CN114195776B - Preparation and application of novel FXR small molecule agonist - Google Patents

Preparation and application of novel FXR small molecule agonist Download PDF

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CN114195776B
CN114195776B CN202010988297.2A CN202010988297A CN114195776B CN 114195776 B CN114195776 B CN 114195776B CN 202010988297 A CN202010988297 A CN 202010988297A CN 114195776 B CN114195776 B CN 114195776B
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赵一爽
张振伟
吴国辉
杨生生
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Kaisi Kaidi Shanghai Pharmaceutical Technology Co ltd
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Abstract

The invention discloses an FXR (farnesol X receptor) small molecule agonist and a preparation method thereof, and the structure is shown as a formula I. Wherein each substituent is defined in the specification and the claims. The compound has the advantages of high FXR agonistic activity, simple synthesis, easily available raw materials and the like, and can be used for medicaments for treating FXR related diseases.

Description

Preparation and application of novel FXR small molecule agonist
Technical Field
The invention belongs to the field of medicines, and relates to preparation and application of a non-steroidal compound serving as an FXR agonist. In particular to a preparation method of organic small molecular compounds which can be used as FXR agonists, and enantiomers, diastereoisomers, tautomers, racemates, hydrates, solvates, prodrugs or pharmaceutically acceptable salts thereof, and application thereof in preparing medicines for treating FXR related diseases.
Background
The farnesoid X receptor (Farnesoid X receptor) is a member of the nuclear receptor superfamily, belongs to ligand-dependent nuclear transcription factors, and is mainly expressed in liver, intestinal tract, kidney, bile duct and other systems; FXR is also called bile acid receptor because it can be activated by endogenous ligand bile acid and participate in important links such as bile acid metabolism and cholesterol metabolism. FXR can be directly involved in regulating expression of more than 300 genes including physiological processes such as lipid metabolism, carbohydrate metabolism, inflammation, fibrosis, liver regeneration, cell differentiation and proliferation. The ligand of the natural environment comprises primary bile acid chenodeoxycholic acid, secondary cholic acid lithocholic acid, deoxycholic acid and the like. For example, FXR activated by endogenous ligand bile acid plays an important role in Triglyceride (TG) metabolism, and FXR can reach steady state balance of TG content in liver and circulating blood by regulating key enzymes, lipoproteins and corresponding receptors for TG metabolism. Therefore, up to now, a number of FXR synthetic ligand molecules have been used in metabolic diseases such as liver.
FXR agonist molecules have shown excellent clinical efficacy in the treatment of liver diseases such as primary biliary cirrhosis (primary biliary cirrhosis, PBC), primary sclerosing cholangitis (primary sclerosing cholangitis, PSC) and non-alcoholic fatty liver disease (nonalcoholic steatohepatitis, NASH). Up to now, the FXR agonist molecule obeticholic acid (OCA), which is the first approved for marketing, has been demonstrated to significantly improve various metabolic symptoms, such as lowering liver fat content, reducing inflammatory response, inhibiting liver fibrosis, and the like. However, OCA has also increasingly revealed a number of clinical shortboards, such as causing itching, lowering of high density lipoprotein (high-density lipoprotein cholesterol, HDLc), raising of low density lipoprotein (low-density lipoprotein cholesterol, LDLc), etc. Therefore, in the aspect of clinical demands, new FXR agonist molecules with good clinical effects and low toxic and side effects are urgently needed.
Furthermore, studies have demonstrated that FXR is closely related to the development and progression of tumors. FXR plays an oncogene-inhibiting role in a variety of tumors. For example, in hepatocellular carcinoma and rectal cancer, FXR is in a low expression state, and after FXR activation, progression of liver cancer or rectal cancer is significantly inhibited by inhibiting the activity of β -catenin. Recent researches indicate that in cholangiocarcinoma, an agonist OCA of FXR can significantly inhibit proliferation, migration, clone formation and the like of intrahepatic cholangiocytes.
Furthermore, FXR agonists can be used as a new antiviral drug candidate, and studies have demonstrated that FXR ligands can be used as a new therapeutic strategy for inhibition of replication of hepatitis b virus (hepatitis B virus, HBV). FXR agonists inhibit HBV surface antigen synthesis, inhibit HBV DNA and RNA replication, and most importantly, inhibit HBV cccDNA production. In the context of hepatitis c virus (hepatitis C virus, HCV), FXR agonist GW4064 can inhibit HCV invasion of liver tissue cells by indirect means. Therefore, agonist molecules of FXR also hold great promise as a development of antiviral drugs.
In view of the above, there is a lack of novel FXR agonist molecules with simple preparation methods and good inhibition effects in the art.
Disclosure of Invention
The invention aims to provide a novel FXR agonist molecule which is simple in preparation method and good in inhibition effect.
In a first aspect of the present invention there is provided a compound of formula I, or an enantiomer, diastereomer, tautomer, racemate, hydrate, solvate, prodrug thereof, or a pharmaceutically acceptable salt thereof.
Figure BDA0002689977370000021
Wherein,,
ar is selected from the group consisting of: substituted or unsubstituted C 6 -C 10 Aryl, substituted or unsubstituted 5-9 membered heteroaryl rings (including monocyclic or fused rings containing 1-3 heteroatoms selected from oxygen, sulfur, and nitrogen);
a is selected from the group consisting of: substituted or unsubstituted C 6 -C 10 Aryl, substituted or unsubstituted 5-9 membered heteroaryl rings (including monocyclic or fused rings containing 1-3 heteroatoms selected from oxygen, sulfur, and nitrogen);
R 1 selected from the group consisting of: substituted or unsubstituted C 1 -C 6 Alkyl, substituted or unsubstituted C 3 -C 6 Cycloalkyl, substituted or unsubstituted 5-9 membered heterocycle (containing 1-3 heteroatoms selected from oxygen, sulfur and nitrogen);
x is selected from the group consisting of: hydrogen or deuterium;
wherein one or more hydrogen atoms of the substituent groups are each independently replaced by a substituent selected from the group consisting of: halogen, halogenated C 1 -C 6 Alkyl, halogenated C 1 -C 6 Alkoxy, C 1 -C 6 Alkyl, C 1 -C 6 Alkoxy, C 3 -C 6 Cycloalkyl, C 3 -C 6 A cycloalkoxy group, a cyano group, or a nitro group.
In another preferred embodiment, R is 1 Selected from the group consisting of: substituted or unsubstituted C 1 -C 6 Alkyl, substituted or unsubstituted C 3 -C 6 Cycloalkyl; wherein the substituents refer to one or more hydrogen atoms on the group each independently replaced by a substituent selected from the group consisting of: halogen, halogenated C 1 -C 6 Alkyl, halogenated C 1 -C 6 Alkoxy, C 1 -C 6 Alkyl, C 1 -C 6 Alkoxy, C 3 -C 6 Cycloalkyl, C 3 -C 6 A cycloalkoxy group, a cyano group, or a nitro group.
In another preferred embodiment, ar is selected from the group consisting of: substituted or unsubstituted C 6 -C 10 Aryl, substituted or unsubstituted 5-9 membered heteroaryl ring, and said substituents are selected from the group consisting of: hydrogen, fluorine, chlorine, bromine, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, trifluoromethyl, or trifluoromethoxy.
In another preferred embodiment, a is selected from the group consisting of: substituted or unsubstituted C 6 -C 10 An aryl, substituted or unsubstituted 5-9 membered heteroaryl ring, wherein the aryl or heteroaryl substituent is selected from the group consisting of: hydrogen, fluorine, chlorine, bromine, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, trifluoromethyl, or trifluoromethoxy.
In another preferred embodiment, ar is selected from the group consisting of: substituted or unsubstituted phenyl, substituted or unsubstituted 5-7 membered heteroaryl ring (including monocyclic or fused rings containing 1-3 heteroatoms selected from oxygen, sulfur and nitrogen).
In another preferred embodiment, a is selected from the group consisting of: substituted or unsubstituted phenyl, substituted or unsubstituted 5-7 membered heteroaryl ring (including monocyclic or fused rings containing 1-3 heteroatoms selected from oxygen, sulfur and nitrogen).
In another preferred embodiment, a is a substituted or unsubstituted benzothiazole.
In another preferred embodiment, each Ar or A is independently selected from the group consisting of substituted and unsubstituted radicals selected from the group consisting of: benzene ring, pyridine ring, pyrimidine ring, pyridazine ring, furan ring, thiophene ring, pyrrole ring, thiazole ring, or imidazole ring.
In another preferred embodiment, R is 1 Selected from the group consisting of: substituted or unsubstituted C 1 -C 4 Alkyl, substituted or unsubstituted cyclopropyl.
In another preferred embodiment, ar is a substituted or unsubstituted benzene ring.
In another preferred embodiment, ar is selected from the group consisting of: 2, 5-dichlorophenyl, 2-methylphenyl, 2-trifluoromethylphenyl, 2-trifluoromethoxyphenyl.
In another preferred embodiment, R is 1 Selected from the group consisting of: methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, cyclopropyl, cyclobutyl, or cyclopentyl.
In another preferred embodiment, the compound of formula (I) has the structure shown below:
Figure BDA0002689977370000031
in another preferred embodiment, the compound of formula (I) has the structure shown below:
Figure BDA0002689977370000032
in another preferred embodiment, the compound is selected from the group consisting of:
Figure BDA0002689977370000033
in a second aspect of the present invention there is provided a process for the preparation of a compound according to the first aspect of the present invention, the process comprising: a compound of formula I is prepared by a method described in route one or route two selected from the group consisting of:
route one:
Figure BDA0002689977370000041
(a) Reacting a compound shown in a general formula II of substituted benzaldehyde with hydroxylamine hydrochloride under the action of alkali to obtain an intermediate, and then chlorinating the intermediate with N-chlorosuccinimide (NCS) to obtain a compound shown in a general formula III;
(b) Then reacting the compound shown in the general formula III with corresponding 3-oxo-propionate under the action of alkali to obtain a compound shown in the general formula IV;
(c) Reducing the ester in the compound shown in the general formula IV into corresponding alcohol, namely the compound shown in the general formula V under the action of a deuterated reducing agent;
(d) Brominating the compound shown in the general formula IV with a bromine reagent to generate a compound shown in VI;
(e) Reacting a compound shown in a general formula VI with a compound shown in VII under the action of alkali to form a compound shown in a general formula VIII;
(f) Reacting a compound shown in a general formula VIII with hydroxylamine hydrochloride under the action of alkali to generate a compound shown in a general formula IX;
(g) The compound shown in the general formula IX reacts under the action of phosgene, triphosgene or carbonyl diimidazole to generate the compound shown in the general formula I,
wherein X is deuterium; r is R 1 Ar, A are as defined in the first aspect of the invention.
Route two:
Figure BDA0002689977370000051
(a) The ester in the compound shown in the general formula IV is reduced into corresponding alcohol, namely the compound shown in the general formula X under the action of a reducing agent;
(b) Oxidizing the compound shown in the general formula X into corresponding aldehyde, namely a compound shown in the general formula XI under the action of an oxidant;
(c) The ester in the compound shown in the general formula XI is reduced into a compound shown in the general formula V under the action of a deuterated reducing agent;
(d) Brominating the compound shown in the general formula V by using a bromine reagent to generate a compound shown in VI;
(e) Reacting a compound shown in a general formula VI with a compound shown in VII under the action of alkali to form a compound shown in a general formula VIII;
(f) Reacting a compound shown in a general formula VIII with hydroxylamine hydrochloride under the action of alkali to generate a compound shown in a general formula IX;
(g) The compound shown in the general formula IX reacts under the action of phosgene, triphosgene or carbonyl diimidazole to generate the compound shown in the general formula I,
in the formula, X is hydrogen; r is R 1 Ar, A are as defined in the first aspect of the invention.
In another preferred embodiment, the compound of formula VII is prepared by the steps of:
Figure BDA0002689977370000052
(k) The compound shown in the general formula XII and the compound shown in the general formula XIII generate a compound shown in the general formula VII under the action of alkali;
in the formulae, A is as defined in the first aspect of the invention.
In another preferred embodiment, when the product is optically isomeric, the preparation is carried out using starting materials of the corresponding optical configuration.
In a third aspect of the present invention, there is provided a pharmaceutical composition comprising a compound of formula I according to the first aspect of the present invention, or an enantiomer, diastereomer, tautomer, racemate, hydrate, solvate, prodrug, pharmaceutically acceptable salt thereof; and a pharmaceutically acceptable carrier.
In a fourth aspect of the invention, there is provided the use of a compound of formula I according to the first aspect of the invention, or an enantiomer, diastereomer, tautomer, racemate, hydrate, solvate, prodrug or a pharmaceutically acceptable salt thereof, for the preparation of a pharmaceutical composition for the treatment of a disease or condition associated with FXR activity or expression level.
In another preferred embodiment, the FXR related disorder is selected from the group consisting of: bile acid metabolism, glycometabolism, lipid metabolism, inflammation, and/or liver fibrosis process-related diseases.
In another preferred example, the FXR related disease is non-alcoholic fatty liver (NASH), primary Biliary Cirrhosis (PBC), primary Sclerosing Cholangitis (PSC), gall stones, non-alcoholic cirrhosis, hepatitis B (HBV), hepatitis C (HCV), liver fibrosis, cholestatic liver disease, hyperlipidemia, hypercholesterolemia, or diabetes.
In another preferred embodiment, the pharmaceutical composition is used as an FXR agonist.
In another preferred embodiment, the pharmaceutical composition is used to reduce the level of ALP, ALT, AST, TBA in serum.
In another preferred embodiment, the pharmaceutical composition is used to reduce the hydroxyproline content in liver tissue.
In another preferred embodiment, the pharmaceutical composition is used to down-regulate α -SMA and Col1 α1mRNA expression in liver tissue.
In another preferred embodiment, the pharmaceutical composition is used for inhibiting HBV surface antigen synthesis, inhibiting HBV DNA and RNA replication, and inhibiting HBV cccDNA production.
In another preferred embodiment, the pharmaceutical composition is for reducing collagen content in the liver.
In another preferred embodiment, the pharmaceutical composition is prepared by the following method: the compound of the formula I is mixed with pharmaceutically acceptable auxiliary materials (such as excipient, diluent and the like) to prepare tablets, capsules, granules or syrup for oral administration.
It is understood that within the scope of the present invention, the above-described technical features of the present invention and technical features specifically described below (e.g., in the examples) may be combined with each other to constitute new or preferred technical solutions. And are limited to a space, and are not described in detail herein.
Detailed Description
The inventor of the application researches widely and intensively to develop a non-steroidal compound which can be used as an FXR agonist and has agonizing capability on FXR at a molecular level and a cellular level, and researches show that the compound can reduce the level of ALP, ALT, AST, TBA in serum, reduce the content of hydroxyproline in liver tissues, down regulate the expression of a-SMA and Col1 a 1mRNA in the liver tissues, reduce the content of collagen in the liver, inhibit HBV surface antigen synthesis, inhibit the replication of HBV DNA and RNA and inhibit the generation of HBV cccDNA. The compound has the advantages of high FXR agonistic activity, simple synthesis, easily available raw materials and the like, and can be used for preparing medicines for treating FXR related diseases. On this basis, the present invention has been completed.
Terminology
In the present invention, unless otherwise indicated, terms used have the ordinary meanings known to those skilled in the art.
In the present invention, the halogen is F, cl, br or I.
In the present invention, the term "C1-C6" means having 1,2,3,4, 5 or 6 carbon atoms, "C3-C6" means having 3,4, 5 or 6 carbon atoms, and so on.
In the present invention, the term "alkyl" means a saturated linear or branched hydrocarbon moiety, for example the term "C1-C6 alkyl" refers to a straight or branched alkyl group having 1 to 6 carbon atoms, including without limitation methyl, ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl, tert-butyl, pentyl, hexyl and the like; ethyl, propyl, isopropyl, butyl, isobutyl, sec-butyl and tert-butyl are preferred.
In the present invention, the term "alkoxy" denotes an-O- (C1-C6 alkyl) group. For example, the term "C1-C6 alkoxy" refers to straight or branched chain alkoxy groups having 1 to 6 carbon atoms, including without limitation methoxy, ethoxy, propoxy, isopropoxy, butoxy and the like.
In the present invention, the term "cycloalkyl" means a saturated cyclic hydrocarbyl moiety, for example the term "C3-C6 cycloalkyl" refers to a cyclic alkyl group having 3 to 6 carbon atoms in the ring, including, without limitation, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, and the like.
In the present invention, the term "cycloalkoxy" means cycloalkyl-O-, cycloalkyl as described above.
In the present invention, the term "aryl" means a hydrocarbyl moiety comprising one or more aromatic rings. Examples of aryl groups include, but are not limited to, phenyl (Ph), naphthyl, pyrenyl, fluorenyl, anthracenyl, and phenanthryl.
In the present invention, the term "heteroaryl" means a moiety comprising one or more aromatic rings having at least one heteroatom (e.g., N, O or S). Examples of heteroaryl groups include furyl, pyrrolyl, thienyl, oxazolyl, imidazolyl, thiazolyl, pyridyl, pyrimidinyl, quinazolinyl, quinolinyl, isoquinolinyl, indolyl, and the like.
Unless otherwise indicated, alkyl, alkoxy, cycloalkyl, cycloalkoxy, aryl, and heteroaryl groups described herein are substituted and unsubstituted groups. Possible substituents on alkyl, alkoxy, cycloalkyl, cycloalkoxy, aryl, and heteroaryl groups include, but are not limited to: hydroxy, amino, nitro, nitrile, halogen, C 1 -C 6 Alkyl, C 2 -C 10 Alkenyl, C 2 -C 10 Alkynyl, C 3 -C 20 Cycloalkyl, C 3 -C 20 Cycloalkenyl, C 1 -C 20 Heterocycloalkyl, C 1 -C 20 Heterocycloalkenyl, C 1 -C 6 Alkoxy, aryl, heteroaryl, heteroaryloxy, C 1 -C 10 Alkylamino, C 1 -C 20 Dialkylamino, arylamino, diarylamino, C 1 -C 10 Alkylsulfamoyl, arylsulfamoyl, C 1 -C 10 Alkylimino, C 1 -C 10 Alkyl sulfo imino, aryl sulfo imino, mercapto, C 1 -C 10 Alkylthio, C 1 -C 10 Alkylsulfonyl, arylsulfonyl, acylamino, or a combination thereof,Aminoacyl, aminothioacyl, guanidino, ureido, cyano, acyl, thioacyl, acyloxy, carboxyl, and carboxylate groups. On the other hand, cycloalkyl, heterocycloalkyl, heterocycloalkenyl, aryl and heteroaryl may also be fused to each other.
In the present invention, the substitution is mono-substitution or poly-substitution, and the poly-substitution is di-substitution, tri-substitution, tetra-substitution, or penta-substitution. The disubstitution means having two substituents and so on.
The pharmaceutically acceptable salts of the present invention may be salts of anions with positively charged groups on the compounds of formula I. Suitable anions are chloride, bromide, iodide, sulfate, nitrate, phosphate, citrate, methylsulfonate, trifluoroacetate, acetate, malate, tosylate, tartrate, fumarate, glutamate, glucuronate, lactate, glutarate or maleate. Similarly, salts may be formed from cations with negatively charged groups on the compounds of formula I. Suitable cations include sodium, potassium, magnesium, calcium and ammonium ions, such as tetramethylammonium.
In another preferred embodiment, "pharmaceutically acceptable salt" refers to the salt of a compound of formula I with an acid selected from the group consisting of: hydrofluoric acid, hydrochloric acid, hydrobromic acid, phosphoric acid, acetic acid, oxalic acid, sulfuric acid, nitric acid, methanesulfonic acid, sulfamic acid, salicylic acid, trifluoromethanesulfonic acid, naphthalenesulfonic acid, maleic acid, citric acid, acetic acid, lactic acid, tartaric acid, succinic acid, oxalacetic acid, pyruvic acid, malic acid, glutamic acid, p-toluenesulfonic acid, naphthalenesulfonic acid, ethanesulfonic acid, naphthalenedisulfonic acid, malonic acid, fumaric acid, propionic acid, oxalic acid, trifluoroacetic acid, stearic acid, pamoic acid, hydroxymaleic acid, phenylacetic acid, benzoic acid, glutamic acid, ascorbic acid, p-aminobenzenesulfonic acid, 2-acetoxybenzoic acid, isethionic acid, and the like; or a sodium, potassium, calcium, aluminum or ammonium salt of a compound of formula I with an inorganic base; or the methylamine, ethylamine or ethanolamine salt of the compounds of the formula I with organic bases.
In another preferred embodiment, the compounds are those wherein A ring, ar, X and R 1 Any one ofThe groups corresponding to the specific compounds described in the examples, respectively.
The compounds of the invention have asymmetric centers, chiral axes and chiral planes and may exist in the form of racemates, R-isomers or S-isomers. Those skilled in the art can resolve the R-isomer and/or S-isomer from the racemate using conventional techniques.
Preparation method
The preparation method of the compound shown in the general formula I comprises the following synthetic route:
route one:
Figure BDA0002689977370000081
(a) Reacting a compound shown in a general formula II of substituted benzaldehyde with hydroxylamine hydrochloride under the action of alkali to obtain an intermediate, and then chlorinating the intermediate with N-chlorosuccinimide (NCS) to obtain a compound shown in a general formula III;
(b) Then reacting the compound shown in the general formula III with corresponding 3-oxo-propionate under the action of alkali to obtain a compound shown in the general formula IV;
(c) Reducing the ester in the compound shown in the general formula IV into corresponding alcohol, namely the compound shown in the general formula V under the action of a deuterated reducing agent;
(d) Brominating the compound shown in the general formula V by using a bromine reagent to generate a compound shown in VI;
(e) Reacting a compound shown in a general formula VI with a compound shown in VII under the action of alkali to form a compound shown in a general formula VIII;
(f) Reacting a compound shown in a general formula VIII with hydroxylamine hydrochloride under the action of alkali to generate a compound shown in a general formula IX;
(g) The compound shown in the general formula IX reacts under the action of phosgene, triphosgene or carbonyl diimidazole to generate the compound shown in the general formula I,
wherein X is deuterium, R 1 Ar, A are as defined in claim 1.
Route two:
Figure BDA0002689977370000091
(h) The ester in the compound shown in the general formula IV is reduced into corresponding alcohol, namely the compound shown in the general formula X under the action of a reducing agent;
(i) Oxidizing the compound shown in the general formula X into corresponding aldehyde, namely a compound shown in the general formula XI under the action of an oxidant;
(j) The ester in the compound shown in the general formula XI is reduced into a compound shown in the general formula V under the action of a deuterated reducing agent;
(d) Brominating the compound shown in the general formula V by using a bromine reagent to generate a compound shown in VI;
(e) Reacting a compound shown in a general formula VI with a compound shown in VII under the action of alkali to form a compound shown in a general formula VIII;
(f) Reacting a compound shown in a general formula VIII with hydroxylamine hydrochloride under the action of alkali to generate a compound shown in a general formula IX;
(g) The compound shown in the general formula IX reacts under the action of phosgene, triphosgene or carbonyl diimidazole to generate the compound shown in the general formula I,
in the formula, X is hydrogen, R 1 Ar, A are as defined in the first aspect of the invention.
In another preferred embodiment, the compound of formula VII is prepared by the steps of:
Figure BDA0002689977370000092
(k) The compound shown in the general formula XII and the compound shown in the general formula XIII generate a compound shown in the general formula VII under the action of alkali;
in the formulae, A is as defined in the first aspect of the invention.
Pharmaceutical composition and therapeutic use thereof
The compound provided by the invention can be singly used or mixed with pharmaceutically acceptable auxiliary materials (such as excipient, diluent and the like) to prepare tablets, capsules, granules or syrups for oral administration. The pharmaceutical composition can be prepared according to a conventional pharmaceutical method. The pharmaceutical compositions of the present invention comprise an active ingredient in a safe and effective amount, and a pharmaceutically acceptable carrier.
The "active ingredient" as used herein refers to the compound of formula I as described herein.
The active ingredient and the pharmaceutical composition are used for preparing medicines for treating FXR related diseases. The invention is that
The "active ingredients" and pharmaceutical compositions are useful as FXR agonists. In another preferred embodiment, the active ingredient may be used for the preparation of a medicament for the prophylaxis and/or treatment of diseases modulated by FXR agonists.
"safe and effective amount" means: the amount of active ingredient is sufficient to significantly improve the condition without causing serious side effects. Typically, the pharmaceutical compositions contain 1-2000mg of active ingredient per dose, more preferably 10-200mg of active ingredient per dose. Preferably, the "one dose" is a tablet.
"pharmaceutically acceptable carrier" means: one or more compatible solid or liquid filler or gel materials which are suitable for human use and must be of sufficient purity and sufficiently low toxicity. "compatibility" as used herein means that the components of the composition are capable of blending with and between the active ingredients of the present invention without significantly reducing the efficacy of the active ingredients. Examples of pharmaceutically acceptable carrier moieties are cellulose and its derivatives (e.g., sodium carboxymethylcellulose, sodium ethylcellulose, cellulose acetate, and the like), gelatin, talc, solid lubricants (e.g., stearic acid, magnesium stearate), calcium sulfate, vegetable oils (e.g., soybean oil, sesame oil, peanut oil, olive oil, and the like), polyols (e.g., propylene glycol, glycerol, mannitol, sorbitol, and the like), emulsifiers (e.g.
Figure BDA0002689977370000101
) Wetting agents (e.g. sodium lauryl sulphate), colouring agents, flavouring agents, stabilisers, antioxidants, preservatives, athermalRaw water, and the like.
The mode of administration of the active ingredient or pharmaceutical composition of the present invention is not particularly limited, and representative modes of administration include, but are not limited to, 5: oral, intratumoral, rectal, parenteral (intravenous, intramuscular, or subcutaneous), and the like.
Solid dosage forms for oral administration include capsules, tablets, pills, powders and granules.
Liquid dosage forms for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups or tinctures. In addition to the active ingredient, the liquid dosage forms may contain inert diluents commonly used in the art such as, for example, water or other solvents, solubilizing agents and emulsifiers such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, propylene glycol, 1, 3-butylene glycol, dimethylformamide and oils, in particular, cottonseed, groundnut, corn germ, olive, castor and sesame oils or mixtures of these substances and the like. In addition to these inert diluents, the compositions can also include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, and perfuming agents.
Suspensions, in addition to the active ingredient, may contain suspending agents as, for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum methoxide and agar or mixtures of these substances, and the like.
Compositions for parenteral injection may comprise physiologically acceptable sterile aqueous or anhydrous solutions, dispersions, suspensions or emulsions, and sterile powders for reconstitution into sterile injectable solutions or dispersions. Suitable aqueous and nonaqueous carriers, diluents, solvents or excipients include water, ethanol, polyols and suitable mixtures thereof.
The compounds of the present invention may be administered alone or in combination with other therapeutic agents, such as hypolipidemic agents.
When a pharmaceutical composition is used, a safe and effective amount of the compound of the present invention is applied to a mammal (e.g., a human) in need of treatment, wherein the dose at the time of administration is a pharmaceutically effective dose, and the daily dose is usually 1 to 2000mg, preferably 20 to 500mg, for a human having a body weight of 60 kg. Of course, the particular dosage should also take into account factors such as the route of administration, the health of the patient, etc., which are within the skill of the skilled practitioner.
The invention will be further illustrated with reference to specific examples. It is to be understood that these examples are illustrative of the present invention and are not intended to limit the scope of the present invention. The experimental procedures, which do not address the specific conditions in the examples below, are generally carried out under conventional conditions (e.g.those described in Sambrook et al, molecular cloning: A laboratory Manual (New York: cold Spring Harbor Laboratory Press, 1989)) or under conditions recommended by the manufacturer. Percentages and parts are weight percentages and parts unless otherwise indicated.
Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. In addition, any methods and materials similar or equivalent to those described herein can be used in the methods of the present invention. The preferred methods and materials described herein are presented for illustrative purposes only.
The instrument and main experimental materials used are as follows:
the reagents and anhydrous solvents used were purchased from chinese commercial company and were used directly unless specified; 1 h and 13 c NMR was performed using Bruker AM-400 and Varian Mercury plus-400 NMR, and Agilent6230 type mass spectrometer, and 200-300 mesh column chromatography silica gel (Qingdao ocean chemical Co., ltd.) and HSGF254 TLC plate (smoke table chemical industry institute).
The compounds of the present invention are prepared by any one of the methods selected from the following routes one and two, using appropriate starting materials:
route one:
Figure BDA0002689977370000111
route two:
Figure BDA0002689977370000121
example intermediate VII-1 synthesis:
Figure BDA0002689977370000122
endo-8-azabicyclo [3.2.1]Octane-3-ol XII (10 g,78.7 mmol) and p-fluorobenzonitrile XIII-1 (78.7 mmol) were dissolved in N, N-dimethylformamide (150 mL), and potassium carbonate (197 mmol) was added in portions at room temperature and reacted at 80℃overnight. Ethyl acetate (500 mL) was added to dilute the reaction, washed with water, and the aqueous phase was extracted with ethyl acetate (300 mL each for 3 times). The organic phase was mixed, washed with saturated brine and concentrated. Column chromatography gave intermediate VII-1 (11 g, yield 61%). 1 H NMR(400MHz,DMSO-d 6 )δ7.51–7.48(m,2H),6.82–6.79(m,2H),4.62(s,1H),4.27(d,J＝5.2Hz,2H),3.78(s,1H),2.28(d,J＝6.8Hz,2H),1.91–1.85(m,4H),1.57(d,J＝14.0Hz,2H).MS(ESI,m/z)：229[M+H] + 。
Synthesis of example compound 1:
Figure BDA0002689977370000131
an aqueous potassium carbonate solution (3N, 182 mmol) was added dropwise to a stirred solution of hydroxylamine hydrochloride (182 mmol) in ethanol (100 mL) at 0℃and 2, 6-dichlorobenzaldehyde II-1 (20 g,114 mmol) was dissolved in 100mL of ethanol, followed by addition to the above reaction solution, and the temperature was raised to 90℃for two hours. The reaction solution was allowed to cool to room temperature and then concentrated to a solid. A solution of water/ethanol (1000 mL/100 mL) was added and the solid was broken up with stirring, filtered and dried overnight under vacuum at 50deg.C to give compound intermediate (18.4 g). This intermediate was dissolved in N, N-dimethylformamide (50 mL), and a solution of N-chlorosuccinimide (97 mmol) in N, N-dimethylformamide (100 mL) was added dropwise at 0℃and stirred overnight. The reaction solution was poured into ice water at 0℃and then extracted with methyl t-butyl ether (200 mL each time for 3 times), and the organic phase was washed with saturated brine and concentrated to give a crude product. N-hexane (600 mL) was added to the flask containing the crude product, and the flask was usedStirring with a magnet, filtration and drying of the solid under vacuum (30 ℃ C.) gives intermediate III-1 (18.3 g, 73% yield). 1 H NMR(400MHz,CDCl 3 )δ7.43–7.39(m,2H),7.39–7.33(m,1H)。
Triethylamine (8.2 g) was added to methyl 3-cyclopropyl-3-oxopropanoate (82 mmol), and stirred for 30 minutes. Then cooled to 10℃and a solution of III-1 (18.3 g,82 mmol) in absolute ethanol (80 mL) was added dropwise thereto (internal temperature not exceeding 30 ℃), and the reaction was allowed to stand overnight at room temperature. Ethyl acetate (100 mL) was added to dilute the reaction, washed with water, and the aqueous phase was extracted with ethyl acetate (100 mL each for 3 times). The organic phase was mixed, washed with saturated brine and concentrated. To the concentrate was added 100mL of diethyl ether with stirring, and the solvent was removed in vacuo to give the solid product IV-1 (21.6 g, yield 84%). 1 H NMR(400MHz,CDCl 3 )δ7.43–7.39(m,2H),7.39–7.33(m,1H),3.72(s,3H),2.21–2.09(m,1H),1.35–1.28(m,2H),1.25–1.18(m,2H)；MS(ESI,m/z)：312[M+H] + 。
Deuterated lithium aluminum (7.3 g) was added to tetrahydrofuran (200 mL), cooled to 0℃and IV-1 (21.6 g,69.2 mmol) tetrahydrofuran (50 mL) was added dropwise thereto (internal temperature not exceeding 5 ℃ C.) and the reaction solution was stirred at room temperature for 2 hours. The reaction was quenched by addition of ice water (9 mL) at 0deg.C, followed by dropwise addition of 15% aqueous sodium hydroxide solution (9 mL) and ice water (27 mL), respectively, and then by addition of anhydrous magnesium sulfate (100 g), and the above mixture was stirred at room temperature for 0.5h, filtered, concentrated, and subjected to column chromatography to give intermediate V-1 (19 g, yield 96%). 1 H NMR(400MHz,DMSO-d 6 )δ7.72–7.59(m,2H),7.58–7.50(m,1H),4.90(s,1H),2.38–2.26(m,1H),1.16–1.03(m,4H).MS(ESI,m/z)：286[M+H] + 。
V-1 (19 g,66.4 mmol) was dissolved in dichloromethane (200 mL), cooled to 0deg.C, phosphorus tribromide (66.4 mmol) was slowly added dropwise to the solution, and the reaction was stirred at room temperature for 2h. The reaction solution was freed from the solvent to give an oil, which was diluted with ethyl acetate (100 mL), the pH of the reaction solution was adjusted to neutrality with a saturated aqueous sodium bicarbonate solution, washed with water, and the aqueous phase was extracted with ethyl acetate (100 mL each for 3 times). The organic phase was mixed, washed with saturated brine and concentrated. Warp yarnColumn chromatography gave intermediate VI-1 (20.2 g, yield 87%). 1 H NMR(400MHz,DMSO-d 6 )δ7.72–7.65(m,2H),7.64–7.56(m,1H),2.48–2.39(m,1H),1.26–1.10(m,4H).MS(ESI,m/z)：348[M+H] + 。
To a solution of VII-1 (1.96 g,8.6 mmol) in anhydrous tetrahydrofuran (150 mL) was added potassium tert-butoxide (21.4 mmol) at 0deg.C, stirred for 15 minutes, then a solution of VI-1 (8.6 mmol) in anhydrous tetrahydrofuran (50 mL) was added dropwise, and the reaction was stirred at room temperature for 4 hours. To the reaction mixture was added water (200 mL), which was extracted with ethyl acetate (200 mL each time, 3 times total), and the organic phase was washed with saturated brine and concentrated, and column chromatography gave intermediate VIII-1 (2.1 g, yield 49%). 1 H NMR(400MHz,DMSO-d 6 )δ7.73-7.63(m,2H),7.64–7.55(m,3H),6.83(d,J＝8.4Hz,2H),4.16(s,2H),3.37(s,1H),2.38–2.31(m,1H),1.83–1.74(m,6H),1.56–1.49(m,2H),1.16–1.06(m,4H).MS(ESI,m/z)：496[M+H] + 。
VIII-1 (2.1 g,4.2 mmol), hydroxylamine hydrochloride (8.4 mmol), absolute ethanol (80 mL) were added to the round bottom flask and stirred, triethylamine (8.4 mmol) was slowly added dropwise and the reaction was heated to 80℃overnight. Cooled to room temperature, the solvent was removed, dissolved in dichloromethane (150 mL), and the organic phase was concentrated by washing with water and saturated brine, and intermediate IX-1 (1.1 g, yield 50%) was obtained by silica gel column chromatography. 1 HNMR(400MHz,DMSO-d 6 )δ7.73–7.63(m,2H),7.64–7.55(m,3H),6.83(d,J＝8.4Hz,2H),4.67(s,2H),3.43–3.35(m,1H),2.39–2.32(m,1H),1.89–1.78(m,6H),1.57–1.48(m,2H),1.16–1.06(m,4H).MS(ESI,m/z)：529[M+H] + 。
IX-1 (1.1 g,2.1 mmol), N, N' -carbonyldiimidazole (3.1 mmol), 1, 4-dioxane (100 mL) was added to a round bottom flask followed by 1, 8-diazabicyclo [5.4.0]Undec-7-ene (3.1 mmol) was reacted for 8 hours by heating to 100 ℃. The reaction solution was cooled to room temperature, diluted with water (100 mL), and the pH was adjusted to about 3 with 1M aqueous hydrochloric acid, followed by extraction with ethyl acetate (100 mL each for 3 times). The organic phases were combined, washed with saturated brine, and the crude product obtained was concentrated and subjected to silica gel column chromatography to give final product 1 (158 mg, yield 13%). 1 HNMR(400MHz,DMSO-d 6 )δ7.65–7.63(m,2H),7.59–7.55(m,3H),6.83(d,J＝8.4Hz,2H),4.17(s,2H),3.38(s,1H),2.37–2.30(m,1H),1.80–1.72(m,6H),1.51(d,J＝14.4Hz,2H),1.16–1.06(m,4H).MS(ESI,m/z)：555[M+H] + 。
Example 2:
Figure BDA0002689977370000141
the preparation of example 2 was prepared by route 1 starting from intermediate VII-2, the synthetic route being as follows:
Figure BDA0002689977370000151
starting from starting material XIII-2, compound VII-2 is synthesized according to the synthesis method for synthesizing intermediate VII-1, and then prepared by scheme 1 to obtain 2, wherein:
white solid VIII-2 yield 77%, 1 HNMR(400MHz,DMSO-d 6 )δ8.21(s,1H),7.70–7.62(m,3H),7.60–7.54(m,1H),7.16(d,J＝8.4Hz,1H),4.29(s,2H),3.43–3.35(m,1H),2.37–2.29(m,1H),1.85-1.65(m,6H),1.63–1.54(m,2H),1.17–1.05(m,4H).MS(ESI,m/z)：497[M+H] + 。
the yield of white solid 2 was 64%, 1 HNMR(400MHz,DMSO-d 6 )δ8.20–8.16(m,1H),7.70(d,J＝8.4Hz,1H),7.67–7.62(m,2H),7.60–7.53(m,1H),7.28–7.21(m,1H),4.28(s,br,2H),3.42–3.40(m,1H),2.38–2.29(m,1H),1.81–1.69(m,6H),1.60–1.52(m,2H),1.16–1.05(m,2H).MS(ESI,m/z)：556[M+H] + 。
example 3:
Figure BDA0002689977370000152
the preparation of example 3 was prepared by route 1 starting from intermediate VII-3, the synthetic route being as follows:
Figure BDA0002689977370000153
starting from starting material XIII-3, compound VII-3 is synthesized according to the synthesis method for synthesizing intermediate VII-1, and then 3 is prepared through scheme 1, wherein:
white solid VIII-3 yield 67%, HNMR (400 MHz, DMSO-d) 6 )δ7.68–7.45(m,4H),6.69(t,J＝12.0Hz,2H),4.18(s,2H),3.46–3.36(m,1H),2.38–2.27(m,1H),1.80–1.66(m,6H),1.60–1.51(m,2H),1.20–1.03(m,4H).MS(ESI,m/z)：497[M+H] + 。
The yield of the white solid 3 is 38%, 1 H NMR(400MHz,DMSO-d 6 )δ7.69–7.42(m,4H),6.74(d,J＝13.6Hz,1H),6.63(d,J＝8.8Hz,1H),4.21(s,2H),3.47–3.43(m,1H),2.35–2.30(m,1H),1.76–1.70(m,6H),1.58–1.50(m,2H),1.20–1.06(m,4H).MS(ESI,m/z)：573[M+H] + 。
example 4:
Figure BDA0002689977370000161
preparation of example 4 reference example 3 preparation 4 was prepared by route 2 starting from intermediate IV-1 as follows:
Figure BDA0002689977370000162
lithium aluminum (420 mg,10 mmol) was added to tetrahydrofuran (8 mL), cooled to 0℃and IV-1 (2 mmol) tetrahydrofuran (2 mL) was added dropwise thereto (internal temperature no more than 5 ℃ C.), and the reaction solution was stirred at room temperature for 2 hours. The reaction was quenched by adding ice water (0.4 mL) at 0deg.C, then 15% aqueous sodium hydroxide solution (0.4 mL) and ice water (1.2 mL) were added dropwise, respectively, and then anhydrous magnesium sulfate (8 g) was added, and the above mixture was stirred at room temperature for 0.5h, filtered, concentrated, and column-chromatographed to give intermediate X-1 (1 g, yield 84%). MS (ESI, m/z): 284[ M+H ]] + 。
X-1 (1 g,3.53 mmol) was added to methylene chloride (20 mL),pyridine chlorochromate (14.14 mmol) was added at room temperature, and the reaction solution was stirred at room temperature for 1h. Filtration, concentration and column chromatography gave intermediate XI-1 (870 mg, 88% yield). MS (ESI, m/z): 282[ M+H ]] + 。
Lithium aluminum (260 mg,6.2 mmol) was added to tetrahydrofuran (4 mL), cooled to 0℃and XI-1 (3.1 mmol) tetrahydrofuran (1 mL) was added dropwise thereto (internal temperature was not more than 5 ℃ C.), and the reaction solution was stirred at room temperature for 2 hours. The reaction was quenched by adding ice water (0.2 mL) at 0deg.C, then 15% aqueous sodium hydroxide solution (0.2 mL) and ice water (0.6 mL) were added dropwise, respectively, and then anhydrous magnesium sulfate (10 g) was added, and the above mixture was stirred at room temperature for 0.5h, filtered, concentrated, and subjected to column chromatography to give intermediate V-2 (730 mg, yield 83%). 1 H NMR(400MHz,DMSO-d 6 )δ7.62-7.60(m,2H),7.56-7.52(m,1H),4.92(d,J＝5.2Hz,1H),4.18(d,J＝5.2Hz,1H),2.35–2.28(m,1H),1.14–1.04(m,4H).MS(ESI,m/z)：285[M+H] + 。
V-2 (730 mg,2.57 mmol) was dissolved in dichloromethane (10 mL), cooled to 0deg.C, phosphorus tribromide (3.08 mmol) was slowly added dropwise to the solution, and the reaction was stirred at room temperature for 2h. The reaction solution was freed from the solvent to give an oil, which was diluted with ethyl acetate (20 mL), the pH of the reaction solution was adjusted to neutrality with saturated aqueous sodium bicarbonate, washed with water, and the aqueous phase was extracted with ethyl acetate (100 mL each for 3 times). The organic phase was mixed, washed with saturated brine and concentrated. Column chromatography gave intermediate VI-2 (720 mg, yield 81%). MS (ESI, m/z): 347[ M+H ]] + 。
To a solution of VII-2 (512 mg,2.08 mmol) in anhydrous tetrahydrofuran (10 mL) was added potassium tert-butoxide (4.16 mmol) at 0deg.C, stirred for 15 min, then a solution of VI-2 (2.08 mmol) in anhydrous tetrahydrofuran (5 mL) was added dropwise and the reaction stirred at room temperature for 4h. To the reaction mixture was added water (20 mL), which was extracted with ethyl acetate (20 mL each for 3 times), and the organic phase was washed with saturated brine and concentrated, followed by column chromatography to give intermediate VIII-4 (420 mg, yield 39%). MS (ESI, m/z): 513[ M+H ]] + 。
VIII-4 (420 m g,0.82 mmol), hydroxylamine hydrochloride (1.64 mmol), absolute ethanol (5 mL) were added to a round bottom flask and stirred, gentlyTriethylamine (1.64 mmol) was slowly added dropwise and reacted overnight with heating to 80 ℃. Cooled to room temperature, the solvent was removed, dissolved in dichloromethane (20 mL), and the organic phase was concentrated by washing with water and saturated brine, and intermediate IX-4 was obtained by silica gel column chromatography (360 m g, yield 81%). MS (ESI, m/z): 546[ M+H ]] + 。
IX-4 (360 mg,0.66 mmol), N, N' -carbonyldiimidazole (0.99 mmol), 1, 4-dioxane (5 mL) was added to a round bottom flask followed by 1, 8-diazabicyclo [5.4.0]Undec-7-ene (0.99 mmol) was heated to 100deg.C and reacted for 8 hours. The reaction solution was cooled to room temperature, diluted with water (10 mL), and the pH was adjusted to about 3 with 1M aqueous hydrochloric acid, followed by extraction with ethyl acetate (10 mL each for 3 times). The organic phases were combined, washed with saturated brine, and the crude product obtained was concentrated and subjected to silica gel column chromatography to give final product 4 (76.7 mg, yield 20%). 1 HNMR(400MHz,DMSO-d 6 )δ7.65-7.62(m,2H),7.58–7.55(m,1H),7.47(t,J＝8.6Hz,1H),6.72–6.65(m,2H),4.22(s,1H),4.17(s,2H),3.39(s,1H),2.36–2.30(m,1H),1.76-1.73(m,6H),1.52(d,J＝14.8Hz,2H),1.14–1.08(m,4H).MS(ESI,m/z)：572[M+H] + 。
Example 5:
Figure BDA0002689977370000171
the preparation of example 5 was prepared by route 1 starting from intermediate VII-4 as follows:
Figure BDA0002689977370000181
starting from starting material XIII-4, compound VII-4 is synthesized according to the synthesis method for synthesizing intermediate VII-1, and then prepared by scheme 1 to give 5, wherein:
the yield of the white solid VIII-5 is 47 percent, 1 HNMR(400MHz,DMSO-d 6 )δ7.68–7.61(m,2H),7.60–7.55(m,1H),7.42(d,J＝8.4Hz,1H),6.71(s,br,1H),6.62(d,J＝8.8Hz,1H),4.17(s,2H),3.41–3.36(m,1H),2.39–2.26(m,4H),1.84–1.66(m,6H),1.58–1.46(m,2H),1.19–1.02(m,4H).MS(ESI,m/z)：510[M+H] + 。
the yield of white solid 3 was 11%, 1 H NMR(400MHz,DMSO-d 6 )δ7.69–7.62(m,2H),7.60–7.35(m,2H),6.75–6.545(m,2H),4.19(s,2H),3.43–3.33(m,1H),2.41–2.29(m,4H),1.88–1.66(m,6H),1.60–1.46(m,2H),1.21–1.03(m,4H).MS(ESI,m/z)：569[M+H] + 。
example 6:
Figure BDA0002689977370000182
the preparation of example 6 was prepared by route 1 starting from intermediate VII-5, the synthetic route being as follows:
Figure BDA0002689977370000183
starting from starting material XIII-5, compound VII-5 is synthesized according to the synthesis method for synthesizing intermediate VII-1, and then prepared by scheme 1 to give 6, wherein:
the yield of the white solid VIII-6 is 32 percent, 1 HNMR(400MHz,DMSO-d 6 )δ7.66–7.61(m,2H),7.60–7.52(m,1H),7.32(d,J＝8.4Hz,1H),6.39–6.29(m,2H),4.22(s,2H),3.83(s,3H),3.41–3.32(m,1H),2.36–2.28(m,1H),1.84–1.66(m,6H),1.60–1.49(m,2H),1.20–1.02(m,4H).MS(ESI,m/z)：526[M+H] + 。
the yield of the white solid was 9%, 1 H NMR(400MHz,DMSO-d 6 )δ7.69–7.52(m,3H),7.37–7.29(m,1H),6.45–6.36(m,2H),4.28(s,2H),3.87(s,3H),3.43–3.32(m,1H),2.42–2.27(m,1H),1.87–1.64(m,6H),1.61–1.47(m,2H),1.21–1.02(m,4H).MS(ESI,m/z)：585[M+H] + 。
example 7:
Figure BDA0002689977370000191
/>
the preparation of example 7 was prepared by route 1 starting from intermediate VII-6 as follows:
Figure BDA0002689977370000192
starting from starting material XIII-6, compound VII-6 is synthesized according to the synthesis method for synthesizing intermediate VII-1, and then 7 is prepared through scheme 1, wherein:
the yield of the white solid VIII-7 is 56 percent, 1 HNMR(400MHz,DMSO-d 6 )δ7.75(d,J＝9.2Hz,1H),7.67–7.52(m,3H),7.08(s,1H),7.02(d,J＝8.8Hz,1H),4.32(s,2H),3.43–3.32(m,1H),2.36–2.28(m,1H),1.78–1.69(m,3H),1.61–1.50(m,2H),1.16–1.06(m,4H).MS(ESI,m/z)：564[M+H] + 。
the yield of white solid 7% was 49%, 1 H NMR(400MHz,DMSO-d 6 )δ7.67–7.47(m,4H),7.11–7.04(m,2H),4.27(s,2H),3.48–3.44(m,1H),2.35–2.31(m,1H),1.85–1.69(m,6H),1.62–1.48(m,2H),1.20–1.01(m,4H).MS(ESI,m/z)：623[M+H] + 。
example 8:
Figure BDA0002689977370000201
the preparation of example 8 was prepared by route 1 starting from starting material II-2 and was synthesized as follows:
Figure BDA0002689977370000202
wherein:
the yield of the colloid V-3 is 98 percent, 1 HNMR(400MHz,DMSO-d 6 )δ7.88(d,J＝7.6Hz,1H),7.81–7.71(m,2H),7.59(d,J＝7.2Hz,1H),4.90(s,1H),2.33-2.26(m,1H),1.12-1.05(m,4H).MS(ESI,m/z)：286[M+H] + 。
the yield of the white solid VIII-8 is 31 percent, 1 HNMR(400MHz,DMSO-d 6 )δ7.90(d,J＝7.6Hz,1H),7.81-7.72(m,2H),7.58(d,J＝7.6Hz,1H),7.50(t,J＝8.4Hz,1H),6.74(dd,J＝13.8,2.2Hz,1H),6.63(dd,J＝8.6,2.2Hz,1H),4.20(s,1H),3.61–3.57(m,1H),3.40–3.32(m,1H),2.33–2.29(m,1H),1.76-1.69(m,6H),1.54(d,J＝14.4Hz,2H),1.18–1.05(m,4H).MS(ESI,m/z)：514[M+H] + 。
the yield of white solid 8 was 29%, 1 HNMR(400MHz,DMSO-d 6 )δ7.90(d,J＝8.0Hz,1H),7.81–7.72(m,2H),7.58(d,J＝7.2Hz,1H),7.47(t,J＝8.6Hz,1H),6.72–6.66(m,2H),4.17(s,2H),3.38(s,1H),2.34–2.28(m,1H),1.77–1.73(m,6H),1.52(d,J＝14.8Hz,2H),1.13–1.05(m,4H).MS(ESI,m/z)：573[M+H] + 。
example 9:
Figure BDA0002689977370000211
the preparation of example 9 was prepared by route 1 starting from starting material II-3 and was synthesized as follows:
Figure BDA0002689977370000212
wherein:
the yield of the colloid V-4 is 76 percent, 1 HNMR(400MHz,DMSO-d 6 )δ7.67-7.63(m,2H),7.53(d,J＝6.8Hz,2H),4.96(s,1H),2.32-2.27(m,1H),1.13-1.04(m,4H).MS(ESI,m/z)：302[M+H] + 。
the yield of the colloid VIII-9 is 47 percent, 1 HNMR(400MHz,DMSO-d 6 )δ7.69–7.61(m,2H),7.56–7.49(m,3H),6.75(d,J＝14.0Hz,1H),6.64(d,J＝8.8Hz,1H),4.21(s,2H),3.45–3.43(m,1H),2.36–2.29(m,1H),1.77–1.69(m,6H),1.56(d,J＝14.8Hz,2H),1.23–1.06(m,4H).MS(ESI,m/z)：530[M+H] + 。
white solid 8 yield 21%, 1 HNMR(400MHz,DMSO-d 6 )δ7.69–7.62(m,2H),7.56–7.46(m,3H),6.72–6.66(m,2H),4.18(s,2H),3.43(s,1H),2.35–2.31(m,1H),1.80–1.75(m,6H),1.54(d,J＝14.4Hz,2H),1.13–1.06(m,4H).MS(ESI,m/z)：589[M+H] + 。
pharmacological experimental examples:
a method for detecting FXR agonistic activity of a compound based on a method for detecting reporter activity:
1.1 construction and preparation of plasmids pGAL4-FXR-LBD and pG5-Luc
pGAL4-FXR-LBD and pG5-Luc plasmids used in the reporter gene detection system were constructed by conventional molecular cloning methods. The method mainly comprises the following steps: inserting FXR (NM_ 001206979.2) cDNA sequence corresponding to FXR-LBD (212-476 AA) amino acid sequence into BamHI and NotI cleavage sites of pGAL4 vector by PCR technology to obtain pGAL4-FXR-LBD; pG5-Luc and phRL-TK plasmids were donated to Shanghai pharmaceutical institute of China academy of sciences; by CaCl 2 The plasmids were transformed into DH 5. Alpha. E.coli, and after further culture amplification, the corresponding plasmid DNA was obtained by purification using a plasmid extraction kit (TIANGEN, #D107).
1.2 plasmid cotransfection of HEK293T cells and Compound treatment
HEK293T cells were plated at 1X 10 the day prior to plasmid transfection 4 Density/well was seeded in 96-well plates. According to transfection reagents
Figure BDA0002689977370000221
Cell transfection was performed in accordance with the instructions of HD (Promega, # E2311). The method mainly comprises the following steps: as an example, the plasmids pGAL4-FXR-LBD, pG5-Luc and phRL-TK were added in a proportion of 20ng, 50ng and 5ng to 10uL of Opti-MEM TM Mixing the above materials in medium I (Gibco, # 11058021); a further 0.25uL of +.>
Figure BDA0002689977370000222
HD, mixing, standing at room temperature for 5min; this 10uL mixture was then added to the cell well containing 100uL of culture broth. 6h after cell cotransfection, the compound is diluted with a gradient of 3 times at the highest concentration of 1uM, 10 total concentrations are added into cell culture solution for 24h, 2 total duplicate wells are divided, and LJN452 compound is used as positive control.
1.3Dual-Glo Luciferase assay
After 24h of treatment of the cells with the compounds, the following are followed
Figure BDA0002689977370000223
Luciferase Assay System (Promega, # E2940) instructions. The method mainly comprises the following steps: 50uL of culture solution is pipetted off per well, and 50uL of +.>
Figure BDA0002689977370000224
Figure BDA0002689977370000225
Luciferase reagent, shake for 10min at room temperature; taking 80uL of the cleavage reaction solution to a white opaque optiPlate-96 well plate, and detecting a luminescence signal value (Firefly-Luc) of Firefly luciferase (Firefly luciferase) by using an MD i3x multifunctional enzyme-labeled instrument; adding 40 uL->
Figure BDA0002689977370000226
Stop&/>
Figure BDA0002689977370000227
Reagent, oscillating for 10min at room temperature; the luminescence signal value (Renilla-Luc) of Renilla luciferase (Renilla luciferase) was detected by an MD i3x multifunctional microplate reader. EC were calculated using GraphPad prism6.0 software to fit a dose-response curve with four parameters using Firefly-Luc/Renilla-Luc ratios as compounds for FXR activation and solvent DMSO group ratios for normalization 50 Values.
2. Results
Experimental data indicate that compounds have a certain FXR agonistic activity, wherein the EC of examples 1,2,3,4 50 All less than 5nM, has very strong FXR agonistic activity. Other examples FXR agonist activity data are shown in table 1.
FXR agonistic Activity of Compounds of Table 1
Figure BDA0002689977370000228
Figure BDA0002689977370000231
＊＊＊＊:EC 50 (nM)<5；＊＊＊:5<EC 50 (nM)<10；＊＊:10<EC 50 (nM)<50；＊:50<EC 50 (nM)。
Figure BDA0002689977370000232
The results show that the compounds of the present invention exhibit superior cellular level activity to the existing FXR agonist compound LJN452 and to the non-deuterated control 1. In particular, the compound of example 3 of the present application was deuterated based on the structure of the non-deuterated reference substance 1, and the activity was significantly improved, suggesting that this position is a critical deuterated site of such a compound.
All documents mentioned in this application are incorporated by reference as if each were individually incorporated by reference. Further, it will be appreciated that various changes and modifications may be made by those skilled in the art after reading the above teachings, and such equivalents are intended to fall within the scope of the claims appended hereto.

Claims (9)

1. A compound represented by the general formula I, or a pharmaceutically acceptable salt thereof,
Figure QLYQS_1
wherein,,
ar is selected from the group consisting of: substituted or unsubstituted C 6 -C 10 Aryl, substituted or unsubstituted 5-9 membered heteroaryl rings, including monocyclic or fused rings, containing 1-3 heteroatoms selected from oxygen, sulfur and nitrogen;
a is selected from the group consisting of: substituted or unsubstituted C 6 -C 10 Aryl, substituted or unsubstituted 5-9 membered heteroaryl rings, including monocyclic or fused rings, containing 1-3 heteroatoms selected from oxygen, sulfur and nitrogen;
R 1 selected from the group consisting of: c (C) 3 -C 6 Cycloalkyl;
x is selected from the group consisting of: hydrogen or deuterium;
wherein the substituents refer to one or more hydrogen atoms on the group each independently replaced by a substituent selected from the group consisting of: halogen, halogenated C 1 -C 6 Alkyl, halogenated C 1 -C 6 Alkoxy, C 1 -C 6 Alkyl, C 1 -C 6 Alkoxy, C 3 -C 6 Cycloalkyl, C 3 -C 6 A cycloalkoxy group, a cyano group, or a nitro group.
2. The compound of claim 1, wherein R 1 Is cyclopropyl.
3. The compound of claim 1, wherein Ar is selected from the group consisting of: substituted or unsubstituted C 6 -C 10 Aryl, substituted or unsubstituted 5-9 membered heteroaryl ring, and said substituents are selected from the group consisting of: hydrogen, fluorine, chlorine, bromine, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, trifluoromethyl, or trifluoromethoxy.
4. The compound of claim 1, wherein a is selected from the group consisting of: substituted or unsubstituted C 6 -C 10 An aryl, substituted or unsubstituted 5-9 membered heteroaryl ring, wherein the aryl or heteroaryl substituent is selected from the group consisting of: hydrogen, fluorine, chlorine, bromine, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, trifluoromethyl, or trifluoromethoxy.
5. The compound of claim 1, wherein said compound is selected from the group consisting of:
Figure QLYQS_2
。
6. a process for the preparation of a compound as claimed in claim 1, wherein said process comprises: a compound of formula I is prepared by a method described in route one or route two selected from the group consisting of:
route one:
Figure QLYQS_3
(a) Reacting a compound shown in a general formula II of substituted benzaldehyde with hydroxylamine hydrochloride under the action of alkali to obtain an intermediate, and then chlorinating the intermediate with N-chlorosuccinimide (NCS) to obtain a compound shown in a general formula III;
(b) Then reacting the compound shown in the general formula III with corresponding 3-oxo-propionate under the action of alkali to obtain a compound shown in the general formula IV;
(c) Reducing the ester in the compound shown in the general formula IV into a compound shown in the general formula V under the action of a deuterated reducing agent;
(d) Brominating the compound shown in the general formula IV with a bromine reagent to generate a compound shown in VI;
(e) Reacting a compound shown in a general formula VI with a compound shown in VII under the action of alkali to form a compound shown in a general formula VIII;
(f) Reacting a compound shown in a general formula VIII with hydroxylamine hydrochloride under the action of alkali to generate a compound shown in a general formula IX;
(g) The compound shown in the general formula IX reacts under the action of phosgene, triphosgene or carbonyl diimidazole to generate the compound shown in the general formula I,
wherein X is deuterium; r is R 1 Ar, A are as defined in claim 1;
route two:
Figure QLYQS_4
(a) The ester in the compound shown in the general formula IV is reduced into corresponding alcohol, namely the compound shown in the general formula X under the action of a reducing agent;
(b) Oxidizing the compound shown in the general formula X into corresponding aldehyde, namely a compound shown in the general formula XI under the action of an oxidant;
(c) Reducing the compound shown in the general formula XI into a compound shown in the general formula V under the action of a deuterated reducing agent;
(d) Brominating the compound shown in the general formula V by using a bromine reagent to generate a compound shown in VI;
(e) Reacting a compound shown in a general formula VI with a compound shown in VII under the action of alkali to form a compound shown in a general formula VIII;
(f) Reacting a compound shown in a general formula VIII with hydroxylamine hydrochloride under the action of alkali to generate a compound shown in a general formula IX;
(g) The compound shown in the general formula IX reacts under the action of phosgene, triphosgene or carbonyl diimidazole to generate the compound shown in the general formula I,
in the formula, X is hydrogen; r is R 1 Ar, A are as defined in claim 1.
7. A pharmaceutical composition comprising a compound of formula I according to claim 1, or a pharmaceutically acceptable salt thereof; and a pharmaceutically acceptable carrier.
8. The use of a compound of formula I, or a pharmaceutically acceptable salt thereof, according to claim 1, for the preparation of a pharmaceutical composition for the treatment of a disease or condition associated with FXR activity or expression level.
9. The use according to claim 8, wherein the FXR related disorder is selected from the group consisting of: bile acid metabolism, glycometabolism, lipid metabolism, inflammation, and/or liver fibrosis process-related diseases.
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CN108064223A (en) * 2014-12-17 2018-05-22 吉利德科学公司 FXR (NR1H4) modulating compound of hydroxyl
WO2019120088A1 (en) * 2017-12-22 2019-06-27 四川科伦博泰生物医药股份有限公司 Isoxazole derivative, preparation method therefor, and use thereof
WO2020156241A1 (en) * 2019-01-31 2020-08-06 中国医药研究开发中心有限公司 Aromatic ring or heteroaromatic ring compounds, preparation method therefor and medical use thereof

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Publication number Priority date Publication date Assignee Title
CN108064223A (en) * 2014-12-17 2018-05-22 吉利德科学公司 FXR (NR1H4) modulating compound of hydroxyl
WO2019120088A1 (en) * 2017-12-22 2019-06-27 四川科伦博泰生物医药股份有限公司 Isoxazole derivative, preparation method therefor, and use thereof
WO2020156241A1 (en) * 2019-01-31 2020-08-06 中国医药研究开发中心有限公司 Aromatic ring or heteroaromatic ring compounds, preparation method therefor and medical use thereof

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