Chromone spiro compound as well as preparation method and application thereof

Abstract

The invention discloses a chromone spiro compound and a preparation method and application thereof.The chromone spiro compound is shown in the formula I. The chromone spiro compound has selective CDK4 / 6 inhibitory activity, can relieve and / or treat myelosuppression caused by chemotherapy, improve weight loss caused by chemotherapy and increase the survival rate and has good medicine safety; the compound is simple in preparation method and convenient for large-scale production, and has great application prospects.

Description

Technical Field

[0001] The invention relates to a chromone spirocyclic compound and also to a preparation method and application of the compound. Background Art

[0002] Chemotherapy-induced myelosuppression (CIM) is a common adverse reaction in cancer chemotherapy, with clinical manifestations of neutropenia, lymphopenia, anemia and thrombocytopenia, which can lead to adverse consequences including infection, sepsis, bleeding and fatigue, and in severe cases, death of patients. At present, the treatment strategies for CIM in clinical practice are symptomatic treatment, and the main drugs are the use of growth factors such as recombinant human granulocyte colony-stimulating factor, erythropoietin and thrombopoietin. However, these drugs only target single lineage blood cells, have slow onset and long duration of medication, and may cause bone marrow depletion and the risk of drug-related adverse reactions. Therefore, the prophylactic use of a drug that can temporarily inhibit the proliferation of bone marrow cells can reduce the sensitivity of bone marrow cells to chemotherapeutic drugs, providing a new strategy for preventing or alleviating CIM.

[0003] Cyclin-dependent kinases (CDKs) play an important role in regulating the cell cycle. CDK4 / 6 belongs to the serine / threonine kinase family. In the early G1 phase of the cell cycle, CDK4 / 6 binds to cyclin D to form a CDK4 / 6-cyclin D complex, which promotes the phosphorylation of retinoblastoma protein (RB), thereby releasing the transcription factor E2F and regulating the transition of the cell cycle from G1 phase to S phase. For hematopoietic stem and progenitor cells (HSPCs), CDK4 / 6 is crucial for the proliferation and regulation of HSPCs.

[0004] CDK4 / 6 is a key initiator protein for the transition of the cell cycle from the G1 phase to the S phase. The inhibition of CDK4 / 6 can block the cell cycle in the G1 phase, thereby reducing cell proliferation and playing an important role in the treatment of cancer and CIM. Currently, five selective CDK4 / 6 inhibitors, Palbociclib, Ribociclib, Abemaciclib, Dalpiciclib and Trilaciclib (TC) have been approved for marketing, of which the first four are used for the treatment of breast cancer, and TC is the only one approved by the US FDA for the treatment of CIM in patients with advanced small cell lung cancer. Intravenous infusion of TC before chemotherapy can significantly induce G1 phase arrest of HSPCs and protect bone marrow cells from toxic damage of chemotherapy drugs.

[0005] Although TC is the only FDA-approved CDK4 / 6 inhibitor for the treatment of CIM, it still has some shortcomings, such as: (1) TC was initially developed as an anticancer drug, and was found to have a therapeutic effect on CIM in later clinical trials, lacking research on drug toxicity and bone marrow penetration; (2) In clinical studies, TC can significantly increase the count of neutrophils and red blood cells, but has little effect on platelets, thereby increasing the risk of bleeding in patients. Therefore, new CDK4 / 6 inhibitors with novel structures, lower toxicity, better effects, and for the treatment of CIM are still highly anticipated. Summary of the invention

[0006] Purpose of the invention: The purpose of the present invention is to provide a chromone spirocyclic compound with selective CDK4 / 6 inhibitory activity, and also provide a method for preparing the above-mentioned compound and its application in the preparation of CDK4 / 6 inhibitors, and drugs for alleviating and / or treating chemotherapy-induced bone marrow suppression.

[0007] Technical solution: The present invention discloses a chromone spirocyclic compound, the structure of which is shown in Formula I:

[0008]

[0009] Wherein, X is selected from O, (CH2) n , C(O), NH or S(O)2, n is 0 or 1;

[0010] R1, R2 are selected from C3-C8 cycloalkyl, C3-C8 cycloalkyloxy or C3-C8 heterocyclyl, wherein the C3-C8 cycloalkyl, C3-C8 cycloalkyloxy or C3-C8 heterocyclyl is independently and optionally substituted with one or more substituents selected from halogen, hydroxyl, amino, nitro, cyano, C1-C6 alkyl, C1-C6 alkoxy, C3-C6 cycloalkyl, C3-C6 cycloalkyloxy or C4-C8 heterocyclyl;

[0011] R3 is hydrogen, halogen, hydroxyl, mercapto, amino, -OR', -SR', -NHR' or -NR'R", wherein R' and R" are C1-C3 alkyl, C2-C6 alkenyl, C3-C6 allyl, C2-C6 alkynyl, C1-C3 haloalkyl, C3-C6 cycloalkyl, C6-C 10 Aryl, C3-C 10 Heteroaryl or C4-C8 heterocyclic group;

[0012] R4 is selected from C3-C8 cycloalkyloxy, C3-C8 heterocyclyl, C3-C8 heterocyclylphenyl or C3-C8 heterocyclylaryl, wherein the C3-C8 cycloalkyloxy, C3-C8 heterocyclyl, C3-C8 heterocyclylphenyl or C3-C8 heterocyclylaryl is independently and optionally substituted by one or more substituents selected from halogen, hydroxyl, amino, nitro, cyano, C1-C6 alkyl, C1-C6 alkoxy, C3-C6 cycloalkyl, C3-C6 cycloalkyloxy or C4-C8 heterocyclyl.

[0013] Wherein, X is C(O) or (CH2) n ; n is 0 or 1; R1 and R2 are C3-C8 cycloalkyl or C3-C8 heterocyclic group; R3 is hydrogen or halogen; R4 is C3-C8 heterocyclic group. Wherein E is glutamic acid, R1 and R2 are organic carbon chains selected from C6-C 24 Alkyl, substituted C6-C 24 Alkyl, C6-C 24 Alkenyl, substituted C6-C 24 Alkenyl, C6-C 24 Alkynyl or substituted C6-C 24 Alkynyl; n is a positive integer ≥1.

[0014] Wherein, X is C(O) or (CH2) n ; n is 0 or 1; R1 and R2 are C4-C6 cycloalkyl or C6 heterocyclic group; R3 is halogen; R4 is C6 heterocyclic group.

[0015] Wherein, the X is selected from: C(O) or (CH2) n ; n is 0 or 1; R1 and R2 are selected from cyclobutyl, cyclopentyl, cyclohexyl, cyclohexyl derivatives, tetrahydropyran, tetrahydrothiothiopyran or N-amide piperidine; R3 is F; R4 is selected from piperidine, piperidine derivatives, morpholine, piperazine or piperazine derivatives.

[0016] Wherein, the above-mentioned chromone spirocyclic compound is specifically selected from the following compounds 1-55:

[0017]

[0018]

[0019] The pharmaceutically acceptable salts of the above-mentioned chromone spirocyclic compounds are also included, specifically the acid-addition salts of the compounds of the general formula (I), wherein the acid used for salt formation includes inorganic acids and organic acids, wherein the inorganic acids include: hydrochloric acid, sulfuric acid, phosphoric acid and methanesulfonic acid; the organic acids include: galactaric acid, D-glucuronic acid, glycerophosphoric acid, hippuric acid, isethionic acid, lactobionic acid, maleic acid, naphthalene-2-sulfonic acid, pivalic acid, terephthalic acid , bile acid, benzenesulfonic acid, citric acid, lactic acid, malic acid, malonic acid, mandelic acid, acetic acid, propionic acid, tartaric acid, succinic acid, formic acid, methanesulfonic acid, picric acid, L-pyroglutamic acid, salicylic acid, gentisic acid, p-toluenesulfonic acid, valeric acid, palmitic acid, stearic acid, lauric acid, adipic acid, 4-benzenesulfonic acid, fumaric acid, 3-hydroxynaphthalene-2-carboxylic acid, ascorbic acid, camphoric acid, camphorsulfonic acid, cinnamic acid, dichloroacetic acid and ethanesulfonic acid.

[0020] The preparation method of the above-mentioned chromone spiro compound is as follows: (1) 1-(4-bromo-2-hydroxyphenyl)ethanone shown in formula II is subjected to a condensation reaction with a cyclic ketone under alkaline conditions to obtain a chromone intermediate compound III; (2) the chromone intermediate compound III is reacted with a bis-pinacol borate in the presence of a palladium catalyst to obtain a compound IV; (3) compound IV is subjected to a coupling reaction with a chloropyrimidine derivative to obtain a compound V; (4) compound V is subjected to a coupling reaction with a 2-aminopyrimidine derivative in the presence of a palladium catalyst and a phosphine ligand to obtain a chromone spiro compound shown in formula I;

[0021] Specifically, 1-(4-bromo-2-hydroxyphenyl)ethanone (II) is first subjected to a Knoevenagel condensation reaction with a differently substituted cyclic ketone under alkaline conditions to obtain a corresponding chromone intermediate compound of formula (III) of each substitution type, and then reacted with a bis-pinacol borate in the presence of a palladium catalyst to obtain a compound of formula (IV), and then subjected to a Suzuki coupling reaction with a chloropyrimidine derivative to obtain a compound of formula (V); the compound of formula (V) is subjected to a Buchwald-Hartwig coupling reaction with a 2-aminopyrimidine derivative in the presence of a palladium catalyst and a phosphine ligand to obtain a compound of formula (I); the synthetic route is as follows:

[0022]

[0023] Wherein, in step (1), compound III is prepared from compound II, and the base used is selected from pyrrolidine, piperidine, tetrahydropyridine, sodium hydroxide, potassium hydroxide, potassium carbonate or sodium carbonate, preferably pyrrolidine. The reaction solvent is selected from methanol, ethanol, isopropanol, perfluoroisopropanol, n-butanol or tert-butanol, preferably methanol. The reaction temperature is 50-100°C, preferably 60-80°C.

[0024] Wherein, in step (2), compound IV is prepared from compound III, and the base used is selected from potassium phosphate, potassium carbonate, sodium carbonate, cesium carbonate, potassium fluoride, potassium acetate, sodium acetate, potassium tert-butoxide or sodium tert-butoxide, preferably potassium acetate. The palladium catalyst used is selected from Pd(dppf)Cl2, Pd(PPh3)2Cl2, Pd(PPh3)4, Pd2(dba)3, PdCl2(dippf)2 or [Pd(allyl)Cl]2, preferably Pd(dppf)Cl2. The reaction solvent is selected from dioxane, N,N-dimethylformamide, N,N-dimethylacetamide, dimethyl sulfoxide, acetonitrile or N-methylpyrrolidone, preferably dioxane. The reaction temperature is 50-100°C, preferably 80-100°C.

[0025] Wherein, in step (3), compound V is prepared from compound IV, and the base used is selected from sodium hydroxide, potassium hydroxide, barium hydroxide, potassium phosphate, potassium carbonate, sodium carbonate, cesium carbonate, potassium fluoride, potassium acetate, sodium acetate, sodium methoxide, potassium methoxide, sodium ethoxide, potassium ethoxide, potassium tert-butoxide or sodium tert-butoxide, preferably sodium carbonate. The palladium catalyst used is selected from Pd(dppf)Cl2, Pd(PPh3)2Cl2, Pd(PPh3)4, Pd2(dba)3, PdCl2(dippf)2 or [Pd(allyl)Cl]2, preferably Pd(PPh3)2Cl2. The reaction solvent is selected from tetrahydrofuran, ethanol, acetonitrile, dioxane, ethylene glycol dimethyl ether, dimethyl sulfoxide or toluene, preferably ethylene glycol dimethyl ether. The volume ratio of the reaction solvent to water is 8:1 to 12:1, preferably 10:1. The reaction temperature is 50-80°C, preferably 60-80°C.

[0026] Wherein, in step (4), compound I is prepared from compound V, and the base used is selected from sodium hydroxide, potassium hydroxide, potassium carbonate, sodium carbonate, cesium carbonate, sodium methoxide, potassium methoxide, sodium ethoxide, potassium ethoxide, potassium tert-butoxide or sodium tert-butoxide, preferably cesium carbonate. The palladium catalyst used is selected from Pd2(dba)3 or Pd(OAc)2, preferably Pd2(dba)3. The phosphine ligand used is selected from PCy3, P(t-Bu)3, P(o-tolyl)3, DPPF, JohnPhos, SPhos, XantPhos or BINAP, preferably BINAP. The reaction solvent is selected from toluene, xylene, dioxane, tetrahydrofuran, ethylene glycol dimethyl ether, N,N-dimethylformamide, N,N-dimethylacetamide or dimethyl sulfoxide, preferably dioxane. The reaction temperature is 60-110°C, preferably 80-100°C.

[0027] The above-mentioned chromone spirocyclic compound can be used in the preparation of CDK4 / 6 inhibitors.

[0028] The above-mentioned chromone spirocyclic compound can also be used in the preparation of drugs for alleviating and / or treating chemotherapy-induced bone marrow suppression.

[0029] The present invention also provides a pharmaceutical composition, which comprises a therapeutically effective amount of a compound of formula (I) or a pharmaceutically acceptable salt or isomer thereof, and a pharmaceutically acceptable carrier.

[0030] Among them, the therapeutically effective amount can be an amount that alleviates one or more symptoms of a disease or condition in a subject to some extent, partially or completely restores one or more physiological or biochemical parameters related to the disease or condition or the cause thereof to normal, and / or reduces the feasibility of the onset of the disease or condition; a pharmaceutically acceptable carrier refers to an excipient or diluent that does not cause significant irritation to an organism and does not interfere with the biological activity and properties of the administered compound.

[0031] Principle of the invention: Currently, the only small molecule drug for the treatment of bone marrow suppression in clinical practice is triclasiclib, which is a selective CDK4 / 6 inhibitor. By analyzing the structures of triclasiclib and other CDK4 / 6 inhibitors on the market, it is found that triclasiclib can play a bone marrow protective role due to the special azaspiro skeleton in its structure. Therefore, based on the structure of triclasiclib, the present invention uses the skeleton replacement principle to replace the azaspiro skeleton in its structure with the chromone spiro skeleton, while retaining the active fragment pyrimidine-NH-pyridine structure of the selective CDK4 / 6 inhibitor, and synthesizes a series of chromone spiro derivatives. This type of derivative not only has a good CDK4 / 6 inhibitory effect, but also has an excellent bone marrow protective effect.

[0032] Beneficial effects: Compared with the prior art, the present invention has the following significant advantages: (1) The chromone spirocyclic compound of the present invention has selective CDK4 / 6 inhibitory activity, can alleviate and / or treat chemotherapy-induced bone marrow suppression, improve chemotherapy-induced weight loss and increase survival rate, and has good drug safety; (2) The synthesis steps of the chromone spirocyclic compound of the present invention are short, the post-processing is simple, and it is convenient for large-scale production, and has great application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0033] Figure 1 is the structural formula of the chromone spiro compound of the present invention;

[0034] Figure 2 Schematic diagram of the interaction between compound 42 and CDK6 (PDB: 5L2S) protein;

[0035] Figure 3Schematic diagram of the in vivo efficacy results of compound 42, wherein Figure A is a time cycle diagram of animals, Figures B to G are the effects of 42 on platelets, red blood cells, hemoglobin, white blood cells, lymphocytes and neutrophils in the blood; Figure H is a schematic diagram of the weight changes of mice after treatment with 5-FU or 42; Figure I is a 14-day survival curve of mice after treatment with 5-FU or 42;

[0036] Figure 4 It is a schematic diagram of the safety evaluation of compound 42, wherein Figures A to B are schematic diagrams of the changes in food intake of female mice and male mice after administration of 42; Figures C to D are diagrams of organ coefficients of female mice and male mice after administration of 42.

[0037] Figure 5 Figure 1 is a schematic diagram of HE staining results, wherein Figure A is the HE staining image of various tissues of female mice, and Figure B is the HE staining image of various tissues of male mice. DETAILED DESCRIPTION

[0038] The technical scheme of the present invention is further described below in conjunction with the examples. The test materials used in the examples can all be purchased through conventional channels.

[0039] Example 1 7-(5-fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,1'-cyclobutyl]-4-one (Compound 1)

[0040]

[0041] 4-Bromo-2-hydroxyacetophenone II (3 g, 23.2 mmol), cyclobutanone (1.96 g, 28 mmol) and pyrrolidine (2.5 g, 35 mmol) were dissolved in 50 mL of methanol. Stirred at 60 °C for 8 h, and the reaction was completed by TLC. The reaction solution was cooled to room temperature, poured into 100 mL of water, extracted with ethyl acetate (50 mL × 3), and the organic phases were combined, washed with water (80 mL) and saturated sodium chloride solution (80 mL), dried over anhydrous Na2SO4, and concentrated under reduced pressure to remove the solvent. The residue was purified by silica gel column chromatography (PE / EA, 40:1-25:1) to obtain compound III-1 as a light yellow liquid (1.61 g, 43.1%). 1 H NMR (300MHz, CDCl3) δ7.71(d,J=8.4Hz,1H),7.20(d,J=1.8Hz,1H),7.14(dd,J=8.4,1.8Hz,1 H),2.90(s,2H),2.42-2.27(m,2H),2.12-2.13(m,2H),2.03-1.88(m,1H),1.81-1.65(m,1H).

[0042] Dissolve III-1 (1.54 g, 5.78 mmol), bis(pyrazol)-2-ylpyrazol (1.76 g, 6.94 mmol), Pd(dppf)2Cl2 (0.21 g, 0.29 mmol) and KOAc (1.7 g, 17.34 mmol) in 20 mL of dioxane. Stir at 80 °C for 24 h under N2 atmosphere. The reaction is complete after TLC detection. After the reaction solution is cooled to room temperature, the insoluble matter is removed by suction filtration, and the filtrate is concentrated under reduced pressure. The resulting residue is purified by silica gel column chromatography (PE / EA, 10:1-6:1) to obtain compound IV-1 as a light yellow liquid (1.55 g, 85.7%). 1 H NMR (300MHz, CDCl3) δ7.84(d,J=7.7Hz,1H),7.46(s,1H),7.41(dd,J=7.7,1.0Hz,1H),2.92(s, 2H),2.39-2.25(m,2H),2.21-2.13(m,2H),2.01-1.87(m,1H),1.82-1.64(m,1H),1.36(s,12H).

[0043] IV-1 (0.95 g, 3 mmol), 2,4-dichloro-5-fluoropyrimidine (0.75 g, 4.5 mmol), Pd(PPh3)2Cl2 (63 mg, 0.29 mmol) and Na2CO3 (0.65 g, 7.5 mmol) were dissolved in a mixed solution of ethylene glycol dimethyl ether (DME) and water (9 mL / 1 mL). Stir at 80 °C for 12 h under N2 atmosphere, and the reaction was completed by TLC. After the reaction solution was cooled to room temperature, the mixture was poured into 30 mL of water, extracted with ethyl acetate (20 mL×3), the organic phases were combined, washed with water (60 mL) and saturated sodium chloride solution (60 mL), dried over anhydrous Na2SO4, concentrated under reduced pressure to remove the solvent, and the residue was purified by silica gel column chromatography (PE / EA, 6:1~2:1) to obtain compound V-1 as a white solid (0.52 g, 53.3%). 1 H NMR (300MHz, CDCl3) δ8.60(d,J=2.9Hz,1H),8.00(d,J=8.2Hz,1H),7.82-7.80(m,1H),7.76(dt,J=8.3 ,1.5Hz,1H),2.98(s,2H),2.46-2.33(m,2H),2.28-2.17(m,2H),2.07-1.91(m,1H),1.84-1.72(m,1H). 13C NMR (151MHz, CDCl3) δ191.94,159.50,156.00,155.69,155.67,154.22,152.94,152.88,149.17,149 .00,138.39,138.35,127.24,122.82,121.45,121.41,119.64,119.59,81.39,46.02,33.07,12.16.

[0044] In a 25 ml sealed tube, add V-1 (220 mg, 0.7 mmol), 5-(4-methylpiperazine-1-yl)-pyridine-2-amino (162 mg, 0.84 mmol), Pd2(dba)3 (64 mg, 0.07 mmol), BINAP (87 mg, 0.14 mmol), Cs2CO3 (456 mg, 1.4 mmol) and dioxane (8 mL) in sequence. Stir at 100 ° C for 12 h under N2 atmosphere, and the reaction is completed by TLC detection. After the reaction solution is cooled to room temperature, it is concentrated under reduced pressure to remove the solvent, and the residue is purified by silica gel column chromatography (DCM / CH3OH, 40:1-10:1) to obtain yellow solid compound 1 (135 mg, 41.4%). 1 H NMR (300MHz, CDCl3) δ8.47(d,J=3.3Hz,1H),8.33(s,1H),8.27(d,J=9.1Hz,1H),8.09(d,J=3.0Hz,1H),8.00(d,J=8.6H z,1H),7.76-7.72(m,2H),7.39(dd,J=9.1,3.0Hz,1H),3.24(t,J=5.0Hz,4H),2.99(s,2H),2.67(t,J=5.0Hz,4H),2.49 -2.35(m,5H),2.29-2.17(m,2H),2.06-1.93(m,2H). 13 C NMR (101MHz, CDCl3) δ192.27,159.49,155.67,155.64,152.08,150.48,150.39,149.53,147.81,147.55,146.19,142.94,140.71,1 40.66,136.83,127.03,126.52,122.11,121.43,121.38,119.35,119.28,112.37,81.21,54.94,49.63,46.10,46.05,33.12,12.16.

[0045] Example 2 7-(5-fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,1'-cyclopentyl]-4-one (Compound 2)

[0046] Compound 2 was prepared by referring to the method of compound 1; 1 H NMR (300MHz, CDCl3) δ8.48-8.44(m,2H),8.27(d,J=9.0Hz,1H),8.10(d,J=3.0 Hz,1H),8.01(d,J=8.2Hz,1H),7.72(dt,J=8.2,1.4Hz,1H),7.69-7.67(m,1H), 7.38(dd,J=9.1,3.0Hz,1H),3.22(t,J=5.0Hz,4H),2.91(s,2H),2.65(t,J=5.0 Hz,4H),2.41(s,3H),2.22-2.10(m,2H),1.98-1.90(m,2H),1.83-1.65(m,4H). 13 C NMR (101MHz, CDCl3) δ192.33,160.23,155.71,155.68,152.04,150.62,150.53,149.50,147.74,147.48,146.24,142.95,140.56,1 40.50,136.81,127.00,126.50,122.07,121.05,120.99,119.46,119.38,112.39,90.37,54.97,49.69,47.08,46.16,37.50,23.90.

[0047] Example 3 7-(5-fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,1'-cyclohexyl]-4-one (Compound 3)

[0048] Compound 3 was prepared by referring to the method of compound 1; 1H NMR (300MHz, CDCl3) δ8.47(d,J=3.3Hz,1H),8.36(s,1H),8.27(d,J=9.1Hz,1H),8.09(d,J=3.0Hz,1H),8.02-7.97(m,1H),7.73-7.69(m,2H),7.37(dd ,J=9.1,3.0Hz,1H),3.23(t,J=5.0Hz,4H),2.78(s,2H),2.66(t,J=5.0Hz, 4H),2.41(s,3H),2.12-1.95(m,3H),1.85-1.64(m,3H),1.62-1.49(m,4H). 13 C NMR (101MHz, CDCl3) δ192.35,159.45,155.66,152.08,150.69,150.60,149.54,147.71,147.44,146.16,142.99,140.66,140.60,1 36.81,126.77,126.45,121.98,120.97,120.91,119.35,119.27,112.40,80.45,54.96,49.68,48.23,46.14,34.75,25.15,21.45.

[0049] Example 4 7-(5-fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)-4'-methylspiro[benzopyran-2,1'-cyclohexyl]-4-one (Compound 4)

[0050] Compound 4 was prepared by referring to the method of compound 1; 1 H NMR (300MHz, CDCl3) δ8.47(d,J=3.3Hz,1H),8.33(s,1H),8.29(d,J=9.1Hz,1 H),8.09(d,J=2.9Hz,1H),8.00(d,J=8.1Hz,1H),7.76-7.68(m,2H),7.37(dd ,J=9.1,3.0Hz,1H),3.25(t,J=5.0Hz,4H),2.74(s,2H),2.71(t,J=5.0Hz,4H ),2.44(s,3H),2.20-2.12(m,2H),1.67-1.37(m,7H),0.99(d,J=3.1Hz,3H). 13CNMR(101MHz, CDCl3)δ192.43,159.38,155.67,155.64,152.06,150.65,150.56,149.51,147.74,147.48,146.20,142.96,140.69,140.63, 136.82,126.79,126.40,122.05,121.04,120.98,119.33,119.26,11 2.40,79.60,54.96,49.67,49.12,46.14,34.44,31.60,29.59,22.24.

[0051] Example 5 4'-ethyl-7-(5-fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,1'-cyclohexyl]-4-one (Compound 5)

[0052] Compound 5 was prepared by referring to the method of compound 1; 1 H NMR (300MHz, CDCl3) δ8.47(d,J=3.3Hz,1H),8.33-8.27(m,2H),8.09(d,J=3.3Hz,1H),8. 00(d,J=8.2Hz,1H),7.74(s,1H),7.71(dt,J=8.1,1.4Hz,1H),7.37(dd,J=9.1,3.0Hz,1H ),3.24(t,J=5.0Hz,4H),2.75(s,2H),2.68(t,J=4.9Hz,4H),2.43(s,3H),2.19(d,J=9.5 Hz,2H),1.67-1.57(m,2H),1.50-1.39(m,4H),1.37-1.19(m,3H),0.93(t,J=7.3Hz,3H). 13 C NMR (101MHz, CDCl3) δ192.45,159.40,155.68,155.65,152.04,150.61,150.52,149.49,147.78,147.52,146.25,142.92,140.70,140.65,136 .81,126.79,126.41,122.06,121.05,120.99,119.31,119.24,112.39, 79.97,54.96,49.65,49.15,46.13,38.26,34.41,29.57,27.26,11.52.

[0053] Example 6 4'-tert-butyl-7-(5-fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,1'-cyclohexyl]-4-one (Compound 6)

[0054] Compound 6 was prepared by referring to the method of compound 1; 1 H NMR(300MHz, CDCl3) δ8.46(d,J=3.3Hz,1H),8.29(d,J=9.7Hz,1H),8.23(s,1H),8.08 (d,J=3.8Hz,1H),8.00(d,J=7.4Hz,1H),7.74-7.68(m,2H),7.36(dd,J=9.1,3.0Hz,1 H),3.24(t,J=5.0Hz,4H),2.74(s,2H),2.69(t,J=5.1Hz,4H),2.43(s,3H),2.24(d,J =12.9Hz,2H),1.68-1.58(m,2H),1.57-1.35(m,4H),1.14-1.03(m,1H),0.92(s,9H). 13 C NMR (101MHz, CDCl3) δ192.46,159.40,155.64,155.61,152.06,150.65,150.56,149.52,147.77,147.51,146.15,142.96,140.69,140.63,136 .80,126.80,126.40,122.07,121.10,121.03,119.17,119.11,112.29, 79.62,54.95,49.65,49.14,47.05,46.13,34.96,32.48,27.57,21.99.

[0055] Example 7 7-(5-fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)-4',4'-dimethylspiro[benzopyran-2,1'-cyclohexyl]-4-one (Compound 7)

[0056] Compound 7 was prepared by referring to the method of compound 1; 1H NMR(300MHz, CDCl3)δ8.46(d,J=3.3Hz,1H),8.28(d,J=9.1Hz,1H),8.24(s,1H),8 .08(d,J=3.0Hz,1H),8.00(d,J=8.0Hz,1H),7.74-7.69(m,2H),7.37(dd,J=9.1,3. 0Hz,1H),3.24(t,J=5.0Hz,4H),2.79(s,2H),2.68(t,J=5.0Hz,4H),2.43(s,3H),2 .03-1.96(m,2H),1.72-1.59(m,4H),1.30-1.24(m,2H),1.02(s,3H),0.97(s,3H). 13 C NMR (126MHz, CDCl3) δ192.30,159.48,155.62,151.91,150.71,150.63,149.87,147.72,147.50,146.17,142.99,140.68,136.94, 126.82,126.46,122.08,121.06,121.02,119.31,119.25,112.40,80.26,54.93,49.60,48.13,46.04,34.20,30.96,29.43,25.37.

[0057] Example 8 4',4'-difluoro-7-(5-fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,1'-cyclohexyl]-4-one (Compound 8)

[0058] Compound 8 was prepared by referring to the method of compound 1; 1 H NMR (300MHz, DMSO-d6) δ9.86 (s, 1H), 8.70 (d, J = 3.4Hz, 1H), 8.02 (s, 1H), 8. 00(d,J=6.5Hz,1H),7.92(d,J=8.2Hz,1H),7.75(t,J=1.2Hz,1H),7.71(dt,J =8.2,1.4Hz,1H),7.41(dd,J=9.1,3.1Hz,1H),3.12(t,J=5.0Hz,4H),2.97(s ,2H),2.50-2.45(m,4H),2.24(s,3H),2.19-1.94(m,6H),1.86-1.72(m,2H). 13CNMR(126MHz,DMSO-d6)δ191.66,158.80,156.43,151.80,150.67,149.78,148.55,145.92,143.25,140.96, 135.95,126.99,125.51,122.09,121.91,119.25,113.72,78.94,54.97,48.93,46.83,46.26,30.79,30.72.

[0059] Example 9 7-(5-Fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,1'-cyclohexyl-4',2"-[1,3]-dioxolane]-4-one (Compound 9)

[0060] Compound 9 was prepared by referring to the method of compound 1; 1 H NMR (300MHz, CDCl3) δ8.47(d,J=3.3Hz,1H),8.27(d,J=9.0Hz,1H),8.25(s,1H),8.08(d ,J=2.9Hz,1H),8.02(d,J=8.6Hz,1H),7.77-7.72(m,2H),7.39(dd,J=9.1,3.0Hz,1H),4 .04-3.95(m,4H),3.25(t,J=5.0Hz,4H),2.80(s,2H),2.69(t,J=5.0Hz,4H),2.44(s,3H ),2.18(d,J=13.8Hz,2H),2.11-1.98(m,2H),1.87-1.74(m,2H),1.66(d,J=13.0Hz,2H). 13 C NMR (101MHz, CDCl3) δ191.88,159.19,155.67,155.64,152.04,150.35,150.26,149.49,147.90,147.63,146.21,142.92,140.90,140.85,136. 75,126.90,126.48,121.93,121.32,121.27,119.34,119.26,112.47,1 07.91,79.20,64.47,64.34,54.94,49.58,47.97,46.10,32.21,30.01.

[0061] Example 10 7-(5-Fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)-5",5"-dimethylspiro[benzopyran-2,1'-cyclohexyl-4',2"-[1,3]-dioxane]-4-one (Compound 10)

[0062] Compound 10 was prepared by referring to the method of compound 1; 1 H NMR(300MHz, CDCl3) δ8.47(d,J=3.3Hz,1H),8.26(d,J=9.1Hz,1H),8.21(s,1H),8.08 (d,J=3.0Hz,1H),8.01(d,J=8.6Hz,1H),7.76-7.72(m,2H),7.38(dd,J=9.1,3.0Hz,1H ),3.57(s,2H),3.50(s,2H),3.25(t,J=5.1Hz,4H),2.78(s,2H),2.69(t,J=5.0Hz,4H ),2.43(s,3H),2.16-2.03(m,4H),1.94-1.83(m,2H),1.79-1.66(m,2H),1.00(s,6H). 13 C NMR (101MHz, CDCl3) δ191.91,159.21,155.68,155.65,152.03,150.42,1 50.33,149.49,147.84,147.59,146.20,142.93,140.86,140.81,136.76 ,126.89,126.47,121.89,121.24,121.19,119.36,119.28,112.44,96.7 5,79.63,70.13,54.95,49.61,48.02,46.12,30.92,30.25,27.38,22.70.

[0063] Example 11 7-(5-Fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)-2',3',5',6'-tetrahydrospiro[benzopyran-2,4'-pyran]-4-one (Compound 11)

[0064] Compound 11 was prepared by referring to the method of compound 1; 1H NMR (300MHz, CDCl3) δ8.50-8.47(m,2H),8.26(d,J=9.1Hz,1H),8.11(d,J=2.4Hz,1H),8.02(dd,J=7.8,0.9Hz,1H),7.78-7.72(m,2H),7.37(dd,J= 9.1,3.0Hz,1H),3.97-3.78(m,4H),3.23(t,J=5.0Hz,4H),2.83(s,2H),2 .66(t,J=5.0Hz,4H),2.41(s,3H),2.09-2.00(m,2H),1.90-1.77(m,2H). 13 C NMR (101MHz, CDCl3) δ191.35,158.93,155.69,155.66,152.06,150.42,150.33,149.51,147.82,147.56,146.12,142.99,140.97,1 40.92,136.81,126.92,126.46,121.88,121.52,121.46,119.26,119.18,112.40,77.59,63.15,54.94,49.62,48.31,46.10,34.79.

[0065] Example 12 7-(5-Fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)-2',3',5',6'-tetrahydrospiro[benzopyran-2,4'-thiopyran]-4-one (Compound 12)

[0066] Compound 12 was prepared by referring to the method of compound 1; 1 H NMR(300MHz, CDCl3)δ8.49(d,J=3.3Hz,1H),8.41(s,1H),8.26(d,J=9.2Hz, 1H),8.10(d,J=2.3Hz,1H),8.01(d,J=8.7Hz,1H),7.77-7.73(m,2H),7.38( dd,J=9.1,3.0Hz,1H),3.24(t,J=5.0Hz,4H),3.20-3.07(m,2H),2.78(s,2H ),2.67(t,J=5.0Hz,4H),2.52-2.43(m,4H),2.42(s,3H),1.95-1.81(m,2H). 13CNMR(126MHz, CDCl3)δ191.28,158.65,155.62,151.93,150.46,150.39,149.89,147.80,147.59,146.14,142.98,140.98,1 37.00,126.93,126.51,121.87,121.55,121.50,119.24,119.18,112.45,78.52,54.88,49.50,48.89,45.93,35.66,23.51.

[0067] Example 13 Tert-butyl-7-(5-fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)-4-oxaspiro[benzopyran-2,4'-piperidine]-1'-carboxylate (Compound 13)

[0068] Compound 13 was prepared by referring to the method of compound 1; 1 H NMR (300MHz, CDCl3) δ8.47(d,J=3.2Hz,1H),8.28-8.23(m,2H),8.08(d,J=2.3Hz,1H),8.02(dd,J=7.9,0.8Hz,1H),7.77-7.72(m,2H),7.37(dd,J=9.1 ,3.0Hz,1H),3.92(s,2H),3.36-3.17(m,6H),2.80(s,2H),2.67(t,J=5.0Hz ,4H),2.42(s,3H),2.09(d,J=13.7Hz,2H),1.72-1.61(m,2H),1.49(s,9H). 13 C NMR (101MHz, CDCl3) δ191.26,158.86,155.68,155.65,154.72,152.05,150.39,150.31,149.51,147.81,147.55,146.10,142.99,140.99,1 40.94,136.75,126.96,126.47,121.86,121.55,121.50,119.22,119 .14,112.39,79.88,78.32,54.93,49.62,48.08,46.11,34.00,28.43.

[0069] Example 14 1'-Acetyl-7-(5-fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 14)

[0070] Compound 14 was prepared by referring to the method of compound 1; 1 H NMR(300MHz, CDCl3)δ8.48(d,J=3.3Hz,1H),8.29-8.22(m,2H),8.08(d,J=2.9Hz,1 H),8.03(d,J=8.1Hz,1H),7.80-7.74(m,2H),7.37(dd,J=9.1,3.0Hz,1H),4.45(d,J =13.7Hz,1H),3.73-3.52(m,2H),3.23(t,J=5.0Hz,4H),3.15(t,J=11.5Hz,1H),2.8 1(s,2H),2.68(t,J=5.0Hz,4H),2.42(s,3H),2.22-2.11(m,5H),1.76-1.60(m,2H). 13 C NMR (101MHz, CDCl3) δ190.93,168.97,158.67,155.73,155.70,152.02,1 50.29,150.21,149.47,147.85,147.59,146.13,142.99,141.11,141.06, 136.74,127.02,126.46,121.77,121.73,121.68,119.15,119.08,112.4 2,78.12,54.93,49.61,48.01,46.11,41.85,36.88,34.56,33.86,21.46.

[0071] Example 15 1'-Acetyl-7-(5-fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)-1'-(methylsulfonyl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 15)

[0072] Compound 15 was prepared by referring to the method of compound 1; 1H NMR(300MHz, CDCl3)δ8.48(d,J=3.2Hz,1H),8.31(s,1H),8.25(d,J=8.4Hz,1H), 8.08(d,J=2.3Hz,1H),8.04(d,J=8.2Hz,1H),7.80-7.73(m,2H),7.37(dd,J=9.1 ,3.0Hz,1H),3.69(d,J=11.9Hz,2H),3.26-3.12(m,6H),2.87(s,3H),2.83(s,2H ),2.66(t,J=4.9Hz,4H),2.42(s,3H),2.24(d,J=13.9Hz,2H),1.93-1.82(m,2H). 13 C NMR (101MHz, CDCl3) δ190.63,158.41,155.71,155.68,152.06,150.32,150.23,149.51,147.78,147.52,145.99,143.08,141.09,141. 04,136.67,127.14,126.42,121.91,121.85,121.82,119.01,118.94,112.40,77.21,54.94,49.60,47.93,46.14,41.35,35.12,33.84.

[0073] Example 16 7-(5-Fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)-1'-propionylspiro[benzopyran-2,4'-piperidin]-4-one (Compound 16)

[0074] Compound 16 was prepared by referring to the method of compound 1; 1 H NMR (300MHz, CDCl3) δ8.48 (d, J = 3.3Hz, 1H), 8.31 (s, 1H), 8.25 (d, J = 9.1Hz, 1H), 8.08 (d, J = 2. 9Hz,1H),8.03(d,J=8.1Hz,1H),7.79-7.74(m,2H),7.37(dd,J=9.1,3.0Hz,1H),4.46(d,J=13. 6Hz,1H),3.71(d,J=13.8Hz,1H),3.54(t,J=12.6Hz,1H),3.26-3.10(m,5H),2.81(s,2H),2.69 -2.62(m,4H),2.45-2.34(m,5H),2.21-2.09(m,2H),1.73-1.59(m,2H),1.18(t,J=7.4Hz,3H).13 C NMR (101MHz, CDCl3) δ190.99,172.29,158.71,155.71,155.68,152.03,15 0.31,150.22,149.48,147.85,147.59,146.12,142.98,141.09,141.04,13 6.75,127.01,126.47,121.78,121.71,121.65,119.16,119.09,112.41,78 .24,54.92,49.60,48.03,46.10,40.89,37.04,34.59,33.96,26.54,9.54.

[0075] Example 17 1'-Butyryl-7-(5-fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 17)

[0076] Compound 17 was prepared by referring to the method of compound 1; 1 H NMR (300MHz, CDCl3) δ8.47(d,J=3.3Hz,1H),8.24(d,J=9.1Hz,1H),8.18(s,1H),8.07(d,J=2.3Hz, 1H),8.03(d,J=8.7Hz,1H),7.80-7.74(m,2H),7.37(dd,J=9.1,3.0Hz,1H),4.46(d,J=13.4Hz,1H) ,3.72(d,J=13.6Hz,1H),3.55(t,J=12.7Hz,1H),3.27-3.09(m,5H),2.81(s,2H),2.67(t,J=5.0Hz ,4H),2.42(s,3H),2.38-2.30(m,2H),2.18-2.10(m,2H),1.74-1.62(m,4H),1.00(t,J=7.4Hz,3H). 13C NMR (101MHz, CDCl3) δ190.97,171.51,158.71,155.70,155.67,152.05,150. 33,150.24,149.51,147.84,147.58,146.10,142.99,141.08,141.02,136.80 ,127.02,126.45,121.80,121.71,121.65,119.16,119.09,112.39,78.25,54.92,49.59,48.04,46.08,41.07,36.96,35.29,34.70,33.98,18.80,14.04.

[0077] Example 18 1'-(Cyclopropylacyl)-7-(5-fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 18)

[0078] Compound 18 was prepared by referring to the method of compound 1; 1 H NMR (300MHz, CDCl3) δ8.47(d,J=3.2Hz,1H),8.26(d,J=9.7Hz,1H),8.19(s,1H),8.07(d,J=2.3H z,1H),8.04(d,J=8.8Hz,1H),7.79-7.75(m,2H),7.37(dd,J=9.1,3.0Hz,1H),4.43(d,J=13.2Hz, 1H),4.06(d,J=13.5Hz,1H),3.67(d,J=13.1Hz,1H),3.33-3.12(m,5H),2.83(s,2H),2.77-2.66( m,4H),2.46(s,3H),2.20-2.09(m,2H),1.83-1.63(m,3H),1.05-0.98(m,2H),0.83-0.75(m,2H). 13CNMR(101MHz,CDCl3)δ191.02,172.00,158.75,155.72,155.69,152.04,15 0.33,150.24,149.50,147.85,147.59,146.10,143.01,141.08,141.03,13 6.76,127.01,126.44,121.81,121.69,121.63,119.19,119.11,112.40,78 .41,54.94,49.62,48.07,46.12,41.02,37.66,34.74,33.96,10.98,7.47.

[0079] Example 19 7-(5-fluoro-2-((5-(4-methylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)-1'-(2,2,2-trifluoroacetyl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 19)

[0080]

[0081] Compound 19 was prepared by referring to the method of compound 1; 1 H NMR (300MHz, CDCl3) δ8.48(d,J=3.3Hz,1H),8.24(d,J=9.1Hz,1H),8.20(s,1H),8.08(d ,J=2.4Hz,1H),8.05(d,J=8.3Hz,1H),7.83–7.76(m,2H),7.37(dd,J=9.1,3.0Hz,1H),4. 45(d,J=13.4Hz,1H),3.92(d,J=14.1Hz,1H),3.67(t,J=13.2Hz,1H),3.39-3.20(m,5H), 2.84(s,2H),2.75-2.66(m,4H),2.44(s,3H),2.26(d,J=14.1Hz,2H),1.83-1.68(m,2H). 13C NMR (126MHz, CDCl3) δ190.33,158.40,155.71,151.90,150.17,149.86,147.85,147.64,145.99,143.11,141.24,141.20,136.86,127. 19,126.39,122.07,122.02,121.78,119.07,119.02,117.65,112.42,77.67,54.94,49.59,47.94,46.07,41.22,39.02,34.66,33.69.

[0082] Example 20 1'-Acetyl-7-(5-fluoro-2-((5-(piperidin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 20)

[0083] In a 25 ml sealed tube, V-14 (220 mg, 0.56 mmol), 5-(piperidin-1-yl)pyridine-2-amino (120 mg, 0.672 mmol), Pd2(dba)3 (52 mg, 0.056 mmol), BINAP (70 mg, 0.112 mmol), Cs2CO3 (365 mg, 1.12 mmol) and dioxane (8 mL) were added in sequence. Under N2 atmosphere, the mixture was stirred at 100 °C for 12 h. The reaction was completed by TLC. After the reaction solution was cooled to room temperature, it was concentrated under reduced pressure to remove the solvent. The residue was purified by silica gel column chromatography (DCM / CH3OH, 40:1-10:1) to obtain yellow solid compound 19 (139 mg, 46.7%). 1 H NMR (300MHz, CDCl3) δ8.47(d,J=3.2Hz,1H),8.33(s,1H),8.24(d,J=8.1Hz,1H),8 .06(d,J=2.8Hz,1H),8.03(dd,J=8.1,0.6Hz,1H),7.80-7.74(m,2H),7.40(dd,J= 9.1,3.0Hz,1H),4.45(d,J=13.5Hz,1H),3.75-3.52(m,2H),3.21-3.09(m,5H),2. 81(s,2H),2.22-2.15(m,2H),2.14(s,3H),1.81-1.72(m,4H),1.71-1.56(m,4H). 13C NMR (101MHz, CDCl3) δ190.92,168.93,158.67,155.76,155.73,152.00,152.24,152.15,149.45,147.87,147.61,145.66,144.01,141.15,141 .09,136.89,127.01,126.97,121.75,121.69,119.16,119.08,112.43, 78.12,51.23,48.03,41.85,36.88,34.55,33.88,25.76,24.01,21.46.

[0084] Example 21 1'-Acetyl-7-(5-fluoro-2-((5-(4-methylpiperidin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 21)

[0085] Compound 21 was prepared by the method of Compound 20, using 5-(4-methylpiperidin-1-yl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino; 1 H NMR (300MHz, CDCl3) δ8.47(d,J=3.3Hz,1H),8.32(s,1H),8.25(d,J=9.1Hz,1H),8.05(s,1H),8.03(d,J= 8.1Hz,1H),7.80-7.75(m,2H),7.42(dd,J=9.1,3.0Hz,1H),4.45(d,J=13.5Hz,1H),3.74-3.51(m,4H),3 .22-3.08(m,1H),2.81(s,2H),2.74(td,J=11.9,2.5Hz,2H),2.22-2.16(m,2H),2.14(s,3H),1.80(d,J= 12.7Hz,2H),1.73-1.60(m,2H),1.58-1.49(m,1H),1.43(td,J=12.1,3.8Hz,2H),1.02(d,J=6.2Hz,3H). 13C NMR (101MHz, CDCl3) δ190.94,168.98,158.66,155.74,155.72,152.00,1 50.24,150.15,149.45,147.86,147.60,145.59,143.72,141.14,141.09 ,136.88,127.01,126.92,121.75,121.69,119.16,119.08,112.46,78.1 1,50.56,48.01,41.86,36.89,34.54,33.98,33.88,30.45,21.89,21.45.

[0086] Example 22 1'-Acetyl-7-(5-fluoro-2-((5-(4-methoxypiperidin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 22)

[0087] Compound 22 was prepared by the method of Compound 20, using 5-(4-methoxypiperidin-1-yl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino; 1 H NMR (300MHz, CDCl3) δ8.47(d,J=3.2Hz,1H),8.34(s,1H),8.27(d,J=9.1Hz,1H),8.05(s,1H), 8.04-8.02(m,1H),7.80-7.75(m,2H),7.44(dd,J=9.1,2.3Hz,1H),4.45(d,J=13.6Hz,1H),3.7 4-3.52(m,2H),3.51-3.42(m,3H),3.41(s,3H),3.22-3.09(m,1H),3.02-2.94(m,2H),2.81(s, 2H),2.23-2.15(m,2H),2.15(s,3H),2.07-2.04(m,1H),1.85-1.72(m,3H),1.71-1.61(m,2H). 13C NMR (101MHz, CDCl3) δ190.92,168.94,158.67,155.72,155.70,152.02,150 .27,150.19,149.47,147.87,147.60,145.86,143.21,141.11,141.06,137 .04,127.02,126.96,121.77,121.74,121.68,119.16,119.08,112.40,78.13,75.55,55.67,48.01,47.75,41.85,36.88,34.56,33.87,30.50,21.46.

[0088] Example 23 Ethyl 1-(6-((4-(1'-acetyl-4-oxospiro[benzopyran-2,4'-piperidin]-7-one)-5-fluoropyrimidin-2-yl)amino)pyridin-3-yl)piperidine-4-carboxylate (Compound 23)

[0089] Compound 23 was prepared by referring to the method of Compound 20, using 1-(6-aminopyridin-3-yl)piperidine-4-carboxylic acid ethyl ester to replace 5-(piperidin-1-yl)pyridin-2-amino; 1 H NMR (300MHz, CDCl3) δ8.48(d,J=3.2Hz,1H),8.40(s,1H),8.26(d,J=9.1Hz,1H),8.06(d,J=2.9Hz,1H), 8.03(d,J=8.2Hz,1H),7.81–7.73(m,2H),7.40(dd,J=9.1,3.0Hz,1H),4.45(d,J=13.6Hz,1H),4.19(q,J =7.1Hz,2H),3.73-3.53(m,4H),3.15(t,J=12.3Hz,1H),2.88-2.77(m,4H),2.51-2.41(m,1H),2.22-2. 15(m,2H),2.14(s,3H),2.08-2.04(m,1H),2.00-1.85(m,3H),1.74-1.61(m,2H),1.30(t,J=7.1Hz,3H). 13C NMR (101MHz, CDCl3) δ190.91,174.70,168.94,158.67,155.69,155.66,152.05,15 0.31,150.22,149.50,147.86,147.60,146.00,143.32,141.08,141.03,137.04,1 27.19,127.03,121.78,121.73,121.68,119.16,119.09,112.43,78.14,60.55,51.83,49.81,48.01,41.85,40.70,40.58,36.88,34.57,33.87,28.00,21.46,14.26.

[0090] Example 24 1'-Acetyl-7-(5-fluoro-2-((5-(4-dimethylaminopiperidin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 24)

[0091] Compound 24 was prepared by the method of Compound 20, using 5-(4-dimethylaminopiperidin-1-yl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino; 1 H NMR (300MHz, CDCl3) δ8.47(d,J=3.3Hz,1H),8.23(d,J=9.1Hz,1H),8.07(s,1H),8.06-8.02( m,2H),7.80-7.74(m,2H),7.38(dd,J=9.1,3.0Hz,1H),4.45(d,J=13.4Hz,1H),3.67(d,J=11 .2Hz,3H),3.64-3.51(m,1H),3.15(t,J=12.2Hz,1H),2.81(s,2H),2.75(dd,J=12.2,2.4Hz, 2H),2.40(s,6H),2.16-2.21(m,2H),2.15(s,3H),2.03(d,J=12.4Hz,2H),1.61-1.80(m,5H). 13C NMR (101MHz, CDCl3) δ190.93,168.97,158.68,155.69,155.66,152.05,150 .29,150.20,149.50,147.85,147.59,145.82,143.25,141.10,141.05,137 .11,127.03,126.88,121.78,121.74,121.68,119.15,119.08,112.36,78.13,61.86,49.80,48.02,41.86,41.64,36.89,34.56,33.88,28.17,21.46.

[0092] Example 25 1'-acetyl-7-(5-fluoro-2-((5-(4-morpholinopiperidin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 25)

[0093] Compound 25 was prepared by the method of Compound 20, using 5-(4-morpholinopiperidin-1-yl)pyridin-2-amino instead of 5-(piperidin-1-yl)pyridin-2-amino; 1 H NMR (300MHz, CDCl3) δ8.47(d,J=3.3Hz,1H),8.23(d,J=9.1Hz,1H),8.10(s,1H),8.07-8. 01(m,2H),7.80-7.74(m,2H),7.37(dd,J=9.1,3.0Hz,1H),4.45(d,J=13.5Hz,1H),3.82( s,4H),3.68(d,J=12.3Hz,3H),3.64-3.52(m,1H),3.15(t,J=12.3Hz,1H),2.81(s,2H),2 .81-2.59(m,6H),2.25-2.15(m,2H),2.15(s,3H),2.09-2.00(m,2H),1.78-1.64(m,5H). 13C NMR (101MHz, CDCl3) δ190.92,168.96,158.67,155.72,155.69,152.02,150. 27,150.18,149.47,147.87,147.61,145.88,143.21,141.11,141.05,137.07 ,127.02,126.84,121.77,121.73,121.67,119.16,119.08,112.38,78.13,67.22,61.74,49.79,49.76,48.01,41.85,36.88,34.55,33.87,27.99,21.46.

[0094] Example 26 1'-Acetyl-7-(5-fluoro-2-((5-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 26)

[0095] Compound 26 was prepared by referring to the method of Compound 20, using 5-(4-(4-methylpiperazin-1-yl)piperidin-1-yl)-pyridine-2-amino to replace 5-(piperidin-1-yl)pyridine-2-amino; 1 H NMR (300MHz, CDCl3) δ8.47(d,J=3.3Hz,1H),8.25-8.19(m,2H),8.07(d,J=2.9Hz,1H),8.03(d,J=8 .1Hz,1H),7.80-7.73(m,2H),7.36(dd,J=9.1,3.0Hz,1H),4.44(d,J=13.5Hz,1H),3.67(d,J=12.0H z,3H),3.63-3.51(m,1H),3.15(t,J=12.0Hz,1H),2.81(s,2H),2.80-2.64(m,6H),2.54(s,4H),2. 43-2.37(m,1H),2.34(s,3H),2.19(s,2H),2.14(s,3H),1.99(d,J=12.4Hz,2H),1.78-1.60(m,4H). 13C NMR (101MHz, CDCl3) δ190.91,168.94,158.66,155.73,155.70,152.01,150.26 ,150.17,149.46,147.86,147.60,145.79,143.18,141.11,141.06,137.03,12 7.01,126.77,121.76,121.73,121.67,119.15,119.07,112.36,78.12,61.46,55.42,49.90,49.03,48.01,46.02,41.85,36.88,34.55,33.87,28.09,21.46.

[0096] Example 27 tert-Butyl 4-(6-((4-(1'-acetyl-4-oxospiro[benzopyran-2,4'-piperidin]-7-one)-5-fluoropyrimidin-2-yl)amino)pyridin-3-yl)piperidine-1-carboxylate (Compound 27)

[0097] Compound 27 was prepared by referring to the method of Compound 20, using 1-(6-aminopyridin-3-yl)piperidine-4-carboxylic acid tert-butyl ester to replace 5-(piperidin-1-yl)pyridin-2-amino; 1 H NMR (300MHz, CDCl3) δ8.57(s,1H),8.52(d,J=3.2Hz,1H),8.36(d,J=8.6Hz,1H),8.21(d,J=2 .3Hz,1H),8.04(dd,J=8.1,0.6Hz,1H),7.80-7.75(m,2H),7.62(dd,J=8.8,2.4Hz,1H),4.45 (d,J=13.5Hz,1H),4.27(s,2H),3.75-3.52(m,2H),3.21-3.08(m,1H),2.92-2.80(m,4H),2. 75-2.64(m,1H),2.22-2.12(m,5H),1.86(d,J=13.0Hz,2H),1.75-1.62(m,4H),1.51(s,9H). 13C NMR (101MHz, CDCl3) δ190.89,168.97,158.69,155.50,155.47,154.83,152 .30,151.19,150.48,150.39,149.74,147.89,147.63,146.40,140.87,140. 82,136.53,134.98,127.08,121.87,121.73,121.68,1119.18,119.11,111.85,79.62,78.18,48.01,41.85,39.64,36.89,34.60,33.84,28.50,21.46.

[0098] Example 28 1'-Acetyl-7-(5-fluoro-2-((5-(piperidin-4-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 28)

[0099] Using 5-(piperidin-4-yl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino, compound 28 was prepared according to the method of compound 20; 1 H NMR (300MHz, DMSO-d6) δ10.12(s,1H),9.20(s,1H),8.76(d,J=3.3Hz,1H),8.20(d,J=2.3Hz,1H) ,8.15(d,J=8.6Hz,1H),7.95-7.90(m,1H),7.75-7.69(m,2H),7.65(dd,J=8.7,2.5Hz,1H),4.12( d,J=13.3Hz,1H),3.73-3.62(m,1H),3.46-3.38(m,2H),3.32(brs,2H),3.07-2.94(m,3H),2.94( s,2H),2.90-2.81(m,1H),2.02(s,3H),1.98-1.86(m,5H),1.84-1.72(m,1H),1.69-1.55(m,1H). 13C NMR(101MHz,DMSO-d6)δ191.73,168.76,158.93,156.13,152.09,152.07,150.00,149.93,148.81,148.59,146.64,140.73,140.68,13 6.33,134.27,126.99,122.15,121.71,119.27,119.22,112.76,79.18,47.30,43.87,41.83,36.92,36.48,34.28,33.48,29.52,21.79.

[0100] Example 29 1'-Acetyl-7-(5-fluoro-2-((5-(1-methylpiperidin-4-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 29)

[0101] Compound 29 was prepared by the method of Compound 20, using 5-(1-methylpiperidin-4-yl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino; 1 H NMR (300MHz, CDCl3) δ8.51(d,J=3.2Hz,1H),8.38(s,1H),8.32(d,J=8.2Hz,1H),8.25(d,J=2. 4Hz,1H),8.04(dd,J=8.0,0.7Hz,1H),7.81-7.74(m,2H),7.63(dd,J=8.6,2.5Hz,1H),4.45(d, J=13.6Hz,1H),3.76-3.50(m,2H),3.22-3.02(m,3H),2.82(s,2H),2.59-2.48(m,1H),2.40(s, 3H),2.22-2.16(m,2H),2.15(s,3H),2.13-2.09(m,1H),1.95-1.83(m,5H),1.76-1.60(m,2H). 13C NMR (101MHz, CDCl3) δ190.90,168.98,158.68,155.66,155.63,152.19,151 .23,150.39,150.30,149.64,147.90,147.64,146.81,140.98,140.92,136 .39,135.35,127.06,121.81,121.76,121.70,119.15,119.08,111.81,78.15,56.17,48.01,46.42,41.86,38.90,36.89,34.58,33.85,33.32,21.45.

[0102] Example 30 1'-Acetyl-7-(5-fluoro-2-((5-morpholinopyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 30)

[0103] Using 5-morpholinopyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino, compound 30 was prepared according to the method of compound 20; 1 H NMR (300MHz, CDCl3) δ8.48(d,J=3.2Hz,1H),8.43(s,1H),8.31(dd,J=9.1,0.7Hz,1H),8.07-8.00(m,2H),7.80-7.75(m,2H),7.39(dd,J=9.2,3 .0Hz,1H),4.44(d,J=13.7Hz,1H),3.95-3.87(m,4H),3.74-3.51(m,2H) ,3.22-3.07(m,5H),2.82(s,2H),2.22-2.10(m,5H),1.76-1.60(m,2H). 13 C NMR (125MHz, CDCl3) δ190.86,168.97,158.73,155.55,151.98,150.47,150.39,149.93,147.79,147.58,146.14,143.09,141.00,140.96, 136.91,127.08,126.46,121.90,121.77,121.72,119.18,119.12,11 2.59,78.20,66.80,49.86,48.06,41.89,36.93,34.64,33.92,21.43.

[0104] Example 31 tert-Butyl 4-(6-((4-(1'-acetyl-4-oxospiro[benzopyran-2,4'-piperidin]-7-one)-5-fluoropyrimidin-2-yl)amino)pyridin-3-yl)piperazine-1-carboxylate (Compound 31)

[0105] Compound 31 was prepared by referring to the method of Compound 20, using 4-(6-aminopyridin-3-yl)piperazine-1-carboxylic acid tert-butyl ester to replace 5-(piperidin-1-yl)pyridin-2-amino; 1 H NMR (300MHz, CDCl3) δ8.48(d,J=3.2Hz,1H),8.39(s,1H),8.31(d,J=9.1Hz,1H),8.07-8.00(m,2H),7.81-7.74(m,2H),7.41(dd,J=9.2,3 .0Hz,1H),4.44(d,J=13.7Hz,1H),3.71-3.55(m,6H),3.21-3.07(m,5H),2.82(s,2H),2.22-2.10(m,5H),1.74-1.64(m,2H),1.51(s,9H). 13 C NMR (101MHz, CDCl3) δ190.92,168.99,158.68,155.63,155.60,154.66,152 .09,150.36,150.27,149.54,147.87,147.61,146.55,142.96,141.01,140 .96,137.08,127.33,127.04,121.80,121.72,121.67,119.16,119.09,112.47,80.09,78.16,49.99,47.99,41.85,36.89,34.57,33.84,28.44,21.45.

[0106] Example 32 1'-Acetyl-7-(5-fluoro-2-((5-(piperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 32)

[0107] Using 5-(piperazin-1-yl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino, compound 32 was prepared according to the method of compound 20; 1H NMR (400MHz, CDCl3) δ8.47(d,J=3.2Hz,1H),8.26-8.22(m,1H),8.09(s,1H),8.06-8.01(m,2H),7.80-7.74(m,2H),7.36(dd,J=9.1,3.0Hz ,1H),4.45(d,J=13.3Hz,1H),3.72-3.54(m,2H),3.23-3.12(m,5H),3.11-3.07(m,4H),2.82(s,2H),2.20-2.13(m,5H),1.70-1.63(m,5H). 13 C NMR (126MHz, DMSO-d6) δ191.72,168.71,158.94,156.38,151.83,149.88,149.81,148.74,148.54,146.33,143.11,140.90,140.85 ,136.33,126.94,126.01,122.12,121.72,119.23,119.18,113.64,79.18,48.50,47.31,44.65,41.82,36.91,34.28,33.50,21.78.

[0108] Example 33 1'-Acetyl-7-(5-fluoro-2-((5-(4-ethylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 33)

[0109] Using 5-(4-ethylpiperazin-1-yl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino, compound 33 was prepared according to the method of compound 20; 1 H NMR(300MHz, CDCl3)δ8.48(d,J=3.3Hz,1H),8.28-8.22(m,2H),8.10-8.07(m,1H),8.03( d,J=8.3Hz,1H),7.80-7.74(m,2H),7.37(dd,J=9.1,3.0Hz,1H),4.45(d,J=13.3Hz,1H),3 .74-3.52(m,2H),3.26(t,J=4.9Hz,4H),3.21-3.09(m,1H),2.81(s,2H),2.73(t,J=4.5Hz ,4H),2.58(q,J=7.2Hz,2H),2.21-2.12(m,5H),1.75-1.61(m,2H),1.20(t,J=7.2Hz,3H). 13C NMR (101MHz, CDCl3) δ190.92,168.94,158.67,155.75,155.72,152.01,1 50.28,150.19,149.46,147.87,147.60,146.12,143.05,141.13,141.07 ,136.74,127.02,126.39,121.76,121.68,119.15,119.08,112.41,78.1 3,52.66,52.37,49.66,48.02,41.85,36.88,34.56,33.87,21.46,11.97.

[0110] Example 34 1'-Acetyl-7-(5-fluoro-2-((5-(4-isopropylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 34)

[0111] Using 5-(4-isopropylpiperazin-1-yl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino, compound 34 was prepared according to the method of compound 20; 1 H NMR (300MHz, CDCl3) δ8.47(d,J=3.2Hz,1H),8.24(d,J=8.8Hz,1H),8.21(s,1H),8.07(d, J=2.3Hz,1H),8.03(d,J=8.2Hz,1H),7.81-7.73(m,2H),7.37(dd,J=9.1,3.0Hz,1H),4.4 5(d,J=13.6Hz,1H),3.75-3.51(m,2H),3.23(t,J=4.9Hz,4H),3.20-3.10(m,1H),2.81(s ,2H),2.77(t,J=5.1Hz,4H),2.21-2.10(m,5H),1.77-1.61(m,3H),1.15(d,J=6.5Hz,6H). 13C NMR (101MHz, CDCl3) δ190.93,168.96,158.67,155.74,155.71,152.01,1 50.27,150.18,149.46,147.86,147.60,146.05,143.13,141.13,141.08 ,136.72,127.01,126.37,121.75,121.69,119.15,119.07,112.41,78.1 2,54.59,50.00,48.57,48.02,41.86,36.89,34.56,33.87,21.46,18.57.

[0112] Example 35 1'-acetyl-7-(5-fluoro-2-((5-(4-(2-alkoxyethyl)piperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 35)

[0113] Compound 35 was prepared by the method of Compound 20, using 5-(4-(2-alkoxyethyl)piperazin-1-yl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino; 1 H NMR(300MHz, CDCl3) δ8.48(d,J=3.3Hz,1H),8.38(s,1H),8.24(d,J=9.2Hz,1H),8.08 (d,J=2.3Hz,1H),8.03(d,J=8.3Hz,1H),7.80-7.73(m,2H),7.36(dd,J=9.1,3.0Hz,1H ),4.44(d,J=13.4Hz,1H),3.73-3.52(m,4H),3.40(s,3H),3.24(t,J=5.0Hz,4H),3.2 1-3.08(m,1H),2.81(s,2H),2.76-2.67(m,6H),2.21-2.10(m,5H),1.75-1.61(m,2H). 13C NMR (101MHz, CDCl3) δ190.94,168.97,158.67,155.73,155.70,152.02,15 0.27,150.18,149.47,147.87,147.61,146.08,143.06,141.12,141.07,13 6.74,127.02,126.40,121.76,121.68,119.15,119.07,112.41,78.12,70. 05,58.98,57.94,53.41,49.51,48.01,41.85,36.89,34.56,33.86,21.45.

[0114] Example 36 1'-Acetyl-7-(5-fluoro-2-((5-(4-cyclopropylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 36)

[0115] Using 5-(4-cyclopropylpiperazin-1-yl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino, compound 36 was prepared according to the method of compound 20; 1 H NMR (300MHz, CDCl3) δ8.48(d,J=3.3Hz,1H),8.43(s,1H),8.24(d,J=9.3Hz,1H),8.09(d,J=2 .3Hz,1H),8.03(d,J=7.9Hz,1H),7.80-7.73(m,2H),7.36(dd,J=9.1,3.0Hz,1H),4.44(d,J= 13.6Hz,1H),3.74-3.52(m,2H),3.17(t,J=5.2Hz,4H),3.12-3.10(m,1H),2.85(t,J=5.0Hz, 4H), 2.81 (s, 2H), 2.19 (s, 1H), 2.17-2.10 (m, 5H), 1.74-1.65 (m, 2H), 0.54 (d, J = 6.2Hz, 4H). 13C NMR (101MHz, CDCl3) δ190.92,168.95,158.67,155.73,155.70,152.02,15 0.29,150.20,149.48,147.85,147.59,146.06,143.17,141.11,141.06,13 6.81,127.03,126.48,121.77,121.74,121.68,119.15,119.08,112.39,78 .13,53.15,49.67,48.02,41.86,38.52,36.88,34.56,33.87,21.46,5.81.

[0116] Example 37 1'-Acetyl-7-(5-fluoro-2-((5-(4-cyclohexylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 37)

[0117] Using 5-(4-cyclohexylpiperazin-1-yl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino, compound 37 was prepared according to the method of compound 20; 1 H NMR (300MHz, CDCl3) δ8.47(d,J=3.3Hz,1H),8.24(d,J=9.1Hz,1H),8.09(s,1H),8.06(d,J=2.9Hz ,1H),8.03(dd,J=8.1,0.6Hz,1H),7.80-7.74(m,2H),7.37(dd,J=9.1,3.0Hz,1H),4.45(d,J=13.5 Hz,1H),3.75-3.51(m,2H),3.29(brs,4H),3.15(t,J=12.2Hz,1H),2.89(brs,4H),2.81(s,2H),2 .12-2.10(m,5H),2.03(brs,2H),1.88(brs,2H),1.75-1.61(m,5H),1.38-1.22(d,J=13.3Hz,4H). 13C NMR (101MHz, CDCl3) δ190.94,168.98,158.67,155.70,155.67,152.04,150. 28,150.19,149.49,147.85,147.59,145.96,143.17,141.11,141.06,136.6 8,127.02,126.34,121.77,121.69,119.14,119.07,112.40,78.12,63.59,50.09,48.84,48.02,41.86,36.89,34.55,33.88,28.91,26.27,25.84,21.45.

[0118] Example 38 1'-Acetyl-7-(5-fluoro-2-((5-(4-(oxetane-3-yl)piperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 38)

[0119] Compound 38 was prepared by the method of Compound 20, using 5-(4-(oxetane-3-yl)piperazin-1-yl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino; 1 H NMR (300MHz, CDCl3) δ8.48(d,J=3.2Hz,1H),8.42(s,1H),8.28(d,J=9.3Hz,1H),8.07(d, J=2.9Hz,1H),8.03(dd,J=8.1,0.6Hz,1H),7.80-7.74(m,2H),7.38(dd,J=9.1,3.0Hz,1H) ,4.79-4.66(m,4H),4.44(d,J=13.4Hz,1H),3.75-3.51(m,3H),3.25(t,J=5.0Hz,4H),3.2 0-3.10(m,1H),2.81(s,2H),2.58(t,J=5.0Hz,4H),2.20-2.12(m,5H),1.76-1.60(m,2H). 13C NMR (101MHz, CDCl3) δ190.91,168.96,158.68,155.65,155.63,152.09,150 .34,150.25,149.54,147.85,147.59,146.25,142.89,141.05,141.00,136 .77,127.04,126.67,121.80,121.73,121.67,119.16,119.09,112.44,78.15,75.36,59.19,49.50,49.42,48.02,41.85,36.88,34.57,33.86,21.46.

[0120] Example 39 1'-acetyl-7-(5-fluoro-2-((5-(4-cyclopropylmethylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 39)

[0121] Using 5-(4-cyclopropylmethylpiperazin-1-yl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino, compound 39 was prepared according to the method of compound 20; 1 H NMR(300MHz, CDCl3)δ8.48(d,J=3.3Hz,1H),8.29-8.22(m,2H),8.08(dd,J=3.0,0.7Hz,1H),8.03 (dd,J=8.1,0.6Hz,1H),7.80-7.74(m,2H),7.37(dd,J=9.1,3.0Hz,1H),4.45(d,J=13.6Hz,1H),3. 74-3.53(m,2H),3.26(t,J=5.0Hz,4H),3.22-3.08(m,1H),2.83-2.74(m,6H),2.40(d,J=6.6Hz,2H ),2.21-2.13(m,5H),1.74-1.62(m,2H),1.02-0.91(m,1H),0.64-0.56(m,2H),0.23-0.16(m,2H). 13C NMR (101MHz, CDCl3) δ190.93,168.97,158.67,155.74,155.71,152.02,15 0.28,150.19,149.47,147.87,147.61,146.09,143.10,141.12,141.07,1 36.74,127.02,126.42,121.76,121.69,119.15,119.07,112.42,78.12,6 3.70,53.05,49.61,48.01,41.86,36.89,34.56,33.87,21.45,8.25,3.98.

[0122] Example 40 1'-Acetyl-7-(5-fluoro-2-((5-(4-acetylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 40)

[0123] Compound 40 was prepared by replacing 5-(piperidin-1-yl)pyridin-2-amino with 5-(4-acetylpiperazin-1-yl)pyridin-2-amino; 1 H NMR (300MHz, CDCl3) δ8.49(d,J=3.2Hz,1H),8.46(s,1H),8.32(d,J=9.1Hz,1H ),8.07-8.01(m,2H),7.79-7.75(m,2H),7.41(dd,J=9.2,3.0Hz,1H),4.44(d, J=13.7Hz,1H),3.86-3.79(m,2H),3.72-3.63(m,3H),3.65-3.52(m,1H),3.20 -3.11(m,5H),2.82(s,2H),2.22-2.16(m,5H),2.14(s,3H),1.76-1.61(m,2H). 13C NMR (101MHz, CDCl3) δ190.92,169.06,169.00,158.68,155.59,155.56,152.14 ,150.41,150.32,149.58,147.85,147.59,146.80,142.62,140.98,140.93,13 7.24,127.41,127.05,121.82,121.72,121.66,119.18,119.11,112.48,78.18,50.35,49.96,47.99,46.16,41.84,41.26,36.88,34.59,33.82,21.46,21.37.

[0124] Example 41 1'-Acetyl-7-(5-fluoro-2-((5-(4-trifluorocarbonylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 41)

[0125] Using 5-(4-trifluoroformylpiperazin-1-yl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino, compound 41 was prepared according to the method of compound 20; 1 H NMR (300MHz, CDCl3) δ8.59(s,1H),8.50(d,J=3.2Hz,1H),8.37(d,J=9.1Hz,1H),8 .07-8.01(m,2H),7.80-7.74(m,2H),7.43(dd,J=9.2,2.9Hz,1H),4.44(d,J=13.6H z,1H),3.90(t,J=5.1Hz,2H),3.83(t,J=5.0Hz,2H),3.74-3.52(m,2H),3.23(t,J= 5.1Hz,4H),3.19-3.08(m,1H),2.82(s,2H),2.22-2.12(m,5H),1.76-1.61(m,2H). 13C NMR (101MHz, CDCl3) δ190.88,168.98,158.69,155.71,155.51,155.48,155.35,1 52.20,150.48,150.39,149.65,147.86,147.59,147.19,142.11,140.90,140.85 ,137.43,127.75,127.07,121.86,121.71,121.65,119.20,119.12,117.83,112.47,78.20,50.38,49.88,47.99,45.69,43.17,41.84,36.88,34.62,33.80,21.47.

[0126] Example 42 1'-Acetyl-7-(5-fluoro-2-((5-(4-cyclopropanecarbonylpiperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 42)

[0127] Using 5-(4-trifluorocarbonylpiperazin-1-yl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino, compound 42 was prepared according to the method of compound 20; 1 H NMR (300MHz, CDCl3) δ8.50 (s, 1H), 8.49 (s, 1H), 8.33 (d, J = 9.1Hz, 1H), 8.07-8.0 1(m,2H),7.81-7.74(m,2H),7.43(dd,J=9.2,3.0Hz,1H),4.44(d,J=13.6Hz,1H), 3.87(brs,4H),3.73-3.53(m,2H),3.27-3.09(m,5H),2.82(s,2H),2.22-2.10(m, 5H),1.84-1.77(m,1H),1.75-1.64(m,2H),1.09-1.01(m,2H),0.89-0.80(m,2H). 13C NMR (101MHz, CDCl3) δ190.93,172.16,169.02,158.68,155.61,155.58,152.12 ,150.38,150.29,149.56,147.87,147.61,146.70,142.69,141.00,140.95,13 7.09,127.30,127.05,121.81,121.72,121.67,119.18,119.10,112.50,78.17,50.42,49.92,47.99,45.38,41.85,36.89,34.58,33.82,21.46,10.98,7.64.

[0128] Example 43 4-(6-((4-(1'-acetyl-4-oxospiro[benzopyran-2,4'-piperidin]-7-one)-5-fluoropyrimidin-2-yl)amino)pyridin-3-yl)piperazine-1-carboxylic acid ethyl ester (Compound 43)

[0129] Compound 43 was prepared by the method of Compound 20, using 4-(6-aminopyridin-3-yl)piperazine-1-carboxylic acid ethyl ester to replace 5-(piperidin-1-yl)pyridin-2-amino; 1 H NMR (300MHz, CDCl3) δ8.49(d,J=3.2Hz,1H),8.43(s,1H),8.30(d,J=9.1Hz,1 H),8.08-8.01(m,2H),7.80-7.74(m,2H),7.40(dd,J=9.1,3.0Hz,1H),4.44( d,J=13.3Hz,1H),4.20(q,J=7.1Hz,2H),3.75-3.52(m,6H),3.22-3.07(m,5H) ),2.82(s,2H),2.22-2.10(m,5H),1.76-1.62(m,2H),1.31(t,J=7.1Hz,3H). 13C NMR (101MHz, CDCl3) δ190.90,168.97,158.68,155.63,155.60,155.45,152. 11,150.39,150.30,149.55,147.85,147.59,146.67,142.90,141.01,140.95 ,137.27,127.39,127.05,121.81,121.72,121.67,119.17,119.09,112.44,78.17,61.61,50.01,48.00,43.56,41.85,36.88,34.59,33.84,21.46,14.71.

[0130] Example 44 1'-Acetyl-7-(5-fluoro-2-((5-(morpholine-4-carbonyl)piperazin-1-yl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 44)

[0131] Compound 44 was prepared by the method of Compound 20, using 5-(morpholine-4-carbonyl)piperazin-1-yl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino; 1 H NMR(300MHz, CDCl3)δ9.05(s,1H),8.60-8.53(m,2H),8.45(dd,J=2.3,0.8Hz,1H),8.08-8.03(m,1H),7.92(dd,J=8.8,2.3Hz,1H),7.81-7.76(m,2 H),4.44(d,J=13.4Hz,1H),3.81-3.65(m,8H),3.59(t,J=11.5Hz,2H),3. 14(t,J=12.7Hz,1H),2.82(s,2H),2.22-2.12(m,5H),1.75-1.62(m,2H). 13 C NMR (101MHz, CDCl3) δ190.87,169.01,168.09,158.73,155.06,153.76,152.60,150.76,150.67,150.03,148.00,147.74,147.35,140.55,1 40.50,138.11,127.20,124.61,121.99,121.71,121.65,119.20,119 .13,111.24,78.25,66.88,47.99,41.85,36.89,34.66,33.76,21.45.

[0132] Example 45 tert-Butyl 4-(6-((4-(1'-acetyl-4-oxospiro[benzopyran-2,4'-piperidin]-7-one)-5-fluoropyrimidin-2-yl)amino)nicotinoyl)piperazine-1-carboxylate (Compound 45)

[0133] Compound 45 was prepared by the method of Compound 20, using 4-(6-aminonicotinoyl)piperazine-1-carboxylic acid tert-butyl ester to replace 5-(piperidin-1-yl)pyridine-2-amino; 1 H NMR (300MHz, CDCl3) δ8.98(s,1H),8.58(d,J=3.1Hz,1H),8.53-8.48(m,2H),8.05(d,J=8.2Hz,1H),7.87(d,J=8.5Hz,1H),7.81-7.75(m,2H),4 .44(d,J=13.4Hz,1H),3.81-3.56(m,6H),3.51(s,4H),3.14(t,J=12.7H z,1H),2.82(s,2H),2.21-2.10(m,5H),1.76-1.63(m,2H),1.50(s,9H). 13 C NMR (101MHz, CDCl3) δ190.85,169.00,168.31,158.73,155.11,155.07,154 .54,153.82,152.58,150.73,150.64,150.01,148.03,147.77,147.44,140. 56,140.51,138.00,127.19,124.75,121.99,121.71,121.65,119.20,119.13,111.19,80.51,78.25,48.00,41.85,36.89,34.66,33.76,28.38,21.46.

[0134] Example 46 1'-Acetyl-7-(5-fluoro-2-((5-(4-methylpiperazine-1-carbonyl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 46)

[0135] Using 5-(4-methylpiperazine-1-carbonyl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino, compound 46 was prepared according to the method of compound 20; 1H NMR (300MHz, CDCl3) δ8.73 (s, 1H), 8.57 (d, J = 3.2Hz, 1H), 8.52-8.45 (m, 2H), 8.05(dd,J=7.9,0.8Hz,1H),7.85(dd,J=8.6,2.4Hz,1H),7.78(dd,J=8.1,1. 2Hz,2H),4.44(d,J=13.6Hz,1H),3.91-3.52(m,6H),3.14(t,J=12.4Hz,1H), 2.82(s,2H),2.55(s,4H),2.41(s,3H),2.23-2.12(m,5H),1.76-1.62(m,2H). 13 C NMR (101MHz, CDCl3) δ190.88,169.01,168.05,158.73,155.17,155.14,153.70,152.54,150.68,150.59,149.97,148.03,147.76,147.50,140. 62,140.56,137.92,127.18,125.07,121.96,121.71,121.65,119.19,1 19.12,111.11,78.23,48.00,46.00,41.85,36.89,34.65,33.76,21.45.

[0136] Example 47 1'-Acetyl-7-(5-fluoro-2-((5-(4-ethylpiperazine-1-carbonyl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 47)

[0137] Compound 47 was prepared by replacing 5-(piperidin-1-yl)pyridin-2-amino with 5-(4-ethylpiperazine-1-carbonyl)pyridin-2-amino; 1 H NMR (300MHz, CDCl3) δ8.55 (d, J = 3.1Hz, 1H), 8.51-8.47 (m, 2H), 8.47-8.44 (m, 1H),8.07-8.03(m,1H),7.85(dd,J=8.8,2.2Hz,1H),7.78(dd,J=8.0,1.2Hz,2H ),4.45(d,J=13.6Hz,1H),3.95-3.52(m,6H),3.14(t,J=12.6Hz,1H),2.82(s, 2H),2.58(s,4H),2.23-2.11(m,5H),1.74-1.63(m,4H),1.18(t,J=7.2Hz,3H). 13C NMR (101MHz, CDCl3) δ190.89,169.02,167.97,158.73,155.17,155.14,1 53.68,152.54,150.67,150.58,149.97,148.03,147.76,147.51,140.62, 140.57,137.94,127.18,125.10,121.96,121.71,121.65,119.19,119.12 ,111.10,78.23,52.26,48.00,41.86,36.90,34.65,33.76,21.45,11.86.

[0138] Example 48 1'-Acetyl-7-(5-fluoro-2-((5-(4-acetylpiperazine-1-carbonyl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 48)

[0139] Compound 48 was prepared by replacing 5-(piperidin-1-yl)pyridin-2-amino with 5-(4-acetylpiperazine-1-carbonyl)pyridin-2-amino; 1 H NMR (300MHz, CDCl3) δ8.95(s,1H),8.58(d,J=3.1Hz,1H),8.54-8.49(m,2H),8.07-8.02(m,1H),7.87(dd,J=8.6,2.4Hz,1H),7.78(dd,J=8.1,1.2Hz,2 H),4.44(d,J=13.6Hz,1H),3.81-3.61(m,7H),3.59-3.53(m,3H),3.21-3. 07(m,1H),2.82(s,2H),2.23-2.15(m,5H),2.14(s,3H),1.76-1.62(m,2H). 13 C NMR (101MHz, CDCl3) δ190.86,169.29,169.02,168.39,158.73,155.08,155.0 5,153.99,152.60,150.77,150.68,150.03,148.03,147.77,147.51,140.53, 140.48,138.06,127.19,124.38,121.99,121.69,121.64,119.21,119.14,111.24,78.26,47.98,46.15,41.84,41.50,36.89,34.65,33.74,21.46,21.42.

[0140] Example 49 1'-Acetyl-7-(5-fluoro-2-((5-(4-cyclopropionylpiperazine-1-carbonyl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 49)

[0141] Using 5-(4-cyclopropylpiperazine-1-carbonyl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino, compound 49 was prepared according to the method of compound 20; 1 H NMR (300MHz, CDCl3) δ8.88(s,1H),8.57(d,J=3.1Hz,1H),8.54(d,J=8.9Hz,1H),8.48(d, J=2.2Hz,1H),8.05(d,J=8.8Hz,1H),7.90(dd,J=8.6,2.1Hz,1H),7.82-7.75(m,2H),4.4 4(d,J=13.5Hz,1H),3.88-3.65(m,8H),3.59(t,J=12.4Hz,2H),3.14(t,J=12.7Hz,1H),2 .83(s,2H),2.23-2.21(m,5H),1.78-1.63(m,3H),1.08-1.02(m,2H),0.89-0.80(m,2H). 13 C NMR (101MHz, CDCl3) δ190.86,172.44,169.01,168.37,158.73,155.09,155 .06,153.94,152.60,150.76,150.67,150.03,148.03,147.77,147.51,140 .54,140.49,138.07,127.19,124.50,121.99,121.71,121.65,119.21,119.14,111.23,78.26,47.99,41.85,36.89,34.66,33.75,21.46,11.06,7.82.

[0142] Example 50 1'-Acetyl-7-(5-fluoro-2-((5-(morpholinomethyl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 50)

[0143] Using 5-(morpholinomethyl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino, compound 50 was prepared according to the method of compound 20; 1H NMR(300MHz, CDCl3)δ8.53(d,J=3.2Hz,1H),8.51(s,1H),8.38(d,J=8.6Hz, 1H),8.31-8.29(m,1H),8.05(d,J=7.9Hz,1H),7.82-7.75(m,3H),4.45(d,J= 13.6Hz,1H),3.75(t,J=4.6Hz,4H),3.69-3.56(m,2H),3.54(s,2H),3.24-3 .09(m,1H),2.82(s,2H),2.52(s,4H),2.23-2.11(m,5H),1.75-1.63(m,2H). 13 C NMR (101MHz, CDCl3) δ190.90,169.00,158.70,155.53,155.50,152.29,1 52.11,150.46,150.37,149.73,148.80,147.94,147.68,140.90,140.84, 139.24,127.09,126.81,121.86,121.74,121.68,119.17,119.10,111.5 5,78.19,66.90,60.21,53.45,48.01,41.85,36.89,34.58,33.84,21.46.

[0144] Example 51 tert-Butyl 4-((6-((4-(1'-acetyl-4-oxospiro[benzopyran-2,4'-piperidin]-7-one)-5-fluoropyrimidin-2-yl)amino)pyridin-3-yl)methyl)piperazine-1-carboxylate (Compound 51)

[0145] Compound 51 was prepared by referring to the method of Compound 20, using 4-((6-aminopyridin-3-yl)methyl)piperazine-1-carboxylic acid tert-butyl ester to replace 5-(piperidin-1-yl)pyridin-2-amino; 1H NMR(300MHz, CDCl3)δ8.53(d,J=3.2Hz,1H),8.44(s,1H),8.37(d,J=8.6Hz,1H), 8.28(d,J=2.2Hz,1H),8.04(dd,J=8.1,0.6Hz,1H),7.81-7.72(m,3H),4.45(d,J =13.7Hz,1H),3.74-3.57(m,2H),3.54(s,2H),3.47(s,4H),3.15(t,J=11.6Hz,1 H),2.82(s,2H),2.45(s,4H),2.23-2.10(m,5H),1.69-1.60(m,2H),1.47(s,9H). 13 C NMR (101MHz, CDCl3) δ190.89,168.98,158.70,155.52,155.49,154.74,152.29 ,152.09,150.45,150.36,149.73,148.75,147.96,147.69,140.89,140.84,13 9.18,127.09,126.94,121.86,121.74,121.68,119.17,119.10,111.54,79.71,78.19,59.83,52.73,48.01,41.85,36.89,34.59,33.84,28.43,26.91,21.46.

[0146] Example 52 1'-Acetyl-7-(5-fluoro-2-((5-((4-methylpiperazin-1-yl)methyl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 52)

[0147] Compound 52 was prepared by the method of Compound 20, using 5-((4-methylpiperazin-1-yl)methyl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino; 1H NMR(300MHz, CDCl3)δ8.52(d,J=3.2Hz,1H),8.43(s,1H),8.36(dd,J=8.5,0.8Hz,1H) ,8.30-8.27(m,1H),8.04(dd,J=8.0,0.6Hz,1H),7.81-7.75(m,2H),7.72(dd,J=8.6, 2.3Hz,1H),4.45(d,J=13.3Hz,1H),3.73-3.57(m,2H),3.52(s,2H),3.15(t,J=12.2H z,1H),2.82(s,2H),2.56(s,8H),2.36(s,3H),2.21-2.13(m,5H),1.75-1.63(m,2H). 13 C NMR (101MHz, CDCl3) δ190.90,168.96,158.70,155.56,155.53,152.26,151 .98,150.39,150.30,149.70,148.78,147.97,147.71,140.93,140.88,139 .17,127.31,127.08,121.85,121.75,121.69,119.16,119.08,111.49,78.18,59.79,55.02,52.85,48.01,45.94,41.86,36.89,34.56,33.86,21.46.

[0148] Example 53 1'-Acetyl-7-(5-fluoro-2-((5-((4-ethylpiperazin-1-yl)methyl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 53)

[0149] Compound 53 was prepared by the method of Compound 20, using 5-((4-ethylpiperazin-1-yl)methyl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino; 1H NMR (300MHz, CDCl3) δ8.56 (s, 1H), 8.53 (d, J = 3.2Hz, 1H), 8.35 (dd, J = 8.5, 0.8Hz, 1H), 8.2 9(dd,J=2.3,0.8Hz,1H),8.04(dd,J=8.1,0.6Hz,1H),7.81-7.75(m,2H),7.72(dd,J=8.6,2 .3Hz,1H),4.45(d,J=13.4Hz,1H),3.74-3.57(m,2H),3.52(s,2H),3.23-3.09(m,1H),2.8 2(s,2H),2.75-2.31(m,10H),2.21-2.13(m,5H),1.75-1.62(m,2H),1.12(t,J=7.2Hz,3H). 13 C NMR (101MHz, CDCl3) δ190.90,168.95,158.70,155.56,155.53,152.26,151. 97,150.38,150.29,149.70,148.80,147.97,147.71,140.93,140.88,139.19 ,127.27,127.07,121.85,121.75,121.69,119.16,119.08,111.49,78.18,59.82,52.80,52.69,52.29,48.02,41.86,36.89,34.56,33.86,21.46,11.89.

[0150] Example 54 1'-Acetyl-7-(5-fluoro-2-((5-((4-acetylpiperazin-1-yl)methyl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 54)

[0151] Compound 54 was prepared by the method of Compound 20, using 5-((4-acetylpiperazin-1-yl)methyl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino; 1H NMR (300MHz, CDCl3) δ8.56-8.52(m,2H),8.38(dd,J=8.5,0.8Hz,1H),8.29(dd ,J=2.3,0.9Hz,1H),8.04(dd,J=8.1,0.6Hz,1H),7.82-7.71(m,3H),4.45(d,J =13.7Hz,1H),3.74-3.58(m,4H),3.55-3.46(m,4H),3.22-3.09(m,1H),2.82( s,2H),2.53-2.42(m,4H),2.22-2.12(m,5H),2.11(s,3H),1.73-1.61(m,2H). 13 C NMR (101MHz, CDCl3) δ190.90,169.01,158.70,155.49,155.46,152.31,152.18 ,150.49,150.40,149.75,148.70,147.95,147.69,140.86,140.80,139.15,12 7.10,126.70,121.87,121.73,121.67,119.18,119.11,111.59,78.20,59.66,52.96,52.54,48.00,46.21,41.85,41.34,36.89,34.58,33.83,21.46,21.36.

[0152] Example 55 1'-Acetyl-7-(5-fluoro-2-((5-((4-cyclopropionylpiperazin-1-yl)methyl)pyridin-2-yl)amino)pyrimidin-4-yl)spiro[benzopyran-2,4'-piperidin]-4-one (Compound 55)

[0153] Compound 55 was prepared by the method of Compound 20, using 5-((4-cyclopropionylpiperazin-1-yl)methyl)pyridine-2-amino instead of 5-(piperidin-1-yl)pyridine-2-amino; 1H NMR (300MHz, CDCl3) δ8.61 (s, 1H), 8.54 (d, J = 3.2Hz, 1H), 8.39 (dd, J = 8.5, 0.8Hz, 1H), 8.33-8.29(m,1H),8.07-8.01(m,1H),7.81-7.73(m,3H),4.44(d,J=13.5Hz,1H),3.78 -3.58(m,6H),3.55(s,2H),3.15(t,J=12.0Hz,1H),2.82(s,2H),2.50(s,4H),2.22-2. 11(m,5H),1.76-1.72(m,1H),1.72-1.61(m,2H),1.02-0.96(m,2H),0.81-0.74(m,2H). 13 C NMR (101MHz, CDCl3) δ190.90,171.99,169.01,158.70,155.51,155.48,152.30,15 2.19,150.47,150.38,149.74,148.74,147.96,147.70,140.87,140.82,139.20,1 27.09,126.68,121.87,121.73,121.68,119.18,119.11,111.60,78.19,59.69,53.15,52.62,48.00,45.35,42.00,41.85,36.89,34.58,33.82,21.46,10.91,7.46.

[0154] Biological evaluation experiments were conducted on the compounds of the present invention:

[0155] Example 56: CDK6 / cyclin D1 inhibitory activity test

[0156] Different concentrations of compounds were added to a 384-well experimental plate (Perkin Elmer, Cat. No. 6007299), and then CDK6 / cyclin D1 (Invitrogen, Cat. No. PV4437) kinase and substrate ULight-4E-BP1 Peptide (Perkin Elmer, Cat. No. TRF0128-M) were added and mixed evenly; after incubation at room temperature for 15 minutes, ATP solution was added to start the reaction; after reaction at room temperature for 3 hours, EDTA was added to terminate the reaction, and the antibody Eu-phos-4E-BP1 (Perkin Elmer, Cat. No. TRF0216-M) was added to generate the detection signal; after incubation at room temperature for 1 hour, it was read in an Envision plate reader (BMGPOLARstar Omega). IC was analyzed using GraphPad Prism 8.0 software (San Diego, USA, RRID: SCR_002798) 50 Values ​​and curve fitting.

[0157] Table 1. IC values ​​of the compounds of the present invention for CDK6 / cyclin D1 kinase inhibitory activity 50 value

[0158]

[0159]

[0160] The experimental results showed that the inhibitory activities of compounds 14, 23, 33, 36-39 and 42 on CDK6 were all lower than 25 nM, showing strong inhibitory activity.

[0161] Example 57: Evaluation of cytotoxicity and cytoprotective effects of active compounds 14, 23, 33, 36-39 and 42

[0162] Cytotoxicity evaluation of compounds: OP9 cells were cultured at 1×10 5 The cells were inoculated at a density of 100 μg / mL in a 96-well plate. After 24 hours, the cells were treated with compounds at different concentrations. A blank control group was set up, and three replicate wells were set up in parallel. After incubation in a 37°C, 5% CO2 incubator for 48 hours, 20 μL of MTT solution (5 mg / mL) was added to each well. After further incubation for 4 hours, the supernatant was discarded, 150 μL of DMSO was added to each well, and the cells were shaken on a shaker for 10 minutes. The absorbance was measured at 490 nm, and CC was calculated using GraphPad Prism 8.0 software (San Diego, USA, RRID: SCR_002798). 50 value.

[0163] The protective effect of the compounds on 5-FU-induced OP9 cells: OP9 cells were cultured at 1×10 5 The cells were inoculated at a density of 100 μg / mL in a 96-well plate. After 24 hours, the cells were treated with compounds at different concentrations. A blank control group was set up, and three replicate wells were set up in parallel. After incubation in a 37°C, 5% CO2 incubator for 8 hours, 20 μL of 5-FU solution (50 μg / mL) was added to each well. After continued cultivation for 48 hours, 20 μL of MTT solution (5 mg / mL) was added to each well. After continued cultivation for 4 hours, the supernatant was discarded, 150 μL of DMSO was added to each well, and the cells were shaken on a shaker for 10 minutes. The absorbance was measured at 490 nm, and the EC was calculated using GraphPad Prism 8.0 software (San Diego, USA, RRID: SCR_002798) 50 value.

[0164] Table 2. CC effects of active compounds 14, 23, 33, 36-39 and 42 of the present invention on OP9 cells 50 EC values ​​for 5-FU-induced OP9 cell injury 50 value

[0165]

[0166] The experimental results showed that compared with the positive control drug Trilaciclib, compound 42 showed minimal cytotoxicity (CC 50 >100μM) and the strongest protective effect (EC 50 =4.02 μM).

[0167] Example 58: Interaction between active compound 42 and CDK6 protein

[0168] The active compound 42 was molecularly docked with CDK6 protein (PDB: 5L2S) using Discovery Studio software. The docking results showed that the chromone spirocyclic skeleton, pyrimidine and secondary amine fragments in compound 42 formed hydrogen bonds with CDK6 ( Figure 2 ). It can be seen that the chromone spirocyclic skeleton, pyrimidine and secondary amine fragments in the active compound of the present invention are the crucial parts of the CDK4 / 6 inhibitory activity of compound 42, and the lack of any part will affect the activity.

[0169] Example 59: Pharmacokinetics of Compound 42

[0170] The pharmacokinetic characteristics of compound 42 were tested. A total of 10 SD rats were used in this experiment. They were divided into two groups (5 rats in each group) and given compound 42 by tail vein injection (2 mg / kg) and oral gavage (50 mg / kg). Blood samples were collected from the retro-orbital venous plexus at predetermined time points (0.05h, 0.083h, 0.167h, 0.25h, 0.5h, 0.75h, 1h, 2h, 4h, 6h, 9h, 12h, 24h), and 150 μL of blood was collected at each time point. The concentration of the compound in the plasma samples of SD rats was determined by LC-MS / MS, and the pharmacokinetic parameters were calculated using WinNolin software.

[0171] Table 3. Pharmacokinetic characteristics test of compound 42

[0172] The experimental results showed that the half-life of the intravenous injection group was 1.54h, the average residence time was 1.55h, and the AUC was 591.10h·ng / mL; the half-life of the oral group was 7.13 hours, Tmax was 6 hours, Cmax was 480.55ng / mL, AUC was 4215.57h·ng / mL, and the oral bioavailability was 28.53%. These results indicate that compound 42 has good drug-like properties.

[0173] Example 60: Pharmacodynamics study of compound 42

[0174] Balb / c mice were randomly divided into 5 groups (n=10): control group, 5-FU group, 5-FU+Trilaciclib group (50 mg / kg, intraperitoneal injection) and 5-FU+42 group (12.5, 25 mg / kg, gavage). 42 or Trilaciclib was administered one day before inducing bone marrow suppression, and then a single intraperitoneal injection of 5-FU (100 mg / kg) was given on the first day. The control group mice were intraperitoneally or gavaged with an equal amount of normal saline at the time of administration or modeling. Blood samples were collected from the fundus at designated time points (4, 6, 8 and 14 days) and analyzed using a BC-5000Vet fully automatic blood analyzer (Mindray, China) ( Figure 3 ). The experimental results showed that compared with the positive control drug Trilaciclib, the 42 treatment group could significantly increase the number of platelets (PLT), red blood cells (RBC), hemoglobin (HGB), white blood cells (WBC), lymphocytes (Lym) and neutrophils (Neu), and could improve the weight loss of mice caused by chemotherapy and increase the survival rate of mice.

[0175] Example 61: Safety evaluation of compound 42

[0176] Acute toxicity test: Half of the male and female Balb / c mice were randomly divided into 4 groups, namely male control group (n=5), male 42 treatment group (n=10), female control group (n=5) and female 42 treatment group (n=10). After a single oral administration of 42 (1000 mg / kg), the mice were weighed every 2 days. After 14 days, the mice were euthanized by cervical dislocation, and the main organs (heart, liver, spleen, lung, kidney) were removed and weighed. The experimental results showed that there was no difference in food intake between the control group and the 42 treatment group ( Figure 4 In addition, the weights of the main organs of the mice, including the heart, liver, spleen, lungs, and kidneys, showed no difference between the control group and the drug-treated group ( Figure 4 ). H&E staining showed that no obvious lesions were found in the main organs of mice in the 1000mg / kg group ( Figure 5 ). The above results show that 42 has good drug safety; therefore, the chromone spirocyclic compound of the present invention has selective CDK4 / 6 inhibitory activity, can alleviate and / or treat chemotherapy-induced bone marrow suppression, improve chemotherapy-induced weight loss and increase survival rate, and has great clinical application prospects.

Claims

1. A chromone spiro compound, characterized in that: The compound has a structure as shown in Formula I: Wherein, X is selected from O, (CH2) n , C(O), NH or S(O)2, n is 0 or 1; R1, R2 are selected from C3-C8 cycloalkyl, C3-C8 cycloalkyloxy or C3-C8 heterocyclyl, wherein the C3-C8 cycloalkyl, C3-C8 cycloalkyloxy or C3-C8 heterocyclyl is independently and optionally substituted with one or more substituents selected from halogen, hydroxyl, amino, nitro, cyano, C1-C6 alkyl, C1-C6 alkoxy, C3-C6 cycloalkyl, C3-C6 cycloalkyloxy or C4-C8 heterocyclyl; R3 is hydrogen, halogen, hydroxyl, mercapto, amino, -OR', -SR', -NHR' or -NR'R", wherein R' and R" are C1-C3 alkyl, C2-C6 alkenyl, C3-C6 allyl, C2-C6 alkynyl, C1-C3 haloalkyl, C3-C6 cycloalkyl, C6-C 10 Aryl, C3-C 10 Heteroaryl or C4-C8 heterocyclic group; R4 is selected from C3-C8 cycloalkyloxy, C3-C8 heterocyclyl, C3-C8 heterocyclylphenyl or C3-C8 heterocyclylaryl, wherein the C3-C8 cycloalkyloxy, C3-C8 heterocyclyl, C3-C8 heterocyclylphenyl or C3-C8 heterocyclylaryl is independently and optionally substituted by one or more substituents selected from halogen, hydroxyl, amino, nitro, cyano, C1-C6 alkyl, C1-C6 alkoxy, C3-C6 cycloalkyl, C3-C6 cycloalkyloxy or C4-C8 heterocyclyl.

2. The chromone spiro compound according to claim 1, characterized in that Said X is C(O) or (CH2) n ; n is 0 or 1; R1 and R2 are C3-C8 cycloalkyl or C3-C8 heterocyclic group; R3 is hydrogen or halogen; R4 is C3-C8 heterocyclic group. Wherein E is glutamic acid, R1 and R2 are organic carbon chains selected from C6-C 24 Alkyl, substituted C6-C 24 Alkyl, C6-C 24 Alkenyl, substituted C6-C 24 Alkenyl, C6-C 24 Alkynyl or substituted C6-C 24 Alkynyl; n is a positive integer ≥1.

3. The chromone spiro compound according to claim 1, characterized in that Said X is C(O) or (CH2) n ; n is 0 or 1; R1 and R2 are C4-C6 cycloalkyl or C6 heterocyclic group; R3 is halogen; R4 is C6 heterocyclic group.

4. The chromone spiro compound according to claim 1, characterized in that The X is selected from: C(O) or (CH2) n ; n is 0 or 1; R1 and R2 are selected from cyclobutyl, cyclopentyl, cyclohexyl, cyclohexyl derivatives, tetrahydropyran, tetrahydrothiothiopyran or N-amide piperidine; R3 is F; R4 is selected from piperidine, piperidine derivatives, morpholine, piperazine or piperazine derivatives.

5. The chromone spiro compound according to claim 1, characterized in that Selected from the following compounds 1-55:

6. The chromone spiro compound according to claim 1, characterized in that Also included are pharmaceutically acceptable salts of the above-mentioned chromone spirocyclic compounds, specifically, salts formed by adding an acid to the compound of general formula (I), wherein the acid used for salt formation includes inorganic acids and organic acids, wherein the inorganic acids include: hydrochloric acid, sulfuric acid, phosphoric acid and methanesulfonic acid; the organic acids include: galactaric acid, D-glucuronic acid, glycerophosphoric acid, hippuric acid, isethionic acid, lactobionic acid, maleic acid, naphthalene-2-sulfonic acid, pivalic acid, terephthalic acid, bile acid Acid, benzenesulfonic acid, citric acid, lactic acid, malic acid, malonic acid, mandelic acid, acetic acid, propionic acid, tartaric acid, succinic acid, formic acid, methanesulfonic acid, picric acid, L-pyroglutamic acid, salicylic acid, gentisic acid, p-toluenesulfonic acid, valeric acid, palmitic acid, stearic acid, lauric acid, adipic acid, 4-benzenesulfonic acid, fumaric acid, 3-hydroxynaphthalene-2-carboxylic acid, ascorbic acid, camphoric acid, camphorsulfonic acid, cinnamic acid, dichloroacetic acid and ethanesulfonic acid.

7. A method for preparing the chromone spiro compound according to claim 1, characterized in that: The preparation method is: (1) 1-(4-bromo-2-hydroxyphenyl)ethanone represented by formula II undergoes condensation reaction with cyclic ketone under alkaline conditions to obtain chromone intermediate compound III; (2) the chromone intermediate compound III reacts with bis-pinacol borate in the presence of a palladium catalyst to obtain compound IV; (3) Compound IV is coupled with a chloropyrimidine derivative to obtain compound V; (4) Compound V is coupled with a 2-aminopyrimidine derivative in the presence of a palladium catalyst and a phosphine ligand to obtain a chromone spirocyclic compound represented by formula I; The synthetic route is as follows, wherein R1, R2, R3, R4 and X are as described in claim 1:

8. The method according to claim 7, characterized in that: In step (1), the base used in the alkaline condition is selected from pyrrolidine, piperidine, tetrahydropyridine, sodium hydroxide, potassium hydroxide, potassium carbonate or sodium carbonate; the reaction solvent is selected from methanol, ethanol, isopropanol, perfluoroisopropanol, n-butanol or tert-butanol; In step (2), the base used in the alkaline condition is selected from potassium phosphate, potassium carbonate, sodium carbonate, cesium carbonate, potassium fluoride, potassium acetate, sodium acetate, potassium tert-butoxide or sodium tert-butoxide; the palladium catalyst is selected from Pd(dppf)Cl2, Pd(PPh3)2Cl2, Pd(PPh3)4, Pd2(dba)3, PdCl2(dippf)2 or [Pd(allyl)Cl]2; the reaction solvent is selected from dioxane, N,N-dimethylformamide, N,N-dimethylacetamide, dimethyl sulfoxide, acetonitrile or N-methylpyrrolidone; In step (3), the base used in the alkaline condition is selected from sodium hydroxide, potassium hydroxide, barium hydroxide, potassium phosphate, potassium carbonate, sodium carbonate, cesium carbonate, potassium fluoride, potassium acetate, sodium acetate, sodium methoxide, potassium methoxide, sodium ethoxide, potassium ethoxide, potassium tert-butoxide or sodium tert-butoxide; the palladium catalyst is selected from Pd(dppf)Cl2, Pd(PPh3)2Cl2, Pd(PPh3)4, Pd2(dba)3, PdCl2(dippf)2 or [Pd(allyl)Cl]2; the reaction solvent is selected from tetrahydrofuran, ethanol, acetonitrile, dioxane, ethylene glycol dimethyl ether, dimethyl sulfoxide or toluene; the volume ratio of the reaction solvent to water is 8:1 to 12:1; In step (4), the base used for the alkaline condition is selected from sodium hydroxide, potassium hydroxide, potassium carbonate, sodium carbonate, cesium carbonate, sodium methoxide, potassium methoxide, sodium ethoxide, potassium ethoxide, potassium tert-butoxide or sodium tert-butoxide; the palladium catalyst is selected from Pd2(dba)3 or Pd(OAc)2; the phosphine ligand is selected from PCy3, P(t-Bu)3, P(o-tolyl)3, DPPF, JohnPhos, SPhos, XantPhos or BINAP; the reaction solvent is selected from toluene, xylene, dioxane, tetrahydrofuran, ethylene glycol dimethyl ether, N,N-dimethylformamide, N,N-dimethylacetamide or dimethyl sulfoxide.

9. Use of the chromone spirocyclic compound according to claim 1 in the preparation of a CDK4 / 6 inhibitor.

10. Use of the chromone spirocyclic compound according to claim 1 in the preparation of a drug for alleviating and / or treating chemotherapy-induced bone marrow suppression.

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