CN118084871A - Compound for targeted degradation of c-Met protein and preparation method and application thereof - Google Patents

Compound for targeted degradation of c-Met protein and preparation method and application thereof Download PDF

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CN118084871A
CN118084871A CN202410525807.0A CN202410525807A CN118084871A CN 118084871 A CN118084871 A CN 118084871A CN 202410525807 A CN202410525807 A CN 202410525807A CN 118084871 A CN118084871 A CN 118084871A
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杨鹏
闵文剑
陈春玲
陈玮娇
王晓
袁凯
王丽萍
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China Pharmaceutical University
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Abstract

The invention discloses a compound for targeted degradation of c-Met protein, a preparation method and application thereof, wherein the structure of the compound is shown as a general formula (I) or pharmaceutically acceptable salt thereof, the compound is formed by connecting a c-Met protein inhibitor and a hydrophobic tag compound through a connecting chain. In addition, the mutant has better inhibition activity on the drug-resistant cell line mutation of the type I c-Met inhibitor D1228N and Y1230H. The invention provides an option for tumor treatment.

Description

Compound for targeted degradation of c-Met protein and preparation method and application thereof
Technical Field
The invention relates to a compound and a preparation method and application thereof, in particular to a compound for targeted degradation of c-Met protein and a preparation method and application thereof, belonging to the field of compounds.
Background
C-Met (MESENCHYMAL EPITHELIAL Transition Factor, a cytoplasmic epithelial transforming factor) plays an important role in normal cells as a member of the receptor tyrosine kinase family, involved in various physiological processes such as embryonic development, organ repair, and regeneration, etc. Aberrant activation or overexpression of c-Met can further lead to initiation of a range of signaling pathways, including PI3K/Akt, MAPK, STAT and Wnt, etc., which promote tumor cell growth, survival proliferation, migration, invasion and angiogenesis. The abnormality of the c-Met signal pathway is reported in various types of tumors such as liver cancer, non-small cell lung cancer, gastric cancer, colon cancer and the like.
Although small molecule inhibitors of c-Met are at the clinical corner of the world, one problem that is not negligible is the problem of resistance to small molecule inhibitors. For acquired drug resistance, the drug resistance source targets can be classified into targeted drug resistance and off-targeted drug resistance. Wherein the mechanism of targeted drug resistance is mainly secondary mutation. Y1230 and D1228 are the most common mutation sites for type I inhibitor-induced acquired resistance.
C-Met has become an important target in anti-tumor therapy. Thus, there is an urgent need for the development of drugs against c-Met targeted degradation for the treatment of tumors.
Disclosure of Invention
The invention aims to: the invention aims to provide a compound for targeted degradation of c-Met protein or pharmaceutically acceptable salt thereof, which can obviously inhibit proliferation of a c-Met abnormal activation cell line, has degradation activity against a c-Met target and has strong antiproliferative activity against the c-Met abnormal activation tumor cell line; and shows better antiproliferative activity against mutant cell lines Ba/F3-TPR-MET-D1228N and Ba/F3-TPR-MET-Y1230H (acquired resistance to c-MetI type inhibitors) of MET; another object of the present invention is to provide a method for preparing a compound for targeted degradation of c-Met protein; it is another object of the invention to provide an application of the c-Met protein in targeted degradation.
The technical scheme is as follows: the invention provides a targeted degradation c-Met compound or pharmaceutically acceptable salt thereof, which has a chemical structure shown in a general formula (I):
The targeted degradation c-Met compound is formed by connecting a c-Met protein inhibitor and a hydrophobic tag compound (HyT) through a connecting chain;
wherein the HyT groups are selected from: ，
Wherein linker is a linking chain which is a straight or branched alkylene chain of 1 to 15 atoms total length, the straight or branched alkylene chain is composed of-CH 2 -; -O-, -CO-, -one or more of CONH-, -N-, alkynylene or cycloalkylene.
Further, the HyT groups are selected from:，
In the Linker, the number of the links is as follows, the straight or branched alkylene chain is composed of-CH 2 -, -O-, -one or more of CO-, -CONH-, -N-.
Further, the HyT groups are selected from:，
in the Linker, the number of the links is as follows, the straight or branched alkylene chain is composed of-CH 2 -; -one or more of CO-, -CONH-.
Further, when HyT groups are any one of the following groups:，
The connecting chain is-COCH 2 -; when HyT groups are the following: ，
the connecting chain is-CH 2CONH-CH2 -; when HyT groups are the following: ，
The connecting chain is a straight or branched alkylene chain of 1 to 15 atoms total length, the straight or branched alkylene chain is composed of-CH 2 -, -O-, -one or more of CO-, -CONH-, -N-.
Further, the compound is selected from the group consisting of formula (H1) to formula (H13):
。
Further, the above compound or a pharmaceutically acceptable salt thereof is an acid addition salt of the compound of the general formula (I), wherein the acid used for salification includes inorganic acids including hydrochloric acid, sulfuric acid, phosphoric acid, and organic acids including acetic acid, trichloroacetic acid, trifluoroacetic acid, propionic acid, butyric acid, maleic acid, p-toluenesulfonic acid, malic acid, methanesulfonic acid, malonic acid, cinnamic acid, citric acid, fumaric acid, camphoric acid, digluconic acid, aspartic acid, and tartaric acid.
In another aspect, the invention provides a method for preparing a compound or pharmaceutically acceptable salt thereof that targets degradation of c-Met protein, comprising the steps of:
Dissolving the compound A and the compound B in N, N-dimethylformamide, adding triethylamine, 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride and 1-hydroxybenzotriazole, stirring the reaction at room temperature, and purifying after TLC monitoring reaction to obtain the compound for targeted degradation of c-Met protein.
In another aspect, the invention provides a method for preparing a compound or pharmaceutically acceptable salt thereof that targets degradation of c-Met protein, comprising the steps of:
Dissolving a compound C and a compound D in N, N-dimethylformamide, adding triethylamine, 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride and 1-hydroxybenzotriazole, stirring the reaction at room temperature, and purifying after TLC monitoring reaction to obtain the compound for targeted degradation of C-Met protein.
In another aspect, the invention provides a pharmaceutical composition comprising a compound of formula (I) or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.
Pharmaceutically acceptable carrier refers to excipients or diluents that do not cause significant irritation to the organism and do not interfere with the biological activity and properties of the compound being administered. The excipient comprises a flavoring agent, a cosolvent, an emulsifying agent, a solubilizer, an adhesive, a preservative, an antioxidant, an osmotic pressure regulator, a colorant filler, a disintegrating agent, a lubricant and the like, and the diluent comprises starch, physiological saline, sucrose, lactose, dextrin and the like.
In another aspect, the present invention provides the use of a compound or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for the prevention and/or treatment of cancer or a tumor-associated disease. The cancer comprises liver cancer, lung cancer, gastric cancer, kidney cancer, colorectal cancer and esophageal cancer.
Preferably, the cancer is liver cancer, lung cancer, stomach cancer, kidney cancer, colorectal cancer or esophageal cancer, and the tumor is a malignant tumor caused by abnormal MET gene and expression.
In some of these embodiments, the tumor is a MET gene amplified malignancy.
The compound of the general formula (I) or the pharmaceutically acceptable salt thereof has degradation activity on c-Met protein and has therapeutic effect on related malignant tumors.
The principle of the invention: when the hydrophobic tag molecule is introduced to the c-Met protein, the compound provided by the invention can cause the protein conformation to be damaged and unstable due to the existence of the hydrophobic group, and is further recognized as misfolded protein by a mechanism, and ubiquitin groups are added on the surface of the protein through an ubiquitin-proteasome system. The ubiquitinated c-Met protein is recognized by the proteasome and eventually degraded. By promoting degradation of c-Met, the function of c-Met can be effectively inhibited. The degradation agent can degrade the c-Met protein level more rapidly, has a strong tumor killing effect, and is expected to overcome the drug resistance problem of the traditional small molecule inhibitor.
Advantageous effects
Compared with the prior art, the invention has the following remarkable advantages: the compound for targeting degradation of c-Met protein prepared by the invention degrades the c-Met protein level, and can effectively resist proliferation of various tumor cells amplified by MET. In addition, preferred compounds exhibit better antiproliferative activity against the D1228N and Y1230H mutations common to type I inhibitors, hopefully overcoming the problem of resistance caused by the D1228N and Y1230H mutations common to type I inhibitors.
Detailed Description
The application is further described below. The following examples are only for more clearly illustrating the technical aspects of the present application, and are not intended to limit the scope of the present application. The experimental methods in the following examples are conventional methods unless otherwise specified. The test materials used in the examples described below, unless otherwise specified, were purchased from conventional biochemical reagent stores. The present application will be described in detail with reference to specific examples.
Synthesis of the intermediate reactant
3- (6-Oxo-1, 6-dihydropyridazin-3-yl) benzonitrile (N-1). 3-Acetylbenzonitrile (25.0 g,172 mmol) was dissolved in 250 mL g of glacial acetic acid, followed by dropwise addition of 23 mL of 50% aqueous glyoxylate and heating at 95 o C overnight. After TLC detection of disappearance of starting material, it was cooled, 500 mL water was added, 19 mL of 80% hydrazine hydrate solution was slowly added dropwise, and the reaction was continued with heating and stirring at 95 o C for 4 hours. Cooling to 60 o C, suction filtering to retain solid, washing with water, washing with acetone, boiling the solid in acetone, and filtering while hot to retain solid. The resulting solid was added to 250 mL glacial acetic acid and stirred with 95 o C heat for 4 hours. The solid obtained after the completion of the reaction was filtered to obtain the objective product (27.2 g, yield 80 %）.1H NMR (400 MHz, DMSO-d6)δ13.37 (s, 1H), 8.32 (t,J= 1.7 Hz, 1H), 8.21 (dt,J= 8.0, 1.1 Hz, 1H), 8.14 (d,J= 10.0 Hz, 1H), 7.92 (dt,J= 7.7, 1.4 Hz, 1H), 7.70 (t,J= 7.9 Hz, 1H), 7.04 (dd,J= 10.0, 1.5 Hz, 1H). ESI-MSm/z: 198.2 [M+H]+.
4- (((5-Chloropyrimidin-2-yl) oxy) methyl) piperidine-1-carboxylic acid tert-butyl ester (N-2). 2-hydroxy-5-chloropyrimidine (13.0 g,100 mmol) and triphenylphosphine (31.4 g,120 mmol) were dissolved in 300 mL anhydrous tetrahydrofuran, stirred under argon for 30min, and diisopropyl azodicarboxylate (DIAD, 23.6 mL,120 mmol) was slowly added dropwise under ice-bath conditions and the reaction was allowed to react overnight at 0 o C. TLC detection of disappearance of starting material, purification of crude silica gel column to give N-2 (24.6 g, yield) 75 %）.1H NMR (400 MHz, Chloroform-d)δ8.28 (s, 2H), 4.19 (s, 2H), 3.90 (m, 2H), 2.75 (m, 2H), 2.07 – 1.92 (m, 1H), 1.85 – 1.77 (m, 2H), 1.47 (s, 9H), 1.36 – 1.29 (m, 2H). ESI-MSm/z: 328.8 [M+H]+.
4- (((5- (3- (Chloromethyl) phenyl) pyrimidin-2-yl) oxy) methyl) piperidine-1-carboxylic acid tert-butyl ester (N-3). N-2 (16.4 g,50 mmol), 3-hydroxymethylphenylboronic acid (8.3 g,55 mmol), potassium carbonate (13.8 g,100 mmol) and Pd (PPh 3)4 (2.8 g,2.5 mmol) were sequentially added to a two-necked flask containing 300mL dioxane/water (4:1), and the reaction was performed overnight at 90℃under argon, TLC detected the disappearance of the starting material, and after purification of the crude silica gel column, the crude silica gel was slowly added to a solution of 200 mL thionyl chloride at 70℃for three hours, the solvent was dried by spin, and the silica gel column was purified to give N-3 (16.0 g, two-step yield 76 %）.1H NMR (400 MHz, Chloroform-d)δ8.46 (s, 2H), 8.38 (m, 1H), 8.31 (m, 1H), 7.48 (m, 2H), 4.68 (s, 2H), 4.20 (s, 2H), 3.95 (d,J= 6.3 Hz, 2H), 2.77 (m, 2H), 2.08 – 1.97 (m, 1H), 1.89 – 1.79 (m, 2H), 1.48 (s, 9H), 1.38 – 1.29 (m, 2H). ESI-MSm/z: 418.5 [M+H]+.
4- (((2- (3- ((3- (3-Cyanophenyl) -6-oxopyridazin-1 (6H) -yl) methyl) phenyl) pyrimidin-5-yl) oxy) methyl) piperidine-1-carboxylic acid tert-butyl ester (N-4). N-3 (16.0 g,38 mmol), N-1 (5.9 g,30 mmol), potassium carbonate (13.8 g,100 mmol) were reacted in 150 mL DMF solvent at 80℃for 6 hours. TLC detection of disappearance of starting material, purification of crude silica gel column to give N-4 (14.3 g, yield) 82 %）.1H NMR (400 MHz, Chloroform-d)δ8.59 (t,J= 1.8 Hz, 1H), 8.48 (s, 2H), 8.30 (dt,J= 8.0, 1.5 Hz, 1H), 8.18 (t,J= 1.8 Hz, 1H), 7.98 (dt,J= 8.0, 1.4 Hz, 1H), 7.70 (dt,J= 7.8, 1.4 Hz, 1H), 7.63 (d,J= 9.7 Hz, 1H), 7.57 (t,J= 7.8 Hz, 2H), 7.45 (t,J= 7.7 Hz, 1H), 7.07 (d,J= 9.7 Hz, 1H), 5.50 (s, 2H), 4.16 (m, 2H), 3.96 (d,J= 6.3 Hz, 2H), 2.75 (m, 2H), 2.06 – 1.95 (m, 1H), 1.88 – 1.80 (m, 2H), 1.47 (s, 9H), 1.37 – 1.27 (m, 2H). ESI-MSm/z: 579.6 [M+H]+.
3- (6-Oxo-1- (3- (5- (piperidin-4-ylmethoxy) pyrimidin-2-yl) benzyl) -1, 6-dihydropyridazin-3-yl) benzonitrile (N-5). N-4 (14.4 g,25 mmol) was added to a 100 mL hydrogen chloride-ethyl acetate solution and reacted at room temperature for 3 hours. Spin-drying the solvent gave N-5 as a white solid (12.0 g, 100% yield). ESI-MSm/z 479.6 [ M+H ] +.
Acetic acid 2- (4- ((2- (3- ((3- (3-cyanophenyl) -6-oxopyridazin-1 (6H) -yl) methyl) phenyl) pyrimidin-5-yl) oxymethyl) piperidin-1-yl) tert-butyl ester (N-6). N-5 (7.2 g,15 mmol), t-butyl bromoacetate (4.4 g,22.5 mmol) and potassium carbonate (4.2 g,30 mmol) were added to the 80 mLDMF solution and reacted at 80℃for 4 hours. Spin-drying the solvent, purifying the crude product with silica gel column to obtain white solid N-6 (8.4 g, yield) 95 %）.1H NMR (400 MHz, Chloroform-d)δ8.57 (t,J= 1.8 Hz, 1H), 8.47 (s, 2H), 8.29 (dt,J= 7.8, 1.5 Hz, 1H), 8.17 (t,J= 1.8 Hz, 1H), 7.99 (dt,J= 8.0, 1.4 Hz, 1H), 7.70 (dt,J= 7.7, 1.4 Hz, 1H), 7.64 (d,J= 9.7 Hz, 1H), 7.60 – 7.53 (m, 2H), 7.45 (t,J= 7.7 Hz, 1H), 7.07 (d,J= 9.7 Hz, 1H), 5.50 (s, 2H), 3.95 (d,J= 5.9 Hz, 2H), 3.76 – 3.65 (m, 1H), 3.14 (m, 3H), 3.03 (m, 2H), 2.24 (m, 2H), 1.85 (m, 3H), 1.48 (s, 9H). ESI-MSm/z: 593.4 [M+H]+.
2- (4- (((2- (3- ((3- (3-Cyanophenyl) -6-oxopyridazin-1 (6H) -yl) methyl) phenyl) pyrimidin-5-yl) oxy) methyl) piperidin-1-yl) acetic acid (N-7). N-6 (8.4 g,14 mmol) was added to a 50 mL dichloromethane solution, 50 mL trifluoroacetic acid was added dropwise, and the reaction was carried out at room temperature for 3 hours. Spin-drying the solvent and recrystallizing the crude product to give N-7 as a white solid (7.1 g, 95% yield). ESI-MSm/z 537.5 [ M+H ] +.
2- ((3 R,5r,7 r) -adamantan-1-yl) -N- (2-aminoethyl) acetamide (N-8 a). N-5 and N-tert-butyloxycarbonyl-1, 2-ethylenediamine are used as raw materials, and are prepared according to the same route as H1, and then deprotected by ethyl acetate-hydrogen chloride solution. ESI-MSm/z 237.4 [ M+H ] +.
2- ((3 R,5r,7 r) -adamantan-1-yl) -N- (4-aminobutyl) acetamide (N-8 b). The N-8 a-N-butyloxycarbonyl-1, 4-butanediamine is used as a raw material and prepared according to the same route. ESI-MSm/z 265.3 [ M+H ] +.
2- ((3 R,5r,7 r) -adamantan-1-yl) -N- (6-aminohexyl) acetamide (N-8 c). N-5 and N-tert-butyloxycarbonyl-1, 6-hexamethylenediamine are used as raw materials and prepared according to the same route as N-8 a. ESI-MSm/z 293.6 [ M+H ] +.
2- ((3 R,5r,7 r) -adamantan-1-yl) -N- (8-aminooctyl) acetamide (N-8 d). The N-5 and N-tert-butyloxycarbonyl-1, 8-octanediamine are used as raw materials and prepared according to the same route as N-8 a. ESI-MSm/z 321.5 [ M+H ] +.
2- ((3 R,5r,7 r) -adamantan-1-yl) -N- (10-aminodecyl) acetamide (N-8 e). The N-8 a-N-butyloxycarbonyl-1, 10-decanediamine is used as a raw material to prepare the N-5-tert-butyloxycarbonyl-1, 10-decanediamine according to the same route. ESI-MSm/z 349.3 [ M+H ] +.
6- (4- (((2- (3- ((3- (3-Cyanophenyl) -6-oxopyridazin-1 (6H) -yl) methyl) phenyl) pyrimidin-5-yl) oxy) methyl) piperidin-1-yl) hexanoic acid (N-9 a). Taking N-5 and bromohexanoic acid tert-butyl ester as raw materials, adding 3 equivalents of DIPEA to react for 8 hours in DMF at 90 o C, removing tert-butyl ester from the product obtained by silica gel column purification in a DCM/TFA=1:1 system, and spin-drying the solvent to obtain N-9a with the yield of 55 percent. ESI-MSm/z 593.7 [ M+H ] +.
4- (4- (((2- (3- ((3- (3-Cyanophenyl) -6-oxopyridazin-1 (6H) -yl) methyl) phenyl) pyrimidin-5-yl) oxy) methyl) piperidin-1-yl) butyric acid (N-9 b). N-5 and tert-butyl bromobutyrate are used as raw materials, and the preparation is carried out according to the same method as N-9a, and the yield is 70%. ESI-MSm/z 565.2 [ M+H ] +.
3- (1- (3- (5- ((1- (2-Aminoethyl) piperidin-4-yl) methoxy) pyrimidin-2-yl) benzyl) -6-oxo-1, 6-dihydropyridazin-3-yl) benzonitrile (N-10). N-5 and N-tert-butyloxycarbonyl-2-aminoacetaldehyde are taken as raw materials, 1.5 equivalent of sodium triacetoxyborohydride is added into a DCM/MeOH=1:1 system to react for three hours at 0 o C, and the obtained product is purified by a silica gel column and deprotected by an ethyl acetate-hydrogen chloride solution to prepare N-10, and the yield is obtained 60 %.1H NMR (400 MHz, DMSO-d6)δ8.66 (m, 2H), 8.50 (s, 2H), 8.43 – 8.35 (m, 2H), 8.29 – 8.21 (m, 2H), 8.19 (d,J= 9.7 Hz, 1H), 7.94 (dt,J= 7.8, 1.3 Hz, 1H), 7.73 (t,J= 7.9 Hz, 1H), 7.53 – 7.45 (m, 2H), 7.17 (d,J= 9.8 Hz, 1H), 5.45 (s, 2H), 4.09 (d,J= 6.4 Hz, 2H), 3.62 (d,J= 11.1 Hz, 2H), 3.06 (q,J= 11.4 Hz, 2H), 2.14 (d,J= 13.5 Hz, 1H), 1.99 (m, 2H), 1.92 (m, 4H), 1.83 – 1.66 (m, 2H), 1.60 (s, 2H). ESI-MSm/z: 522.4 [M+H]+.
2. Synthesis of compounds H1 to H13:
Example 1
Synthesis of 3- (1- (3- (5- ((1- (3, 3-diphenylpropionyl) piperidin-4-yl) methoxy) pyrimidin-2-yl) benzyl) -6-oxo-1, 6-dihydropyridazin-3-yl) benzonitrile (H1):
N-5 (479 mg,1 mmol), 3-diphenylpropionic acid (226 mg,1 mmol), EDCI (573 mg,3 mmol), HOBT (202 mg,1.5 mmol) and 0.7 mL triethylamine were added to the 5mL DMF solution and reacted at room temperature for 5 hours. Purifying the crude product by silica gel column (426, 426 mg, yield) 62 %）.1H NMR (400 MHz, Chloroform-d) δ 8.59 (d, J = 1.8 Hz, 1H), 8.47 (s, 2H), 8.30 (dt, J = 7.9, 1.6 Hz, 1H), 8.19 (t, J = 1.8 Hz, 1H), 7.97 (dt, J = 8.0, 1.6 Hz, 1H), 7.73 – 7.67 (m, 1H), 7.63 (d, J = 9.7 Hz, 1H), 7.60 – 7.53 (m, 2H), 7.45 (t, J = 7.7 Hz, 1H), 7.31 – 7.26 (m, 8H), 7.19 (tdd, J = 6.5, 3.9, 2.0 Hz, 2H), 7.07 (d, J = 9.7 Hz, 1H), 5.50 (s, 2H), 4.69 (t, J = 7.5 Hz, 2H), 3.94 – 3.79 (m, 3H), 3.18 – 2.97 (m, 2H), 2.94 – 2.85 (m, 1H), 2.55 – 2.44 (m, 1H), 2.05 – 1.93 (m, 1H), 1.77 (t, J = 14.2 Hz, 2H), 1.13 (qd, J = 12.3, 4.2 Hz, 1H), 0.90 – 0.84 (m, 1H). 13C NMR (101 MHz, DMSO-d6) δ 168.55, 158.67, 155.73, 151.46, 144.79, 144.74, 144.05, 141.86, 137.23, 136.85, 135.32, 132.76, 130.71, 130.27, 130.14, 130.03, 129.58, 129.34, 128.88, 128.18, 128.15, 127.67, 127.60, 126.83, 126.29, 125.89, 118.44, 112.05, 72.37, 54.53, 46.92, 44.59, 40.74, 37.46, 35.12, 28.65, 27.88. HRMS (ESI) m/z calcd. for C43H38N6O3 [M+H]+ 687.3078, found: 687.3076. Purity: 97.2 %.
Example 2
Synthesis of 3- (1- (3- (5- ((1- (2- (9H-fluoro-9-yl) acetyl) piperidin-4-yl) methoxy) pyrimidin-2-yl) benzyl) -6-oxo-1, 6-dihydropyridazin-3-yl) benzonitrile (H2):
h2 (478 mg, yield) was prepared by the same route as H1 using N-5 and 9-fluoreneacetic acid as raw materials 70 %）.1H NMR (400 MHz, Chloroform-d)δ8.59 (t,J= 1.8 Hz, 1H), 8.48 (s, 2H), 8.30 (dt,J= 7.9, 1.4 Hz, 1H), 8.19 (d,J= 1.8 Hz, 1H), 7.97 (ddd,J= 8.0, 1.9, 1.1 Hz, 1H), 7.77 (d,J= 7.5 Hz, 2H), 7.70 (dt,J= 7.8, 1.3 Hz, 1H), 7.63 (d,J= 9.7 Hz, 1H), 7.57 (tt,J= 7.5, 1.9 Hz, 4H), 7.45 (t,J= 7.7 Hz, 1H), 7.42 – 7.34 (m, 2H), 7.30 (tdd,J= 7.4, 4.1, 1.2 Hz, 2H), 7.07 (d,J= 9.7 Hz, 1H), 5.50 (s, 2H), 4.93 (d,J= 13.1 Hz, 1H), 4.66 (t,J= 7.2 Hz, 1H), 3.95 (qd,J= 9.0, 6.2 Hz, 2H), 3.79 (d,J= 13.6 Hz, 1H), 3.06 – 2.93 (m, 1H), 2.82 – 2.65 (m, 3H), 2.17 – 2.04 (m, 1H), 1.97 (d,J= 13.5 Hz, 1H), 1.82 (d,J= 13.2 Hz, 1H), 1.42 (qd,J= 12.7, 4.0 Hz, 1H), 1.20 (td,J= 12.5, 4.1 Hz, 1H). 13C NMR (101 MHz, DMSO-d6)δ168.96, 158.58, 155.64, 151.37, 146.97, 146.93, 143.94, 141.76, 140.02, 137.15, 136.75, 135.21, 132.66, 130.58, 130.16, 130.03, 129.93, 129.48, 129.23, 128.78, 126.98, 126.88, 126.76, 126.22, 124.72, 124.68, 119.71, 118.35, 111.96, 72.27, 54.45, 44.46, 43.21, 40.97, 36.69, 35.08, 28.58, 27.93. HRMS (ESI)m/zcalcd. for C43H36N6O3[M+H]+685.2922, found: 685.2922. Purity: 96.4 %.
Example 3
Synthesis of 3- (1- (3- (5- ((1- (2- (((1 r,2s,5 r) -2-isopropyl-5-methylcyclohexyl) oxy) acetyl) piperidin-4-yl) methoxy) pyrimidin-2-yl) benzyl) -6-oxo-1, 6-dihydropyridazin-3-yl) benzonitrile (H3):
preparation of H3 from N-5 and (-) -Montoxyacetic acid as starting materials according to the same route as H1 (398 mg, yield) 59 %）.1H NMR (400 MHz, Chloroform-d)δ8.59 (t,J= 1.8 Hz, 1H), 8.49 (s, 2H), 8.30 (dt,J= 7.9, 1.5 Hz, 1H), 8.19 (d,J= 1.8 Hz, 1H), 7.98 (dt,J= 8.0, 1.4 Hz, 1H), 7.70 (dt,J= 7.7, 1.3 Hz, 1H), 7.63 (d,J= 9.7 Hz, 1H), 7.60 – 7.52 (m, 2H), 7.45 (t,J= 7.7 Hz, 1H), 7.07 (d,J= 9.7 Hz, 1H), 5.50 (s, 2H), 4.64 (d,J= 14.8 Hz, 1H), 4.24 (dd,J= 12.6, 6.5 Hz, 1H), 4.18 – 4.07 (m, 2H), 3.96 (q,J= 8.9, 7.5 Hz, 2H), 3.19 (tt,J= 10.5, 3.9 Hz, 1H), 3.08 (t,J= 12.9 Hz, 1H), 2.64 (dt,J= 14.0, 10.4 Hz, 1H), 2.32 – 2.07 (m, 3H), 1.92 (t,J= 14.0 Hz, 2H), 1.83 – 1.69 (m, 2H), 1.64 (tq,J= 9.6, 3.1 Hz, 2H), 1.47 – 1.32 (m, 3H), 1.27 – 1.22 (m, 1H), 1.00 – 0.82 (m, 9H), 0.77 (dd,J= 13.3, 7.0 Hz, 3H). 13C NMR (101 MHz, DMSO-d6)δ167.80, 159.21, 156.30, 152.02, 144.56, 142.37, 137.79, 137.37, 135.86, 133.28, 131.21, 130.79, 130.67, 130.58, 130.13, 129.86, 129.39, 127.43, 126.86, 118.97, 112.62, 78.79, 72.93, 67.85, 55.08, 48.27, 44.92, 41.20, 35.73, 34.46, 31.31, 29.20, 28.35, 25.31, 23.12, 22.67, 21.33, 16.66, 16.31. HRMS (ESI)m/zcalcd. for C40H46N6O4[M+H]+675.3653, found: 675.3654. Purity: 97.0 %.
Example 4
Synthesis of N- (bicyclo [2.2.1] hept-5-en-2-ylmethyl) -2- (4- ((2- (3- ((3- (3-cyanophenyl) -6-oxopyridazin-1 (6H) -yl) methyl) phenyl) pyrimidin-5-yl) oxymethyl) piperidin-1-yl) acetamide (H4):
preparation of H4 (513 mg, yield) from N-7 and 5-norbornene-2-methylamine as raw materials according to the same route as H1 80 %）.1H NMR (400 MHz, Chloroform-d)δ8.60 (t,J= 1.8 Hz, 1H), 8.50 (s, 2H), 8.30 (dt,J= 7.7, 1.4 Hz, 1H), 8.19 (t,J= 1.7 Hz, 1H), 7.97 (dt,J= 8.3, 1.5 Hz, 1H), 7.70 (dt,J= 7.7, 1.4 Hz, 1H), 7.63 (d,J= 9.9 Hz, 1H), 7.60 – 7.52 (m, 2H), 7.45 (t,J= 7.7 Hz, 1H), 7.22 (s, 1H), 7.07 (d,J= 9.8 Hz, 1H), 6.19 (dd,J= 5.8, 3.0 Hz, 1H), 5.98 (dd,J= 5.7, 2.8 Hz, 1H), 5.51 (s, 2H), 3.98 (d,J= 5.7 Hz, 2H), 3.15 – 2.96 (m, 4H), 2.92 (d,J= 11.5 Hz, 2H), 2.81 (s, 2H), 2.34 – 2.16 (m, 3H), 1.96 – 1.81 (m, 4H), 1.54 – 1.41 (m, 3H), 1.27 (d,J= 8.3 Hz, 1H), 0.60 (ddd,J= 11.6, 4.4, 2.5 Hz, 1H). 13C NMR (101 MHz, DMSO-d6)δ169.46, 159.23, 156.25, 152.09, 144.56, 142.41, 137.81, 137.40, 137.36, 136.99, 136.79, 135.88, 131.26, 130.82, 130.69, 130.59, 130.12, 129.89, 129.42, 127.40, 126.85, 118.99, 112.62, 73.36, 62.24, 55.09, 53.46, 49.41, 44.20, 42.70, 42.31, 38.96, 35.33, 30.07, 28.79. HRMS (ESI)m/zcalcd. for C38H39N7O3[M+H]+642.3182, found: 642.3186. Purity: 96.6 %.
Example 5
Synthesis of 3- (1- (3- (5- ((1- (2- ((3 r,5r,7 r) -adamantan-1-yl) acetyl) piperidin-4-yl) methoxy) pyrimidin-2-yl) benzyl) -6-oxo-1, 6-dihydropyridazin-3-yl) benzonitrile (H5):
Preparation of H5 (513 mg, yield) from N-5 and adamantaneacetic acid as raw materials according to the same route as H1 60 %）.1H NMR (400 MHz, Chloroform-d)δ8.58 (t,J= 1.8 Hz, 1H), 8.49 (s, 2H), 8.29 (dt,J= 7.8, 1.5 Hz, 1H), 8.19 (t,J= 1.7 Hz, 1H), 7.98 (dt,J= 8.0, 1.5 Hz, 1H), 7.70 (dt,J= 7.8, 1.4 Hz, 1H), 7.64 (d,J= 9.7 Hz, 1H), 7.61 – 7.51 (m, 2H), 7.45 (t,J= 7.7 Hz, 1H), 7.08 (d,J= 9.7 Hz, 1H), 5.50 (s, 2H), 4.85 – 4.72 (m, 1H), 4.06 (d,J= 13.7 Hz, 1H), 3.97 (qd,J= 9.0, 6.2 Hz, 2H), 3.72 (pd,J= 6.7, 4.1 Hz, 1H), 3.29 – 3.11 (m, 1H), 3.14 – 2.99 (m, 1H), 2.59 (td,J= 12.9, 2.8 Hz, 1H), 2.22 (d,J= 13.4 Hz, 1H), 2.16 – 2.07 (m, 1H), 2.00 – 1.86 (m, 5H), 1.72 (d,J= 2.8 Hz, 2H), 1.52 – 1.41 (m, 6H), 1.39 – 1.20 (m, 4H), 0.91 – 0.84 (m, 1H). 13C NMR (101 MHz, DMSO-d6)δ168.20, 158.67, 155.72, 151.50, 144.07, 141.88, 137.23, 136.86, 135.33, 132.77, 130.73, 130.29, 130.16, 130.04, 129.57, 129.35, 128.88, 126.82, 126.29, 118.43, 112.06, 72.39, 54.52, 53.46, 45.81, 45.00, 42.01, 36.30, 35.21, 32.90, 28.90, 28.09, 27.97. HRMS (ESI)m/zcalcd. for C40H42N6O3[M+H]+655.3391, found: 655.3389. Purity: 95.3 %.
Example 6
Synthesis of 2- ((3 r,5r,7 r) -adamantan-1-yl) -N- (2- (2- (4- (((2- (3- ((3- (3-cyanophenyl) -6-oxopyridazin-1 (6H) -yl) methyl) phenyl) pyrimidin-5-yl) oxy) methyl) piperidin-1-yl) acetamido) ethyl) acetamide (H6):
N-7 (537 mg,1 mmol), N-8a (237 mg,1 mmol), EDCI (573 mg,3 mmol), HOBT (202 mg,1.5 mmol) and 0.7 mL triethylamine were added to a solution of 5mL DMF and reacted at room temperature for 5 hours. Purifying the crude product by silica gel column (324 mg, yield) 43 %）.1H NMR (400 MHz, Chloroform-d)δ8.59 (t,J= 1.8 Hz, 1H), 8.50 (s, 2H), 8.30 (dt,J= 7.9, 1.5 Hz, 1H), 8.19 (t,J= 1.8 Hz, 1H), 7.98 (dt,J= 8.1, 1.5 Hz, 1H), 7.70 (dt,J= 7.8, 1.4 Hz, 1H), 7.66 – 7.53 (m, 4H), 7.45 (t,J= 7.7 Hz, 1H), 7.07 (d,J= 9.7 Hz, 1H), 6.18 (s, 1H), 5.51 (s, 2H), 3.97 (d,J= 5.8 Hz, 2H), 3.51 – 3.35 (m, 4H), 3.02 (s, 2H), 2.89 (d,J= 11.1 Hz, 2H), 2.24 (t,J= 11.5 Hz, 2H), 1.96 (t,J= 3.2 Hz, 3H), 1.92 (s, 2H), 1.91 – 1.81 (m, 3H), 1.81 – 1.65 (m, 6H), 1.62 (m, 6H), 1.49 (m, 2H). 13C NMR (101 MHz, DMSO-d6)δ170.74, 170.09, 159.22, 156.31, 152.12, 144.56, 142.41, 137.83, 137.38, 135.89, 133.28, 131.22, 130.81, 130.68, 130.58, 130.13, 129.87, 129.39, 127.41, 126.86, 118.97, 112.63, 73.44, 62.16, 55.08, 53.55, 50.53, 42.53, 39.01, 38.73, 36.86, 35.25, 32.55, 28.70, 28.50. HRMS (ESI)m/zcalcd. for C44H50N8O4[M+H]+755.4028, found: 755.4029. Purity: 96.2 %.
Example 7
Synthesis of 2- ((3 r,5r,7 r) -adamantan-1-yl) -N- (4- (2- (4- (((2- (3- ((3- (3-cyanophenyl) -6-oxopyridazin-1 (6H) -yl) methyl) phenyl) pyrimidin-5-yl) oxy) methyl) piperidin-1-yl) acetamido) butyl) acetamide (H7):
The N-7 and N-8b are used as raw materials to synthesize H7 by the same method as H6, and the yield is high 42 %.1H NMR (400 MHz, Chloroform-d)δ8.60 (d,J= 1.9 Hz, 1H), 8.50 (s, 2H), 8.30 (dt,J= 7.8, 1.5 Hz, 1H), 8.20 (t,J= 1.7 Hz, 1H), 7.97 (dt,J= 8.0, 1.5 Hz, 1H), 7.70 (dt,J= 7.8, 1.4 Hz, 1H), 7.64 (d,J= 9.7 Hz, 1H), 7.62 – 7.49 (m, 3H), 7.46 (t,J= 7.7 Hz, 1H), 7.07 (d,J= 9.7 Hz, 1H), 5.64 (s, 1H), 5.51 (s, 2H), 3.99 (d,J= 5.9 Hz, 2H), 3.30 (m, 4H), 3.08 (m, 4H), 2.36 (s, 2H), 1.93 (m, 8H), 1.74 – 1.61 (m, 10H), 1.60 – 1.50 (m, 8H). ESI-MSm/z: 783.6 [M+H]+. 13C NMR (101 MHz, DMSO-d6)δ170.15, 159.24, 156.34, 152.09, 144.61, 142.45, 137.82, 137.42, 135.92, 133.32, 131.28, 130.85, 130.72, 130.60, 130.15, 129.91, 129.43, 127.39, 126.86, 118.98, 112.64, 73.20, 55.09, 53.30, 50.54, 42.59, 38.51, 38.44, 36.91, 32.58, 29.03, 28.49, 27.24, 27.19. HRMS (ESI)m/zcalcd. for C46H54N8O4[M+H]+783.4341, found: 783.4341. Purity: 95.6 %.
Example 8
Synthesis of 2- ((3 r,5r,7 r) -adamantan-1-yl) -N- (6- (2- (4- (((2- (3- ((3- (3-cyanophenyl) -6-oxopyridazin-1 (6H) -yl) methyl) phenyl) pyrimidin-5-yl) oxy) methyl) piperidin-1-yl) acetamido) hexyl) acetamide (H8):
The H8 is synthesized by the same method of H6 by taking N-7 and N-8c as raw materials, and the yield is increased 31 %.1H NMR (400 MHz, Chloroform-d)δ8.59 (d,J= 1.8 Hz, 1H), 8.50 (s, 2H), 8.30 (dt,J= 7.9, 1.5 Hz, 1H), 8.19 (t,J= 1.7 Hz, 1H), 7.98 (dt,J= 8.0, 1.5 Hz, 1H), 7.70 (dt,J= 7.7, 1.4 Hz, 1H), 7.64 (d,J= 9.7 Hz, 1H), 7.57 (t,J= 7.9 Hz, 2H), 7.45 (t,J= 7.7 Hz, 1H), 7.29 (s, 1H), 7.07 (d,J= 9.7 Hz, 1H), 5.50 (s, 2H), 5.49 (s, 1H), 3.98 (d,J= 5.8 Hz, 2H), 3.32 – 3.19 (m, 4H), 3.05 (s, 2H), 2.95 (d,J= 9.7 Hz, 2H), 2.27 (d,J= 11.2 Hz, 2H), 2.01 – 1.86 (m, 10H), 1.74 – 1.61 (m, 8H), 1.58 – 1.41 (m, 8H), 1.38 – 1.32 (m, 4H). 13C NMR (101 MHz, DMSO-d6)δ170.09, 159.24, 156.28, 152.10, 144.59, 142.44, 137.81, 137.42, 135.90, 133.33, 131.29, 130.85, 130.72, 130.60, 130.14, 129.91, 129.44, 127.38, 126.85, 118.99, 112.62, 73.28, 55.09, 53.39, 50.55, 42.58, 38.60, 38.56, 36.91, 32.58, 29.65, 28.49, 26.58, 26.51. HRMS (ESI)m/zcalcd. for C48H58N8O4[M+H]+811.4654, found: 811.4652. Purity: 97.9 %.
Example 9
Synthesis of 2- ((3 r,5r,7 r) -adamantan-1-yl) -N- (8- (2- (4- (((2- (3- ((3- (3-cyanophenyl) -6-oxopyridazin-1 (6H) -yl) methyl) phenyl) pyrimidin-5-yl) oxy) methyl) piperidin-1-yl) acetamido) octyl) acetamide (H9):
The H9 is synthesized by the same method of H6 by taking N-7 and N-8d as raw materials, and the yield is increased 28 %.1H NMR (400 MHz, Chloroform-d)δ8.60 (d,J= 1.8 Hz, 1H), 8.50 (s, 2H), 8.30 (dt,J= 7.8, 1.5 Hz, 1H), 8.20 (t,J= 1.7 Hz, 1H), 7.98 (dt,J= 8.0, 1.5 Hz, 1H), 7.70 (dt,J= 7.7, 1.4 Hz, 1H), 7.64 (d,J= 9.7 Hz, 1H), 7.60 – 7.53 (m, 2H), 7.45 (t,J= 7.7 Hz, 1H), 7.35 (s, 1H), 7.07 (d,J= 9.7 Hz, 1H), 5.50 (s, 2H), 5.38 (m, 1H), 3.99 (d,J= 5.9 Hz, 2H), 3.31 – 3.20 (m, 4H), 3.20 – 2.83 (m, 4H), 2.34 (s, 2H), 2.00 – 1.83 (m, 9H), 1.73 – 1.60 (m, 10H), 1.58 – 1.42 (m, 7H), 1.31 (s, 8H).13C NMR (101 MHz, DMSO-d6)δ170.08, 159.25, 156.33, 152.10, 144.64, 142.46, 137.82, 137.43, 135.93, 133.34, 131.31, 130.87, 130.73, 130.61, 130.15, 129.93, 129.44, 127.39, 126.86, 118.98, 112.64, 55.09, 50.56, 42.60, 38.65, 36.93, 32.59, 29.64, 29.16, 29.11, 28.51, 26.83, 26.75. HRMS (ESI)m/zcalcd. for C50H62N8O4[M+H]+839.4967, found: 839.4963. Purity: 98.6 %.
Example 10
Synthesis of 2- ((3 r,5r,7 r) -adamantan-1-yl) -N- (10- (2- (4- (((2- (3- ((3- (3-cyanophenyl) -6-oxopyridazin-1 (6H) -yl) methyl) phenyl) pyrimidin-5-yl) oxy) methyl) piperidin-1-yl) acetamido) decyl) acetamide (H10):
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the H10 is synthesized by the same method of H6 by taking N-7 and N-8e as raw materials, and the yield is increased 30 %.1H NMR (400 MHz, Chloroform-d)δ8.60 (t,J= 1.8 Hz, 1H), 8.50 (s, 2H), 8.30 (dt,J= 7.9, 1.5 Hz, 1H), 8.20 (t,J= 1.8 Hz, 1H), 7.98 (dt,J= 8.0, 1.5 Hz, 1H), 7.70 (dt,J= 7.7, 1.4 Hz, 1H), 7.64 (d,J= 9.7 Hz, 1H), 7.57 (t,J= 7.9 Hz, 2H), 7.45 (t,J= 7.7 Hz, 1H), 7.07 (d,J= 9.7 Hz, 1H), 5.50 (s, 2H), 5.38 (m, 1H), 3.98 (d,J= 5.8 Hz, 2H), 3.34 – 3.20 (m, 4H), 3.16 – 2.87 (m, 4H), 2.30 (s, 2H), 1.99 – 1.87 (m, 9H), 1.74 – 1.61 (m, 10H), 1.50 (m, 7H), 1.28 (m, 12H). 13C NMR (101 MHz, DMSO-d6)δ170.08, 159.22, 156.33, 152.08, 144.57, 142.41, 137.82, 137.38, 135.90, 133.29, 131.23, 130.81, 130.68, 130.58, 130.11, 129.88, 129.38, 127.44, 126.86, 118.96, 112.64, 73.25, 55.08, 53.36, 50.57, 42.60, 38.65, 38.63, 36.93, 32.58, 29.65, 29.63, 29.43, 29.39, 29.17, 28.53, 26.89, 26.81. HRMS (ESI)m/zcalcd. for C52H66N8O4[M+H]+867.5280, found: 867.5277. Purity: 97.3 %.
Example 11
Synthesis of N- (((3 r,5r,7 r) -adamantan-1-yl) -methyl) -6- (4- (((2- (3- ((3- (3-cyanophenyl) -6-oxopyridazin-1 (6H) -yl) methyl) phenyl) pyrimidin-5-yl) oxy) methyl) piperidin-1-yl) hexanamide (H11):
The N-9a and the 1-adamantane methylamine are used as raw materials to synthesize H11 by the same method of H6, and the yield is increased 30 %.1H NMR (400 MHz, DMSO-d6)δ8.63 (s, 2H), 8.38 (dd,J= 4.4, 2.2 Hz, 2H), 8.30 – 8.20 (m, 2H), 8.17 (d,J= 9.8 Hz, 1H), 7.93 (dt,J= 7.7, 1.4 Hz, 1H), 7.72 (t,J= 7.9 Hz, 1H), 7.59 (t,J= 6.3 Hz, 1H), 7.53 – 7.43 (m, 2H), 7.16 (d,J= 9.7 Hz, 1H), 5.44 (s, 2H), 4.03 (d,J= 5.9 Hz, 2H), 2.94 – 2.80 (m, 2H), 2.75 (d,J= 6.2 Hz, 2H), 2.23 (t,J= 7.4 Hz, 2H), 2.09 (t,J= 7.3 Hz, 2H), 1.95 – 1.82 (m, 5H), 1.73 (ms, 3H), 1.68 – 1.61 (m, 3H), 1.60 – 1.49 (m, 5H), 1.46 – 1.36 (m, 8H), 1.34 – 1.20 (m, 4H). ESI-MSm/z: 740.4 [M+H]+. 13C NMR (101 MHz, DMSO-d6)δ172.68, 159.25, 156.26, 152.15, 144.61, 142.46, 137.83, 137.42, 135.93, 133.33, 131.30, 130.87, 130.73, 130.61, 129.93, 129.43, 127.39, 126.85, 118.98, 112.64, 73.45, 58.75, 55.08, 53.38, 50.46, 40.33, 37.02, 35.92, 35.81, 34.08, 28.88, 28.17, 27.15, 26.74, 25.95. HRMS (ESI)m/zcalcd. for C45H53N7O3[M+H]+740.4283, found: 740.4284. Purity: 99.6 %.
Example 12
Synthesis of N- (((3 r,5r,7 r) -adamantan-1-yl) -methyl) -4- (4- (((2- (3- ((3- (3-cyanophenyl) -6-oxopyridazin-1 (6H) -yl) methyl) phenyl) pyrimidin-5-yl) oxy) methyl) piperidin-1-yl) butanamide (H12):
H12 is synthesized by the same method of H6 by taking N-9b and 1-adamantanamine as raw materials, and the yield is increased 30 %.1H NMR (400 MHz, DMSO-d6)δ8.63 (s, 2H), 8.42 – 8.34 (m, 2H), 8.28 – 8.20 (m, 2H), 8.17 (d,J= 9.8 Hz, 1H), 7.92 (dt,J= 7.7, 1.4 Hz, 1H), 7.71 (t,J= 7.9 Hz, 1H), 7.61 (t,J= 6.3 Hz, 1H), 7.52 – 7.45 (m, 2H), 7.16 (d,J= 9.8 Hz, 1H), 5.44 (s, 2H), 4.03 (d,J= 5.8 Hz, 2H), 2.87 (m, 2H), 2.76 (d,J= 6.2 Hz, 2H), 2.26 (t,J= 7.2 Hz, 2H), 2.12 (t,J= 7.3 Hz, 2H), 1.97 – 1.83 (m, 5H), 1.82 – 1.70 (m, 3H), 1.65 (m, 5H), 1.56 (m, 3H), 1.41 (m, 6H), 1.37 – 1.26 (m, 2H).13C NMR (101 MHz, DMSO-d6)δ172.61, 159.23, 156.24, 152.12, 144.57, 142.43, 137.81, 137.40, 135.89, 133.32, 131.28, 130.84, 130.71, 130.60, 130.11, 129.91, 129.42, 127.39, 126.84, 118.99, 112.62, 73.40, 58.16, 55.09, 53.32, 50.47, 40.31, 37.02, 35.87, 34.11, 33.85, 31.44, 28.85, 28.16, 26.80, 23.38. HRMS (ESI)m/zcalcd. for C43H49N7O3[M+H]+712.3970, found: 712.3970. Purity: 95.3 %.
Example 13
Synthesis of 2- ((3 r,5r,7 r) -adamantan-1-yl) -N- (2- (4- (((2- (3- ((3- (3-cyanophenyl) -6-oxopyridazin-1 (6H) -yl) methyl) phenyl) pyrimidin-5-yl) oxy) methyl) piperidin-1-yl) ethyl) acetamide (H13):
h13 is synthesized by the same method of H6 by taking N-10 and adamantane acetic acid as raw materials, and the yield is increased 20 %.1H NMR (400 MHz, Chloroform-d)δ8.59 (d,J= 1.8 Hz, 1H), 8.49 (s, 2H), 8.29 (d,J= 7.8 Hz, 1H), 8.19 (d,J= 1.9 Hz, 1H), 7.98 (d,J= 7.9 Hz, 1H), 7.70 (d,J= 7.7 Hz, 1H), 7.64 (d,J= 9.7 Hz, 1H), 7.57 (t,J= 8.0 Hz, 2H), 7.45 (t,J= 7.7 Hz, 1H), 7.30 (d,J= 8.1 Hz, 1H), 7.07 (d,J= 9.7 Hz, 1H), 5.50 (s, 2H), 3.97 (d,J= 5.9 Hz, 2H), 3.40 (q,J= 5.6 Hz, 2H), 3.04 (d,J= 11.2 Hz, 2H), 2.58 (d,J= 6.1 Hz, 2H), 2.21 – 2.11 (m, 2H), 1.97 (s, 4H), 1.94 – 1.85 (m, 4H), 1.73 – 1.63 (m, 10H), 1.49 (m, 1H), 1.25 (s, 3H). 13C NMR (101 MHz, DMSO-d6)δ169.71, 158.67, 155.69, 151.54, 144.02, 141.88, 137.24, 136.85, 135.33, 132.77, 130.73, 130.29, 130.16, 130.04, 129.56, 129.35, 128.88, 126.80, 126.28, 118.43, 112.05, 72.69, 57.06, 54.52, 52.56, 50.05, 41.98, 36.36, 35.72, 34.91, 32.04, 27.95. HRMS (ESI)m/zcalcd. for C42H47N7O3[M+H]+698.3813, found: 698.3814. Purity: 99.7 %.
3. Biological evaluation experiment:
(1) In vitro antitumor cell proliferation activity assay:
The inhibition of MET gene amplified hepatocellular carcinoma mhc 97H by the compounds was determined according to CCK-8 method (Cell Counting Kit-8) and half inhibitory concentration IC 50 values of the compounds for inhibition of cell proliferation activity were obtained. Inoculating cells in logarithmic growth phase at 3000-5000 cells/hole in 96-well plate, and culturing at 37deg.C under 5% CO 2 for 12-24 hr; adding 100 mu L of gradient diluted compound solutions to be tested with different concentrations into the culture plate, and incubating the culture plate for 72 hours under the conditions of 37 ℃ and 5% CO 2 incubator; before the end of incubation, 4 h. Mu.L of CCK-8 solution (5 mg/mL) was added to each well. After the incubation, OD 450 was measured with an enzyme-labeled instrument, inhibition rate = (control OD value-experimental OD value)/control OD value x 100%; after the data is obtained, GRAPHPAD PRISM fits yield IC 50.
From the experimental results, it can be seen that the compounds of the present invention have strong antiproliferative activity on MET gene-amplified hepatocellular carcinoma mhc 97H, and the results are shown in table 1, wherein the ICs 50 of the compounds are classified according to the description:
"A" means IC 50 measured as 50 or less nM;
"B" means IC 50 measured 500 or less and nM or more than 50 nM;
"C" means IC 50 measured at 1 μM or less and greater than 500 nM;
"D" means IC 50 measured greater than 1. Mu.M;
"-" indicates that IC 50 was not measured.
(2) C-Met protein degradation Activity assay:
MHCC97H cells (5X 105 cells/well) were seeded into 6-well plates and treated with different concentrations of the compound of interest for 24 hours. Cells were collected and lysed in 100 μl RIPA lysis buffer (Thermo FISHER SCIENTIFIC, USA), after 15 minutes the lysate was centrifuged to collect the supernatant. Protein concentration was measured using a Bradford protein assay kit and microplate reader (Bio-Tek, USA). Then, the same mass of protein (5-10. Mu.g) was added to the prepared buffer and denatured by boiling in 100℃water. Proteins were separated by 10% SDS-PAGE and transferred to PVDF membrane. Membranes were blocked and further incubated overnight with antibody. The next day was rinsed and after one hour incubation with secondary antibodies of the same species was rinsed. Western blot images were taken using an imager.
From the experimental results, it can be seen that some of the example compounds of the present invention have better degradation activity on c-Met protein in the mhc c97H cell line, and the results are shown in table 1, where the degradation rate of the compounds on c-Met protein is 1 μm:
"A" represents a degradation rate measurement value of 50% or more;
"B" represents a degradation rate measurement value of less than 50% and 10% or more;
"C" means that the degradation rate measurement value is less than 10%;
"-" indicates that the degradation rate was not measured.
TABLE 1 antiproliferative activity and c-Met degrading Activity of the inventive Compounds on MHCC97H cell lines
(3) In vitro Activity assay of mutant cell lines:
TABLE 2 antiproliferative Activity of Compound H11 of the invention on MET fusions and various mutant cells
The inhibition of the proliferation of the engineered cell lines Ba/F3-TPR-MET, ba/F3-TPR-MET-D1228N and Ba/F3-TPR-MET-Y1230H by the compounds was determined according to CELLTITER GLO and half inhibition concentration IC 50 values of the cell proliferation inhibition activity of the compounds were obtained. Inoculating cells in logarithmic growth phase at 5000 cells/hole in 96-well plate, and culturing at 37deg.C under 5% CO 2 for 12-24 hr; 100. Mu.L of gradient diluted solutions of the test compounds at different concentrations were added to the plates, and the plates were incubated at 37℃for 72 hours in a 5% CO 2 incubator. The assay plates were equilibrated to room temperature prior to measurement, 50 [ mu ] L CELLTITER Glo cube reagent was added to each well, the contents were mixed on an orbital shaker for 2 minutes to induce cell lysis, followed by incubation at room temperature for 10 minutes to stabilize the luminescence signal, and the luminescence value (RLU) on SpectraMax Paradigm was recorded by an enzyme-labeled instrument. Inhibition (Inh%) was calculated relative to vehicle (DMSO) -treated control wells using the following formula: inhibition ratio (Inh%) =100- (compound group RLU-blank group RLU)/(control group RLU-blank group RLU) ×100%; after the data is obtained, GRAPHPAD PRISM fits yield IC 50.

Claims (10)

1. A compound or a pharmaceutically acceptable salt thereof, characterized in that it has a chemical structure according to formula (I):
wherein HyT groups are selected from: ；
Wherein linker is a linking chain which is a straight or branched alkylene chain of 1 to 15 atoms total length, the straight or branched alkylene chain is composed of-CH 2 -; -O-, -CO-, -one or more of CONH-, -N-, alkynylene or cycloalkylene.
2. A compound or pharmaceutically acceptable salt thereof according to claim 1,
The HyT groups are selected from:；
In the Linker, the number of the links is as follows, the straight or branched alkylene chain is composed of-CH 2 -, -O-, -one or more of CO-, -CONH-, -N-.
3. A compound or pharmaceutically acceptable salt thereof according to claim 1,
The HyT groups are selected from:；
in the Linker, the number of the links is as follows, the straight or branched alkylene chain is composed of-CH 2 -; -one or more of CO-, -CONH-.
4. A compound or pharmaceutically acceptable salt thereof according to claim 1,
When HyT groups are any one of the following groups:，
The connecting chain is-COCH 2 -;
when HyT groups are the following:
，
The connecting chain is-CH 2CONH-CH2 -;
when HyT groups are the following:
，
The connecting chain is a straight or branched alkylene chain of 1 to 15 atoms total length, the straight or branched alkylene chain is composed of-CH 2 -, -O-, -one or more of CO-, -CONH-, -N-.
5. The compound according to claim 1, or a pharmaceutically acceptable salt thereof, selected from any one of the compounds of structural formulas H1 to H13:
。
6. The compound of claim 1, or a pharmaceutically acceptable salt thereof, wherein the pharmaceutically acceptable salt is an acid addition salt of a compound of formula (I), wherein the acid used for salt formation is an inorganic acid, which is hydrochloric acid, sulfuric acid, or phosphoric acid, or an organic acid, which is acetic acid, trichloroacetic acid, trifluoroacetic acid, propionic acid, butyric acid, maleic acid, p-toluenesulfonic acid, malic acid, methanesulfonic acid, malonic acid, cinnamic acid, citric acid, fumaric acid, camphoric acid, digluconic acid, aspartic acid, or tartaric acid.
7. A method of preparing a compound of claim 1, or a pharmaceutically acceptable salt thereof, comprising the steps of:
Dissolving the compound A and the compound B in an organic solvent, adding triethylamine, 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride and 1-hydroxybenzotriazole, and obtaining the compound for targeted degradation of c-Met protein after the reaction is finished.
8. A method of preparing a compound of claim 1, or a pharmaceutically acceptable salt thereof, further comprising the steps of: Dissolving the compound C and the compound D in an organic solvent, adding triethylamine, 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride and 1-hydroxybenzotriazole, and obtaining the compound for targeted degradation of C-Met protein after the reaction is finished.
9. A pharmaceutical composition comprising a compound of formula (I) according to claim 1 or a pharmaceutically acceptable salt thereof, and a pharmaceutically acceptable carrier.
10. Use of a compound according to claim 1 or a pharmaceutically acceptable salt thereof for the manufacture of a medicament for the prevention and/or treatment of cancer or a tumor-related disease.
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Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101687857A (en) * 2007-07-12 2010-03-31 默克专利有限公司 Pyridazinone derivates
CN102264727A (en) * 2008-12-23 2011-11-30 默克专利有限公司 Pyridazinone derivatives
CN116283918A (en) * 2023-03-16 2023-06-23 泰比棣医药科技(石家庄)有限公司 Bifunctional compound for degrading receptor tyrosine kinase and application thereof
CN116496253A (en) * 2022-08-19 2023-07-28 中国人民解放军军事科学院军事医学研究院 c-MET protein targeted degradation agent and medical application thereof

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101687857A (en) * 2007-07-12 2010-03-31 默克专利有限公司 Pyridazinone derivates
CN102264727A (en) * 2008-12-23 2011-11-30 默克专利有限公司 Pyridazinone derivatives
CN116496253A (en) * 2022-08-19 2023-07-28 中国人民解放军军事科学院军事医学研究院 c-MET protein targeted degradation agent and medical application thereof
CN116283918A (en) * 2023-03-16 2023-06-23 泰比棣医药科技(石家庄)有限公司 Bifunctional compound for degrading receptor tyrosine kinase and application thereof

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