z C 21 H 23 Calculated value of ClN3O4[M+H]+: 416.1372, measured value: 416.1375.

[0192] 2-(2-(4-(2-chlorodibenzo[b,f][1,4]thiazepine-11-yl)piperazine-1-yl)ethoxy)ethane-1-ol (compound 7) [ka] The compound was synthesized from 2,11-dichlorodibenzo[b,f][1,4]thiazepine (119 mg, 0.425 mmol) and 1-[2-(2-hydroxyethoxy)ethyl]piperazine (147 μL, 0.850 mmol) according to GP7. Purification of the crude product by preparative HPLC yielded the title compound as a brown oily substance and formate (156 mg, 79%). 1H NMR (300 MHz, CDCl3) δ 8.37 (s, 1H), 7.46-7.41 (m, 1H), 7.38 (dd, J = 7.7, 1.5 Hz, 1H), 7.33-7.27 (m, 2H), 7.29 (br s, 1H), 7.23-7.17 (m, 1H), 7.06 (dd, J = 8.0, 1.5 Hz, 1H), 6.93 (td, J = 7.3, 1.5 Hz, 1H), 3.88 (br s, 2H), 3.75 (t, J = 5.1 Hz, 2H), 3.70 (t, J = 4.8 Hz, 2H), 3.58 (t, J = 4.8 Hz, 13C[1H] NMR (75 MHz, CDCl3) δ 166.7, 158.9, 148.2, 138.1, 134.9, 134.8, 133.4, 132.9, 131.2, 129.4, 128.6, 127.3, 125.3, 123.6, 72.6, 66.1, 61.4, 57.2, 52.0, 45.1;HPLC purity (254nm):95%;MALDI-HRMS:m / z C 21 H 25 Calculated value of ClN3O2S[M+H]+: 418.1351, measured value: 418.1350.

[0193] 2-(2-(4-(2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine-11-yl)piperazine-1-yl)ethoxy)acetic acid (compound 8) [ka] The compound was synthesized according to GP8 from 11-(piperazin-1-yl)-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine (75 mg, 0.102 mmol), 2-(2-chloroethoxy)acetic acid (28 mg, 0.205 mmol), and triethylamine (0.14 mL, 1.02 mmol). Purification of the crude product by flash chromatography (DCM / MeOH 95:5 → 9:1) yielded the title compound as a colorless solid (28 mg, 61%). 1 H NMR (400 MHz, CDCl3) δ 7.66 (dd, J = 8.5, 2.0 Hz, 1H), 7.56-7.56 (m, 1H), 7.31 (d, J = 8.5 Hz, 1H), 7.12-7.03 (m, 3H), 6.97 (td, J = 7.6, 1.8 Hz, 1H), 4.21 (s, 2H), 3.73-3.43 (m, 12H); 13C[1H] NMR (101 MHz, CDCl3) δ 168.7, 163.4, 158.8, 151.5, 139.7, 128.5, 127.8, 127.4, 126.3, 125.4, 124.1, 123.6, 122.5, 120.5, 73.8, 69.7, 61.8, 44.3, 41.7;HPLC purity (254nm):98%;MALDI-HRMS:m / z C 22 H 23 The calculated value of F3N3O4[M+H]+ is 450.1635, and the measured value is 450.1637.

[0194] 2-(2-(4-(2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine-11-yl)-1,4-diazepan-1-yl)ethoxy)acetic acid (compound 9) [ka] The compound was synthesized according to GP8 from 11-(4-methyl-1,4-diazepan-1-yl)-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine (40 mg, 0.111 mmol), 2-(2-chloroethoxy)acetic acid (31 mg, 0.221 mmol), and triethylamine (0.16 mL, 1.11 mmol). Purification of the crude product by flash chromatography (DCM / MeOH 95:5 → 9:1) yielded the title compound as a colorless solid (20 mg, 39%). 1 H NMR (400 MHz, CDCl3) δ 7.95-7.91 (m, 1H), 7.80-7.77 (m, 1H), 7.59 (t, J = 8.8 Hz, 1H), 7.22-7.18 (m, 1H), 7.11-6.93 (m, 3H), 4.63 13C[1H] NMR (101 MHz, CDCl3) δ 168.6, 162.4, 157.6, 150.7, 140.3, 129.9, 126.6, 126.3, 126.0, 126.0, 123.7, 123.6, 123.7, 122.4, 120.2, 72.5, 69.1, 68.9, 68.2, 67.6, 63.2, 60.0, 44.0;HPLC purity (254nm):95%;MALDI-HRMS:m / z C 23 H 25 The calculated value of F3N3O4[M+H]+ is 464.1792, and the measured value is 464.1788.

[0195] 5-((4-(2-chlorodibenzo[b,f][1,4]oxazepine-11-yl)piperazine-1-yl)methyl)-1,2-dihydro-3H-1,2,4-triazole-3-one (compound 10) [ka] The compound was synthesized according to GP8 from commercially available amoxapine (108 mg, 0.334 mmol), 3-(chloromethyl)-1H-1,2,4-triazole-5(4H)-one (66 mg, 0.478 mmol), and triethylamine (0.47 mL, 3.34 mmol). Purification of the crude product by flash chromatography (Â15 / MeOH 9:1) yielded the title compound as a colorless solid (136 mg, 99%). 1 H NMR (400 MHz, DMSO-d6) δ 11.36 (br s, 1H), 11.26 (br s, 1H), 7.61 (dd, J = 8.7, 2.6 Hz, 1H), 7.42-7.39 (m, 2H), 7.18 (dd, J = 7.9, 1.2 13C[1H] NMR (101 MHz, CDCl3) δ 158.7, 158.0, 156.2, 151.2, 144.3, 139.9, 133.0, 129.5, 128.7, 126.5, 125.8, 124.4, 124.2, 123.1, 120.2, 53.2, 52.0, 48.6;HPLC purity (254nm):99%;MALDI-HRMS:m / z C 20 H 20 Calculated value of ClN6O2[M+H]+: 411.1331, measured value: 411.1327.

[0196] 5-((4-(2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine-11-yl)piperazine-1-yl)methyl)-1,2-dihydro-3H-1,2,4-triazole-3-one (compound 11) [ka] Compound 13 (20.0 mg, 0.0576 mmol), 3-(chloromethyl)-1H-1,2,4-triazole-5(4H)-one (15.9 mg, 0.115 mmol), and triethylamine (80.3 μL, 0.576 mmol) were synthesized according to GP8. The title compound was obtained by purification of the crude product by flash chromatography (DCM / MeOH 95:5 → 8:2) (24.0 mg, 94%). 1 H NMR (500 MHz, DMSO-d6) δ 11.37 (s, 1H), 11.26 (s, 1H), 7.95 (dd, J = 8.7, 2.1 Hz, 1H), 7.70 (d, J = 2.2 Hz, 1H), 7.60 (d, J = 8.5 Hz, 1H), 7.23 (dd, J = 7.9, 1.3 Hz, 1H), 7.13-7.06 (m, 2H), 7.04-7.00 (m, 1H), 3.46 (br s, 4H), 3.35 (s, 2H), 2.53 (br s, 4H). 13 C{ 1 H} (126 MHz, DMSO-d6) δ 162.6, 158.1, 156.2, 150.8, 144.3, 139.9, 130.3, 126.8, 126.6, 126.2, 126.1, 123.6, 124.4, 123.6, 122.6, 120.4, 53.3, 52.0, 46.7;HPLC purity (254nm):99%;MALDI-HRMS:m / z C 21 H 20 Calculated value of F3N6O2[M+H]+: 445.1594, measured value: 445.1592.

[0197] 2-(2-(4-(2-chlorodibenzo[b,f][1,4]thiazepine-11-yl)piperazine-1-yl)ethoxy)acetic acid (compound 12) [ka] The compound was synthesized according to GP8 from 2-chloro-11-(piperazin-1-yl)dibenzo[b,f][1,4]thiazepine (45 mg, 0.136 mmol), 2-(2-chloroethoxy)acetic acid (37.7 mg, 0.272 mmol), and triethylamine (191 μL, 1.36 mmol). The title compound was obtained by purification of the crude product by preparative HPLC (19.6 mg, 33%). 1 H NMR (300 MHz, CDCl3) δ 7.46 (d, J = 8.3 Hz, 1H), 7.39 (dd, J = 7.7, 1.3 Hz, 1H), 7.35-7.31 (m, 2H), 7.24-7.18 (m, 1H), 7.07 (dd, J = 7.9, 1.2 Hz, 1H), 6.94 (td, J = 7.6, 1.4 Hz, 1H), 4.25 (s, 2H), 3.84-3.36 (m, 12H). 13 C{ 1 H} NMR (75 MHz, CDCl3) δ 168.5, 159.2, 148.1, 138.3, 135.1, 134.8, 133.5, 132.2, 131.3, 129.4, 128.5, 127.4, 125.3, 123.7, 73.6, 70.0, 61.7, 44.1, 41.5;HPLC purity (254nm):99%;MALDI-HRMS:m / z C 21 H 23 ClN3O3S[M+H] + Calculated value: 432.1143, measured value: 432.1148.

[0198] 11-(piperazin-1-yl)-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepane (compound 13) [ka] The compound was synthesized from 11-chloro-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine (267 mg, 0.897 mmol) and piperazine (155 mg, 1.79 mmol) according to GP7. Purification of the crude product by flash chromatography (DCM / MeOH 95:5 → 9:1) yielded the title compound as a yellow solid (240 mg, 77%). 1 H NMR (400 MHz, CDCl3) δ 8.77 (s, 2H), 8.47 (br s, 1H), 7.73 (d, J = 8.0 Hz, 1H), 7.64-7.60 (m, 1H), 7.37 (d, J = 8.0 Hz, 1H), 7.18-7.03 (m, 4H), 3.77-3.23 (m, 8H).

[0199] 11-(1,4-diazepan-1-yl)-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine (compound 14) [ka] The compound was synthesized from 11-chloro-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine (267 mg, 0.897 mmol) and homopiperazine (183 mg, 1.79 mmol) according to GP7. Purification of the crude product by flash chromatography (DCM / MeOH 95:5 → 9:1) yielded the title compound as a yellow solid (178 mg, 55%). 1 H NMR (400 MHz, CDCl3) δ 7.69 (dd, J = 8.6, 2.0, 1H), 7.63 (d, J = 1.9 Hz, 1H), 7.36 (d, J = 8.5 Hz, 1H), 7.13-7.06 (m, 3H), 7.99-6.95 (m, 1H), 3.98-3.13 (m, 9H), 2.14-1.90 (m, 2H).

[0200] 3-((4-(2-chlorodibenzo[b,f][1,4]oxazepine-11-yl)piperazine-1-yl)methyl)-1,2,4-oxadiazole-5(4H)-one (compound 15) [ka] To a solution of compound 17 (22.0 mg, 0.0570 mmol) in EtOH (2.0 mL), sodium methoxide solution in MeOH (5 M, 39.1 μL, 0.171 mmol) and diethyl carbonate (27.9 μL, 0.228 mmol) were added at room temperature, and the reaction mixture was heated under reflux overnight. The crude product was purified by preparative HPLC to obtain the title compound (11.0 mg, 47%). 1 H NMR (400 MHz, DMSO-d6) δ 7.62 (dd, J = 8.7, 2.6 Hz, 1H), 7.43 (d, J = 2.4 Hz, 1H), 7.40 (d, J = 9.6 Hz, 1H), 7.20-7.17 (m, 1H), 7.12-6.98 (m, 3H), 3.53-3.48 (m, 6H), 2.60 (br s, 4H). 13 C{ 1 H} (101 MHz, DMSO-d6) δ 159.9, 158.7, 158.0., 157.4, 151.2, 139.9, 133.0, 129.5, 128.7, 126.5, 125.8, 124.4, 124.3, 123.1, 120.2, 51.9, 51.2, 46.6. MALDI-HRMS:m / z C 20 H 19 Calculated value of ClN5O3[M+H]+: 412.1171, measured value: 412.1171.

[0201] 2-(4-(2-chlorodibenzo[b,f][1,4]oxazepine-11-yl)piperazine-1-yl)acetonitrile (compound 16) [ka] The compound was synthesized from amoxapine (150 mg, 0.464 mmol), bromoacetonitrile (66.6 μL, 0.927 mmol), and triethylamine (653 μL, 4.64 mmol) according to GP8. The title compound was obtained by purification of the crude product by flash chromatography (DCM / MeOH 99:1 → 95:5) (75.1 mg, 46%). 1 H NMR (400 MHz, CDCl3) δ 7.39 (dd, J = 8.5, 2.6 Hz, 1H), 7.32 (d, J = 2.2 Hz, 1H), 7.20-7.15 (m, 2H), 7.12-7.06 (m, 2H), 7.03-6.98 (m, 1H), 3.59-3.51 (m, 6H), 2.71 (s, 4H). 13 C{ 1 H} (101 MHz, CDCl3) δ 159.5, 159.0, 152.0, 139.6, 132.9, 130.5, 129.1, 127.2, 126.0, 125.1, 124.8, 122.9, 120.3, 114.6, 51.6, 47.2, 46.1. MALDI-HRMS:m / z C 19 H 18 Calculated value of ClN4O[M+H]+: 353.1164, measured value: 353.1165.

[0202] 2-(4-(2-chlorodibenzo[b,f][1,4]oxazepine-11-yl)piperazine-1-yl)-N'-hydroxyacetimidoamide (compound 17) [ka] To a solution of compound 16 (50.0 mg, 0.140 mmol) in EtOH (1.0 mL), an aqueous hydroxylamine solution (50 wt%, 23.2 μL, 0.379 mmol) was added at room temperature, and the reaction mixture was heated under reflux overnight. The crude product was purified by flash chromatography (DCM / MeOH 99:1 → 95:5) to obtain the title compound (35.4 mg, 65%). 1H NMR (400 MHz, DMSO-d6) δ 9.07 (br s, 1H), 7.61 (dd, J = 8.7, 2.6 Hz, 1H), 7.41-7.37 (m, 2H), 7.20-7.17 (m, 1H), 7.10-6.98 (m, 3H), 5.30 (s, 2H), 3.47 (br s, 4H), 2.92 (s, 2H), 2.54-2.46 (m, 4H). 13 C{ 1 H} (101 MHz, DMSO-d6) δ 158.6, 158.1, 151.2, 149.9, 139.9, 132.9, 129.5, 128.7, 126.5, 125.8, 124.5, 124.2, 123.1, 120.2, 57.8, 52.1, 46.8. MALDI-HRMS:m / z C 19 H 21 Calculated value of ClN5O2[M+H]+: 386.1378, measured value: 386.1371.

[0203] 5-((4-(8-fluoro-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine-11-yl)piperazine-1-yl)methyl)-1,2-dihydro-3H-1,2,4-triazole-3-one (compound 18) [ka] Compound 187 (36.0 mg, 0.0985 mmol), 3-(chloromethyl)-1H-1,2,4-triazole-5(4H)-one (27.1 mg, 0.197 mmol), and triethylamine (137 μL, 0.985 mmol) were synthesized according to GP8. The title compound was obtained by purification of the crude product by preparative HPLC (13.2 mg, 29%). 1H NMR (500 MHz, DMSO-d6) δ 11.36 (s, 1H), 11.26 (s, 1H), 7.97 (dd, J = 8.7, 2.1 Hz, 1H), 7.72 (d, J = 2.1 Hz, 1H), 7.61 (d, J = 8.5 Hz, 1H), 7.28-7.24 (m, 1H), 6.87-6.80 (m, 2H), 3.49 (br s, 4H), 3.36 (s, 2H), 2.60-2.44 (m, 4H). 13 C{ 1 H} (126 MHz, DMSO-d6) δ 162.5, 160.7, 158.7, 158.5, 156.2, 147.1, 144.3, 141.48, 128.7, 126.3, 124.6, 123.3, 122.5, 121.4, 112.3, 110.3, 53.2, 51.9, 45.7. MALDI-HRMS:m / z C 21 H 19 The calculated value of F4N6O2[M+H]+ is 463.1500, and the measured value is 463.1499.

[0204] 5-((4-(2-chloro-8-fluorodibenzo[b,f][1,4]oxazepine-11-yl)piperazine-1-yl)methyl)-1,2-dihydro-3H-1,2,4-triazole-3-one (compound 19) [ka] Compound 175 (50.0 mg, 0.151 mmol), 3-(chloromethyl)-1H-1,2,4-triazole-5(4H)-one (41.5 mg, 0.115 mmol), and triethylamine (210 μL, 1.51 mmol) were synthesized according to GP8. The title compound was obtained by purification of the crude product by preparative HPLC (38.9 mg, 60%). 1H NMR (400 MHz, DMSO-d6) δ 11.38 (s, 1H), 11.26 (s, 1H), 7.44-7.16 (m, 5H), 6.83-6.76 (m, 2H), 3.56 (s, 4H), 3.35 (s, 2H), 2.59-2.41 (m, 4H). 13 C{ 1 H} (101 MHz, DMSO-d6) δ 160.8, 158.5, 158.4, 156.2, 147.5, 144.3, 141.8, 133.1, 129.6, 128.8, 124.2, 123.0, 121.6, 112.2, 110.1, 53.2, 52.0, 46.5. MALDI-HRMS:m / z C 20 H 19 Calculated value of ClFN6O2[M+H]+: 429.1237, measured value: 429.1239.

[0205] 5-((4-(2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine-11-yl)-1,4-diazepan-1-yl)methyl)-1,2-dihydro-3H-1,2,4-triazole-3-one (compound 20) [ka] Compound 14 (20.0 mg, 0.0553 mmol), 3-(chloromethyl)-1H-1,2,4-triazole-5(4H)-one (15.2 mg, 0.111 mmol), and triethylamine (77.9 μL, 0.553 mmol) were synthesized according to GP8. The title compound was obtained by purification of the crude product by flash chromatography (DCM / MeOH 95:5 → 8:2) (16.8 mg, 67%). 1H NMR (500 MHz, DMSO-d6) δ 11.29 (s, 1H), 11.20 (s, 1H), 7.91 (dd, J = 8.6, 2.1 Hz, 1H), 7.77 (d, J = 2.0 Hz, 1H), 7.58 (d, J = 8.5 Hz, 1H), 7.20 (dd, J = 8.0, 1.4 Hz, 1H), 7.08-7.01 (m, 2H), 6.96-6.92 (m, 1H), 3.60 (s, 4H), 3.44 (s, 2H), 2.80-2.64 (m, 4H), 1.98-1.75 (m, 2H). 13 C{ 1 H} (126 MHz, DMSO-d6) δ 162.2, 157.7, 156.2, 150.6, 145.2, 140.6, 129.7, 126.7, 126.4, 126.0, 125.7, 123.9, 123.7, 123.3, 122.3, 120.2. MALDI-HRMS:m / z C 22 H 22 The calculated value of F3N6O2[M+H]+ is 459.1751, and the measured value is 459.1752.

[0206] 2-(4-(8-fluoro-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine-11-yl)piperazine-1-yl)ethane-1-ol (compound 21) [ka] Compound 232 (42.0 mg, 0.133 mmol) and 1-(2-hydroxyethyl)piperazine (33.3 μL, 0.266 mmol) were synthesized according to GP7. The title compound was obtained by purification of the crude product by flash chromatography (DCM / MeOH 99:1 → 95:5) (24.4 mg, 44%). 1H NMR (400 MHz, CDCl3) δ 7.72 (dd, J = 8.5, 1.8 Hz, 1H), 7.61 (d, J = 2.3 Hz, 1H), 7.36 (d, J = 8.8 Hz, 1H), 7.07-7.04 (m, 1H), 6.85-6.82 (m, 1H), 6.71-6.67 (m, 1H), 3.62 (br s, 4H), 3.17-2.93 (m, 5H). 13 C{ 1 H} (101 MHz, CDCl3) δ 163.2, 160.5, 159.4, 147.7, 141.4, 129.9, 127.7, 127.1, 123.6, 124.1, 122.2, 120.9, 113.4, 110.4, 59.5, 57.9, 52.8, 47.5. MALDI-HRMS:m / z C 20 H 20 Calculated value of F4N3O2[M+H]+: 410.1486, measured value: 410.1485

[0207] 5-Chloro-2-(2-nitrophenoxy)methyl benzoate (compound 22) [ka] The compound was synthesized from methyl 5-chlorosalicylate (2.80 g, 15.0 mmol), 2-fluoronitrobenzene (1.06 mL, 10.0 mmol), and potassium carbonate (2.07 g, 15 mmol) according to GP1. Purification of the crude product by flash chromatography (n-hexane / siRNA 9:1 → 8:2) yielded the title compound as a colorless solid (3.07 g, 99%). 1H NMR (250 MHz, DMSO-d6) δ 8.07 (dd, J = 8.1, 1.7 Hz, 1H) 7.92 (d, J = 2.7 Hz, 1H), 7.75 (dd, J = 8.8, 2.8 Hz, 1H) 7.68-7.60 (m, 1H), 7.37-7.30 (m, 1H), 7.27 (d, J = 8.8 Hz, 1H), 7.00 (d, J = 8.4, 1.1 Hz, 1H), 3.70 (s, 3H).

[0208] 2-(2-nitrophenoxy)-5-(trifluoromethyl)methyl benzoate (compound 23) [ka] The compound was synthesized from methyl 2-hydroxy-5-(trifluoromethyl)benzoate (991 mg, 4.50 mmol), 2-fluoronitrobenzene (317 μL, 3.00 mmol), and potassium carbonate (622 mg, 4.50 mmol) according to GP1. Purification of the crude product by flash chromatography (hexane / siRNA 9:1 → 8:2) yielded the title compound as a colorless solid (615 mg, 60%). 1 H NMR (250 MHz, CDCl3) δ 8.27 (s, 1H) 8.03 (d, J = 8.2 Hz, 1H), 7.75 (d, J = 8.6 Hz, 1H), 7.56 (t, J = 8.2 Hz, 1H), 7.29 (t, J = 8.2 Hz, 1H), 7.08 (d, J = 8.6 Hz, 1H), 6.98 (d, J = 8.3 Hz, 1H), 3.85 (s, 3H).

[0209] 2-(2-aminophenoxy)-5-chlorobenzoate methyl (compound 24) [ka] The compound was synthesized from methyl 5-chloro-2-(2-nitrophenoxy)benzoate (954 mg, 3.10 mmol) and tin(II) chloride dihydrate (2.80 g, 12.4 mmol) according to GP2. Purification of the crude product by flash chromatography (hexane / siRNA 9:1 → 8:2) yielded the title compound as a colorless solid (635 mg, 74%). 1 H NMR (300 MHz, CDCl3) δ 7.82 (d, J = 2.6 Hz, 1H), 7.33 (dd, J = 8.7, 2.6 Hz, 1H), 7.04-6.97 (m, 1H), 6.88-6.80 (m, 3H), 6.71 (td, J = 7.7, 1.5 Hz, 1H), 3.89 (s, 3H), 3.64 (br s, 2H).

[0210] 2-(2-aminophenoxy)-5-(trifluoromethyl)methyl benzoate (compound 25) [ka] The compound was synthesized from methyl 2-(2-nitrophenoxy)-5-(trifluoromethyl)benzoate (334 mg, 0.980 mmol) and tin(II) chloride dihydrate (900 mg, 3.91 mmol) according to GP2. Purification of the crude product by flash chromatography (hexane / siRNA 9:1 → 8:2) yielded the title compound as a colorless solid (171 mg, 56%). 1 H NMR (250 MHz, CDCl3) δ 8.12 (d, J = 2.1 Hz, 1H), 7.61 (dd, J = 8.7, 2.4 Hz, 1H), 7.10-7.02 (m, 1H), 6.98-6.93 (m, 2H), 6.84 (dd, J = 7.9, 1.5 Hz, 1H), 6.79-6.72 (m, 1H), 3.99 (br s, 2H), 3.94 (s, 3H).

[0211] 2-Chlorodibenzo[b,f][1,4]oxazepine-11(10H)-one (compound 26) [ka] The compound was synthesized from methyl 2-(2-aminophenoxy)-5-chlorobenzoate (630 mg, 2.27 mmol) and concentrated sulfuric acid (150 μL, 2.81 mmol) according to GP3. After filtration, the title compound was obtained as a colorless solid (508 mg, 90%). 1 H NMR (250 MHz, DMSO-d6) δ 10.66 (s, 1H), 7.72 (d, J = 2.5 Hz, 1H) 7.67 (dd, J = 8.6, 2.8 Hz, 1H), 7.40 (d, J = 8.6 Hz, 1H) 7.34 (dt, J = 7.1, 1.2 Hz, 1H), 7.21-7.10 (m, 3H).

[0212] 2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine-11(10H)-one (compound 27) [ka] The compound was synthesized from methyl 2-(2-aminophenoxy)-5-(trifluoromethyl)benzoate (166 mg, 0.533 mmol) and concentrated sulfuric acid (35.3 μL, 0.663 mmol) according to GP3. After filtration, the title compound was obtained as a colorless solid (73 mg, 49%). 1 H NMR (250 MHz, DMSO-d6) δ 10.77 (s, 1H), 8.05-7.98 (m, 2H) 7.59 (d, J = 8.3 Hz, 1H), 7.39 (dt, J = 7.3, 1.2 Hz, 1H), 7.23-7.13 (m, 3H).

[0213] 2-Fluoro-N-(5-Fluoro-2-hydroxyphenyl)-5-(trifluoromethyl)benzamide (Compound 28) [ka] The compound was synthesized according to GP4 from 2-fluoro-5-(trifluoromethyl)benzoic acid (1.50 g, 7.06 mmol), thionyl chloride (1.04 mL, 14.1 mmol), 2-amino-4-fluorophenol (925 mg, 7.06 mmol), and triethylamine (1.98 mL, 14.1 mmol). Purification of the crude product by flash chromatography (n-hexane / siRNA 99:1 → 2:3) yielded the title compound as a red solid (1.43 g, 64%). 1 H NMR (400 MHz, DMSO-d6) δ 8.17-8.13 (m, 1H), 8.05-7.91 (m, 2H), 7.66-7.59 (m, 1H), 6.93-6.80 (m, 2H).

[0214] 8-Fluoro-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine-11(10H)-one (compound 29) [ka] The compound was synthesized from 2-fluoro-N-(5-fluoro-2-hydroxyphenyl)-5-(trifluoromethyl)benzamide (196 mg, 0.618 mmol) and sodium hydroxide (25 mg, 0.618 mmol) according to GP5. After filtration, the title compound was obtained as a light brown solid (168 mg, 92%). 1 H NMR (250 MHz, DMSO-d6) δ 10.85 (s, 1H), 8.04-8.00 (m, 2H), 7.61-7.58 (m, 1H), 7.48-7.40 (m, 1H), 7.06-6.97 (m, 2H).

[0215] 2,11-Dichlorodibenzo[b,f][1,4]oxazepane (compound 30) [ka] The compound was synthesized according to GP6 from 2-chlorodibenzo[b,f][1,4]oxazepine-11(10H)-one (558 mg, 2.27 mmol), phosphorus oxychloride, and N,N-dimethylaniline (173 μL, 1.36 mmol). After work-up, the title compound was obtained as a brown solid (598 mg, 99%).

[0216] 11-Chloro-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine (compound 32) [ka] The compound was synthesized according to GP6 from 2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine-11(10H)-one (73 mg, 0.261 mmol), phosphorus oxychloride, and N,N-dimethylaniline (20.3 μL, 0.160 mmol). After work-up, the title compound was obtained as a brown solid (64 mg, 82%).

[0217] 2,11-Dichlorodibenzo[b,f][1,4]thiazepine (compound 33) [ka] The compound was synthesized according to GP6 from commercially available 2-chlorodibenzo[b,f][1,4]thiazepine-11(10H)-one (125 mg, 0.478 mmol), phosphorus oxychloride, and N,N-dimethylaniline (36.8 μL, 0.290 mmol). After work-up, the title compound was obtained as a brown solid.

[0218] 11-Chloro-8-fluoro-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine (compound 34) [ka] The compound was synthesized according to GP6 from 8-fluoro-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine-11(10H)-one (120 mg, 0.404 mmol), phosphorus oxychloride, and N,N-dimethylaniline (31.0 μL, 0.242 mmol). After work-up, the title compound was obtained as a light brown solid.

[0219] 5-Chloro-2-(4-fluoro-2-nitrophenoxy)methyl benzoate (compound 35) [ka] The compound was synthesized from methyl 5-chlorosalicylate (700 mg, 3.75 mmol), 2,5-difluoronitrobenzene (271 μL, 2.50 mmol), and potassium carbonate (524 mg, 3.75 mmol) according to GP1. The title compound was obtained by purification of the crude product by flash chromatography (n-hexane / siRNA 9:1 → 8:2) (811 mg, 99%). 1 H NMR (250 MHz, CDCl3) δ 7.97 (d, J = 2.7 Hz, 1H), 7.73 (dd, J = 7.7, 3.1 Hz, 1H), 7.50 (dd, J = 8.8, 2.7 Hz, 1H), 7.27-7.19 (m, 1H), 6.99 (d, J = 8.7 Hz, 1H), 6.87 (dd, J = 9.2, 4.4 Hz, 1H), 3.81 (s, 3H).

[0220] 2-(2-amino-4-fluorophenoxy)-5-chlorobenzoate methyl (compound 36) [ka] Compound 35 (1.02 g, 3.14 mmol) and tin(II) chloride dihydrate (2.89 g, 12.6 mmol) were synthesized according to GP2. The title compound was obtained by purification of the crude product by flash chromatography (n-hexane / siRNA 9:1 → 8:2) (630 mg, 68%). 1H NMR (250 MHz, CDCl3) δ 7.80 (d, J = 2.7 Hz, 1H), 7.34 (dd, J = 8.9, 2.7 Hz, 1H), 6.87-6.80 (m, 2H), 6.52 (dd, J = 9.9, 2.9 Hz, 1H), 6.44-6.35 (m, 1H), 3.97 (br s, 2H), 3.91 (s, 3H).

[0221] 2-Chloro-8-fluorodibenzo[b,f][1,4]oxazepine-11(10H)-one (compound 37) [ka] Compound 36 (625 mg, 2.11 mmol) and concentrated sulfuric acid (141 μL, 2.64 mmol) were synthesized according to GP3I. After workup, the title compound was obtained (499 mg, 90%). 1 H NMR (250 MHz, DMSO-d6) δ 10.73 (br s, 1H), 7.72 (d, J = 2.3 Hz, 1H), 7.68 (dd, J = 8.4, 1.4 Hz, 1H), 7.43-7.36 (m, 2H), 7.04-6.94 (m, 2H).

[0222] 2,11-Dichloro-8-fluorodibenzo[b,f][1,4]oxazepine (compound 38) [ka] Compound 37 (150 mg, 0.569 mmol), phosphorus oxychloride, and N,N-dimethylaniline (43.1 μL, 0.340 mmol) were synthesized according to GP6. After workup, the title compound was obtained (138 mg, 86%).

[0223] 2-Chloro-8-fluoro-11-(piperazin-1-yl)dibenzo[b,f][1,4]oxazepine (compound 39) [ka] Compound 38 (94.6 mg, 0.335 mmol) and piperazine (57.8 mg, 0.671 mmol) were synthesized according to GP7. The title compound was obtained by purification of the crude product by flash chromatography (DCM / MeOH 99:1 → 8:2) (74.1 mg, 67%). 1 H NMR (400 MHz, CDCl3) δ 7.72 (dd, J = 8.5, 1.8 Hz, 1H), 7.61 (d, J = 2.3 Hz, 1H), 7.36 (d, J = 8.8 Hz, 1H), 7.07-7.04 (m, 1H), 6.85-6.82 (m, 1H), 6.71-6.67 (m, 1H), 3.62 (br s, 4H), 3.17-2.93 (m, 5H). 13 C{ 1 H} (101 MHz, CDCl3) δ 161.5, 159.2, 159.1, 148.0, 141.2, 133.1, 130.7, 129.0, 124.5, 122.8, 120.8, 120.7, 112.2, 46.7, 44.6. MALDI-HRMS:m / z C 17 H 16 Calculated value of ClFN3[M+H]+: 332.0960, measured value: 322.0959.

[0224] 8-Fluoro-11-(piperazin-1-yl)-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine (compound 40) [ka] Compound 34 (128 mg, 0.404 mmol) and piperazine (69.6 mg, 0.808 mmol) were synthesized according to GP7. The title compound was obtained by purification of the crude product by flash chromatography (DCM / MeOH 99:1 → 8:2) (77.3 mg, 52%). 1H NMR (400 MHz, CDCl3) δ 7.72 (dd, J = 8.5, 1.8 Hz, 1H), 7.61 (d, J = 2.3 Hz, 1H), 7.36 (d, J = 8.8 Hz, 1H), 7.07-7.04 (m, 1H), 6.85-6.82 (m, 1H), 6.71-6.67 (m, 1H), 3.62 (br s, 4H), 3.17-2.93 (m, 5H). 13 C{ 1 H} (101 MHz, CDCl3) δ 163.3, 162.4, 159.4, 147.7, 141.4, 130.1, 127.8, 127.1, 124.7, 124.0, 122.5, 122.2, 120.9, 112.3, 45.3, 29.9. MALDI-HRMS:m / z C 18 H 16 The calculated value of F4N3O[M+H]+ is 366.1224, and the measured value is 366.1226.

[0225] 2-Fluoro-N-(5-Fluoro-2-hydroxyphenyl)-5-(trifluoromethyl)benzamide (Compound 41) [ka] The compound was synthesized according to GP4 from 2-fluoro-5-(trifluoromethyl)benzoic acid (1.50 g, 7.06 mmol), thionyl chloride (1.0 mL, 14.1 mmol), 2-amino-4-fluorophenol (925 mg, 7.06 mmol), and triethylamine (2.0 mL, 14.1 mmol). The title compound was obtained by purification of the crude product by flash chromatography (n-hexane / siRNA 99:1 → 4:6) (1.43 g, 64%). 1 H NMR (250 MHz, CDCl3) δ 8.15 (dd, J = 6.4, 2.1 Hz, 1H), 8.05-7.98 (m, 1H), 7.94 (dd, J = 10.4, 2.5 Hz, 1H), 7.63 (t, J = 9.6 Hz, 1H), 6.94-6.80 (m, 2H).

[0226] 8-Fluoro-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine-11(10H)-one (compound 42) [ka] Compound 41 (196 mg, 0.618 mmol) and sodium hydroxide (25 mg, 0.618 mmol) were synthesized according to GP5. After workup, the title compound was obtained (168 mg, 92%). 1 H NMR (250 MHz, DMSO-d6) δ 10.85 (br s, 1H), 8.05-8.00 (m, 2H), 7.60 (d, J = 8.1 Hz, 1H), 7.48-7.40 (m, 1H), 7.06-6.97 (m, 2H).

[0227] 11-Chloro-8-fluoro-2-(trifluoromethyl)dibenzo[b,f][1,4]oxazepine (compound 43) [ka] Compound 42 (120 mg, 0.404 mmol), phosphorus oxychloride, and N,N-dimethylaniline (31.0 μL, 0.242 mmol) were synthesized according to GP6. After workup, the title compound was obtained (50.4 mg, 39%).

[0228] Example 3: Functional Activity Test The functional activity of human Slack was screened in vitro using a cell-based FluxOR assay compared to roxapine as a reference activator. For this purpose, cultured HEK293 cells that stably express human Slack (referred to herein as HEK-Slack cells) were prepared by preventing compound-independent Slack activation by monovalent cations using Na. + The compound was incubated with the buffer solution that did not contain the compound. 7、19 .

[0229] In the initial experiment, the inventors confirmed that the modified FluxOR assay could detect roxapine-induced Slack activation (Figure 2). Furthermore, 2 mM Ca 2+ , 2 mM Mg 2+ , and Na containing 0.03% DMSO + In a buffer that does not contain [the specified substance], pre-stimulation with 25 μM roxapine induced Slack inhibitor "Compound 31" (References) 18 Dose-response experiments using ) [ka] (Compound 31) Compound 31 inhibits the F / F (baseline) ratio in a dose-dependent manner (IC 50 We clarified (=2.1μM) and confirmed that the readings depended on Slack. Subsequently, EC 50 and E MAX A dose-response curve was created to determine the values.

[0230] In detail, these experiments determined the Slack-activating efficacy and titer of novel compounds using the commercially available FluxOR® potassium ion channel assay (Invitrogen). HEK293 cells stably expressing human Slack (referred to herein as HEK-Slack cells) were plated at a density of 50,000 cells / well in each well of a poly-D-lysine-coated (75 μg / mL, Sigma Aldrich) 96-well, black-walled, clear-bottom microplate (Greiner Bio-One) containing 10% FBS and 1% penicillin / streptomycin 24 hours prior to the assay.

[0231] On the day of the experiment, the cells were loaded with FluxOR® dye according to the manufacturer's protocol. Na was added to a solution containing 140 mM choline chloride, 5 mM KCl, 2 mM CaCl2, 2 mM MgSO4, 10 mM HEPES, and 5.55 mM glucose, adjusted to pH 7.4 with KOH. +Assay buffer that did not contain was used. To reduce background fluorescence, 10% BackDrop™ Background Suppressor (Invitrogen) was added to all assay components. To reduce background fluorescence, 10% BackDrop™ Background Suppressor (Invitrogen) was added to all assay components.

[0232] After the washing step, cells were incubated with the compounds at 37°C for 30 minutes. All compounds were prepared as 33.3 mM DMSO stocks and diluted to final concentrations of 100 μM, 50 μM, 25 μM, 12.5 μM, 6.25 μM, and 3.125 μM in assay buffer containing 0.03% DMSO on the experimental day. 50 μM roxapine was used as a positive control. After stimulation with Tl2SO4 solution, fluorescence was measured over 100 seconds using an Infinite M200 Microplate Reader (Tecan) (excitation 485 nm, emission 525 nm). The assay was performed in triplicate, and the ratio to baseline fluorescence was calculated for all time points. Dose-response curves including the six concentrations were obtained, showing the fluorescence / fluorescence ratio at 100 seconds. ベースライン The formulas were created in triplicates using ratios. The values ​​were calculated for roxapine within each individual experiment as follows: 100 × ((F / F B (Compound)-F / F B (vehicle 平均 )) / (F / F B (Roxapine)-F / F B (vehicle 平均 )) EC 50 and E MAX Standard logistic nonlinear regression analysis using GraphPad Prism 9.0 was used to generate the values.

[0233] Of the 125 novel compounds screened, 45 functioned as Slack activators to varying degrees. The most interesting candidates showed titers ranging from 3.2 μM to 62.5 μM, while roxapine activated Slack at a titer of 20.7 μM (Figures 3A, 3B, and Table 3). Efficacy (EMAX The values ​​ranged from 63.1% to 264% compared to roxapine (Figures 3A, 3B, and Table 3). This indicates that roxapine exerted a Slack-dependent effect in mice at low doses. 19 , and the FluxOR assay inhibits Slack activity with 2mM Ca 2+ The procedure was performed in a buffer containing the following: 13 Considering this, the EC of the new compound 50 Although the values ​​are in the micromolar range, the inventors concluded that the novel compounds can sufficiently activate Slack in vivo at standard doses. For compounds 10, 11, and 18-21, the corresponding time-dependent fluorescence / fluorescence ベースライン The ratios are shown in Figure 3E in comparison to roxapine and the vehicle.

[0234] TIFF2026509430000155.tif241170

[0235] EC in the lower micromolar range obtained by the FluxOR assay 50 In interpreting the values, these experiments showed that 2 mM Ca inhibits Slack activity. 2+ It must be taken into consideration that the assay was performed using an assay buffer containing [specific component]. Under these conditions, EC that activates Slack 50 Achieving values ​​in the lower nanomolar range is virtually impossible. However, these values ​​are still useful for estimating the efficacy of the compounds. As shown in further experiments on the inhibition of scratching behavior in mice after administration of compounds 6 and 10 (comparative example 9), compounds with EC50s shown in Table 3 sufficiently activate Slack in vivo.

[0236] Evaluation of analogues of this series of Slack activators in the FluxOR assay suggests that activity is improved by substituting the chloro substituent of B with a trifluoromethyl group (Compound 2). The addition of fluoro substituents to other aryl rings (A) increased efficacy but decreased titer (Compound 4). In some cases, sulfur analogues showed comparable activity (Compound 7). Substitution of piperazine with homopiperazine resulted in EC 50 The value improved, but E MAX The value decreased (compound 3). Modification of alkyl residues of E was generally well tolerated and held potential for adjusting Slack activity and pharmacological properties. However, these conservative substituents did not alter the high lipophilicity of the original skeleton, likely leading to CNS exposure similar to roxapine. Substitution of methyl groups on the piperazine ring offered an opportunity to significantly increase polarity. Derivatives with aliphatic hydroxyl groups had acceptable EC values. 50 The compound showed good efficacy in terms of values ​​(Compound 5). Finally, we were able to incorporate an ionizable carboxyl group (Compound 6).

[0237] Example 4: Slack activation by novel compounds was validated using whole-cell patch-clamp recordings in HEK-Slack cells. Typically, in these experiments, HEK-Slack cells were plated onto poly-D-lysine-coated (100 μg / mL, Sigma Aldrich) coverslips one day prior to the experiment and cultured in DMEM containing 10% FBS and 1% penicillin / streptomycin at 37°C in 5% CO2. Whole-cell voltage clamp recordings were obtained using an EPC 9 amplifier and Patchmaster software (HEKA Electronics, Lambrecht / Palatinate, Germany). Current was sampled at 20 kHz and filtered at 5 kHz. Data analysis was performed using Fitmaster software (HEKA Electronics). Membrane potential was maintained at -70 mV, with outward K + Current (I κThe depolarization was induced by a depolarization process (duration 500 milliseconds) increasing in 20 mV increments from -120 mV to +120 mV. The pipette solution contained 140 mM KCl, 2 mM MgCl2, 5 mM EGTA, and 10 mM HEPES, and was adjusted to pH 7.4 with KOH. The extracellular solution contained 140 mM NaCl, 5 mM KCl, 2 mM CaCl2, 2 mM MgCl2, and 10 mM HEPES, and was adjusted to pH 7.4 with NaOH. The osmotic pressure of all solutions was adjusted to 290 mOsmol / L to 300 mOsmol / L with glucose. The patch pipette resistance was 6 MΩ to 8 MΩ, obtained from borosilicate glass capillaries (Science Products) using a conventional puller (DMZ-Universal Puller, Zeitz Instruments).

[0238] After baseline measurement, a novel compound or roxapine (both with a final concentration of 50 μM) dissolved in an external solution containing 0.03% DMSO was added to the bath without continuous perfusion, and within 5 minutes, K + Current was measured. Roxapine was used as the positive control, while a vehicle containing 0.03% DMSO was used as the negative control. A series of test pulses with a length of 500 milliseconds were applied at 20 mV intervals from -120 mV to +120 mV at a holding potential of -70 mV.

[0239] The multiplier in the patch-clamp experiment was determined by calculating the baseline current density relative to the current density after compound application. From these patch-clamp records, the novel compounds (all except compound 3) were found to be 1 compared to the vehicle. κ It was found that the amplitude was significantly increased (Figures 3C and 4A, representative I κ The trace is shown in Figure 4B.

[0240] At a voltage of +80mV, the current density of the novel compound increased by 2.04 to 14.77 times compared to baseline (Figure 3C). Notably, the effectiveness of the novel compound in patch-clamp recordings (i.e., the multiplier increase in current density) was significantly correlated with the effectiveness in the FluxOR assay (Figure 3D). In addition to the results shown in Figures 3C and 4A, the following results were monitored for compound 10 in patch-clamp experiments performed similarly as described above: At a concentration of compound 10 of 25 μM, a 7.74 ± 1.525-fold increase compared to baseline was monitored, and at a concentration of compound 10 of 50 μM, a 12.54 ± 1.395-fold increase compared to baseline was monitored.

[0241] Overall, FluxOR assays and patch-clamp analyses in HEK-Slack cells demonstrate that the compounds according to the present invention activate Slack in vitro.

[0242] Example 5: Microsomal stability assay of compound 10 in liver microsomes The metabolic stability of compound 10 in liver microsomes, as shown in Table 4, was measured by the following assay. The test compound was dissolved in DMSO (1 mM). The solubilized test compound (5 μL, final concentration 10 μM) was pre-incubated at 37°C in 432 μL of phosphate buffer (0.1 M, pH=7.4) with 50 μL of NADPH regeneration system (30 mM glucose-6-phosphate, 4 U / mL glucose-6-phosphate dehydrogenase, 10 mM NADP, 30 mM MgCl2). After 5 minutes, the reaction was started by adding 13 μL of a Sprague-Dawley rat liver microsome mix (Invitrogen, 20 mg of protein per 1 mL of 0.1 M phosphate buffer) in a shaking water bath at 37°C. The reaction was stopped by adding 500 μL of ice-cold methanol at 0, 15, 30, and 60 minutes. The samples were centrifuged at 4°C and 5000×g for 5 minutes, and the test compound was quantified from the supernatant by HPLC. The mobile phase composition was adjusted to match the test compound, ranging from 40% to 90% MeOH and 10% to 60% water (0.1% formic acid). Flow rate: 1 mL / min, stationary phase: Purospher STAR, RP18, 5 μm, 125×4, pre-column: Purospher STAR, RP18, 5 μm, 4×4, detection wavelengths: 254 nm and 280 nm, injection volume: 50 μL. Control samples were prepared to confirm the stability of the test compound in the reaction mixture. The first control was free of NADPH necessary for the enzymatic activity of microsomes, the second control contained inactivated microsomes (incubated at 90°C for 20 minutes), and the third control did not contain the test compound (to determine the baseline). The amount of the test compound was quantified using an external calibration curve. Data are expressed as the mean ± SEM of residual compound amounts obtained from three independent experiments. In vitro half-lives were calculated by logarithmically transforming the residual amount of non-metabolized compounds with time.

[0243] TIFF2026509430000156.tif42170

[0244] Example 6: Blood-brain barrier-specific parallel artificial membrane permeability assay (PAMPA-BBB) BBB permeability was estimated using a BBB-specific parallel artificial membrane permeability assay (PAMPA-BBB). PAMPA-BBB is a non-cell-based permeability model that predicts the passive permeability of drug molecules across phospholipid membranes using porcine polar brain lipid extracts. 20 .

[0245] Procedure described by Mueller et al. 21 According to a slightly modified version, log P is used to predict passive permeability of the blood-brain barrier (BBB). e The effective permeability value was determined. A 5 v / v% DMSO solution of each compound was prepared in PBS (phosphate-buffered saline; 0.01 M, pH=7.4) buffer to a final nominal concentration of 100 μM. If the test compound was insoluble at this concentration, these solutions were sonicated and then centrifuged. log P e Since it depends only on the relative concentration, a saturated homogeneous solution (C D It is sufficient to proceed with (0).

[0246] log P e A BBB-specific PAMPA system was used to determine membrane retention (MR). Each well of the top plate (MultiScreen MAIPNTR10; Millipore (Billerica, USA)) was carefully coated with 5 μL of porcine polar brain lipid extract (PBLE; Avanti Polar Lipids (Birmingham, USA)) solution, and then C D(0) 150 μL of solution was placed on the membrane. The bottom plate (MultiScreen MDCPN2M50; Millipore (Billerica, USA)) was filled with 2050 μL of PBS. The PBLE solution consisted of 10 μL of n-dodecane and 1 mg of PBLE per 30 μL of n-hexane. The donor plate was placed on the acceptor plate and covered with a wet paper towel and the plate lid. This sandwich system was incubated at 37°C for 4 hours. The plates were shaken and incubated using a Thermo Scientific® MaxQ® 4000 Benchtop Orbital Shaker (Hale, Belgium). After incubation, the PAMPA sandwich plate was separated and the donor solution (C D (t)) and acceptor solution (C A The compound concentration in (t) was determined by UPLC-MS (Waters (Milford, USA)). The concentration of the donor solution at time zero (C) D (0)) was determined using the supernatant after centrifugation. Under isopH conditions, the effective permeability and membrane retention of the drug were calculated using the following formula. 22 :

number

[0247] In the formula, P e is the effective transmittance coefficient (cm / s), and A is the filter area (0.3 cm²). 2 ) and V D and V A This is the donor phase (0.15 cm). 3 ) and acceptor phase (2.05 cm) 3 ) is the capacity, t is the incubation time (seconds), and τ SS This is the time (seconds) to reach a steady state (240 seconds), and C D (t) is the concentration of the compound in the donor phase at time t (mol / cm³). 3 ) and C A (t) is the concentration of the compound in the acceptor phase at time t (mol / cm³). 3 ) and C D(0) is the concentration of the compound in the donor phase at time 0 (mol / cm³). 3 ) and MR is the estimated membrane retention coefficient (estimated mole fraction of solute lost to the membrane):

number

[0248] As shown in Figure 5, the novel compounds exhibit various effective BBB permeability (log P e The values ​​and membrane retention coefficient (MR; indicating the fraction of solute bound to the membrane) were shown. The evaluation of a series of compounds in this assay revealed that substitution in the aromatic core, introduction of carboxylic acid linkers, and introduction of hydroxy linkers had little effect on permeability. The absence of a decrease in passive permeability in amphoteric carboxylic acid derivatives may be due to intramolecular charge neutralization, as is known with cetirizine. 23 In contrast, replacing piperazine with homopiperazine increased permeability. A high MR coefficient corresponds to log P e Considering the limitations on the predictive validity of the values, from in vitro PAMPA-BBB data, compound 6 and compound 8 (both log P e <-4.57 and MR≦0) and compound 10 (log P e The inventors concluded that values ​​of -4.492 and MR=0.02 are most likely to have low BBB permeability.

[0249] TIFF2026509430000159.tif83170

[0250] Example 7: Pharmacokinetic properties of a novel Slack activator obtained from in vivo studies The pharmacokinetic properties of the compound were investigated in vivo by administering 1 mg / kg intraperitoneally (ip) to mice and determining plasma and brain levels over 8 hours by LC-MS.

[0251] Pharmacokinetic studies were conducted by Pharmacelsus (Saarbrücken, Germany). A total of 27 male C57BL / 6 mice (8 weeks old, weighing 23g-31g, purchased from Janvier Labs (France)) were used. The animals were housed in temperature-controlled rooms (20°C-24°C) and maintained on a 12-hour light / dark cycle. Food and water were freely available. All experimental procedures were approved and carried out in accordance with the regulations of the local animal welfare authority (Landesamt fuer Gesundheit und Verbraucherschutz, Abteilung Lebensmittel-und Veterinaerwesen) (Saarbrücken).

[0252] The animals were divided into three groups (9 mice per group), and cassette administration was performed. In this procedure, mice were simultaneously administered intraperitoneally in single doses of 3 to 4 compounds (dissolved in 0.9% NaCl containing 20% ​​cyclodextrin) at a dose of 1 mg per kg of body weight. Two blood samples were taken from each animal under short-term isoflurane anesthesia from the retroorbital venous plexus (at 0.25 and 0.5 hours from 3 mice, at 1 and 2 hours from 3 mice, and at 4 and 8 hours from 3 mice). Li-heparin was used as an anticoagulant. Plasma samples were obtained by centrifugation at 3000 × g for 10 minutes at 4°C. Immediately after the last blood sample collection, the animals were sacrificed by cervical dislocation, and the brains were removed and frozen in liquid nitrogen. Plasma and brain samples were stored at -80°C until LC-MS analysis.

[0253] LC-MS analysis was performed as follows: For roxapine, compound 2, and compound 6, the HPLC system consisted of a Surveyor Pump Plus pump and a Surveyor Plus Autosampler (Thermo Fisher Scientific, USA), and mass spectrometry was performed on a TSQ Quantum Discovery MAX mass spectrometer (Thermo Fisher Scientific, USA) equipped with a positive SRM mode ESI (electrospray ionization) interface, connected to a PC running standard software Xcalibur 2.0.7. For all other compounds, the HPLC system consisted of a U-HPLC pump (Accela) and an AS Open autosampler (Thermo Fisher Scientific, USA), and mass spectrometry was performed on a Q Exactive (Orbitrap) high-precision mass spectrometer (Thermo Fisher Scientific, USA) equipped with a heated electrospray (H-ESI) interface, connected to a PC running standard software Chromeleon 7.2. For all compounds, the HPLC pump flow rate was set to 600 μl / min, and the compounds were separated using an analytical column with an appropriate pre-column. Pharmacokinetic analysis was performed using Kinetica 5.0 software (Thermo Scientific, Waltham, USA) with a non-compartmental model. All given parameters were obtained by trapezoidal area calculation.

[0254] All compounds reached their peak in plasma within 30 minutes after injection (Figure 6A). Peak (C max Plasma concentrations range from 25.6 ng / ml to 315.5 ng / ml, and the terminal phase elimination half-life (t 1 / 2zThe incubation period ranged from 1.2 hours to 3.9 hours (Table 6). In the brain, the peak levels were measured within 30 minutes after injection, reaching a maximum of 369 ng / ml (Figure 6A). The brain / plasma ratio is shown in Figure 6B. Notably, compounds 6 and 10 had low brain / plasma ratios of 0.3 or less, suggesting that the permeability of these compounds into the brain is limited.

[0255] TIFF2026509430000160.tif116170

[0256] Example 8: Off-target profile of compound 6 Roxapine is a first-generation antipsychotic that has substantial binding affinity to many receptors. 24 The inventors used the SafetyScreen44 panel (Eurofins), which screens for the interaction of compounds with 44 targets, to investigate the pharmacological profiles of compound 6 and roxapine (both 10 μM) in vitro.

[0257] In this panel, roxapine (10 μM) is equivalent to adrenaline (α 1A and α 2A ), dopamine (D1 and D 2S ), histamine (H1 and H2), muscarine (M1, M2 and M3), serotonin (5-HT) 1A , 5-HT 1B , 5-HT 2A , 5-HT 2B and 5-HT3 receptor, Na + Of the 44 targets, including channels, norepinephrine transporters (NETs), and serotonin transporters (SETs), 17 showed substantial binding inhibition (over 50%) (Figure 7), confirming the "dirty" nature of first-generation antipsychotics. Notably, compound 6 (10 μM) showed considerably less off-target activity, and inhibited the binding of 8 targets (α 1A , D1, D 2S H1, 5-HT 2A , 5-HT 2BCompound 10 (10 μM) showed substantial binding to NET and SET, and showed substantial binding to five targets (CB1, D1, H1, 5-HT). 2A and 5-HT 2B It showed substantial binding inhibition against ). Most of the targets affected by compound 6 or compound 10 are mainly expressed in the CNS (especially D1, D 2S , 5-HT 2B NET and SET 25、26 Considering that both compounds exhibit limited brain permeability (as reported by www.proteinatlas.org), these data suggest that the pharmacological profiles of compounds 6 and 10 are improved compared to roxapine. Furthermore, compounds 6 and 10 (like roxapine) are effective in off-target screening against human ether-a-go-go (hERG) potassium channels or voltage-gated (K) receptors. V The compound did not show measurable binding to potassium channels (Figure 7), indicating that it does not act as a nonspecific regulator of potassium channels.

[0258] Example 9: Compounds 6 and 10 significantly inhibit pruritus-related behavior in mice. Further studies were conducted to evaluate the in vivo efficacy of compounds 6 and 10 in mice. Specifically, experiments were performed on 8- to 16-week-old C57BL / 6N mice (Charles River, Sulzfeld, Germany) and animals with Slack gene deletion (Slack ― / ― mouse 27The experiments were conducted in either male or female animals. The animals were kept in a 12-hour light / dark cycle with free access to food and water. All experiments were conducted in compliance with the International Association for the Study of Pain guidelines and the ARRIVE (Animal Research: Reporting on In Vivo Experiments) guidelines, and were approved and carried out in accordance with the regulations of the local animal welfare authority (Darmstadt Administrative District Office (Regierungspraesidium Darmstadt) (Germany)). All behavioral studies were conducted during a daytime light cycle at room temperature (20°C-24°C) by observers blinded to the treatment of the animals and / or their genotypes.

[0259] The accelerated rotorod and vertical pole tests were conducted as follows: Mice were placed on a rotorod treadmill (Ugo Basile, Italy) at increasing speeds (4 rpm to 40 rpm over 300 seconds) and trained for 4 to 5 consecutive days. Only mice that reached 300 seconds without falling were included in the experiment. On the day of the test, a compound in a vehicle consisting of 0.9% saline (Sigma-Aldrich) containing 10% 2-hydroxypropyl-β-cyclodextrin (ITW Reagents) was administered intraperitoneally, and the latency until the mouse fell from the accelerated rotorod was measured 15 minutes later. Immediately afterward, the mouse was placed head-up on top of a vertical pole (1 cm in diameter, 40 cm in height) with a rough surface, and the time until the animal reached the ground was recorded (cutoff time 20 seconds). For further analysis, the average of three trials at each time point was calculated.

[0260] Treatment with compounds 6 and 10 (3 mg to 30 mg intraperitoneally) did not impair motor function in the accelerated rotarod test, a standard model of motor ability (Figure 8A), nor in the vertical pole test, which assesses basal ganglia-related motor impairments (Figure 8B). In contrast, roxapine at doses of 0.39 mg / kg or higher significantly reduced the ability of mice in both models (Figures 8A and 8B), which is consistent with its high CNS permeability and confirms previous reports. 28.

[0261] Acute pruritic behavior in mice was monitored using histamine-based, chloroquine-based, and SLIGRL-based acute pruritic behavior models. In these models, the hair on the dorsal outer side of the neck was shaved under short-term isoflurane anesthesia 3-4 days prior to the experiment. On the day of the experiment, mice were acclimatized for 30 minutes in individual plexiglass cylinders (30 cm in diameter). The compounds were dissolved in 0.9% saline containing 10% 2-hydroxypropyl-β-cyclodextrin and administered intraperitoneally in volumes of up to 200 μL. Fifteen minutes later, the pruritic substances chloroquine (200 μg), SLIGRL (100 μg), or histamine (800 μg), all dissolved in 0.9% saline (20 μL), were subcutaneously injected into the nape of the neck. The number of scratching behaviors on the nape of the neck was evaluated over a 30-minute period by video recording.

[0262] Antipruritic efficacy of compounds 6 and 10 in a histamine-independent pruritus model induced by the antimalarial drug chloroquine: Importantly, systemic treatment of mice with compound 6 (3 mg / kg, 10 mg / kg, or 30 mg / kg intraperitoneally) or compound 10 (3 mg / kg, 10 mg / kg, or 30 mg / kg intraperitoneally) 15 minutes before subcutaneous injection of chloroquine into the nape of the neck resulted in a dose-dependent reduction of scratching behavior in a histamine-independent pruritus model induced by the antimalarial drug chloroquine (Figure 8C). Further experiments showed that compound 6 (10 mg / kg) was effective in animals with a Slack gene deletion (Slack ― / ― Since it did not significantly alter chloroquine-induced scratching in mice, the conclusion that the antipruritic effect is mediated by Slack activation was supported (Figure 8D). The hypothesis that Slack is the main receptor that mediates the activity in vivo is supported by Slack ― / ― This is supported by observations that the effects of compound 6 were significantly reduced in mice and blocked by compound 31, a Slack inhibitor (Figure 8H).

[0263] These data, supported by genetic and pharmacological evidence, suggest that treatment with compound 6 or compound 10 reduces chloroquine-induced itching by activating Slack.

[0264] Antipruritic efficacy of compound 6 in histamine-independent itching induced by SLIGRL injection: Chloroquine-induced scratching in mice is caused by activation of MrgprA3 in the NP2 population of sensory neurons. 4、29 To further investigate the antipruritic efficacy of compound 6 in histamine-independent itching, the inventors evaluated the behavioral response after subcutaneous injection of SLIGRL (the peptide Ser-Leu-Ile-Gly-Arg-Leu). SLIGRL is expressed in the NP2 and NP3 populations as MrgprC11 (References) 30 ) is activated, which induces scratching. 4 Similar to the chloroquine model, compound 6 (10 mg / kg intraperitoneal) significantly inhibited SLIGRL-induced scratching behavior (Figure 8E). In contrast, subcutaneous histamine-induced scratching was unaffected by compound 6 (Figure 8F). This indicates that Slack's function in processing histamine-dependent itching is limited. Furthermore, this finding provides further evidence that the antipruritic effect of compound 6 observed in the histamine-independent model is not a result of motor impairment.

[0265] These studies did not observe any significant behavioral differences between sexes (Figures 9 and 11).

[0266] Furthermore, the effect of compound 6 on chronic itching was investigated in a model of allergic contact dermatitis. 31In short, the hair on the dorsal outer side of the neck was shaved under short-term isoflurane anesthesia. Three to four days later, 100 μL of 0.15% 2,4-dinitrofluorobenzene (DNFB) in acetone / olive oil (3:1) was applied to the shaved skin area. Ten to eleven days later, the skin was shaved again, and the DNFB solution was applied three to four days later (i.e., two application sessions at 14-day intervals). Ninety minutes after the second application, the mice were acclimatized for 15 minutes in individual plexiglass cylinders (30 cm in diameter).

[0267] This treatment resulted in persistent itching behavior, manifested as scratching and head shaking of the applied area, which was significantly correlated with scratching.

[0268] The compound or vehicle (10% 2-hydroxypropyl-β-cyclodextrin in 0.9% physiological saline) was administered intraperitoneally, and spontaneous scratching was videotaped 15 minutes later for 30 minutes (Figure 10A). Intraperitoneal treatment with compound 6 after a second DNFB exposure significantly inhibited the number of scratching behaviors and head shaking compared to animals treated with the vehicle (Figures 10B and 10C).

[0269] Similarly, the effects of compound 6 were investigated in the MC903 model of skin inflammation exhibiting several characteristics of allergic contact dermatitis and atopic dermatitis. 32 .

[0270] Under short-term isoflurane anesthesia, the hair on the dorsal outer side of the neck was shaved. Five days later, 20 μL of MC903 (0.2 mM; calcipotriol, Tocris) dissolved in anhydrous ethanol was applied to the skin on the back of the neck for seven consecutive days under short-term anesthesia. On day 8, the mice were acclimatized for 30 minutes in individual plexiglass cylinders (30 cm in diameter). The compound or vehicle (10% 2-hydroxypropyl-β-cyclodextrin in 0.9% saline) was administered intraperitoneally, and 15 minutes later, spontaneous scratching over a 30-minute period was videotaped (Figure 10D). In these studies, intraperitoneal administration of compound 6 significantly reduced both scratching and head-shaking behavior (Figures 10E and 10F). These findings are significant because they demonstrate that Slack activators can reduce already established pruritus.

[0271] Finally, potential side effects of compound 6 were evaluated by measuring cardiovascular function using pulse oximetry. A pulse oximeter (MouseOX Plus, Starr Life Sciences Corp) was used on conscious, freely moving mice. After acclimatizing the mice to color clips for at least 30 minutes continuously for 3 days, baseline measurements were performed. Subsequently, the compound was administered intraperitoneally, and after a 5-minute rest period, heart rate, respiratory rate, and arterial O2 saturation were recorded over 30 minutes. All datasets were sampled at 1 Hz, error-corrected according to the manufacturer's instructions, and averaged over 300 seconds for each time point.

[0272] In these pulse oximetry measurements in conscious, freely moving mice, intraperitoneal administration of compound 6 (10 mg / kg) did not affect either heart rate (Figure 12A) or respiratory rate (Figure 12B) during the 30-minute observation period. In contrast, both parameters were significantly reduced after intraperitoneal administration of morphine (10 mg / kg), which was used as a positive control in this experiment.

[0273] In summary, these results highlight that compound 6 effectively inhibits pruritic behavior in mice at acceptable doses in multiple models.

[0274] Example 9: Compound 6 inhibits itch-sensitive sensory neurons. To determine whether the antipruritic effect of compound 6 occurs directly at the neuronal level, whole-cell patch-clamp electrophysiological analysis was used to measure the excitability of DRG neurons.

[0275] Primary DRG neuronal cell cultures were prepared and stimulated overnight with an inflammatory soup to induce hyperexcitability. Briefly, C57BL / 6N naive mice (4-8 weeks old) were sacrificed by CO2 inhalation, and lumbar (L1-L5) DRGs were transferred to Hanks equilibrium salt solution (Gibco, Thermo Fisher Scientific). They were incubated with 500 U / mL collagenase IV and 2.5 U / mL dispase II (both Sigma Aldrich) for 60 minutes, with careful shaking every 20 minutes during that time. The DRGs were then washed and gently triturated twice using a flame-polished Pasteur pipette in neurobasal medium (Gibco, Thermo Fisher Scientific) containing 10% FBS and 0.5 mM GlutMax (Gibco, Thermo Fisher Scientific). Dissociated DRGs were seeded onto poly-D-lysine-coated (100 μg / mL) coverslips and cultured at 37°C in 5% CO2 in Neurobasal medium supplemented with 2% B27 (Gibco, Thermo Fisher Scientific), 1% penicillin / streptomycin, and 0.5 mM GlutMax.

[0276] To simulate the pathological state, DRG neuron cultures were prepared using an inflammatory soup (histamine: 10 μM, PGE2: 10 μM, serotonin: 10 μM, bradykinin: 10 μM). 33Incubate overnight with the mouse pruritus-sensitive neuron population (NP1-NP3). 4 Cells that bind to isolectin B4, a marker for non-peptidogenic C-fiber neurons, were recorded. Prior to recording, DRG neurons were pre-incubated with 10 μg / mL FITC-coupled IB4 (Sigma-Aldrich) for 5–10 minutes to select Slack-expressing neurons.

[0277] Whole-cell current-clamp recordings were obtained using an EPC 9 amplifier in combination with Patchmaster software (HEKA Electronics, Lambrecht / Palatinate, Germany). General settings, pipettes, and extracellular solutions were used as described above. To determine electrophysiological parameters, induced action potentials (APs) were triggered by 10-millisecond current injections, starting from 0 pA and injecting in 20 pA increments. Action potential (AP) firing was induced by depolarizing current pulses (200 pA to 950 pA in 150 pA increments, with a duration of 1000 milliseconds). Recordings were performed at baseline and after a 1-minute incubation with 50 μM compound 6 or vehicle (external solution containing 0.03% DMSO). The number of APs before and after compound application was counted in the same current injection process.

[0278] DRG neurons from the cultures described above showed marked hyperexcitability, as indicated by spontaneous action potential (AP) firing after current injection (200 pA–950 pA) (Figure 13A). Notably, application of compound 6 (50 μM) resulted in a significant decrease in the number of fired APs (Figure 13B). Furthermore, after injecting small amounts of current (0 pA–220 pA), the rheobase (i.e., the amount of current required to generate APs) significantly increased in the presence of compound 6 (Figure 13D), and the AP amplitude was significantly lower (Figure 13E). These findings confirm that compound 6 inhibits pruritus-sensitive DRG neurons in mice.

[0279] Example 10: Chemicals / Drugs and Cell Culture Drugs / Medicines: Roxapine, pregabalin, chloroquine, histamine, PGE2, serotonin, and bradykinin were purchased from Sigma-Aldrich. All stated concentrations refer to pure substances. Reagents and solvents for the synthesis of roxapine derivatives were obtained from Acros Organics (Hale, Belgium), Alfa Aesar GmbH & Co KG (Karlsruhe, Germany), BLDPharm Inc. (NuiNan, China), Fluorochem Ltd. (Hadfield, UK), Sigma-Aldrich (Munich, Germany), and TCI Europe NV (Zweindrecht, Belgium).

[0280] Cell Culture: HEK293 cells (hereinafter referred to as HEK-Slack cells; SB-HEK-KCa4.1; SB Drug Discovery (Lanarkshire, UK)) stably transfected with human Kcnt1 were maintained at 37°C in 5% CO2 in Dulbecco's modified Eagle medium-Glutamax containing 10% fetal bovine serum and 1% penicillin / streptomycin, supplemented with 0.6 mg / mL of G-418 (all Gibco / Thermo Fisher Scientific). Cells were passaged every 4-5 days from P11 to P35 depending on confluence.

[0281] Statistical analysis. Statistical analysis was performed using Prism 9 (GraphPad). Normal distribution data are presented as mean ± standard error (SEM), and nonparametric data are presented as median and interquartile range. The ROUT method with a Q of 1% (desired maximum false detection rate) was used to identify and remove potential outliers. The statistical tests and replication numbers for each analysis are shown in the legend of the figures or in the text. For all tests, a probability value of P < 0.05 was considered statistically significant. No statistical methods were used to pre-determine sample size.

[0282] Example 11: Antinociceptive effect of Slack activator (compound 6) in a mouse model of inflammatory pain. Accumulated evidence indicates that the sodium-activated potassium channel Slack is highly expressed in populations of nociceptive neuronals that detect painful stimuli, suggesting that Slack contributes to pain processing. Here, we evaluated whether systemic administration of compound 6, a novel Slack activator, affects persistent inflammatory pain. Interestingly, in a mouse model of inflammatory pain induced by intraplantar injection of total Freund's adjuvant (CFA), compound 6 significantly improved CFA-induced foot hypersensitivity. These data suggest that Slack activators may be effective treatments for various pain disorders in humans.

[0283] Animals: Experiments were conducted in C57BL / 6N mice (Charles River, Sulzfeld, Germany) aged 8 to 16 weeks, either male or female. Animals were housed in a 12-hour light / dark cycle with free access to food and water. All experiments were conducted in accordance with ARRIVE (Animal Research: Reporting on In Vivo Experiments) guidelines and approved and carried out according to the regulations of the local animal welfare authority (Darmstadt Administrative District (Regierungspraesidium Darmstadt) (Germany); approval number V54-19c20 / 15-FR / 2011). All behavioral studies were conducted during a daytime light cycle at room temperature (20°C to 24°C) by observers blinded to the treatment of the animals and / or their genotypes.

[0284] Behavioral Assessment: The mechanical sensitivity of the hind paw was assessed using a dynamic planter estasiometer (Ugo Basile (Italy, VA, Comerio)). The animals were placed on a wire mesh grid and acclimatized in the device chamber for 1 hour. A thin probe (0.5 mm in diameter) was applied from below to the surface of the sole of the foot, increasing the force from 0 g to 5 g within 10 seconds, and a constant force of 5 g was applied for another 10 seconds until a strong retraction occurred. The retraction latency of the foot was automatically recorded and calculated as the average of 6 to 8 measurements. After baseline measurements, heat-sterilized Mycobacterium tuberculosum (Mycobacterium) was measured in complete Freund's adjuvant (CFA, 85% paraffin oil and 15% monooleate mannide). Sigma-Aldrich (Darmstadt, Germany) 20 μL containing 1 mg / mL of tuberculosis fertilizer was injected into the plantar surface of the hind foot. Mechanical sensitivity was re-determined 24 hours after CFA injection. Immediately thereafter, the drug or vehicle (10% cyclodextrin in 0.9% NaCl) was administered intraperitoneally, and mechanical sensitivity was determined over 3 hours.

[0285] Statistical Analysis: Statistical analysis was performed using Prism 9 (GraphPad). The Kolmogorov-Smirnov test was used to assess the normal distribution of data within each group. Normally distributed data were analyzed using two-way repeated measures ANOVA and Sidac's post-hoc test, and expressed as mean ± standard error (SEM). In all statistical tests, a probability value of P < 0.05 was considered statistically significant.

[0286] To investigate the in vivo antinociceptive effect of compound 6, a Slack activator, the inventors used a mouse model of inflammatory pain induced by intraplantar injection of CFA into the hind paw. Intraplantar injection of CFA induces paw edema and hypersensitivity to mechanical stimuli. The results are shown in Figure 14. Compound 6 (10 mg / kg) or a vehicle was administered intraperitoneally 24 hours after CFA injection, and mechanical hypersensitivity in mice was assessed by testing the paw retraction latency using a dynamic planter estasiometer. As shown in Figure 1, CFA injection induced mechanical hypersensitivity, as indicated by a decrease in paw retraction latency 24 hours after CFA. Notably, at 0.5 and 1 hour after administration of compound 6, paw retraction latency was significantly increased compared to vehicles-treated animals, demonstrating an antinociceptive effect. These data suggest that persistent inflammatory pain may be ameliorated in vivo by compound 6.

[0287] Example 12: Topical application of Slack activator demonstrates efficacy in a mouse model of pruritus. Various disorders are accompanied by histamine-independent (non-histaminergic) itching, which is often resistant to currently available treatments. Recently, we demonstrated that pharmacological activation of Slack, a potassium channel highly expressed in pruritic sensory neurons, has therapeutic potential for treating itching. In previous studies, we showed that intraperitoneal or oral administration of novel Slack activators inhibited scratching behavior in multiple models of acute histamine-independent and chronic itching in mice without motor side effects. Here, we investigated whether topical application of Slack activating compounds exhibits antipruritic efficacy. Interestingly, topical application of two Slack activators, compound 6 and compound 21, inhibited scratching behavior in a mouse model of chloroquine-induced histamine-independent itching. These data suggest that topically administered Slack activators may be effective treatments for various pruritic disorders in humans.

[0288] Animals: Experiments were conducted in C57BL / 6N mice (Charles River, Sulzfeld, Germany) aged 8 to 16 weeks, either male or female. Animals were housed in a 12-hour light / dark cycle with free access to food and water. All experiments were conducted in accordance with the ARRIVE (Animal Research: Reporting on In Vivo Experiments) guidelines and approved and carried out according to the regulations of the local animal welfare authority (Darmstadt Administrative District (Regierungspraesidium Darmstadt) (Germany); approval number V54-19c20 / 15-FR / 2011). All behavioral studies were conducted during a daytime light cycle at room temperature (20°C to 24°C) by blinded observers regarding the treatment of the animals.

[0289] Drug formulations: Cream formulations of compound 6 and compound 21 were prepared by mixing 5% of each Slack activator with a hydrophilic cream (Basiscreme DAC).

[0290] Behavioral evaluation: Three to four days prior to the experiment, the hair on the outer dorsal side of the neck was shaved under short-term isoflurane anesthesia. On the day of the experiment, under short-term isoflurane anesthesia, compound 6 cream, compound 21 cream, or cream vehicle was applied topically to the shaved area of ​​the neck, and immediately thereafter the mice were acclimatized in a plexiglass cylinder (30 cm in diameter) for 30 minutes. After acclimatization, chloroquine (200 μg; dissolved in 20 μL of 0.9% physiological saline), an pruritic substance, was subcutaneously injected into the nape of the neck. The number of times the mice scratched the nape of their neck was evaluated over a 30-minute period by video recording.

[0291] Statistical Analysis: Statistical analysis was performed using Prism 9 (GraphPad). The Kolmogorov-Smirnov test was used to assess the normal distribution of data within each group. Normally distributed data were analyzed using one-way ANOVA and Dunnett's post-hoc test, and expressed as mean ± standard error (SEM). In all statistical tests, a probability value of P < 0.05 was considered statistically significant.

[0292] The inventors evaluated the antipruritic efficacy of compounds 6 and 21, which are Slack activators, in a histamine-independent pruritus model induced by the antimalarial drug chloroquine. Importantly, the inventors found that topical application of compound 6 (5%) and compound 21 (5%) 30 minutes prior to subcutaneous injection of chloroquine into the nape of the neck significantly improved scratching behavior compared to vehicle-treated animals (Figure 15). These data suggest that topical treatment with Slack activators improves histamine-independent itching.

[0293] References The references are as follows: 1 Carstens, E., Follansbee, T. & Iodi Carstens, M. The Challenge of Basic Itch Research. Acta Derm. Venereol. 100, adv00023 (2020). 2 Yosipovitch, G., Rosen, JD & Hashimoto, T. Itch: From mechanism to (novel) therapeutic approaches. J. Allergy Clin. Immunol. 142, 1375-1390 (2018). 3 Chen, XJ & Sun, YG Central circuit mechanisms of itch. Nat. Commun. 11, 3052 (2020). 4 Usoskin, D., et al. Unbiased classification of sensory neuron types by large-scale single-cell RNA sequencing. Nat. Neurosci. 18, 145-153 (2015). 5Meixiong, J. & Dong, X. Mas-Related G Protein-Coupled Receptors and the Biology of Itch Sensation. Annu. Rev. Genet. 51, 103-121 (2017). 6 Tsantoulas, C. & McMahon, S.B. Opening paths to novel analgesics: the role of potassium channels in chronic pain. Trends Neurosci. 37, 146-158 (2014). 7 Yuan, A., et al. The sodium-activated potassium channel is encoded by a member of the Slo gene family. Neuron 37, 765-773 (2003). 8 Budelli, G., et al. SLO2 Channels Are Inhibited by All Divalent Cations That Activate SLO1 K+ Channels. J. Biol. Chem. 291, 7347-7356 (2016). 9 Sharma, N., et al. The emergence of transcriptional identity in somatosensory neurons. Nature 577, 392-398 (2020). 10 Martinez-Espinosa, P.L., et al. Knockout of Slo2.2 enhances itch, abolishes KNa current, and increases action potential firing frequency in DRG neurons. Elife 4 (2015). 11Kupari, J., et al. Single cell transcriptomics of primate sensory neurons identifies cell types associated with chronic pain. Nat. Commun. 12, 1510 (2021). 12 Tavares-Ferreira, D., et al. Spatial transcriptomics of dorsal root ganglia identifies molecular signatures of human nociceptors. Sci. Transl. Med. 14, eabj8186 (2022). 13 Biton, B., et al. The antipsychotic drug Loxapine is an opener of the Na+-activated potassium channel Slack (slo2.2). J. Pharmacol. Exp. Ther. 340, 706-715 (2011). 14 Chakrabarti, A., et al. Loxapine for schizophrenia. Cochrane Database Syst. Rev., CD001943 (2007). 15 Schmiedl, S., et al. Loxapine for Treatment of Patients With Refractory, Chemotherapy-Induced Neuropathic Pain: A Prematurely Terminated Pilot Study Showing Efficacy But Limited Tolerability. Front Pharmacol 10, 838 (2019). 16Tavares-Ferreira, D., et al. Spatial transcriptomics of dorsal root ganglia identifies molecular signatures of human nociceptors. Sci. Transl. Med. 14, eabj8186 (2022). 17 Kupari, J., et al. Single cell transcriptomics of primate sensory neurons identifies cell types associated with chronic pain. Nat. Commun. 12, 1510 (2021). 18 Griffin, A.M., et al. Discovery of the First Orally Available, Selective KNa1.1 Inhibitor: In Vitro and In Vivo Activity of an Oxadiazole Series. ACS Med Chem Lett 12, 593-602 (2021). 19 Lu, R., et al. Slack channels expressed in sensory neurons control neuropathic pain in mice. J. Neurosci. 35, 1125-1135 (2015). 20 Avdeef, A. The rise of PAMPA. Expert Opin. Drug Metab. Toxicol. 1, 325-342 (2005). 21 Muller, J., Esso, K., Dargo, G., Konczol, A. & Balogh, G.T. Tuning the predictive capacity of the PAMPA-BBB model. Eur. J. Pharm. Sci. 79, 53-60 (2015). 22 Avdeef, A. Permeability - PAMPA. in Absorption and Drug Development: Solubility, Permeability, and Charge State, Second Edition 319-498 (Wiley, 2012). 23 Pagliara, A., et al. Molecular properties and pharmacokinetic behavior of cetirizine, a zwitterionic H1-receptor antagonist. J. Med. Chem. 41, 853-863 (1998). 24 Popovic, D., Nuss, P. & Vieta, E. Revisiting Loxapine: a systematic review. Ann Gen Psychiatry 14, 15 (2015). 25 Sjostedt, E., et al. An atlas of the protein-coding genes in the human, pig, and mouse brain. Science 367 (2020). 26 Karlsson, M., et al. A single-cell type transcriptomics map of human tissues. Sci Adv 7 (2021). 27 Lu, R., et al. Slack channels expressed in sensory neurons control neuropathic pain in mice. J. Neurosci. 35, 1125-1135 (2015). 28Popovic, D., Nuss, P. & Vieta, E. Revisiting Loxapine: a systematic review. Ann Gen Psychiatry 14, 15 (2015). 29 Liu, Q., et al. Sensory neuron-specific GPCR Mrgprs are itch receptors mediating chloroquine-induced pruritus. Cell 139, 1353-1365 (2009). 30 Liu, Q., et al. The distinct roles of two GPCRs, MrgprC11 and PAR2, in itch and hyperalgesia. Sci. Signal. 4, ra45 (2011). 31 Kitamura, A., Takata, R., Aizawa, S., Watanabe, H. & Wada, T. A murine model of atopic dermatitis can be generated by painting the dorsal skin with hapten twice 14 days apart. Sci Rep 8, 5988 (2018). 32 Li, M., et al. Topical vitamin D3 and low-calcemic analogs induce thymic stromal lymphopoietin in mouse keratinocytes and trigger an atopic dermatitis. Proc. Natl. Acad. Sci. U. S. A. 103, 11736-11741 (2006). 33Grundy, L., et al. Chronic linaclotide treatment reduces colitis-induced neuroplasticity and reverses persistent bladder dysfunction. JCI Insight 3, e121841 (2018).

Claims

1. A compound, or a salt, complex, diastereomer, enantiomer and / or tautomer thereof, wherein the compound is of formula I: 【Chemistry 1】 (In the formula, A is -H, -F, -Cl, -Br, -I, -CF ３ , -OCF ３ , -CF ２ H, and -OCF ２ One or two substituents independently selected from H, B is -F, -Cl, -Br, -I, -CF ３ , -OCF ３ , -CF ２ H, and -OCF ２ One or two substituents independently selected from H, X is selected from -S- and -O-, m is between 0 and 3. Y is, -H; Nitrile group; A 5- to 6-membered unsaturated heterocycle comprising three heteroatoms selected from O and N, preferably two or three of which are nitrogen, and the heterocycle being monosubstituted or disubstituted with =S or =O, preferably with =O; or, Linear-C １～３ an alkyl, wherein the linear-C １～３ alkyl's -(CH ２ )-, preferably one -(CH ２ )- is optionally mono-substituted with -OH, and the terminal -CH １～３ of the linear-C ３ alkyl is -OR １ or -SR １ and is preferably mono-substituted with -OR １ , and R １ is H, -(CH ２ )-COOH or -(CH ２ )-(CH ２ )-OH, a linear-C １～３ alkyl; or, - (CH ２ ) ｉ - CX １ R ２ (Here, X １ is, =NR ３ Or = O, R ３ is -H or -OH, i is either 0 or 1, R ２ ha-OR ４ or -NR ５ R ６ And R ４ is -H or -C １～２ It is alkyl, R ５ and R ６ (is independently selected from -H or -methyl); Selected from, Here, it is referred to as "Z", 【Chemistry 2】 is an aliphatic group containing one or two heterocycles, the heterocycles together containing two nitrogen atoms. Of the two nitrogen atoms, the first nitrogen atom is Z in formula I 【Transformation 3】 Linked to, the second nitrogen atom of the two nitrogen atoms is Z in formula I 【Chemistry 4】 Preferably, 【Transformation 5】 It connects to this, and here, Each of the two nitrogen atoms is a heteroatom in one of two linked four-membered heterocycles (each heterocycle has only one heteroatom), and the two interconnected four-membered heterocycles form a spirocycle, preferably, the four-membered heterocycle having the first nitrogen atom is linked to the four-membered heterocycle having the second nitrogen atom via a shared carbon atom, thereby forming a spirocycle having two heteroatoms which are two nitrogen atoms, or The first of the two nitrogen atoms is linked to a four-membered heterocycle having one heteroatom, the second of the two nitrogen atoms is the heteroatom in the four-membered heterocycle to which the first nitrogen atom is linked, and the first nitrogen atom is further substituted with methyl, or The two nitrogen atoms are heteroatoms in a 6- to 7-membered heterocycle having two heteroatoms, and the heterocycle is optionally substituted with methyl atoms, preferably monosubstituted. The compound, 【Transformation 6】 A compound, or a salt, complex, diastereomer, enantiomer, and / or tautomer thereof, that is not any of the compounds mentioned above.

2. The compound according to claim 1, wherein B is a substituent at carbon position 2 and carbon position 3, preferably a substituent -Cl at carbon position 2 and carbon position 3, and the carbon position numbers are as follows. 【Transformation 7】

3. A and B each consist of one substituent, preferably the compound is of formula II: 【Transformation 8】 The compound according to claim 1, having the following characteristics.

4. (i) A is selected from -H or -F, (ii) B is -Cl or -CF ３ Selected from, (iii) m is 0 to 2, more preferably m is 0 or 1, most preferably m is 1. (iv) Y is, Selected from the group consisting of, Z is A compound according to any one of claims 1 to 3, selected from the group consisting of the following.

5. The aforementioned compound is selected from the group consisting of compound 2 to compound 14. Preferably, the compound is selected from the group consisting of compound 6, compound 8, compound 9, compound 10, compound 11, compound 12, compound 18, compound 19, compound 20, and compound 21, and more preferably, the compound is selected from the group consisting of compound 6, compound 8, compound 9, and compound 10, according to any one of claims 1 to 4.

6. The compound is a potassium channel in cells, preferably a potassium channel Slack(K) Ｎａ A compound according to any one of claims 1 to 5, which has the activity of an activator as described in 1.1), and preferably activates potassium channels at an EC50 of less than 100 μM, preferably 0.5 μM to 80 μM.

7. The aforementioned compound, When administered to animals, it does not cross the blood-brain barrier (BBB), or crosses the BBB less effectively than reference compounds such as roxapine. A compound according to any one of claims 1 to 6, which does not bind to human dopamine receptors, or more preferably binds to human dopamine receptors less than a reference compound such as roxapine.

8. The compound according to any one of claims 1 to 7, wherein the compound is isolated and / or has a purity of at least 75%, optionally at least 90%, and optionally at least 95%.

9. A pharmaceutical composition comprising a compound according to any one of claims 1 to 8, or a salt, solvate, or ester thereof, and a pharmaceutically acceptable carrier or excipient.

10. The pharmaceutical composition according to claim 9, wherein the compound, or a salt, solvate, or ester thereof, is present in an amount of 0.1% w / w to 10% w / w, optionally 1% w / w to 10% w / w.

11. A compound or composition for the treatment of a disease in a subject, wherein the compound or composition contains an effective amount of the compound described in any one of claims 1 to 8, or a salt, complex, diastereomer, enantiomer and / or tautomer thereof, preferably the compound described in any one of claims 1 to 8, or a salt, solvate or ester thereof, wherein the compound is of formula I: 【Chemistry 9】 (In the formula, A is -H, -F, -Cl, -Br, -I, -CF ３ , -OCF ３ , -CF ２ H, and -OCF ２ One or two substituents independently selected from H, B is -F, -Cl, -Br, -I, -CF ３ , -OCF ３ , -CF ２ H, and -OCF ２ One or two substituents independently selected from H, X is selected from -S- and -O-, m is between 0 and 3. Y is, -H; Nitrile group; A 5- to 6-membered unsaturated heterocycle comprising three heteroatoms selected from O and N, preferably two or three of which are nitrogen, and the heterocycle being monosubstituted or disubstituted with =S or =O, preferably with =O; or, Linear-C １～３ Alkyl, and the linear-C １～３ Alkyl - (CH ２ ) - preferably one - (CH ２ )- is arbitrarily monosubstituted with -OH, and the linear-chain -C １～３ alkyl terminal -CH ３ However, -OR １ Or -SR １ And preferably -OR １ It is monosubstituted with R １ H, -(CH ２ )-COOH or-(CH ２ )-(CH ２ )-OH, a straight chain -C １～３ alkyl; or, - (CH ２ ) ｉ - CX １ R ２ (Here, X １ is, =NR ３ Or = O, R ３ is -H or -OH, i is either 0 or 1, R ２ ha-OR ４ or -NR ５ R ６ And R ４ is -H or -C １～２ It is alkyl, R ５ and R ６ (is independently selected from -H or -methyl); Selected from, Here, it is referred to as "Z", 【Chemistry 10】 is an aliphatic group containing one or two heterocycles, the heterocycles together containing two nitrogen atoms. Of the two nitrogen atoms, the first nitrogen atom is Z in formula I 【Chemistry 11】 Linked to, the second nitrogen atom of the two nitrogen atoms is Z in formula I 【Chemistry 12】 Preferably, 【Chemistry 13】 It connects to this, and here, Each of the two nitrogen atoms is a heteroatom in one of two linked four-membered heterocycles (each heterocycle has only one heteroatom), and the two interconnected four-membered heterocycles form a spirocycle, preferably, the four-membered heterocycle having the first nitrogen atom is linked to the four-membered heterocycle having the second nitrogen atom via a shared carbon atom, thereby forming a spirocycle having two heteroatoms which are two nitrogen atoms, or The first of the two nitrogen atoms is linked to a four-membered heterocycle having one heteroatom, the second of the two nitrogen atoms is the heteroatom in the four-membered heterocycle to which the first nitrogen atom is linked, and the first nitrogen atom is further substituted with methyl, or The two nitrogen atoms are heteroatoms in a 6- to 7-membered heterocycle having two heteroatoms, and the heterocycle is optionally substituted with methyl atoms, preferably monosubstituted. Preferably, the composition is the pharmaceutical composition according to claim 9 or 10, wherein the disease is treatable by activating potassium channels in cells associated with the pathology of the disease, and the treatment comprises administering the compound or composition to a subject in need of treatment.

12. The compound or composition according to claim 11, wherein the disease is a pruritic disorder such as acute or chronic itching, and optionally the pruritic disorder is related to a secondary pathological condition such as one selected from the following: Dermatological disorders such as xerosis or xerosis (dry skin), dermatitis or eczema (e.g., atopic dermatitis), psoriasis (e.g., psoriasis vulgaris), prurigo (e.g., nodular prurigo), urticaria (e.g., chronic idiopathic urticaria), connective tissue disorders (e.g., dermatomyositis), and post-burn pruritus; Renal impairment (e.g., chronic kidney disease, chronic renal failure, or end-stage renal disease), dialysis (e.g., hemodialysis), uremic pruritus; Hepatobiliary disorders (e.g., cholestasis, primary biliary cholangitis, primary sclerosing cholangitis, secondary sclerosing cholangitis, hepatitis, toxic liver disease, chronic liver disease, or cirrhosis), cholestatic pruritus; Endocrine disorders (e.g., hyperthyroidism or diabetes mellitus); Metabolic disorders (e.g., iron deficiency or iron overload); Benign or malignant neoplasms (e.g., solid tumors, carcinomas, or hematological malignancies (e.g., Hodgkin lymphoma, non-Hodgkin lymphoma, myeloproliferative disorders, or polycythemia vera)); Infectious diseases (e.g., viral infections such as herpes simplex, herpes zoster, chickenpox, human immunodeficiency virus (HIV), and infections associated with hepatitis; bacterial infections or parasitic diseases); Neurological disorders (e.g., neurodegenerative diseases, multiple sclerosis, brain tumors, postherpetic neuralgia, small fiber neuropathy, brachioradialis pruritus, or dorsal paresthesia), neurogenic itching, neurogenic pruritus; Mental illnesses (e.g., depression, obsessive-compulsive disorder, delusional disorder, eating disorders, or anxiety disorders); Drug-induced pruritus (e.g., caused by opioids, antibiotics, antimalarial drugs, ACE inhibitors, angiotensin receptor blockers, antiarrhythmics, antidepressants, antidiabetic drugs, antihypertensive drugs, anticonvulsants, anti-inflammatory drugs, beta-blockers, bronchodilators, calcium channel blockers, diuretics, hormones, immunosuppressants, antilipids, nerve blockers, plasma expanders, tranquilizers, or uric acid-lowering drugs); Age-related pruritus; Pregnancy pruritus; and / or, Chronic idiopathic pruritus.

13. The aforementioned disease is pain or a pain-related adverse sensation in the subject, preferably the pain is Neuropathic pain induced by traumatic nerve injury, cancer and cancer treatment (e.g., chemotherapy), neurological disorders (e.g., multiple sclerosis), neurodegenerative diseases (e.g., Parkinson's disease), trigeminal neuralgia, diabetic peripheral neuropathy, stroke, herpes zoster, HIV, leprosy, Guillain-Barré syndrome, vascular diseases, vascular malformations, and autoimmune diseases. acute postoperative pain, Inflammatory pain, rheumatoid arthritis, osteoarthritis, and / or Pain dysregulation, A compound or composition according to claim 11 or 12, selected from the above.

14. The compound or composition according to any one of claims 11 to 13, wherein the administration includes intravenous administration, intraperitoneal administration, subcutaneous administration, intramuscular administration, intrathecal administration, epidural administration, local administration, oral administration, gastric administration and / or rectal administration.

15. A method for synthesizing a compound according to any one of claims 1 to 8, preferably comprising a synthesis step of synthesizing a lactam core, preferably having the following structure: 【Chemistry 14】 A method comprising (wherein A, X, and B are defined in any one of claims 1 to 8).

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