CN113912601A

CN113912601A - Novel imidazo [1,2-a ] pyridine derivatives, preparation method and medical application thereof

Info

Publication number: CN113912601A
Application number: CN202110779116.XA
Authority: CN
Inventors: 李文明; 刘宁; 刘彪; 刘浩淼; 余健; 邹昊; 祝伟; 李正涛
Original assignee: Shanghai Tuojie Biomedical Technology Co ltd
Current assignee: Shanghai Tuojie Biomedical Technology Co ltd
Priority date: 2020-07-10
Filing date: 2021-07-09
Publication date: 2022-01-11
Anticipated expiration: 2041-07-09
Also published as: CN113912601B

Abstract

The present disclosure relates to a novel class of imidazo [1,2-a]Pyridine derivatives, preparation method and medical application thereof. Specifically, the substituent of the derivative comprises a fused ring structure, the structural formula of the fused ring structure is shown as the following, wherein R is₁、R₂、R₃、R₄、R₅X and Y are defined as in the specification.

Description

Novel imidazo [1,2-a ] pyridine derivatives, preparation method and medical application thereof

Technical Field

The disclosure relates to the field of medicine, in particular to a novel imidazo [1,2-a ] pyridine derivative, a preparation method thereof and application thereof in medicine.

Background

The P2X receptor is a family of cation permeable, ligand-gated ion channels that open in response to binding of extracellular adenosine 5' -triphosphate (ATP). They belong to a larger family of receptors, called purinergic receptors. The P2X receptor is present in a variety of organisms including humans, mice, rats, rabbits, chickens, zebrafish, bullfrogs, flukes and amoebas. Seven independent genes encoding the P2X subunit have been identified and designated P2X1 through P2X 7. The different subunits exhibit different sensitivities to purinergic agonists and antagonists.

The P2X3 receptor has 4 ATP binding sites on a single subunit, consisting of 2 transmembrane domains, N-and C-termini located intracellularly, conserved sequences located in extracellular loop structures. High expression of the P2X3 receptor was found in specific medium and small diameter neurons that were associated with injury information. At the same time, the P2X3 receptor is also involved in the transmission of some non-nociceptive sensations. It has been demonstrated that the P2X3 receptor is involved in bladder sensory function, is a key receptor-mediated bladder sensory signal, and is expressed in the mucosal tissues of the bladder, which are rich in sensory nerve fibers. The expression of P2X3 is also found in sensory nerve fibers of the pharyngeal mucosa, and is involved in the conduction and formation of taste sensation.

When an organism is injured or damaged by nerves, a large amount of ATP is released to activate a presynaptic membrane P2X3 receptor, so that a large amount of Ca2+ flows in, the intracellular calcium concentration is increased to activate Protein Kinase A (PKA) and Protein Kinase C (PKC), the PKA and the PKC are phosphorylated, simultaneously, the release of glutamate is promoted, the NMDA receptor is further activated, the excitatory postsynaptic current is generated, and the central sensitization is caused. Many studies have shown that upregulation of P2X3 receptor expression can lead to the development of pain sensitivity and is involved in pain signaling.

MK-7264 is a P2X3 receptor activity antagonist with IC50 values of-30 nM and 100 nM for human homologous recombination hP2X3 and hP2X2/3, respectively, and its use for chronic cough treatment has progressed to stage III.

Disclosure of Invention

The present disclosure provides compounds of formula (I), or a pharmaceutically acceptable salt thereof, or an isomer thereof,

wherein R is₁Selected from hydrogen, deuterium, C optionally substituted by halogen or deuterium₁-C₆Alkyl, halogen, C optionally substituted by halogen or deuterium₁-C₆Alkoxy radical, C₁-C₆A hydroxyalkyl group;

R₂selected from hydrogen, deuterium, C optionally substituted by halogen or deuterium₁-C₆Alkyl, halogen, C optionally substituted by halogen or deuterium₁-C₆Alkoxy radical, C₁-C₆A hydroxyalkyl group;

R₃and R₄Independently selected from hydrogen, halogen, C optionally substituted by halogen₁-C₄Alkyl, or R₃And R₄Together with the carbon atom to which they are both attached form C optionally substituted by halogen₃-C₆Cycloalkylene radical, or C on adjacent carbon atoms, optionally substituted by halogen₁-C₄R of alkyl₃And R₄Connecting to form a ring;

R₅selected from the group consisting of: optionally halogen-or cyano-substituted C₁-C₆Alkyl, C optionally substituted by halogen or cyano₃-C₆Cycloalkyl, heterocyclyl optionally substituted by halogen or cyano, C optionally substituted by halogen or cyano₁-C₆Alkoxy, optionally alkyl-substituted amino;

y is selected from 3-18 membered aromatic or non-aromatic heterocyclyl, preferably 7-15 membered aromatic or non-aromatic heterocyclyl, more preferably 9-12 membered bicyclic or tricyclic heterocyclyl, said heterocyclyl being optionally substituted;

x is selected from C, N and O.

In some embodiments, Y is selected from 3-18 membered aromatic or non-aromatic heterocyclyl, preferably 7-15 membered aromatic or non-aromatic heterocyclyl, more preferably 9-12 membered bicyclic or tricyclic heterocyclyl, said heterocyclyl being optionally substituted with alkylamide, halogen, oxygen, alkyl, alkoxy.

In some embodiments, Y is selected from 3-18 membered aromatic or non-aromatic heterocyclyl, preferably 7-15 membered aromatic or non-aromatic heterocyclyl, more preferably 9-12 membered bicyclic or tricyclic heterocyclyl, said heterocyclyl being optionally substituted with methylamide, fluoro, chloro, methyl, ethyl, oxygen.

In some embodiments of the present invention, the substrate is,

y is selected from

J₁Selected from N, C, O, S, J₂Selected from N, C, ring a is a cyclic compound containing-NH-C (═ O) -or-NH-S (═ O)₂-a heterocyclic group of (a);

R₆independently selected from halogen, C₁-C₄An alkyl group;

R₇independently selected from halogen, C₁-C₄Alkyl radical, C₃-C₆Cycloalkylene radicals, or adjacent R₇Are connected into a ring, and the ring optionally has halogen or C₁-C₄Alkyl substitution;

m is an integer from 0 to 4;

n is an integer from 0 to 3;

a is selected from an integer of 1 to 4.

In some embodiments, in the compounds of formula (I),

R₁selected from hydrogen, deuterium, halogen, C optionally substituted by halogen or deuterium₁-C₆An alkyl group, a carboxyl group,

R₂selected from the group consisting of hydrogen, deuterium, halogen,

y is selected from

R₆independently selected from halogen, C₁-C₄An alkyl group;

x is selected from C, N and O;

m is an integer from 0 to 4;

n is an integer from 0 to 3;

a is selected from an integer of 1 to 4.

In some embodiments, in the compounds of formula (I),

R₂selected from the group consisting of hydrogen, deuterium, halogen,

R₅is selected from C₁-C₆Alkyl or C₁-C₆An alkoxy group,

y is selected from

R₆is independently selected fromHalogen, C₁-C₄An alkyl group;

x is selected from C, N and O;

m is an integer from 0 to 4;

n is an integer from 0 to 3;

a is selected from an integer of 1 to 4.

In some embodiments, in the compounds of formula (I),

R₂selected from the group consisting of hydrogen, deuterium, halogen,

R₃and R₄Selected from hydrogen, halogen, or R₃And R₄Together with the carbon atom to which they are both attached form C optionally substituted by halogen₃-C₆A cyclic-alkylene group,

R₅is selected from C₁-C₆Alkyl or C₁-C₆An alkoxy group,

x is selected from the group consisting of C, O,

y is selected from

R₆independently selected from halogen, C₁-C₄An alkyl group;

m is an integer from 0 to 4;

n is an integer from 0 to 3;

a is selected from an integer of 1 to 4.

In some embodiments, in the compounds of formula (I),

R₁selected from hydrogen, deuterium, halogen, C optionally substituted by halogen or deuterium₁-C₆An alkyl group;

R₂selected from hydrogen, deuterium, halogen;

R₃and R₄Selected from hydrogen, halogen, or R₃And R₄Together with the carbon atom to which they are both attached form C optionally substituted by halogen₃-C₆A cycloalkylene group;

R₅is selected from C₁-C₆Alkyl or C₁-C₆An alkoxy group;

x is selected from C, O;

y is selected from

J₁Selected from N, C, O, S, J₂Selected from N, C, ring a is a cyclic compound containing-NH-C (═ O) -or-NH-S (═ O)₂-5-13 membered heterocyclyl;

R₆independently selected from halogen, C₁-C₄An alkyl group;

m is an integer from 0 to 4;

n is an integer from 0 to 3;

a is selected from an integer of 1 to 4.

In some embodiments, in the compounds of formula (I),

R₁selected from hydrogen, deuterium, fluoro, chloro, or optionally deuterium, fluoro substituted methyl;

R₂selected from hydrogen, fluorine, chlorine;

R₃and R₄Selected from hydrogen, fluorine, or R₃And R₄Together with the carbon atom to which they are both attached form a cyclopropylene group optionally substituted with halogen;

R₅is selected from C₁-C₆Alkyl or C₁-C₆An alkoxy group;

x is selected from C, O;

y is selected from

R₆independently selected from halogen, C₁-C₄An alkyl group;

m is an integer from 0 to 4;

n is an integer from 0 to 3;

a is selected from an integer of 1 to 4.

In some embodiments, in the compounds of formula (I),

y is selected from

R₆Independently selected from halogen, C₁-C₄An alkyl group;

m is an integer from 0 to 4;

n is an integer from 0 to 3;

R₁selected from hydrogen, deuterium, C optionally substituted by halogen or deuterium₁-C₆Alkyl, halogen, C optionally substituted by halogen or deuterium₁-C₆Alkoxy radical, C₁-C₆A hydroxyalkyl group;

x is selected from C, N and O.

In some embodiments, in the compounds of formula (I),

y is selected from

Or one or more of F in the foregoing structures is substituted with Cl or H.

In some embodiments, in the compounds of formula (I),

y is selected from

In some embodiments, in the compounds of formula (I),

R₂selected from hydrogen, fluorine, chlorine;

R₅is selected from C₁-C₆Alkyl or C₁-C₆An alkoxy group;

x is selected from C, O;

y is selected from

R₆Independently selected from halogen, C₁-C₄An alkyl group;

m is an integer from 0 to 4;

n is an integer from 0 to 3.

In some embodiments, in the compounds of formula (I),

R₂selected from hydrogen, fluorine, chlorine;

R₅is selected from C₁-C₆Alkyl or C₁-C₆An alkoxy group;

x is selected from C, O;

y is selected from

Or one or more of F in the foregoing structures is substituted with Cl or H.

In some embodiments, in the compounds of formula (I),

R₂selected from hydrogen, fluorine, chlorine;

R₅is selected from C₁-C₆Alkyl or C₁-C₆An alkoxy group;

x is selected from C, O;

y is selected from

The present disclosure also provides compounds of formula (I-1), or a pharmaceutically acceptable salt thereof, or an isomer thereof,

y is selected from

R₆independently selected from halogen, C₁-C₄An alkyl group;

m is an integer from 0 to 4;

n is an integer from 0 to 3;

a is selected from an integer of 1 to 4.

In some embodiments, in the compounds of formula (I-1),

y is selected from

J₁Selected from N, C, O, S, J₂Selected from N, C, ring a is a cyclic compound containing-NH-C (═ O) -or-NH-S (═ O)₂-a 5-13 membered heterocyclic group of (a),

R₆independently selected from halogen, C₁-C₄An alkyl group;

m is an integer from 0 to 4;

n is an integer from 0 to 3;

a is selected from an integer of 1 to 4.

In some embodiments, in the compounds of formula (I-1), Y is selected from

R₆Independently selected from halogen, C₁-C₄An alkyl group;

m is an integer from 0 to 4;

n is an integer from 0 to 3.

In some embodiments, in the compounds of formula (I-1), Y is selected from

In some embodiments, in the compounds of formula (I-1), Y is selected from

Or one or more of F in the foregoing structures is substituted with Cl or H.

The present disclosure also provides a compound as shown below, or a pharmaceutically acceptable salt thereof, or an isomer thereof,

the present disclosure provides a process for preparing a compound of formula (I) or a pharmaceutically acceptable salt, or isomer thereof, comprising the steps of:

carrying out cyclization reaction on the compound shown in the formula (I-a) and the compounds shown in the formula (I-b) and the formula (I-c) under the condition of a catalyst to obtain the compound shown in the formula (I);

the catalyst is selected from palladium/carbon, raney nickel, tetra-triphenylphosphine palladium, palladium dichloride, palladium acetate, [1,1' -bis (diphenylphosphino) ferrocene ] palladium dichloride, 1' -bis (dibenzylphosphine) dichloropentairon palladium, tris (dibenzylideneacetone) dipalladium or 2-dicyclohexylphosphine-2 ',6' -dimethoxybiphenyl, [1,1' -bis (di-tert-butylphosphino) ferrocene ] dichloropalladium (II), cuprous iodide, cuprous bromide, cuprous chloride and copper trifluoromethanesulfonate;

R₁、R₂、R₃、R₄、R₅x, Y are as defined for compounds of formula (I).

The present disclosure provides a process for preparing a compound of formula (I-1) or a pharmaceutically acceptable salt thereof, or an isomer thereof, comprising the steps of:

carrying out cyclization reaction on the compound shown in the formula (I-d) and the compounds shown in the formula (I-e) and the formula (I-c) under the condition of a catalyst to obtain a compound shown in the formula (I-1);

y is as defined for the compound of formula (I-1).

carrying out cyclization reaction on the compound shown in the formula (I-d) and the compound shown in the formula (I-a) to obtain a compound shown in the formula (I-e); halogenating the compound shown in the formula (I-e) to obtain a compound shown in a formula (I-f); carrying out C-C coupling reaction on the compound of the formula (I-f) under an alkaline condition to obtain a compound shown in the formula (I);

the catalyst is selected from palladium/carbon, raney nickel, tetrakis-triphenylphosphine palladium, palladium dichloride, palladium acetate, [1,1' -bis (diphenylphosphino) ferrocene ] palladium dichloride, 1' -bis (dibenzylideophosphonium) dichlorodipentaerythritol palladium, tris (dibenzylideneacetone) dipalladium or 2-dicyclohexylphosphine-2 ',6' -dimethoxybiphenyl, [1,1' -bis (di-tert-butylphosphino) ferrocene ] dichloropalladium (II);

R₁、R₂、R₃、R₄、R₅x, Y are as defined for compounds of formula (I-1);

z' and Z are selected from the group consisting of halogen, sulfonyl and sulfinyl.

carrying out cyclization reaction on the compound shown in the formula (I-a) and the compound shown in the formula (I-g) to obtain the compound shown in the formula (I);

R₁、R₂、R₃、R₄、R₅x, Y are as defined for compounds of formula (I);

z is selected from the group consisting of halogen, sulfonyl and sulfinyl.

carrying out cyclization reaction on the compound shown in the formula (I-a) and the compound shown in the formula (I-h) to obtain a compound shown in the formula (I-I); carrying out addition reduction and dehydroxylation on the compound shown in the formula (I-I) and the compound shown in the formula (I-j) to obtain a compound shown in the formula (I);

z is selected from the group consisting of halogen, sulfonyl and sulfinyl.

The present disclosure also provides a process for preparing the compound or a pharmaceutically acceptable salt thereof, or an isomer thereof. In particular, the preparation is carried out by the method of the examples.

The present disclosure also relates to a pharmaceutical composition comprising a compound described in the present disclosure or a pharmaceutically acceptable salt thereof, or an isomer thereof. Typically, the composition further comprises at least one pharmaceutically acceptable carrier, diluent or excipient.

The disclosure also provides the use of the compound or a pharmaceutically acceptable salt thereof, or an isomer thereof, or a pharmaceutical composition comprising the same, in the preparation of a medicament for treating a disease associated with P2X3 activity. In some embodiments, the disease associated with P2X3 activity refers to a disease associated with P2X3 hyperactivity. The compounds of the present disclosure are highly selective for P2X3, which can avoid taste loss. In some embodiments, the compounds of the present disclosure antagonize P2X3 cognate receptors more than 20-fold stronger than antagonizing P2X2/3 heteromeric receptors. In some embodiments, the compounds of the present disclosure antagonize P2X3 cognate receptors more than 30-fold stronger than antagonizing P2X2/3 heteromeric receptors. In some embodiments, the compounds of the present disclosure antagonize P2X3 cognate receptors more than 50-fold stronger than antagonizing P2X2/3 heteromeric receptors. In some embodiments, the compounds of the present disclosure antagonize P2X3 cognate receptors more than 100-fold stronger than antagonizing P2X2/3 heteromeric receptors.

In some embodiments, the compounds of the present disclosure or pharmaceutically acceptable salts thereof, or isomers thereof, or pharmaceutical compositions comprising the same, are useful for the treatment of pain, urinary tract disorders, cough, and the like. Pain may be, for example, chronic pain, neuropathic pain, acute pain, back pain, cancer pain, pain caused by rheumatoid arthritis, migraine, and visceral pain. Urinary tract disorders such as overactive bladder (also known as urinary incontinence), pelvic hypersensitivity, and urethritis.

In some embodiments, a compound of the present disclosure or a pharmaceutically acceptable salt thereof, or an isomer thereof, or a pharmaceutical composition comprising the same, is useful for treating gastrointestinal disorders, including, for example, constipation and functional gastrointestinal disorders (e.g., irritable bowel syndrome or functional dyspepsia); can be used for treating cancer; can be used for the treatment of cardiovascular disorders or for cardioprotection following myocardial infarction; can be used as an immunomodulator, particularly in the treatment of autoimmune diseases (e.g. arthritis), in skin transplantation, organ transplantation, or similar surgical needs, in collagen diseases, in allergies, or as an anti-tumor or anti-viral agent; can be used for treating multiple sclerosis, Parkinson's disease, and Huntington's chorea; can be used for treating depression, anxiety, stress-related disorders (e.g., post-traumatic stress disorder, panic disorder, social phobia, or obsessive compulsive disorder), premature ejaculation, psychosis, traumatic brain injury, stroke, Alzheimer's disease, spinal injury, drug addiction (e.g., treatment of alcohol, nicotine, opioid, or other drug abuse), or sympathetic nervous system disorders (e.g., hypertension); can be used for treating diarrhea; can be used for treating pulmonary disorders such as, for example, asthma, cough, or pulmonary edema.

The compounds of the present disclosure or pharmaceutically acceptable salts thereof, or isomers thereof, can be formulated in a dosage form suitable for oral, buccal, vaginal, rectal, via inhalation, via insufflation, intranasal, sublingual, topical, or parenteral (e.g., intramuscular, subcutaneous, intraperitoneal, intrathoracic, intravenous, epidural, intrathecal, intracerebroventricular, or by injection into the joints) administration.

The term "treatment" refers to the administration of a pharmaceutical composition for prophylactic and/or therapeutic purposes. By "preventing a disease" is meant prophylactically treating a subject who is not yet ill, but is susceptible to, or at risk for, a particular disease. By "treating a disease" is meant treating a patient already suffering from the disease to improve or stabilize the patient's condition.

Any isotopically-labeled (or "radiolabeled") derivative of a compound described in the present disclosure or a pharmaceutically acceptable salt thereof, or an isomer thereof, is encompassed by the present disclosure. Such derivatives are where one or more atoms are replaced by an atom having an atomic mass or mass number different from the atomic mass or mass number usually found in nature. Examples of radionuclides that may be incorporated include²H (also written as "D", i.e. deuterium),³H (also written as "T", i.e. tritium),¹¹C、¹³C、¹⁴C、¹³N、¹⁵N、¹⁵O、¹⁷O、¹⁸O、¹⁸F、³⁶Cl、⁸²Br、⁷⁵Br、⁷⁶Br、⁷⁷Br、¹²³I、¹²⁴I、¹²⁵I、³¹P、³²P、³⁵S, and¹³¹I. the radionuclide used will depend on the particular application of the radiolabeled derivative. For example, for in vitro receptor labeling and competition assays,³h or¹⁴C is often useful. For the application of radiological imaging,¹¹c or¹⁸F is often useful. In some embodiments, the radionuclide is³H. In some embodiments, the radionuclide is¹⁴C. In some embodiments, the radionuclide is¹¹C. And in some embodiments the radionuclide is¹⁸F。

Unless stated to the contrary, the following terms used in the specification and claims have the following meanings.

"halogen" refers to fluorine, chlorine, bromine and iodine.

"alkyl" means a straight or branched chain alkyl group such as methyl, ethyl, n-propyl, isopropyl, and the like.

"alkoxy" refers to an alkyloxy group such as methoxy, ethoxy, and the like.

"heterocyclyl" refers to a stable 3-to 18-membered aromatic or non-aromatic ring group containing 1 to 12 carbon atoms and 1 to 6 heteroatoms selected from nitrogen, oxygen, and sulfur. Unless otherwise specified in the specification, a heterocyclic group may be a monocyclic, bicyclic, tricyclic or tetracyclic ring system, which may include a spiro or bridged ring system; and the nitrogen, carbon or sulfur atoms in the heterocyclic group may be optionally oxidized; the nitrogen atoms may optionally be quaternized; and heterocyclic groups may be partially or fully saturated. "comprises-NH-C (═ O) -or-NH-S (═ O)₂Heterocyclyl "to" is not to be interrupted by "-NH-C (═ O) -" and "-NH-S (═ O) on the cyclic structure₂The other moiety, besides the "moiety, optionally contains a heteroatom.

"cycloalkyl" refers to a stable non-aromatic monocyclic or polycyclic hydrocarbon radical consisting solely of carbon and hydrogen atoms, which may contain a spiro or bridged ring system, having 3 to 15 carbon atoms, 3 to 10 carbon atoms, or 5 to 7 carbon atoms, and which is saturated or unsaturated and is linked to the rest of the molecule by a single bond. Monocyclic cycloalkyl groups include non-bridged cycloalkyl groups such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, and cyclooctyl.

"Cyclohydrocarbylene" refers to a divalent cyclic hydrocarbyl group derived from a cyclic hydrocarbyl group. For example,

and the like.

"optional" or "optionally" means that the subsequently described event or circumstance may, but need not, occur, and that the description includes instances where the event or circumstance occurs or does not. For example "C optionally substituted by halogen or cyano₁-C₆Alkyl "means that halogen or cyano may, but need not, be present, and the description includes the case where alkyl is substituted with halogen or cyano and the case where alkyl is not substituted with halogen and cyano.

Detailed Description

The structure of the compounds is determined by Nuclear Magnetic Resonance (NMR) or/and Mass Spectrometry (MS). NMR shift (. delta.) of 10^-6The units in (ppm) are given. NMR was measured using a Bruker AVANCE-400 NMR spectrometer using deuterated dimethyl sulfoxide (DMSO-d)₆) Deuterated chloroform (CDCl)₃) And deuterated methanol (CD)₃OD), internal standard Tetramethylsilane (TMS).

HPLC measurements were performed using Waters ACQUITY ultra high Performance LC, Shimadzu LC-20A systems, Shimadzu LC-2010HT series, or Agilent 1200 LC HPLC (ACQUITY UPLC BEH C181.7 μm 2.1X 50mm column, Ultimate XB-C183.0X 150mm column, or XTate C182.1X 30mm column).

MS was determined using a Waters SQD2 mass spectrometer, scanning in positive/negative ion mode with a mass scan range of 100-1200.

Chiral HPLC analytical determination used Chiralpak IC-3100X 4.6mm I.D., 3 μm, Chiralpak AD-3150X 4.6mm I.D., 3 μm, Chiralpak AD-350X 4.6mm I.D., 3 μm, Chiralpak AS-3150X 4.6mm I.D., 3 μm, Chiralpak AS-3100X 4.6mm I.D., 3 μm, Chiralpak OD-3150X 4.6mm I.D., 3 μm, Chiralcel OD-3100X 4.6mm I.D., 3 μm, Chiralcel OJ-H150X 4.6mm I.D., 5 μm, Chiralcel OJ-3150X 4.6mm I.D., 3 μm chromatography columns.

The thin layer chromatography silica gel plate adopts HSGF254 of tobacco yellow sea or GF254 of Qingdao, the specification of the silica gel plate used by Thin Layer Chromatography (TLC) is 0.15 mm-0.2 mm, and the specification of the thin layer chromatography separation and purification product is 0.4 mm-0.5 mm.

The column chromatography generally uses 100-200 mesh, 200-300 mesh or 300-400 mesh silica gel of Tibet Huanghai silica gel as a carrier.

The chiral preparative column used DAICEL CHIRALPAK IC (250 mm. times.30 mm,10 μm) or Phenomenex-Amylose-1(250 mm. times.30 mm,5 μm).

Known starting materials of the present disclosure may be synthesized using or according to methods known in the art, or may be purchased from companies such as ABCR GmbH & co.kg, Acros Organics, Aldrich Chemical Company, nephelo Chemical science and technology (Accela ChemBio Inc), dare chemicals, and the like.

In the examples, the reaction can be carried out in an argon atmosphere or a nitrogen atmosphere, unless otherwise specified.

Argon atmosphere or nitrogen atmosphere means that the reaction flask is connected with an argon or nitrogen balloon having a volume of about 1L.

The hydrogen atmosphere refers to a reaction flask connected with a hydrogen balloon with the volume of about 1L.

The pressure hydrogenation reaction used a hydrogenation apparatus of Parr 3916EKX type and a hydrogen generator of Qinglan QL-500 type or a hydrogenation apparatus of HC2-SS type.

The hydrogenation reaction was usually evacuated and charged with hydrogen and repeated 3 times.

The microwave reaction was carried out using a CEM Discover-S908860 type microwave reactor.

In the examples, the solution means an aqueous solution unless otherwise specified.

In the examples, the reaction temperature is, unless otherwise specified, from 20 ℃ to 30 ℃ at room temperature.

The monitoring of the progress of the reaction in the examples employed Thin Layer Chromatography (TLC), a developing solvent used for the reaction, a system of eluents for column chromatography used for purifying compounds and a developing solvent system for thin layer chromatography including: a: dichloromethane/methanol system, B: n-hexane/ethyl acetate system, C: petroleum ether/ethyl acetate system, D: the volume ratio of petroleum ether/ethyl acetate/methanol and solvent is regulated according to the different polarity of the compound, and a small amount of basic or acidic reagents such as triethylamine, acetic acid and the like can be added for regulation.

Example 1

Methyl (S) -2- ((7-methyl-2- (1-oxoisoindolin-5-yl) imidazo [1,2-a ] pyridin-3-yl) methyl) morpholine-4-carboxylate (1)

The method comprises the following steps: 5-Vinylisoindoline-1-one (1a)

Adding 5-bromo-2, 3-dihydro-1H-isoindol-1-one (500.0mg,2.36mmol) and 4,4,5, 5-tetramethyl-2-vinyl-1, 3, 2-dioxaborolane under the protection of nitrogen(480. mu.L, 2.83mmol) of a mixed solution of 1, 4-dioxane (10mL) and water (2.5mL) to which Cs was added₂CO₃(1536.6mg,4.72mmol) and Pd (dppf) Cl₂(174.9mg,0.24mmol), followed by stirring at 90 ℃ for 12 hours. TLC showed the reaction was complete, the reaction mixture was concentrated under reduced pressure and the residue was purified by silica gel column chromatography (ethyl acetate/methanol 0-5%) to give the title compound 1a (330.0mg, 79.1%).

¹H NMR(400MHz,CD₃OD)δ7.73(d,J＝8.0Hz,1H),7.63(s,1H),7.57(d,J＝8.0Hz,1H),6.85(dd,J＝11.2,17.6Hz,1H),5.94(d,J＝17.6Hz,1H),5.38(d,J＝11.2Hz,1H),4.44(s,2H).

Step two: 1-oxoisoindoline-5-carbaldehyde (1b)

To a mixed solution of 1a (150.0mg,0.85mmol) of 1, 4-dioxane (6mL) and water (1.2mL) was added osmium tetroxide (32.3mg,0.13mmol), and the mixture was stirred at 15 ℃ for 10 minutes. Sodium periodate (997.0mg,3.39mmol) was then added and stirred at 15 ℃ for 1 hour. TLC showed the reaction was complete. The reaction mixture was poured into water and extracted with ethyl acetate (5X 30mL), and the combined organic phases were taken over Na₂SO₄After drying, filtration and concentration the title compound 1b (120.0mg, 79.0%) was obtained.

¹H NMR(400MHz,DMSO-d₆)δ10.14(s,1H),8.89(br s,1H),8.10(s,1H),8.02(d,J＝7.6Hz,1H),7.89-7.84(m,1H),7.87(d,J＝7.6Hz,1H),4.48(s,2H).

Step three: methyl (S) -2- ((7-methyl-2- (1-carbonylisoindolin-5-yl) imidazo [1,2-a ] pyridin-3-yl) methyl) morpholine-4-carboxylate (1)

In a microwave tube, (2S) -2-ethynylmorpholine-4-carboxylic acid methyl ester (100.0mg,0.59mmol), 1b (95.3mg,0.59mmol) and 4-methylpyridin-2-amine (63.9mg,0.59mmol) were dissolved in toluene (5mL) and N, N-dimethylacetamide (16. mu.L, 0.177mmol), cuprous chloride (17.5mg,0.177mmol) and copper triflate (64.1mg,0.177mmol) were added. After bubbling the reaction solution with nitrogen for 3 minutes, it was heated to 140 ℃ under microwave (1bar) for 5 hours. LCMS showed complete reaction, the residue obtained after filtration and concentration of the reaction mixture was subjected to preparative HPLC (column: Phenomenex Gemini-NX 75X 30mm X3 μm; mobile phase: acetonitrile/water (0.05% NH)₃·H₂O), 14% -54%) to give the title compound 1(11.7mg, yield: 4.5%).

MS(ESI)m/z＝421.2[M+H]⁺.

¹H NMR(400MHz,CD₃OD)δ8.34(d,J＝6.0Hz,1H),7.99-7.79(m,3H),7.33(br s,1H),6.84(d,J＝6.8Hz,1H),4.55(s,2H),4.01(d,J＝12.0Hz,1H),3.84(d,J＝12.0Hz,2H),3.76(br.s,1H),3.69(s,3H),3.41(t,J＝10.8Hz,1H),3.28(d,J＝6.0Hz,2H),3.30-3.25(m,1H),2.99(br s,1H),2.44(s,3H).

Example 2

Methyl (S) -2- ((7-methyl-2- (3-oxoisoindolin-5-yl) imidazo [1,2-a ] pyridin-3-yl) methyl) morpholine-4-carboxylate (2)

The method comprises the following steps: 6-Vinylisoindoline-1-one (2a)

See example 1, step one for the synthesis of compound 2 a. Wherein 6-bromo-2, 3-dihydro-1H-isoindol-1-one is used instead of 5-bromo-2, 3-dihydro-1H-isoindol-1-one.

MS(ESI)m/z＝160.1[M+H]⁺.

¹H NMR(400MHz,CDCl₃)δ7.94(s,1H),7.61(dd,J＝1.6,8.0Hz,1H),7.44(d,J＝8.0Hz,1H),6.84-6.77(m,1H),6.65(br s,1H),5.86(d,J＝17.6Hz,1H),5.35(d,J＝10.8Hz,1H),4.47(s,2H)；ES-MS m/z 160.1[M+H]⁺.

Step two: 1-oxoisoindoline-6-carbaldehyde (2b)

See example 1, step two for the synthesis of compound 2 b. In which 1a is replaced by 2 a.

¹H NMR(400MHz,DMSO-d₆)δ10.11(s,1H),8.79(br s,1H),8.18(s,1H),8.11(dd,J＝1.2,8.0Hz,1H),7.79(d,J＝8.0Hz,1H),4.49(s,2H).

Step three: methyl (S) -2- ((7-methyl-2- (3-oxoisoindolin-5-yl) imidazo [1,2-a ] pyridin-3-yl) methyl) morpholine-4-carboxylate (2)

See example 1, step three, for the synthetic procedure for compound 2. Wherein 1b is replaced by 2 b.

MS(ESI)m/z＝421.2[M+H]⁺.

¹H NMR(400MHz,CD₃OD)δ8.37-8.25(m,2H),8.15-8.02(m,2H),7.64(br d,J＝7.6Hz,1H),7.34(br s,1H),6.78(br d,J＝7.2Hz,1H),4.42(s,2H),3.89(br d,J＝13.2Hz,1H),3.80-3.70(m,3H),3.60(s,3H),3.41-3.28(m,3H),2.98-2.88(m,1H),2.79-2.76(m,1H),2.39(s,3H).

Example 3

Methyl (S) -2- ((7-methyl-2- (2-methyl-1H-benzo [ d ] imidazol-5-yl) imidazo [1,2-a ] pyridin-3-yl) methyl) morpholine-4-carboxylate (3)

The method comprises the following steps: 5-bromo-2-methyl-1H-benzo [ d ] imidazole (3a)

To a solution of 4-bromobenzene-1, 2-diamine (1.00g,5.35mmol) in acetic acid (30mL) was added concentrated hydrochloric acid (134. mu.L, 1.60mmol), and the reaction mixture was stirred at 100 ℃ for 12 hours. LCMS showed reaction completion, the reaction mixture was concentrated under reduced pressure, and the resulting residue was poured into saturated NaHCO₃In solution (30mL), ethyl acetate (30mL) was extracted and the organic phase was washed with Na₂SO₄Drying, filtration and concentration of the filtrate under reduced pressure gave the title compound 3a (600.0mg, 53.2%).

¹H NMR(400MHz,CD₃OD)δ7.62(d,J＝1.2Hz,1H),7.41-7.35(m,1H),7.29(dd,J＝1.6,8.4Hz,1H),2.55(s,3H).

Step two: 2-methyl-1H-benzo [ d ] imidazole-5-carbaldehyde (3b)

To a suspension of NaH (41.7mg,1.04mmol) in dry THF (25mL) at 0 deg.C under nitrogen was added 5-bromo-2-methyl-1H-1, 3-benzodiazole (100.0mg,0.47mmol) and the mixture was stirred at 0 deg.C for 30 min. The mixture was then cooled to-78 deg.C, N-butyllithium (2.4mL,5.92mmol) was added and after stirring for 1 hour, N-dimethylformamide (92)6 μ L,11.84 mmol). After the reaction mixture was stirred at-78 ℃ for 1 hour, TLC showed the reaction was complete. Saturated NH for reaction mixture₄The Cl solution (20mL) was quenched and extracted with ethyl acetate (3X 30mL), and the combined organic phases were Na filtered₂SO₄Drying, filtration, and concentration of the filtrate under reduced pressure gave a residue which was purified by silica gel column chromatography (DCM/MeOH ═ 0-5%) to give the title compound 3b (120.0mg, 31.6%).

¹H NMR(400MHz,CD₃OD)δ10.00(s,1H),8.05(s,1H),7.79(dd,J＝1.6,8.4Hz,1H),7.61(d,J＝8.4Hz,1H),2.61(s,3H).

Step three: methyl (S) -2- ((7-methyl-2- (2-methyl-1H-benzo [ d ] imidazol-5-yl) imidazo [1,2-a ] pyridin-3-yl) methyl) morpholine-4-carboxylate (3)

See example 1, step three, for the synthetic procedure for compound 3. In which 1b is replaced by 3 b.

MS(ESI)m/z＝420.3[M+H]⁺.

¹H NMR(400MHz,CD₃OD)δ8.31(d,J＝7.2Hz,1H),7.87(s,1H),7.58(s,2H),7.32(s,1H),6.81(d,J＝6.8Hz,1H),3.96(d,J＝12.8Hz,1H),3.84(d,J＝12.4Hz,2H),3.72(s,1H),3.67(s,3H),3.44-3.38(m,1H),3.26(d,J＝4.8Hz,2H),2.98(br s,1H),2.76(br s,1H),2.60(s,3H),2.44(s,3H).

Example 4

Methyl (S) -2- ((2- (4, 6-difluoro-2-methyl-1H-benzo [ d ] imidazol-5-yl) -7-methylimidazo [1,2-a ] pyridin-3-yl) methyl) morpholine-4-carboxylate (4)

The method comprises the following steps: 4, 6-difluoro-2-methyl-1H-benzo [ d ] imidazole (4a)

See example 3, step one for the synthesis of compound 4 a. Wherein 3, 5-difluorobenzene-1, 2-diamine is used for replacing 4-bromobenzene-1, 2-diamine.

¹H NMR(400MHz,CD₃OD)δ7.00(dd,J＝2.0,8.4Hz,1H),6.78(dt,J＝2.4,10.4Hz,1H),2.54(s,3H).

Step two: 4, 6-difluoro-2-methyl-1H-benzo [ d ] imidazole-5-carbaldehyde (4b)

See example 3, step two for the synthesis of compound 4 b. In which 3a is replaced by 4 a.

¹H NMR(400MHz,CD₃OD)δ10.32(s,1H),7.15-7.10(m,1H),2.59(s,3H).

Step three: methyl (S) -2- ((2- (4, 6-difluoro-2-methyl-1H-benzo [ d ] imidazol-5-yl) -7-methylimidazo [1,2-a ] pyridin-3-yl) methyl) morpholine-4-carboxylate (4)

See example 1, step three, for the synthetic procedure for compound 4. In which 1b is replaced by 4 b.

MS(ESI)m/z＝456.2[M+H]⁺.

¹H NMR(400MHz,CD₃OD)δ8.40(d,J＝7.2Hz,1H),7.33(s,1H),7.21(d,J＝9.6Hz,1H),6.86(d,J＝7.2Hz,1H),3.77(d,J＝11.6Hz,3H),3.62(s,3H),3.59-3.51(m,1H),3.39-3.32(m,2H),3.16-2.99(m,2H),2.83(br s,1H),2.61(s,3H),2.46(s,3H).

Example 5

Methyl (S) -2- ((2- (5, 7-difluoro-2-carbonyl-2, 3-dihydrobenzo [ d ] oxazol-6-yl) -7-methylimidazo [1,2-a ] pyridin-3-yl) methyl) morpholine-4-carboxylate (5)

The method comprises the following steps: 5, 7-Difluorobenzo [ d ] oxazol-2 (3H) -one (5a)

To a solution of 2-amino-4, 6-difluorophenol (828.6mg,5.71mmol) in tetrahydrofuran (20mL) was added N, N-carbonyldiimidazole (1.1g,6.85mmol), the reaction solution was stirred at room temperature for 2 hours, the reaction was completed, the reaction solution was concentrated, and the crude product was purified by silica gel column chromatography (ethyl acetate/petroleum ether: 5-45%, flow rate: 20mL/min) to give compound 5a (869.5mg, 89.0%).

MS(ESI)m/z＝172.3[M+H]⁺.

Step two: 5, 7-difluoro-2-carbonyl-2, 3-dihydrobenzo [ d ] oxazole-6-carbaldehyde (5b)

Lithium diisopropylamide (3.7mL,5.60mmol) was added dropwise to 5a (869.5mg,5.08mmol) in anhydrous tetrahydrofuran (10mL) at-78 ℃, after completion of the addition, stirring was performed for 20 minutes while maintaining-78 ℃, then N, N-dimethylformamide (0.44mL,5.60mmol) was added dropwise, followed by heating to 0 ℃ to react for 30 minutes, completion of the reaction, quenching was added with a saturated aqueous ammonium chloride solution, extraction was performed using dichloromethane and water, the organic phase was concentrated by drying, and the crude product was purified by silica gel column chromatography (ethyl acetate/petroleum ether ═ 10 to 55%, flow rate: 20mL/min) to give the title compound 5b (677.3mg, 67.0%).

MS(ESI)m/z＝200.4[M+H]⁺.

Step three: methyl (S) -2- ((2- (5, 7-difluoro-2-carbonyl-2, 3-dihydrobenzo [ d ] oxazol-6-yl) -7-methylimidazo [1,2-a ] pyridin-3-yl) methyl) morpholine-4-carboxylate (5)

See example 1, step three, for the synthetic procedure for compound 5. In which 1b is replaced by 5 b.

MS(ESI)m/z＝459.4[M+H]⁺.

¹H NMR(400MHz,DMSO-d₆)δ12.59(br s,1H),8.83(br d,J＝6.8Hz,1H),7.73(s,1H),7.37(br d,J＝6.3Hz,1H),7.26(d,J＝8.5Hz,1H),3.90(br d,J＝13.3Hz,1H),3.24(br d,J＝12.3Hz,3H),3.18-3.06(m,2H),2.83(br s,1H),2.68-2.59(m,1H),2.55(s,4H),2.44-2.32(m,1H),1.36-1.04(m,1H).

Example 6

Methyl (S) -2- ((2- (6, 8-difluoro-3-carbonyl-3, 4-dihydro-2H-benzo [ b ] [1,4] oxazin-7-yl) -7-methylimidazo [1,2-a ] pyridin-3-yl) methyl) morpholine-4-carboxylate (6)

The method comprises the following steps: 6, 8-difluoro-2H-benzo [ b ] [1,4] oxazin-3 (4H) -one (6a)

To a solution of 2-amino-4, 6-difluorophenol (828.6mg,5.71mmol) and potassium carbonate (2.4g,17.13mmol) in tetrahydrofuran (20mL) was added chloroacetyl chloride (0.5mL,6.28mmol), followed by stirring at 40 ℃ for 1 hour, completion of the reaction, extraction with dichloromethane and water, drying of the organic phase for concentration, and purification of the crude product via silica gel column chromatography (ethyl acetate/petroleum ether ═ 10-45%, flow rate: 30mL/min) to give the title compound 6a (919.7mg, 87.0%).

MS(ESI)m/z＝186.3[M+H]⁺.

Step two: 6, 8-difluoro-3-carbonyl-3, 4-dihydro-2H-benzo [ b ] [1,4] oxazine-7-carbaldehyde (6b)

See example 5, step two, for the synthesis of compound 6b, wherein compound 6a is substituted for 5 a.

MS(ESI)m/z＝214.5[M+H]⁺.

Step three: methyl (S) -2- ((2- (6, 8-difluoro-3-carbonyl-3, 4-dihydro-2H-benzo [ b ] [1,4] oxazin-7-yl) -7-methylimidazo [1,2-a ] pyridin-3-yl) methyl) morpholine-4-carboxylate (6)

See example 1, step three, for the synthetic procedure for compound 6. In which 1b is replaced by 6 b.

MS(ESI)m/z＝473.6[M+H]⁺.

¹H NMR(400MHz,DMSO-d₆)δ10.98(s,1H),8.53(d,J＝7.3Hz,1H),7.41(s,1H),7.00-6.82(m,1H),6.74-6.55(m,2H),5.92-5.70(m,1H),4.72(s,2H),3.98(br s,2H),3.55(br s,3H),3.48-3.39(m,1H),3.48-3.39(m,2H),3.29-3.16(m,3H),2.40(s,3H).

Biological evaluation

The FLIPR assay screens compounds for antagonistic activity at hP2X3 and hP2X2/3 receptors (changes in calcium flux signaling indicate the effect of the compound on ion channels).

1. Laboratory instruments and materials

2. Experimental procedure

1321N1 cells (adherent cells) stably transfected with hP2X3 and hP2X2/3 receptor were digested, centrifuged, resuspended and counted in plating medium (DMEM + 10% DFBS), and cells were adjusted to 2X 10⁵cells/mL, 50. mu.L of cells per well in 384-well Assay Plate, placed in 5% CO₂And culturing in an incubator at 37 ℃ for 16-24 hours. 180-fold higher concentration of test Compound (20mM DMSO stock) was made up in DMSO, 500nL per well was added to 384-well Compound Plate, and 30. mu.L of FLIPR buffer (containing 1.26mM Ca) was supplemented²⁺1 HBSS +2mM CaCl₂+20mM HEPES), shaken for 20-40min to mix. 3-fold desired concentration of agonist α, β -meATP (500 nM final hP2X3 cells and 1000nM final hP2X2/3 cells) was formulated with a FLIPR buffer, adding 35. mu.L of agonist per well to another 384-well Compound Plate. The cell plates, which were plated for 16-24 hours, were removed, the cell supernatants were aspirated, and 30. mu.L of Dye (E) was added to each well

Calcium 4Assay Kit, FLIPR buffer dilution), incubated for 1 hour. mu.L of compound was added to each well (sample was added to the FLIPR instrument) and after 15 minutes, 22.5. mu.L of agonist was added to each well and the fluorescence signal was detected (excitation wavelength 470nm-495nm, emission wavelength 515nm-575 nm). Taking the difference value between the peak value and the valley value of the signal as basic data, taking the data of the highest concentration of the positive drug as the 100% inhibition rate and taking DMSO data as the 0% inhibition rate, fitting an inhibition effect curve of the compound on software Graphpad Prism 6 and calculating IC₅₀The value is obtained.

Compound numbering	hP2X3(IC₅₀,nM)	hP2X2/3(IC₅₀,nM)
			MK-7264	35.4	116.2
1	>10000	NA
			2	>10000	NA
3	>10000	NA
			4	639.5	NA
5	221.2	NA
			6	47.0	1232

Claims

1. A compound of formula (I), or a pharmaceutically acceptable salt thereof, or an isomer thereof,

R₂selected from hydrogen, deuterium, optionally substituted by halogen or deuteriumSubstituted C₁-C₆Alkyl, halogen, C optionally substituted by halogen or deuterium₁-C₆Alkoxy radical, C₁-C₆A hydroxyalkyl group;

y is selected from the group consisting of 3-18 membered aromatic or non-aromatic heterocyclyl, preferably 7-15 membered aromatic or non-aromatic heterocyclyl, more preferably 9-12 membered bicyclic or tricyclic heterocyclyl, said heterocyclyl being optionally substituted, preferably said heterocyclyl is substituted with alkylamide, halogen, oxygen, alkyl or alkoxy;

x is selected from C, N and O.

2. A compound of formula (I) according to claim 1,

y is selected from

R₆independently selected from halogen, C₁-C₄An alkyl group;

R₇independently selected from halogen, C₁-C₄Alkyl radical, C₃-C₆A cycloalkylene radical, orAdjacent R₇Are connected into a ring, and the ring optionally has halogen or C₁-C₄Alkyl substitution;

m is an integer from 0 to 4;

n is an integer from 0 to 3;

a is selected from an integer of 1 to 4.

3. A compound of formula (I) according to claim 1,

R₂selected from hydrogen, deuterium, halogen;

R₃and R₄Each independently selected from hydrogen, halogen, or R₃And R₄Together with the carbon atom to which they are both attached form C optionally substituted by halogen₃-C₆A cycloalkylene group;

R₅is selected from C₁-C₆Alkyl or C₁-C₆An alkoxy group;

x is selected from C, O;

y is selected from

R₆、R₇m, n, a are as defined in claim 1.

4. A compound of formula (I) according to claim 1,

R₁selected from hydrogen, deuterium, fluorine, chlorine, or methyl optionally substituted with deuterium or fluorine;

R₂selected from hydrogen, fluorine, chlorine;

R₃and R₄Each independently selected from hydrogen, fluorine, or R₃And R₄Together with the carbon atom to which they are both attached form an optionally halogen-substituted radicalCyclopropylene of (a);

R₅is selected from C₁-C₆Alkyl or C₁-C₆An alkoxy group;

x is selected from C, O;

y is selected from

R₆、R₇m, n, a are as defined in claim 1.

5. A compound of formula (I) according to claim 1,

y is selected from

R₆Independently selected from halogen, C₁-C₄An alkyl group;

m is an integer from 0 to 4;

n is an integer from 0 to 3;

R₁、R₂、R₃、R₄、R₅x is as defined in claim 1.

6. A compound of formula (I) according to claim 1,

y is selected from

Or one or more F in the foregoing structures is substituted with Cl or H;

R₁、R₂、R₃、R₄、R₅x is as defined in claim 1.

7. A compound of formula (I) according to claim 1,

R₁、R₂、R₃、R₄、R₅x is as defined in claim 1,

y is selected from

8. A compound of formula (I-1), or a pharmaceutically acceptable salt thereof, or an isomer thereof,

y is selected from

R₆independently selected from halogen, C₁-C₄An alkyl group;

m is an integer from 0 to 4;

n is an integer from 0 to 3;

a is selected from an integer of 1 to 4.

9. The compound of formula (I-1) according to claim 8,

y is selected from

R₆Independently selected from halogen, C₁-C₄An alkyl group;

m is an integer from 0 to 4;

n is an integer from 0 to 3.

Preferably, Y is selected from

Or the foregoing structureWherein one or more F is substituted with Cl or H.

More preferably, Y is selected from

10. A compound represented by the following formula, or a pharmaceutically acceptable salt thereof, or an isomer thereof,

11. a method of treating a compound of formula (I) or a pharmaceutically acceptable salt thereof, or an isomer thereof, comprising the steps of:

R₁、R₂、R₃、R₄、R₅x, Y are as defined in claim 1.

12. A process for preparing a compound of formula (I) as claimed in claim 1 or a pharmaceutically acceptable salt thereof, or an isomer thereof, comprising the steps of:

R₁、R₂、R₃、R₄、R₅x, Y are as defined in claim 1;

13. A process for preparing a compound of formula (I) as claimed in claim 1 or a pharmaceutically acceptable salt thereof, or an isomer thereof, comprising the steps of:

R₁、R₂、R₃、R₄、R₅x, Y are as defined in claim 1;

z is selected from the group consisting of halogen, sulfonyl and sulfinyl.

14. A process for preparing a compound of formula (I) as claimed in claim 1 or a pharmaceutically acceptable salt thereof, or an isomer thereof, comprising the steps of:

R₁、R₂、R₃、R₄、R₅x, Y are as defined in claim 1;

z is selected from the group consisting of halogen, sulfonyl and sulfinyl.

15. A pharmaceutical composition comprising a compound of claims 1-10, or a pharmaceutically acceptable salt thereof, or an isomer thereof, and at least one pharmaceutically acceptable carrier, diluent, or excipient.

16. Use of a compound according to claims 1-10, or a pharmaceutically acceptable salt, or isomer thereof, or a pharmaceutical composition of claim 24, for the manufacture of a medicament for the treatment of a disease associated with P2X3 activity; preferably the disorder is selected from pain, urinary tract disorders or cough.