CN116554165A - Tetrahydropyridopyrimidine derivatives and their use - Google Patents

Abstract

The invention belongs to the field of chemical medicine and provides a class of tetrahydropyridopyrimidine derivatives. The present invention provides a class of novel small-molecule compounds with tetrahydropyridopyrimidine as the backbone. Such derivatives exhibit good anti-tumor activity on enzymes, cells and animals, and can be used as ATR kinase inhibitors for cancer treatment. In particular, it has shown good anticancer effects on colorectal cancer and mantle cell lymphoma, providing a new option for the treatment of these diseases.

Description

Tetrahydropyridopyrimidine derivatives and uses thereof

Technical Field

The invention relates to tetrahydropyridopyrimidine derivatives and uses thereof, belonging to the field of chemical medicine.

Background Art

Ataxia telangiectasia and Rad3-related (ATR) is an atypical serine/threonine protein kinase that plays a key role in the DNA replication stress response. ATR, along with ataxia telangiectasia mutated kinase (ATM) and DNA-dependent protein kinase (DNA-PK), belongs to the phosphatidylinositol 3-kinase-like kinase (PIKK) family and is an essential component of the DNA damage response (DDR); the DDR has evolved to recognize, signal, and promote the repair of damaged DNA.

Unlike ATR, which responds to DNA replication stress, ATM and DNA-PK respond primarily to DNA double-strand breaks. Normally, the DDR network coordinates the recognition, signaling, and repair of damaged DNA, thereby ensuring genomic stability. Numerous studies have shown that tumor cells with ATM mutations or loss of function, or with other DDR defects that promote replicative stress, are often more dependent on ATR for survival. Based on the principle of synthetic lethality, this provides the possibility of using ATR inhibitors to kill cancer cells while sparing normal non-cancerous cells.

Therefore, the development of ATR kinase inhibitors for the treatment of related cancers has important application value.

Summary of the invention

The present invention develops a new type of tetrahydropyrido-pyrimidine derivative ATR inhibitor and a preparation method thereof, which exhibits good anti-tumor activity on enzymes, cells and animals and can be used to prepare ATR inhibitor drugs.

The present invention first provides a compound represented by formula I or a pharmaceutically acceptable salt thereof:

R ₁ is selected from R ₄ and R ₅ are independently selected from substituted or unsubstituted C1-C8 alkyl, substituted or unsubstituted 3-8 membered cycloalkyl, substituted or unsubstituted C1-C8 alkenyl, substituted or unsubstituted 6-10 membered aryl; in R ₁ , the substituents of the substituted C1-C8 alkyl and substituted C1-C8 alkenyl are selected from halogen, C1-C8 acyl, C1-C8 alkoxy, and cyano, and the substituents of the substituted 3-10 membered cycloalkyl and substituted 6-10 membered aryl are selected from C1-C8 alkyl, halogen, C1-C8 acyl, C1-C8 alkoxy, and cyano;

R ₂ and R ₃ are independently selected from H, substituted or unsubstituted C1-C8 alkyl, substituted or unsubstituted 3-10-membered azacycloalkyl, and R ₂ and R ₃ are not H at the same time, or R ₂ and R ₃ are connected with N to form a substituted or unsubstituted 4-10-membered heterocycloalkyl, the 3-10-membered azacycloalkyl contains 1-2 N, and the 4-10-membered heterocycloalkyl contains 0-2 heteroatoms in addition to N shown in formula I, and the heteroatoms are N, O or S; in R ₂ and R ₃ , the substituent of the substituted C1-C8 alkyl is selected from C1-C8 alkoxycarbonyl, amino, and sulfo; the substituent of the substituted 4-10-membered azacycloalkyl and substituted 4-10-membered heterocycloalkyl is selected from C1-C8 alkyl, C1-C8 alkoxycarbonyl, amino, and sulfo;

_R4 is indolyl.

Among them, in the above compounds, R ₄ is

Wherein, in the above compounds, R ₄ and R ₅ are independently selected from substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted 6-10 membered aryl, n＝0～3; in R ₄ and R ₅ , the substituent of the substituted C1～C6 alkyl is selected from halogen, C1～C6 acyl, C1～C6 alkoxy and cyano, and the substituent of the substituted 6～10 membered aryl is selected from C1～C6 alkyl, halogen, C1～C6 acyl, C1～C6 alkoxy and cyano; R ₆ is selected from H, halogen, cyano, halogen substituted or unsubstituted C1～C6 alkyl; R ₇ is selected from H, halogen and C1～C6 alkyl; R ₈ is selected from H and C1～C6 alkyl; R ₉ is selected from H, C1～C6 alkyl and C1～C6 alkoxy.

Preferably, in the above compounds, R ₄ and R ₅ are independently selected from substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted 6-membered aryl, n＝0～3; in R ₄ and R ₅ , the substituent of the substituted C1～C6 alkyl is selected from halogen, C1～C4 acyl, C1～C4 alkoxy and cyano, and the substituent of the substituted 6～10 membered aryl is selected from C1～C4 alkyl, halogen, C1～C4 acyl, C1～C4 alkoxy and cyano; R ₆ is selected from H, halogen, cyano, halogen substituted or unsubstituted C1～C4 alkyl; R ₇ is selected from H, halogen and C1～C4 alkyl; R ₈ is selected from H and C1～C4 alkyl; R ₉ is selected from H, C1～C4 alkyl and C1～C4 alkoxy.

More preferably, in the above compounds, R ₄ and R ₅ are independently selected from substituted or unsubstituted C1-C6 alkyl, phenyl, n＝0～3; in R ₄ and R ₅ , the substituent of the substituted C1～C6 alkyl is selected from F, Cl, C1～C4 acyl; R ₆ is selected from H, F, cyano, unsubstituted C1～C4 alkyl, trifluoromethyl; R ₇ is selected from H, F, C1～C4 alkyl; R ₈ is selected from H, C1～C4 alkyl; R ₉ is selected from H, C1～C4 alkyl, C1～C4 alkoxy.

More preferably, in the above compounds, _R4 is selected from substituted or unsubstituted C1-C6 alkyl, phenyl, n＝0～3; in R ₄ , the substituent of the substituted C1～C6 alkyl is selected from F, Cl, C1～C4 acyl; R ₆ is selected from H, F, cyano, unsubstituted C1～C4 alkyl, trifluoromethyl; R ₇ is selected from H, F, C1～C4 alkyl; R ₈ is selected from H, C1～C4 alkyl; R ₉ is selected from H, C1～C4 alkyl, C1～C4 alkoxy; R ₅ is fluorine-substituted or unsubstituted C1～C4 alkyl.

Most preferably, in the above compounds, _R4 is selected from methyl, difluoromethyl, trifluoromethyl, acetylmethyl, isopropyl, tert-butyl, isobutyl, Vinyl, Phenyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, R ₅ is selected from trifluoromethyl and methyl.

Wherein, in the above compounds, R ₂ and R ₃ are independently selected from H, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted 3-8 membered azacycloalkyl, and R ₂ and R ₃ are not H at the same time, and the 3-8 membered azacycloalkyl contains 1-2 N; in R ₂ and R ₃ , the substituent of the substituted C1-C6 alkyl is selected from C1-C6 alkoxycarbonyl, -NHR ₁₀ , and -SO ₂ R ₁₁ ; the substituent of the substituted 3-8 membered azacycloalkyl is selected from C1-C6 alkyl, C1-C6 alkoxycarbonyl, -NHR ₁₂ , and -SO ₂ R ₁₃ ; R ₁₀ and R ₁₂ are independently selected from H and C1-C6 alkyl; R ₁₁ and R ₁₃ are independently selected from C1-C6 alkyl.

Preferably, in the above compounds, R ₂ and R ₃ are independently selected from H, substituted or unsubstituted C1-C4 alkyl, substituted or unsubstituted 5-6-membered azacycloalkyl, and R ₂ and R ₃ are not H at the same time, and the 5-6-membered azacycloalkyl contains 1 N; in R ₂ and R ₃ , the substituent of the substituted C1-C4 alkyl is selected from C1-C4 alkoxycarbonyl, -NHR ₁₀ , and -SO ₂ R ₁₁ ; the substituent of the substituted 5-6-membered azacycloalkyl is selected from C1-C4 alkyl, C1-C4 alkoxycarbonyl, -NHR ₁₂ , and -SO ₂ R ₁₃ ; R ₁₀ and R ₁₂ are independently selected from H and C1-C4 alkyl; R ₁₁ and R ₁₃ are independently selected from C1-C4 alkyl.

More preferably, in the above compounds, R ₂ and R ₃ are independently selected from H, C1-C4 alkoxycarbonyl substituted or unsubstituted C1-C4 alkyl, unsubstituted 5-6 membered azacycloalkyl, and R ₂ and R ₃ are not H at the same time, and the 5-6 membered azacycloalkyl contains 1 N.

Most preferably, in the above compounds, R ₂ and R ₃ are independently selected from H, methyl,

The present invention also provides a compound having a structure as shown in Formula II or a pharmaceutically acceptable salt thereof:

The nitrogen heterocycle A is selected from substituted or unsubstituted 5- to 8-membered heterocycloalkyl groups, wherein the 5- to 8-membered heterocycloalkyl groups contain 0 to 1 heteroatoms in addition to N as shown in formula II, and the heteroatoms are N, O or S; the substituents of the substituted 5- to 8-membered heterocycloalkyl groups are selected from C1-C6 alkyl groups, C1-C6 alkoxycarbonyl groups, -NHR ₁₄ , and -SO ₂ R ₁₅ ; R ₁₄ is selected from H and C1-C6 alkyl groups; and R ₁₅ is selected from C1-C6 alkyl groups.

Among them, in the above compounds, the nitrogen heterocyclic ring A is selected from X is selected from O, NR ₂₀ , CHR ₂₁ , SO ₂ ; R ₁₆ , R ₁₇ , R ₁₉ are independently selected from H, C1-C4 alkyl, C1-C4 alkoxycarbonyl, -NHR ₁₄ , -SO ₂ R ₁₅ ; R ₁₄ is selected from H, C1-C4 alkyl; R ₁₅ is C1-C4 alkyl; R ₁₈ , R ₂₀ , R ₂₁ are independently selected from H, C1-C4 alkoxycarbonyl, -SO ₂ R ₂₂ ; R ₂₂ is selected from C1-C4 alkyl.

Preferably, in the above compounds, the nitrogen heterocycle A is selected from R ₁₇ is -NHR ₁₄ ; R ₂₃ , R ₂₄ , R ₂₅ and R ₂₆ are independently selected from H, C1-C4 alkyl, C1-C4 alkoxycarbonyl and -NHR ₁₄ ; R ₁₄ is selected from H and C1-C4 alkyl; R ₂₇ is -SO ₂ R ₁₅ ; R ₁₅ is selected from C1-C4 alkyl.

Most preferably, in the above compounds, the nitrogen heterocycle A is selected from

Among the above compounds, for for for for for for

The present invention also provides some specific compounds, the structural formula of which is as follows:

The present invention also provides a class of pharmaceutical compositions, which are composed of the above-mentioned compounds or pharmaceutically acceptable salts thereof as active ingredients and pharmaceutically acceptable auxiliary ingredients.

The present invention also provides the use of the above compound or its pharmaceutically acceptable salt, and the above pharmaceutical composition in the preparation of ATR kinase inhibitors.

The present invention also provides the use of the above compound or its pharmaceutically acceptable salt, and the above pharmaceutical composition in the preparation of a drug for treating cancer; preferably, the cancer is colorectal cancer or mantle cell lymphoma.

The present application also provides a method for preparing the above compound, comprising the following steps:

Synthesis route 1:

Synthesis reagents and reaction conditions of compound 5-X in synthesis route 1: a. boron reagent of R ₄ , tetrakistriphenylphosphine palladium, potassium carbonate, 1,4-dioxane/water (V:V=50/1), 95°C, 12h; b. substituted or unsubstituted cyclic amine, DIPEA, DMSO, 130°C, 8h; c. dioxane hydrochloride solution (4M), DCM; d. acyl chloride of R ₁ , TEA, DCM; or acid of R ₁ , HATU, TEA, DCM.

Synthesis route 2:

Synthesis reagents and reaction conditions of compound 5-X in synthesis route 2: boron reagent for R ³ , palladium tetrakistriphenylphosphine, potassium carbonate, 1,4-dioxane/water (V:V=50/1), 95°C, 12h; c. 1,4-dioxane hydrochloric acid solution (4M), DCM; d. acid for R ₁ , HATU, TEA, DCM, or acid chloride for R ₁ , TEA, DCM; e. different amines, DIPEA, DMSO, 130°C, 8h (Note: the protected amine needs to be further deprotected).

Beneficial effects:

The present invention provides a novel small molecule compound with a tetrahydropyridopyrimidine as a skeleton. The derivatives can be used as ATR kinase inhibitors for cancer treatment and exhibit good anti-cancer effects on colorectal cancer and mantle cell lymphoma.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG1 shows the anti-tumor effect of compound 5-30 on mantle cell lymphoma cells Granta-519.

DETAILED DESCRIPTION

The scheme of the present invention will be explained below in conjunction with the embodiments. It will be appreciated by those skilled in the art that the following embodiments are only used to illustrate the present invention and should not be considered as limiting the scope of the present invention. Where specific techniques or conditions are not indicated in the embodiments, the techniques or conditions described in the literature in this area or the product specifications are used. The reagents or instruments used are not indicated by the manufacturer and are all conventional products that can be obtained commercially.

Target molecule synthesis example 1:

Preparation of intermediate 4-(1-(tert-butyloxycarbonyl)-1H-indol-3-yl)-2-chloro-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-carboxylic acid tert-butyl ester 2:

The raw materials 2,4-dichloro-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-carboxylic acid tert-butyl ester 1 (21.37 mmol, 1.0 eq) and (1-(tert-butyloxycarbonyl)-1H-indol-3-yl)boric acid (25.64 mmol, 1.2 eq) were weighed and dissolved in 213 mL of dioxane/water (v/v=50:1), and potassium carbonate (42.74 mmol, 2 eq) and tetrakis(triphenylphosphine)palladium (3.21 mmol, 0.15 eq) were added. Under nitrogen protection, the reaction was carried out at 95°C for 12 h, and the reaction was monitored by TLC to be complete. Post-treatment: 400 mesh crude silica gel was directly added to the system, the sample was concentrated and mixed, and separated by prep-CC (PE:EA=3:1) to obtain a white solid product 2 with a yield of 60%.

Preparation of intermediate 4-(1-(tert-butyloxycarbonyl)-1H-indol-3-yl)-2-morpholino-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-carboxylic acid tert-butyl ester 3:

Take a 100mL round-bottom flask, dissolve 4-(1-(tert-butyloxycarbonyl)-1H-indol-3-yl)-2-chloro-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-carboxylic acid tert-butyl ester 2 (4.74mmol) and morpholine (23.71mmol, 5eq) in 47ml DMSO, and then add DIPEA (23.71mmol, 5eq). Under argon protection, gradually heat to 130℃, react for 5h, and monitor the reaction by TLC spot plate. Post-treatment: After the reaction is cooled to room temperature, the sample is mixed with 400 mesh coarse silica gel, and separated by prep-CC (PE:EA＝3:1) to obtain a white solid product 3 with a yield of 67%.

Preparation of intermediate 4-(4-(1H-indol-3-yl)-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidin-2-yl)morpholine 4:

Take a 100mL round-bottom flask, add 4-(1-(tert-butyloxycarbonyl)-1H-indol-3-yl)-2-morpholino-5,8-dihydropyrido[3,4-d]pyrimidine-7(6H)-carboxylic acid tert-butyl ester 3 (4.29mmol), dissolve in 5mL DCM, and then slowly add 5mL of 4mol/L hydrochloric acid dioxane solution. React at room temperature overnight, and monitor the reaction completion by TLC spot plate. Post-treatment: The reaction solution is directly concentrated and dried under reduced pressure to obtain a white solid powder of hydrochloride, which is directly used in the next step.

Preparation of the final product 4-(4-(1H-indol-3-yl)-7-R ₁ -5,6,7,8-tetrahydropyrido[3,4-d]pyrimidin-2-yl)morpholine 5-X:

Method 1: Take a 25mL round-bottom flask, dissolve 4-(4-(1H-indol-3-yl)-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidin-2-yl)morpholine 4 (149.07umol) in 4mL DCM, add the corresponding anhydride or acid chloride (163.98umol), place the flask in an ice-water bath, and then add triethylamine (223.61mmol), react for 5h, and monitor the reaction completion by TLC spot plate. Post-treatment: The reaction solution is concentrated and separated by prep-CC (PE:EA=2:1-0:1) to obtain the product 5-X with a yield of 60-95%.

Method 2: Take a 25mL round-bottom flask, dissolve 4-(4-(1H-indol-3-yl)-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidin-2-yl)morpholine 4 (149.07umol) and the corresponding acid (149.07umol) in 4mL DCM, add HATU (223.61mmol) and triethylamine (298.14mmol). React for two hours at room temperature, and monitor the completion of the reaction by TLC spot plate. Post-treatment: The reaction solution is concentrated and separated by prep-CC (PE:EA=2:1-0:1) to obtain the product 5-X with a yield of 60-95%.

Example molecule and NMR data:

The final product 1-(4-(1H-indol-3-yl)-2-morpholino-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)-2,2-difluoroethane-1-one (5-1) was a white solid with a yield of 78%. ¹ H NMR (400MHz, DMSO-d ₆ ) δ 11.76 (s, 1H), 8.28 (t, J = 8.0Hz, 1H), 7.92 (d, J = 26.1Hz, 1H), 7.48 (d, J = 7.8Hz, 1H), 7.17 (dt, J = 20.3, 7.0Hz, 2H), 6.88 (td, J = 5 2.7, 14.0Hz, 1H), 4.62 (s, 1H), 4.55 (s, 1H), 3.74 (d, J = 17.0Hz, 10H), 2.97 (t, J = 5.1Hz, 1H), 2.91 (t, J = 5.3Hz, 1H).

The final product 1-(4-(1H-indol-3-yl)-2-morpholino-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)ethan-1-one (5-2) was a white solid with a yield of 82%. ¹ H NMR (400MHz, DMSO-d ₆ ) δ11.77(d,J＝9.3Hz,1H),8.28(d,J＝5.1Hz,1H),7.92(d,J＝7.5Hz,1H),7.48(d,J＝7.6Hz,1H),7.16(dt,J＝20.6,7.1Hz,3H),4.51( d,J＝18.9Hz,2H),3.87–3.54(m,12H),2.95(s,1H),2.83(s,1H),2.13(d,J＝8.0Hz,3H). ¹³ CNMR(101MHz,DMSO-d ₆ )δ169.05,162.40,160.14,136.64,129.96,129.74,126.87,122.52,122.20,120.85,113.08,112.23,66.56,50.54,46.86,44.74,44.18,26.96, 22.16,21.58.

The final product 1-(4-(1H-indol-3-yl)-2-morpholino-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)prop-2-en-1-one (5-3) was an off-white solid with a yield of 72%. ¹ H NMR (400MHz, DMSO-d ₆ ) δ11.74(s,1H),8.28(d,J＝6.8Hz,1H),7.93(d,J＝12.1Hz,1H),7.47(d,J＝7.9Hz,1H),7.16(dt,J＝20.5,7.0Hz,2H),6.93(dt,J＝17 .1,8.5Hz,1H),6.26–6.11(m,1H),5.75(t,J＝10.7Hz,1H),4.62(d,J＝32.7Hz,2H),3.74(d,J＝15.9Hz,10H),2.91(d,J＝21.0Hz,2H).

The final product 1-(4-(1H-indol-3-yl)-2-morpholino-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)-2-methylpropan-1-one (5-4) was an off-white solid with a yield of 75%. ¹ H NMR (400MHz, DMSO-d ₆ ) δ 11.74 (s, 1H), 8.28 (d, J = 7.0Hz, 1H), 7.95 (s, 1H), 7.47 (d, J = 7.7Hz, 1H), 7.16 (dt, J = 20.5, 7.0Hz, 2H), 4.54 (d, J = 40.2Hz, 2H), 3.73(d,J＝13.3Hz,10H),3.06–2.90(m,2H),2.83(s,1H),1.06(dd,J＝17.1,6.5Hz,H).

The final product (4-(1H-indol-3-yl)-2-morpholino-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(cyclopropyl)methanone (5-5) was an off-white solid with a yield of 80%. ¹ H NMR (400MHz, DMSO-d ₆ ) δ11.74(s,1H),8.29(s,1H),7.94(d,J＝2.5Hz,1H),7.48(d,J＝7.8Hz,1H),7.16(dt,J＝20.3,7.0Hz,2H),4.77(s,1H),4.51(s,1H ), 3.91 (s, 1H), 3.73 (d, J = 16.4Hz, 11H), 2.97 (s, 1H), 2.84 (s, 1H), 2.09 (s, 1H), 0.77 (d, J = 8.0Hz, 7H).

The final product 1-(4-(1H-indol-3-yl)-2-morpholino-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)-2,2-dimethylpropan-1-one (5-6) was an off-white solid with a yield of 78%. ¹ H NMR (400MHz, DMSO-d ₆ ) δ11.74(s,1H),8.31(d,J＝7.8Hz,1H),7.97(d,J＝2.8Hz,1H),7.48(d,J＝7.9Hz,1H),7.16(dt,J＝20.0,6.9Hz,2H),4.55(s,2H),3. 81(t,J＝5.3Hz,2H),3.73(d,J＝9.6Hz,8H),2.90(t,J＝5.0Hz,2H),2.51(s,1H),1.27(s,9H).

The final product (4-(1H-indol-3-yl)-2-morpholino-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(phenyl)methanone (5-7) was an off-white solid with a yield of 70%. ¹ H NMR (400 MHz, DMSO-d ₆ ) δ 11.74 (s, 1H), 8.29 (d, J=7.7 Hz, 1H), 7.97 (s, 1H), 7.60–7.44 (m, 6H), 7.16 (dt, J=20.0, 7.0 Hz, 2H), 4.65 (s, 1H), 4.45 (s, 1H), 3.97–3.44 (m, 10H), 2.95 (s, 2H).

The final product 4-(4-(1H-indol-3-yl)-7-(methylsulfonyl)-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidin-2-yl)morpholine (5-8) was a light yellow solid with a yield of 74%. ¹ H NMR (400 MHz, DMSO-d ₆ ) δ 11.91 (s, 1H), 8.30 (d, J=7.7 Hz, 1H), 7.99 (s, 1H), 7.49 (d, J=7.8 Hz, 1H), 7.16 (dt, J=19.8, 7.0 Hz, 2H), 4.24 (s, 2H), 3.73 (d, J=10.9 Hz, 8H), 3.40 (d, J=16.0 Hz, 2H), 3.01 (s, 5H).

The final product 4-(4-(1H-indol-3-yl)-7-((trifluoromethyl)sulfonyl)-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidin-2-yl)morpholine (5-9) was a light yellow solid with a yield of 83%. ¹ H NMR (400 MHz, DMSO-d ₆ ) δ 11.80 (s, 1H), 8.27 (d, J=7.8 Hz, 1H), 7.99 (d, J=2.8 Hz, 1H), 7.48 (d, J=7.9 Hz, 1H), 7.17 (dt, J=19.5, 6.9 Hz, 2H), 4.53 (s, 2H), 3.74 (dt, J=17.3, 10.1 Hz, 19H), 3.03 (s, 2H).

The final product (R)-4-(4-(1H-indol-4-yl)-7-trifluoroacetyl-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidin-2-yl)-3-methylmorpholine (5-10) was an off-white solid with a yield of 75%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.29(s,1H),7.51(d,J＝7.9Hz,1H),7.41(s,1H),7.18(t,J＝7.6Hz,1H),7.10(d,J＝7.0Hz,1H),6.37(s,1H),4.66(td,J＝18.0,7.4Hz,3H),4.26(d,J＝12.7Hz,1H),3 .90(d,J＝11.2Hz,1H),3.72(dd,J＝18.4,9.5Hz,3H),3.57(d,J＝10.7Hz,1H),3.42(t,J＝11.6Hz,1H),3.16(t,J＝12.2Hz,1H),2.72–2.56(m,2H),1.21(d, J=6.7Hz,3H).

The final product (R)-4-(4-(1H-indol-4-yl)-7-acetyl-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidin-2-yl)-3-methylmorpholine (5-11) was a white solid with a yield of ⁷⁰ %. NMR(400MHz,Chloroform-d)δ8.59(s,1H),7.47(d,J＝7.9Hz,1H),7.26(s,1H),7.16(d,J＝7.0Hz,1H),6.52(s,1H),4.80(d,J＝16.6Hz,1H),4.60(s,1H),4.40(d, J＝13.3Hz,1H),3.97(d,J＝6.8Hz,1H),3.73(t,J＝18.7Hz,3H),3.66–3.51(m,3H),3.37–3.21(m,1H),2.65(t,J＝18.9Hz,2H),2.20(d,J＝28.9Hz,3H),1. 33(d,J=6.7Hz,3H).

The final product (R)-4-(4-(1H-indol-4-yl)-7-methanesulfonyl-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidin-2-yl)-3-methylmorpholine (5-12) was a white solid with a yield of 70%. ¹ H NMR(400MHz,Chloroform-d)δ8.45(s,1H),7.48(d,J＝8.0Hz,1H),7.24(s,1H),7.16(d,J＝7.2Hz,1H),6.49(s,1H),4.84–4.71(m,1H),4.43(d,J＝15.9Hz,2H), 3.93(dd,J＝27.3,10.8Hz,2H),3.78–3.67(m,3H),3.56(s,1H),3.46(dd,J＝11.7,5.6Hz,1H),3.34(dd,J＝21.1,14.5Hz,3H),2.89(s,3H),1.33(d,J＝6.7 Hz,3H).

The final product (R)-4-(4-(1H-indol-3-yl)-7-trifluoroacetyl-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidin-2-yl)-3-methylmorpholine (5-13) was an off-white solid with a yield of 78%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.75(s,1H),8.29(dd,J＝7.4,3.6Hz,1H),7.96(dd,J＝7.7,2.9Hz,1H),7.49(d,J＝8.3Hz,1H),7.17(dt,J＝20.6,7.8Hz,2H),4.76–4.67(m,1H),4.66–4.59(m,1H),4.30(d,J＝16.9Hz,1H),4.0 0–3.92(m,1H),3.91–3.79(m,2H),3.76(d,J＝13.0Hz,1H),3.63(dd,J＝12.7,3.4Hz,1H),3.48(t,J＝10.3Hz,1H),3.42–3.36(m,1H),3.27–3.17(m,1H) ,3.05–2.94(m,2H),1.26(d,J＝6.8Hz,3H).

The final product (R)-4-(4-(1H-indol-3-yl)-7-difluoroacetyl-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidin-2-yl)-3-methylmorpholine (5-14) was an off-white solid with a yield of 72%. ¹ H NMR (400 MHz, Chloroform-d) δ8.48 (s, 1H), 8.29 (d, J = 7.9 Hz, 1H), 7.56 (s, 1H), 7.45 (d, J = 8.1 Hz, 1H), 7.29 (dd, J = 13.8, 5.1 Hz, 2H), 6.20 (td, J = 53.7, 52.9, 9.3 Hz, 1H), 4.87–4.80 (m, 1H), 4.77–4.66 (m, 2H), 4.65–4. 59(m,1H),4.47–4.41(m,1H),4.01(dd,J＝11.2,2.3Hz,1H),3.87–3.82(m,1H),3.77(dd,J＝10.6,3.7Hz,1H),3.60(td,J＝11.6,3.0Hz,1H),3.39–3.33( m,1H),3.32–3.27(m,1H),3.00–2.92(m,2H),1.29–1.24(m,4H).

The final product (R)-4-(4-(1H-indol-3-yl)-7-methylsulfonyl-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidin-2-yl)-3-methylmorpholine (5-15) was a light yellow solid with a yield of 74%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.72(s,1H),8.31(dd,J＝7.4,3.6Hz,1H),7.94(dd,J＝7.7,2.9Hz,1H),7.48(d,J＝8.3Hz,1H),7.14(dt,J＝20.6,7.8Hz,2H),4.74–4.67(m,1H),4.68–4 .59(m,1H),4.40(d,J＝16.9Hz,1H),4.00–3.9 2(m,1H),3.91–3.79(m,2H),3.76(d,J＝13.0Hz,1H),3.63(dd,J＝12.7,3.4Hz,1H),3.48(t,J＝10.3Hz,1H),3.42–3.36(m,1H),3.27–3.17(m,1H),3.08 (s,5H), 3.05–2.94(m,2H),1.26(d,J＝6.8Hz,3H).

The final product (R)-1-(4-(1H-indol-3-yl)-2-(3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)butane-1,3-dione (5-16) was an off-white solid with a yield of 63%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.67(s,1H),8.31(d,J＝7.9Hz,1H),7.90(d,J＝2.6Hz,1H),7.47(d,J＝8.0Hz,1H),7.14(dt,J＝23.1,6.9Hz,2H),4.68(d,J＝4.6Hz,1H),4.27(d,J＝13.3Hz,1H),3.94( d,J＝11.4Hz,1H),3.74(d,J＝6.2Hz,3H),3.64–3.59(m,1H),3.52–3.44(m,6H),3.17(t,J＝11.1Hz,1H),2.96(h,J＝6.7Hz,2H),2.81–2.71(m,2H),1.23( d,J＝6.6Hz,3H).

The final product (R)-4-(4-(1H-indol-3-yl)-7-ethoxyacryloyl-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidin-2-yl)-3-methylmorpholine (5-17) was an off-white solid with a yield of 65%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.73(s,1H),8.30(d,J＝7.7Hz,1H),7.92(d,J＝2.2Hz,1H),7.48(dd,J＝9.7,7.2Hz,2H),7.16(dt,J＝21.3,6.9Hz,2H),6.00(t,J＝11.1Hz,1H),4.76–4. 69(m,1H),4.57(q,J＝23.7,20.7Hz,2H),4.3 1(d,J＝12.5Hz,1H),3.98(p,J＝7.3Hz,3H),3.76(td,J＝17.2,7.0Hz,3H),3.63(dd,J＝11.2,2.6Hz,1H),3.47(td,J＝11.8,2.5Hz,1H),3.21(td,J＝13.0,3. 1Hz, 1H), 2.88 (s, 2H), 1.26 (d, J = 8.4Hz, 6H).

The final product (R)-4-(4-(1H-indol-3-yl)-7-isopropanoyl-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidin-2-yl)-3-methylmorpholine (5-18) was an off-white solid with a yield of 72%. ¹ H NMR (400 MHz, DMSO-d ₆ ) δ 11.74 (s, 1H), 8.30 (d, J = 6.8 Hz, 1H), 7.95 (s, 1H), 7.48 (d, J = 7.6 Hz, 1H), 7.16 (dt, J = 21.0, 7.1 Hz, 2H), 4.71 (s, 1H), 4.59 (s, 1H), 4.55–4.41 (m, 1H), 4.30 (d, J = 12.5 Hz, 1H), 3.96 (d ,J＝10.8Hz,1H),3.76(d,J＝10.5Hz,2H),3.66(dd,J＝24.8,8.9Hz,2H),3.47(t,J＝10.8Hz,1H),3.28–3.13(m,1H),3.04–2.79(m,3H),1.28–1.22(m,3H), 1.06(dd,J=16.6,6.2Hz,6H).

The final product (R)-1-(4-(1H-indol-3-yl)-2-(3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)-2-methylprop-2-en-1-one (5-19) was a white solid with a yield of 80%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.75(s,1H),8.32(d,J＝7.7Hz,1H),7.97(s,1H),7.49(d,J＝7.8Hz,1H),7.17(dt,J＝20.5,7.1Hz,1H),5.30(s,1H),5.15(s,1H),4.77–4.66(m,2H) ,4.58(d,J＝16.8Hz,1H),4.30(d, J＝13.0Hz,1H),3.95(d,J＝9.1Hz,1H),3.76-3.61(m,3H),3.47(t,J＝10.8Hz,1H),3.37(t,J＝10.8Hz,1H),3.21(t,J＝12.6Hz,1H),3.3-2.91(m,2H),1.93 (s,3H),1.26(d,J=6.6Hz,3H).

The final product (R)-1-(4-(1H-indol-3-yl)-2-(3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)-2-fluoro-2-methylpropane-1-one (5-20) was a white solid with a yield of 73%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.75(s,1H),8.32(d,J＝7.7Hz,1H),7.97(s,1H),7.49(d,J＝7.8Hz,1H),7.17(dt,J＝20.5,7.1Hz,1H),4.87–4.70(m,2H),4.62-4.46(m,1H),4.30(d ,J＝13.0Hz,1H),3.96-3.70(m,4H),3.66-3.60(m,1H),3.46(t,J＝10.8Hz,1H),3.21(t,J＝12.6Hz,1H),3.03-2.91(m,2H),δ1.61(d,J＝21.3Hz,6H),1. 26(d,J=5.3Hz,3H).

The final product (R)-(4-(1H-indol-3-yl)-2-(3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(1-fluorocyclopropylmethanone (5-21) was a white solid with a yield of 83%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.74(s,1H),8.31(d,J＝7.8Hz,1H),7.95(s,1H),7.49(d,J＝8.0Hz,1H),7.17(dt,J＝20.5,6.9Hz,2H),4.72(d,J＝4.6Hz,2H),4.51(m,1H),4.31(d,J ＝12.3Hz,1H),4.02-3.72(m,4H),3.63(dd,J＝11.3,2.7Hz,1H),3.47(t,J＝11.7Hz,1H),3.30–3.15(m,1H),3.05-2.95(d,2H),1.40–1.17(m,7H).

The final product 1-(4-(1H-indol-3-yl)-2-((R)-3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)-2-methylbutan-1-one (5-22) was an off-white solid with a yield of 64%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.75(s,1H),8.32(t,J＝7.4Hz,1H),7.95(d,J＝4.8Hz,1H),7.50(d,J＝7.2Hz,1H),7.17(dt,J＝20.4,7.0Hz,2H),4.72(s,1H),4.52(dd,J＝42.0,18.9Hz, 2H),4.31(d,J＝11.9Hz,1H),3.95(d,J＝11.0Hz,1H),3.86–3.58(m,4H ),3.47(t,J＝11.3Hz,1H),3.28–3.15(m,1H),2.93(s,1H),2.81(dd,J＝13.3,6.5Hz,2H),1.63(dt,J＝12.8,6.4Hz,1H),1.40–1.31(m,1H),1.25(q,J＝8. 2, 6.9Hz, 3H), 1.04 (dd, J = 18.3, 7.3Hz, 3H), 0.84 (dt, J = 12.2, 7.1Hz, 3H).

The final product (R)-(4-(1H-indol-3-yl)-2-(3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(cyclobutyl)methanone (5-23) was an off-white solid with a yield of 78%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.73(s,1H),8.29(d,J＝7.0Hz,1H),7.92(d,J＝11.4Hz,1H),7.49(d,J＝7.9Hz,1H),7.16(dt,J＝20.8,7.0Hz,2H),4.76–4.64(m,1H),4.57–4.37(m,2H),4.30(d,J＝13.4Hz,1H),3.9 5(d,J＝11.1Hz,1H),3.75(d,J＝11.0Hz,1H),3.69–3.41(m,4H),3.26–3.13(m,1H),2.98–2.76(m,2H),2.31–2.07(m,4H),2.00–1.87(m,1H),1.85-1 .74(m,1H),1.25(d,J＝6.5Hz,6H).

The final product (R)-(4-(1H-indol-3-yl)-2-(3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(cyclopentyl)methanone (5-24) was an off-white solid with a yield of 83%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.74(s,1H),8.30(d,J＝7.1Hz,1H),7.94(s,1H),7.49(d,J＝7.8Hz,1H),7.16(dt,J＝21.0,7.0Hz,2H),4.76-4.67(m,1H),4.50(dd,J＝42.9,18.8Hz,2H),4.35-4.26(m,1H),3.95(d,J＝1 1.0Hz,1H),3.84–3.57(m,4H),3.47(t,J＝11.5Hz,1H),3.20(td,J＝14.9,12.3,5.6Hz,1H),3.10(p,J＝7.4Hz,1H),2.99–2.87(m,1H),2.83(s,1H),1.8 9–1.48(m,8H),1.25(d,J＝6.5Hz,3H).

The final product (R)-(4-(1H-indol-3-yl)-2-(3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(cyclohexyl)methanone (5-25) was an off-white solid with a yield of 75%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.74(s,1H),8.29(d,J＝7.2Hz,1H),7.95(s,1H),7.48(d,J＝7.8Hz,1H),7.16(dt,J＝21.4,7.0Hz,2H),4.71(s,1H),4.60–4.38(m,2H),4.29(d,J＝12.0Hz,1H),3.96(d,J＝11.0Hz,1H),3 .81–3.57(m,3H),3.47(t,J＝11.6Hz,1H),3.36(t,J＝11.6Hz,1H),3.23–3.16(m,1H),2.96–2.80(m,2H),2.71(t,J＝8.4Hz,1H),1.71(dd,J＝21.4,8.1Hz ,5H),1.32(dt,J＝47.1,8.6Hz,8H).

The final product (R)-1-(4-(1H-indol-3-yl)-2-(3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)-2,2-dimethylpropane-1-one (5-26) was an off-white solid with a yield of 65%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.73(s,1H),8.32(d,J＝7.8Hz,1H),7.97(d,J＝2.6Hz,1H),7.48(d,J＝7.9Hz,1H),7.16(dt,J＝21.2,7.1Hz,2H),4.71(d,J＝6.4Hz,1H),4.55(q,J＝18. 2Hz,2H),4.30(d,J＝12.7Hz,1H), 3.96(d,J＝11.1Hz,1H),3.90–3.82(m,1H),3.76(d,J＝11.5Hz,2H),3.63(d,J＝11.2Hz,1H),3.47(t,J＝11.7Hz,1H),3.20(t,J＝12.8Hz,1H),2.90(d,J＝4 .9Hz, 2H), 1.26 (d, J = 10.3Hz, 12H).

The final product (R)-1-(4-(1H-indol-3-yl)-2-(3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)-3-chloro-2,2-dimethylpropane-1-one (5-27) was an off-white solid with a yield of 68%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.74(s,1H),8.33(d,J＝7.7Hz,1H),7.96(s,1H),7.49(d,J＝8.0Hz,1H),7.17(dt,J＝21.1,6.9Hz,2H),4.76–4.64(m,1H),4.55(q,J＝18.2Hz,2H),4. 31(d,J＝12.6Hz,1H),3.99–3.72(m,6H),3.63(d,J＝13.4Hz,1H),3.47(t,J＝10.5Hz,1H),3.21(t,J＝12.8Hz,1H),3.01-2.84(m,2H),1.37(s,6H),1.26 (d,J=6.7Hz,3H).

The final product (R)-1-(4-(1H-indol-3-yl)-2-(3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)-2,2-dimethylbutan-1-one (5-28) was an off-white solid with a yield of 68%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.76(s,1H),8.32(d,J＝7.6Hz,1H),7.96(s,1H),7.48(d,J＝7.8Hz,1H),7.16(dt,J＝21.2,6.9Hz,2H),4.75–4.64(m,1H),4.55(q,J＝18.3Hz,2H),4. 30(d,J＝12.9Hz,1H),3.95(d,J＝9.5Hz,1H),3.87(d,J＝13.2Hz, 1H),3.76(d,J＝10.5Hz,1H),3.63(d,J＝10.2Hz,1H),3.47(t,J＝11.0Hz,1H),3.41–3.37(t,J＝11.0Hz,1H),3.20(t,J＝12.3Hz,1H),3.02–2.80(m,2H),1 .66(d,J＝7.2Hz,2H),1.37–1.15(m,9H),0.81(t,J＝7.0Hz,3H).

The final product (R)-(4-(1H-indol-3-yl)-2-(3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(1-methylcyclopropyl)methanone (5-29) was an off-white solid with a yield of 75%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.75(s,1H),8.31(d,J＝7.7Hz,1H),7.97(s,1H),7.48(d,J＝8.0Hz,1H),7.16(dt,J＝21.3,6.9Hz,3H),4.71-4.70(m,1H),4.65-4.465(m,1H),4.30 (d,J＝12.5Hz,1H),3.95(d,J＝10.9Hz,1H),3.91-3.8 1(m,1H),3.76(d,J＝11.2Hz,2H),3.63(d,J＝11.1Hz,1H),3.47(t,J＝12.7Hz,1H),3.40(d,J＝7.0Hz,1H),3.20(t,J＝14.4Hz,1H),2.93(m,2H),1.31(s,3H ), 1.25 (d, J = 6.6Hz, 3H), 0.88 (s, 2H), 0.61 (s, 2H).

The final product (R)-(4-(1H-indol-3-yl)-2-(3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(1-(trifluoromethyl)cyclopropyl)methanone (5-30) was an off-white solid with a yield of 64%. ¹ H NMR (400MHz, DMSO-d ₆ ) δ11.77(s,1H),8.31(d,J＝7.8Hz,1H),7.95(s,1H),7.49(d,J＝7.9Hz,1H),7.17(dt,J＝20.9,7.1Hz,2H),4.76–4.45(m,3H),4.30( d,J＝12.7Hz,1H),4.02–3.71(m,4H),3.63(d,J＝11.2Hz,1H),3.53–3.32(m,1H),3.21(t,J＝12.8Hz,1H),3.06–2.84(m,2H),1.26(q,J＝15.4,11.6Hz,7H) .

The final product (R)-1-(4-(1H-indol-3-yl)-2-(3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)-3,3,3-trifluoro-2,2-dimethylpropan-1-one (5-31) was an off-white solid with a yield of 77%. ¹ H NMR (400 MHz, DMSO-d ₆ ) δ 11.76 (s, 1H), 8.33 (d, J = 7.7 Hz, 1H), 7.97 (s, 1H), 7.49 (d, J = 7.9 Hz, 1H), 7.17 (dt, J = 20.6, 7.0 Hz, 2H), 4.77-4.67 (m, 1H), 4.57 (q, J = 18.2 Hz, 2H), 4.31 (d, J = 12.6 Hz,1H),4.05–3.84(m,2H),3.84–3.74(m,2H),3.69–3.59(m,1H),3.53–3.43(m,1H),3.21(t,J＝14.3Hz,1H),3.04–2.87(m,2H),1.56(s,6H),1.26(d ,J=6.6Hz,3H).

The final product (R)-(4-(1H-indol-3-yl)-2-(3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(2,2,3,3-tetramethylcyclopropyl)methanone (5-32) was a white solid with a yield of 72%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.73(s,1H),8.31(d,J＝5.6Hz,1H),7.95(d,J＝2.7Hz,1H),7.48(t,J＝7.5Hz,1H),7.16(dt,J＝21.1,7.0Hz,2H),4.71(d,J＝6.4Hz,1H),4.56(s,1H),4 .53–4.39(m,1H),4.30(d,J＝13.0Hz,1H),3.96(d,J＝10.8Hz,1H), 3.76(d,J＝9.1Hz,2H),3.63(d,J＝10.4Hz,2H),3.48(t,J＝13.1Hz,1H),3.21(t,J＝12.6Hz,1H),3.02–2.90(m,1H),2.81(s,1H),1.25(d,J＝5.0Hz,3H),1 .19(d,J＝9.9Hz,6H), 1.13(d,J＝3.5Hz,3H), 1.05(d,J＝20.5Hz,3H).

The final product (4-(1H-indol-3-yl)-2-((R)-3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(2,2-difluoro-1-methylcyclopropyl)methanone (5-33) was a white solid with a yield of 79%. NMR (400MHz, DMSO-d6) δ11.88(s,1H),8.31(d,J＝7.8Hz,1H),7.97(d,J＝7.5Hz,1H),7.49(t,J＝8.0Hz,1H),7.16(dt,J＝21.4,7.2Hz,2H),4.74–4.67(m,1H),4 .60(dt,J＝20.7,9.6Hz,1H),4.52–4.39(m,1H),4.35–4.2 6(m,1H),3.96(d,J＝10.5Hz,1H),3.76(d,J＝11.2Hz,1H),3.66–3.42(m,4H),3.19(dd,J＝23.8,11.2Hz,2H),2.93–2.83(m,1H),1.85(d,J＝7.0Hz,1H),1 .65(d,J＝6.1Hz,1H),1.45(d,J＝23.3Hz,3H),1.28–1.22(m,3H).

The final product (R)-1-(4-(1H-indol-3-yl)-2-(3-methylmorpholino)-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidine-7-carbonyl)cyclopropane-1-carbonitrile (5-34) was a white solid with a yield of 64%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.92(s,1H),8.29(d,J＝7.8Hz,1H),7.91(s,1H),7.50(d,J＝7.9Hz,1H),7.16(dt,J＝21.3,7.3Hz,2H),4.71(d,J＝7.0Hz,2H),4.66-4.45(m,1H),4.31(d,J＝13.0Hz,1H),4.11 –3.89(m,2H),3.76(d,J＝11.2Hz,1H),3.63(dd,J＝11.2,2.7Hz,1H),3.52–3.41(m,2H),3.21(q,J＝9.8,7.8Hz,1H),3.12–2.85(m,2H),1.70–1.56(m,4H ), 1.26 (d, J = 6.7Hz, 3H).

The final product (4-(1H-indol-3-yl)-2-((R)-3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(adamantan-1-yl)methanone (5-35) was an off-white solid with a yield of 84%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.72(s,1H),8.33(d,J＝7.8Hz,1H),7.98(s,1H),7.49(d,J＝8.0Hz,1H),7.16(dt,J＝21.4,6.8Hz,2H),4.75–4.64(m,1H),4.56(q,J＝18.3Hz,2H),4. 30(d,J＝12.3Hz,1H),4.00–3.89(m,2H), 3.83–3.78(m,1H),3.76(d,J＝11.2Hz,1H),3.63(d,J＝13.7Hz,1H),3.47(t,J＝11.6Hz,1H),3.20(t,J＝12.8Hz,1H),3.01-2.84(m,2H),1.98(d,J＝6.3Hz, 9H), 1.71 (s, 6H), 1.25 (d, J = 6.7Hz, 3H).

The final product (S)-(4-(1H-indol-3-yl)-2-(3-methylmorpholino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(1-(trifluoromethyl)cyclopropyl)methanone (5-36) was a light yellow solid with a yield of 35%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ12.25(s,1H),8.31(d,J＝7.7Hz,1H),8.12(d,J＝2.5Hz,1H),7.52(d,J＝7.7Hz,1H),7.20(dt,J＝15.5,6.8Hz,2H),4.89–4.70(m,2H),4.39(d,J＝13.0Hz,1H),3.98 (d,J＝11.0Hz,1H),3.90(s,1H),3.78(d,J＝11.4Hz,1H),3.65(d,J＝13.6Hz,1H),3.50(t,J＝11.5Hz,1H),3.33(d,J＝15.8Hz,1H),2.96(s,2H),1.31(d,J＝ 6.7Hz,3H).

The final product (4-(1H-indol-3-yl)-2-morpholino-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(1-(trifluoromethyl)cyclopropyl)methanone (5-37) was obtained as an off-white solid with a yield of 32%. ¹ H NMR (400MHz, DMSO-d ₆ ) δ12.18(s,1H),8.30(d,J＝7.6Hz,1H),8.10(d,J＝2.7Hz,1H),7.52(d,J＝7.7Hz,1H),7.20(dt,J＝15.7,6.8Hz,2H),4.70(s,2H),3 .74(d,J＝4.4Hz,10H),2.96(s,2H),1.37(d,J＝4.0Hz,2H),1.32(s,2H).

The final product (S)-(2-(3-aminopiperidin-1-yl)-4-(1H-indol-3-yl)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(1-(trifluoromethyl)cyclopropyl)methanone (5-38) was an off-white solid with a yield of 38%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ12.06(s,1H),8.42(s,2H),8.29(d,J＝6.8Hz,1H),8.04(d,J＝2.4Hz,1H),7.51(d,J＝8.2Hz,1H),7.25–7.15(m,2H),4.65–4.60(m,2H),4.37(d,J＝13.3Hz,1H),3 .86(s,2H),3.44–3.32(m,1H),3.31–3.17(m,2H),2.95(s,2H),2.08(s,1H),1.87(d,J＝12.7Hz,1H),1.72(q,J＝9.3Hz,1H),1.64–1.50(m,1H),1.43 –1.25(m,4H).

The final product (R)-(2-(3-aminopiperidin-1-yl)-4-(1H-indol-3-yl)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(1-(trifluoromethyl)cyclopropyl)methanone (5-39) was an off-white solid with a yield of 42%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ12.21(s,1H),8.54(s,3H),8.35–8.24(m,1H),8.06(d,J＝2.6Hz,1H),7.57–7.46(m,1H),7.20(dt,J＝9.7,5.3Hz,2H),4.86–4.55(m,3H),4.39(d,J＝12.9Hz,1H),3.86(s,3H),3 .45–3.33(m,1H),3.33–3.15(m,2H),2.95(s,2H),2.55(s,1H),2.10(d,J＝8.7Hz,1H),1.91–1.82(m,1H),1.80–1.68(m,1H),1.57(q,J＝13.5,12.0 Hz, 1H), 1.35 (d, J = 20.9Hz, 4H).

The final product (4-(1H-indol-3-yl)-2-(methyl(pyrrolidin-3-yl)amino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(1-(trifluoromethyl)cyclopropyl)methanone (5-40) was an off-white solid with a yield of 46%. ¹ H NMR (400MHz, DMSO-d ₆ ) δ12.02 (s, 1H), 8.41–8.29 (m, 1H), 7.95 (d, J = 2.6Hz, 1H), 7.55–7.45 (m, 1H), 7.27–7.05 (m, 2H), 5.64–5.49 (m, 1H), 4.73–4.47 (m,3H),3.85(s,3H),3.18(d,J＝20.3Hz,6H),2.93(s,1H),2.18–2.10(m,1H),2.05–1.93(m,1H),1.39–1.28(m,4H).

The final product (R)-(4-(1H-indol-3-yl)-2-(3-(methylamino)pyrrolidin-1-yl)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(1-(trifluoromethyl)cyclopropyl)methanone (5-41) was an off-white solid with a yield of 38%. ¹ HNMR (400MHz, DMSO-d ₆ ) δ11.97(s,1H),8.49(d,J＝7.8Hz,1H),7.96(d,J＝2.8Hz,1H),7.49(d,J＝7.8Hz,1H),7.16(dt,J＝18.5,6.8Hz,2H),4.58(s,2H),4 .00–3.73(m,6H),3.64(d,J＝8.5Hz,1H),2.94(s,2H),2.60(s,3H),2.33(ddt,J＝27.4,12.8,6.6Hz,2H),1.34(d,J＝26.6Hz,4H).

The final product (S)-(2-(3-aminopyrrolidin-1-yl)-4-(1H-indol-3-yl)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(1-(trifluoromethyl)cyclopropyl)methanone (42) was obtained as an off-white solid with a yield of 38%. ¹ H NMR (400MHz, DMSO-d ₆ ) δ11.93(s,1H),8.50(d,J＝7.9Hz,1H),7.97(s,1H),7.49(d,J＝7.9Hz,1H),7.16(dt,J＝22.4,7.0Hz,2H),4.58(s,2H),3.99–3.6 3(m,7H),2.94(s,2H),2.34(dq,J＝14.1,7.4Hz,1H),2.17(dd,J＝12.1,6.6Hz,1H),1.35(d,J＝28.0Hz,4H).

The final product (R)-(4-(1H-indol-3-yl)-2-(methyl(pyrrolidin-3-yl)amino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(1-(trifluoromethyl)cyclopropyl)methanone (5-43) was an off-white solid with a yield of 28%. ¹ HNMR (400MHz, DMSO-d ₆ ) δ11.76(s,1H),8.36(d,J＝8.1Hz,1H),7.93(s,1H),7.48(d,J＝7.8Hz,1H),7.22–7.11(m,2H),5.46–5.39(m,1H),4.67–4.46(m,3 H),3.90–3.78(m,2H),3.08(q,J＝11.7,10.9Hz,4H),2.96–2.79(m,4H),2.03–1.97(m,1H),1.84–1.75(m,1H),1.41–1.28(m,4H).

The final product (2-(1,1-dioxothiomorpholino)-4-(1H-indol-3-yl)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(1-(trifluoromethyl)cyclopropyl)methanone (5-44) was obtained as an off-white solid with a yield of 48%. ¹ H NMR (400MHz, DMSO-d ₆ ) δ11.84(s,1H),8.21(d,J＝7.6Hz,1H),7.98(s,1H),7.49(d,J＝7.8Hz,1H),7.17(ddt,J＝14.4,7.4,3.7Hz,2H),4.61(s,2H),4.29 (s,4H),3.85(s,3H),3.21(s,3H),2.96(s,2H),2.55(s,2H),1.41–1.28(m,4H).

The final product (2-(((3aR, 6aS)-hexahydropyrrolo[3,4-c]pyrrol-2(1H)-yl)-4-(1H-indol-3-yl)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(1-(trifluoromethyl)cyclopropyl)methanone (5-45) was an off-white solid with a yield of 42%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ11.85(s,1H),8.48(d,J＝7.7Hz,1H),7.95(s,1H),7.48(d,J＝7.8Hz,1H),7.16(dt,J＝19.0,7.0Hz,2H),4.56(s,2H),3.96–3.73(m,5H),3.49(s,3H) ,3.13(dd,J＝10.4,6.2Hz,2H),2.93(s,4H),2.81(d,J＝10.9Hz,2H),1.34(d,J＝26.2Hz,4H).

The final product (4-(1H-indol-3-yl)-7-(1-(trifluoromethyl)cyclopropane-1-carbonyl)-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidin-2-yl)piperidine-4-carboxylic acid methyl ester (5-46) was an off-white solid with a yield of 38%. ¹ H NMR (400 MHz, DMSO-d ₆ )δ12.15(s,1H),8.30(d,J＝7.5Hz,1H),8.19–8.11(m,1H),7.53(d,J＝7.8Hz,1H),7.22(dt,J＝14.0,6.4Hz,2H),4.83–4.56(m,4H),3.63(s,3H),3.46(d ,J＝9.9Hz,1H),3.25(t,J＝11.5Hz,2H),2.96(s,2H),2.76(t,J＝11.2Hz,1H),2.71(s,1H),1.98(d,J＝12.5Hz,2H),1.69–1.57(m,2H),1.34(dd,J＝25.9, 6.5Hz, 4H).

The final product (2-(((2R,6S)-2,6-dimethylmorpholino)-4-(1H-indol-3-yl)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(1-(trifluoromethyl)cyclopropyl)methanone (5-47) was an off-white solid with a yield of 33%. ¹ HNMR (400 MHz, DMSO-d ₆ )δ11.76(s,1H),8.28(d,J＝7.8Hz,1H),7.95(d,J＝2.6Hz,1H),7.49(d,J＝7.9Hz,1H),7.17(dt,J＝20.4,6.9Hz,2H),4.57(d,J＝12.2Hz,4H),3.85(s,2H), 3.62(dd,J＝9.5,4.1Hz,2H),2.92(s,2H),2.55(s,2H),1.41–1.27(m,4H),1.19(d,J＝6.1Hz,6H).

The final product (4-(1H-indol-3-yl)-2-(pyrrolidin-3-ylamino)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(1-(trifluoromethyl)cyclopropyl)methanone (5-48) was obtained as an off-white solid with a yield of 35%. ¹ H NMR (400MHz, DMSO-d ₆ ) δ11.85(s,1H),8.52(d,J＝7.8Hz,1H),7.94(s,1H),7.48(d,J＝7.9Hz,1H),7.16(dt,J＝22.3,7.0Hz,2H),4.55(s,2H),3.89–3.58 (m,6H),2.92(s,2H),2.11(dt,J＝11.7,6.0Hz,1H),1.78(dt,J＝12.0,6.0Hz,1H),1.43–1.27(m,4H).

The final product (4-(1H-indol-3-yl)-2-(4-(methylsulfonyl)piperazin-1-yl)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(1-(trifluoromethyl)cyclopropyl)methanone (5-49) was an off-white solid with a yield of 43%. ¹ HNMR (400MHz, DMSO-d ₆ ) δ11.93(s,1H),8.28(d,J＝7.7Hz,1H),7.96(s,1H),7.50(d,J＝7.6Hz,1H),7.24–7.11(m,2H),4.59(s,2H),3.89(d,J＝30.7Hz,6 H), 3.23 (s, 4H), 2.93 (s, 2H), 2.90 (s, 3H), 1.36 (dd, J = 22.3, 12.1Hz, 4H).

The final product (2-((3-azabicyclo[3.1.0]hexane-6-yl)amino)-4-(1H-indol-3-yl)-5,8-dihydropyrido[3,4-d]pyrimidin-7(6H)-yl)(1-(trifluoromethyl)cyclopropyl)methanone (5-50) was an off-white solid with a yield of 38%. ¹ H NMR (400MHz, DMSO-d ₆ ) δ11.87(s,1H),8.42(d,J＝7.8Hz,1H),7.94(s,1H),7.48(d,J＝7.8Hz,1H),7.16(dt,J＝21.4,7.3Hz,2H),4.65–4.49(m,2H),3.8 4(s,2H),3.20(s,4H),2.93(d,J＝6.9Hz,2H),2.03–1.95(m,1H),1.49–1.27(m,6H).

The final product, methyl N-(4-(1H-indol-3-yl)-7-(1-(trifluoromethyl)cyclopropane-1-carbonyl)-5,6,7,8-tetrahydropyrido[3,4-d]pyrimidin-2-yl)-N-methylglycine methyl ester (5-51), was an off-white solid with a yield of 44%. ¹ HNMR (400MHz, DMSO-d ₆ ) δ11.72(s,1H),8.42(d,J＝7.8Hz,1H),7.94(d,J＝2.6Hz,1H),7.48(d,J＝8.0Hz,1H),7.16(dt,J＝21.2,6.9Hz,2H),4.57(s,2H),3 .85(d,J＝11.4Hz,2H),3.43–3.35(m,3H),3.20(s,3H),2.93(s,2H),2.70(s,2H),1.35(d,J＝25.6Hz,4H).

Note: The synthesis of the example molecules is selected from route 1 or route 2.

Evaluation of pharmacological activity of compounds

In vitro kinase assay of tetrahydropyridopyrimidine derivatives

The in vitro kinase assay was performed using the Kinase Profiler service provided by Eurofins. The mixture of the test compound 5-X and the kinase ATR/ATRIP was incubated with 50 nM GST-CMyc-p53 and ATP buffer containing Mg ions, and the reaction was initiated by Mg/ATP. After incubation at room temperature for 30 minutes, the reaction was terminated by adding a stop buffer containing EDTA, and then a detection buffer containing a monoclonal antibody against GST with a d2 tag and a monoclonal antibody against phosphorylated p53 with a Europium tag was added. The reaction plate was placed in a microplate reader, and the time-resolved fluorescence (HTRF) mode was used to calculate the HTRF signal value according to the formula HTRF = (Em665nm/Em620nm)*10000.

Test results: The inhibition of compound 5-X on ATR activity was determined. The results are shown in Table 1 (A represents IC ₅₀ <100 nM, B represents 500 nM>IC ₅₀ >100 nM, C represents 1000 nM>IC ₅₀ >500 nM, and D represents IC ₅₀ >1000 nM).

Table 1 Kinase activity data

Kinase selectivity test of compounds 5-30

Table 2 Inhibition data of compounds 5-30 on 4 off-target targets

Kinase IC ₅₀ (μM) Kinase IC ₅₀ (μM) AAK1 2.326 PI3K (p110d/p85a) 4.945 mTOR/FKBP12 >10 PI3K (p110a/p85a) >10

In the kinase selectivity test, the present invention found that compound 5-30 at a concentration of 10 μM only inhibited 4 kinase targets among the 412 recombinant human protein kinases tested by more than 50%, namely kinase AAK1, PI3K (p110d/p85a), PI3K (p110a/p85a) and mTOR/FKBP12, showing excellent selectivity. The IC ₅₀ values of these 4 kinase targets were further tested, and the results are shown in Table 2. The kinase activity of compound 5-30 was only within 10 μM for 2 of the targets. In general, compound 5-30 has excellent kinase selectivity for 412 recombinant human protein kinases.

In vitro cell experiments on tetrahydropyridopyrimidine derivatives

(1) Cell culture consumables:

Cell culture medium: DMEM medium (Gibco), RPMI-1640 (Gibco); fetal bovine serum (FBS, Hyclone); penicillin solution and streptomycin solution (Invitrogen Biotechnology Company)

Cells: HT-29 (human colorectal adenocarcinoma cells), LoVo (human colorectal adenocarcinoma cells), NCI-H23 (human non-small cell lung cancer cells), A549 (human lung cancer cells), HL60 (human myeloid leukemia cells) were purchased from the American Type Culture Collection (ATCC), and mantle cell lymphoma cells were purchased from Granta519.

Cell culture consumables: trypsin (Invitrogen Biotechnology Company); 6-well plates, 24-well plates, 96-well plates, centrifuge tubes, gun tips, etc. (Corning Company), 20, 100 mm cell culture dishes (WHB Company).

(2) Cell culture:

HT-29 and LoVo cells were cultured in DMEM medium (containing 10% fetal bovine serum, 100U/mL penicillin and 100μg/mL streptomycin); NCI-H23, A549, and HL60 cells were cultured in RPMI1640 medium (containing 10% fetal bovine serum, 100U/mL penicillin and 100μg/mL streptomycin). At the same time, all cells were cultured in a humidified cell incubator at 37°C and 5% _CO2 . During the cell culture process, the cells were digested with trypsin and passaged reasonably to ensure the appropriate cell density. All experiments used cells in the logarithmic growth phase.

(3) MTT assay

A. Solution Configuration

5 mg/mL MTT solution: Dissolve MTT powder in physiological saline to prepare a 5 mg/mL solution. Filter through a 0.22 μm filter membrane and store at 4°C in the dark. Use within 1 month.

20% acidic SDS solution: Weigh 80g of SDS powder and add ultrapure water to 320mL. Ultrasonicate until dissolved, then add 400μL of concentrated hydrochloric acid, and finally dilute to 400mL with ultrapure water.

B. Treatment of adherent cells (HT-29, LoVo, A549)

Cells in the logarithmic growth phase were inoculated in 96-well plates at 100 μL per well (the inoculation numbers of LoVo, HT-29 and A549 cells were 3500, 2000 and 3000 per well, respectively). A blank group and a cell control group were set aside, 200 μL of injection saline was added to the side wells, and the wells were cultured in a cell incubator overnight. 100 μL of culture medium containing different concentrations of compounds was added to the wells inoculated with cells, and 3 replicates were set for each group. Incubate in a cell incubator for 72 hours.

C. Treatment of suspension cells (NCI-H23 and HL60)

After adding 100 μL of culture medium containing different concentrations of compounds to the well plate seeded with cells, the cells in the logarithmic growth phase were seeded in a 96-well plate at 100 μL per well (the seeding numbers of NCI-H23 and HL60 cells were 8000 and 10000 per well, respectively). Three replicate wells were set up for each group, leaving out a blank group and a cell control group. 200 μL of injection saline was added to the side wells and incubated in a cell incubator for 72 h.

D. Color

Add 20 μL of 5 mg/mL MTT staining solution to each well and incubate in a cell incubator for 2-4 hours. Confirm that the cells are completely stained under a microscope, add 50 μL of 20% acidic SDS solution, and continue to culture in the cell incubator overnight. Finally, use a multifunctional microplate reader to detect the absorbance value at 570 nm.

E. Calculation of _IC50

The absorbance of each well was used to calculate the cell survival rate under different concentrations of the compound, with the compound concentration as the abscissa and the survival rate as the ordinate, and the _IC50 was calculated using GraphPad Prism 5.0 software. Survival rate = 100-100% × (control group - drug group) / (control group - blank group).

Lovo cell test results: The inhibition of ATR activity by compound 5-X was determined. The results are shown in Table 3.

Table 3 LoVo cell activities of representative tetrahydropyridopyrimidine derivatives

Cell experiments were performed on compounds with ATR kinase activity within 0.5 μM. MTT tests were performed using ATM-deficient LoVo cell lines. The cell data are shown in Table 3. Compound 5-30 has the strongest ability to inhibit the proliferation of human colorectal cancer LoVo cells, with an IC ₅₀ of 0.270 μM; compound 5-27 is second, with an IC ₅₀ of 0.330 μM; the other compounds have an inhibitory effect on the proliferation of LoVo cells at 1.0 μM.

In order to find tumor cell lines that are more sensitive to tetrahydropyrido-pyrimidine compounds. The present invention has carried out a large number of cell screenings and found that the series of compounds are relatively sensitive to the ATM mutated mantle cell lymphoma cell Granta519 cell line. Through the MTT experiment, we tested the ability of 8 representative compounds to inhibit the proliferation of mantle cell lymphoma cells Granta519. The test results are shown in Table 4. Compounds 5-30 showed excellent anti-proliferative activity in mantle cell lymphoma cells, with an IC ₅₀ of 0.089 μM.

Table 4 Activity of representative tetrahydropyridopyrimidine derivatives in 519 cells

Compound 5-26 5-22 5-19 5-28 5-30 5-31 5-33 5-36 Cell IC ₅₀ (μM) 0.308 0.377 0.600 0.300 0.089 0.263 0.708 0.768

In order to evaluate the in vivo anti-tumor activity of compound 5-30, the present invention uses female NOD/SCID mice to establish a mantle cell lymphoma cell Granta-519 subcutaneous tumor xenograft model. Compound 5-30 was orally administered at 37.5 or 75 mg/kg twice a day (BID), and the control group was orally administered with the same volume of solvent. After 13 consecutive days of administration, it was found that the mantle cell lymphoma volume of the control group exceeded 2000 mm ³ , and the experiment was stopped. The animal results are shown in Figure 1. In all treatment groups, compound 5-30 slowed down tumor growth in a dose-dependent manner. Compared with the blank group, oral administration of 75 mg/kg compound 5-30 twice a day (BID) can significantly control the growth of mantle cell lymphoma; at the same time, the body weight of mice in the administration group did not change significantly during the administration period, which preliminarily showed that compound 5-30 was non-toxic in mice and had a certain safety.

Claims (11)

1. The compound represented by formula I or a pharmaceutically acceptable salt thereof, characterized in that: the structure is as follows: R ₁ is selected from R ₄ and R ₅ are independently selected from substituted or unsubstituted C1-C8 alkyl, substituted or unsubstituted 3-8 membered cycloalkyl, substituted or unsubstituted C1-C8 alkenyl, substituted or unsubstituted 6-10 membered aryl; in R ₁ , the substituents of the substituted C1-C8 alkyl and substituted C1-C8 alkenyl are selected from halogen, C1-C8 acyl, C1-C8 alkoxy, and cyano, and the substituents of the substituted 3-10 membered cycloalkyl and substituted 6-10 membered aryl are selected from C1-C8 alkyl, halogen, C1-C8 acyl, C1-C8 alkoxy, and cyano; R ₂ and R ₃ are independently selected from H, substituted or unsubstituted C1-C8 alkyl, substituted or unsubstituted 3-10-membered azacycloalkyl, and R ₂ and R ₃ are not H at the same time, or R ₂ and R ₃ are connected with N to form a substituted or unsubstituted 4-10-membered heterocycloalkyl, the 3-10-membered azacycloalkyl contains 1-2 N, and the 4-10-membered heterocycloalkyl contains 0-2 heteroatoms in addition to N shown in Formula I, and the heteroatoms are N, O or S; in R ₂ and R ₃ , the substituent of the substituted C1-C8 alkyl is selected from C1-C8 alkoxycarbonyl, amino, and sulfo; the substituent of the substituted 4-10-membered azacycloalkyl and substituted 4-10-membered heterocycloalkyl is selected from C1-C8 alkyl, C1-C8 alkoxycarbonyl, amino, and sulfo; _R4 is indolyl. 2. The compound according to claim 1, characterized in that: R ₄ is 3. The compound according to claim 1 or 2, characterized in that: R ₄ and R ₅ are independently selected from substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted 6-10 membered aryl, n＝0～3； _R4 、 _R5 , the substituent of the substituted C1～C6 alkyl is selected from halogen, C1～C6 acyl, C1～C6 alkoxy, cyano, the substituent of the substituted 6～10 membered aryl is selected from C1～C6 alkyl, halogen, C1～C6 acyl, C1～C6 alkoxy, cyano; _R6 is selected from H, halogen, cyano, halogen substituted or unsubstituted C1～C6 alkyl; _R7 is selected from H, halogen, C1～C6 alkyl; _R8 is selected from H, C1～C6 alkyl; _R9 is selected from H, C1～C6 alkyl, C1～C6 alkoxy; Preferably, R ₄ and R ₅ are independently selected from substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted 6-membered aryl, n＝0～3； _R4 、 _R5 , the substituent of the substituted C1～C6 alkyl is selected from halogen, C1～C4 acyl, C1～C4 alkoxy, cyano, the substituent of the substituted 6～10 membered aryl is selected from C1～C4 alkyl, halogen, C1～C4 acyl, C1～C4 alkoxy, cyano; _R6 is selected from H, halogen, cyano, halogen substituted or unsubstituted C1～C4 alkyl; _R7 is selected from H, halogen, C1～C4 alkyl; _R8 is selected from H, C1～C4 alkyl; _R9 is selected from H, C1～C4 alkyl, C1～C4 alkoxy; More preferably, R ₄ and R ₅ are independently selected from substituted or unsubstituted C1-C6 alkyl, phenyl, n＝0～3; in R ₄ and R ₅ , the substituent of the substituted C1～C6 alkyl is selected from F, Cl, C1～C4 acyl; R ₆ is selected from H, F, cyano, unsubstituted C1～C4 alkyl, trifluoromethyl; R ₇ is selected from H, F, C1～C4 alkyl; R ₈ is selected from H, C1～C4 alkyl; R ₉ is selected from H, C1～C4 alkyl, C1～C4 alkoxy; More preferably, _R4 is selected from substituted or unsubstituted C1-C6 alkyl, phenyl, n＝0～3；In _R4 , the substituent of the substituted C1～C6 alkyl is selected from F, Cl, C1～C4 acyl; _R6 is selected from H, F, cyano, unsubstituted C1～C4 alkyl, trifluoromethyl; _R7 is selected from H, F, C1～C4 alkyl; _R8 is selected from H, C1～C4 alkyl; _R9 is selected from H, C1～C4 alkyl, C1～C4 alkoxy; _R5 is fluorine-substituted or unsubstituted C1～C4 alkyl; Most preferably, _R4 is selected from methyl, difluoromethyl, trifluoromethyl, acetylmethyl, isopropyl, tert-butyl, isobutyl, Vinyl, Phenyl, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, R ₅ is selected from trifluoromethyl and methyl. 4. The compound according to any one of claims 1 to 3, characterized in that: R ₂ and R ₃ are independently selected from H, substituted or unsubstituted C1-C6 alkyl, substituted or unsubstituted 3-8-membered azacycloalkyl, and R ₂ and R ₃ are not H at the same time, and the 3-8-membered azacycloalkyl contains 1-2 N; in R ₂ and R ₃ , the substituent of the substituted C1-C6 alkyl is selected from C1-C6 alkoxycarbonyl, -NHR ₁₀ , and -SO ₂ R ₁₁ ; the substituent of the substituted 3-8-membered azacycloalkyl is selected from C1-C6 alkyl, C1-C6 alkoxycarbonyl, -NHR ₁₂ , and -SO ₂ R ₁₃ ; R ₁₀ and R ₁₂ are independently selected from H and C1-C6 alkyl; R ₁₁ and R ₁₃ are independently selected from C1-C6 alkyl; Preferably, R ₂ and R ₃ are independently selected from H, substituted or unsubstituted C1-C4 alkyl, substituted or unsubstituted 5-6-membered azacycloalkyl, and R ₂ and R ₃ are not H at the same time, and the 5-6-membered azacycloalkyl contains 1 N; in R ₂ and R ₃ , the substituent of the substituted C1-C4 alkyl is selected from C1-C4 alkoxycarbonyl, -NHR ₁₀ , and -SO ₂ R ₁₁ ; the substituent of the substituted 5-6-membered azacycloalkyl is selected from C1-C4 alkyl, C1-C4 alkoxycarbonyl, -NHR ₁₂ , and -SO ₂ R ₁₃ ; R ₁₀ and R ₁₂ are independently selected from H and C1-C4 alkyl; R ₁₁ and R ₁₃ are independently selected from C1-C4 alkyl; More preferably, R ₂ and R ₃ are independently selected from H, C1-C4 alkoxycarbonyl substituted or unsubstituted C1-C4 alkyl, unsubstituted 5-6-membered azacycloalkyl, and R ₂ and R ₃ are not H at the same time, and the 5-6-membered azacycloalkyl contains 1 N; Most preferably, R ₂ and R ₃ are independently selected from H, methyl, 5. The compound according to any one of claims 1 to 4, characterized in that: the structure is as shown in Formula II: The nitrogen heterocycle A is selected from substituted or unsubstituted 5- to 8-membered heterocycloalkyl groups, wherein the 5- to 8-membered heterocycloalkyl groups contain 0 to 1 heteroatoms in addition to N as shown in formula II, and the heteroatoms are N, O or S; the substituents of the substituted 5- to 8-membered heterocycloalkyl groups are selected from C1-C6 alkyl groups, C1-C6 alkoxycarbonyl groups, -NHR ₁₄ , and -SO ₂ R ₁₅ ; R ₁₄ is selected from H and C1-C6 alkyl groups; and R ₁₅ is selected from C1-C6 alkyl groups. 6. The compound according to claim 5, characterized in that: The nitrogen heterocycle A is selected from X is selected from O, NR ₂₀ , CHR ₂₁ , SO ₂ ; R ₁₆ , R ₁₇ , R ₁₉ are independently selected from H, C1-C4 alkyl, C1-C4 alkoxycarbonyl, -NHR ₁₄ , -SO ₂ R ₁₅ ; R ₁₄ is selected from H, C1-C4 alkyl; R ₁₅ is selected from C1-C4 alkyl; R ₁₈ , R ₂₀ , R ₂₁ are independently selected from H, C1-C4 alkoxycarbonyl, -SO ₂ R ₂₂ ; R ₂₂ is selected from C1-C4 alkyl; Preferably, the nitrogen heterocycle A is selected from R ₁₇ is -NHR ₁₄ ; R ₂₃ , R ₂₄ , R ₂₅ , and R ₂₆ are independently selected from H, C1-C4 alkyl, C1-C4 alkoxycarbonyl, and -NHR ₁₄ ; R ₁₄ is selected from H, C1-C4 alkyl; R ₂₇ is -SO ₂ R ₁₅ ; R ₁₅ is selected from C1-C4 alkyl; Most preferably, the nitrogen heterocycle A is selected from 7. The compound according to claim 6, characterized in that: for for for for for for 8. The compound according to claims 1 to 7, characterized in that the structural formula is as follows: 9. A pharmaceutical composition, comprising the compound according to any one of claims 1 to 8 or a pharmaceutically acceptable salt thereof as an active ingredient, and pharmaceutically acceptable auxiliary ingredients. 10. Use of the compound according to any one of claims 1 to 8 or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition according to claim 9 in the preparation of an ATR kinase inhibitor. 11. Use of the compound according to any one of claims 1 to 8 or a pharmaceutically acceptable salt thereof, or the pharmaceutical composition according to claim 9 in the preparation of a drug for treating cancer; preferably, the cancer is colorectal cancer or mantle cell lymphoma.