CN109734700B - SMO inhibitor containing benzoyl piperidine structure and preparation method and application thereof - Google Patents

SMO inhibitor containing benzoyl piperidine structure and preparation method and application thereof Download PDF

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CN109734700B
CN109734700B CN201910032140.XA CN201910032140A CN109734700B CN 109734700 B CN109734700 B CN 109734700B CN 201910032140 A CN201910032140 A CN 201910032140A CN 109734700 B CN109734700 B CN 109734700B
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徐云根
纪德重
张晶晶
朱启华
梁停停
柏英
王致斌
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China Pharmaceutical University
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Abstract

The invention discloses benzoyl piperidine derivatives, a preparation method and application thereof, wherein the benzoyl piperidine derivatives are compounds containing an N-methyl-N- (1- (2-aminobenzoyl) piperidine-4-yl) amide structure, and the structure of the benzoyl piperidine derivatives is shown as a formula I. The invention also discloses a preparation method of the benzoyl piperidine derivative. The invention also discloses application of the benzoyl piperidine derivative in preparation of an SMO protein inhibitor and application of a pharmaceutical composition containing the benzoyl piperidine derivative in preparation of an anti-tumor medicament.

Description

SMO inhibitor containing benzoyl piperidine structure and preparation method and application thereof
Technical Field
The invention belongs to the technical field of pharmaceutical chemistry, and particularly relates to SMO protein inhibitors containing an N-methyl-N- (1- (2-aminobenzoyl) piperidine-4-yl) amide structure, preparation methods thereof, pharmaceutical compositions containing the compounds, and application of the pharmaceutical compositions in preparing antitumor drugs.
Technical Field
Tumors are a serious disease that endangers human health. With the annual increase in morbidity and mortality, malignancies have become the leading cause of death and an important public health problem worldwide. Chemotherapy is one of the main means for clinical treatment of tumors, so the research of antitumor drugs is always the focus and hot spot of the research in the pharmaceutical industry.
Stem cells are an emerging new field of biomedical research in recent years and have great clinical application value. Since the 21 st century, with the continuous improvement of the tumor stem cell theory, researchers have had new insights into why tumors are difficult to cure completely. A great deal of evidence indicates that the tumor stem cells existing in the tumor tissues are insensitive to the existing chemotherapy and radiotherapy measures, and the cells have the capacity of re-differentiating to form tumor cells, so that the targeted killing of the tumor stem cells becomes a key factor for completely curing the cancer.
Given that the basic characteristics of tumor stem cells are very similar to those of adult stem cells, targeting a key pathway for somatic cell development can become a means of killing tumor stem cells. Current research on developmental-related pathway Notach, Hedgehog, and Wnt inhibitors has yielded a series of results, with inhibitors against the key protein SMO in the Hedgehog pathway being FDA-approved for marketing for the treatment of basal cell carcinomas. However, clinical studies found that patients quickly developed resistance to marketed drugs due to mutations in the SMO protein (D473H). Therefore, the development of second generation SMO inhibitors against SMO mutant resistance presents new opportunities and challenges to the current research field.
Disclosure of Invention
The purpose of the invention is as follows: the invention provides a benzoyl piperidine derivative containing an N-methyl-N- (1- (2-aminobenzoyl) piperidine-4-yl) amide structure, a specific preparation method of the derivative and pharmaceutical application of the derivative in preparing SMO protein inhibitors.
The technical scheme is as follows: the invention discloses a benzoyl piperidine derivative shown as a general formula I, or pharmaceutically acceptable salt thereof:
Figure BDA0001944617740000011
wherein:
R1selected from substituted or unsubstituted phenyl, wherein substituted phenyl is phenyl substituted with one, two or more substituents selected from: halogen, -NO2、-NH2-CN, C1-C6 alkoxy, -CHO, -CF3、-CONH2C1-C6 alkanoyl,-SO2CH3、-NHCOCH3、-NHCOCH2CH3、-CH2CONH2or-NHCONH2；
R2Selected from substituted or unsubstituted five-membered or six-membered aromatic heterocyclic group, or substituted or unsubstituted C6-C12 aromatic hydrocarbon group, wherein, the substituted five-membered or six-membered aromatic heterocyclic group is a five-membered or six-membered aromatic heterocyclic group substituted by one, two or more substituents selected from the following group: halogen, C1-C6 alkyl, -CN, -NH2、-NHCOCH3C1-C6 alkanoyl, C1-C6 alkoxy or-COOH; substituted C6-C12 aromatic hydrocarbon groups are C6-C12 aromatic hydrocarbon groups substituted with one, two or more substituents selected from the group consisting of: halogen, C1-C6 alkyl, -CN, -NH2、-NHCOCH3C1-C6 alkanoyl, C1-C6 alkoxy or-COOH.
Preferably, the method comprises the following steps:
R1is selected from
Figure BDA0001944617740000021
Wherein R is3And R4Each independently selected from-H, halogen, -NO2、-NH2、-CN、-OCH3、-CHO、-CF3、-CONH2、-COCH3、-SO2CH3、-NHCOCH3、-NHCOCH2CH3、-CH2CONH2or-NHCONH2；
R2Selected from substituted or unsubstituted five-membered or six-membered N-containing aromatic heterocyclic group, or substituted or unsubstituted phenyl or biphenyl group, wherein the substituted five-membered or six-membered N-containing aromatic heterocyclic group is a five-membered or six-membered N-containing aromatic heterocyclic group substituted by one, two or more substituents selected from the group consisting of: halogen, -CH3、-CH2CH3、-CH2CH2CH3、-CN、-NH2、-NHCOCH3、-COCH3、-COCH2CH3、-COCH2CH2CH3、-OCH3、-OCH2CH3、-OCH2CH2CH3or-COOH; substituted phenyl or biphenyl radicals by one, twoPhenyl or biphenyl substituted with one or more substituents selected from: halogen, -CH3、-CH2CH3、-CH2CH2CH3、-CN、-NH2、-NHCOCH3、-COCH3、-COCH2CH3、-COCH2CH2CH3、-OCH3、-OCH2CH3、-OCH2CH2CH3or-COOH.
Further preferably:
R1is selected from
Figure BDA0001944617740000022
Wherein R is3Selected from-H, halogen, -NO2、-NH2、-CN、-OCH3、-CHO、-CF3、-CONH2、-COCH3、-SO2CH3、-NHCOCH3、-NHCOCH2CH3、-CH2CONH2or-NHCONH2；R4Is selected from-H or-CF3；
R2Selected from substituted or unsubstituted five-membered or six-membered N-containing aromatic heterocyclic group, or substituted or unsubstituted phenyl or biphenyl group, wherein the substituted five-membered or six-membered N-containing aromatic heterocyclic group is a five-membered or six-membered N-containing aromatic heterocyclic group substituted by one, two or more substituents selected from the group consisting of: halogen, -CH3、-CN、-NH2、-NHCOCH3、-COCH3、-OCH3or-COOH; substituted phenyl or biphenyl is phenyl or biphenyl substituted with one, two or more substituents selected from the group consisting of: halogen, -CH3、-CN、-NH2、-NHCOCH3、-COCH3、-OCH3or-COOH.
More preferably:
R1is selected from
Figure BDA0001944617740000031
Figure BDA0001944617740000032
R2Is selected from
Figure BDA0001944617740000033
Figure BDA0001944617740000034
The above preferred or further preferred or more preferred radicals do not necessarily represent a simultaneous preference for these radicals, such as, in preference, R1May be preferred only, R2May not be preferred, of course R1And R2May also be preferred.
The term "aromatic hydrocarbon group" refers to aromatic hydrocarbon compound groups such as phenyl, biphenyl, naphthyl, anthryl, phenanthryl and the like.
The term "arylheterocyclyl" refers to an aromatic heterocyclic group containing a heteroatom such as N, S or O, such as phenanthridinyl, carbazole, dibenzofuran, pyrrolyl, triazolyl, pyridyl, pyrazinyl, indolyl, pyrimidinyl, quinolinyl, and the like.
The term "C1-C6 alkyl" refers to a straight or branched chain alkyl group having 1 to 6 carbon atoms, such as methyl, ethyl, propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, and the like.
The term "C1-C6 alkanoyl" refers to straight or branched chain alkanoyl of 1 to 6 carbon atoms, e.g. C1 alkanoyl refers to acetyl and C2 alkanoyl refers to propionyl.
The term "C1-C6 alkoxy" refers to straight or branched chain alkoxy groups having 1 to 6 carbon atoms, such as methoxy, ethoxy, propoxy, isopropoxy, n-butoxy, isobutoxy, sec-butoxy, tert-butoxy and the like.
The term "substituted" refers to mono-or polysubstitution.
Preferably, the benzoyl piperidine derivative is selected from I-1 to I-17:
Figure BDA0001944617740000035
Figure BDA0001944617740000041
Figure BDA0001944617740000051
the preparation method of the benzoyl piperidine derivative comprises the following steps of preparing a compound I by a condensation reaction of a compound IV and an IX:
Figure BDA0001944617740000061
preferably, the preparation method of the benzoyl piperidine derivative comprises the following steps:
(1) the compound II and the compound III are subjected to Buchwald-Hartwig reaction to prepare an intermediate IV;
(2) preparing VI from the compound V through reductive amination reaction; then carrying out condensation reaction on the compound VI and the compound VII to prepare a compound VIII; then removing the protecting group from the compound VIII to prepare a compound IX;
(3) and (3) preparing a compound I by condensation reaction of a compound IX and an intermediate IV:
Figure BDA0001944617740000062
or, the preparation method of the benzoyl piperidine derivative comprises the following steps of preparing a compound I by Buchwald-Hartwig reaction of compounds XI and XII:
Figure BDA0001944617740000071
wherein A is halogen.
Preferably, the method comprises the steps of preparing a compound XI from a compound IX-1 and X through condensation reaction; then, the compound I is prepared by Buchwald-Hartwig reaction of the compound XI and the compound XII.
Figure BDA0001944617740000072
Wherein A is halogen.
For example, it can be prepared by the following method:
preparation method of the compound of the general formula I:
Figure BDA0001944617740000073
wherein R is1Represents
Figure BDA0001944617740000074
Wherein R is1`represents-NO2、-OCH3、-F、-CN、-CF3、-COCH3or-SO2CH3。
Further, the process for preparing the compound I by the condensation reaction of the compound IV and IX comprises the following steps: the condensing agent used is selected from benzotriazol-1-yl-oxytripyrrolidinophosphonium hexafluorophosphate (PyBOP), 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (EDCI), Dicyclohexylcarbodiimide (DCC) or N, N' -Carbonyldiimidazole (CDI), preferably EDCI; the solvent used is selected from dichloromethane, N-Dimethylformamide (DMF), N-Dimethylacetamide (DMAC) or Dimethylsulfoxide (DMSO), preferably dichloromethane; the reaction temperature is selected from 0 ℃ to 50 ℃, preferably 20 ℃ to 30 ℃; the reaction time is 4-48 h, preferably 8-16 h. The compound IV: compound IX: the ratio of the condensing agent (molar ratio) is 1:1: 1-1: 10:10, preferably 1:1: 1-1: 2: 2.
A second process for the preparation of a compound of formula I:
Figure BDA0001944617740000081
wherein R is2Represents
Figure BDA0001944617740000082
Wherein R is2`represents-H, -CN, -OCH3or-COOH.
Further, the procedure for preparing compound I from compound XI and XII by Buchwald-Hartwig reaction: the compound XII is aniline, 4-cyanoaniline, 2-cyanoaniline, 4-methoxyaniline, 2-methoxyaniline, 4-aminopyridine, 2-aminopyridine, 4-phenylaniline and 4-aminobenzoic acid; the solvent is selected from toluene, xylene, Tetrahydrofuran (THF), ethylene glycol dimethyl ether (DME), dioxane, N-Dimethylformamide (DMF), N-methylpyrrolidone (NMP), dimethyl sulfoxide (DMSO) or a mixed solvent of any two or three of the solvents, preferably toluene; the catalyst is selected from tris (dibenzylideneacetone) dipalladium (Pd)2(dba)3) Or palladium acetate (Pd (OAc)2) Preferably Pd (OAc)2(ii) a The ligand is selected from 2,2 '-bis- (diphenylphosphino) -1,1' -Binaphthyl (BINAP), tri (2-tolyl) phosphine (P (o-tolyl)3) Or 4, 5-bis (diphenylphosphino) -9, 9-dimethylxanthene (Xantphos), preferably BINAP; the reaction temperature is selected from 70-110 ℃, preferably 100-110 ℃; the reaction time is 4-48 h, preferably 8-16 h. The compound XI: compound XII: catalyst: the ratio of the ligands (molar ratio) is 1:1:0.01: 1-1: 10:0.5:5, preferably 1:1:0.1: 2-1: 2:0.2: 3.
The invention relates to a preparation method of a key intermediate IV in a preparation method of a compound with a general formula I, which comprises the following steps:
Figure BDA0001944617740000083
further, the procedure for preparing compound IV from compound II and III by Buchwald-Hartwig reaction: the catalyst is selected from tris (dibenzylideneacetone) dipalladium (Pd)2(dba)3) Palladium acetate (Pd (OAc)2) Or copper iodide (CuI), preferably CuI; the base is selected from sodium tert-butoxide (NaOtBu), lithium hexamethyldisilazide (LiHMDS), potassium carbonate (K)2CO3) Potassium phosphate (K)3PO4) Or cesium carbonate (Cs)2CO3) Preferably potassium carbonate; the solvent is selected from toluene, xylene, Tetrahydrofuran (THF), ethylene glycol dimethyl ether (DME),Dioxane, N-Dimethylformamide (DMF), N-methylpyrrolidone (NMP), dimethyl sulfoxide (DMSO) or a mixed solvent of any two or three of them, preferably DMF; the reaction temperature is selected from 70-110 ℃, preferably 100-105 ℃; the reaction time is 4-48 h, preferably 8-16 h. The compound II: compound III: catalyst: the ratio of the alkali (molar ratio) is 1:1:0.1: 1-1: 10:1:10, preferably 1:1:0.2: 2-1: 1.5:0.5: 5.
The invention relates to a preparation method of a key intermediate IX in a preparation method of a compound with a general formula I, which comprises the following steps:
Figure BDA0001944617740000091
wherein R is1Represents
Figure BDA0001944617740000092
Wherein R is1`represents-NO2、-OCH3、-F、-CN、-CF3、-COCH3or-SO2CH3。
Further, the process for preparing compound VI from compound V by reductive amination: the reducing agent is selected from sodium borohydride (NaBH)4) Sodium triacetoxyborohydride (NaBH (OAc))3) Or sodium cyanoborohydride (NaBH)3CN), preferably sodium cyanoborohydride; the solvent is selected from ethyl acetate, methanol, tetrahydrofuran, chloroform, acetonitrile, dichloromethane, toluene or a mixed solvent of any two or three of the solvents, preferably methanol; the reaction temperature is selected from 0 ℃ to 50 ℃, preferably 20 ℃ to 30 ℃; the reaction time is 4-24 h, preferably 8-16 h. The compound V: methylamine: the ratio of the reducing agent (molar ratio) is 1:1: 1-1: 10:5, preferably 1:1.5: 1.5-1: 3: 3.
Process for preparing compound VIII by condensation of compounds VI and VII: the compound VII is 2-trifluoromethyl-4-fluorobenzoic acid, 4-nitrobenzoic acid, 4-methoxybenzoic acid, 4-fluorobenzoic acid, 4-cyanobenzoic acid, 4-trifluoromethylbenzoic acid, 4-formylbenzoic acid and 4-methylsulfonylbenzoic acid; the chlorinating agent is selected from thionyl chloride (SOCl)2) Phosphorus oxychloride (POCl)3) Phosphorus pentachloride (PCl)5) Or oxalyl chloride ((COCl)2) Preferably oxalyl chloride; the acid-binding agent is selected from triethylamine, N-Diisopropylethylamine (DIEA), 4-Dimethylaminopyridine (DMAP), pyridine, sodium acetate, sodium carbonate or potassium carbonate, preferably triethylamine; the solvent is selected from methanol, ethanol, dichloromethane, acetone, ethyl acetate, tetrahydrofuran or a mixed solvent of any two or three of the solvents, preferably dichloromethane; the reaction temperature is selected from 0 ℃ to 50 ℃, preferably 0 ℃ to 30 ℃; the reaction time is 0.5h to 5h, preferably 1h to 2 h. The compound VI: compound VII: a chlorination reagent: the acid binding agent is in a molar ratio of 1:1:1: 2-5: 1:5:10, preferably 1:1:1: 2-2: 1:2: 4.
Procedure for preparation of compound IX from compound VIII deprotection: the deprotection reagent used is selected from hydrogen chloride, hydrogen bromide, sulfuric acid or trifluoroacetic acid, preferably trifluoroacetic acid; the solvent is methanol, ethanol, dichloromethane, acetone, ethyl acetate, tetrahydrofuran or a mixed solvent of any two, preferably dichloromethane; the reaction temperature is selected from 0 ℃ to 50 ℃, preferably 20 ℃ to 30 ℃; the reaction time is 0.5h to 5h, preferably 1h to 2 h.
The invention relates to a method for preparing a key intermediate XI in a preparation method of a compound with a general formula I, which comprises the following steps:
Figure BDA0001944617740000101
further, the process for preparing compound XI from compound X and IX-1 by condensation reaction: the chlorinating agent is selected from thionyl chloride (SOCl)2) Phosphorus oxychloride (POCl)3) Phosphorus pentachloride (PCl)5) Or oxalyl chloride ((COCl)2) Preferably oxalyl chloride; the acid-binding agent is selected from triethylamine, N-Diisopropylethylamine (DIEA), 4-Dimethylaminopyridine (DMAP), pyridine, sodium acetate, sodium carbonate or potassium carbonate, preferably triethylamine; the solvent is selected from methanol, ethanol, dichloromethane, acetone, ethyl acetate, tetrahydrofuran or a mixed solvent of any two or three of the solvents, preferably dichloromethane; the reaction temperature is selected from 0 ℃ to 50 ℃, preferably 0The temperature is between 30 and C; the reaction time is 1 to 10 hours, preferably 3 to 6 hours. The compound X: compound IX-1: a chlorination reagent: the acid binding agent is in a molar ratio of 1:1:1: 1-5: 1:5:5, preferably 1:1:1: 1-2: 1:2: 2.
The invention also discloses a pharmaceutical composition which contains the compound of the general formula I or pharmaceutically acceptable salt thereof and pharmaceutically acceptable auxiliary materials. The compound can be added with pharmaceutically acceptable adjuvants to make into common medicinal preparations, such as tablet, capsule, syrup, suspending agent, and optionally flavoring agent, sweetener, liquid or solid filler or diluent.
The invention relates to application of benzoyl piperidine derivatives with a general formula I or pharmaceutically acceptable salts thereof in preparation of SMO protein inhibitor drugs.
The application of the benzoyl piperidine derivative with the general formula I and the stereoisomer, hydrate, solvate or crystal thereof in preparing SMO protein inhibitor drugs is also within the protection scope of the invention.
The application of the benzoyl piperidine derivative with the general formula I or the pharmaceutically acceptable salt thereof in preparing the antitumor drugs is also within the protection scope of the invention.
Furthermore, the SMO protein inhibitor is used for preparing a medicament for treating malignant tumors.
Has the advantages that: the invention discloses a novel benzoyl piperidine derivative shown in a general formula I, and pharmacological experiments show that the compound I can generate a good inhibition effect on SMO protein and can be used for preparing a medicament for treating malignant tumors with excessive activation of a Hedgehog pathway; the invention also discloses a preparation method of the benzoyl piperidine derivative.
Detailed Description
The present application will be described in detail with reference to specific examples.
Example 1
Synthesis of N-methyl-N- (1- (2- ((1-methyl-1H-pyrazol-5-yl) amino) benzoyl) piperidin-4-yl) -2-trifluoromethyl-4-fluorobenzamide (I-1)
Synthesis of 2- ((1-methyl-1H-pyrazol-5-yl) amino) -benzoic acid (IV)
O-iodobenzoic acid (II) (5.00g,20.00mmol), 1-methyl-1-H-5-aminopyrazole (III) (1.90g,20.00mmol), potassium carbonate (5.50g,40.00mmol) and CuI (0.76g,4.00mmol) were dissolved in DMF (20.00ml) and stirred at 100-105 ℃ for 8H. And detecting the completion of the reaction of the raw material II by TLC (petroleum ether: ethyl acetate ═ 1: 1), stopping heating, and cooling to room temperature. Pouring the reaction solution into water (60.00ml), stirring for 20min, adjusting the pH value of the reaction system to 5-6 by using 1.00mol/L HCl, precipitating brown solid, performing suction filtration, and drying to obtain 3.10g of brown solid, wherein the yield is 70%, and the m.p.134-139 ℃.
1H-NMR(300MHz,CDCl3)δ(ppm):10.77-8.78(m,2H,NH,ArH),8.14-7.30(m,3H,ArH),6.89(s,1H,ArH),6.46(s,1H,ArH),3.85(s,3H,C 3H).
Synthesis of tert-butyl 4- (methylamino) piperidine-1-carboxylate (VI)
N-Boc-piperidin-4-one (V) (1.00g,5.00mmol) and a methylamine alcohol solution (1.00ml) were dissolved in methanol (10.00ml), adjusted to pH 5 to 6 with acetic acid, and stirred at room temperature for 1 hour. Sodium cyanoborohydride (0.48g,7.60mmol) was added portionwise in an ice bath and reacted at room temperature for 12 h. TLC (dichloromethane: methanol ═ 10:1) detected that starting material V was reacted completely. Methanol was dried by spinning, pH adjusted to 9 with saturated NaOH solution, extracted with dichloromethane (10.00ml x 3), dried over anhydrous sodium sulfate, filtered and solvent dried by spinning to give 1.00g of light yellow oil, 100% yield.
1H-NMR(300MHz,CDCl3)δ(ppm):3.98(d,J＝10.20Hz,2H,Boc-NC 2H),2.74(t,J＝12.20Hz,2H,BocNC 2H),2.48-2.43(m,1H,NHCH),2.38(s,3H,NHC 3H),1.79(d,J＝12.20Hz,2H,NHCHC 2H),1.39(s,9H,Boc-H),1.17(dd,J＝20.20,10.20Hz,2H,NHCHC 2H).
Synthesis of 4- (4-fluoro-N-methyl-2- (trifluoromethyl) benzoylamino) piperidine-1-carboxylic acid tert-butyl ester (VIII-1)
2-trifluoromethyl-4-fluorobenzoic acid (VII-1) (3.00g, 14.40mmol) was dissolved in dichloromethane (5.00ml), 2 drops of DMF were added, and then cooled to 0 deg.C, oxalyl chloride (1.50ml, 17.30mmol) was added dropwise, and the mixture was stirred at room temperature for 30min, and dichloromethane was dried to obtain acid chloride.
N-Boc-4-methylaminopiperidine (VI) (3.70g, 17.30mmol) was dissolved in methylene chloride (20.00ml), triethylamine (3.00ml, 21.60mmol) was added thereto, the mixture was cooled to 0 ℃ and a methylene chloride solution of acid chloride was added dropwise thereto, followed by stirring at room temperature for 1 hour. The reaction was complete by TLC (petroleum ether: ethyl acetate 3: 1). The reaction mixture was washed with 1.00mol/L HCl, saturated aqueous sodium bicarbonate solution, and saturated brine, dried over anhydrous sodium sulfate, and separated by column chromatography (petroleum ether: ethyl acetate: 6: 1) to obtain 5.33g of a colorless oil with a yield of 91.30%.
Major rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.44-7.39(m,1H,ArH),7.32-7.29(m,2H,ArH),4.73(m,1H,CH3NCH),4.25(m,2H,BocNC 2H),2.88-2.82(m,2H,BocNC 2H),2.63(s,3H,NC 3H),1.79-1.63(m,4H,CH3N(C 2 2H)),1.46(s,9H,Boc-H).
Minor rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.44-7.39(m,1H,ArH),7.32-7.29(m,2H,ArH),4.15-4.11(m,2H,BocNC 2H),3.23(s,1H,CH3NCH),2.98(s,3H,NC 3H),2.51-2.35(m,2H,BocNC 2H),1.79-1.63(m,4H,CH3N(C 2 2H)),1.43(s,9H,Boc-H).
Synthesis of 4-fluoro-N-methyl-N- (piperidin-4-yl) -2-trifluoromethylbenzamide (IX-1)
Compound VIII-1(4.33g,10.70mmol) was dissolved in methylene chloride (20.00ml), and trifluoroacetic acid (5.00ml) was added thereto at room temperature, followed by stirring at room temperature for 1 hour. TLC (dichloromethane: methanol 10:1) detected the starting material had reacted, the solvent was dried, water (15.00ml) was added, the pH adjusted to 9 with saturated NaOH solution, extracted with dichloromethane (15.00ml x 3), dried over anhydrous sodium sulphate to give 3.20g of a pale yellow oil in 98% yield.
Major rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.46-7.41(m,1H,ArH),7.33-7.30(m,2H,ArH),5.66(br,1H,NH),4.78(m,1H,CH3NCH),3.44-3.40(m,2H,HNC 2H),2.98-2.90(m,2H,BocNC 2H),2.68(s,3H,NC 3H),2.09-1.58(m,4H,CH3N(C 2 2H)).
Minor rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.46-7.41(m,1H,ArH),7.33-7.30(m,2H,ArH),5.66(brs,1H,NH),3.27-3.13(m,3H,CH3NCHCH2C 2HNH),3.02(s,3H,NC 3H),2.51-2.37(m,2H,BocNC 2H),2.09-1.58(m,4H,CH3N(C 2 2H)).
Synthesis of N-methyl-N- (1- (2- ((1-methyl-1H-pyrazol-5-yl) amino) benzoyl) piperidin-4-yl) -2-trifluoromethyl-4-fluorobenzamide (I-1)
Intermediate IV (500.00mg,2.30mmol) and intermediate IX-1(699.00mg,2.30mmol) were dissolved in dichloromethane (20.00ml), HOBt (372.00mg,2.76mmol) was added, EDCI (530.00mg,2.76mmol) was added in portions and stirred at room temperature for 12 h. TLC (dichloromethane: methanol 25: 1) detects that the raw material IX-1 has reacted completely, stops the reaction, and is washed with 1.00mol/L HCl (10.00mlx2), saturated sodium bicarbonate (10.00ml x2), and saturated brine (10.00ml x2) in this order, dried over anhydrous sodium sulfate, filtered, the filtrate is concentrated to dryness under reduced pressure, and the residue is separated by column chromatography (dichloromethane: methanol 200: 1 to 100: 1) to obtain 0.74g of dark green solid, yield 64.00%, m.p.206 to 207 ℃.
Major rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.47-7.41(m,2H,ArH),7.34-7.29(m,3H,ArH),7.27-7.12(m,2H,ArH,NH),6.91-6.75(m,2H,ArH),6.06(s,1H,ArH),5.00-4.81(m,1H,CH3NCH),4.80-4.12(m,2H,CON(C 2 2H)),3.72(s,3H,CH3),3.29-3.03(m,2H,CON(C 2 2H)),2.67(s,3H,CH3),1.84(m,4H,NCH(C 2 2H)).
Minor rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.47-7.41(m,2H,ArH)7.34-7.29(m,3H,ArH),7.27-7.12(m,2H,ArH),6.91-6.75(m,2H,ArH),6.06(s,1H,ArH),4.80-4.12(m,2H,CON(C 2 2H)),3.70(s,3H,CH3),3.43-3.60(m,1H,CH3NCH),3.02(s,3H,CH3),2.64-2.55(m,2H,CON(C 2 2H)),1.84(m,4H,NCH(C 2 2H)).
Major rotamer:13C-NMR(75MHz,CDCl3)δ(ppm):169.54,167.52,161.65(d,J＝251.50Hz),143.59,138.09,130.89,128.91,128.80,128.50,128.40,127.68,122.30(q,J＝276.20Hz),119.18,118.65(d,J＝36.10Hz),118.47,114.64,113.95(ddd,J＝25.30,9.20,4.70Hz),98.11,50.54,34.46,31.11,28.64,27.95.
Minor rotamer:13C-NMR(75MHz,CDCl3)δ(ppm):169.54,167.52,161.65(d,J＝251.50Hz),143.59,138.09,131.02,128.91,128.80,128.50,128.40,127.54,122.30(q,J＝276.20Hz),119.18,118.65(d,J＝36.10Hz),118.47,114.64,113.95(ddd,J＝25.30,9.20,4.70Hz),98.25,56.13,34.46,29.41,28.99,27.02.
HRMS(ESI):m/z[M+H]+.Calcd for C25H26F4N5O2:504.2023；found 504.2008.
IR(cm-1):3345.86,1630.91,1589.00,1559.42,1503.80,1467.52,1445.43,1422.09,1367.50,1330.25,1319.31,1271.16,1210.59,1171.33,1074.76,1040.48,1008.04,904.29,859.78,760.33,659.16,606.26.
Example 2
Synthesis of N-methyl-N- (1- (2- ((1-methyl-1H-pyrazol-5-yl) amino) benzoyl) piperidin-4-yl) -4-nitrobenzamide (I-2)
Synthesis of 4- (N-methyl-4-nitrobenzamido) piperidine-1-carboxylic acid tert-butyl ester (VIII-2)
Using 4-nitrobenzoic acid (VII-2) (2.40g, 14.40mmol) as the starting material, the same procedure as VIII-1 was followed, and column chromatography (petroleum ether: ethyl acetate: 6: 1) was performed to give 4.30g of a pale yellow solid, yield: 82 percent and m.p.152-153 ℃.
1H-NMR(300MHz,CDCl3)δ(ppm):8.31(d,J＝7.90Hz,2H,ArH),7.58(d,J＝7.20Hz,2H,ArH),4.72(s,1H,CONCH),4.26(brs,2H,CONCHC 2H),3.01(brs,2H,CONCHC 2H),1.77(s,3H,CONC 3H),1.49(brs,4H,BocN(C 2 2H)),1.49(s,9H,Boc-H).
Synthesis of N-methyl-4-nitro-N- (piperidin-4-yl) benzamide (IX-2)
Starting from VIII-2(4.89g,10.70mmol), the same procedure as IX-1 gave 2.70g of a pale yellow oil, yield: 97 percent. Synthesis of N-methyl-N- (1- (2- ((1-methyl-1H-pyrazol-5-yl) amino) benzoyl) piperidin-4-yl) -4-nitrobenzamide (I-2)
Using IV (500.00mg,2.30mmol), IX-2(605.00mg,2.30mmol) as raw materials, the same procedure as I-1 was followed, and column chromatography (dichloromethane: methanol 200: 1-100: 1) was performed to obtain 659.00mg of pale yellow solid, yield: 62 percent and m.p.91-92 ℃.
1H-NMR(300MHz,CDCl3)δ(ppm):8.31(d,J＝8.40Hz,2H,ArH),7.58(d,J＝8.40Hz,2H,ArH),7.47(s,1H,ArH),7.30-7.16(m,3H,ArH,NH),6.90-6.81(m,2H,ArH),6.04(s,1H,ArH),4.85(s,1H,CH3NCH),4.69-4.30(m,2H,CONC 2H),3.73(s,3H,CH3),3.14-3.01(m,2H,CONC 2H),2.84(s,3H,CH3),1.89-1.74(m,4H,CONCH(C 2 2H)).
13C-NMR(75MHz,CDCl3)δ(ppm):170.06,169.34,148.38,144.18,142.61,138.58,131.44,128.11,127.87,127.74,127.67,123.95,118.91,115.14,98.42,51.62,34.94,32.08,29.97,29.02.
HRMS(ESI):m/z[M+H]+.Calcd for C24H27N6O4:463.2094；found 463.2081.
IR(cm-1):3243.39,2934.42,1625.07,1557.39,1522.37,1445.61,1351.24,1328.59,1274.16,1070.27,1016.01,924.94,856.70,759.52,698.26.
Example 3
Synthesis of N-methyl-N- (1- (2- ((1-methyl-1H-pyrazol-5-yl) amino) benzoyl) piperidin-4-yl) 4-methoxybenzamide (I-3)
Synthesis of 4- (4-methoxy-N-methylbenzamido) piperidine-1-carboxylic acid tert-butyl ester (VIII-3)
Using 4-methoxybenzoic acid (VII-3) (2.20g, 14.40mmol) as the starting material, the procedure was as described for VIII-1, and column chromatography (petroleum ether: ethyl acetate: 6: 1) separated to give 4.40g of a white solid, yield: 88% and m.p.116-117 ℃.
1H-NMR(300MHz,CDCl3)δ(ppm):7.39(d,J＝8.50Hz,2H,ArH),6.94(d,J＝8.60Hz,2H,ArH),4.62(brs,1H,CONCH),4.25-4.21(m,2H,CONCHC 2H),3.87(s,3H,OCH3),2.91(s,3H,NCH3),2.77(brs,1H,ONCHC 2H),1.73(brs,4H,BocN(C 2 2H)),1.51(s,9H,Boc-H).
Synthesis of 4-methoxy-N-methyl-N- (piperidin-4-yl) benzamide (IX-3)
Starting from VIII-3(3.70g,10.70mmol), the same procedure as IX-1 gave 2.56g of a colorless oil, yield: 96.30 percent. Synthesis of N-methyl-N- (1- (2- ((1-methyl-1H-pyrazol-5-yl) amino) benzoyl) piperidin-4-yl) 4-methoxybenzamide (I-3)
Using IV (500.00mg,2.30mmol), IX-3(570.00mg,2.30mmol) as raw materials, the same procedure as I-1 was followed, and column chromatography (dichloromethane: methanol 200: 1-100: 1) was performed to obtain 740.00mg of pale yellow solid, yield: 72% and m.p.162-163 ℃.
1H-NMR(300MHz,CDCl3)δ(ppm):7.47(d,J＝1.80Hz,1H,ArH),7.38(d,J＝8.70Hz,2H,ArH),7.29(s,1H,NH),7.24-7.21(m,2H,ArH),6.94(d,J＝8.70Hz,2H,ArH),6.87(t,J＝7.40Hz,1H,ArH),6.81(d,J＝8.20Hz,1H,ArH),6.05(d,J＝1.70Hz,1H,ArH),4.76-4.32(m,3H,NCH3CH,CON(C 2 2H)),3.86(s,3H,OCH3),3.73(s,3H,CH3),3.16-2.96(m,2H,CON(C 2 2H)),2.91(s,3H,CH3),1.93-1.74(m,4H,NCH3CH(C 2 2H)).
13C-NMR(75MHz,CDCl3)δ(ppm):171.63,169.97,160.73,144.08,139.87,138.59,131.30,128.94,128.68,128.11,119.88,118.90,115.04,113.78,98.49,55.33,50.97,34.95,29.74,29.43,27.61.
HRMS(ESI):m/z[M+H]+.Calcd for C25H30N5O3:448.2349；found 448.2338.
IR(cm-1):3469.03,3242.52,2959.32,2926.25,1618.90,1589.14,1562.60,1512.59,1442.84,1429.01,1362.50,1325.73,1303.34,1255.14,1168.93,1068.02,1031.51,1007.47,926.08,846.61,762.88,593.99.
Example 4
Synthesis of N-methyl-N- (1- (2- ((1-methyl-1H-pyrazol-5-yl) amino) benzoyl) piperidin-4-yl) -4-fluorobenzamide (I-4)
Synthesis of 4- (4-fluoro-N-methylbenzamido) piperidine-1-carboxylic acid tert-butyl ester (VIII-4)
Using 4-fluorobenzoic acid (VII-4) (2.00g, 14.40mmol) as a starting material, the same procedure as VIII-1 was followed, and column chromatography (petroleum ether: ethyl acetate: 6: 1) was performed to give 3.68g of a pale yellow oil, yield: 76 percent.
1H-NMR(300MHz,CDCl3)δ(ppm):7.53-7.34(m,2H,ArH),7.15-7.08(m,2H,ArH),4.67(s,1H,CONCH),4.23(s,2H,CONCHC 2H),3.27-3.00(m,2H,CONCHC 2H),2.88(s,3H,NCH3),1.72(brs,4H,ONCHC 2H),1.49(s,9H,Boc-H).
Synthesis of 4-fluoro-N-methyl-N- (piperidin-4-yl) benzamide (IX-4)
The same procedure as for IX-1 was carried out using VIII-4(3.60g,10.70mmol) as the starting material to give 2.47g of a pale yellow oil, yield: 98 percent. Synthesis of N-methyl-N- (1- (2- ((1-methyl-1H-pyrazol-5-yl) amino) benzoyl) piperidin-4-yl) -4-fluorobenzamide (I-4)
Using IV (500.00mg,2.30mmol), IX-4(543.00mg,2.30mmol) as raw materials, the same procedure as I-1, and separating by column chromatography (dichloromethane: methanol 200: 1-100: 1) to obtain a grey solid, yield: 58% and m.p.87-88 ℃.
1H-NMR(300MHz,CDCl3)δ(ppm):7.47-7.42(m,3H,ArH),7.28-7.20(m,3H,ArH,NH),7.15-7.09(m,2H,ArH),6.89-6.80(m,2H,ArH),6.04(s,1H,ArH),5.02-4.60(m,1H,CH3NCH),4.60-4.20(m,2H,CON(C 2 2H)),3.72(s,3H,CH3),3.23-2.97(m,2H,CON(C 2 2H)),2.89(s,3H,CH3),1.92-1.80(m,4H,CH(C 2 2H)).
13C-NMR(75MHz,CDCl3)δ(ppm):170.75,170.01,163.34(d,J＝250.20Hz),144.12,139.86,138.57,132.59,131.35,129.00,128.11,119.73,118.91,115.63(d,J＝21.70Hz),115.09,98.46,45.05,34.94,29.66,29.36,28.83.
HRMS(ESI):m/z[M+H]+.Calcd for C24H27FN5O2:436.2149；found 436.2141.
IR(cm-1):3522.58,3271.66,2939.39,2859.62,1624.01,1557.39,1445.79,1368.77,1325.66,1274.19,1222.40,1158.03,1065.48,1005.75,926.05,894.43,848.71,760.00,729.00,583.85.
Example 5
Synthesis of N-methyl-N- (1- (2- ((1-methyl-1H-pyrazol-5-yl) amino) benzoyl) piperidin-4-yl) -4-cyanobenzamide (I-5)
Synthesis of 4- (4-cyano-N-methylbenzoylamino) piperidine-1-carboxylic acid tert-butyl ester (VIII-5)
Using 4-cyanobenzoic acid (VII-5) (2.10g, 14.4mmol) as a starting material, the same procedure as VIII-1 was followed, and separation was performed by column chromatography (petroleum ether: ethyl acetate: 6: 1) to obtain 3.50g of a white solid, yield: 71% and m.p.120-121 ℃.
1H-NMR(300MHz,CDCl3)δ(ppm):7.76(d,J＝8.00Hz,2H,ArH),7.51(d,J＝7.30Hz,2H,ArH),4.71(s,1H，CONCH),4.27(brs,2H,CONCHC 2H),3.00-2.89(m,1H,CONCHC 2H),2.81(s,3H,NCH3),2.51(s,1H,CONCHC 2H),1.77-1.71(m,4H,BocN(C 2 2H)),1.50(s,9H,Boc-H).
Synthesis of 4-cyano-N-methyl-N- (piperidin-4-yl) benzamide (IX-5)
The same procedure as for IX-1 was carried out using VIII-5(3.67g,10.70mmol) as the starting material to give 2.40g of a colorless oil, yield: 92 percent. Synthesis of N-methyl-N- (1- (2- ((1-methyl-1H-pyrazol-5-yl) amino) benzoyl) piperidin-4-yl) -4-cyanobenzamide (I-5)
Using IV (500.00mg,2.30mmol), IX-5(559.00mg,2.30mmol) as raw materials, the same procedure as I-1, and separating by column chromatography (dichloromethane: methanol 200: 1-100: 1) to obtain 559.00mg of yellow-green solid, yield: 55 percent and m.p.204-205 ℃.
1H-NMR(300MHz,CDCl3)δ(ppm):7.74(d,J＝7.50Hz,2H,ArH),7.61-7.35(m,3H,ArH),7.29-7.20(m,3H,ArH,NH),6.9-6.81(m,2H,ArH),6.04(s,1H,ArH),4.92-4.72(m,1H,CH3NCH),4.69-4.24(m,2H,CON(C 2 2H)),3.72(s,3H,CH3),3.22-2.97(m,2H,CON(C 2 2H)),2.83(s,3H,CH3),1.98-1.81(m,4H,CH(C 2 2H)).
13C-NMR(75MHz,CDCl3)δ(ppm):170.04,169.58,144.17,140.84,139.84,138.58,132.49,131.42,128.11,127.57,127.48,118.91,117.94,115.13,113.58,98.43,51.55,34.94,32.14,29.04,27.93.
HRMS(ESI):m/z[M+H]+.Calcd for C25H27N6O2:443.2195；found 443.2185.
IR(cm-1):3294.45,2938.25,2868.83,2230.93,1504.66,1586.66,1556.58,1447.14,1368.27,1329.95,1203.61,1069.66,1021.24,856.78,757.43,656.91,558.69.
Example 6
Synthesis of N-methyl-N- (1- (2- ((1-methyl-1H-pyrazol-5-yl) amino) benzoyl) piperidin-4-yl) -4-trifluoromethylbenzamide (I-6)
Synthesis of 4- (N-methyl-4- (trifluoromethyl) benzoylamino) piperidine-1-carboxylic acid tert-butyl ester (VIII-6)
Using 4-trifluoromethylbenzoic acid (VII-6) (2.70g, 14.40mmol) as a starting material, the same procedure as VIII-1 was followed, and separation was performed by column chromatography (petroleum ether: ethyl acetate ═ 6: 1) to obtain 3.89g of a white solid, yield: 70 percent and m.p.122-123 ℃.
1H-NMR(300MHz,CDCl3)δ(ppm):7.71(d,J＝8.00Hz,2H,ArH),7.52(d,J＝7.60Hz,2H,ArH),4.73(s,1H,CONCH),4.27(s,2H,CONCHC 2H),3.00(s,1H,CONCHC 2H),2.81(s,3H,NCH3),2.52(s,1H,CONCHC 2H),1.77(brs,4H,BocN(C 2 2H)),1.49(s,9H,Boc-H).
Synthesis of 4-trifluoromethyl-N-methyl-N- (piperidin-4-yl) benzamide (IX-6)
The same procedure as for IX-1 was carried out starting from VIII-6(4.10g,10.70mmol), giving 2.88g of a colorless oil, yield: 94 percent. Synthesis of N-methyl-N- (1- (2- ((1-methyl-1H-pyrazol-5-yl) amino) benzoyl) piperidin-4-yl) -4-trifluoromethylbenzamide (I-6)
Using IV (500.00mg,2.30mmol), IX-6(658.00mg,2.30mmol) as raw materials, the same procedure as I-1, and separating by column chromatography (dichloromethane: methanol 200: 1-100: 1) to obtain 703.00mg of yellow-green solid, yield: 63% and m.p.187-188 ℃.
1H-NMR(300MHz,CDCl3)δ(ppm):7.70(d,J＝7.40Hz,2H,ArH),7.60-7.38(m,3H,ArH),7.35-7.20(m,3H,ArH,NH),6.89-6.80(m,2H,ArH),6.04(s,1H,ArH),5.04-4.74(m,1H,CH3NCH),4.72-4.26(m,2H,CON(C 2 2H)),3.72(s,3H,CH3),3.30-3.02(m,2H,CON(C 2 2H)),2.85(s,3H,CH3),2.06-1.82(m,4H,CH(C 2 2H)).
13C-NMR(75MHz,CDCl3)δ(ppm):172.68,170.02,144.14,140.08,139.85,138.57,131.39,128.10,127.19,126.98,125.69,121.35(q,J＝271.20Hz),119.55,118.90,115.10,98.45,51.34,34.93,32.05,30.03,29.14.
HRMS(ESI):m/z[M+H]+.Calcd for C25H27F3N5O2:486.2117；found 486.2101.
IR(cm-1):3326.03,2940.53,1624.30,1582.28,1556.33,1446.57,1369.62,1325.64,1276.02,1170.31,1123.75,1071.10,1016.11,925.88,854.57,757.17,663.81.
Example 7
Synthesis of N-methyl-N- (1- (2- ((1-methyl-1H-pyrazol-5-yl) amino) benzoyl) piperidin-4-yl) -4-acetylbenzamide (I-7)
Synthesis of 4- (4-acetyl-N-methylbenzamido) piperidine-1-carboxylic acid tert-butyl ester (VIII-7)
Using 4-formylbenzoic acid (VII-7) (2.40g, 14.40mmol) as raw material, the same procedure as VIII-1 was followed, and column chromatography (petroleum ether: ethyl acetate: 6: 1) was performed to obtain 2.70g of pale yellow solid, yield: 52 percent and m.p.101-102 ℃.
1H-NMR(300MHz,CDCl3)δ(ppm):8.02(d,J＝8.10Hz,2H,ArH),7.48(d,J＝7.60Hz,2H,ArH),4.72(s,1H,CONCH),4.25(brs,2H,CONCHC 2H),2.94-2.89(m,1H,CONCHC 2H),2.80(s,3H,NCH3),2.64(s,3H,COCH3),2.50-2.43(m,1H,CONCHC 2H),1.76(brs,4H,BocN(C 2 2H)),1.48(s,9H,Boc-H).
Synthesis of 4-acetyl-N-methyl-N- (piperidin-4-yl) -benzamide (IX-7)
The same procedure as for IX-1 was carried out using VIII-7(3.85g,10.70mmol) as the starting material to give 2.58g of a pale yellow oil, yield: 93 percent. Synthesis of N-methyl-N- (1- (2- ((1-methyl-1H-pyrazol-5-yl) amino) benzoyl) piperidin-4-yl) -4-acetylbenzamide (I-7)
Using IV (500.00mg,2.30mmol), IX-7(598.00mg,2.30mmol) as raw materials, the same procedure as I-1 was followed, and column chromatography (dichloromethane: methanol 200: 1-100: 1) was performed to obtain 538.00mg of brown solid, yield: 51% and m.p.177-178 ℃.
1H-NMR(300MHz,CDCl3)δ(ppm):8.02(d,J＝8.00Hz,2H,ArH),7.50-7.40(m,3H,ArH),7.40-7.13(m,3H,ArH,NH),6.89-6.80(m,2H,ArH),6.04(s,1H,ArH),5.03-4.72(m,1H,CH3NCH),4.69-4.28(m,2H,CON(C 2 2H)),3.72(s,3H,CH3),3.30-3.02(m,2H,CON(C 2 2H)),2.84(s,3H,CH3),2.64(s,3H,COCH3),2.03-1.70(m,4H,CH(C 2 2H)).
13C-NMR(75MHz,CDCl3)δ(ppm):172.63,170.56,170.01,144.15,140.92,139.85,138.58,137.82,131.38,128.58,128.10,127.00,126.77,118.90,115.10,98.48,51.36,34.94,32.04,29.67,29.19,26.62.
HRMS(ESI):m/z[M+H]+.Calcd for C26H30N5O3:460.2349；found 460.2338.
IR(cm-1):3327.23,2944.82,2865.06,1683.88,1616.87,1556.60,1504.68,1468.81,1366.06,1327.17,1265.23,1079.76,1065.61,1013.66,863.06,851.80,757.45,602.99.
Example 8
Synthesis of N-methyl-N- (1- (2- ((1-methyl-1H-pyrazol-5-yl) amino) benzoyl) piperidin-4-yl) -4-methylsulfonylbenzamide (I-8)
Synthesis of 4- (N-methyl-4- (methylsulfonyl) benzoylamino) piperidine-1-carboxylic acid tert-butyl ester (VIII-8)
Using 4-methylsulfonylbenzoic acid (VII-8) (3.10g, 14.40mmol) as the starting material, the same procedure as VIII-1 was followed, and column chromatography (petroleum ether: ethyl acetate: 3: 1) was performed to obtain 3.90g of a white solid, yield: 69 percent and m.p.196-197 ℃.
1H-NMR(300MHz,CDCl3)δ(ppm):8.01(d,J＝8.10Hz,2H,ArH),7.57(d,J＝7.40Hz,2H,ArH),4.69(m,1H,CONCH),4.44-4.00(m,2H,CONCHC 2H),3.07(s,3H,SO2C 3H),3.03-2.81(m,2H,CONCHC 2H),2.77(s,3H,NCH3),1.85-1.60(m,4H,BocN(C 2 2H)),1.46(s,9H,Boc-H).
Synthesis of 4-methylsulfonyl-N-methyl-N- (piperidin-4-yl) benzamide (IX-8)
Starting from VIII-8(4.20g,10.70mmol), the same procedure as IX-1 gave 3.00g of a colorless oil, yield: 96 percent. Synthesis of N-methyl-N- (1- (2- ((1-methyl-1H-pyrazol-5-yl) amino) benzoyl) piperidin-4-yl) -4-methylsulfonylbenzamide (I-8)
Using IV (500.00mg,2.30mmol), IX-8(681.00mg,2.30mmol) as starting material, the same procedure as I-1 was followed, separation by column chromatography (dichloromethane: methanol ═ 150: 1) gave 831.00mg of a dark green solid, yield: 73 percent and m.p.107-108 ℃.
1H-NMR(300MHz,CDCl3)δ(ppm):8.03(d,J＝7.70Hz,2H,ArH),7.59(d,J＝7.80Hz,2H,ArH),7.47(s,1H,NH),7.32-7.23(m,3H,ArH),6.89-6.80(m,2H,ArH),6.04(s,1H,ArH),4.99-4.81(m,1H,CH3NCH),4.68-4.23(m,2H,CON(C 2 2H)),3.72(s,3H,SO2CH3),3.23-2.95(m,5H,CONCH3,CON(C 2 2H)),2.83(s,3H,CH3),2.06-1.62(m,4H,CH(C 2 2H)).
13C-NMR(125MHz,CDCl3)δ(ppm):170.03,169.66,144.11,141.92,141.59,139.90,138.53,131.43,128.13,128.01,127.88,127.76,118.96,115.16,98.40,51.53,44.39,34.94,32.21,29.67,29.02.
HRMS(ESI):m/z[M+H]+.Calcd for C25H30N5O4S:496.2019；found 496.2004.
IR(cm-1):3457.23,3343.06,2927.76,1624.62,1586.54,1556.50,1446.95,1428.07,1369.76,1326.92,1312.89,1277.30,1152.58,1065.37,1006.97,957.64,853.66,785.69,752.34,682.56,553.70.
Example 9
Synthesis of N-methyl-N- (1- (2- (phenylamino) benzoyl) piperidin-4-yl) -2-trifluoromethyl-4-fluorobenzamide (I-9) Synthesis of 4-fluoro-N- (1- (2-iodophenyl) piperidin-4-yl) -N-methyl-2- (trifluoromethyl) benzamide (XI)
Compound X (2.00g,8.00mmol) was dissolved in dichloromethane (10.00ml), 2 drops of DMF were added and cooled to 0 deg.C, oxalyl chloride (0.80ml, 9.60mmol) was added dropwise, stirring was carried out for 30min after displacement to room temperature, and dichloromethane was spin-dried to give the acid chloride for use.
IX-1(2.40g,8.00mmol) was dissolved in dichloromethane (20ml), triethylamine (1.70ml,12.00mmol) was added, cooling to 0 ℃ was effected, a dichloromethane solution of acid chloride was added dropwise, and the mixture was stirred at room temperature for 5 hours. The reaction was complete by TLC (petroleum ether: ethyl acetate 3: 1). Washing the reaction solution with 1N HCl, saturated sodium bicarbonate water solution and saturated brine respectively, drying with anhydrous sodium sulfate, filtering, spin-drying the solvent, and separating by column chromatography (petroleum ether: ethyl acetate: 8: 1) to obtain a light yellow solid 3.70g, yield 86%, m.p.85-86 ℃.
Major rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.91-7.81(m,1H,ArH),7.45-7.36(m,2H,ArH),7.36-7.28(m,2H,ArH),7.24-7.17(m,1H,ArH),7.14-7.02(m,1H,ArH),5.00-4.75(m,1H,CH3NCH),4.23(brs,2H,NCHC 2H),3.07(br,2H,NCHC 2H),2.58(s,3H,CH3),1.75(m,4H,CON(C 2 2H)).
Minor rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.91-7.81(m,1H,ArH),7.45-7.36(m,2H,ArH),7.36-7.28(m,2H,ArH),7.24-7.17(m,1H,ArH),7.14-7.02(m,1H,ArH),4.23(brs,2H,NCHC 2H),3.38(m,1H,CH3NCH),3.02(s,3H,CH3),2.41(brs,2H,NCHC 2H),1.75(m,4H,CON(C 2 2H)).
Synthesis of N-methyl-N- (1- (2- (phenylamino) benzoyl) piperidin-4-yl) -2-trifluoromethyl-4-fluorobenzamide (I-9)
XI (500.00mg,0.90mmol), aniline (XII-1) (84.00mg,0.90mmol), palladium acetate (20.00mg,0.09mmol) and BINAP (112.00mg,0.18mmol) were dissolved in toluene (10.00ml) and heated to 100 ℃ under nitrogen for 12 h. TLC (dichloromethane: methanol ═ 35: 1) detected complete reaction of starting materials. Cooling the reaction solution, performing suction filtration on the reaction solution by using kieselguhr, spin-drying the solvent, and separating by using column chromatography (an eluent: dichloromethane: methanol: 300: 1-100: 1) to obtain 300.00mg of light yellow solid, wherein the yield is as follows: 67%, m.p.164-165 ℃.
Major rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.45-7.38(m,2H,ArH),7.30-7.20(m,6H,ArH),7.14-7.03(m,2H,ArH),6.96-6.87(m,2H,ArH),4.88-4.78(m,1H,CH3NCH),4.67-4.00(m,2H,CON(C 2 2H)),3.16-2.91(m,2H,CON(C 2 2H)),2.47(s,3H,CH3),1.76-1.58(m,4H,NCH(C 2 2H)).
Minor rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.45-7.38(m,2H,ArH),7.30-7.20(m,6H,ArH),7.14-7.03(m,2H,ArH),6.96-6.87(m,2H,ArH),4.67-4.00(brs,2H,CON(C 2 2H)),3.4.-3.30(m,1H,CH3NCH),2.83(s,3H,CH3),2.72-2.56(m,2H,CON(C 2 2H)),1.76-1.58(m,4H,NCH(C 2 2H)).
Major rotamer:13C-NMR(75MHz,CDCl3)δ(ppm):169.35,167.44,161.60(d,J＝250.10Hz),142.01,141.26,141.22,129.94,128.95,128.88,128.79,128.47,128.36,127.31,123.64,120.78,119.68,118.97(d,J＝21.30Hz),117.66,113.89(dq,J＝9.40,4.20Hz),50.35,30.78,28.45,27.69.
Minor rotamer:13C-NMR(75MHz,CDCl3)δ(ppm):169.33,167.44,161.60(d,J＝250.10Hz),142.01,141.26,141.22,130.03,128.95,128.88,128.79,128.47,128.36,127.15,123.64,120.97,119.68,118.97(d,J＝21.30Hz),117.66,113.89(dq,J＝9.40,4.20Hz),56.08,29.18,28.75,26.81.
HRMS(ESI):m/z[M+H]+.Calcd for C27H26F4N3O2:500.1961；found 500.1949.
IR(cm-1):3044.25,2943.95,2867.26,1635.00,1594.22,1511.76,1450.29,1325.89,1279.31,1173.89,1136.30,1040.50,995.21,906.81,877.35,877.35,691.71,500.18.
Example 10
Synthesis of N-methyl-N- (1- (2- ((4-cyano) phenylamino) benzoyl) piperidin-4-yl) -2-trifluoromethyl-4-fluorobenzamide (I-10)
Using XI (500.00mg,0.90mmol), 4-cyanoaniline (XII-2) (106.00mg,0.90mmol) as a starting material, the same procedure as I-9 was followed, and column chromatography (eluent: dichloromethane: methanol 300: 1 to 100: 1) was performed to obtain 297.00mg of white powder, yield 63%, m.p.226 to 227 ℃.
Major rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.53(s,1H,ArH),7.48-7.45(m,3H,ArH),7.41-7.34(m,2H,ArH),7.29-7.27(m,2H,ArH),7.09(t,J＝6.00Hz,1H,ArH),7.04-7.01(m,2H,ArH),4.84-4.75(m,1H,CH3NCH),4.68-3.70(m,2H,CON(C 2 2H)),3.24-2.90(m,2H,CON(C 2 2H)),2.46(s,3H,CH3),1.76-1.69(m,2H,NCH(C 2 2H)),1.69-1.58(m,2H,NCH(C 2 2H)).
Minor rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.53(s,1H,ArH),7.48-7.45(m,3H,ArH),7.41-7.34(m,2H,ArH),7.29-7.27(m,2H,ArH),7.09(t,J＝6.00Hz,1H,ArH),7.04-7.01(m,2H,ArH),4.68-3.70(m,2H,CON(C 2 2H)),3.42-3.27(m,1H,CH3NCH),2.85(s,3H.CH3),2.72-2.57(m,2H,CON(C 2 2H)),1.76-1.69(m,2H,NCH(C 2H)2),1.69-1.58(m,2H,NCH(C 2 2H)).
Major rotamer:13C-NMR(125MHz,CDCl3)δ(ppm):169.05,167.96,162.14(d,J＝251.40Hz),147.27,138.81,133.73,131.63,131.31,130.56,129.31(d,J＝8.00Hz),128.50(q,J＝7.50Hz),128.04,124.9(q,J＝281.20Hz),122.8,121.05,119.52(d,J＝21.20Hz),119.51,115.67,114.43(d,J＝22.50Hz),102.13,50.82,31.27,28.88,28.23.
Minor rotamer:13C-NMR(125MHz,CDCl3)δ(ppm):169.05,167.53,162.21(d,J＝250.00Hz),146.95,139.02,133.73,131.63,131.31,130.70,129.31(d,J＝8.00Hz),128.50(q,J＝7.50Hz),127.96,124.9(q,J＝281.20Hz),122.63,120.64,119.52(d,J＝21.20Hz),119.51,115.86,114.43(d,J＝22.50Hz),102.45,56.42,28.88,28.23,27.26.
HRMS(ESI):m/z[M+H]+.Calcd for C28H25F4N4O2:525.1914found 525.1898.
IR(cm-1):3492.63,3290.46,2955.75,2926.25,2215.65,1633.13,1583.44,1516.89,1449.56,1411.71,1367.00,1319.46,1280.89,1161.66,1140.82,1040.35,1009.21,904.82,873.92,846.23,740.45,544.11.
Example 11
Synthesis of N-methyl-N- (1- (2- ((2-cyanophenyl) amino) benzoyl) piperidin-4-yl) -2-trifluoromethyl-4-fluorobenzamide (I-11)
Using XI (500.00mg,0.90mmol), 2-cyanoaniline (XII-3) (106.00mg,0.90mmol) as the starting material, the same procedure as I-9 was followed, and column chromatography was performed (eluent: dichloromethane: methanol 200: 1 to 100: 1) to obtain 240.00mg of white powder, 51% yield, m.p.187 to 188 ℃.
Major rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.54-7.49(m,1H,ArH),7.45-7.35(m,4H,ArH),7.33-7.27(m,4H,ArH),7.13-7.08(m,1H,ArH),6.97-6.89(m,1H,ArH),4.88-4.78(m,1H,CH3NCH),4.60-3.46(m,2H,CON(C 2 2H)),3.25-2.95(m,2H,CON(C 2 2H)),2.55(s,3H,CH3),1.90-1.57(m,4H,NCH(C 2 2H)).
Minor rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.54-7.49(m,1H,ArH),7.45-7.35(m,4H,ArH),7.33-7.27(m,4H,ArH),7.13-7.08(m,1H,ArH),6.97-6.89(m,1H,ArH),4.60-3.46(m,2H,CON(C 2 2H)),3.40-3.28(m,1H,CH3NCH),2.89(s,3H,CH3),2.70-2.64(m,2H,CON(C 2 2H)),1.90-1.57(m,4H,NCH(C 2 2H)).
Major rotamer:13C-NMR(75MHz,CDCl3)δ(ppm):168.25,167.45,161.59(d,J＝251.10Hz),145.71,138.20,133.40,132.75,131.29,129.96,128.94,128.83,128.48,128.38,127.70,125.32(q,J＝183.70Hz),122.50,120.31,120.04,119.84,119.00(d,J＝21.50Hz),116.63,113.87(dq,J＝14.70,5.00Hz),50.35,30.93,28.67,27.76.
Minor rotamer:13C-NMR(75MHz,CDCl3)δ(ppm):168.25,167.45,161.59(d,J＝251.10Hz),145.71,138.20,133.47,132.83,131.29,130.06,128.94,128.83,128.48,128.38,127.50,125.32(q,J＝183.70Hz),122.42,120.31,120.04,119.84,119.00(d,J＝21.50Hz),116.63,113.87(dq,J＝14.70,5.00Hz),56.04,28.67,27.76,26.86.
HRMS(ESI):m/z[M+H]+.Calcd for C28H25F4N4O2:525.1914；found 525.1899.
IR(cm-1):3461.31,3300.38,2935.60,2219.24,1633.64,1589.08,1516.10,1448.85,1422.55,1366.81,1326.76,1166.45,1122.68,1040.00,1005.65,905.71,876.54,757.01.
Example 12
Synthesis of N-methyl-N- (1- (2- ((4-methoxyphenyl) amino) benzoyl) piperidin-4-yl) -2-trifluoromethyl-4-fluorobenzamide (I-12)
Using XI (500.00mg,0.90mmol), 4-methoxyaniline (XII-4) (111.00mg,0.90mmol) as raw material, proceeding with I-9, separating by column chromatography (eluent: dichloromethane: methanol: 200: 1-100: 1) to obtain 476.00mg of white powder, yield 61%, m.p.165-166 ℃.
Major rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.45-7.39(m,1H,ArH),7.31(d,J＝6.50Hz,2H,ArH),7.25-7.00(m,5H,ArH),6.85(d,J＝8.90Hz,2H,ArH),6.82-6.74(m,1H,ArH),4.91-4.80(m,1H,CH3NCH),4.72-4.20(m,2H,CON(C 2 2H)),3.78(s,3H,OCH3),3.20-2.96(m,2H,CON(C 2 2H)),2.57(s,3H,CH3),1.81-1.59(m,4H,NCH(C 2 2H)).
Minor rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.45-7.39(m,1H,ArH),7.31(d,J＝6.50Hz,2H,ArH),7.25-7.00(m,5H,ArH),6.85(d,J＝8.90Hz,2H,ArH),6.82-6.74(m,1H,ArH),4.72-4.20(m,2H,CON(C 2 2H)),3.78(s,3H,OCH3),3.41-3.30(m,1H,CH3NCH),2.92(s,3H,CH3),2.80-2.62(m,2H,CON(C 2 2H)),1.81-1.59(m,4H,NCH(C 2 2H)).
Major rotamer:13C-NMR(125MHz,CDCl3)δ(ppm):170.18,167.94,162.12(d,J＝250.00Hz),155.47,144.10,135.03,131.83,130.60,129.40,129.34,127.89,122.76(q,J＝270.00Hz),122.48,121.74,119.46(d,J＝21.60Hz),118.51,115.73,114.74,114.40(d,J＝24.80Hz),55.56,51.00,31.42,29.05,28.39.
Minor rotamer:13C-NMR(125MHz,CDCl3)δ(ppm):170.25,167.58,162.18(d,J＝249.00Hz),155.57,144.13,134.88,131.91,130.69,129.40,129.34,127.73,122.76(q,J＝270.00Hz),122.48,121.50,119.46(d,J＝21.60Hz),118.51(s),115.73,114.74,114.40(d,J＝24.80Hz),56.66,55.56,29.84,29.39,27.39.
HRMS(ESI):m/z[M+H]+.Calcd for C28H28F4N3O3:530.2067；found 530.2056.
IR(cm-1):3374.40,2934.04,1636.33,1587.02,1513.36,1367.21,1328.60,1318.48,1247.04,1170.35,1136.40,1038.64,1004.04,905.98,876.43,767.36,749.28,519.68.
Example 13
Synthesis of N-methyl-N- (1- (2- ((2-methoxyphenyl) amino) benzoyl) piperidin-4-yl) -2-trifluoromethyl-4-fluorobenzamide (I-13)
Using XI (500.00mg,0.90mmol), 2-methoxyaniline (XII-5) (111.00mg,0.90mmol) as raw material, proceeding with I-9, separating by column chromatography (eluent: dichloromethane: methanol 300: 1-100: 1) to obtain 276.00mg of white powder, yield 58%, m.p. 219-220 ℃.
Major rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.45-7.35(m,2H,ArH),7.29-7.25(m,4H,ArH),7.17-7.00(m,1H,ArH),7.00-6.91(m,1H,ArH),6.88-6.84(m,3H,ArH),5.10-4.10(m,3H,CH3NCH,CON(C 2 2H)),3.86(s,3H,OCH3),3.16-2.90(m,2H,CON(C 2 2H)),2.44(s,3H,CH3),1.85-1.47(m,4H,NCH(C 2 2H)).
Minor rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):7.45-7.35(m,2H,ArH),7.29-7.25(m,4H,ArH),7.17-7.00(m,1H,ArH),7.00-6.91(m,1H,ArH),6.88-6.84(m,3H,ArH),5.08-4.09(m,2H,CON(C 2 2H)),3.86(s,3H,OCH3),3.38-3.23(m,1H,CH3NCH),2.77(s,3H,CH3),2.69-2.61(m,2H,CON(C 2 2H)),1.85-1.47(m,4H,NCH(C 2 2H)).
Major rotamer:13C-NMR(75MHz,CDCl3)δ(ppm):169.07,167.40,161.58(d,J＝250.80Hz),148.41,140.33,131.75,129.70,128.91,128.80,128.35,127.28,127.16,120.24,120.08,120.10,120.04,118.98(d,J＝21.10Hz),118.09,114.90,113.87(ddd,J＝13.80,8.80,4.10Hz),110.24,55.21,50.34,30.66,28.36,27.59.
Minor rotamer:13C-NMR(75MHz,CDCl3)δ(ppm):169.07,167.40,161.58(d,J＝250.80Hz),148.41,140.33,131.75,129.77,128.91,128.80,128.35,127.16,127.12,120.24,120.08,120.10,120.04,118.98(d,J＝21.10Hz),118.09,114.90,113.87(ddd,J＝13.80,8.80,4.10Hz),110.32,56.08,55.21,29.09,28.63,26.65.
HRMS(ESI):m/z[M+H]+.Calcd for C28H28F4N3O3:530.2067；found 530.2058.
IR(cm-1):3355.27,2991.15,1639.17,1589.19,1516.19,1420.47,1367.24,1327.27,1278.32,1244.99,1169.44,1130.41,1074.06,1039.91,906.12,874.10,751.72,642.40,606.02,560.94.
Example 14
Synthesis of N-methyl-N- (1- (2- (pyridin-4-ylamino) benzoyl) piperidin-4-yl) -2-trifluoromethyl-4-fluorobenzamide (I-14) starting from XI (500.00mg,0.90mmol), 4-aminopyridine (XII-6) (85.00mg,0.90mmol) by column chromatography (eluent: dichloromethane: methanol: 150: 1 to 100: 1) to give 198.00mg of pale yellow powder, 44% yield, m.p.136 to 137 ℃.
Major rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):8.40-8.19(m,2H,ArH),7.49-7.29(m,7H,ArH,NH),7.21-7.15(m,1H,ArH),6.93-6.78(m,2H,ArH),5.07-4.48(m,2H,CH3NCH,CON(C 2 2H)),4.19-3.68(m,1H,CON(C 2 2H)),3.25-2.91(m,2H,CON(CH2)2),2.41(s,3H,CH3),1.92-1.43(m,4H,NCH(C 2 2H)).
Minor rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):8.40-8.19(m,2H,ArH),7.49-7.29(m,7H,ArH,NH),7.21-7.15(m,1H,ArH),6.93-6.78(m,2H,ArH),5.07-4.48(m,1H,CON(C 2 2H)),4.19-3.68(m,1H,CON(C 2 2H)),3.42-3.27(m,1H,CH3NCH),2.85(s,3H,CH3),2.69-2.58(m,2H,CON(C 2 2H)),1.92-1.43(m,4H,NCH(C 2 2H)).
Major rotamer:13C-NMR(75MHz,CDCl3)δ(ppm):168.88,167.95,162.13(d,J＝247.60Hz),150.24,131.63,130.42,129.35,129.24,128.89(d,J＝7.90Hz),127.85,123.53,123.32,122.33,122.00,119.45(d,J＝21.20Hz),114.38(d,J＝20.20Hz),109.94,109.80,50.70,31.17,28.81,28.15.
Minor rotamer:13C-NMR(75MHz,CDCl3)δ(ppm):168.88,167.95,162.13(d,J＝247.60Hz),150.24,131.63,130.57,129.35,129.24,128.89(d,J＝7.90Hz),127.85,123.53,123.32,122.33,122.00,119.45(d,J＝21.20Hz),114.38(d,J＝20.20Hz),109.94,109.80,56.39,29.58,29.19,27.24.
HRMS(ESI):m/z[M+H]+.Calcd for C26H25F4N4O2:501.1914；found 501.1900.
IR(cm-1):3263.74,2930.64,1631.34,1592.68,1515.28,1422.00,1410.90,1370.34,1321.60,1282.23,1267.24,1214.83,1171.03,1134.24,1072.83,1041.27,997.24,905.95,817.03,757.23,528.80.
Example 15
Synthesis of N-methyl-N- (1- (2- (pyridin-2-ylamino) benzoyl) piperidin-4-yl) -2-trifluoromethyl-4-fluorobenzamide (I-15) starting from XI (500.00mg,0.90mmol), 2-aminopyridine (XII-7) (85.00mg,0.90mmol) by column chromatography (eluent: dichloromethane: methanol: 150: 1 to 100: 1) to give 162.00mg of pale yellow powder, 36% yield, m.p.155 to 156 ℃.
Major rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):8.30-8.16(m,1H,ArH),8.01(d,J＝8.3Hz,1H,ArH),7.84-7.66(m,2H,ArH),7.57-7.25(m,5H,ArH,NH),7.17-6.96(m,1H,ArH),6.96-6.67(m,2H,ArH),4.89-4.77(m,1H,CH3NCH),4.74-3.65(m,2H,CON(C 2 2H)),3.21-2.93(m,2H,CON(C 2 2H)),2.50(s,3H,CH3),1.93-1.50(m,4H,NCH(C 2 2H)).
Minor rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):8.30-8.16(m,1H,ArH),8.01(d,J＝8.30Hz,1H,ArH),7.84-7.66(m,2H,ArH,NH),7.57-7.25(m,5H,ArH),7.17-6.96(m,1H,ArH),6.96-6.67(m,2H,ArH),4.74-3.65(m,2H,CON(C 2 2H)),3.42-3.23(m,1H,CH3NCH),2.84(s,3H,CH3),2.69-2.61(m,2H,CON(C 2 2H)),1.93-1.50(m,4H,NCH(C 2 2H)).
Major rotamer:13C-NMR(75MHz,CDCl3)δ(ppm):122.64,169.54,162.13(d,J＝258.70Hz),155.39,148.07,148.01,138.77,137.66,130.43,129.38,129.28,128.96,128.86,127.50,121.82,121.35,119.42(d,J＝20.90Hz),115.37,114.50(d,J＝4.50Hz),109.98,50.84,31.26,28.94,28.16.
Minor rotamer:13C-NMR(75MHz,CDCl3)δ(ppm):122.64,169.54,162.13(d,J＝258.70Hz),155.39,148.07,148.01,138.77,137.75,130.52,129.38,129.28,128.96,128.86,127.37,121.82,121.35,119.42(d,J＝20.90Hz),115.54,114.17(d,J＝4.10Hz),109.98,56.50,29.16,28.45,27.23.
HRMS(ESI):m/z[M+H]+.Calcd for C26H25F4N4O2:501.1914；found 501.1903.
IR(cm-1):3392.33,2920.35,2861.36,2861.36,1592.16,1511.12,1434.99,1367.46,1329.28,1307.24,1173.57,1132.21,1075.27,1038.87,907.50,862.63,759.68,644.57,603.31,523.19.
Example 16
Synthesis of N-methyl-N- (1- (2- ([1, 1' -diphenyl ] -4-ylamino) benzoyl) piperidin-4-yl) -2-trifluoromethyl-4-fluorobenzamide (I-16)
Starting from XI (500.00mg,0.90mmol), 4-phenylaniline (XII-8) (152.00mg,0.90mmol), the procedure is as for I-9, and the product is separated by column chromatography (eluent: dichloromethane: methanol 300: 1 to 100: 1) to give 340.00mg of yellow powder, 66% yield, m.p.112 to 113 ℃.
Major rotamer:1H-NMR(500MHz,CDCl3)δ(ppm):7.62-7.49(m,5H,ArH),7.49-7.38(m,3H,ArH),7.38-7.25(m,5H,ArH,NH),7.19-7.16(m,3H,ArH),6.99(t,J＝7.50Hz,1H,ArH),4.91-4.84(m,1H,CH3NCH),4.82-3.79(m,2H,CON(C 2 2H)),3.30-2.95(m,2H,CON(C 2 2H)),2.52(s,3H,CH3),1.84-1.73(m,2H,NCH(C 2 2H)),1.69-1.62(m,2H,NCH(C 2 2H)).
Minor rotamer:1H-NMR(500MHz,CDCl3)δ(ppm):7.62-7.49(m,5H,ArH),7.49-7.38(m,3H,ArH),7.38-7.25(m,5H,ArH,NH),7.19-7.16(m,3H,ArH),6.99(t,J＝7.50Hz,1H,ArH),4.82-3.79(m,2H,CON(C 2 2H)),3.47-3.28(m,1H,CH3NCH),2.89(s,3H,C 3H),2.73-2.64(m,2H,CON(C 2 2H)),1.84-1.73(m,2H,NCH(C 2 2H)),1.69-1.62(m,2H,NCH(C 2 2H)).
Major rotamer:13C-NMR(125MHz,CDCl3)δ(ppm):169.81,167.92,162.09(d,J＝249.87Hz),141.99,141.62,140.69,134.08,131.77(dd,J＝4.30,2.30Hz),130.48,129.32(d,J＝8.10Hz),128.92(dd,J＝16.10,7.30Hz),128.72,127.97,127.88,126.67,126.46,122.99(q,J＝261.25Hz),120.33,119.43(d,J＝20.80Hz),118.49,118.28,118.24,114.35(dd,J＝24.80,4.60Hz),50.89,31.27,28.94,28.21.
Minor rotamer:13C-NMR(125MHz,CDCl3)δ(ppm):169.77,167.54,162.21(d,J＝250.25Hz),141.81,141.66,140.63,134.31,131.77(dd,J＝4.30,2.30Hz),130.58,129.32(d,J＝8.10Hz),128.92(dd,J＝16.10,7.30Hz),128.77,128.03,127.71,126.75,126.50,122.99(q,J＝261.250Hz),120.33,119.43(d,J＝20.80Hz),118.49,118.28,118.24,114.35(dd,J＝24.80,4.60Hz),56.59,29.72,29.31,27.27.
HRMS(ESI):m/z[M+H]+.Calcd for C33H30F4N3O2:576.2274；found 576.2257.
IR(cm-1):3376.30,3028.90,2930.00,2858.81,1631.67,1596.56,1523.77,1423.06,1369.58,1320.85,1279.21,1170.45,1134.23,1072.03,1040.87,1005.23,905.16,838.02,762.81,698.13,501.94.
Example 17
Synthesis of N-methyl-N- (1- (2- ((4-formylphenyl) amino) benzoyl) piperidin-4-yl) -2-trifluoromethyl-4-fluorobenzamide (I-17)
Using XI (500.00mg,0.90mmol), 4-aminobenzoic acid (XII-9) (123.00mg,0.90mmol) as raw material, the same procedure as I-9 was followed, and column chromatography (eluent: dichloromethane: methanol: 150: 1 to 100: 1) was performed to obtain 210.00mg of white powder, yield 43%, m.p.178 to 179 ℃.
Major rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):8.16-7.92(m,2H,ArH),7.55-7.51(m,2H,ArH),7.46-7.30(m,4H,ArH),7.22-7.01(m,3H,ArH),4.87-4.79(m,1H,CH3NCH),4.40-3.54(m,2H,CON(C 2 2H)),3.17-2.93(m,2H,CON(C 2 2H)),2.41(s,3H,CH3),1.93-1.41(m,4H,NCH(C 2 2H)).
Minor rotamer:1H-NMR(300MHz,CDCl3)δ(ppm):8.16-7.92(m,2H,ArH),7.55-7.51(m,2H,ArH),7.46-7.30(m,4H,ArH),7.22-7.01(m,3H,ArH),4.40-3.54(m,2H,CON(C 2 2H)),3.39-3.32(m,1H,CH3NCH),3.17-2.93(m,2H,CON(C 2 2H)),2.80(s,3H,CH3),1.93-1.41(m,4H,NCH(C 2 2H)).
Major rotamer:13C-NMR(75MHz,CDCl3)δ(ppm):170.84,169.34,167.99,162.09(d,J＝250.50Hz),156.98,148.45,139.09,132.16,130.46,129.36,129.26,128.98,127.81,124.42(q,J＝206.30Hz),122.78,121.53,120.43,119.46(d,J＝20.80Hz),114.88,114.37(d,J＝20.60Hz),50.73,31.16,28.81,28.10.
Minor rotamer:13C-NMR(75MHz,CDCl3)δ(ppm):170.72,169.34,167.99,162.09(d,J＝250.50Hz),156.98,148.45,139.09,132.16,130.58,129.36,129.26,128.98,127.68,124.42(q,J＝206.30Hz),122.78,121.32,120.74,119.46(d,J＝20.80Hz),114.88,114.37(d,J＝20.60Hz),56.41,29.63,29.15,27.20.
HRMS(ESI):m/z[M+H]+.Calcd for C28H26F4N3O4:544.1859；found 544.1845.
IR(cm-1):3474.93,3307.19,3067.85,2933.28,2507.37,1707.76,1522.02,1593.82,1451.29,1411.14,1370.89,1326.58,1280.08,1173.88,1134.95,1072.94,1041.42,905.55,844.43,769.54,750.47,660.97,500.78.
The pharmacological experiments and results of the partial compounds prepared in the above are as follows:
first, Gli-luciferase experiment
Principle of experiment
A firefly luciferase gene firefly driven by 12XGLI-binding site was introduced into NIH/3T3 cells by lentivirus stable transfection, while a Renilla luciferase gene renilla driven by CMV promoter was introduced. Wherein the firefly is used for detecting the transcriptional activity of a Hedgehog signal, and the renilla is used as an internal reference gene of a reporter gene. The Hedgehog ligand can be used for remarkably improving the Hedgehog pathway transcriptional activity of the cell line.
② Experimental materials
ShhFlashII/NIH3T3 cell line: life Technologies
96-well plate: corning Costar #3595
DMEM medium: life Technologies
Figure BDA0001944617740000251
Luciferase Substrate：Promega
Stop&
Figure BDA0001944617740000252
Substrate：Promega
An enzyme-labeling instrument: themo Electro Co.
CO2 incubator: ESCO Micro pe.
A cell counter: themo Electro Co.
(iii) Experimental methods
(1) Cell counts were performed on commercially available ShhFlashII/NIH3T3 single cell suspensions containing pancreatin.
(2)96 well plates were inoculated with 100. mu.l of 2X10 per well4A number of cells.
(3) Cells were incubated at 37 ℃ with 95% humidity, 5% CO2Cultured in an incubator for 3 days.
(4) And (3) preparing a DMEM culture solution containing the substance to be detected or the shh ligand.
(5) The cell culture fluid is aspirated, and the culture fluid of the object to be tested is added for culturing for 48 h.
(6) The culture medium was removed and 50. mu.l/well was added
Figure BDA0001944617740000253
Luciferase Substrate, cultured in the dark for 10 min.
(7) The Gli-luciferase reading was read with a microplate reader.
(8) Add 50. mu.l of Stop&
Figure BDA0001944617740000254
Substrate, cultured in the dark for 10 min.
(9) Renilla Luciferase readings were taken with a microplate reader.
(10) The Firefoy readings were normalized to those of internal standard Renilla, and IC was calculated50。
Experimental results
The partial compounds of the present invention were screened for inhibitory activity of the Hedeghog pathway, and the results are shown in Table 1.
TABLE 1 IC of part of the test compounds on Gli-luciferase50Value of
Figure BDA0001944617740000255
The results in Table 1 show that the compounds of the present invention have better inhibitory activity on the Hedeghog pathway, with compound I-9 having the best activity and IC on Gli-luc reporter50The value was 34.07 nM.
Doay cell line viability assay
Firstly, the tested medicine
Compounds I-1, I-9, I-13, LY2940680
② Experimental materials
Fetal Bovine Serum (FBS) (GIBCO, Invitrogen Corporation, NY, USA), DMEM medium (GIBCO, Invitrogen Corporation, NY, USA), penicillin (Sigma, st.louis, MO, USA), streptomycin (Sigma, st.louis, MO, USA), MTT reagent, piceatian institute of biotechnology.
A FORMA700 model ultra-low temperature refrigerator, Thermo corporation; YC-300L type medicine storage cabinet, Mitsubishi low temperature science and technology, Inc., of China; Direct-Q with pump type ultrapure water instrument, Millopore corporation; SW-CJ-2FD type superclean bench: suzhou clarification plant, Inc.; forma 3111 type water jacket CO2An incubator: thermo Electron company; berthold LB941 microplate type multifunctional microplate reader, Berthold Co.
Doay cell lines were purchased from the American Tissue Culture Collection (ATCC, Rockville, Md., USA).
(iii) Experimental methods
Test grouping and drug concentration selection
Selection of the concentration of the compound of class I (Compounds I-1, I-9, I-13): 0. mu.M, 0.01. mu.M, 0.10. mu.M, 1. mu.M, 5. mu.M, 10. mu.M.
All compounds were solubilized with DMSO at 0. mu.M in an equal volume of DMSO as a blank, at a final DMSO concentration of no more than 0.10% (v/v).
Cell culture conditions
The cells are subcultured for 10-15 generations under the condition of DMEM medium containing penicillin (final concentration of 100U/ml), streptomycin (final concentration of 100. mu.g/ml) and 10% FBS, when the cells are fused to 90%, the old medium is discarded, the cells are washed with 2ml of PBS for 2 times, 2ml of 0.25% (w/v) Trypsin-0.53mM EDTA mixed digestive juice is added after the PBS is discarded, the mixture is observed under a microscope for about 30s, when the cells are rounded, 2ml of complete medium is rapidly added to stop digestion, the cells are lightly blown and beaten, and the cells are collected. Centrifuging at 800rpm and 4 deg.C for 5min, discarding supernatant, suspending cells with complete culture medium, culturing in bottles, and changing the culture medium every other day.
MTT assay
Cells in logarithmic growth phase were grown at 1X 105cells/well were seeded in 96-well plates at 37 ℃ with 5% CO2The cells were cultured under the condition until the cells were 90% confluent, and then incubated for 2h with serum-free DMEM medium to synchronize the cells. Subsequently, the supernatant was discarded, and HG-class I compounds at concentrations of (0. mu.M, 0.01. mu.M, 0.10. mu.M, 1. mu.M, 5. mu.M, 10. mu.M) were added to the respective solutions and incubated for 72 hours in complete DMEM. 4h before the end of incubation, 20. mu.l MTT solution (5mg/ml) was added to each well. IncubationAfter the completion of the reaction, the supernatant of each well was discarded, 150. mu.l DMSO was added to each well, the mixture was shaken on a cell shaker for 10min, and OD was measured with a microplate reader after the crystals were sufficiently dissolved570。
The inhibition rate (control OD value-experimental OD value)/control OD value × 100%.
Experimental results
Human medulloblastoma cells (Daoy) were tested for their antiproliferative activity in vitro on a portion of the compounds of the invention, and the results are shown in table 2.
TABLE 2 antiproliferative effect of test compounds on human medulloblastoma cells Daoy
Figure BDA0001944617740000271
The results show that the tested compounds have better inhibition effect on the proliferation of human medulloblastoma cells Daoy, wherein the antiproliferative effect of the compounds I-1, I-9 and I-13 is better than that of the positive control LY 2940680.

Claims (4)

1. The benzoyl piperidine derivative is characterized by being a compound I-9 and a compound I-13 shown as follows:
Figure DEST_PATH_IMAGE001
。
2. a pharmaceutical composition comprising the benzoylpiperidine derivative according to claim 1 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable excipient.
3. Use of the benzoylpiperidine derivative of claim 1, or a pharmaceutically acceptable salt thereof, for the manufacture of a medicament for the treatment of an SMO protein inhibitor.
4. Use of the benzoylpiperidine derivative or its pharmaceutically acceptable salt of claim 1 for the preparation of an anti-tumor agent, wherein the tumor is medulloblastoma.
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