SPIRO RING-CONTAINING QUINAZOLINE COMPOUNDS

Abstract

The present invention relates to a spiro ring-containing quinazoline compound, a preparation method therefor, and use of the compound as a K-Ras G12C inhibitor in preparing antitumor medicaments.

##STR00001##

Claims

1. A compound with a structure as shown in general formula (1), isomers thereof, crystalline forms thereof, pharmaceutically acceptable salts thereof, hydrates thereof or solvates thereof: ##STR00559## wherein in formula (1): R.sup.1 is H, halogen, C1-C3 alkyl, C2-C4 alkenyl, C2-C4 alkynyl or C3-C6 cycloalkyl; R.sup.2 is C1-C3 alkoxy, C1-C3 haloalkoxy or —NR.sup.aR.sup.b, wherein R.sup.a and R.sup.b are independently H, C1-C3 alkyl or C1-C3 haloalkyl, or R.sup.a and R.sup.b, together with a N atom, form a 4-7 membered heterocycloalkyl group, wherein the heterocycloalkyl group may be substituted with 1-3 halogen atoms; R.sup.3 is ##STR00560## wherein R.sup.c is H or F; R.sup.d is H, F, Cl or Me; R.sup.e is H, F, Cl or Me; R.sup.f is F, NH.sub.2, Me or cyclopropyl; R.sup.x1, R.sup.x2, R.sup.x3, R.sup.x4, R.sup.x5, R.sup.x6 and R.sup.x7 are independently H, F, Cl, OH, OMe, NH.sub.2, CF.sub.3, C1-C3 alkyl or C3-C6 cycloalkyl; R.sup.4 is H, halogen, CN, C1-C3 alkyl, C1-C3 haloalkyl or heteroaryl; and when R.sup.3 is ##STR00561## and R.sup.4 is H, R.sup.5 is: ##STR00562## wherein n.sub.1, n.sub.2, n.sub.3, m.sub.1, m.sub.2 and m.sub.3 are independently integers of 1 or 2; R.sup.g is C1-C3 alkyl, C3-C6 cycloalkyl, (C1-C3)alkoxy-(C2-C3)alkyl-, (halogenated C1-C3)alkoxy-(C2-C3)alkyl-, (C3-C6)cycloalkyl-(C1-C3)alkyl-, heterocycloalkyl, heterocycloalkyl-(C1-C3)alkyl-, C1-C3 haloalkyl or cyano-substituted C1-C3 alkyl; R.sup.h is ##STR00563## when R.sup.3 is ##STR00564## and R.sup.4 is halogen, CN, C1-C3 alkyl, C1-C3 haloalkyl or heteroaryl; or, when R.sup.3 is ##STR00565## ##STR00566## wherein n.sub.1, n.sub.2, n.sub.3, m.sub.1, m.sub.2 and m.sub.3 are independently integers of 1 or 2; R.sup.g is C1-C3 alkyl, C3-C6 cycloalkyl, (C1-C3)alkoxy-(C2-C3)alkyl-, (halogenated C1-C3)alkoxy-(C2-C3)alkyl-, (C3-C6)cycloalkyl-(C1-C3)alkyl-, heterocycloalkyl, heterocycloalkyl-(C1-C3)alkyl-, C1-C3 haloalkyl or cyano-substituted C1-C3 alkyl; R.sup.h is ##STR00567## R.sup.i is H, halogen, methyl or cyano.

2. The compound according to claim 1, wherein in the general formula (1), R.sup.1 is H, F, Cl, Me, Et, isopropyl, vinyl, ethynyl or cyclopropyl.

3. The compound according to claim 1, wherein in the general formula (1), R.sup.2 is CH.sub.3O—, CH.sub.3CH.sub.2O—, CF.sub.3CH.sub.2O—, CHF.sub.2CH.sub.2O—, ##STR00568##

4. The compound according to claim 1, wherein in the general formula (1), R.sup.3 is ##STR00569## ##STR00570##

5. The compound according to claim 1, wherein in the general formula (1), R.sup.4 is H, F, CN, Me, CF.sub.3, ##STR00571##

6. The compound according to claim 1, wherein in the general formula (1), when R.sup.3 is ##STR00572## and R.sup.4 is H, R.sup.5 is: ##STR00573## ##STR00574## ##STR00575## ##STR00576## ##STR00577## ##STR00578## ##STR00579##

7. The compound according to claim 1, wherein in the general formula (1), when R.sup.3 is ##STR00580## and R.sup.4 is F, CN, Me, CF.sub.3, ##STR00581## or, when R.sup.3 is ##STR00582## ##STR00583## R.sup.5 is: ##STR00584## ##STR00585## ##STR00586## ##STR00587## ##STR00588## ##STR00589## ##STR00590## ##STR00591## ##STR00592##

8. The compound according to claim 1, wherein in the general formula (1), R.sup.5 is: ##STR00593## ##STR00594## ##STR00595## ##STR00596## ##STR00597##

9. The compound, the isomers thereof, the crystalline forms thereof, the pharmaceutically acceptable salts thereof, the hydrates thereof or the solvates thereof according to claim 1, wherein the compound has one of the following structures: ##STR00598## ##STR00599## ##STR00600## ##STR00601## ##STR00602## ##STR00603## ##STR00604## ##STR00605## ##STR00606## ##STR00607## ##STR00608## ##STR00609## ##STR00610## ##STR00611## ##STR00612## ##STR00613## ##STR00614## ##STR00615## ##STR00616## ##STR00617## ##STR00618## ##STR00619## ##STR00620## ##STR00621## ##STR00622## ##STR00623## ##STR00624## ##STR00625## ##STR00626## ##STR00627## ##STR00628## ##STR00629## ##STR00630## ##STR00631## ##STR00632## ##STR00633## ##STR00634## ##STR00635## ##STR00636## ##STR00637## ##STR00638## ##STR00639## ##STR00640## ##STR00641## ##STR00642## ##STR00643## ##STR00644## ##STR00645## ##STR00646## ##STR00647## ##STR00648## ##STR00649## ##STR00650## ##STR00651## ##STR00652## ##STR00653## ##STR00654## ##STR00655## ##STR00656## ##STR00657## ##STR00658## ##STR00659## ##STR00660## ##STR00661## ##STR00662## ##STR00663## ##STR00664## ##STR00665## ##STR00666## ##STR00667## ##STR00668## ##STR00669## ##STR00670## ##STR00671## ##STR00672##

10. A compound with a structure as shown in general formula (2), isomers thereof, crystalline forms thereof, pharmaceutically acceptable salts thereof, hydrates thereof or solvates thereof: ##STR00673## wherein in general formula (2): R.sup.1a is ##STR00674## R.sup.2a is CH.sub.3O—, CH.sub.3CH.sub.2O—, CF.sub.3CH.sub.2O— or CHF.sub.2CH.sub.2O—; R.sup.3a is ##STR00675## wherein R.sup.c is H or F, R.sup.d is H, F, Cl or Me, R.sup.e is H, F, Cl or Me, and R.sup.f is F, NH.sub.2, Me or cyclopropyl; R.sup.4a is H or F; and R.sup.5a is: H, ##STR00676## wherein n.sub.1, n.sub.2, n.sub.3, m.sub.1, m.sub.2 and m.sub.3 are independently integers of 1 or 2; v is an integer of 1, 2 or 3; R.sup.g is C1-C3 alkyl, C3-C6 cycloalkyl, (C1-C3)alkoxy-(C2-C3)alkyl-, (halogenated C1-C3)alkoxy-(C2-C3)alkyl-, (C3-C6)cycloalkyl-(C1-C3)alkyl-, heterocycloalkyl, heterocycloalkyl-(C1-C3)alkyl-, C1-C3 haloalkyl or cyano-substituted C1-C3 alkyl; is independently halogen, CN, SO.sub.2Me, C1-C3 alkyl, C1-C3 haloalkyl, C1-C3 alkoxy, C1-C3 haloalkoxy, hydroxy-substituted C1-C3 alkyl, cyano-substituted C1-C3 alkyl, C3-C6 cycloalkyl or ##STR00677## R.sup.k is independently halogen, CN, OH, C1-C3 alkyl, C1-C3 alkoxy, C3-C6 cycloalkyl or ##STR00678## R.sup.n is independently halogen, CN, OH, C1-C3 alkyl, C1-C3 alkoxy or C3-C6 cycloalkyl, two R.sup.n groups, together with one carbon atom, form a spiro ring, or two R.sup.n groups, together with different carbon atoms, form a bridged ring; R.sup.l and R.sup.m are independently C1-C3 alkyl, C1-C3 haloalkyl, hydroxy-substituted C1-C3 alkyl, cyano-substituted C1-C3 alkyl, C3-C6 cycloalkyl, (C1-C3)alkoxy-(C2-C3)alkyl-, (halogenated C1-C3)alkoxy-(C2-C3)alkyl-, (C3-C6)cycloalkyl-(C1-C3)alkyl-, or R.sup.l and R.sup.m, together with a N atom, form a 3-8 membered heterocycloalkyl group, wherein the 3-8 membered heterocycloalkyl group may be substituted with 1-3 groups selected from OH, halogen, cyano, C1-C3 alkyl, C3-C6 cycloalkyl, heterocycloalkyl, (C1-C3)alkoxy or (halogenated C1-C3)alkoxy.

11. The compound according to claim 10, wherein in the general formula (2), R.sup.3a is ##STR00679##

12. The compound according to claim 10, wherein in the general formula (2), R.sup.5a is: H, ##STR00680## ##STR00681## ##STR00682## ##STR00683## ##STR00684## ##STR00685## ##STR00686## ##STR00687## ##STR00688## ##STR00689## ##STR00690## ##STR00691## ##STR00692##

13. The compound, the isomers thereof, the crystalline forms thereof, the pharmaceutically acceptable salts thereof, the hydrates thereof or the solvates thereof according to claim 10, wherein the compound has one of the following structures: ##STR00693## ##STR00694## ##STR00695## ##STR00696## ##STR00697## ##STR00698## ##STR00699## ##STR00700## ##STR00701## ##STR00702## ##STR00703## ##STR00704## ##STR00705## ##STR00706## ##STR00707## ##STR00708## ##STR00709## ##STR00710## ##STR00711## ##STR00712## ##STR00713## ##STR00714## ##STR00715## ##STR00716## ##STR00717## ##STR00718## ##STR00719## ##STR00720## ##STR00721## ##STR00722## ##STR00723## ##STR00724## ##STR00725## ##STR00726## ##STR00727## ##STR00728## ##STR00729## ##STR00730## ##STR00731##

14. A compound with a structure as shown in general formula (3), isomers thereof, crystalline forms thereof, pharmaceutically acceptable salts thereof, hydrates thereof or solvates thereof: ##STR00732## wherein, R.sup.5b is: ##STR00733## wherein n.sub.1, n.sub.2, n.sub.3, m.sub.1, m.sub.2 and m.sub.3 are independently integers of 1 or 2; R.sup.g is C1-C3 alkyl, C3-C6 cycloalkyl, (C1-C3)alkoxy-(C2-C3)alkyl-, (halogenated C1-C3)alkoxy-(C2-C3)alkyl-, (C3-C6)cycloalkyl-(C1-C3)alkyl-, heterocycloalkyl, heterocycloalkyl-(C1-C3)alkyl-, C1-C3 haloalkyl or cyano-substituted C1-C3 alkyl; R.sup.h is ##STR00734## R.sup.i is H, halogen, methyl or cyano; or R.sup.5b is: H, ##STR00735## wherein n.sub.1, n.sub.2, n.sub.3, m.sub.1, m.sub.2 and m.sub.3 are independently integers of 1 or 2; v is an integer of 1, 2 or 3; R.sup.g is C1-C3 alkyl, C3-C6 cycloalkyl, (C1-C3)alkoxy-(C2-C3)alkyl-, (halogenated C1-C3)alkoxy-(C2-C3)alkyl-, (C3-C6)cycloalkyl-(C1-C3)alkyl-, heterocycloalkyl, heterocycloalkyl-(C1-C3)alkyl-, C1-C3 haloalkyl or cyano-substituted C1-C3 alkyl; R.sup.j is independently halogen, CN, SO.sub.2Me, C1-C3 alkyl, C1-C3 haloalkyl, C1-C3 alkoxy, C1-C3 haloalkoxy, hydroxy-substituted C1-C3 alkyl, cyano-substituted C1-C3 alkyl, C3-C6 cycloalkyl or ##STR00736## R.sup.k is independently halogen, CN, OH, C1-C3 alkyl, C1-C3 alkoxy, C3-C6 cycloalkyl or ##STR00737## R.sup.n is independently halogen, CN, OH, C1-C3 alkyl, C1-C3 alkoxy or C3-C6 cycloalkyl, two R.sup.n groups, together with one carbon atom, form a spiro ring, or two R.sup.n groups, together with different carbon atoms, form a bridged ring; R.sup.l and R.sup.m are independently C1-C3 alkyl, C1-C3 haloalkyl, hydroxy-substituted C1-C3 alkyl, cyano-substituted C1-C3 alkyl, C3-C6 cycloalkyl, (C1-C3)alkoxy-(C2-C3)alkyl-, (halogenated C1-C3)alkoxy-(C2-C3)alkyl-, (C3-C6)cycloalkyl-(C1-C3)alkyl-, or R.sup.l and R.sup.m, together with a N atom, form a 3-8 membered heterocycloalkyl group, wherein the 3-8 membered heterocycloalkyl group may be substituted with 1-3 groups selected from OH, halogen, cyano, C1-C3 alkyl, C3-C6 cycloalkyl, heterocycloalkyl, (C1-C3)alkoxy or (halogenated C1-C3)alkoxy.

15. The compound according to claim 14, wherein R.sup.5b is: H, ##STR00738## ##STR00739## ##STR00740## ##STR00741## ##STR00742## ##STR00743## ##STR00744## ##STR00745## ##STR00746## ##STR00747## ##STR00748## ##STR00749## ##STR00750## ##STR00751## ##STR00752## ##STR00753## ##STR00754## ##STR00755## ##STR00756## ##STR00757## ##STR00758## ##STR00759## ##STR00760## ##STR00761## ##STR00762## ##STR00763## ##STR00764## ##STR00765## ##STR00766##

16. A pharmaceutical composition comprising a pharmaceutically acceptable excipient or carrier, and the compounds of general formula (1), the isomers thereof, the crystalline forms thereof, the pharmaceutically acceptable salts thereof, the hydrates thereof or the solvates thereof according to claim 1 as active ingredients.

17. (canceled)

18. A pharmaceutical composition comprising a pharmaceutically acceptable excipient or carrier, and the compounds of general formula (2), the isomers thereof, the crystalline forms thereof, the pharmaceutically acceptable salts thereof, the hydrates thereof or the solvates thereof according to claim 10 as active ingredients.

19. A pharmaceutical composition comprising a pharmaceutically acceptable excipient or carrier, and the compounds of general formula (3), the isomers thereof, the crystalline forms thereof, the pharmaceutically acceptable salts thereof, the hydrates thereof or the solvates thereof according to claim 14 as active ingredients.

Description

BRIEF DESCRIPTION OF THE DRAWING

[0086] FIG. 1 shows the inhibition of the phosphorylated ERK (pERK) level in cells by compounds.

DETAILED DESCRIPTION

Example 1: Synthesis of 1-(7-(6-Cyclopropyl-8-ethoxy-2-ol-(2-methoxyethyl)piperidin-4-yl)oxy)-7-(5-methyl-1H-indazol-4-yl)quinazolin-4-yl)-2,7-diazaspiro[3.5]nonan-2-yl)-2-fluoroprop-2-en-1-one (Compound 1)

[0087] ##STR00217## ##STR00218##

Step 1: Synthesis of Compound 1-3

[0088] Compound 1-1 (5.5 g, 13.1 mmol) was suspended in dioxane (80 mL). DIPEA (10.1 g, 78.6 mmol) was added under ice bath, followed by 1-2 (3.0 g, 13.1 mmol). The mixture was stirred for 30 min, and stirred at room temperature for 1 h. The reaction was completed as detected by TLC. Water was added, followed by EA for extraction. The organic phase was dried and concentrated, and the residue was slurried with EA to obtain a yellow solid 1-3 (4.5 g, 56% yield).

[0089] .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ: 8.26 (d, J=1.5 Hz, 1H), 3.79 (s, 4H), 3.65 (s, 4H), 1.86 (t, J=5.3 Hz, 4H), 1.39 (s, 9H); MS(ESI): MS (ESI): 611.2 [M+1].sup.+.

Step 2: Synthesis of Compound 1-4

[0090] Compound 1-3 (4.5 g, 7.4 mmol) was dissolved in a mixed solution of DMF (40 mL) and THF (40 mL). 1-(2-Methoxyethyl)-4-hydroxypiperidine (2.4 g, 14.8 mmol) and DABCO (0.2 g, 1.5 mmol) were added. The mixture was stirred at room temperature overnight. After the reaction was completed, water was added, followed by EA for extraction. The organic phase was dried and concentrated, and the residue was subjected to column chromatography to obtain compound 1-4 (4.1 g, 76% yield).

[0091] .sup.1H NMR (400 MHz, DMSO-d.sub.6) δ: 8.14 (s, 1H), 4.99 (ddd, J=11.7, 8.5, 3.6 Hz, 1H), 3.66 (s, 9H), 3.44 (t, J=5.8 Hz, 2H), 3.24 (s, 3H), 3.17 (d, J=5.2 Hz, 1H), 2.77 (dt, J=9.5, 8.8 Hz, 2H), 2.26 (t, J=9.8 Hz, 2H), 2.00 (d, J=12.0 Hz, 2H), 1.84 (d, J=4.3 Hz, 4H), 1.74-1.61 (m, 2H), 1.39 (s, 9H); MS(ESI): 734.2 [M+1].sup.+.

Step 3: Synthesis of Compound 1-5

[0092] Trifluoroethanol (0.9 g, 8.4 mmol) was dissolved in anhydrous DMF (10 mL). NaH was added under ice bath. The mixture was stirred at room temperature for 5 min to obtain sodium trifluoroethoxide. Compound 1-4 (4.1 g, 5.6 mmol) was dissolved in anhydrous THF (40 mL). The solution of sodium trifluoroethoxide in DMF prepared above was added. The mixture was stirred at room temperature overnight. After the reaction was completed, water was added, followed by EA for extraction. The organic phase was dried and concentrated, and the residue was subjected to column chromatography to obtain compound 1-5 (4.5 g, 99% yield). MS (ESI): 814.2 [M+1].sup.+.

Step 4: Synthesis of Compound 1-6

[0093] To a single-necked flask were added compound 1-5 (4.1 g, 5.5 mmol), cyclopropylboronic acid (0.5 g, 6.1 mmol), Pd(dppf)Cl.sub.2 (0.9 g, 1.1 mmol) and K.sub.3PO.sub.4 (0.4 g, 1.7 mmol), followed sequentially by MeCN (40 mL), dioxane (40 mL) and H.sub.2O (16.5 mL). The mixture was stirred under nitrogen at 100° C. for 5 h. After the reaction was completed, the mixture was subjected to column chromatography to obtain compound 1-6 (2.5 g, 62% yield). MS (ESI): 728.3 [M+1].sup.+.

Step 5: Synthesis of Compound 1-7

[0094] To a single-necked flask were added compound 1-6 (2.5 g, 3.4 mmol), 5-methyl-1H-indazole-4-boronic acid (0.9 g, 5.1 mmol), Pd.sub.2(dba).sub.3 (0.3 g, 0.4 mmol), Xatphos (0.3 g, 0.7 mmol) and K.sub.3PO.sub.4 (2.2 g, 10.2 mmol), followed by dioxane (40 mL) and H.sub.2O (4 mL). The mixture was stirred under nitrogen at 120° C. overnight. After the reaction was completed, the mixture was subjected to column chromatography to obtain compound 1-7 (1 g, 38% yield). MS (ESI): 780.4 [M+1].sup.+.

Step 6: Synthesis of Compound 1-8

[0095] Compound 1-7 (1 g, 1.3 mmol) was dissolved in DCM (15 mL). TFA (5 mL) was added. The mixture was stirred at room temperature for 2 h. After the reaction was completed, the mixture was concentrated, basified with saturated sodium carbonate, and extracted with EA. The organic phase was dried and concentrated to obtain compound 1-8 (0.9 g, 99% yield). MS(ESI): 680.4 [M+1].sup.+.

Step 7: Synthesis of Compound 1

[0096] Compound 1-8 (150 mg, 0.2 mmol) and 2-fluoroacrylic acid (20 mg, 0.22 mmol) were dissolved in DCM (15 mL). DIPEA (52 mg, 0.4 mmol) and HATU (114 mg, 0.3 mmol) were added under ice bath. The mixture was stirred overnight. After the reaction was completed, the reaction mixture was washed with saturated brine. The organic phase was dried and concentrated, and the residue was subjected to column chromatography to obtain compound 1 (30 mg, 20% yield).

[0097] .sup.1H NMR (400 MHz, CD.sub.3OD) δ: 7.47-7.37 (m, 2H), 7.30 (d, J=8.6 Hz, 1H), 7.24 (s, 1H), 5.52 (d, J=3.4 Hz, 0.5H), 5.40 (d, J=3.4 Hz, 0.5H), 5.15 (d, J=3.4 Hz, 0.5H), 5.11 (d, J=3.4 Hz, 0.5H), 4.54 (dq, J=17.7, 8.8 Hz, 1H), 4.23-4.12 (m, 3H), 3.83 (s, 2H), 3.73-3.59 (m, 4H), 3.54 (t, J=5.2 Hz, 2H), 3.28 (s, 3H), 3.10 (dd, J=10.1, 6.2 Hz, 2H), 2.88 (s, 2H), 2.85-2.72 (m, 2H), 2.13 (d, J=26.2 Hz, 5H), 1.97 (dd, J=11.9, 6.8 Hz, 6H), 1.36 (dt, J=14.0, 6.6 Hz, 1H), 1.27-1.17 (m, 5H); MS (ESI): 752.4 [M+1].sup.+.

Example 2-341: Synthesis of Compound 2-341

[0098] The target compound 2-341 was obtained using different starting materials according to a synthesis method similar to that in Example 1.

TABLE-US-00001 TABLE 1 Compound Compound structure [M + H].sup.+ 2 [00219] 698.4 3 [00220] 734.4 4 [00221] 684.4 5 [00222] 738.3 6 [00223] 792.4 7 [00224] 747.4 8 [00225] 691.4 9 [00226] 727.4 10 [00227] 694.3 11 [00228] 712.3 12 [00229] 728.3 13 [00230] 708.3 14 [00231] 722.4 15 [00232] 736.4 16 [00233] 718.3 17 [00234] 748.4 18 [00235] 759.3 19 [00236] 802.3 20 [00237] 812.4 21 [00238] 812.4 22 [00239] 800.4 23 [00240] 801.4 24 [00241] 799.4 25 [00242] 801.4 26 [00243] 706.3 27 [00244] 734.4 28 [00245] 767.4 29 [00246] 786.3 30 [00247] 770.4 31 [00248] 766.4 32 [00249] 752.3 33 [00250] 756.3 34 [00251] 756.3 35 [00252] 734.4 36 [00253] 756.3 37 [00254] 738.3 38 [00255] 756.3 39 [00256] 735.4 40 [00257] 753.3 41 [00258] 761.3 42 [00259] 779.3 43 [00260] 762.3 44 [00261] 780.3 45 [00262] 714.3 46 [00263] 732.3 47 [00264] 713.4 48 [00265] 731.3 49 [00266] 781.3 50 [00267] 799.2 51 [00268] 698.3 52 [00269] 715.3 53 [00270] 716.3 54 [00271] 734.3 55 [00272] 748.3 56 [00273] 715.3 57 [00274] 714.3 58 [00275] 730.3 59 [00276] 746.3 60 [00277] 746.3 61 [00278] 708.3 62 [00279] 750.3 63 [00280] 766.4 64 [00281] 694.3 65 [00282] 738.3 66 [00283] 736.3 67 [00284] 674.3 68 [00285] 692.3 69 [00286] 687.3 70 [00287] 705.3 71 [00288] 731.4 72 [00289] 749.3 73 [00290] 701.3 74 [00291] 719.3 75 [00292] 745.4 76 [00293] 763.4 77 [00294] 715.4 78 [00295] 733.4 79 [00296] 741.4 80 [00297] 759.4 81 [00298] 771.4 82 [00299] 789.4 83 [00300] 755.4 84 [00301] 773.4 85 [00302] 783.3 86 [00303] 801.3 87 [00304] 754.4 88 [00305] 772.4 89 [00306] 757.4 90 [00307] 775.4 91 [00308] 759.4 92 [00309] 777.4 93 [00310] 773.4 94 [00311] 791.4 95 [00312] 773.4 96 [00313] 791.4 97 [00314] 701.3 98 [00315] 719.3 99 [00316] 745.4 100 [00317] 763.4 101 [00318] 773.4 102 [00319] 791.4 103 [00320] 759.4 104 [00321] 777.4 105 [00322] 773.4 106 [00323] 791.4 107 [00324] 787.4 108 [00325] 805.4 109 [00326] 662.3 110 [00327] 680.3 111 [00328] 704.3 112 [00329] 722.3 113 [00330] 706.3 114 [00331] 724.3 115 [00332] 706.3 116 [00333] 723.3 117 [00334] 750.4 118 [00335] 768.3 119 [00336] 746.4 120 [00337] 764.4 121 [00338] 746.4 122 [00339] 764.4 123 [00340] 702.3 124 [00341] 720.3 125 [00342] 746.3 126 [00343] 764.3 127 [00344] 744.3 128 [00345] 762.3 129 [00346] 730.4 130 [00347] 748.4 131 [00348] 774.4 132 [00349] 792.4 133 [00350] 772.4 134 [00351] 790.4 135 [00352] 730.4 136 [00353] 748.4 137 [00354] 773.4 138 [00355] 792.4 139 [00356] 693.3 140 [00357] 719.4 141 [00358] 737.3 142 [00359] 687.3 143 [00360] 705.3 144 [00361] 717.3 145 [00362] 735.3 146 [00363] 717.3 147 [00364] 735.3 148 [00365] 745.4 149 [00366] 763.4 150 [00367] 702.3 151 [00368] 720.3 152 [00369] 717.3 153 [00370] 735.3 154 [00371] 745.4 155 [00372] 763.4 156 [00373] 702.3 157 [00374] 727.4 158 [00375] 729.3 159 [00376] 715.4 160 [00377] 759.4 161 [00378] 729.4 162 [00379] 715.4 163 [00380] 729.4 164 [00381] 743.4 165 [00382] 689.3 166 [00383] 703.4 167 [00384] 743.3 168 [00385] 705.3 169 [00386] 723.3 170 [00387] 731.4 171 [00388] 700.4 172 [00389] 719.3 173 [00390] 719.3 174 [00391] 731.4 175 [00392] 731.4 176 [00393] 737.3 177 [00394] 715.4 178 [00395] 733.3 179 [00396] 745.4 180 [00397] 751.3 181 [00398] 700.4 182 [00399] 700.4 183 [00400] 715.4 184 [00401] 715.4 185 [00402] 689.4 186 [00403] 689.4 187 [00404] 703.4 188 [00405] 703.4 189 [00406] 717.4 190 [00407] 717.4 191 [00408] 733.4 192 [00409] 733.4 193 [00410] 733.4 194 [00411] 733.4 195 [00412] 745.4 196 [00413] 745.4 197 [00414] 745.4 198 [00415] 745.4 199 [00416] 751.3 200 [00417] 751.3 201 [00418] 719.3 202 [00419] 719.3 203 [00420] 731.4 204 [00421] 731.3 205 [00422] 737.3 206 [00423] 737.3 207 [00424] 729.4 208 [00425] 729.4 209 [00426] 731.4 210 [00427] 731.4 211 [00428] 747.4 212 [00429] 747.4 213 [00430] 759.4 214 [00431] 759.4 215 [00432] 715.4 216 [00433] 729.4 217 [00434] 701.4 218 [00435] 715.4 219 [00436] 705.3 220 [00437] 719.3 221 [00438] 687.3 222 [00439] 701.4 223 [00440] 691.3 224 [00441] 705.3 225 [00442] 677.4 226 [00443] 688.4 227 [00444] 703.4 228 [00445] 707.3 229 [00446] 705.3 230 [00447] 719.4 231 [00448] 714.3 232 [00449] 703.4 233 [00450] 717.4 234 [00451] 719.4 235 [00452] 733.4 236 [00453] 721.4 237 [00454] 701.4 238 [00455] 712.3 239 [00456] 785.3 240 [00457] 717.3 241 [00458] 765.3 242 [00459] 726.3 243 [00460] 717.3 244 [00461] 730.4 245 [00462] 715.4 246 [00463] 715.4 247 [00464] 715.4 248 [00465] 715.4 249 [00466] 726.3 250 [00467] 726.3 251 [00468] 731.4 252 [00469] 731.4 253 [00470] 759.4 254 [00471] 759.4 255 [00472] 744.4 256 [00473] 744.4 257 [00474] 726.3 258 [00475] 726.3 259 [00476] 731.4 260- [00477] 731.4 261 [00478] 735.3 262 [00479] 735.3 263 [00480] 749.4 264 [00481] 749.4 265 [00482] 731.4 266 [00483] 745.4 267 [00484] 737.3 268 [00485] 729.4 269 [00486] 743.4 270 [00487] 727.4 271 [00488] 717.3 272 [00489] 717.3 273 [00490] 719.3 274 [00491] 719.3 275 [00492] 731.4 276 [00493] 731.4 277 [00494] 744.4 278 [00495] 744.4 279 [00496] 688.3 280 [00497] 731.4 281 [00498] 731.4 282 [00499] 688.3 283 [00500] 702.3 284 [00501] 690.3 285 [00502] 697.3 286 [00503] 715.4 287 [00504] 727.4 288 [00505] 741.4 289 [00506] 741.4 290 [00507] 755.4 291 [00508] 755.4 292 [00509] 755.4 293 [00510] 755.4 294 [00511] 729.4 295 [00512] 729.4 296 [00513] 715.4 297 [00514] 715.4 298 [00515] 729.4 299 [00516] 729.4 300 [00517] 729.4 301 [00518] 729.4 302 [00519] 741.4 303 [00520] 741.4 304 [00521] 690.3 305 [00522] 706.3 306 [00523] 721.3 307 [00524] 750.3 308 [00525] 702.3 309 [00526] 746.3 310 [00527] 751.3 311 [00528] 753.3 312 [00529] 751.3 313 [00530] 751.3 314 [00531] 737.3 315 [00532] 737.3 316 [00533] 739.3 317 [00534] 753.3 318 [00535] 768.3 319 [00536] 768.3 320 [00537] 687.3 321 [00538] 701.3 322 [00539] 701.3 323 [00540] 715.3 324 [00541] 729.3 325 [00542] 729.3 326 [00543] 741.4 327 [00544] 733.4 328 [00545] 733.4 329 [00546] 745.4 330 [00547] 661.4 331 [00548] 661.4 332 [00549] 673.4 333 [00550] 697.4 334 [00551] 697.4 335 [00552] 709.4 336 [00553] 673.3 337 [00554] 687.4 338 [00555] 745.4 339 [00556] 701.4 340 [00557] 701.4 341 [00558] 713.4

Example 342: Chiral Resolution of Compound 142

[0099] The compounds of the present application may have axial chirality. Compounds with axial chirality can be resolved to obtain two chiral isomers.

[0100] Compound 142 (50 mg) was dissolved in ethanol (2 mL) at a concentration of 25 mg/mL. The volume for each injection was 500 μL. Conditions for preparative chromatography: CHIRALPAK AD-H (20×250 mm, 5 μm) chromatography column; mobile phase: ethanol-n-hexane (40/60); flow rate: 12 mL/min; wavelength of detection: 254 nm. The stepwise eluate was concentrated by rotary evaporation and dried to obtain two chiral isomers 142-a and 142-b of compound 142:

[0101] a first chiral isomer: 142-a; retention time on the chromatography column: 6.662 min; and

[0102] a second chiral isomer: 142-b; retention time on the chromatography column: 10.831 min.

[0103] Compounds 171, 174 and 270 were chirally resolved using a similar resolution procedure to obtain their two chiral isomers 171-a/171-b, 174-a/174-b and 270-a/270-b, respectively. Their retention times on the chromatography column are as follows:

TABLE-US-00002 TABLE 2 Conditions for and results of chiral resolution of compounds 171, 174 and 270 Conditions Compound for resolution Results of resolution 171 Same as those for a first chiral isomer: 171-a; retention time on the compound 142 chromatography column: 7.481 min; and a second chiral isomer: 171-b; retention time on the chromatography column: 12.770 min. 174 Same as those for a first chiral isomer: 174-a; retention time on the compound 142 chromatography column: 8.994 min; and a second chiral isomer: 174-b; retention time on the chromatography column: 14.583 min. 270 Same as those for a first chiral isomer: 270-a; retention time on the compound 142 except that chromatography column: 7.280 min; and the mobile phase is ethanol- a second chiral isomer: 270-b; retention time on n-hexane (30/70) the chromatography column: 12.962 min.

[0104] Other compounds in the present application can also be chirally resolved using a similar method.

Example 343: pERK and ERK Protein Content Assay in 11358 Cells by Compounds

[0105] H358 cells were seeded in a 24-well plate. After one day of growth, a test compound (at a concentration of 1 μM) was added. After 24 h of action of the compound, the cells were lysed, and the cell lysate was transferred to a 96-well ELISA plate. The levels of pERK and ERK in the lysate were measured using an ELISA kit (abcam 176660). The ratio of pERK to ERK was calculated and compared with that of the DMSO group, and the percentage of inhibition of pERK activity by the compound was calculated. The results are shown in Table 3 below.

TABLE-US-00003 TABLE 3 Inhibitory activity of the compounds of the present invention against the pERK level in H358 cells Inhibition rate Inhibition rate Inhibition rate Compound (%) Compound (%) Compound (%) 1 +++ 2 +++ 3 +++ 4 +++ 5 +++ 6 +++ 7 ++ 8 ++ 9 +++ 10 ++ 11 +++ 12 +++ 13 +++ 14 +++ 15 +++ 16 ++ 17 +++ 18 ++ 19 ++ 20 +++ 21 +++ 22 ++ 23 +++ 24 ++ 25 +++ 26 ++ 27 ++ 28 ++ 29 +++ 30 +++ 31 +++ 32 +++ 33 +++ 34 +++ 35 +++ 36 +++ 37 ++ 38 +++ 39 +++ 40 +++ 41 +++ 42 +++ 43 +++ 44 +++ 45 +++ 46 +++ 47 +++ 48 +++ 49 +++ 50 +++ 51 +++ 52 +++ 53 +++ 54 +++ 55 +++ 56 +++ 57 +++ 58 +++ 59 +++ 60 +++ 61 +++ 62 +++ 63 +++ 64 ++ 65 +++ 66 ++ 67 ++ 68 ++ 69 +++ 70 +++ 71 +++ 72 +++ 73 +++ 74 +++ 75 +++ 76 +++ 77 ++ 78 ++ 79 ++ 80 ++ 81 +++ 82 +++ 83 +++ 84 +++ 85 +++ 86 ++ 87 +++ 88 ++ 89 +++ 90 +++ 91 +++ 92 +++ 93 +++ 94 +++ 95 +++ 96 +++ 97 ++ 98 ++ 99 +++ 100 +++ 101 +++ 102 +++ 103 +++ 104 ++ 105 +++ 106 ++ 107 ++ 108 ++ 109 +++ 110 +++ 111 +++ 112 ++ 113 +++ 114 +++ 115 ++ 116 ++ 117 +++ 118 +++ 119 ++ 120 ++ 121 ++ 122 ++ 123 +++ 124 +++ 125 +++ 126 +++ 127 +++ 128 +++ 129 +++ 130 +++ 131 +++ 132 +++ 133 +++ 134 +++ 135 ++ 136 ++ 137 +++ 138 +++ 139 +++ 140 +++ 141 +++ 142 +++ 143 +++ 144 +++ 145 +++ 146 +++ 147 +++ 148 ++ 149 ++ 150 +++ 151 +++ 152 +++ 153 +++ 154 +++ 155 ++ 156 +++ 157 +++ 158 +++ 159 +++ 160 +++ 161 +++ 162 +++ 163 +++ 164 +++ 165 +++ 166 +++ 167 +++ 168 +++ 169 ++ 170 ++ 171 +++ 172 +++ 173 +++ 174 +++ 175 +++ 176 ++ 177 +++ 178 +++ 179 +++ 180 +++ 181 +++ 182 +++ 183 +++ 184 +++ 185 +++ 186 +++ 187 +++ 188 +++ 189 +++ 190 +++ 191 +++ 192 +++ 193 +++ 194 +++ 195 +++ 196 +++ 197 +++ 198 +++ 199 +++ 200 +++ 201 +++ 202 +++ 203 +++ 204 +++ 205 ++ 206 ++ 207 +++ 208 +++ 209 +++ 210 +++ 211 +++ 212 +++ 213 ++ 214 ++ 215 +++ 216 +++ 217 +++ 218 +++ 219 +++ 220 +++ 221 +++ 222 +++ 223 +++ 224 +++ 225 +++ 226 +++ 227 +++ 228 +++ 229 +++ 230 ++ 231 ++ 232 +++ 233 +++ 234 +++ 235 +++ 236 +++ 237 +++ 238 +++ 239 ++ 240 +++ 241 ++ 242 +++ 243 +++ 244 +++ 245 +++ 246 +++ 247 +++ 248 +++ 249 ++ 250 ++ 251 +++ 252 +++ 253 +++ 254 +++ 255 +++ 256 +++ 257 +++ 258 +++ 259 +++ 260 +++ 261 +++ 262 +++ 263 +++ 264 +++ 265 +++ 266 +++ 267 +++ 268 +++ 269 +++ 270 +++ 271 +++ 272 +++ 273 +++ 274 +++ 275 +++ 276 +++ 277 +++ 278 +++ 279 +++ 280 +++ 281 +++ 282 +++ 283 ++ 284 +++ 285 ++ 286 +++ 287 +++ 288 +++ 289 +++ 290 +++ 291 +++ 292 +++ 293 +++ 294 +++ 295 +++ 296 +++ 297 +++ 298 +++ 299 +++ 300 +++ 301 +++ 302 +++ 303 +++ 304 ++ 305 ++ 306 +++ 307 +++ 308 +++ 309 +++ 310 ++ 311 +++ 312 +++ 313 +++ 314 +++ 315 +++ 316 +++ 317 +++ 318 +++ 319 +++ 320 +++ 321 +++ 322 +++ 323 +++ 324 +++ 325 +++ 326 +++ 327 +++ 328 +++ 329 +++ 330 +++ 331 +++ 332 +++ 333 +++ 334 +++ 335 +++ 336 +++ 337 +++ 338 +++ 339 +++ 340 +++ 341 +++ 142-a +++ 142-b ++ 171-a +++ 171-b +++ 174-a +++ 174-b ++ 270-a +++ 270-b +++ B +++ C +++ + indicates an inhibition rate less than or equal to 50% ++ indicates an inhibition rate from 50% to 90% +++ indicates an inhibition rate greater than 90%.

Example 344: Antiproliferative Activity of Compounds Against 11358 Cells

[0106] 2500 H358 cells were seeded in a 96-well ultra-low attachment plate (corning, 7007). After one day of growth, a serially diluted compound (a maximum concentration of 5 μM, 5-fold dilution, a total of five doses) was added. Three days after the addition of the compound, Cell Titer Glow (Promega, G9681) was added to evaluate pellet growth, and the IC.sub.50 value was calculated. The results are shown in Table 4 below.

TABLE-US-00004 TABLE 4 Antiproliferative activity of the compounds of the present invention against H358 cells Compound IC.sub.50 Compound IC.sub.50 Compound IC.sub.50 1 +++ 2 +++ 3 +++ 4 +++ 5 +++ 6 +++ 7 ++ 8 ++ 9 +++ 10 ++ 11 +++ 12 +++ 13 ++ 14 ++ 15 +++ 16 ++ 17 +++ 18 ++ 19 ++ 20 +++ 21 +++ 22 ++ 23 +++ 24 ++ 25 +++ 26 ++ 27 ++ 28 ++ 29 +++ 30 +++ 31 +++ 32 +++ 33 +++ 34 +++ 35 +++ 36 +++ 37 ++ 38 ++ 39 +++ 40 +++ 41 +++ 42 ++ 43 +++ 44 +++ 45 +++ 46 +++ 47 +++ 48 ++ 49 +++ 50 +++ 51 +++ 52 +++ 53 +++ 54 +++ 55 +++ 56 +++ 57 +++ 58 +++ 59 +++ 60 +++ 61 +++ 62 +++ 63 +++ 64 ++ 65 +++ 66 ++ 67 ++ 68 ++ 69 +++ 70 ++ 71 +++ 72 +++ 73 +++ 74 +++ 75 +++ 76 +++ 77 ++ 78 ++ 79 ++ 80 ++ 81 +++ 82 +++ 83 +++ 84 +++ 85 ++ 86 ++ 87 +++ 88 ++ 89 +++ 90 +++ 91 +++ 92 +++ 93 +++ 94 +++ 95 +++ 96 +++ 97 ++ 98 ++ 99 +++ 100 +++ 101 +++ 102 +++ 103 +++ 104 ++ 105 +++ 106 ++ 107 ++ 108 ++ 109 +++ 110 +++ 111 +++ 112 ++ 113 +++ 114 +++ 115 ++ 116 ++ 117 +++ 118 +++ 119 +++ 120 ++ 121 +++ 122 ++ 123 +++ 124 +++ 125 +++ 126 +++ 127 +++ 128 +++ 129 +++ 130 +++ 131 +++ 132 +++ 133 +++ 134 +++ 135 ++ 136 ++ 137 +++ 138 +++ 139 +++ 140 +++ 141 +++ 142 +++ 143 +++ 144 +++ 145 +++ 146 +++ 147 +++ 148 ++ 149 ++ 150 +++ 151 ++ 152 +++ 153 +++ 154 +++ 155 ++ 156 +++ 157 +++ 158 +++ 159 +++ 160 +++ 161 +++ 162 +++ 163 +++ 164 +++ 165 +++ 166 +++ 167 ++ 168 +++ 169 ++ 170 ++ 171 +++ 172 +++ 173 +++ 174 +++ 175 +++ 176 ++ 177 +++ 178 +++ 179 +++ 180 ++ 181 +++ 182 +++ 183 +++ 184 +++ 185 +++ 186 +++ 187 +++ 188 +++ 189 +++ 190 +++ 191 +++ 192 +++ 193 +++ 194 +++ 195 +++ 196 +++ 197 +++ 198 +++ 199 ++ 200 ++ 201 +++ 202 +++ 203 ++ 204 ++ 205 ++ 206 ++ 207 +++ 208 +++ 209 +++ 210 +++ 211 +++ 212 +++ 213 ++ 214 ++ 215 +++ 216 +++ 217 +++ 218 +++ 219 +++ 220 +++ 221 +++ 222 +++ 223 +++ 224 +++ 225 +++ 226 +++ 227 +++ 228 +++ 229 +++ 230 ++ 231 ++ 232 +++ 233 +++ 234 +++ 235 +++ 236 +++ 237 +++ 238 ++ 239 ++ 240 +++ 241 ++ 242 +++ 243 +++ 244 +++ 245 +++ 246 +++ 247 +++ 248 +++ 249 ++ 250 ++ 251 +++ 252 +++ 253 ++ 254 ++ 255 +++ 256 +++ 257 ++ 258 ++ 259 +++ 260 +++ 261 +++ 262 +++ 263 +++ 264 +++ 265 +++ 266 +++ 267 +++ 268 +++ 269 +++ 270 +++ 271 +++ 272 +++ 273 +++ 274 +++ 275 +++ 276 +++ 277 +++ 278 +++ 279 +++ 280 +++ 281 +++ 282 +++ 283 ++ 284 +++ 285 ++ 286 +++ 287 +++ 288 +++ 289 +++ 290 +++ 291 +++ 292 +++ 293 +++ 294 +++ 295 +++ 296 +++ 297 +++ 298 +++ 299 +++ 300 +++ 301 +++ 302 +++ 303 +++ 304 ++ 305 ++ 306 +++ 307 +++ 308 +++ 309 +++ 310 ++ 311 +++ 312 +++ 313 +++ 314 +++ 315 +++ 316 +++ 317 +++ 318 +++ 319 +++ 320 +++ 321 +++ 322 +++ 323 +++ 324 +++ 325 +++ 326 +++ 327 +++ 328 +++ 329 +++ 330 +++ 331 +++ 332 +++ 333 +++ 334 +++ 335 +++ 336 +++ 337 +++ 338 +++ 339 +++ 340 +++ 341 +++ 142-a +++ 142-b ++ 171-a +++ 171-b +++ 174-a +++ 174-b ++ 270-a +++ 270-b +++ B +++ C +++ + indicates the IC.sub.50 of the compound is greater than 1 μM ++ indicates the IC.sub.50 of the compound is from 0.3 to 1 μM +++ indicates the IC.sub.50 of the compound is less than 0.3 μM.

[0107] As can be seen from the data in Tables 3 and 4, the antiproliferative activity of most of the compounds of the present invention against H358 cells is less than 0.3 μM, and when R.sup.5 (or R.sup.5a or R.sup.5b) is a spiro ring or other substituted heterocyclic ring, the compounds have very high K-RAS G12C inhibitory activity. Compounds 131, 142 and 171 all have good antiproliferative activity against H358 cells, with their IC.sub.50 values being 1.5 nM, 2.5 nM and 1.4 nM, respectively, while the IC.sub.50 values of the reference compounds B and C were 4.6 nM and 5.1 nM, respectively, indicating that the cell activity of the compounds was greatly improved after cyclization of the amino groups on the side chains of the compounds. In addition, when position 2 (substituent R.sup.4) of acrylamide is substituted with a F atom that is small in size, the compounds also have very high K-RAS G12C inhibitory activity.

Example 345: Pharmacokinetic Evaluation in Mice

[0108] The compounds were administered by intravenous injection at a dose of 2 mg/kg and oral gavage at a dose of 10 mg/kg (0.5% CMC-Na suspension). 15 male ICR mice were selected for each group, and each mouse was subjected to blood collection at 3 discrete time points, with 3 mice per time point. The time points of sampling were as follows: before the administration, and at 5 min, 15 min, 30 min, 1 h, 3 h, 5 h, 8 h, 12 h and 24 h after the administration. 80 μL of blood was collected from the eye sockets or the hearts of the mice at each of the time points after the administration. All whole blood samples were collected in tubes containing EDTA K.sub.2 and centrifuged (1500-1600 rmp) at 4° C. for 10 min to isolate plasma, which was then stored in a refrigerator at −90 to −60° C. for sample analysis. The compound concentration in the plasma was determined by liquid chromatography-tandem mass spectrometry, and the corresponding pharmacokinetic parameters were obtained according to a plasma concentration-time curve.

TABLE-US-00005 TABLE 5 Pharmacokinetic parameters of compounds in mice Route of Dose t.sub.1/2 T.sub.max C.sub.max AUC.sub.0-t Vss Cl F Compound administration (mg/kg) (h) (h) (ng/mL) (ng .Math. h/L) (mL/kg) (mL/h/kg) (%)  1 iv 2 4.31 1 2557 13987 782 140 NA po 10 6.94 4 1543 20304 NA NA 29.0 131 iv 2 4.12 0.5 2346 11340 2104  112 NA po 10 5.86 2 1824 22315 NA NA 39.4 142 iv 2 3.31 0.083 13067 51995 187   38.5 NA po 10 3.89 2 6730 45952 NA NA .sup. 17.7% 171 iv 2 4.85 0.083 4910 26500 505   75.6 NA po 10 3.93 2 3320 35700 NA NA 26.9 B iv 2 4.47 0.083 2883 7822 1010  252 NA po 10 3.74 2 1300 10348 -NA  NA 26.5 C iv 2 4.02 0.083 3210 20200 481  99 NA po 10 3.38 0.5 3110 24800 NA NA 24.6 NA indicates data are not available;

[0109] As can be seen from the above table, compared to compound B, compound 131 has good oral absorption properties, and has improved metabolic parameters such as half-life (t.sub.1/2), maximum plasma concentration (C.sub.max), area under the drug-time curve (AUC.sub.0-t), and oral bioavailability. It should be particularly noted that, compared to the reference compound C in the patent (Example 65 of WO2018/143315), compound 171 has better metabolic parameters, and compound 142 also has significantly improved metabolic parameters such as C.sub.max and AUC.sub.0-t, indicating that the metabolic properties of the compound are well improved after the amino groups on the side chain are cyclized. The metabolic properties of the compounds similar to compounds 131 and 171 in the present application are also significantly improved. Good oral absorption properties are of great significance in improving the efficacy of drugs, reducing the dose of administration and reducing the costs.

Example 346: Evaluation of Antitumor Activity in Mice

[0110] Human pancreatic cancer Mia PaCa-2 cells were cultured conventionally in 1640 medium containing 10% fetal bovine serum in a 37° C./5% CO.sub.2 incubator. After passage, the cells were collected when they reached the desired amount. 1×10.sup.7 Mia PaCa-2 cells were injected into the left dorsal side of each nude mouse, and the animals were randomly grouped for administration after tumors grew to 150 mm3. The groups are as follows: 1) a solvent control group of 8 mice; and 2) compound 1 group, compound 2 group, compound 5 group, compound 31 group, compound 131 group, compound 142 group, compound 171 group, compound B group and compound C group, with 8 mice per group. Mice in the solvent control group were subjected to intragastric administration of 0.5% CMC-Na once daily; mice in compound 1 group, compound 2 group, compound 5 group, compound 31 group, compound 131 group, compound 142 group, compound 171 group, compound B group and compound C group were subjected to intragastric administration of a suspension of a compound in 0.5% CMC-Na once daily. On Tuesday and Thursday each week, tumor volumes and body weight of the mice were measured, and the nude mice were sacrificed on day 21 of administration. The test results are shown in Table 6 below.

TABLE-US-00006 TABLE 6 Experimental therapeutic effects of compounds on graft tumors of human pancreatic cancer Mia PaCa-2 in nude mice Dose Administration Anti-tumor Compound (mg/kg) regimen effect 1 10 qd*21 39% regression 2 10 qd*21 23% regression 5 10 qd*21 30% regression 31 10 qd*21 32% regression 131 10 qd*21 37% regression 142 10 qd*21 35% regression 171 10 qd*21 38% regression B 10 qd*21 25% regression C 10 qd*21  8% regression

[0111] As can be seen from the data in the table above, the compounds of the present invention have high in vivo antitumor activity; a tumor can regress after 21 consecutive days of administration at 10 mg/kg/day; compounds 1, 5, 31, 131, 142 and 171 have higher in vivo activity than reference compound B and compound C, and compounds 142 and 171 have significantly higher in vivo activity than compound C, indicating the in vivo activity of the compound is also greatly improved after the amino groups on the side chain of the compound are cyclized.

Example 346: pERK Level Assay by Western Blot

[0112] H358 cells were plated on to a 24-well plate at 2×10.sup.5 cells/well. Serially diluted compounds including AMG510, MRTX849, compound 142 and compound 171 were added. After overnight incubation, cells were lysed, and proteins were quantified and subjected to gel electrophoresis. The results of the phosphorylated ERK (pERK) level assay by western blot are shown in FIG. 1. As can be seen from the results in FIG. 1, the compounds 142 and 171 of the present invention shows stronger inhibition of the phosphorylated ERK (pERK) level in cells than the reference drugs AMG510 and MRTX849 when at the same concentration.