CN112794860B

CN112794860B - Oxazole pyrimidone amide compound or medicinal salt thereof, preparation method and application

Info

Publication number: CN112794860B
Application number: CN202110314841.XA
Authority: CN
Inventors: 钟燕; 王永临
Original assignee: Shanghai Zhaoyu Medicine Technology Co ltd
Current assignee: Shanghai Zhaoyu Medicine Technology Co ltd
Priority date: 2021-03-24
Filing date: 2021-03-24
Publication date: 2021-06-29
Anticipated expiration: 2041-03-24
Also published as: CN112794860A

Abstract

The invention discloses an oxazole pyrimidone amide compound or a medicinal salt thereof, a preparation method and application thereof, wherein the oxazole pyrimidone amide compound has the following structural general formula (I):

wherein R is₁，R₂Each independently selected from hydrogen and C_1‑3An alkyl group; r₃Is selected from C_1‑3An alkyl group; r₄，R₅Each independently selected from 5-6 membered aryl, the aryl is phenyl or an heteroaryl containing 1-3 heteroatoms, the heteroatoms are taken from oxygen or nitrogen atoms, and the position of the heteroatoms is any position on the heteroaryl; the aromatic group being unsubstituted or at least substituted by one or more halogens or C_1‑3Alkyl radical, C_1‑3Polyhaloalkyl, C_1‑3Alkoxy substitution. The invention also discloses the application of the TRPA1 antagonist in preparing medicaments for treating or preventing diseases, symptoms and/or disorders modulated by TRPA1, such as pain and the like.

Description

Oxazole pyrimidone amide compound or medicinal salt thereof, preparation method and application

Technical Field

The present invention relates to antagonists of the Transient Receptor Potential (TRP) ion channel family. In particular, the invention relates to oxazolopyrimidinone amides having a high active antagonistic effect on TRPA1, including their preparation and their use as TRPA1 channel antagonists.

Background

Transient Receptor Potential (TRP) channels are a type of ion channel that is widely found in the human body. This family is commonly used as receptors for multiple chemical and physical stimuli. At least 28 human TRP channels are known to exist, of which the transient receptor potential a1 channel (TRPA 1) is a non-selective cation-conducting channel, widely distributed in neural and non-neural cells, the former mainly including the vagus nerve ganglion, trigeminal ganglion and dorsal root ganglion; the latter include, for example, vascular endothelial cells, pancreatic islet cells, cardiac myocytes, inner ear hair cells, hepatocytes, gastrointestinal mucosa, pancreatic cells, renal epithelial cells, prostate epithelial cells, breast cells, B lymphocytes, T lymphocytes, lung fibroblasts, melanocytes, dental pulp fibroblasts, mast cells, and enterochromaffin cells and keratinocytes, among others.

The TRPA1 channel has important significance for maintaining normal physiological functions of organisms, regulates membrane potential through sodium, potassium and calcium ion flow, plays an important role in functions of temperature sensing, mechanical sensing, pain sensation, vision, smell, hearing and the like, and a plurality of physiological and pathological processes of asthma, pruritus and the like, and also participates in regulating and controlling cell proliferation and differentiation and cell immune response, and regulates a plurality of pathological processes of respiratory diseases, bladder diseases, gastrointestinal diseases, cardiovascular diseases, skin diseases and the like.

TRPA1 plays an important role in the pathological process of body development of pain and increased pain sensitivity. As the most well-defined cold pain receptor, TRPA1 is involved in the regulation of many different types of pain processes in the body, such as visceral pain, neuropathic pain, migraine, dental pain, etc. It can be activated by various irritant and pro-pain compounds, including noxious low temperatures, intracellular Ca²⁺\\ endogenous substances (e.g., bradykinin), irritant natural substances (e.g., mustard, allicin, and allyl isothiocyanate, AITC), environmental irritants (e.g., acrolein), amphiphilic molecules (e.g., trinitrophenol and chlorpromazine), and pharmaceutical molecules (e.g., URB 597). In addition, amino acid point mutations of TRPA1 cause Pain Syndrome of dominant Familial outbreaks of human autosomal dominant Pain Syndrome (FEPS).

Studies have demonstrated that modulation of TRPA1 is highly correlated with pain relief. Nociception (perception of pain) represents the encoding and processing of noxious stimuli in the nervous system, which are sensed by nociceptive neurons of the nervous system and transmit pain signals. TRPA1 is highly expressed in nociceptive neurons and is activated by a variety of stimuli that cause pain, playing the role of a "chemosensor". Knockout of TRPA1 results in a loss of the ability of the body to sense stimuli, see Zappia K J et al, Eneuro, 2017, 4 (1): ENEURO.0069-16.2017.

Dysfunction of TRPA1 may lead to the development of a variety of human diseases. When the function of TRPA1 is up-regulated, the body can generate uncomfortable symptoms such as pain, pruritus, asthma and the like, and can maintain or even aggravate inflammatory reaction, generate mechanical pain hypersensitivity so as to aggravate peripheral neuropathy and the like, and the down-regulation of the function of TRPA1 can play a therapeutic role in relevant diseases. At the same time, TRPA1 is also a peripheral target, the activity of which in the brain is not essential for pain transmission, and thus therapeutic effects can be achieved even if only peripheral local delivery is blocked.

Many TRPA1 agonists have been shown to cause pain, irritation and neurogenic inflammation in humans and animals, and it is expected that antagonists of TRPA1, as well as substances that block the biological effects of the channel activators, may play a therapeutic role in related diseases, such as pain, itch, inflammation, asthma, cough, and the like.

Thus, modulation against TRPA1 has many industrial and therapeutic uses. For example, antagonists of TRPA1 may fulfill the need in the art for novel analgesic drugs for the treatment and/or prevention of nociceptive and neuropathic pain in mammals, especially humans. Estimated by the World Health Organization (WHO), 6.9 million people die of opioid overdose and 1500 million people are opioid addicts every year worldwide. These drugs have only been used in the past for acute and cancer pain management, but in recent years, the use of large amounts in the relief of non-cancer chronic pain, such as back pain, has led to opioid abuse. Opioid crisis has become a public health incident in the united states, resulting in the loss of tens of thousands of people each year with an annual economic burden exceeding $ 1 trillion. The development of non-opiate and non-addiction substitute drugs is supported to be one of the principles for solving the crisis. Small molecule inhibitors of TRPA1 are one of the effective strategies.

Disclosure of Invention

It is an object of the present invention to provide a highly potent TRPA1 antagonist that can effectively block the binding of TRPA1 to its ligand.

In order to achieve the above objects, the present invention provides an oxazolopyrimidinone amide compound or a pharmaceutically acceptable salt thereof, the oxazolopyrimidinone amide compound has the following structural formula (i):

wherein R is₁，R₂Each independently selected from hydrogen and C_1-3An alkyl group; r₃Is selected from C_1-3An alkyl group; r₄，R₅Each independently selected from 5-6 membered aryl, the aryl is phenyl or an heteroaryl containing 1-3 heteroatoms, the heteroatoms are taken from oxygen or nitrogen atoms, and the positions of the heteroatoms are any positions on the aryl; the aryl group is substituted or unsubstituted.

Preferably, the aromatic group comprises at least one phenyl group, pyridyl group, pyrimidyl group, oxazolyl group, oxadiazolyl group or triazolyl group.

Preferably, R₄Unsubstituted or at least substituted by one or more halogens or C_1-3Alkyl radical, C_1-3Polyhaloalkyl, C_1-3Alkoxy substitution, and/or, R₅Unsubstituted or at least substituted by one or more halogens or C_1-3Alkyl radical, C_1-3Polyhaloalkyl, C_1-3Alkoxy substitution.

The invention also provides a preparation method of the oxazole pyrimidone amide compound or the pharmaceutically acceptable salts thereof, and the reaction route of the method comprises the following steps:

carrying out condensation reaction on an amine compound shown in a formula (II) and a carboxylic acid compound shown in a formula (III) under the action of a coupling agent to prepare a compound shown in a general structural formula (I); or the like, or, alternatively,

the compound of the general structural formula (I) is prepared by subjecting an amine compound of the formula (II) to an aminoacylation reaction with a carboxylic acid derivative of the formula (IIIa).

The invention also provides an application of the oxazole pyrimidone amide compound or the pharmaceutically acceptable salts thereof, wherein the oxazole pyrimidone amide compound or the pharmaceutically acceptable salts thereof is used for preparing a medicine for treating or preventing diseases or symptoms which respond to TRPA1 regulation.

Preferably, the disease or condition includes any one or more of the disorders/conditions associated with various pains, tissue damage, oxidative stress, nicotine addiction, eye irritation, skin irritation, spasticity, pruritus, respiratory diseases, gastrointestinal diseases or urinary system.

Preferably, the pain-related disorder/condition includes, but is not limited to:

any one or any more of various acute pain, chronic pain, mild pain, moderate pain, severe pain, widespread pain, or local pain; and/or the presence of a gas in the gas,

any one or any more of allodynia, dysesthesia, hyperesthesia, hyperalgesia, allodynia, nociceptive pain, inflammatory pain, muscle pain, skeletal pain, post-operative pain, neuropathic pain, pain associated with a neuritic condition, pain associated with cancer; and or (b) a,

chronic tendinitis, complex regional pain syndrome, carpal tunnel syndrome, dental pain, headache, migraine, lumbar pain, back pain, piriformis syndrome, visceral pain, pelvic allergic pain, sciatica pain, menstrual pain, causalgia, cauda equina syndrome, diabetic pain, post-stroke pain, multiple sclerosis pain, ischemic pain, chemoradiotherapy pain.

Preferably, the inflammatory pain includes, but is not limited to, pain associated with arthritis, wherein pain associated with arthritis includes, but is not limited to, rheumatoid arthritis, ankylosing spondylitis, osteoarthritis, reactive arthritis, psoriatic arthritis, traumatic arthritis, infectious arthritis, tumor-associated arthritis, gouty arthritis, pseudogouty arthritis; the neuropathic pain includes, but is not limited to, central pain, central and peripheral neuralgia, central and peripheral neuropathic pain; pain associated with the neuritic condition includes, but is not limited to, pain caused by hypothermia, infection, chemical injury, radiation exposure, disease, or a defective condition.

Preferably, the disorders/conditions associated with respiratory disease include, but are not limited to, asthma, pertussis, chronic obstructive pulmonary disease, bronchiectasis or bronchoconstriction; disorders/conditions associated with gastrointestinal diseases include, but are not limited to, gastroesophageal reflux disease, ulcers, irritable bowel syndrome or inflammatory bowel disease; disorders/pain associated with the urinary system include, but are not limited to, incontinence, micturition disorders, renal colic and cystitis.

The invention also provides a pharmaceutical composition which comprises the oxazole pyrimidone amide compound (I) or the pharmaceutically acceptable salts thereof.

Since TRPA1 is a major molecular site for pain pathway activation and is highly expressed in human primary and peripheral sensory neurons, TRPA1 is a promising new target for pain therapy. The compound of the structural general formula (I) and the acceptable salt thereof provided by the invention can strongly inhibit TRPA1, and can be used for preparing novel analgesic drugs.

There are many reviews of TRPA1 as drug targets, for example, Rech et al (Future Med. chem. 2010: 843-. TRPA1 is highly expressed in a subset of C-fiber nociceptors in the peripheral nervous system (Kobayashi et al 2005, J Comp Neurol 493 (4): 596-. Activated TRPA1 open cation (mainly Ca)²⁺And Na⁺) Enter the cell, depolarize the membrane potential and affect calcium homeostasis in primary afferents. Depolarizing primary nerve endings leads to opening of action potentials, which enhances both hyperalgesia and hyperalgesia (Jiang and Gebhart 1998, Pain 77 (3): 305-13; Cervero and Laird 1996, Pain 68 (1): 13-23). AITC is known to activate TRPA1 concentration-dependently, produce stimulation signals and afferent nerve fibers to trigger pain and hyperalgesia (Reeh 1986, Brain Res 384: 42-50); and are equally effective in topical applications (Namer et al 2005, neuroreport 6 (9): 955-. However, TRPA1 knockout mice had almost lost sensitivity to AITC and showed severe impairment of bradykinin pain response signaling (Kwan 2006, Neuron50 (2): 277-. Formalin can also effectively activate TRPA1, and can be widely used for relieving pain in rodent modelAnd (6) evaluating. Likewise, formalin has little effect on TRPA1 knockout mice (McNamara et al 2007, Proc Natl Acad Sci USA 104 (33): 13525-_。These results demonstrate the feasibility of TRPA1 as a target for pain therapy. A number of TRPA1 small molecule antagonists have been reported demonstrating the effectiveness of TRPA1 antagonist drugs in the treatment of pain. The TRPA1 antagonist from Hydra corporation (HC 030031) significantly reduced the formalin-induced pain response in a dose-dependent manner (W02007/073505 a 2). The compounds of Abbott have an in vitro effect on TRPA1 and show good analgesic efficacy in vivo in a rat osteoarthritis Pain model (US 2009/0176883 and Mc Garaughty et al, Molecular Pain 20106: 14). Several compounds from Glenmark also show significant in vivo efficacy in several whole animal pain models (US 2009/0325987). Therefore, the compound with the structural general formula (I) and the acceptable salt thereof provided by the invention can inhibit TRPA1 in vitro, and can reasonably predict the potential in vivo of the compound as an analgesic drug.

TRPA1 is also known as a "gatekeeper of inflammation". When tissue damage or inflammation occurs, it plays a key role as a nociceptor in conducting pain sensations. Products of tissue injury, inflammation and oxidative stress, such as 4-hydroxynonenal, histamine, substance P, 5-hydroxytryptamine and related substances have all been shown to activate the TRPA1 channel. These findings provide yet another rationale for the treatment of various diseases with small molecule TRPA1 antagonists. These diseases include diseases associated with tissue damage, oxidative stress and smooth muscle contraction, such as asthma, Chronic Obstructive Pulmonary Disease (COPD), spasticity, and pulmonary inflammation, among others.

Asthma is a disease caused by chronic airway inflammation. When the respiratory tract is stimulated by external stimuli such as aldehydes, chlorine, cigarette smoke, etc., TRPA1 in respiratory tract primary sensory neurons is activated, causing Ca²⁺Internal flow, high intracellular concentration of Ca²⁺Can promote the release of substance P, calcitonin gene-related peptide and neurokinin A, etc. (Yang H, et al, Medical Science Monitor International Medical Journal of Experimental)& Clinical Research, 2016, 22: 2917-2923). These substances act on effector cells of the respiratory tract to cause bronchoconstriction, and lymphocytes release inflammatory factors, thereby causing symptoms such as cough and bronchospasm. TRPA1 has also been shown to be a "switch" for coughing. TRPA1 is widely distributed on nerve endings on the surface of the lung, and can be activated by a stimulus which enters the lung along with air, so that cough is caused, and therefore, the antagonism of TRPA1 is helpful for treating diseases related to chronic cough and the like. When inflammation occurs in the bladder, overactive bladder such as frequent urination and urgent urination may occur, and pain may accompany the bladder. It has been shown that down-regulation of TRPA1 function significantly reduces overactive bladder and pain caused by interstitial cystitis (Chen D et al, relative Neuroscience, 2016,7 (1): 133-138). In addition, TRPA1 plays an important role in cardiovascular and gastrointestinal regulation (Pan Y et al, Pflugers Archiv-European Journal of Physiology, 2017: 1-8, Fothergill J et al, Nutrients, 2016, 8 (10): 623). Therefore, the compounds of the general structural formula (I) and acceptable salts thereof provided by the invention can be used for preparing medicines for treating or preventing tissue damage, oxidative stress, respiratory diseases, gastrointestinal diseases or related disorders/symptoms of the urinary system.

Functional upregulation of TRPA1 also induces pruritus, such as chronic pruritus caused by allergic dermatitis (Fernandes E S et al, fast Journal of the Publication of American society for Experimental Biology, 2013, 27 (4): 1664). Itching is mainly classified into histamine dependence and non-histamine dependence, and the former can be treated by anti-histamine drugs. TRPA1 plays an important role in histamine-independent pruritus. TRPA1 has been shown to mediate pruritus caused by BAM8-22, SLIGRL, IL-31, etc. (Roberson D P et al, Nature Neuroscience, 2013, 16 (7): 910; Cevikbas F et al, Journal of Allergy & Clinical Immunology, 2014, 133 (2): 448 and 460), and in particular TRPA1 knockout mice were found to have a marked reduction in the intense pruritus caused by chloroquine. In addition, TRPA1 is expressed in both pain and itch sensory neurons of the eye and plays an important role in the non-histamine itch pathway of itching of the eye. Therefore, the compound with the structural general formula (I) and the acceptable salt thereof can be used for preparing medicines for treating or preventing pruritus and eye irritation.

TRPA1 can also be activated by nicotine and its structural analogs. Certain smokers experience skin irritation and even allergy when using nicotine replacement therapy to quit smoking, indicating an association with the TRPA1 channel. Therefore, the compound with the structural general formula (I) and the acceptable salt thereof provided by the invention can be used for preparing medicines for treating or preventing nicotine addiction and skin irritation.

Briefly, the compounds of the present invention are TRPA1 antagonists useful in the treatment or prevention of any disorder, condition, or disease mediated by or associated with the activity of TRPA1, including any one or more of the disorders/conditions associated with various forms of pain, tissue damage, oxidative stress, nicotine addiction, ocular irritation, skin irritation, spasticity, pruritis, respiratory disorders, gastrointestinal disorders, or urinary system disorders.

In the context of the present invention, the various pain conditions/disorders include, but are not limited to, various acute pain, chronic pain, mild pain, moderate pain, severe pain, widespread pain, localized pain; such as allodynia, dysesthesia, hyperesthesia, hyperalgesia, allodynia, nociceptive pain, inflammatory pain, muscle pain, skeletal pain, postoperative pain, neuropathic pain, pain associated with a neuritic condition, pain associated with cancer. Specifically, including but not limited to chronic tendinitis, complex regional pain syndrome, carpal tunnel syndrome, dental pain, headache, migraine, lumbar pain, back pain, piriformis syndrome, visceral pain, pelvic allergic pain, sciatica pain, menstrual pain, causalgia, cauda equina syndrome, pain associated with arthritis, including but not limited to rheumatoid arthritis, ankylosing spondylitis, osteoarthritis, reactive arthritis, psoriatic arthritis, traumatic arthritis, infectious arthritis, tumor-associated arthritis, gouty arthritis, pseudogouty arthritis, bladder disease-related disorders/pain, including but not limited to incontinence, micturition disorders, renal colic, and cystitis, and including but not limited to pain associated with various neuritic disorders resulting from hypothermia, infection, chemical injury, radiation exposure, disease or deficiency disorder, such as hypothyroidism, fibromyalgia, referred pain.

In the context of the present invention, neuropathic pain includes, but is not limited to: central pain, central and peripheral neuropathic pain, such as pain associated with autonomic neuropathy, Peripheral Nervous System (PNS) injury or Central Nervous System (CNS) injury or disease, polyneuropathy (e.g., Diabetic Peripheral Neuropathy (DPN) and chemotherapy-induced neuropathy) pain, polyradiculopathies of the cervical, lumbar or sciatica type, post-herpetic neuralgia, spinal cord injury pain, fibromyalgia, post-stroke pain, multiple sclerosis pain, ischemic pain, chronic musculoskeletal pain, Chronic Inflammatory Demyelinating Polyneuropathy (CIDP), vasculitic neuropathy, mechanical hyperalgesia, and cold allodynia.

Compounds of formula (I), including but not limited to those specified in the examples, solvates, pharmaceutically acceptable salts, prodrugs, salts of prodrugs, or any combination thereof may be used as described in Bautista, d, et al, Cell 2006, 124, 1269-1282; trevisani, M.et al, Proceedings of the National Academy of Sciences USA 2007, 104, 13519-; dai, Y, et al, Journal of Clinical Investigation 2007, 117, 1979-; diogens, A. et al, Journal of Dental Research 2007, 86, 550-555; katsura, H. et al, Journal of Neurochemistry 2007, 102, 16 show treatment or prevention of inflammatory, nociceptive and neuropathic pain.

Compounds of formula (I), including but not limited to those specified in the examples, solvates, pharmaceutically acceptable salts, prodrugs, salts of prodrugs, or any combination thereof, are useful for treating or preventing Inflammatory Bowel Disease (IBD), such as esophagitis, colitis and crohn's disease, as indicated by Kimball, e.s. et al, neuro astrology & Motility 2007, 19, 90-400.

Compounds of formula (I), including but not limited to those specified in the examples, solvates, pharmaceutically acceptable salts, prodrugs, salts of prodrugs, or any combination thereof, are useful for treating or preventing gastrointestinal disorders such as gastroesophageal reflux disease (GERD), ulcers, Irritable Bowel Syndrome (IBS), as shown by Penuelas, a. et al, European Journal of Pharmacology 2007, 576, 143-.

Compounds of formula (I), including but not limited to those specified in the examples, solvates, pharmaceutically acceptable salts, prodrugs, salts of prodrugs, or any combination thereof, are useful, for example, in Andre et al, Journal of Clinical Investigation 2008, 118, 2574-2582; bessac et al, Journal of Clinical investment 2008, 118, 1899-; and Simon and Liedtke, Journal of Clinical Investigation 2008, 118, 2383-.

Compounds of formula (I), including but not limited to those specified in the examples, solvates, pharmaceutically acceptable salts, prodrugs, salts of prodrugs, or any combination thereof, are useful for treating or preventing cold hyperalgesia or cold sensitivity as shown in Story, g.m. Current Neuropharmacology 2006, 4, 183-196 and Story, g.m. and Gereau, r.w. Neuron 2006, 50, 177-180.

The compounds of the present invention may be used in the form of pharmaceutically acceptable salts. The term "pharmaceutically acceptable salt" refers to salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit/risk ratio.

Pharmaceutically acceptable salts are well known in the art. For example, S.M. Berge et al describe in detail pharmaceutically acceptable salts in (J. Pharmaceutical Sciences, 1977, 66: let seq).

The compounds of the present invention contain basic functional groups and can be converted into pharmaceutically acceptable salts using suitable acids, if desired. The salts may be prepared in situ during the final isolation and purification of the compounds of the invention.

Acids used to form pharmaceutically acceptable salts include inorganic acids such as hydrochloric acid, hydrobromic acid, sulfuric acid, phosphoric acid and the like, and organic acids such as acetic acid, fumaric acid, maleic acid, 4-methylbenzenesulfonic acid, succinic acid, citric acid and the like. Representative acid addition salts include, but are not limited to: acetate, adipate, citrate, aspartate, benzoate, benzenesulfonate, bisulfate, butyrate, camphorate, camphorsulfonate, digluconate, glycerophosphate, hemisulfate, heptanoate, hexanoate, fumarate, hydrochloride, hydrobromide, hydroiodide, 2-hydroxyethanesulfonate (isethionate), lactate, malate, maleate, methanesulfonate, nicotinate, 2-naphthalenesulfonate, oxalate, palmitate (palmitate), pectate (pectate), persulfate, 3-phenylpropionate, picrate, pivalate, propionate, succinate, tartrate, thiolate, phosphate, glutamate, bicarbonate, p-toluenesulfonate, and undecanoate.

The compounds of the invention may be used in the manufacture of a medicament for the treatment or prevention of any of the diseases disclosed herein. The compounds and pharmaceutical compositions described herein are useful for modulating the TRPA1 ion channel, where modulation is believed to be associated with various disease states.

The compounds of the present invention are useful in pharmaceutical compositions containing at least one compound described herein.

In conclusion, the invention provides a novel oxazolopyrimidinone amide compound as a TRPA1 antagonist. In particular, the compounds described herein are suitable for the treatment or prevention of diseases, conditions and/or disorders modulated by TRPA1, such as various types of pain. The invention also provides methods of making the compounds described herein, and intermediates used in the synthesis.

The invention has the beneficial effects that: provides a novel oxazolopyrimidinone amide compound which has obvious antagonism on TRPA1 channel, and can be used for preparing medicaments or pharmaceutical compositions for diseases or symptoms which are responsive to TRPA1 antagonism, especially for treating or preventing various disorders/symptoms related to pain, tissue injury, oxidative stress or smooth muscle contraction.

Detailed Description

The compounds described in the present invention may form salts, and non-limiting examples of pharmaceutically acceptable salts that form part of the present invention include salts of inorganic acids, salts of organic acids, salts of natural amino acids, and salts of unnatural amino acids.

Certain compounds of the general formula (I) of the present invention can exist in stereoisomeric forms (e.g., diastereomers and/or enantiomers). The present invention includes these stereoisomeric forms (including diastereomers and/or enantiomers) and mixtures thereof. The various stereoisomeric forms of the compounds of the invention may be separated from each other by methods known in the art, or a given isomer may be obtained by stereospecific or asymmetric synthesis. Tautomeric forms and mixtures of the compounds of formula (1) of the invention are also contemplated.

General preparation method

The compounds described herein are prepared using techniques known to those skilled in the art, such as the preparative routes shown in scheme one, scheme two and scheme three, as well as by other methods. Wherein R is₁、R₂、R₃、R₄、R₅As defined in the specification. Furthermore, reference to specific acids, bases, reagents, coupling agents, solvents, etc., in the following synthetic schemes should be understood that other suitable acids, bases, reagents, coupling agents, solvents, etc., may be used and are included within the scope of the invention. The compounds obtained by using the general reaction scheme may not be sufficiently pure, and these compounds may be purified by using any method for purifying organic compounds known to those skilled in the art, for example, crystallization or silica gel or alumina column chromatography using different solvents in suitable proportions.

All ratios and percentages referred to herein are by volume.

The general technique for the synthesis of the target compounds of the oxazolopyrimidinone amide class (I) is shown in scheme one:

scheme one

According to the first synthetic route, an amino-containing oxazolopyrimidinone amine derivative intermediate shown in the general formula (II) and a carboxylic acid derivative intermediate shown in the general formula (III) are subjected to condensation reaction in the presence of a suitable peptide coupling agent, a suitable solvent and a suitable base to generate an oxazolopyrimidinone amide target compound shown in the general formula (I). For example, a general coupling method is to stir the oxazolopyrimidinone amine intermediate (II) and the appropriate arylcarboxylic acid intermediate (III) in an aprotic solvent such as Dichloromethane (DCM), N-Dimethylformamide (DMF) or the like, which does not adversely affect the reaction, in the presence or absence of an additive such as 1-hydroxybenzotriazole (HOBt), in the presence or absence of a condensing agent represented by 1-ethyl-3- (3' -dimethylaminopropyl) carbodiimide hydrochloride (EDCI), in the presence or absence of a base such as triethylamine or the like, overnight at room temperature or with heating. The reaction solution is diluted by water and extracted by ethyl acetate, washed by water and saturated saline in turn, dried by magnesium sulfate, filtered, concentrated, recrystallized or purified by silica gel column chromatography, and eluted by ethyl acetate and hexane to obtain the coupling condensation target amide product (I).

Alternatively, the carboxylic acid intermediate of formula (III) is converted to the corresponding acid chloride intermediate (IIIa) by an acylating agent such as sulfuryl chloride, thionyl chloride, etc. in an aprotic solvent such as DCM, DMF, etc. which does not adversely affect the reaction, and then subjected to an aminoacylation reaction with the amino group-containing oxazolopyrimidinone amine derivative (II) to obtain the target amide compound (I).

Part of the amine derivative intermediate (II) used in the synthesis can be obtained commercially, for example, 6-amino-7-isopropyl-2, 3-dimethyl-5H-oxazolo [3,2-a ] pyrimidin-5-one (II-1); or prepared by the method described in scheme two. The partial carboxylic acid derivative of formula (III) can be obtained by commercially available methods, such as 6- (2, 6-difluorophenyl) -5-fluoropicolinic acid (III-4), 6- (2-fluorophenyl) picolinic acid (III-5), 6- (2, 3-difluorophenyl) picolinic acid (III-6), 6- (2, 4-difluorophenyl) picolinic acid (III-7), 6- (2, 5-difluorophenyl) picolinic acid (III-8), 6- (3-fluoro-4-methylphenyl) picolinic acid (III-9), or prepared by the methods described in scheme three.

The peptide coupling agent used is selected from, but not limited to, the following agents: n, N ' -Dicyclohexylcarbodiimide (DCC), 1-ethyl-3- (3 ' -dimethylaminopropyl) carbodiimide (WSC), Diisopropylcarbodiimide (DIC), 1-ethyl-3- (3 ' -dimethylaminopropyl) carbodiimide hydrochloride (EDCI), 1-hydroxybenzotriazole (HOBt), 4-Dimethylaminopyridine (DMAP), 1-benzotriazolyloxytris (dimethylamino) hexafluorophosphate (BOP), N- [ 1-H-benzotriazol-1-yl ] (dimethylamino) methylene ] -N-methylmethanium (methanaminium) -hexafluorophosphate-N-oxide (HBTU), N- [ (1-H-benzotriazol) dimethylamino ] methylene ] N-methylmethanium tetrafluoroborate-N-oxide (TBTU), O- (benzotriazol-1-yl) -1, 3-dimethyl-1, 3-dimethyleneurea Hexafluorophosphate (HBMDU), O- (benzotriazol-1-yl) -1, 1, 3, 3-bis (tetramethylene) urea hexafluorophosphate (HBPyU), O- (benzotriazol-1-yl) -1, 1, 3, 3-bis (pentamethylene) urea hexafluorophosphate (HBPipU), 3-hydroxy-4-oxo-3, 4-dihydro-1, 2, 3-benzotriazine (HODhbt), O- (3, 4-dihydro-4-oxo-1, 2, 3-benzotriazin-3-yl) -1, 1, 3, 3-tetramethyluronium hexafluorophosphate, O- (3, 4-dihydro-4-oxo-1, 2, 3-benzotriazin-3-yl) -1, 1, 3, 1-tetramethyluronium tetrafluoroborate, [3, 4-dihydro-4-oxo-1, 2, 3-benzotriazin-3-yl ] oxy ] tris (pyrrolidinyl) hexafluorophosphate, 1-hydroxy-7-azabenzotriazole (HAPyU).

Suitable solvents include, but are not limited to, DCM, dichloroethane, Tetrahydrofuran (THF), ethyl acetate, ethanol, DMF and the like which do not interfere with the reaction.

Suitable bases include, but are not limited to: triethylamine, pyridine, piperidine, and the like.

The general technique for the synthesis of oxazolopyrimidinone amine derivatives (II) is shown in scheme two.

Scheme two

As shown in the second synthetic route, the chloro-ketone derivative shown in formula (IIa) is condensed with urea to form an amino oxazole derivative shown in formula (IIb), and then cyclized with ethyl acetoacetate shown in formula (IIc) to form an oxazolopyrimidinone derivative shown in formula (IId), and the oxazolopyrimidinone nitro derivative shown in formula (IIe) is obtained by nitration under the action of concentrated sulfuric acid and concentrated nitric acid, and then the nitro is reduced to amino by using a suitable reducing agent, such as metallic iron or zinc, to obtain the amino-containing oxazolopyrimidinone amine derivative (II).

The general technique for the synthesis of the aromatic carboxylic acid intermediate derivative (III) is shown in scheme three.

Scheme three

Containing R₄And R₅The intermediate (IIIb) of (1) can be prepared by a conventional method of organic synthesis. For example, R₄And R₅Covalent bonds between can be generated by a number of effective metal-catalyzed coupling methods, such as the method known as Suzuki coupling. In this embodiment, either of the two groups to be attached may be a boronic acid/boronic ester or a halogen/pseudo-halogen, while the corresponding other group is a halogen/pseudo-halogen or boronic acid/boronic ester, respectively. The coupling condensation is carried out under conditions comprising heating the boronic acid derivative and the aryl halide compound in the presence of an organic or inorganic base in a polar solvent mixture and using a palladium catalyst.

In the following synthetic schemes, -LG represents a suitable leaving group, such as a chlorine atom, a bromine atom, an iodine atom, a trifluoromethanesulfonyloxy group, etc.; -B (OR)₂represents-B (OH)₂Or suitable boronic acid derivatives (IVb, IVd, Vb) such as pyrocatechol borane, pinacol borane and N-methyliminodiacetic acid borate.

As shown in scheme III, R containing a suitable leaving group is reacted in a suitable solvent₄Derivatives (IVa) and corresponding compounds containing R₅The boric acid derivative (Vb) is subjected to condensation reaction under the action of alkali and a transition metal catalyst to generate an aryl derivative intermediate (IIIb). In the reaction, it may be added or not addedAdding metal salt or metal ion complexing ligand.

As shown in scheme IV, R with a suitable leaving group can also be reacted in a suitable solvent₅Derivative (Va) and corresponding R₄The boric acid derivative (IVb) generates condensation reaction under the action of alkali and transition metal catalyst to generate an aryl derivative intermediate (IIIb). In the reaction, a metal salt or a metal ion complexing ligand may or may not be added.

As shown in scheme five, R which contains a carboxylate group and at the same time has a suitable leaving group can also be reacted in a suitable solvent₄Derivative (IVc) or R having a carboxylate group₄Boronic acid derivatives (IVd), each with the corresponding R₅Boronic acid derivative (Vb) or R containing a suitable leaving group₅And (5) carrying out coupling condensation reaction on the derivative (Va) under the action of alkali and a transition metal catalyst to generate an aryl carboxylic ester intermediate (IIIc). The reaction may be carried out with or without addition of metal salts or metal ion complexing ligands. Then ester group hydrolysis is carried out under the action of acid or alkali to generate aromatic carboxylic acid intermediate (III). The acid used for hydrolysis includes inorganic or organic acids such as hydrochloric acid, sulfuric acid, trifluoroacetic acid, etc.; the base includes inorganic or organic bases such as LiOH, NaOH, piperidine, pyridine, and the like.

In scheme three, scheme four, and scheme five, suitable solvents for the coupling condensation reaction include, but are not limited to, solvents that do not interfere with the reaction, such as 1, 4-dioxane, toluene, tetrahydrofuran, acetonitrile, or butanol, with or without the addition of co-solvents such as water; the base used includes organic or inorganic bases such as triethylamine, pyridine, sodium hydroxide, sodium carbonate, potassium carbonate, sodium hydrogencarbonate, potassium hydrogencarbonate, tripotassium phosphate and the like; catalysts include, but are not limited to [1, 1' -bis (diphenylphosphino) ferrocene ] dichloropalladium (II), tris (dibenzylideneacetone) dipalladium (0), tetrakis (triphenylphosphine) palladium (0), palladium (II) acetate, and the like; metal salts with or without addition include, but are not limited to, copper acetate; metal ion complexing ligands with or without addition include, but are not limited to, 2,4, 6-triisopropyl-2' - (dicyclohexylphosphino) biphenyl.

As shown in scheme six, the intermediate derivative (IIId) containing a methyl group on the aromatic ring is used in a strong oxidizing agent such as KMnO₄Can be converted into the corresponding aromatic carboxylic acid intermediate (III).

General procedure for the preparation of intermediate (II): taking preparation II-1 as an example, the general procedure is illustrated as follows:

example 1

Step 1: preparation of IIb-1

Urea (0.2 mol) was added to 150 mL of 2-propanol containing 3-chloro-butyl-2-one (22 g, 0.2 mol), stirred under reflux for 5 minutes, the precipitate was washed with 2-propanol and dissolved in water, adjusted to pH =11 with sodium carbonate solution, extracted with dichloromethane, dried over sodium sulfate, and the solvent was recovered under reduced pressure, and the resulting product lib-1 was used in the next reaction without purification. Yield: 52 percent. MS m/z (ESI): 115.1 [ M +1 ]]⁺。

Step 2: preparation of IId-1

Stirring a mixed solution of 85% phosphoric acid (10 g, 0.09 mol) and anhydrous phosphoric acid (12 g, 0.09 mol) at 120 ℃ for 2 hours, cooling the prepared polyphosphoric acid to 35 ℃, slowly dropwise adding IIc-1 (0.05 mol), stirring for 10 minutes, adding 0.05mol IIb-1, slowly heating the mixed solution to 120 ℃ and 130 ℃, and stirring for 4 hours. Cooling, adding 100 ml of water, quenching, adjusting pH to 5-6 with sodium carbonate solution, collecting precipitate, washing with water, and recrystallizing to obtain IId-1. The yield thereof was found to be 56%. MS m/z (ESI): 207.1 [ M +1 ]]⁺。

And step 3: preparation of IIe-1

IId-1 (0.05 mol) is dissolved in 40 mL of 96% sulfuric acid at the temperature of 10-15 ℃, 97% nitric acid (0.09 mol) is slowly dropped into the solution while stirring, and the temperature is slowly raised to 23-25 ℃ for reaction for 1 hour. And pouring the reaction solution into ice cubes, collecting precipitates, washing with water and recrystallizing to obtain IIe-1. The yield thereof was found to be 30%. MS m/z (ESI): 252.1 [ M +1 ]]⁺。

And 4, step 4: preparation of (II-1) intermediate

To a three-necked flask, 500 mL of 2-propanol and 120 mL of water were added, and under vigorous stirring, 20 g (0.35 mol) of reduced iron powder was added, followed by slowly dropping 6 g (0.6 mol) of 35% hydrochloric acid. The solution was heated to 75 ℃ for 5 minutes, then cooled to room temperature, IIe-1 (0.09 mol) was added, 12.3 g (0.12 mol) of 35% hydrochloric acid was slowly added dropwise, the temperature was raised to 75 to 80 ℃ and stirred for 4 hours, 9.5 g (0.09 mol) of saturated sodium carbonate solution was added, and stirring was continued at 75 to 80 ℃ for 1 hour. The precipitate was filtered while hot and washed successively with hot 2-propanol and chloroform. The washing liquid and the filtrate are combined and the solvent is recovered under reduced pressure10-15 mL of mother liquor is remained, chloroform extraction is carried out, sodium sulfate is dried, the solvent is recovered under reduced pressure until the solvent is dried, and the residue is recrystallized by 2-propanol to obtain II-1. The yield thereof was found to be 65%. MS m/z (ESI): 222.1 [ M +1 ]]⁺。

The other intermediates (II-2) to (II-7) were prepared in a similar manner to that described above, optionally using the respective starting materials.

Table 1: structure and structural identification data of selected intermediate (II)

General procedure for the preparation of intermediate (III), representative preparation of which is shown in scheme seven. Using preparation III-1 as an example, the general preparation procedure is illustrated below.

Example 2

Synthesis of intermediate III-1

Lithiation is carried out on 5-methyl-1, 3 difluorobenzene, then the lithiated 5-methyl-1, 3 difluorobenzene and 2-isopropoxy-4, 4, 5, 5-tetramethyl-1, 3, 2-dioxaborolane are reacted to obtain borate, then the borate and bromofluoropyridine formate (IVc-1) are subjected to Suzuki coupling under the catalysis of palladium to obtain an aryl carboxylic ester intermediate (IIIc-1), and then ester hydrolysis is carried out under alkaline conditions to obtain an aryl formic acid intermediate (III-1). The dosage of alkali and the reaction time in the hydrolysis are adjusted according to different reactants.

Step 1: synthesis of 6-bromo-5-fluoropicolinic acid

2-bromo-3-fluoro-6-methylpyridine (1.0 eq.) and potassium permanganate (1.0 eq.) were added to 30 mL of water and heated at 100 ℃ for 5 hoursThen 1.0 equivalent of potassium permanganate is added, and heating is continued for 48 hours. The reaction solution was filtered through celite and washed with water. The aqueous phases were combined and acidified to pH =4 with 1N hydrochloric acid, extracted with ethyl acetate (200 mL), washed with saturated brine, dried over magnesium sulfate, filtered and concentrated to give 6-bromo-5-fluoropicolinic acid as a white solid in 15% yield. MS m/z (ESI): 219.9 [ M +1 ]]⁺。

Step 2: synthesis of methyl 6-bromo-5-fluoropicolinate (IVc-1)

Sulfuric acid (4.2 equiv.) is added to a methanol solution (0.2M) of 6-bromo-5-fluoropicolinic acid (1.0 equiv.), stirred at room temperature for 2 hours, diluted with a proper amount of ethyl acetate, and slowly added with a saturated aqueous solution of sodium bicarbonate to quench the reaction. Extraction with ethyl acetate and drying of the organic phase over magnesium sulfate, filtration and concentration in vacuo gave IVc-1 as a white solid. The yield thereof was found to be 94%. MS m/z (ESI): 233.9 [ M +1 ]]⁺。

And step 3: synthesis of Vb-1

To a solution of 1, 3-difluoro-5-methylbenzene (1.0 eq, 0.2M) in dry THF at-78 deg.C under a nitrogen atmosphere was slowly added n-butyllithium (1 eq, 1.6M in hexanes). After stirring at-78 ℃ for 2 hours, 2-isopropoxy-4, 4, 5, 5-tetramethyl-1, 3, 2-dioxaborolane (1.15 eq.) was added and the reaction was allowed to warm slowly to room temperature. After the reaction, saturated aqueous sodium bicarbonate solution was added to quench the reaction, and ethyl acetate extraction was performed. The organic phase was washed with saturated brine, dried over sodium sulfate, filtered and concentrated to give Vb-1 as a white solid with a yield of 89%.¹HNMR（400MHz，CDCl₃δppm 6.63（dd，J= 9.35，0.76 Hz，2H），2.36（s，3H），1.38（s，12H）, MS m/z（ESI）：255.1 [M+1]⁺。

And 4, step 4: synthesis of methyl 6- (2, 6-difluoro-4-methylphenyl) -5-fluoropicolinate (IIIc-1)

To a THF/water (10: 1) mixed solution of IVc-1 (1.0 equiv., 0.1M) were added Vb-1 (1.75 equiv.) and potassium fluoride (3.3 equiv.). The reaction is degassed by nitrogen and added with Pd₂（dba）₃(0.25 eq.) and tri-isobutylphosphine (0.5 eq.) and warmed to 80 ℃ for 1 hour. Cool to room temperature, concentrate in vacuo and purify on silica gel column eluting with ethyl acetate/hexane (0% -30% ethyl acetate) to give IIIc-1 as a white solid in 81% yield. MS m/z (ESI): 282.0 [ M +1 ]]⁺。

And 5: synthesis of 6- (2, 6-difluoro-4-methylphenyl) -5-fluoropicolinic acid (III-1):

to a solution of IIIc-1 (1.0 equiv., 0.1M) in THF was added an aqueous solution of LiOH (5.5 equiv., 2M) and the mixture was stirred at room temperature for 4 hours. The organic solvent was distilled off under reduced pressure, and the residual aqueous phase was acidified with 2N hydrochloric acid to pH = 4. The precipitate was collected by filtration and dried to give III-1 as a pale yellow solid in a yield of 72%.

Example 3

Synthesis of intermediate III-2

Step 1: synthesis of methyl 6- (2, 6-difluoro-4-methoxyphenyl) -5-fluoropicolinate (IIIc-2)

To a THF/water (10: 1) mixed solution of IVc-1 (1.0 equiv., 0.1M) were added 2, 6-difluoro-4-methoxyphenylboronic acid Vb-2 (2.5 equiv.) and potassium fluoride (3.3 equiv.). Under the protection of nitrogen, adding Pd₂（dba）₃(0.25 equiv.) and tri-isobutylphosphine (0.5 equiv.), maintained at 80 ℃ for 1 hour. Cooling to room temperature, vacuum concentrating, purifying with silica gel column, and adding ethyl acetateHexane (0% -30% ethyl acetate) elution gave IIIc-2 as a white solid in 85% yield. MS m/z (ESI): 298.1 [ M +1 ]]⁺。

Step 2: synthesis of 6- (2, 6-difluoro-4-methoxyphenyl) -5-fluoropicolinic acid (III-2)

To a mixed solution of IIIc-2 (1.0 equiv., 0.09M) in THF/MeOH (2: l) was added an aqueous LiOH solution (l.5 equiv.), stirred at room temperature for 1 hour and then quenched with 1N hydrochloric acid, extracted with ethyl acetate, washed with saturated brine, dried over sodium sulfate, filtered, and concentrated to give III-2 in 86% yield.

Example 4

Synthesis of intermediate 2- (2, 6-difluorophenyl) pyrimidine-4-carboxylic acid (III-3)

2-Chloropyrimidine-4-carboxylic acid (1.0 eq.) is dissolved in appropriate amounts of DMF and 2M Na₂CO₃To the mixed solution were added 2, 6-difluorophenylboronic acid Vb-3 (1.3 eq.) and Pd (dppf) Cl₂DCM (0.05 eq.) was heated in a microwave reactor for 30 min at 120 ℃. The mixture was diluted with ethyl acetate and 1N aqueous NaOH solution was added. The organic phase was extracted three times with 1N NaOH and once with 6N NaOH. The aqueous phases were combined and acidified to pH =1 by addition of concentrated hydrochloric acid and extracted with ethyl acetate. The organic layer was dried over magnesium sulfate, filtered, and concentrated to give III-3 in 78% yield.

Example 5

Synthesis of intermediate 2- (2, 6-difluorophenyl) -3-fluoro-6-methylpyridine (III-4)

Step 1: synthesis of IIId-4

To a solution of 2-bromo-3-fluoro-6-methylpyridine (1.0 eq, 0.1M) in a THF/water (10: 1) mixture was added 2, 6-difluorophenylboronic acid Vb-3 (2.0 eq) and potassium fluoride (3.3 eq). The reaction was degassed for 10 minutes and Pd was added₂（dba）₃(0.05 eq.) and tri-tert-butylphosphine (0.1 eq.) and stirred at 60 ℃ for 1 hour. Cooled to room temperature, extracted with ethyl acetate, dried over sodium sulfate, filtered and concentrated. The residue was diluted to 0.1M with ethanol and 0.5 eq NaBH added₄To reduce dba. Stir at room temperature for 1 hour, quench with water, and concentrate under vacuum to remove ethanol. Extraction with ether, washing with saturated brine, drying the organic phase over sodium sulfate, filtration and concentration in vacuo. The residue was purified by silica gel column chromatography eluting with hexane and ethyl acetate (0% to 10% ethyl acetate). IIId-4 was obtained as a pale yellow oil in 82% yield. MS m/z (ESI): 224.1 [ M +1 ]]⁺。

Step 2: synthesis of 6- (2, 6-difluorophenyl) -5-fluoropicolinic acid (III-4)

KMnO was added to an aqueous solution of IIId-4 (1.0 eq.) in water₄(2.0 eq.) and heated to reflux for 10 hours. 2.0 equivalents of KMnO were added₄Stirring under reflux was continued for 8 hours. The solution was cooled to room temperature, filtered through celite, and washed with water. The filtrate was acidified with 6N hydrochloric acid to pH =3 and a white precipitate was collected. The filtrate was further acidified to pH =1 and the precipitate was collected by filtration again. The filtrate was extracted with ethyl acetate, washed with saturated brine, dried over magnesium sulfate, filtered, and concentrated. The residue was dissolved in ethyl acetate, washed with 1N aqueous NaOH, and the aqueous layer was acidified to pH =1 to collect a white precipitate. The solids were combined to give the product III-4 as a white solid in 32% yield.

Structural information and characterization data for selected intermediate (III) are shown in table 2.

Table 2: structure and structural identification data for selected intermediates (III)

Preparation of the Compounds of the general formula (I)

Exemplary examples of the preferred embodiments described herein are synthesized using scheme one described herein to couple oxazolopyrimidinone amine intermediate (II) with the appropriate aryl carboxylic acid intermediate (III) or aryl carboxylic acid derivative intermediate (IIIa), or using other methods known in the art, with reference to the following examples.

Examples 6-14 (method a):

the appropriate amine intermediate (II) (1.0 equiv., 0.05M) was dissolved in the DMF/ethanol (1/5) mixed solution and the appropriate picolinic acid derivative (III) (1.3 equiv.), HOBt (1.3 equiv.) and EDCI (1.3 equiv.) were added. The mixture was stirred at room temperature for 6 hours, extracted with ethyl acetate, washed with water, 1N NaOH and saturated brine in that order, dried over magnesium sulfate, filtered, concentrated, the residue was dissolved in ethanol, and Cs was added₂CO₃(1.0 eq.) and stirred at 60 ℃ for 2 hours. Distilling under reduced pressure to remove ethanol, diluting with water, and extracting with ethyl acetate. The organic phase was washed with saturated brine, dried over magnesium sulfate, filtered and concentrated to give the product.

The target compounds I-1, I-2, I-3, I-4, I-5, I-6, I-7, I-8 and I-9 are synthesized by a method similar to the method A.

Examples 15 to 21 (method B)：

To a solution of the carboxylic acid derivative intermediate (III) (1.0 eq) in 1, 2-dichloroethane were added EDCI (1.2 eq), HOBt (0.3 eq) and 4-dimethylaminopyridine (DMAP, 0.l eq) and the mixture was stirred at room temperature for 10-15 minutes. The appropriate amine intermediate (II) (1.0 eq) was added and stirring continued at room temperature for 48 hours. Concentrated under reduced pressure, and methanol was added to the residue, followed by stirring at room temperature for 30 minutes. Filtering to collect solid, and recrystallizing and purifying by isopropanol or methanol to obtain the target product.

Synthesizing the target compounds I-10, I-11, I-12, I-13, I-14, I-15 and I-16 by a method similar to the method B.

Examples 22 to 27 (method C)：

Carboxylic acid derivative intermediate (III) (1.0 equiv.) was added to a THF/DMF (3: 1) mixture, EDCI (2.0 equiv.) was added with stirring, and stirring was continued for 30 minutes. The appropriate amine intermediate (II) (1.0 equiv.) and DMAP (0.2 equiv.) were added and stirred at 80 ℃ for 24 hours. Most of the THF was evaporated under reduced pressure and the residue was purified by recrystallization or by using silica gel column chromatography (methanol-chloroform elution).

Synthesizing the target compounds I-17, I-18, I-22, I-23, I-24 and I-25 according to the method C.

Examples 28 to 30 (method D)：

The appropriate amine intermediate (II) (1.0 equiv.) is added to dry toluene, the carboxylic acid halide intermediate (IIIa) (1.0 equiv.) is added with stirring, and the mixture is stirred at room temperature for 30 minutes and then heated under reflux for an additional 2 hours. After cooling, it was purified by recrystallization or silica gel column chromatography using methanol-chloroform as an eluent.

Synthesizing the target compounds I-19, I-20 and I-21 according to the method D.

Table 3: structure and Structure identification data of Compound (I) of some examples

Antagonistic Activity of the Compounds of the present invention against TRPA1

Binding of TRPA1 to its agonist causes the ion channel to activate and open, causing intracellular Ca²⁺The concentration increases sharply, and the addition of TRPA1 antagonist will cause Ca²⁺The concentration is reduced. Thus by monitoring Ca in cells²⁺At the same time, the antagonistic activity of the test compound against TRPA1 channel can be detected.

In actual tests, to detect Ca²⁺The change in concentration requires the loading of Ca to the cells²⁺Sensitive fluorescent dye to cause a change in fluorescence in the cell with Ca therein²⁺The concentration changes correspondingly. Since TRPA1 antagonists inhibit Ca induced by the addition of TRPA1 agonists²⁺And (4) flowing inwards, so that the fluorescence in the cells is reduced or even not generated. By detecting the change of fluorescence in the cells, Ca can be reflected indirectly²⁺The change in concentration, thereby calculating the antagonistic activity of the test compound against the TRPA1 channel.

hTRPA1 cell culture

The human TRPA1 gene was cloned into pT-REx-Dest30 inducible vector and stably transfected into T-Rex-293 cells (purchased from Invitrogen). hTRPA 1/HEK 293 cells (hereinafter "hTRPA 1 cells") were maintained under standard sterile cell culture conditions. The culture medium was DMEM (Gibco BRL, Invitrogen) containing 0.5 g/L geneticin (Gibco), 5 mg/L blasticidin (Invitrogen), 14.6 g/L-glutamine (200 mM; Gibco), 5 g/L penicillin/streptomycin (5X 10)^-6IU/LGibco), 5.5 g/L pyruvate (Gibco) and 10% fetal calf serum (Hyclone, Logan UT, USA).

²⁺Determination of cell antagonistic Activity of Compounds Using Ca fluorescence

The method uses the FLIPR calcium detection kit (R8033; Molecular Devices, Sunnyvale, Calif.) to measure Ca on a fluorescence imaging plate reader (FLIPR)²⁺And (4) flow rate.

HEK293 cells expressing hTRPA1 were grown in 384-well microtiter plates (microtiter plates). Prior to the start of the assay, a 4 Xsolution of the test compound (i.e., the target compound (I) according to the present invention) was prepared in HEPES buffer (HBSS/HEPES), and Ca was added according to the manufacturer's instructions²⁺The fluorescent indicator dye Fluo-4 was dissolved in Hanks Balanced salt solution supplemented with 20 mM HEPES buffer. For detection, the medium was aspirated and the cells were loaded with 100 μ l of Fluo-4 solution at room temperature for about 2-3 hours, then 50 μ l of test compound solution was added to the cells at a delivery rate of 10 μ l/sec at the 10 second time point, and after 3 minutes the TRPA1 agonist AITC (allyl isothiocyanate) was added to activate and open the TRPA1 channel. The change in fluorescence over time was measured during the experiment with a FLIPR.

Data were analyzed with GraphPad Prism Software (GraphPad Software, San Diego, Calif.) using four parameter logistic Hill equations to curve fit concentration-effect data and derive IC₅₀The value is obtained. IC (integrated circuit)₅₀Indicating the concentration of test compound required to inhibit 50% of the response, a smaller value indicates a greater antagonism of the TRPA1 channel, and an activity at nM indicates that the compound is able to achieve effective antagonism of the TRPA1 channel. Part of the test results are shown in table 4.

Table 4: partial examples antagonistic Activity of Compound (I) of interest on human TRPA1

As shown in the table, the compounds I-1 to I-25 prepared by the invention have good TRPA1 antagonistic action and can inhibit the activation of a TRPA1 channel at nM concentration.

While the present invention has been described in detail with reference to the preferred embodiments, it should be understood that the above description should not be taken as limiting the invention. Various modifications and alterations to this invention will become apparent to those skilled in the art upon reading the foregoing description. Accordingly, the scope of the invention should be determined from the following claims.

Claims

1. The oxazole pyrimidone amide compound or the pharmaceutically acceptable salts thereof is characterized in that the oxazole pyrimidone amide compound has the following structural general formula (I):

wherein R is₁，R₂Each independently selected from hydrogen and C_1-3An alkyl group; r₃Is selected from C_1-3An alkyl group; r₄，R₅Each independently selected from 5-6 membered aryl, the aryl is phenyl or an heteroaryl containing 1-3 heteroatoms, the heteroatoms are taken from oxygen or nitrogen atoms, and the position of the heteroatoms is any position on the heteroaryl; the aromatic group being unsubstituted or at least substituted by one or more halogens or C_1-3Alkyl radical, C_1-3Polyhaloalkyl, C_1-3Alkoxy substitution.

2. An oxazolpyrimidone amide compound or a pharmaceutically acceptable salt thereof as claimed in claim 1, wherein the aromatic group is selected from phenyl, pyridyl, pyrimidyl, oxazolyl, oxadiazolyl or triazolyl.

3. A process for the preparation of an oxazolopyrimidinone amide compound or a pharmaceutically acceptable salt thereof according to claim 1 or 2 in a reaction scheme comprising:

4. Use of an oxazolopyrimidinone amide or a pharmaceutically acceptable salt thereof according to claim 1 or 2 for the preparation of a medicament for the treatment or prevention of a disease or condition responsive to TRPA1 modulation.

5. The use according to claim 4, wherein the disease or condition is selected from any one or more of pain, ocular irritation, skin irritation, pruritis, respiratory diseases associated with sensory disorders, gastrointestinal diseases associated with sensory disorders.

6. Use according to claim 5, wherein the disorder/condition associated with pain is selected from: hyperalgesia, pain caused by physical or chemical factors, infectious pain, neuropathic pain, diabetic-induced peripheral neuralgia, cancer-related pain, osteoarthritis-associated pain with rheumatoid arthritis, muscle spastic pain, renal colic, ureteral colic, bladder inflammatory pain.

7. Use according to claim 5, wherein the respiratory diseases associated with sensory disorders are selected from asthma, pertussis, chronic obstructive pulmonary disease, bronchitis; the gastrointestinal disorder associated with sensory disorders is selected from gastroesophageal reflux disease, irritable bowel syndrome or inflammatory bowel disease.

8. The use according to claim 6, wherein the pain caused by physical or chemical factors comprises: pain due to mechanical injury, and pain associated with radiotherapy and chemotherapy.

9. A pharmaceutical composition comprising an oxazolopyrimidinone amide compound according to claim 1 or 2 or a pharmaceutically acceptable salt thereof.