CN113912601B

CN113912601B - Novel imidazo [1,2-a ] pyridine derivatives, preparation method and medical application thereof

Info

Publication number: CN113912601B
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: 2023-06-06
Anticipated expiration: 2041-07-09
Also published as: CN113912601A

Abstract

The present disclosure relates to a novel class of imidazo [1,2-a]Pyridine derivatives, a preparation method and medical application thereof. Specifically, the substituent group of the derivative comprises a parallel ring structure, the structural formula is shown as the following, and R is ₁ 、R ₂ 、R ₃ 、R ₄ 、R ₅ The definitions of X and Y are described in the specification.

Description

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

Technical Field

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

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, known as purinergic receptors. The P2X receptor is present in a variety of organisms including humans, mice, rats, rabbits, chickens, zebra fish, bullfrog, flukes and amoebas. Seven independent genes encoding the P2X subunits have been identified and designated P2X1 to P2X7. 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 intracellular, conserved sequences located in the extracellular loop structure. High expression of the P2X3 receptor is found in both specific small and medium diameter neurons associated with nociceptive 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, a key receptor-mediated bladder sensory signal, expressed in bladder mucosal tissue containing abundant sensory nerve fibers. There is also expression of P2X3 in sensory nerve fibers of the pharyngeal mucosa, which is involved in taste transduction, formation.

When the organism is injured or nerve injured, a large amount of ATP is released, a presynaptic membrane P2X3 receptor is activated, a large amount of Ca < 2+ > is caused to flow in, and intracellular calcium concentration is increased to activate Protein Kinase A (PKA), protein Kinase C (PKC), so that PKA and PKC are phosphorylated, glutamate release is promoted, an NMDA receptor is further activated, and excitatory postsynaptic current is generated to cause central sensitization. Many studies have shown that upregulation of P2X3 receptor expression can lead to nociception, involved in pain signaling.

MK-7264 is an antagonist of P2X3 receptor activity with IC50 values of 30nM and 100-250nM for human homologous recombination hP2X3 and hP2X2/3, respectively, and its use for treatment of chronic cough patients has been advanced to clinical stage III.

Disclosure of Invention

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

wherein R is ₁ Selected from hydrogen, deuterium, optionally halogen-or deuterium-substituted C ₁ -C ₆ Alkyl, halogen, optionally halogen-or deuterium-substituted C ₁ -C ₆ Alkoxy, C ₁ -C ₆ A hydroxyalkyl group;

R ₂ selected from hydrogen, deuterium, optionally halogen-or deuterium-substituted C ₁ -C ₆ Alkyl, halogen, optionally halogen-or deuterium-substituted C ₁ -C ₆ Alkoxy, C ₁ -C ₆ A hydroxyalkyl group;

R ₃ and R is ₄ Independently selected from hydrogen, halogen, optionally halogen-substituted C ₁ -C ₄ Alkyl, or R ₃ And R is ₄ Together with the carbon atoms to which they are attached form an optionally halogen-substituted C ₃ -C ₆ Cyclic hydrocarbylene radicals, or C selected from the group consisting of optionally halogen-substituted C on adjacent carbon atoms ₁ -C ₄ R of alkyl ₃ And R is ₄ Connecting the two components to form a ring;

R ₅ selected from the group consisting of: c optionally substituted by halogen or cyano ₁ -C ₆ Alkyl, optionally halogen-or cyano-substituted C ₃ -C ₆ Cycloalkyl, optionally halogen-or cyano-substituted heterocyclyl, optionally halogen-or cyano-substituted C ₁ -C ₆ Alkoxy, amino optionally substituted with alkyl;

y is selected from 3-18 membered aromatic or non-aromatic heterocyclic groups, preferably 7-15 membered aromatic or non-aromatic heterocyclic groups, more preferably 9-12 membered bicyclic or tricyclic heterocyclic groups, said heterocyclic groups 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 optionally being 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 optionally being substituted with methylamide, fluoro, chloro, methyl, ethyl, oxygen.

In some embodiments of the present invention, in some embodiments,

y is selected from

J ₁ Selected from N, C, O, S, J ₂ Selected from N, C, ring A is a ring containing-NH-C (=O) -or-NH-S (=O) ₂ -a heterocyclyl group;

R ₆ independently selected from halogen, C ₁ -C ₄ An alkyl group;

R ₇ independently selected from halogen, C ₁ -C ₄ Alkyl, C ₃ -C ₆ Cyclic hydrocarbylene, or adjacent R ₇ Connected in a ring, optionally having halogen or C ₁ -C ₄ Alkyl substitution;

m is selected from integers from 0 to 4;

n is an integer from 0 to 3;

a is selected from integers from 1 to 4.

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

R ₁ selected from hydrogen, deuterium, halogen, optionally halogen-or deuterium-substituted C ₁ -C ₆ An alkyl group, a hydroxyl group,

R ₂ selected from the group consisting of hydrogen, deuterium, and halogen,

y is selected from

R ₆ independently selected from halogen, C ₁ -C ₄ An alkyl group;

x is selected from C, N and O;

m is selected from integers from 0 to 4;

n is an integer from 0 to 3;

a is selected from integers from 1 to 4.

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

R ₂ selected from the group consisting of hydrogen, deuterium, and halogen,

R ₅ selected from C ₁ -C ₆ Alkyl or C ₁ -C ₆ An alkoxy group, an amino group,

y is selected from

R ₆ independently selected from halogen, C ₁ -C ₄ An alkyl group;

x is selected from C, N and O;

m is selected from integers from 0 to 4;

n is an integer from 0 to 3;

a is selected from integers from 1 to 4.

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

R ₂ selected from the group consisting of hydrogen, deuterium, and halogen,

R ₃ and R is ₄ Selected from hydrogen, halogen, or R ₃ And R is ₄ Together with the carbon atoms to which they are commonly attached to formOptionally halogen-substituted C ₃ -C ₆ A cyclic alkylene group, an alkylene group,

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

y is selected from

R ₆ independently selected from halogen, C ₁ -C ₄ An alkyl group;

m is selected from integers from 0 to 4;

n is an integer from 0 to 3;

a is selected from integers from 1 to 4.

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

R ₁ selected from hydrogen, deuterium, halogen, optionally halogen-or deuterium-substituted C ₁ -C ₆ An alkyl group;

R ₂ selected from hydrogen, deuterium, halogen;

R ₃ and R is ₄ Selected from hydrogen, halogen, or R ₃ And R is ₄ Together with the carbon atoms to which they are attached form an optionally halogen-substituted C ₃ -C ₆ A cyclic alkylene group;

R ₅ selected from C ₁ -C ₆ Alkyl or C ₁ -C ₆ An alkoxy group;

x is selected from C, O;

y is selected from

J ₁ Selected from N, C, O, S, J ₂ Selected from N, C, ring A is a ring containing-NH-C (=O) -or-NH-S (=O) ₂ -a 5-13 membered heterocyclyl;

R ₆ independently selected from halogen, C ₁ -C ₄ An alkyl group;

m is selected from integers from 0 to 4;

n is an integer from 0 to 3;

a is selected from integers from 1 to 4.

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

R ₁ selected from hydrogen, deuterium, fluorine, chlorine, or methyl optionally substituted with deuterium, fluorine;

R ₂ selected from hydrogen, fluorine, chlorine;

R ₃ and R is ₄ Selected from hydrogen, fluorine, or R ₃ And R is ₄ Together with the carbon atoms to which they are attached form an optionally halogen-substituted cyclopropylene group;

R ₅ selected from C ₁ -C ₆ Alkyl or C ₁ -C ₆ An alkoxy group;

x is selected from C, O;

y is selected from

R ₆ independently selected from halogen, C ₁ -C ₄ An alkyl group;

m is selected from integers from 0 to 4;

n is an integer from 0 to 3;

a is selected from integers from 1 to 4.

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

y is selected from

R ₆ Independently selected from halogen, C ₁ -C ₄ An alkyl group;

m is selected from integers from 0 to 4;

n is an integer from 0 to 3;

R ₁ selected from hydrogen, deuterium, optionally halogen-or deuterium-substituted C ₁ -C ₆ Alkyl, halogen, optionally halogen-or deuterium-substituted C ₁ -C ₆ Alkoxy, C ₁ -C ₆ A hydroxyalkyl group;

R ₅ selected from the followingThe group consisting of: c optionally substituted by halogen or cyano ₁ -C ₆ Alkyl, optionally halogen-or cyano-substituted C ₃ -C ₆ Cycloalkyl, optionally halogen-or cyano-substituted heterocyclyl, optionally halogen-or cyano-substituted C ₁ -C ₆ Alkoxy, amino optionally substituted with alkyl;

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 F in the foregoing structure is replaced 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),

R ₂ selected from hydrogen, fluorine, chlorine;

R ₅ selected from C ₁ -C ₆ Alkyl or C ₁ -C ₆ An alkoxy group;

x is selected from C, O;

y is selected from

R ₆ Independently selected from halogen, C ₁ -C ₄ An alkyl group;

m is selected from integers from 0 to 4;

n is an integer from 0 to 3.

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

R ₂ selected from hydrogen, fluorine, chlorine;

R ₅ selected from C ₁ -C ₆ Alkyl or C ₁ -C ₆ An alkoxy group;

x is selected from C, O;

y is selected from

Or one or more F in the foregoing structure is replaced with Cl or H.

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

R ₂ selected from hydrogen, fluorine, chlorine;

R ₅ selected from C ₁ -C ₆ Alkyl or C ₁ -C ₆ An 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

R ₆ independently selected from halogen, C ₁ -C ₄ An alkyl group;

m is selected from integers from 0 to 4;

n is an integer from 0 to 3;

a is selected from integers from 1 to 4.

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

y is selected from

J ₁ Selected from N, C, O, S, J ₂ Selected from N, C, ring A is a ring containing-NH-C (=O) -or-NH-S (=O) ₂ -a 5-to 13-membered heterocyclic group,

R ₆ independently selected from halogen, C ₁ -C ₄ An alkyl group;

m is selected from integers from 0 to 4;

n is an integer from 0 to 3;

a is selected from integers from 1 to 4.

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

R ₆ Independently selected from halogen, C ₁ -C ₄ An alkyl group;

m is selected from integers from 0 to 4;

n is an integer from 0 to 3.

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

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

Or one or more F in the foregoing structure is replaced 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:

cyclizing a compound shown in the formula (I-a) with a compound shown in the formula (I-b) and a compound shown in the formula (I-c) under the condition of a catalyst to obtain a 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) iron palladium dichloride, 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 triflate;

R ₁ 、R ₂ 、R ₃ 、R ₄ 、R ₅ x, Y are as defined for the compounds of formula (I).

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

cyclizing a compound shown in the formula (I-d) with a compound shown in the formula (I-e) and a compound shown in 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 compounds of formula (I-1).

cyclizing a compound shown in the formula (I-d) with a compound shown in the formula (I-a) to obtain a compound shown in the formula (I-e); carrying out halogenation reaction on a compound shown in the formula (I-e) to obtain a compound shown in the formula (I-f); the compound shown in the formula (I-f) is subjected to C-C coupling reaction under alkaline condition 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) dichloro iron palladium, tris (dibenzylideneacetone) dipalladium or 2-dicyclohexylphosphine-2 ',6' -dimethoxybiphenyl, [1,1' -bis (di-tert-butylphosphino) ferrocene ] dichloropalladium (II);

R ₁ 、R ₂ 、R ₃ 、R ₄ 、R ₅ x, Y are defined as compounds of formula (I-1);

z' and Z are selected from halogen, sulfonyl and sulfinyl.

cyclizing a compound shown in the formula (I-a) with a compound shown in the formula (I-g) to obtain a compound shown in the formula (I);

R ₁ 、R ₂ 、R ₃ 、R ₄ 、R ₅ x, Y are defined as compounds of formula (I);

z is selected from halogen, sulfonyl and sulfinyl.

cyclizing a compound shown in the formula (I-a) with a compound shown in the formula (I-h) to obtain a compound shown in the formula (I-I); the compound shown in the formula (I-I) is subjected to addition reduction dehydroxylation with the compound shown in the formula (I-j) to obtain the compound shown in the formula (I);

z is selected from halogen, sulfonyl and sulfinyl.

The present disclosure also provides methods of preparing the compounds, or pharmaceutically acceptable salts thereof, or isomers thereof. In particular, the preparation was carried out by the method of 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. Generally, 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 containing 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 excessive P2X3 activity. The compounds of the present disclosure are highly selective for P2X3 and may avoid loss of taste. In some embodiments, the compounds of the present disclosure antagonize P2X3 cognate receptors more than 20-fold more than P2X2/3 heteromeric receptors. In some embodiments, the compounds of the present disclosure antagonize P2X3 cognate receptors more than 30-fold more than P2X2/3 heteromeric receptors. In some embodiments, the compounds of the present disclosure antagonize P2X3 cognate receptors more than 50-fold more than P2X2/3 heteromeric receptors. In some embodiments, the compounds of the present disclosure antagonize P2X3 cognate receptors by more than 100-fold over P2X2/3 heteromeric receptors.

In some embodiments, the compounds of the present disclosure, or pharmaceutically acceptable salts, or isomers thereof, or pharmaceutical compositions comprising the same, are useful in the treatment of pain, urinary tract disorders, cough, and the like. The 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, may be used to treat 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 treating cardiovascular diseases or for cardioprotection after myocardial infarction; can be used as immunomodulator, in particular for the treatment of autoimmune diseases (e.g. arthritis), for skin grafts, organ transplants, or similar surgical needs, for collagen diseases, for allergies, or as antitumor or antiviral agents; can be used for treating multiple sclerosis, parkinson's disease (Huntington's chorea); can be used for the treatment of 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 cord injury, drug addiction (e.g., treatment of alcohol, nicotine, opioid, or other drug abuse), or sympathological disorders (e.g., hypertension); can be used for treating diarrhea; can be used for treating pulmonary disorders such as asthma, cough or pulmonary oedema, for example.

The compounds of the present disclosure, or pharmaceutically acceptable salts, or isomers thereof, may 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, lateral intraventricular, or by injection into a joint).

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

Any isotopically-labeled (or "radiolabeled") derivative of a compound of the disclosure or a pharmaceutically acceptable salt thereof, or an isomer thereof, is covered by the present disclosure. Such derivatives are those in which one or more atoms are replaced by an atom having an atomic mass or mass number different from that commonly found in nature. Examples of radionuclides that may be incorporated include ² H (also written as "D", deuterium), ³ H (also written as "T", tritium), ¹¹ C、 ¹³ C、 ¹⁴ C、 ¹³ N、 ¹⁵ N、 ¹⁵ O、 ¹⁷ O、 ¹⁸ O、 ¹⁸ F、 ³⁶ Cl、 ⁸² Br、 ⁷⁵ Br、 ⁷⁶ Br、 ⁷⁷ Br、 ¹²³ I、 ¹²⁴ I、 ¹²⁵ I、 ³¹ P、 ³² P、 ³⁵ S, S ¹³¹ I. The radionuclide used will depend on the specific application of the radiolabeled derivative. For example, for in vitro receptor labeling and competition assays, ³ h or ¹⁴ C is often useful. For radiological imaging applications, ¹¹ c or ¹⁸ F is often useful. In some embodiments, the radionuclide is ³ H. In some embodiments, the radionuclide is ¹⁴ C. In some embodiments, the radionuclide is ¹¹ C. And in some embodiments, the radionuclide is ¹⁸ F。

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" refers to straight or branched chain alkyl groups 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 specifically indicated in the present specification, a heterocyclyl group may be a monocyclic, bicyclic, tricyclic or tetracyclic ring system, which may include spiro or bridged ring systems; and the nitrogen, carbon or sulfur atoms in the heterocyclyl may optionally be oxidized; the nitrogen atom may optionally be quaternized; and the heterocyclic group may be partially or fully saturated. "comprising-NH-C (=O) -or-NH-S (=O) ₂ -heterocyclic "ring structure is removed" -NH-C (=O) - "and" -NH-S (=O) ₂ In addition to the "moiety, the other moieties optionally contain heteroatoms.

"cycloalkyl" refers to a stable, non-aromatic, monocyclic or polycyclic hydrocarbon group consisting of only 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 linked to the remainder 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, the number of the cells to be processed,

etc.

"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 cyano ₁ -C ₆ Alkyl "means that halogen or cyano may be, but need not be, present, and this 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 ^-6 Units of (ppm) are given. NMR was performed using Bruker AVANCE-400 nuclear magnetic resonance apparatus with deuterated dimethyl sulfoxide (DMSO-d) ₆ ) Deuterated chloroform (CDCl) ₃ ) And deuterated methanol (CD) ₃ OD), internal standard is Tetramethylsilane (TMS).

HPLC was performed using a Waters ACQUITY ultra high performance LC, shimadzu LC-20A systems, shimadzu LC-2010HT series or Agilent 1200 LC high pressure liquid chromatograph (ACQUITY UPLC BEH C18.7 μm 2.1 x 50mm column, ultimate XB-C18.0 x 150mm column or Xtime C18.1 x 30mm column).

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

Chiral HPLC analysis was performed using a chiral HPLC analysis of 3 μm, chiral pak AD-3X 4.6mm I.D.,3 μm, chiral pak AS-3X 4.6mm I.D.,3 μm, chiral Cel OD-3X 4.6mm I.D.,3 μm, chiral Cel OJ-H150X 4.6mm I.D.,5 μm, chiral Cel OJ-3X 4.6mm I.D.,3 μm, chiral Cel OD-3X 4.6mm I.D.

The thin layer chromatography silica gel plate uses a smoke table yellow sea HSGF254 or Qingdao GF254 silica gel plate, the specification of the silica gel plate used by the 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.

Column chromatography generally uses 100-200 mesh, 200-300 mesh or 300-400 mesh of yellow sea silica gel as a carrier.

The chiral preparation column used DAICEL CHIRALPAK IC (250 mm. Times.30 mm,10 μm) or Phenomnex-Amylose-1 (250 mm. Times.30 mm,5 μm).

Known starting materials of the present disclosure may be synthesized using or following methods known in the art, or may be purchased from ABCR GmbH & co.kg, acros Organics, aldrich Chemical Company, shaog chemical technology (Accela ChemBio Inc), dary chemicals, and the like.

The examples are not particularly described, and the reaction can be carried out under an argon atmosphere or a nitrogen atmosphere.

An argon atmosphere or nitrogen atmosphere means that the reactor flask is connected to a balloon of argon or nitrogen of about 1L volume.

The hydrogen atmosphere is defined as the reaction flask being connected to a hydrogen balloon of about 1L volume.

The pressure hydrogenation reaction uses a Parr 3916 model EKX hydrogenometer and a clear blue QL-500 type hydrogen generator or HC2-SS type hydrogenometer.

The hydrogenation reaction is usually vacuumized, filled with hydrogen and repeatedly operated for 3 times.

The microwave reaction used was a CEM Discover-S908860 type microwave reactor.

The examples are not specifically described, and the solution refers to an aqueous solution.

The reaction temperature is room temperature and is 20-30 deg.c without specific explanation in the examples.

The monitoring of the progress of the reaction in the examples employed Thin Layer Chromatography (TLC), the developing reagent used for the reaction, the system of eluent for column chromatography employed for purifying the compound and the developing reagent system of thin layer chromatography included: 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 is regulated according to the polarity of the compound, and can be regulated by adding a small amount of alkaline or acidic reagents such as triethylamine, acetic acid and the like.

Example 1

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

Step one: 5-Vinylisoindolin-1-one (1 a)

To a mixture of 5-bromo-2, 3-dihydro-1H-isoindol-1-one (500.0 mg,2.36 mmol) and 4, 5-tetramethyl-2-vinyl-1, 3, 2-dioxaborolan (480. Mu.L, 2.83 mmol) in 1, 4-dioxan (10 mL) and water (2.5 mL) under nitrogen was added Cs ₂ CO ₃ (1536.6 mg,4.72 mmol) and Pd (dppf) Cl ₂ (174.9 mg,0.24 mmol) and then stirred 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 (330.0 mg, 79.1%).

¹ H NMR(400MHz,CD ₃ OD)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-oxo-isoindoline-5-carbaldehyde (1 b)

To a mixture of 1a (150.0 mg,0.85 mmol) of 1, 4-dioxane (6 mL) and water (1.2 mL) was added osmium tetroxide (32.3 mg,0.13 mmol), and the mixture was stirred at 15℃for 10 minutes. Sodium periodate (997.0 mg,3.39 mmol) 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 30 mL) and the combined organic phases were taken up over Na ₂ SO ₄ Drying, filtration and concentration gave the title compound 1b (120.0 mg, 79.0%).

¹ H NMR(400MHz,DMSO-d ₆ )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-ethynyl morpholine-4-carboxylic acid methyl ester (100.0 mg,0.59 mmol), 1b (95.3 mg,0.59 mmol) and 4-methylpyridin-2-amine (63.9 mg,0.59 mmol) were dissolved in toluene (5 mL) followed by N, N-dimethylacetamide (16. Mu.L, 0.177 mmol), cuprous chloride (17.5 mg,0.177 mmol) and copper triflate (64.1 mg,0.177 mmol). After bubbling the reaction solution with nitrogen for 3 minutes, it was heated to 140℃under microwaves (1 bar) to react for 5 hours. LCMS showed complete reaction, and the residue obtained after filtration and concentration of the reaction mixture was purified by preparative HPLC (column Phenomenex Gemini-nx75x30mm×3μm; mobile phase acetonitrile/water (0.05% nh) ₃ ·H ₂ O), 14% -54%) to give the title compound 1 (11.7 mg, yield: 4.5%).

MS(ESI)m/z＝421.2[M+H] ⁺ .

¹ H NMR(400MHz,CD ₃ OD)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)

Step one: 6-Vinylisoindolin-1-one (2 a)

The synthetic procedure for compound 2a is described in step one of example 1. Wherein 6-bromo-2, 3-dihydro-1H-isoindol-1-one is substituted for 5-bromo-2, 3-dihydro-1H-isoindol-1-one.

MS(ESI)m/z＝160.1[M+H] ⁺ .

¹ H NMR(400MHz,CDCl ₃ )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-oxo-isoindoline-6-carbaldehyde (2 b)

The synthesis procedure for compound 2b is described in example 1, step two. Wherein 1a is replaced with 2 a.

¹ H NMR(400MHz,DMSO-d ₆ )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)

The synthetic procedure for compound 2 is described in step three of example 1. Wherein 1b is replaced with 2 b.

MS(ESI)m/z＝421.2[M+H] ⁺ .

¹ H NMR(400MHz,CD ₃ OD)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)

Step one: 5-bromo-2-methyl-1H-benzo [ d ] imidazole (3 a)

To a solution of 4-bromobenzene-1, 2-diamine (1.00 g,5.35 mmol) in acetic acid (30 mL) was added concentrated hydrochloric acid (134. Mu.L, 1.60 mmol), and the reaction mixture was stirred at 100deg.C for 12 hours. LCMS showed complete reaction, the reaction mixture was concentrated under reduced pressure,the resulting residue was poured into saturated NaHCO ₃ In solution (30 mL), ethyl acetate (30 mL) was extracted and the organic phase was taken up in Na ₂ SO ₄ Drying, filtration, and concentration of the filtrate under reduced pressure gave the title compound 3a (600.0 mg, 53.2%).

¹ H NMR(400MHz,CD ₃ OD)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 (3 b)

To a suspension of NaH (41.7 mg,1.04 mmol) in anhydrous THF (25 mL) under nitrogen at 0deg.C was added 5-bromo-2-methyl-1H-1, 3-benzodiazole (100.0 mg,0.47 mmol) and the mixture was stirred at 0deg.C for 30 min. The mixture was then cooled to-78 ℃, N-butyllithium (2.4 ml,5.92 mmol) was added and after stirring for 1 hour N, N-dimethylformamide (926 μl,11.84 mmol) was added. After the reaction mixture was stirred at-78 ℃ for 1 hour, TLC showed the reaction was complete. The reaction mixture was saturated with NH ₄ Cl solution (20 mL) was quenched and extracted with ethyl acetate (3X 30 mL) and the combined organic phases were taken up over Na ₂ SO ₄ Drying, filtration, and concentration of the filtrate under reduced pressure gave a residue that was purified by silica gel column chromatography (DCM/meoh=0-5%) to give the title compound 3b (120.0 mg, 31.6%).

¹ H NMR(400MHz,CD ₃ OD)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)

The synthetic procedure for compound 3 is described in step three of example 1. Wherein 1b is replaced with 3 b.

MS(ESI)m/z＝420.3[M+H] ⁺ .

¹ H NMR(400MHz,CD ₃ OD)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)

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Step one: 4, 6-difluoro-2-methyl-1H-benzo [ d ] imidazole (4 a)

The synthesis procedure for compound 4a is described in example 3, step one. Wherein 3, 5-difluorobenzene-1, 2-diamine is used for replacing 4-bromobenzene-1, 2-diamine.

¹ H NMR(400MHz,CD ₃ OD)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 (4 b)

The synthesis procedure for compound 4b is described in example 3, step two. Wherein 3a is replaced with 4 a.

¹ H NMR(400MHz,CD ₃ OD)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)

The synthetic procedure for compound 4 is described in step three of example 1. Wherein 1b is replaced with 4 b.

MS(ESI)m/z＝456.2[M+H] ⁺ .

¹ H NMR(400MHz,CD ₃ OD)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)

Step one: 5, 7-difluorobenzo [ d ] oxazol-2 (3H) -one (5 a)

To a solution of 2-amino-4, 6-difluorophenol (828.6 mg,5.71 mmol) in tetrahydrofuran (20 mL) was added N, N-carbodiimidazole (1.1 g,6.85 mmol), the reaction 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: 20 mL/min) to give compound 5a (869.5 mg, 89.0%).

MS(ESI)m/z＝172.3[M+H] ⁺ .

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

Lithium diisopropylamide (3.7 mL,5.60 mmol) was added dropwise to 5a (869.5 mg,5.08 mmol) of anhydrous tetrahydrofuran (10 mL) at-78deg.C, stirring was maintained at-78deg.C for 20 min, then N, N-dimethylformamide (0.44 mL,5.60 mmol) was added dropwise, then the reaction was allowed to proceed to 0deg.C for 30 min, quenched with saturated aqueous ammonium chloride solution, extracted with dichloromethane and water, and concentrated by organic drying, and the crude product was purified by silica gel column chromatography (ethyl acetate/petroleum ether=10-55%, flow rate: 20 mL/min) to give the title compound 5b (677.3 mg, 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)

The synthetic procedure for compound 5 is described in step three of example 1. Wherein 1b is replaced with 5 b.

MS(ESI)m/z＝459.4[M+H] ⁺ .

¹ H NMR(400MHz,DMSO-d ₆ )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)

Step one: 6, 8-difluoro-2H-benzo [ b ] [1,4] oxazin-3 (4H) -one (6 a)

To a solution of 2-amino-4, 6-difluorophenol (828.6 mg,5.71 mmol) and potassium carbonate (2.4 g,17.13 mmol) in tetrahydrofuran (20 mL) was added chloroacetyl chloride (0.5 mL,6.28 mmol), and then the reaction was stirred at 40℃for 1 hour, after which the reaction was completed, extracted with dichloromethane and water, the organic phase was concentrated by dryness, and the crude product was purified by silica gel column chromatography (ethyl acetate/petroleum ether=10-45%, flow rate: 30 mL/min) to give the title compound 6a (919.7 mg, 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 (6 b)

The synthesis of compound 6b is described in example 5, step two, wherein compound 6a is substituted for 5a.

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)

The synthetic procedure for compound 6 is described in step three of example 1. Wherein 1b is replaced with 6 b.

MS(ESI)m/z＝473.6[M+H] ⁺ .

¹ H NMR(400MHz,DMSO-d ₆ )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 their antagonistic activity against hp2x3 and hp2x2/3 receptors (the effect of the compounds on ion channels is indicated by changes in calcium flux signals).

1. Experimental instrument and materials

2. Experimental procedure

1321N1 cells stably transfected with hP2X3 and hP2X2/3 receptor (adherent cells) were digested, centrifuged, resuspended in plating medium (DMEM+10% DFBS) and counted to adjust the cells to 2X 10 ⁵ mu.L of cells per well in 384-well Assay Plate were plated at 5% CO per mL ₂ Culturing in a 37 ℃ incubator for 16-24 hours. Test compounds were prepared in DMSO at 180-fold desired concentrations (20 mM DMSO stock solution), 500nL per well was added to 384-well Compound Plate, and 30. Mu.L of FLIPR buffer (containing 1.26mM Ca) ²⁺ Hbss+2mm CaCl ₂ +20mM HEPES), and shaken for 20-40min to mix well. The agonist α, β -meATP (500 nM for hP2X3 cells and 1000nM for hP2X2/3 cells) was formulated in FLIPR buffer at 3-fold concentrations and 35 μl of agonist was added to another 384-well Compound Plate per well. Taking out the cell plate paved with 16-24 hours of culture, sucking out the cell supernatant, adding 30 mu L Dye per well

Calcium 4Assay Kit,FLIPR buffer dilution), for 1 hour. mu.L of the compound (FLIPR instrument) was added to each well of cells, and after 15 minutes, 22.5. Mu.L of the agonist was added to each well, and fluorescence signals were detected (excitation wavelength: 470nm to 495nm, emission wavelength: 515nm to 575 nm). Taking the difference between the peak value and the valley value of the signal as basic data, taking the highest concentration data of the positive medicine as 100% inhibition rate, taking the DMSO data as 0% inhibition rate, fitting an inhibition effect curve of the compound on software Graphpad Prism 6 and calculating IC ₅₀ Values.

Numbering of compounds	hP2X3(IC ₅₀ ,nM)	hP2X2/3(IC ₅₀ ,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,

wherein R is ₁ Selected from hydrogen, deuterium, C optionally substituted by halogen or deuterium ₁ -C ₆ Alkyl, halogen, C optionally substituted by halogen or deuterium ₁ -C ₆ Alkoxy and C ₁ -C ₆ A hydroxyalkyl group;

R ₂ selected from hydrogen, deuterium, C optionally substituted by halogen or deuterium ₁ -C ₆ Alkyl, halogen, C optionally substituted by halogen or deuterium ₁ -C ₆ Alkoxy and C ₁ -C ₆ A hydroxyalkyl group;

R ₃ and R is ₄ Independently selected from hydrogen, halogen and C optionally substituted with halogen ₁ -C ₄ An alkyl group;

R ₅ selected from the group consisting of: c optionally substituted by halogen or cyano ₁ -C ₆ Alkyl, C optionally substituted by halogen or cyano ₃ -C ₆ Cyclic hydrocarbon radicals, optionally halogen-or cyano-substituted C ₁ -C ₆ Alkoxy and-NH ₂ ；

Y is selected from

X is selected from O.

2. The compound of formula (I) or a pharmaceutically acceptable salt thereof according to claim 1,

R ₁ selected from hydrogen, deuterium, halogen and optionally halogen or halogen-substitutedDeuterium substituted C ₁ -C ₆ An alkyl group;

R ₂ selected from hydrogen, deuterium, and halogen;

R ₃ and R is ₄ Each independently selected from hydrogen and halogen;

R ₅ selected from C ₁ -C ₆ Alkyl and C ₁ -C ₆ An alkoxy group;

x is selected from O.

3. The compound of formula (I) or a pharmaceutically acceptable salt thereof according to claim 1,

R ₁ selected from hydrogen, deuterium, fluorine, chlorine and methyl optionally substituted with deuterium or fluorine;

R ₂ selected from hydrogen, fluorine and chlorine;

R ₃ and R is ₄ Each independently selected from hydrogen and fluorine;

R ₅ selected from C ₁ -C ₆ Alkyl and C ₁ -C ₆ An alkoxy group;

x is selected from O.

4. A compound as shown below, or a pharmaceutically acceptable salt thereof,

5. a process for the preparation of a compound of formula (I) as defined in claim 1, or a pharmaceutically acceptable salt thereof, comprising the steps of:

the catalyst is cuprous chloride or copper triflate;

R ₁ 、R ₂ 、R ₃ 、R ₄ 、R ₅ x, Y are defined as in claim 1.

6. A pharmaceutical composition comprising a compound of any one of claims 1-4, or a pharmaceutically acceptable salt thereof, and at least one pharmaceutically acceptable carrier, diluent, or excipient.

7. Use of a compound according to any one of claims 1-4, or a pharmaceutically acceptable salt thereof, or a pharmaceutical composition according to claim 6, in the manufacture of a medicament for the treatment of a disease associated with P2X3 activity.

8. The use according to claim 7, wherein the disease is selected from pain, urinary tract disease or cough.