Pyrazoloquinazoline compounds, their preparation methods and uses

Abstract

This invention discloses a pyrazoloquinazoline compound, its preparation method, and its uses. The invention provides a pyrazoloquinazoline compound as shown in Formula I, its pharmaceutically acceptable salt, its solvate, or a solvate of a pharmaceutically acceptable salt thereof. This compound exhibits significant inhibitory effects on PLK1.

Description

[Technical Field]

[0001] The invention relates to a pyrazoloquinazoline compound, its preparation method and uses. [Previous Technology]

[0002] PLKs (Polo-like kinases) belong to the serine / threonine kinase family. To date, five subtypes have been identified, namely PLK1-5. PLKs are mainly expressed in dividing cells and play a crucial role in cell cycle control and mitosis. Among the PLK family, PLK1 is the most studied and detailed, and its biological activity is best described. The scientific community generally believes that PLK1 is a key kinase guiding mitotic entry, centrosome maturation and separation, bipolar spindle formation, the transition from metaphase to anaphase, and the initiation of cytokinesis. Studies have found that PLK1 is not only highly expressed in various tumor tissues but is also activated by DNA damage and replication stress, which is consistent with the abnormally active proliferation of tumor cells. Knocking out the PLK1 gene using chemical small molecule inhibitors or biological methods can result in a large number of tumor cells arresting in the G2 / M phase, further leading to tumor cell apoptosis. Therefore, PLK1 is considered a promising anti-tumor target, especially given its high expression in various tumor tissues, which suggests potential for broad-spectrum anti-tumor activity. Furthermore, PLK1 expression levels are low in normal cells, suggesting that developing PLK1 inhibitors as anti-tumor drugs may have a favorable safety profile.

[0003] Compared to the high expression of PLK1 in tumor cells, PLK2 and PLK3 are expressed more abundantly in post-mitotic cells such as nerve cells. Therefore, developing PLK1-specific inhibitors may avoid the toxic side effects caused by simultaneous inhibition of PLK2 / 3. The PLK1 inhibitor Volasetib (BI-6727) developed by BI (Boehringer Ingelheim) produced severe toxic side effects in clinical trials due to its lack of selectivity for PLK2 / 3, limiting its efficacy and leading to its failure in phase III clinical trials. As a PLK1-specific inhibitor, the small molecule inhibitor Onvansertib developed by Cardiff Oncology has shown good anti-tumor activity and manageable toxic side effects in several proof-of-concept phase II clinical trials. However, Onvansertib still has many areas for improvement, such as poor membrane permeability resulting in low bioavailability. Moreover, its long half-life (t1 / 2 greater than 24 hours) necessitates intermittent administration during treatment cycles, which, in addition to causing significant toxic side effects, further limits its therapeutic efficacy. [Summary of the Invention]

[0004] The technical problem to be solved by the present invention is the defects of the existing PLK1 inhibitors, such as simple structure and poor inhibitory effect. The present invention provides a pyrazoloquinazoline compound, its preparation method and use. The compound has a significant inhibitory effect on PLK1.

[0005] The present invention solves the above-mentioned technical problems through the following technical solutions.

[0006] This invention provides a pyrazoloquinazoline compound as shown in Formula I, a pharmaceutically acceptable salt thereof, a solvate thereof, or a solvate of a pharmaceutically acceptable salt thereof; wherein R1 is a C1-C4 alkoxy, a C1-C4 alkoxy substituted with one or more R1-1, or a 3-6 membered cycloalkyloxy; R1-1 is independently a halogen or a 3-6 membered cycloalkane; R2 is H, a halogen, or a C1-C4 alkyl; R3 is a 5-10 membered heterocyclic alkyl or a 5-10 membered heterocyclic alkyl substituted with one or more R3-1; wherein the heteroatom in the 5-10 membered heterocyclic alkyl is selected from one or more of N, O, and S, and the number of heteroatoms is 1-2; R3-1 is independently a C1-C4 alkyl; R4 is a cyano group, a 3- to 6-membered cycloalkane, a 3- to 6-membered cycloalkane substituted with one or more R4-1 groups, a 3- to 6-membered heterocycloalkane, or a 3- to 6-membered heterocycloalkane substituted with one or more R4-2 groups; a C1- to C4 alkyl group substituted with one or more R4-3 groups; wherein the heteroatom is selected from one or more of N, O, and S, and the number of heteroatoms is 1 to 2; R4-1 is independently a halogen, a hydroxyl group, or a C1- to C4 alkyl group; R4-2 is independently a halogen or a C1- to C4 alkyl group; R4-3 is independently a halogen, a cyano group, or a hydroxyl group; R5 is H, a hydroxyl group, a C1- to C4 alkyl group, or a C1- to C4 alkoxy group.

[0007] In one embodiment, certain groups in the pyrazoloquinazoline compound, its pharmaceutically acceptable salt, its solvate, or a solvate of a pharmaceutically acceptable salt thereof are defined as follows, and the remaining groups are defined as in any of the preceding embodiments (hereinafter referred to as "in one embodiment"):

[0008] In a certain scheme, R1-1 is independently F, Cl, Br or I, for example F.

[0009] In one embodiment, R1-1 is independently a 3- to 6-membered cycloalkane, such as cyclopropane, cyclobutane, or cyclopentane.

[0010] In one embodiment, when R1 is a C1-C4 alkoxy group, the C1-C4 alkoxy group is a methoxy group, an ethoxy group, an isopropoxy group, or a tert-butoxy group, such as a methoxy group.

[0011] In one embodiment, when R1 is a C1-C4 alkoxy group substituted by one or more R1-1, the C1-C4 alkoxy group may be methoxy, ethoxy, isopropoxy or tert-butoxy, for example methoxy.

[0012] In one embodiment, when R1 is a 3-6 membered cycloalkyloxy group, the 3-6 membered cycloalkoxy group is cyclopropyloxy, cyclobutyloxy or cyclopentyloxy, for example cyclopropyloxy or cyclobutyloxy.

[0013] In a certain scheme, R1 can be , , , , , or, for example, , , or.

[0014] In a certain scheme, when R2 is a halogen, the halogen can be F, Cl, Br or I.

[0015] In one embodiment, when R2 is a C1-C4 alkyl group, the C1-C4 alkyl group may be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl, for example methyl.

[0016] In one embodiment, when R3-1 is a C1-C4 alkyl group, the C1-C4 alkyl group may be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl, for example methyl.

[0017] In one embodiment, when R3 is a 5- to 10-membered heterocyclic alkyl group substituted with one or more R3-1, the 5- to 10-membered heterocyclic alkyl group may be a 6- to 9-membered heterocyclic alkyl group, such as a 6- to 9-membered heterocyclic alkyl group containing two N, or for example, pirazine, hexahydropyridazine, hexahydropyrimidinyl, 3,8-diazabicyclo[3.2.1]octyl, octahydropyrrole[1,2-a]pyrazine, 2,6-diazaspiro[3.4]octyl, 3,6-diazabicyclo[3.2.0]heptyl, 1,6-diazaspiro[3.4]octyl or octahydropyrrole[3,4-c]pyrrole.

[0018] In a certain scheme, R3 is preferably , , , , , , , or (b is the connection site of and ).

[0019] In a certain scheme, R3 can be , , , , , , , or.

[0020] In a certain scheme, R3 can be , , , , , , , or.

[0021] In one embodiment, R4 is a cyano group, a 3-6 membered cycloalkane substituted with one or more R4-1 groups, a 3-6 membered heterocycloalkane substituted with one or more R4-2 groups.

[0022] In one embodiment, R4-1 is a halogen or a hydroxyl group.

[0023] In one embodiment, R4-2 is a C1~C4 alkyl group.

[0024] In a certain scheme, R5 is H.

[0025] In one scheme, when R4-1 is a halogen, the halogen is F, Cl, Br or I, for example F.

[0026] In one embodiment, when R4-1 is a C1-C4 alkyl group, the C1-C4 alkyl group may be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl, for example methyl or ethyl.

[0027] In one embodiment, when R4-2 is a C1-C4 alkyl group, the C1-C4 alkyl group may be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl, for example methyl or ethyl.

[0028] In one scheme, when R4-2 is a halogen, the halogen is F, Cl, Br or I, for example F.

[0029] In one embodiment, when R4 is a 3- to 6-membered cycloalkane, the 3- to 6-membered cycloalkane may be cyclopropane, cyclobutane or cyclopentane, for example cyclopropane or cyclobutane.

[0030] In one embodiment, when R4 is a 3- to 6-membered cycloalkane substituted by one or more R4-1, the 3- to 6-membered cycloalkane may be cyclopropane, cyclobutane or cyclopentane, for example cyclopropane or cyclobutane.

[0031] In a certain scheme, when R4 is a 3-6 membered cycloalkane substituted by one or more R4-1, the 3-6 membered cycloalkane substituted by one R4-1 may be , or .

[0032] In a certain scheme, when R4 is a 3-6 member heterocyclic alkane, the 3-6 member heterocyclic alkane can be , or .

[0033] In one embodiment, when R4 is a 3- to 6-membered heterocyclic alkane substituted with an R4-2, the 3- to 6-membered heterocyclic alkane may be a 4-membered heterocyclic alkane containing an N or an O or a 5-membered heterocyclic alkane containing an N or an O.

[0034] In one embodiment, when R4 is a 3- to 6-membered heterocyclic alkane substituted with one or more R4-2, the 3- to 6-membered heterocyclic alkane substituted with one or more R4-2 is , or.

[0035] In one embodiment, when R4 is a 3- to 6-membered heterocyclic alkane substituted with one or more R4-2, the 3- to 6-membered heterocyclic alkane substituted with one or more R4-2 is or.

[0036] In one embodiment, when R4 is a C1-C4 alkyl group substituted with one or more R4-3, the C1-C4 alkyl group substituted with one or more R4-3 is...

[0037] In one embodiment, R4 is a cyano group, a 3-6 membered cycloalkane substituted with one or more R4-1 groups, a 3-6 membered heterocycloalkane substituted with one or more R4-2 groups, or a C1-C4 alkyl group substituted with one or more R4-3 groups.

[0038] In one embodiment, R4 is a 3- to 6-membered cycloalkane substituted by one or more R4-1, wherein R4-1 is a halogen or a hydroxyl group.

[0039] In one embodiment, R4 is a 3- to 6-membered heterocyclic alkane substituted by one or more R4-2, wherein R4-2 is a C1- to C4 alkyl group.

[0040] In one embodiment, R4 is a C1-C4 alkyl group substituted with one or more R4-3, wherein R4-3 is a hydroxyl group.

[0041] In a certain scheme, R4 is selected from the following group: cyano, , , , and.

[0042] In one embodiment, R5 is H or a hydroxyl group.

[0043] In one embodiment, R5 is a C1-C4 alkyl group, which may be methyl, ethyl, isopropyl or tert-butyl, for example methyl or ethyl.

[0044] In one embodiment, R5 is a C1-C4 alkoxy group, which is a methoxy, ethoxy, isopropoxy, or tert-butoxy group, for example, a methoxy group.

[0045] The present invention also provides a pyrazoloquinazoline compound as shown in Formula Ia, a pharmaceutically acceptable salt thereof, a solvate thereof, or a solvate of a pharmaceutically acceptable salt thereof:

[0046] In one embodiment, the present invention provides any of the following pyrazoloquinazoline compounds, pharmaceutically acceptable salts thereof, solvates thereof, or solvates of pharmaceutically acceptable salts thereof:

[0047] The present invention also provides a pyrazoloquinazoline compound as shown in Formula II, a pharmaceutically acceptable salt thereof, a solvate thereof, or a solvate of a pharmaceutically acceptable salt thereof: wherein R1 is a C1-C4 alkoxy group substituted with one or more R1-1; R1-1 is independently a halogen; R2 is H, a halogen, or a C1-C4 alkyl group; R3' is a 7-9 membered heterocyclic alkyl group or a 7-9 membered heterocyclic alkyl group substituted with one R3-1; the 7-9 membered heterocyclic alkyl group is a 7-9 membered heterospirocyclic alkyl group or a 7-9 membered heterobridged alkyl group; the heteroatom is selected from one or more of N, O, and S, and the number of heteroatoms is 1 to 2; R3-1 is a C1-C4 alkyl group; R4' is independently a C1-C4 alkyl group substituted with an R4-3; R4-3 is a hydroxyl group or a halogen; R5 is H, a hydroxyl C1-C4 alkyl group or a C1-C4 alkoxy group.

[0048] In a certain scheme, R1-1 is F, Cl, Br or I, for example F.

[0049] In one embodiment, when R1 is a C1-C4 alkoxy group substituted by one or more R1-1, the C1-C4 alkoxy group may be methoxy, ethoxy, isopropoxy or tert-butoxy, for example methoxy.

[0050] In a certain scheme, R1 is , , , for example.

[0051] In one embodiment, when R2 is a C1-C4 alkyl group, the C1-C4 alkyl group may be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl, for example methyl.

[0052] In one embodiment, R3-1 may be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl, for example methyl.

[0053] In one embodiment, when R3' is a 7-9 member heterospirocycloalkyl group, the 7-9 member heterospirocycloalkyl group may be a 7-9 member heterospirocycloalkyl group containing two nitrogen atoms, for example, or.

[0054] In one embodiment, when R3' is a 7-9 member heterospirocycloalkyl substituted with one R3-1, the 7-9 member heterospirocycloalkyl substituted with one R3-1 or unsubstituted may be a 7-9 member heterospirocycloalkyl substituted with one R3-1 containing two nitrogen atoms, for example or.

[0055] In one embodiment, when R3' is a 7-9 member heterobridged cycloalkyl group, the 7-9 member heterobridged cycloalkyl group may be a 7-9 member heterobridged cycloalkyl group containing two N atoms, for example.

[0056] In one embodiment, when R3' is a 7-9 member heterobridged cycloalkyl substituted with one R3-1, the 7-9 member heterobridged cycloalkyl substituted with one R3-1 may be a 7-9 member heterobridged cycloalkyl substituted with one R3-1 containing two nitrogen atoms, for example, or.

[0057] In a certain scheme, when R4-3 is a halogen, the halogen is F, Cl, Br or I.

[0058] In one embodiment, when R4' is independently a C1-C4 alkyl group substituted with an R4-3, the C1-C4 alkyl group may be methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl, for example methyl.

[0059] In one scheme, R4 can be -CH2OH.

[0060] In one embodiment, when R5 is a C1-C4 alkyl group, the C1-C4 alkyl group may be methyl, ethyl, isopropyl or tert-butyl, for example methyl or ethyl.

[0061] In one embodiment, when R5 is a C1-C4 alkoxy group, the C1-C4 alkoxy group is a methoxy group, an ethoxy group, an isopropoxy group, or a tert-butoxy group, such as a methoxy group.

[0062] In one embodiment, the pyrazoloquinazoline compound as shown in Formula II, its pharmaceutically acceptable salt, its solvate, or a solvate of its pharmaceutically acceptable salt may be any of the following:

[0063] The present invention also provides a pyrazoloquinazoline compound as shown in Formula III, a pharmaceutically acceptable salt thereof, a solvate thereof, or a solvate of a pharmaceutically acceptable salt thereof: wherein R1 is a C1-C4 alkoxy or a C1-C4 alkoxy substituted with one or more R1-1; R1-1 is independently a halogen or a 3-6 membered cycloalkane; R2 is H, a halogen, or a C1-C4 alkyl; R3 is a 5-10 membered heterocyclic alkyl or a 5-10 membered heterocyclic alkyl substituted with one or more R3-1; wherein the heteroatom in the 5-10 membered heterocyclic alkyl is selected from one or more of N, O, and S, and the number of heteroatoms is 1-2; R3-1 is independently a C1-C4 alkyl; R4 is a cyano group, a 3- to 6-membered cycloalkane, a 3- to 6-membered cycloalkane substituted with one or more R4-1 groups, a 3- to 6-membered heterocycloalkane, or a 3- to 6-membered heterocycloalkane substituted with one or more R4-2 groups; a C1- to C4 alkyl group substituted with one or more R4-3 groups; wherein the heteroatom is selected from one or more of N, O, and S, and the number of heteroatoms is 1 to 2; R4-1 is independently a halogen, a hydroxyl group, or a C1- to C4 alkyl group; R4-2 is independently a halogen or a C1- to C4 alkyl group; R4-3 is independently a halogen, a cyano group, or a hydroxyl group; R5 is H, a hydroxyl group, a C1- to C4 alkyl group, or a C1- to C4 alkoxy group.

[0064] In one embodiment, R1 is a C1-C4 alkoxy group, such as methoxy, ethoxy, isopropoxy or tert-butoxy.

[0065] In one embodiment, R1 is a C1-C4 alkoxy group substituted by one or more R1-1, wherein the C1-C4 alkoxy group may be methoxy, ethoxy, isopropoxy or tert-butoxy, for example methoxy.

[0066] In one embodiment, R1 is a C1-C4 alkoxy group substituted with one or more R1-1, wherein R1-1 is independently F, Cl, Br or I, for example F.

[0067] In a certain scheme, R1 can be selected from , , , , or.

[0068] In a certain scheme, R1 can be selected from , or .

[0069] In a certain scheme, R2 is H.

[0070] In one scheme, R2 is a halogen, which is F, Cl, Br or I.

[0071] In one embodiment, R2 is a C1-C4 alkyl group, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl.

[0072] In one embodiment, R3 is a 5- to 10-membered heterocyclic alkyl group, such as a 6- to 9-membered heterocyclic alkyl group, such as a 6- to 9-membered heterocyclic alkyl group containing two N atoms, or for example, pirazine, hexahydropyridazine, hexahydropyrimidinyl, 3,8-diazabicyclo[3.2.1]octyl, octahydropyrrole[1,2-a]pyrazine, 2,6-diazaspiro[3.4]octyl, 3,6-diazabicyclo[3.2.0]heptyl, 1,6-diazaspiro[3.4]octyl or octahydropyrrole[3,4-c]pyrrole.

[0073] In one embodiment, R3 is a 5- to 10-membered heterocyclic alkyl group substituted by one or more R3-1, wherein the 5- to 10-membered heterocyclic alkyl group is, for example, a 6- to 9-membered heterocyclic alkyl group, such as a 6- to 9-membered heterocyclic alkyl group containing two N atoms, or, for example, pirazine, hexahydropyridazine, hexahydropyrimidinyl, 3,8-diazabicyclo[3.2.1]octyl, octahydropyrrole[1,2-a]pyrazine, 2,6-diazaspiro[3.4]octyl, 3,6-diazabicyclo[3.2.0]heptyl, 1,6-diazaspiro[3.4]octyl or octahydropyrrole[3,4-c]pyrrole, wherein R3-1 is a C1- to C4 alkyl group, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl.

[0074] In a certain scheme, R3 is , , , , , , , or.

[0075] In a certain scheme, R3 is , , , , , , , or.

[0076] In a certain scheme, R3 is , , , , , , , or.

[0077] In one scheme, R4 is a cyano group.

[0078] In one scheme, R4 is a 3- to 6-membered cycloalkane, such as cyclopropane, cyclobutane or cyclopentane.

[0079] In one embodiment, R4 is a 3- to 6-membered cycloalkane substituted by one or more R4-1, wherein the 3- to 6-membered cycloalkane is, for example, cyclopropane, cyclobutane or cyclopentane, and R4-1 is a halogen, such as F, Cl, Br or I.

[0080] In one embodiment, R4 is a 3- to 6-membered cycloalkane substituted by one or more R4-1, wherein the 3- to 6-membered cycloalkane is, for example, cyclopropane, cyclobutane or cyclopentane, and R4-1 is a C1 to C4 alkyl group, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl.

[0081] In a certain scheme, R4 is , , or .

[0082] In one embodiment, R4 is a 3- to 6-membered heterocyclic alkane, such as a 4-membered heterocyclic alkane containing one N or O or a 5-membered heterocyclic alkane containing one N or O.

[0083] In one embodiment, R4 is a 3- to 6-membered heterocyclic alkane substituted with one or more R4-2, such as a 4-membered heterocyclic alkane containing one N or O or a 5-membered heterocyclic alkane containing one N or O, wherein R4-2 is a halogen, such as F, Cl, Br or I.

[0084] In a certain scheme, R4 is OR.

[0085] In one embodiment, R4 is a 3- to 6-membered heterocyclic alkane substituted with one or more R4-2, such as a 4-membered heterocyclic alkane containing one N or O or a 5-membered heterocyclic alkane containing one N or O, wherein R4-2 is a C1- to C4 alkyl group, such as methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl or tert-butyl.

[0086] In one embodiment, R4 is a C1-C4 alkyl group substituted with one or more R4-3, wherein the C1-C4 alkyl group is methyl, ethyl, isopropyl or tert-butyl, and R4-3 is a halogen, such as F, Cl, Br or I.

[0087] In one embodiment, R4 is a C1-C4 alkyl group substituted with one or more R4-3, wherein the C1-C4 alkyl group is methyl, ethyl, isopropyl or tert-butyl, and R4-3 is cyano.

[0088] In one embodiment, R4 is a C1-C4 alkyl group substituted with one or more R4-3, wherein the C1-C4 alkyl group is methyl, ethyl, isopropyl or tert-butyl, and R4-3 is hydroxyl.

[0089] In a certain scheme, R4 is.

[0090] In one embodiment, R5 is H or a hydroxyl group.

[0091] In one embodiment, R5 is a C1-C4 alkyl group, such as methyl, ethyl, isopropyl or tert-butyl.

[0092] In one embodiment, R5 is a C1-C4 alkoxy group, such as methoxy, ethoxy, isopropoxy or tert-butoxy.

[0093] In one embodiment, the pyrazoloquinazoline compound as shown in Formula III is:

[0094] The present invention also provides a method for preparing a pyrazoloquinazoline compound as shown in Formula I, a pharmaceutically acceptable salt thereof, a solvate thereof, or a solvate of a pharmaceutically acceptable salt thereof, comprising the following steps: in an organic solvent, in a base reagent, and under the action of a palladium catalyst, 1 g of compound I and a ligand reagent are coupled with R3-H to obtain compound I; wherein, R1, R2, R3, R4, or R5 are defined as described above; X is a halogen; L is a hydrochloride salt.

[0095] In this coupling reaction, the palladium catalyst can be tridibenzylacetone dipalladium.

[0096] In this coupling reaction, the organic solvent can be a conventional solvent for this type of reaction in the art, such as tetradioxane.

[0097] In this coupling reaction, the base reagent can be cesium carbonate.

[0098] In this coupling reaction, the ligand reagent may be 4,5-bis(diphenylphosphine)-9,9-dimethyloxanthracene.

[0099] The present invention also provides a pyrazoloquinazoline compound as shown in Formula II, a pharmaceutically acceptable salt thereof, a solvate thereof, or a solvate of a pharmaceutically acceptable salt thereof, comprising the following steps: in an organic solvent, under the action of a palladium catalyst, 1 g of the compound, a ligand reagent, and R3-L are coupled together to obtain compound II; wherein, R1, R2, R3', R4', or R5 are defined as described above; X is a halogen; L is a hydrochloride salt.

[0100] In this coupling reaction, the palladium catalyst can be tridibenzylacetone dipalladium.

[0101] In this coupling reaction, the organic solvent can be a conventional solvent for this type of reaction in the art, such as tetradioxane.

[0102] In this coupling reaction, the base can be cesium carbonate.

[0103] In this coupling reaction, the ligand reagent can be 4,5-bis(diphenylphosphine)-9,9-dimethyloxanthracene.

[0104] The present invention provides the use of a pyrazoloquinazoline compound as shown in Formula I, a pharmaceutically acceptable salt thereof, a solvate thereof, or a solvate of a pharmaceutically acceptable salt thereof, in the preparation of a PLK1 inhibitor; the inhibitor is preferably an inhibitor for in vitro use.

[0105] The present invention also provides a pharmaceutical composition comprising a pyrazoloquinazoline compound as shown in Formula I, a pharmaceutically acceptable salt thereof, a solvate thereof, or a solvate of a pharmaceutically acceptable salt thereof, and pharmaceutical excipients.

[0106] The present invention also provides the use of pyrazoloquinazoline compounds, pharmaceutically acceptable salts thereof, solvates thereof, or solvates of pharmaceutically acceptable salts thereof as shown in Formula I as described above in the preparation of a medicament;

[0107] This drug is for the treatment of at least one of the following diseases: breast cancer, prostate cancer, lung cancer, colorectal cancer, liver cancer, pancreatic cancer, stomach cancer, esophageal cancer, melanoma, multiple myeloma, leukemia and lymphoma.

[0108] The present invention provides the use of a pyrazoloquinazoline compound as shown in Formula II, a pharmaceutically acceptable salt thereof, a solvate thereof, or a solvate of a pharmaceutically acceptable salt thereof in the preparation of a PLK1 inhibitor; the inhibitor is preferably an inhibitor for in vitro use.

[0109] The present invention also provides a pharmaceutical composition comprising a pyrazoloquinazoline compound as shown in Formula I, a pharmaceutically acceptable salt thereof, a solvate thereof, or a solvate of a pharmaceutically acceptable salt thereof, and a pharmaceutical excipient.

[0110] The present invention also provides the use of the pyrazoloquinazoline compound as shown in Formula II as described above, its pharmaceutically acceptable salt, its solvate or a solvate of its pharmaceutically acceptable salt in the preparation of a medicament;

[0111] This drug is for the treatment of at least one of the following diseases: breast cancer, prostate cancer, lung cancer, colorectal cancer, liver cancer, pancreatic cancer, stomach cancer, esophageal cancer, ovarian cancer, melanoma, osteosarcoma, multiple myeloma, leukemia and lymphoma.

[0112] The present invention also provides a pharmaceutical composition comprising the compound of the present invention, a pharmaceutically acceptable salt thereof, a solvate thereof or a solvate of a pharmaceutically acceptable salt thereof, and one or more chemotherapeutic agents.

[0113] In one embodiment, the one or more chemotherapeutic agents are used simultaneously, separately or sequentially with the compounds of the present invention, pharmaceutically acceptable salts thereof, solvates thereof or solvates of pharmaceutically acceptable salts thereof.

Implementation Method

[0114] Terminology Explanation

[0115] In various parts of this specification, the substituents of the disclosed compounds are disclosed according to the type or range of groups. In particular, the invention includes each independent sub-combination of the members of these types and ranges of groups. For example, the term "C1-C4 alkyl" specifically refers to the independently disclosed methyl, ethyl, C3 alkyl (i.e., propyl, including n-propyl and isopropyl), and C4 alkyl (i.e., butyl, including n-butyl, isobutyl, sec-butyl, and tert-butyl).

[0116] When the listed substituents do not specify which atom they are attached to in connection with a compound included but not specifically mentioned in the general chemical formula, such substituents may be bonded through any of their atoms. Combinations of substituents and / or their variants are permitted only if such combinations produce stable compounds.

[0117] When the listed groups do not explicitly indicate that they have substituents, such groups refer only to those that are not substituted. For example, when there is no "substituted or unsubstituted" limitation before "C1~C4 alkyl", it refers only to "C1~C4 alkyl" itself or "unsubstituted C1~C4 alkyl".

[0118] The term "Ci-Cj" represents a range of carbon atoms, where i and j are integers and j is greater than i, and the range of carbon atoms includes the endpoints (i.e., i and j) and every integer point between the endpoints. For example, C1 to C6 represent a range of 1 to 6 carbon atoms, including 1 carbon atom, 2 carbon atoms, 3 carbon atoms, 4 carbon atoms, 5 carbon atoms, and 6 carbon atoms. In some embodiments, the term "C1-C12" represents 1 to 12, particularly 1 to 10, particularly 1 to 8, particularly 1 to 6, particularly 1 to 5, particularly 1 to 4, particularly 1 to 3, or particularly 1 to 2 carbon atoms.

[0119] The term "alkyl" refers to a saturated straight-chain or branched hydrocarbon group. The term "Ci-Cj alkyl" refers to an alkyl group having i to j carbon atoms. In some embodiments, the alkyl group comprises 1 to 12 carbon atoms. In some embodiments, the alkyl group comprises 1 to 11 carbon atoms, 1 to 10 carbon atoms, 1 to 9 carbon atoms, 1 to 8 carbon atoms, 1 to 7 carbon atoms, 1 to 6 carbon atoms, 1 to 5 carbon atoms, 1 to 4 carbon atoms, 1 to 3 carbon atoms, or 1 to 2 carbon atoms. Examples of alkyl groups include, but are not limited to, methyl, ethyl, 1-propyl (n-propyl), 2-propyl (isopropyl), 1-butyl (n-butyl), 2-methyl-1-propyl (isobutyl), 2-butyl (neobutyl), 2-methyl-2-propyl (tert-butyl), 1-pentyl (n-pentyl), 2-pentyl, 3-pentyl, 2-methyl-2-butyl, 3-methyl-2-butyl, 3-methyl-1-butyl, 2-methyl-1-butyl, 1-hexyl, 2-hexyl, 3-hexyl, 2-methyl-2-pentyl, 3-methyl-2-pentyl, 4-methyl-2-pentyl, 3-methyl-3-pentyl, 2-methyl-3-pentyl, 2,3-dimethyl-2-butyl, 3,3-dimethyl-2-butyl, 1-heptyl, 1-octyl, etc.

[0120] The term "alkoxy" refers to an alkyl group as defined above, which is attached to a parent molecule through an oxygen atom. The term "Ci-Cj alkoxy" refers to an alkyl portion of an alkoxy group having i to j carbon atoms. In some embodiments, the alkoxy group contains 1 to 12 carbon atoms. In some embodiments, the alkoxy group contains 1 to 11 carbon atoms. In some embodiments, the alkoxy group contains 1 to 10 carbon atoms, 1 to 9 carbon atoms, 1 to 8 carbon atoms, 1 to 7 carbon atoms, 1 to 6 carbon atoms, 1 to 5 carbon atoms, 1 to 4 carbon atoms, 1 to 3 carbon atoms, or 1 to 2 carbon atoms.

[0121] The terms "cycloalkanes" or "cycloalkyl groups" refer to saturated monocyclic cyclic groups consisting only of carbon atoms and having a specified number of carbon atoms (e.g., C3 to C6). Cycloalkyl groups include, but are not limited to, cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, etc.

[0122] The term "heterocyclic alkyl" refers to a cyclic group having a specified number of ring atoms (e.g., 5 to 10), a specified number of heteroatoms (e.g., 1, 2, or 3), and a specified type of heteroatom (one or more of N, O, and S), which is monocyclic, bridged, or spirocyclic, and each ring is saturated. A bridged ring is a polycyclic ring that shares two or more atoms between monocyclic rings. A spirocyclic ring is a polycyclic ring that shares one atom between monocyclic rings. Heterocyclic alkyl groups include, but are not limited to, nitrogen-containing heterocyclic butyl, tetrahydropyrrolyl, tetrahydrofuranyl, morpholinyl, and piracetamyl.

[0123] The term "pharmaceutically acceptable salt" refers to a salt obtained by reacting a compound with a pharmaceutically acceptable (relatively non-toxic, safe, and suitable for patient use) acid or base. When a compound contains a relatively acidic functional group, a base addition salt can be obtained by contacting the free form of the compound with a sufficient amount of a pharmaceutically acceptable base in a suitable inert solvent. Pharmaceutically acceptable base addition salts include, but are not limited to, sodium salts, potassium salts, calcium salts, aluminum salts, magnesium salts, bismuth salts, and ammonium salts. When a compound contains a relatively basic functional group, an acid addition salt can be obtained by contacting the free form of the compound with a sufficient amount of a pharmaceutically acceptable acid in a suitable inert solvent. Pharmaceutically acceptable acid addition salts include, but are not limited to, hydrochloride salts, sulfate salts, and mesylate salts. See Handbook of Pharmaceutical Salts: Properties, Selection, and Use (P. Heinrich Stahl, 2002) for details.

[0124] The term "solvate" refers to a substance formed by the crystallization of a compound with a solvent (including but not limited to water, methanol, ethanol, etc.). Solvates are classified into stoichiometric solvates and non-stoichiometric solvates.

[0125] The term "pharmaceuticalally acceptable salt solvate" refers to a substance formed by the combination of a compound with a pharmaceutically acceptable (relatively non-toxic, safe, and suitable for patient use) acid or base, or a solvent (including but not limited to: water, methanol, ethanol, etc.). Here, "pharmaceutically acceptable salt" has the same meaning as the term "pharmaceutically acceptable salt" above, and the solvent may be stoichiometric or non-stoichiometric. Pharmaceutically acceptable salt solvates include, but are not limited to, hydrochloride monohydrates.

[0126] The term "pharmaceutical excipients" refers to excipients and additives used in the production of pharmaceuticals and the dispensing of prescriptions. They are all substances included in pharmaceutical preparations, excluding the active ingredient. See the Pharmacopoeia of the People's Republic of China (2020 edition) or Handbook of Pharmaceutical Excipients (Raymond C Rowe, 2009) for details.

[0127] Unless otherwise specified, all technical and scientific terms used herein have the standard meaning in the field to which the claimed subject matter pertains. Where multiple definitions exist for a term, the definition herein shall prevail.

[0128] It should be understood that the singular forms used in this invention, such as "a" or "an," include plural references unless otherwise specified. Furthermore, the term "comprising" is an open-ended limitation, not a closed one, meaning it includes the contents specified in this invention but does not exclude other aspects.

[0129] Unless otherwise stated, the present invention uses traditional methods of mass spectrometry and elemental analysis, and the steps and conditions can be referred to conventional operating procedures and conditions in the field.

[0130] Unless otherwise specified, the present invention employs standard nomenclature and standard laboratory procedures and techniques of analytical chemistry, organic synthetic chemistry, and optics. In some cases, standard techniques are used in chemical synthesis, chemical analysis, and performance testing of light-emitting devices.

[0131] In addition, it should be noted that, unless otherwise explicitly stated, the descriptive phrase "...independently" used in this invention should be interpreted broadly, meaning that the described individuals are independent of each other and can independently be the same or different specific groups. More specifically, the descriptive phrase "...independently" can mean that in different groups, the specific options expressed by the same symbols do not affect each other; or it can mean that in the same group, the specific options expressed by the same symbols do not affect each other.

[0132] Those skilled in the art will understand that, according to the conventions used in the art, the use of "" in the structural formula of the group described in this application means that the corresponding group is connected to other fragments or groups in the compound through that site.

[0133] Without violating common sense in the field, the above-mentioned preferred conditions can be combined arbitrarily to obtain various preferred embodiments of the present invention.

[0134] The reagents and raw materials used in this invention are all commercially available.

[0135] The positive and progressive effects of the present invention are that the present invention provides a pyrazoloquinazoline compound, its preparation method and use, the compound having at least one of the following advantages: significant inhibition effect on PLK1 and ability to further regulate drug half-life.

[0136] Detailed Implementation

[0137] The present invention will be further described below by way of embodiments, but the invention is not limited to the scope of the embodiments described herein. Experimental methods not specifically described in the following embodiments were performed according to conventional methods and conditions, or as selected according to the product instructions.

[0138] Example 1

[0139] In the first step, at room temperature, compound 1a (480 mg, 2.0 mmol) and 2-hydrazinoethanol (152 mg, 2.0 mmol) were dissolved in 5 mL of glacial acetic acid and stirred for 5 hours. After the reaction was completed, most of the glacial acetic acid was removed, and the crude product was dispersed in ethyl acetate (20 mL) and water (30 mL). The aqueous phase was then extracted with ethyl acetate (20 mL x 1), and the organic phase was washed successively with saturated sodium bicarbonate solution (20 mL x 1) and saturated brine (20 mL x 1), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain compound 1b (430 mg, yield: 85%).

[0140] In the second step, under nitrogen protection, a solution of compound 1b (800 mg, 3.2 mmol) and dimethylformamide di-tert-butyl acetal (709 mg, 3.5 mmol) in dimethylformamide (20 mL) was heated to 60 °C and reacted for 2 hours. The reaction was then complete. After cooling, water (50 mL) was added to the reaction solution, and the mixture was extracted with ethyl acetate (20 mL x 2). The organic phase was washed successively with water (20 mL x 1) and saturated brine (20 mL x 1), dried over anhydrous sodium sulfate, filtered, and concentrated to obtain compound 1c (750 mg, yield: 76%).

[0141] In the third step, under nitrogen protection, a solution of compound 1c (750 mg, 2.4 mmol) and compound 1d (728 mg, 2.4 mmol) in dimethylformamide (10 mL) was heated to 120 °C and stirred for 2 hours. The reaction was then complete. After cooling, 30 mL of water was added, and the mixture was extracted with ethyl acetate (20 mL x 2). The organic phase was washed successively with water (20 mL x 1) and saturated brine (20 mL x 1), dried over anhydrous sodium sulfate, filtered, concentrated, and the crude product was subjected to silica gel column chromatography (methanol / dichloromethane = 0 / 100%) to give compound 1e (780 mg, yield: 59%). LCMS (M+H)+ m / z: 542 / 544.

[0142] In the fourth step, at room temperature, a potassium hydroxide ethanol solution (1.5 M, 3 mL) was added to a 10 mL ethanol solution of compound 1e (780 mg, 1.4 mmol). The resulting reaction mixture was stirred for 4 hours. The reaction was then complete. The mixture was filtered to give compound 1f (650 mg, yield: 82%).

[0143] In step 5, under room temperature and nitrogen protection, 2-(7-azabenzotriazole)-N,N,N',N'-tetramethylurea hexafluorophosphate (684 mg, 1.8 mmol) was added to a solution of compound 1f (650 mg, 1.2 mmol), ammonium chloride (80 mg, 1.5 mmol), and triethylamine (310 mg, 3.6 mmol) in dimethylformamide (5 mL), and the reaction was carried out for 2 hours. The reaction was then terminated. The reaction solution was diluted with water (20 mL), extracted with ethyl acetate (20 mL x 3), and the organic phase was washed successively with water (10 mL x 1) and saturated brine (10 mL x 1), dried over anhydrous sodium sulfate, filtered, and concentrated to give 1 g of compound (510 mg, yield: 83%). LCMS (M+H)+ m / z: 513 / 515.

[0144] In step six, under nitrogen protection, a solution of 1 g (51 mg, 0.1 mmol) of compound 1, 2 mL of 1,4-dioxane dihydrochloride (24 mg, 0.12 mmol), cesium carbonate (130 mg, 0.4 mmol), 4,5-bis(diphenylphosphine)-9,9-dimethyloxanthracene (24 mg, 0.04 mmol), and tris(dibenzylacetone)palladium (18 mg, 0.02 mmol) was heated under reflux for 4 hours. The reaction was then complete. After cooling, water (10 mL) was added to the reaction solution, and ethyl acetate (15 mL x 3) was added for extraction. The organic phase was washed successively with water (10 mL x 1) and saturated brine (10 mL x 1), dried over anhydrous sodium sulfate, filtered, concentrated, and the crude product was purified by preparative HPLC to obtain compound 1 (1.5 mg, yield: 27%). LCMS(M+H)+ m / z: 559.

[0145] Example 2

[0146] In the first step, compound 2 (3.0 mg) was obtained from 1 g of compound according to the method of Example 1. LCMS (M+H)+ m / z: 559.

[0147] Example 3

[0148] In the first step, compound 3 (1.6 mg) was obtained from 1 g of compound according to the method of Example 1. LCMS (M+H)+ m / z: 545.

[0149] Example 4

[0150] In the first step, compound 4 (1.3 mg) was obtained from 1 g of compound according to the method of Example 1. LCMS (M+H)+ m / z: 545.

[0151] Example 5

[0152] In the first step, compound 5 (2.0 mg) was obtained from 1 g of compound according to the method of Example 1. LCMS (M+H)+ m / z: 545.

[0153] Example 6

[0154] In the first step, 1,2-cyclohexanedione 6F_1 (100 g, 0.892 mol) was dissolved in a mixed solvent of toluene (700 mL) and MeOH (500 mL). p-Toluenesulfonic acid (15.4 g, 89.2 mmol) was added, and the solution was stirred at 100 °C for 48 hours. The solvent was evaporated, and the residue was dissolved in DCM (500 mL) and washed with a saturated solution of NaHCO3 (500 mL). The organic phase was dried over Na2SO4 and concentrated. Purification was performed by silica gel column chromatography (0%–30% EA in PE) to obtain an oily 2-methoxycyclohexyl-2-en-1-one 6F_2 (42 g, 37.3%). ESI-MS (M+H)+=127.

[0155] In the second step, under a N2 atmosphere, LiHMDS (51 mL, 51.0 mmol, 1M in THF) was added dropwise to a solution of 2-methoxycyclohexyl-2-en-1-one (6.40 g, 45.7 mmol) in 60 mL of THF at -50 °C. After stirring at -50 °C for 30 minutes, diethyl oxalate (7.47 g, 51.0 mmol) was added. The solution was stirred at 25 °C for 16 hours. After the reaction was complete, water (100 mL) was added, and the pH was adjusted to 4-5 with 1N HCl. The resulting solution was extracted with EA (50 mL * 3). The organic layer was dried over Na2SO4 and evaporated to dryness. The crude product was purified by column chromatography (0%–15% EA in PE) to give 8.2 g (71.4%) of pale yellow oily ethyl 2-(3-methoxy-2-oxocyclohexyl-3-en-1-yl)-2-oxoethyl acetate 6F_3. ¹H NMR (400 MHz, CDCl3) δ H 14.86 (s, 1H), 5.89 (t, 1H), 4.35 (q, 2H), 3.66 (s, 3H), 2.90 (t, 2H), 2.45–2.38 (m, 2H), 1.38 (t, 3H).

[0156] In the third step, hydroxyethyl hydrazine (0.6 mL, 0.012 mmol) was added to an ethanol (30 mL) solution of ethyl 2-(3-ethoxy-2-oxocyclohexyl-3-en-1-yl)-2-oxoethyl 6F_3 (3.0 g, 0.012 mmol) at 25 °C. The reaction was stirred at 80 °C for 5 hours. The solvent was then evaporated and the residue was redissolved with DCM (30 mL). The organic layer was washed with water, dried over Na2SO4, and concentrated. The residue was dissolved in THF (12 mL) and HCl (12 mL, 1 mol / L) and stirred at 25 °C for 2 hours. The reaction was quenched with saturated NaHCO3 solution (50 mL) and effluent (EA) (50 mL x 3). The combined organic layers were washed with brine, dried over anhydrous sodium sulfate, filtered, and evaporated under reduced pressure to obtain ethyl 1-(2-hydroxyethyl)-7-oxo-4,5,6,7-tetrahydro-1H-indazole-3-carboxylate 6F_3 (2.4 g, crude product), which was a brown oil. ESI-MS (M+H)+ = 253.

[0157] In the fourth step, 6F_3 (4 g, 15.9 mmol) was dissolved in DMF (24 mL) at 25 °C, and then 8 mL of DMFDMA was added. The resulting mixture was stirred at 80 °C for 20 h, then cooled to 25 °C, concentrated to remove the solvent, and the crude product was purified by column chromatography (20%~50% EA in PE) to obtain 6F (3.25 g, 66.7%) as a yellow solid. ¹H NMR (600 MHz, DMSOd6) δH 7.49 (s, 1H), 4.83 (t, 1H), 4.60 (t, 2H), 4.26 (q, 2H), 3.72 (q, 2H), 3.12 (s, 6H), 2.95-2.80 (m, 4H), 1.29 (t, 3H).

[0158] Step 5: 5-Bromo-4-fluoro-2-methoxyaniline 6A (7.5 g, 34.3 mmol) was dissolved in EtOH (90 mL), and Ac2O (8.7 g, 85.6 mmol) was added. The mixture was stirred at 25 °C for 2.5 h, and the solvent was evaporated to obtain N-(5-bromo-4-fluoro-2-methoxyphenyl)acetamide 6A_1 (8 g crude product) as a brown solid. ESI-MS (M+H)+=262.

[0159] In step six, N-(5-bromo-4-fluoro-2-methoxyphenyl)acetamide 6A_1 (1.5 g, 5.74 mmol), Davephos (90 mg, 0.223 mmol), Pd2(dba)3 (105 mg, 0.115 mmol), and THF (7.5 mL) were added to a round-bottom flask. The flask was evacuated and backfilled with nitrogen. LiHMDS (1 mol / L in tetrahydrofuran, 12.63 mL) and N-methylpiperazine (690 mg, 6.89 mmol) were added dropwise, and the reaction was refluxed at 70 °C for 1 h. The reaction mixture was added to a saturated NH4Cl aqueous solution (10 mL), extracted with EA (10 mL * 3), the organic layer was washed with brine, dried over anhydrous sodium sulfate, filtered, and evaporated under reduced pressure. The residue was purified by column chromatography (2.5%–5% methanol, dichloromethane) to give N-(4-fluoro-2-methoxy-5-(4-methylpiperazin-1-yl)phenyl)acetamide 6A_2 (400 mg, 24.8%) as a pale yellow solid. LCMS (M+H)+=282.

[0160] In step seven, N-(4-fluoro-2-methoxy-5-(4-methylpiperazin-1-yl)phenyl)acetylamine 6A_2 (400 mg, 1.67 mmol) was dissolved in EtOH (4 mL) and stirred at room temperature. HCl (1.2 mL, 12 mol / L) was added to the solution. The mixture was stirred at 80 °C for 6 h. After the reaction was complete, 2 mol / L NaOH was added to adjust the pH of the reaction mixture to 8-9. The aqueous phase was extracted with EA (10 mL × 3), the organic layer was washed with brine, dried over Na2SO4, and concentrated. The residue was purified by column chromatography (1%-50% EA in PE) to give 4-fluoro-2-methoxy-5-(4-methylpiperazin-1-yl)aniline 6B (300 mg, 88.2%) as a yellow solid. 1HNMR (400 MHz, DMSO-d6) δ H 6.68 (d, 1H), 6.37 (d, 1H), 4.47 (s, 2H), 3.69 (s, 3H), 2.85 (brs, 4H), 2.43 (brs, 4H), 2.20 (s, 3H).

[0161] In step eight, 4-fluoro-2-methoxy-5-(4-methylpiperazin-1-yl)aniline 6B (150 mg, 0.514 mmol) was dissolved in hydrochloric acid (6 mol / L, 1 mL), cyanamide (345 mg, 4.11 mmol, 50% in H2O) was added, and the reaction was stirred at 80 °C for 48 H. The mixture was cooled to 25 °C, diluted with water (3 mL), and extracted with dichloromethane. The pH of the aqueous phase was adjusted to >11 by adding 2 mol / L NaOH. The aqueous phase was extracted with EA (10 mL * 3), dried over Na2SO4, and concentrated to obtain 1-(4-fluoro-2-methoxy-5-(4-methylpiperazin-1-yl)phenyl)guanidine 6C (150 mg, crude product) as a brown solid.

[0162] In step nine, 1-(4-fluoro-2-methoxy-5-(4-methylpiperazin-1-yl)phenyl)guanidine 6C (300 mg, 0.98 mmol) and 6-((dimethylamino)methylene)-1-(2-hydroxyethyl)-7-oxo-4,5,6,7-tetrahydro-1H-indazole-3-carboxylic acid ethyl ester 6F were stirred in DMF solution (3.5 mL) at 25 °C. After stirring at 80 °C for 12 hours, the reaction mixture was evaporated under reduced pressure. The residue was purified by column chromatography (2%~5% MeOH in DCM solution) to give ethyl 8-((4-fluoro-2-methoxy-5-(4-methylpiperazin-1-yl)phenyl)amino)-1-(2-hydroxyethyl)-4,5-dihydro-1H-pyrazolo[4,3-h]quinoxaline-3-carboxylic acid ethyl ester 6D (150 mg, 26.7%) as a pale yellow solid. LCMS(M+H)+=526.

[0163] In step ten, the solution of ethyl 8-((4-fluoro-2-methoxy-5-(4-methylpiperazin-1-yl)phenyl)amino)-1-(2-hydroxyethyl)-4,5-dihydro-1H-pyrazolo[4,3-h]quinoxaline-3-carboxylic acid ethyl ester 6D (110 mg, 0.222 mmol) in NH3 MeOH solution (1.5 mL, 7 M) was stirred in a 10 mL sealed tube at 80 °C for 36 hours. After the reaction was completed, the reaction mixture was evaporated under reduced pressure, and the residue was purified by column chromatography (DCM:MeOH=10:1) to give 8-((4-fluoro-2-methoxy-5-(4-methylpiperazin-1-yl)phenyl)amino)-1-(2-hydroxyethyl)-4,5-dihydro-1H-pyrazolo[4,3-h]quinazolin-3-methamide 6 (12.0 mg, 11.5%) as a light yellow solid. 1HNMR (400 MHz, CD3OD) δ H 8.27 (s, 1H), 7.70 (d, 1H), 6.88 (d, 1H), 4.74 (s, 2H), 3.87 (t, 2H), 3.84 (s, 3H), 3.13-3.03 (m, 6H), 2.91-2.86 (m, 2H), 2.65 (brs, 4H), 2.36 (s, 3H). LCMS(M+H)+=497.2.

[0164] Example 7

[0165] In the first step, under a N2 atmosphere, guanidine carbonate (849.4 mg, 4.64 mmol) was added to a solution of (Z)-6-((dimethylamino)methylene)-1-(2-hydroxyethyl)-7-oxo-4,5,6,7-tetrahydro-1H-indazole-3-carboxylate 6F (570 mg, 1.88 mmol) in DMF (50 mL). After stirring at 110°C for 16 hours, the reaction mixture was added to water (50 mL), filtered, and dried under vacuum. The residue was purified by column chromatography (20%–50% EA / PE) to give ethyl 8-amino-1-(2-hydroxyethyl)-4,5-dihydro-1H-pyrazolo[4,3-h]quinazolin-3-carboxylate 7F_1 (307 mg, 53.8%) as a pale yellow solid. ESI-MS (M+H)+ = 304.

[0166] In the second step, under N2, cuprous iodide (57.1 mg, 0.300 mmol), iodine (126.5 mg, 0.500 mmol), cesium iodide (259 mg, 1.00 mmol), and isoamyl nitrite (176 mg, 1.50 mmol) were added to a THF (30 mL) solution of ethyl 8-amino-1-(2-hydroxyethyl)-4,5-dihydro-1H-pyrazolo[4,3-h]quinazolin-3-carboxylate 7F_1 (303 mg, 1.00 mmol). The reaction mixture was stirred at 80 °C for 12 hours. The reaction mixture was diluted with DCM (50 mL), washed with 20% NH3·H2O (80 mL), saturated Na2S2O3 aqueous solution (80 mL), and brine (80 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by column chromatography (2.5%–10% MeOH in DCM solution) to give ethyl 1-(2-hydroxyethyl)-8-iodo-4,5-dihydro-1H-pyrazolo[4,3-h]quinazolin-3-carboxylate 7F_2 (300 mg, 52%) as a yellow solid. ESI-MS (M+H)+=415.

[0167] In the third step, 27 g (104 mmol) of 1-bromo-2-fluoro-4-(trifluoromethoxy)benzene 7A was placed in a round-bottom flask, and potassium nitrate (14 g, 138 mmol) and concentrated sulfuric acid (60 ml) were added under N2. The reaction mixture was stirred at 25°C for 2 hours. The reaction mixture was quenched with water (200 mL), extracted with DCM (50 mL*3), and the combined organic layers were washed with saturated NaHCO3 aqueous solution (100 mL) and brine (100 mL), dried over anhydrous sodium sulfate, filtered, and evaporated under reduced pressure. The residue was purified by column chromatography (0%~5% EA in PE) to give 30 g (95%) of 1-bromo-2-fluoro-5-nitro-4-(trifluoromethoxy)benzene 7B, which was a pale yellow oil. ESI-MS (M+H)+=303.

[0168] In the fourth step, under N2, iron powder (22.0 g, 0.393 mol) was added to a solution of 1-bromo-2-fluoro-5-nitro-4-(trifluoromethoxy)benzene 7B (25.0 g, 12.1 mmol) in AcOH (50 mL) and EtOH (50 mL). The reaction was stirred at 25 °C for 3 hours. The reaction mixture was concentrated and diluted with water (100 mL), extracted with EA (30 mL*3), the combined organic layers were washed with brine, dried over anhydrous sodium sulfate, filtered, and evaporated under reduced pressure. The residue was purified by column chromatography (5%~20% EA in PE) to give 5-bromo-4-fluoro-2-(trifluoromethoxy)aniline 7C (21.2 g, 93.8%) as a yellow solid. 1HNMR (400 MHz, CDCl3) δ H 7.00-6.95 (m, 2H), 3.79 (brs, 2H). ESI-MS(M+H)+=274.1.

[0169] In the fifth step, compound 7D was obtained from compound 7C through two steps of synthesis, following the synthesis method of Example 6.

[0170] Step 6: Under N2, 4-fluoro-5-(4-methylpiperazin-1-yl)-2-(trifluoromethoxy)aniline 7D (60 mg, 0.144 mmol), Pd(OAc)2 (3.2 mg, 0.0144 mmol), BINAP (18.0 mg, 0.0289 mmol), and K2CO3 (59.6 mg, 0.432 mmol) were added to a solution of 1-(2-hydroxyethyl)-8-iodo-4,5-dihydro-1H-pyrazolo[4,3-h]quinazolin-3-carboxylic acid ethyl ester 7F_2 (126.6 mg, 0.432 mmol) in DMF (6 ml). The reaction mixture was stirred at 80 °C for 4 hours. The reaction mixture was concentrated and diluted with water (20 mL), extracted with EA (10 mL * 3), the combined organic layers were washed with brine, dried over anhydrous sodium sulfate, filtered, and evaporated under reduced pressure. The residue was purified by column chromatography (10% MeOH in DCM) to give ethyl 8-((4-fluoro-5-(4-methylpiperazin-1-yl)-2-(trifluoromethoxy)phenyl)amino)-1-(2-hydroxyethyl)-4,5-dihydro-1H-pyrazolo[4,3-h]quinazolin-3-carboxylate 7F (75 mg, 16%) as a pale yellow solid. ESI-MS (M+H)+=580.

[0171] Step 7: Following the synthesis method of Example 6, compound 7 (5.2 mg, 7.3%) was obtained from compound 7F (75 mg, 0.129 mmol) via a one-step reaction. ¹H NMR (400 MHz, CD3OD) δH 8.27 (s, 1H), 7.45 (d, 1H), 7.17 (d, 1H), 4.78–4.74 (m, 2H), 3.81 (t, 2H), 3.26–3.21 (m, 4H), 3.10–3.05 (m, 2H), 2.90–2.83 (m, 2H), 2.67–2.62 (m, 4H), 2.36 (s, 3H). LCMS (M+H)+ m / z = 551.2.

[0172] Example 8

[0173] In the first step, compound 8A was obtained from compound 8-1 through three steps, following the synthesis method of Example 6. ¹H NMR (400 MHz, DMSO d6) δ H 8.15 (s, 1H), 7.95 (d, 1H), 7.26-7.15 (m, 9H), 7.05 (d, 6H), 6.87 (d, 1H), 4.23 (q, 2H), 3.80 (s, 3H), 3.00 (t, 2H), 2.88-2.81 (m, 4H), 2.67-2.58 (m, 2H), 2.53-2.35 (m, 7H), 1.24 (t, 3H).

[0174] In the second step, TFA (0.5 mL) was added to 8-((4-fluoro-5-(4-methylpiperazin-1-yl)-2-(trifluoromethoxy)phenyl)amino)-1-(2-hydroxyethyl)-4,5-dihydro-1H-pyrazolo[4,3-h]quinazolin-3-carboxylic acid ethyl ester 8A (100 mg, 0.138 mmol) in DCM (5 mL) solution at 25 °C, and stirred at 25 °C for 4 hours. The reaction was quenched by adding saturated NaHCO3 aqueous solution (30 mL), extracted with DCM (15 mL * 2), the organic layer was washed with brine, dried over anhydrous sodium sulfate, filtered, evaporated under reduced pressure, and the residue was purified by column chromatography (2%–8% MeOH in DCM solution) to give ethyl 8-((4-fluoro-2-methoxy-5-(4-methylpiperazin-1-yl)phenyl)amino)-4,5-dihydro-1H-pyrazolo[4,3-h]quinazolin-3-carboxylate 8B (40 mg, 60.2%). ESI-MS (M+H)+ = 482.

[0175] The third step is to obtain compound 8C by starting from compound 8B through a one-step reaction, following the synthesis method of Example 6.

[0176] In the third step, at 0°C, Cs₂CO₃ (32.6 mg, 0.1 mmol) and 2-chloroacetonitrile (6.0 mg, 0.08 mmol in 1 mL DMF) were added to a DMF solution of 8-((4-fluoro-5-(4-methylpiperazin-1-yl)-2-(trifluoromethoxy)phenyl)amino)-1-(2-hydroxyethyl)-4,5-dihydro-1H-pyrazolo[4,3-h]quinazolin-3-methamide 8C (30 mg, 0.066 mmol). After stirring at 25°C for 4 hours, the reaction mixture was quenched with water (10 mL). The mixture was extracted with EA (15 mL * 2), the combined organic layers were washed with brine, dried over anhydrous sodium sulfate, filtered, and evaporated under reduced pressure. The residue was purified by column chromatography (DCM:MeOH=15:1) to give 1-(cyanomethyl)-8-((4-fluoro-2-methoxy-5-(4-methylpiperazin-1-yl)phenyl)amino)-4,5-dihydro-1H-pyrazolo[4,3-h]quinazolin-3-methamide 8 (16.7 mg, 51.2%) as a pale yellow solid. 1HNMR (400 MHz, CD3OD) δ H 8.31 (s, 1H), 7.68 (d, 1H), 6.88 (d, 1H), 5.88 (s, 2H), 3.84 (s, 3H), 3.15-3.05 (m, 6H) ), 2.90 (t, 2H), 2.63-2.58 (m, 4H), 2.33 (s, 3H). LCMS [M+H]+: 492.3.

[0177] Example 9

[0178] Compound 9 was obtained from compounds 8-3 and 7D via a four-step reaction according to the synthetic method of Example 8. ¹H NMR (400 MHz, CD3OD) δH 8.30 (s, 1H), 7.45 (d, 1H), 7.14 (d, 1H), 5.80 (s, 2H), 3.18–3.05 (m, 6H), 2.90 (t, 2H), 2.68–2.60 (m, 4H), 2.36 (s, 3H). LCMS [M+H]+: 446.3.

[0179] Example 10

[0180] Compound 10 was obtained from compounds 8-3 and 10-1 via a four-step reaction following the synthetic methods of Examples 7 and 8. ¹H NMR (400 MHz, CD3OD) δH 8.37 (s, 1H), 7.54 (s, 1H), 7.26 (d, 1H), 6.85 (d, 1H), 5.87 (s, 2H), 3.31–3.25 (m, 4H), 3.14 (t, 2H), 2.96 (t, 2H), 2.71–2.63 (m, 4H), 2.38 (s, 3H). LCMS [M+H]+: 528.2.

[0181] Example 11

[0182] According to the synthesis method of Example 9, compound 11 was obtained from compound 10C through a one-step reaction. ¹H NMR (400 MHz, CD3OD) δH 8.31 (s, 1H), 7.45 (s, 1H), 7.25-7.20 (m, 1H), 6.85-6.77 (m, 1H), 5.10-5.02 (m, 2H), 4.62-4.51 (m, 2H), 4.34-4.28 (m, 1H), 3.26 (brs, 4H), 3.15-3.08 (m, 2H), 2.94-2.86 (m, 2H), 2.61 (brs, 4H), 2.55-2.49 (m, 1H), 2.36 (s, 3H). 2.35-2.29 (m, 1H). LCMS [M+H]+: 559.2.

[0183] Example 12

[0184] In the first step, methyl 1-hydroxycyclopropane-1-carboxylate 12A (3.0 g, 25.8 mmol) was dissolved in MeOH (15 mL), and hydrazine hydrate (6.37 g, 258 mmol) was added. The mixture was stirred at 60 °C for 16 hours, and the solvent was evaporated. The residue was ground together with EA (30 mL) for 0.5 hours and filtered to obtain a white solid 1-hydroxycyclopropane-1-carboxyhydrazine 12E_1 (2.8 g, 93.3%). LCMS (M+H)+=117.

[0185] In the second step, under a N2 atmosphere and at -5-0°C, a suspension of 1-hydroxycyclopropane-1-methylhydrazine 12E_1 (3.0 g, 258 mmol) in THF (15 mL) was added dropwise to BH3·THF (258 mL, 258 mmol, 1 M). The reaction mixture was stirred at 60°C for 21 hours. MeOH (50 mL) was added dropwise to the reaction mixture under N2 protection at -5-0°C, and then stirred at 20-25°C for 0.5 hours. The reaction mixture was concentrated to obtain a residue. The residue was dissolved in 50 mL of MeOH and stirred at 80°C for 3 hours. The mixture was cooled to 25°C and evaporated to obtain a residue. The residue was ground together with DCM (30 mL) for 0.5 hours and filtered. The filtrate was evaporated to give a pale yellow oily crude product, 1-(hydrazylmethyl)cyclopropane-1-ol 12E_2 (2.2 g, 75%). 1H NMR (400 MHz, CDCl3) d2.91(s, 2H), 0.83 (t, J = 6.0 Hz, 2H), 0.55-0.50 (m, 2H).

[0186] The third step followed the synthesis method of Example 9, starting from compound 12E_2 and compound 6F_3, and obtained compound 12 through a 4-step reaction. 1H NMR (400 MHz, CD3OD) δ H 8.28 (s, 1H), 7.44 (s, 1H), 7.20 (d, 1H), 6.79 (d, 1H), 4.74 (s, 2H), 3.30-3.24 (m, 4H), 3.12-3.05 (m, 2H), 2.90-2.84 (m, 2H), 2.63-2.58 (m, 4H), 2.33 (s, 3H), 0.59-0.54 (m, 2H), 0.49-0.46 (m, 2H). LCMS [M+H]+: 559.2.

[0187] Example 13

[0188] According to the synthesis method of Example 9, compound 13 was obtained from compound 10C through a one-step reaction. 1H NMR (400 MHz, DMSO-d6) δ 9.12 (s, 1H), 8.38 (s, 1H), 7.49 – 7.21 (m, 5H), 6.92 (dd, J = 9.1, 3.0 Hz, 1H), 4.79 (d, J = 7.0 Hz, 2H), 4.40 (t, J = 7.0 Hz, 2H), 4.31 (t, J = 6.2 Hz, 2H), 3.56 (d, J = 12.3 Hz, 3H), 3.20 (ddt, J = 26.3, 15.4, 7.4 Hz, 4H), 3.00 (t, J = 7.5 Hz, 4H), 2.90 (s, 4H), 2.83 (t, J = 7.7 Hz, 3H).

[0189] Example 14

[0190] According to the synthesis method of Example 9, compound 14 was obtained from compound 10C through a one-step reaction. 1H NMR (400 MHz, CD3OD-d4) δ ppm 8.51 (s, 1H), 7.53 (d, J = 2.9 Hz, 1H), 7.36 – 7.21 (m, 1H), 6.87 (dd, J = 9.1, 3.0 Hz, 1H), 4.76 (d, J = 12.7 Hz, 2H), 4.50 (d, J = 13.2 Hz, 1H), 4.42 (d, J = 12.7 Hz, 1H), 3.65 (d, J = 2.0 Hz, 2H), 3.26 (dt, J = 18.0, 6.2 Hz, 6H), 3.06 (t, J = 7.7 Hz, 2H), 2.67 (t, J = 5.0 Hz, 4H), 2.40 (s, 3H), 1.42 (s, 3H). LCMS [M+H]+: 573.2.

[0191] Example 15

[0192] Compound 15 was obtained from compound 10C via a one-step reaction according to the synthetic method of Example 9. ¹H NMR (400 MHz, CD3OD) δH 8.35 (s, 1H), 7.53 (s, 1H), 7.26 (d, 1H), 6.85 (d, 1H), 5.00–4.88 (m, 1H), 4.65–4.58 (m, 1H), 3.29 (s, 4H), 3.15–3.08 (m, 2H), 2.93 (t, 2H), 2.80 (s, 4H), 2.49 (s, 3H), 2.21–2.12 (m, 1H), 1.39–1.32 (m, 2H). LCMS [M+H]+: 579.2.

[0193] Example 16

[0194] In the first step, at 0°C, Cs₂CO₃ (650 mg, 2 mmol) and (2-bromoethoxy)tert-butyldimethylsilane (480 mg, 2 mmol) were added to a DMF (5 mL) solution of 16A (193 mg, 1 mmol). The mixture was stirred overnight at 25°C, and the reaction mixture was quenched with water (10 mL). The mixture was extracted with EA (15 mL x 2), the combined organic layers were washed with brine, dried over anhydrous sodium sulfate, filtered, and evaporated under reduced pressure. The residue was purified by column chromatography (PE:EA = 5:1) to give 16B (220 mg, 64%) as a white solid.

[0195] In the second step, 16B 510 mg was treated with tris(dimethylamino)methane (2 mL), and the reaction was stirred overnight at 90 °C. The volatiles were removed under reduced pressure, and the residue was used without further purification.

[0196] The third step is to obtain compound 16D by starting from compound 16B and compound 15-1 through a one-step reaction, according to the synthesis method of Example 10.

[0197] In the fourth step, methyl 9-(2-((tert-butyldimethylsilyl)oxy)ethyl)-2-((5-(4-methylpiperazin-1-yl)-2-(trifluoromethoxy)phenyl)amino)-6,9-dihydro-5H-pyrrolo[3,2-h]quinazolin-7-carboxylic acid methyl ester 16D (330 mg, 0.5 mmol) was suspended in dioxane (5 mL) and treated overnight at reflux with 2N NaOH solution (5 mL, 10 mmol). H2O (50 mL) was added and the solution was acidified to pH 4 with 2N HCl. Concentrate under reduced pressure, and the residue was purified by column chromatography (DCM:MeOH 10:1) to give a yellow solid 9-(2-hydroxyethyl)-2-((5-(4-methylpiperazin-1-yl)-2-(trifluoromethoxy)phenyl)amino)-6,9-dihydro-5H-pyrrolo[3,2-h]quinazolin-7-carboxylic acid (240 mg, 90% yield). LC-MS (M+H)+: 533.

[0198] In the fifth step, 9-(2-hydroxyethyl)-2-((5-(4-methylpiperazin-1-yl)-2-(trifluoromethoxy)phenyl)amino)-6,9-dihydro-5H-pyrrolo[3,2-h]quinazolin-7-carboxylic acid (240 mg, 0.45 mmol) was dissolved in DMF. HOBt (121 mg, 0.9 mmol) and EDCI (172 mg, 0.9 mmol) were added to the reaction mixture, and the mixture was stirred for 0.5 h. Subsequently, NH4OH (4 eq) was added. The reaction mixture was stirred at room temperature for 14 hours, and then concentrated under reduced pressure. The mixture was then subjected to preparative HPLC to obtain 9-(2-hydroxyethyl)-2-((5-(4-methylpiperazin-1-yl)-2-(trifluoromethoxy)phenyl)amino)-6,9-dihydro-5H-pyrrolo[3,2-h]quinazolin-7-methamide, a white solid (106 mg, 47% yield). 1H NMR (400 MHz, DMSO-d6) δ 8.52 (s, 1H), 8.14 (s, 1H), 7.55 (s, 1H), 7.24 (d, J = 3.0 Hz, 1H), 7.21 (dq, J = 9.1, 1.4 Hz, 1H), 6.79 (d, J = 3.0 Hz, 1H), 6.77 (d, J = 3.0 Hz, 1H), 4.71 (t, J = 5.3 Hz, 1H), 4.35 (t, J = 5.0 Hz, 2H), 3.46 – 3.40 (m, 3H), 3.15 (dd, J = 6.4, 3.7 Hz, 4H), 2.98 (t, J = 7.7 Hz, 2H), 2.72 (t, J = 7.7 Hz, 2H), 2.46 (t, J = 5.0 Hz, 4H), 2.24 (s, 3H).

[0199] Example 17

[0200] In the first step, at 5°C, LiHMDS (1M THF solution, 6.06 mL, 9.09 mmol) was added dropwise to a THF (24 mL) solution of cyclopropylmethanol 17A (721.00 μL, 9.09 mmol). After 30 minutes, 4-fluoro-3-nitrobobromobenzene (1.11 mL, 9.09 mmol) was rapidly added, and the reaction mixture was allowed to warm naturally to room temperature and stirred overnight. The reaction mixture was quenched with water and extracted with EA. The organic layer was collected, washed with brine, dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by silica gel column chromatography (PE / EA, 100:0 to 20:1) to give 3.02 g of 4-bromo-1-(cyclopropylmethoxy)-2-nitrobenzene 17B. LCMS(M+H)+=272.

[0201] In the second step, under a N2 atmosphere, 4-bromo-1-(cyclopropylmethoxy)-2-nitrobenzene (587 mg, 2.16 mmol), XANTPHOS (0.37 g, 0.65 mmol), Pd2(dba)3 (0.39 g, 0.43 mmol, Aldrich), Cs2CO3 (1.4 g, 4.3 mmol) and 1-methylpiperazine (0.43 g, 4.3 mmol) were added to a dry 1,4-dioxane (20 mL). After heating at 90°C overnight, the reaction was quenched with water, extracted with ethyl acetate (50 mL × 3), dried over sodium sulfate, concentrated, and purified by column chromatography (dichloromethane / methanol 20:1) to give 1-(4-(cyclopropylmethoxy)-3-nitrophenyl)-4-methylpiperazine 17C (383 mg, 1.32 mmol, 61%) as a brown solid. ESI-MS (M+H)+=292.

[0202] In the third step, 1-(4-(cyclopropylmethoxy)-3-nitrophenyl)-4-methylpiperazine 17C (291 mg, 1 mmol) was dissolved in a mixed solvent of acetic acid (10 mL) and ethanol (10 mL), and zinc powder (650 mg, 10 mmol) was added. The mixture was stirred at 80 °C for 3 h, filtered through diatomaceous earth, concentrated, and purified by column chromatography (dichloromethane / methanol 20:1) to obtain 2-(cyclopropylmethoxy)-5-(4-methylpiperazine-1-yl)aniline 17D (185 mg, 0.71 mmol, 71%). ESI-MS (M+H)+=262.

[0203] In the fourth step, following the synthesis method of Example 7, compound 17,2-((2-(cyclopropylmethoxy)-5-(4-methylpiperazin-1-yl)phenyl)amino)-9-(2-hydroxyethyl)-6,9-dihydro-5H-pyrrolo[3,2-h]quinazolin-7-methamide was obtained through a two-step reaction starting from compound 17D and 7F_2. 1H NMR (400 MHz, DMSO-d6) δ 8.43 (s, 1H), 8.05 (s, 1H), 7.74 (d, J = 2.8 Hz, 1H), 7.50 (s, 1H), 7.29 (s, 1H), 6.94 (d, J = 8.9 Hz, 1H), 6.60 (dd, J = 8.8, 2.9 Hz, 1H), 4.78 (t, J = 5.6 Hz, 2H), 3.85 (d, J = 6.9 Hz, 2H), 3.80 (t, J = 5.6 Hz, 2H), 3.05 (t, J = 5.0 Hz, 4H), 3.00 (d, J = 7.7 Hz, 2H), 2.83 (t, J = 7.6 Hz, 2H), 2.49 (t, J = 4.9 Hz, 4H), 2.25 (s, 3H), 1.29 – 1.18 (m, 1H), 0.60 – 0.51 (m, 2H), 0.36 – 0.28 (m, 2H). LCMS [M+H]+: 519.3.

[0204] Example 18

[0205] 18A was prepared by the method described in patent CN101563351B. A stirred solution of 18A (100 mg, 0.178 mmol) in 2 mL of THF was added to a 50 mL sealed tube containing 28 mg, 0.712 mmol, of LiAlH4. After stirring at -78°C for 24 hours, the reaction mixture was cooled to 0°C, and then 20 mL of water was added. The mixture was extracted with EA (20 mL x 3), the combined organic layers were washed with brine, dried over anhydrous sodium sulfate, filtered, and evaporated under reduced pressure. The residue was purified by column chromatography (2.5%–10% MeOH in DCM solution) to give compound 18 (15 mg, 16.2%) as a pale yellow solid. 1HNMR (400 MHz, CD3OD) δH 8.27 (s, 1H), 7.54 (d, 1H), 7.20 (d, 1H), 6.79 (d, 1H), 4.71 (t, 2H), 4.60 (s, 2H),3.79 (t, 2H), 3.28-3.21 (m,4H), 2.90-2.80 (m, 4H), 2.66 (m, 4H), 2.38 (s, 3H). LC-MS[M+H]+: 520.20.

[0206] Example 19

[0207] 18A (0.15 g, 0.267 mmol) was stirred in a 6 mL DCM solution at 25 °C in a 50 mL sealed tube with EDCI (70.5 mg, 0.347 mmol), DMAP (16.3 mg, 0.134 mmol), and O-methyl-hydroxylamine chloride (35.3 mg, 0.4 mmol). The mixture was stirred at 30 °C for 2 hours, concentrated to remove the solvent, and the crude product was purified by column chromatography (DCM:MeOH = 10:1) to give 19 (30 mg). 1HNMR (400 MHz, CD3OD) δH 8.30 (s, 1H), 7.51 (d, 1H), 7.21 (d, 1H), 6.79 (d, 1H), 4.76 (t, 2H), 3.82 (t, 2H), 3.79 (s, 3H), 3.30-3.22 (m, 4H), 3.07 (t, 2H), 2.88 (t, 2H), 2.64-2.61 (m, 4H), 2.35 (s, 3H). LC-MS[M+H]+: 563.30.

[0208] Example 20

[0209] In a round-bottom flask, 17A (5 g, 0.023 mol), cyclopropanol (1.32 g, 0.023 mol), K2CO3 (9.42 g, 0.068 mol), and DMF (10 mL) were charged. The reaction mixture was stirred overnight at 25 °C. The reaction mixture was quenched by adding H2O (100 mL), extracted with EA (100 mL × 3), the combined organic layers were washed with brine, dried over anhydrous sodium sulfate, filtered, and evaporated under reduced pressure. The residue was purified by column chromatography (5%–10% EA in PE) to give 20A (5.3 g, 90.4%) as a yellow solid.

[0210] Compound 14 was obtained from compound 20A by a four-step reaction according to the synthesis method of Example 17. 1HNMR (400 MHz, CD3OD) δH 8.30 (s, 1H), 7.82 (d, 1H), 7.24 (d, 1H), 6.72 (dd, 1H), 4.90-4.88 (m, 2H), 3.94 (t, 2H), 3.85-3.82 (m, 1H), 3.15-3.12 (m, 4H), 3.07 (t, 2H), 2.87 (t, 2H), 2.71-2.66 (m, 4H), 2.39 (s, 3H), 0.79-0.77 (m, 2H), 0.74-0.72 (m, 2H). LC-MS[M+H]+: 505.30.

[0211] Example 21

[0212] PLK1 kinase activity test

[0213] Experimental materials: PLK1 Active was purchased from CARNA; Casein Protein was purchased from SignalChem; ADP-Glo ​​Kinase Assay was purchased from Promega; Kinase assay buffer III was purchased from SignalChem; Nivo multilabel analyzer (PerkinElmer).

[0214] Experimental Method:

[0215] Use the kinase buffer provided in the kit to dilute the enzyme, substrate, ATP, and inhibitor. Dilute the test compound to 1 mM with 100% DMSO as the first concentration, and then perform a 5-fold dilution to the 8th concentration using a multi-channel pipette, i.e., from 1 mM to 0.013 μM. Prepare working solutions containing 5% DMSO by 20-fold dilution of each concentration point of the compound using 1X kinase buffer. Add 1 μL of the working solution for each concentration gradient to a microplate and set up duplicate wells. Add 2 μl of PLK1 enzyme (15 ng), 2 μl of a mixture of substrate and ATP (20 μM ATP, 0.2 μg / μl Casein protein) to the microplate. At this point, the final concentration gradient of the compound is 10 μM diluted to 0.13 nM. Incubate the reaction system at 25°C for 60 minutes. After the reaction was complete, 5 μl of ADP-Glo ​​reagent was added to each well, and the reaction was continued at 25 degrees Celsius for 40 minutes. After the reaction was completed, 10 μL of kinase detection reagent was added to each well, and the reaction was continued at 25 degrees Celsius for 30 minutes. The chemiluminescence was then read using a PerkinElmer Nivo multilabel analyzer with an integration time of 0.5 seconds.

[0216] Data Analysis:

[0217] The original data is converted into inhibition rate by using the equation (Sample-Min) / (Max-Min)*100%. The IC50 value can then be obtained by curve fitting using four parameters (obtained in the log(inhibitor) vs. response -- Variable slope mode in GraphPad Prism).

[0218] shows the PLK1 kinase activity test results of the compounds of this application. Compound numbering PLK1 IC 50 (nM) 6 47.17 7 101.8 8 824.9 9 526 10 9.55 11 8.57 12 9.67 13 18.94 14 50.12 15 14.62 16 6.39 17 9.71

[0219] In vitro cell proliferation analysis

[0220] The inhibitory activity of the compound on tumor cell proliferation was determined using the MTT assay. HT-29 tumor cells in the logarithmic growth phase were seeded into culture plates at a certain cell volume and cultured for 24 h. Different concentrations of inhibitor were added, and the cells were cultured for another 48 h at 37 °C and 5% CO2. 20 μL of MTT solution was added to each well and the cells were cultured for another 4 h. The cells were then dissolved and crystallized in DMSO, and the OD value was measured at 570 nm using an enzyme-linked immunosorbent assay (ELISA) instrument to calculate the IC50.

[0221] shows the test results of the HT-29 tumor cell proliferation inhibition activity of the compound of this application. Compound numbering HT-29 IC 50 (nM) 6 269 10 111 11 232.2 12 75 15 71

Claims

1. A pyrazoloquinazoline compound as shown in Formula I, or a pharmaceutically acceptable salt thereof: wherein, R1 is a C1-C4 alkoxy, a C1-C4 alkoxy substituted with one or more R1-1, or a 3-6 membered cycloalkyloxy; R1-1 is independently a halogen or a 3-6 membered cycloalkane; R2 is H, a halogen, or a C1-C4 alkyl; R3 is a 5-10 membered heterocyclic alkyl or a 5-10 membered heterocyclic alkyl substituted with one or more R3-1; in this 5-10 membered heterocyclic alkyl, the heteroatom is selected from one or more of N, O, and S, and the number of heteroatoms is 1-2; R3-1 is independently a C1-C4 alkyl. R4 is a cyano group, a 3- to 6-membered cycloalkane, a 3- to 6-membered cycloalkane substituted with one or more R4-1 groups, a 3- to 6-membered heterocycloalkane, or a 3- to 6-membered heterocycloalkane substituted with one or more R4-2 groups; wherein the heteroatom is selected from one or more of N, O, and S, and the number of heteroatoms is 1 to 2; R4-1 is independently a halogen, a hydroxyl group, or a C1 to C4 alkyl group; R4-2 is independently a halogen or a C1 to C4 alkyl group; R5 is H, a hydroxyl group, a C1 to C4 alkyl group, or a C1 to C4 alkoxy group.

2. A pyrazoloquinazoline compound of formula I as described in claim 1, or a pharmaceutically acceptable salt thereof, wherein, R1-1 is independently F, Cl, Br, or I; and / or, when R1-1 is independently a 3- to 6-membered cycloalkane, the 3- to 6-membered cycloalkane is cyclopropane, cyclobutane, or cyclopentane; and / or, when R1 is a C1- to C4 alkoxy group, the C1- to C4 alkoxy group is methoxy, ethoxy, isopropoxy, or tert-butoxy; and / or, when R1 is a C1- to C4 alkoxy group substituted by one or more R1-1 groups, the C1- to C4 alkoxy group is methoxy, ethoxy, isopropoxy, or tert-butoxy; and / or, when R1 is a 3- to 6-membered cycloalkyloxy group, the 3- to 6-membered cycloalkoxy group is cyclopropyloxy, cyclobutyloxy, or cyclopentyloxy; and / or, when R2 is a halogen, the halogen is F, Cl, Br, or I. And / or, when R2 is a C1-C4 alkyl group, the C1-C4 alkyl group is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, or tert-butyl; and / or, when R3-1 is a C1-C4 alkyl group, the C1-C4 alkyl group is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, or tert-butyl; and / or, when R3 is a 5-10 membered heterocyclic alkyl group substituted by one or more R3-1 groups, the 5-10 membered heterocyclic alkyl group is a 6-9 membered heterocyclic alkyl group; and / or, when R4-1 is a halogen, the halogen is F, Cl, Br, or I; and / or, when R4-1 is a C1-C4 alkyl group, the C1-C4 alkyl group is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, or tert-butyl; And / or, when R4-2 is a C1-C4 alkyl group, the C1-C4 alkyl group is methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, or tert-butyl; and / or, when R4-2 is a halogen, the halogen is F, Cl, Br, or I; and / or, when R4 is a 3-6 membered cycloalkane, the 3-6 membered cycloalkane is cyclopropane, cyclobutane, or cyclopentane; and / or, when R4 is a 3-6 membered cycloalkane substituted with one or more R4-1 groups, the 3-6 membered cycloalkane is cyclopropane, cyclobutane, or cyclopentane; and / or, when R4 is a 3-6 membered heterocyclic alkane substituted with one or more R4-2 groups, the 3-6 membered heterocyclic alkane is a 4-membered heterocyclic alkane containing one N or O or a 5-membered heterocyclic alkane containing one N or O; And / or, when R5 is a C1-C4 alkyl group, the C1-C4 alkyl group is methyl, ethyl, isopropyl or tert-butyl; and / or, when R5 is a C1-C4 alkoxy group, the C1-C4 alkoxy group is methoxy, ethoxy, isopropoxy or tert-butoxy.

3. A pyrazoloquinazoline compound of formula I as described in claim 1 or 2, or a pharmaceutically acceptable salt thereof, wherein, R1-1 is independently F; and / or, when R1-1 is independently a 3- to 6-membered cycloalkane, the 3- to 6-membered cycloalkane is cyclopropane or cyclobutane; and / or, when R1 is a C1- to C4 alkoxy, the C1- to C4 alkoxy is methoxy; and / or, when R1 is a C1- to C4 alkoxy substituted by one or more R1-1, the C1- to C4 alkoxy is methoxy; and / or, when R1 is a 3- to 6-membered cycloalkyloxy, the 3- to 6-membered cycloalkoxy is cyclopropyloxy or cyclobutyloxy; and / or, when R2 is a halogen, the halogen is F; and / or, when R2 is a C1- to C4 alkyl, the C1- to C4 alkyl is methyl; and / or, when R3-1 is a C1- to C4 alkyl, the C1- to C4 alkyl is methyl; And / or, when R3 is a 5- to 10-membered heterocyclic alkyl group substituted with one or more R3-1, the 5- to 10-membered heterocyclic alkyl group is a 6- to 9-membered heterocyclic alkyl group containing two N atoms; and / or, when R4-1 is a halogen, the halogen is F; and / or, when R4-1 is a C1- to C4 alkyl group, the C1- to C4 alkyl group is methyl or ethyl; and / or, when R4-2 is a C1- to C4 alkyl group, the C1- to C4 alkyl group is methyl or ethyl; and / or, when R4-2 is a halogen, the halogen is F; and / or, when R4 is a 3- to 6-membered cycloalkane, the 3- to 6-membered cycloalkane is cyclopropane or cyclobutane; and / or, when R4 is a 3- to 6-membered cycloalkane substituted with one or more R4-1, the 3- to 6-membered cycloalkane is cyclopropane or cyclobutane; And / or, when R4 is a 3- to 6-membered heterocyclic alkane substituted with one or more R4-2, the 3- to 6-membered heterocyclic alkane is a 4-membered heterocyclic alkane containing one O; and / or, when R5 is a C1- to C4 alkyl, the C1- to C4 alkyl is methyl or ethyl; and / or, when R5 is a C1- to C4 alkoxy, the C1- to C4 alkoxy is methoxy.

4. A pyrazoloquinazoline compound of formula I as described in claim 3, or a pharmaceutically acceptable salt thereof, wherein, When R3 is a 5- to 10-membered heterocyclic alkyl group substituted with one or more R3-1 groups, the 5- to 10-membered heterocyclic alkyl group is pirazinyl, hexahydropyridinyl, hexahydropyrimidinyl, 3,8-diazabicyclo[3.2.1]octyl, octahydropyrrole[1,2-a]pyrazinyl, 2,6-diazaspiro[3.4]octyl, 3,6-diazabicyclo[3.2.0]heptyl, 1,6-diazaspiro[3.4]octyl, or octahydropyrrole[3,4-c]pyrroleyl.

5. A pyrazoloquinazoline compound of formula I as described in claim 1, or a pharmaceutically acceptable salt thereof, wherein, R1 is , , , , , or; and / or, R3 is , , , , , , , or; and / or, R3 is , , , , , , , or; and / or, R3 is , , , , , , , or; and / or, when R4 is a 3- to 6-membered cycloalkane substituted with one or more R4-1, the 3- to 6-membered cycloalkane substituted with one or more R4-1 is , , or; and / or, when R4 is a 3- to 6-membered heterocycloalkane, the 3- to 6-membered heterocycloalkane is , or; and / or, when R4 is a 3- to 6-membered heterocycloalkane substituted with one or more R4-2, the 3- to 6-membered heterocycloalkane substituted with one or more R4-2 is , or.

6. A pyrazoloquinazoline compound of formula I as described in claim 5, or a pharmaceutically acceptable salt thereof, wherein, R1 can be , , or .

7. A pyrazoloquinazoline compound of formula I as described in claim 1, or a pharmaceutically acceptable salt thereof, wherein, R4 is a cyano group, a 3- to 6-membered cycloalkane substituted with one or more R4-1 groups, a 3- to 6-membered heterocycloalkane substituted with one or more R4-2 groups; and / or, R4-1 is a halogen or a hydroxyl group; and / or, R4-2 is a C1- to C4 alkyl group; and / or, R5 is H, a hydroxyl group, or a C1- to C4 alkoxy group.

8. A pyrazoloquinazoline compound of formula I as described in claim 1, or a pharmaceutically acceptable salt thereof, wherein, R4 is a cyano group, a 3- to 6-membered cycloalkane substituted with one or more R4-1 groups, a 3- to 6-membered heterocycloalkane substituted with one or more R4-2 groups; R4-1 is a halogen or a hydroxyl group; R4-2 is a C1- to C4 alkyl group; R5 is H, a hydroxyl group, or a C1- to C4 alkoxy group.

9. A pyrazoloquinazoline compound of formula I as described in claim 8, or a pharmaceutically acceptable salt thereof, wherein R4 is selected from the group consisting of cyano, , , , and.

10. A pyrazoloquinazoline compound or a pharmaceutically acceptable salt thereof as shown in Formula I as claimed in claim 1, having the following formula: .

11. A pyrazoloquinazoline compound of formula I as described in claim 10, or a pharmaceutically acceptable salt thereof, wherein R1 is a C1-C4 alkoxy group substituted with one or more R1-1.

12. A pyrazoloquinazoline compound of formula I as described in claim 10, or a pharmaceutically acceptable salt thereof, wherein R2 is H or a halogen.

13. A pyrazoloquinazoline compound of formula I as described in claim 10, or a pharmaceutically acceptable salt thereof, wherein R4 is selected from the group consisting of cyano, , , , and.

14. A pyrazoloquinazoline compound or a pharmaceutically acceptable salt thereof, wherein, The pyrazoloquinazoline compound is selected from any of the following:

15. A method for preparing a pyrazoloquinazoline compound as shown in Formula I or a pharmaceutically acceptable salt thereof, comprising the following steps: in an organic solvent, under the action of a palladium catalyst, and in a base reagent, coupling 1 g of the compound, a ligand reagent, and R3-H to obtain compound I; wherein, R1, R2, R3, R4, or R5 are defined as described in claims 1-9; X is a halogen.

16. The preparation method as described in claim 15, wherein, The palladium catalyst is tridibenzylacetone dipalladium; and / or, the organic solvent is tetradioxane; and / or, the base reagent is cesium carbonate; and / or, the ligand reagent is 4,5-bis(diphenylphosphine)-9,9-dimethyloxanthracene.

17. Use of any pyrazoloquinazoline compound or a pharmaceutically acceptable salt thereof as described in any one of claims 1 to 14 above in the preparation of a PLK1 inhibitor.

18. The use as described in claim 17, wherein the inhibitor is an inhibitor used in vitro.

19. A pharmaceutical composition, wherein, It comprises pyrazoloquinazoline compounds as described in claims 1 to 14, or pharmaceutically acceptable salts thereof, and pharmaceutical excipients.

20. Use of a pyrazoloquinazoline compound or a pharmaceutically acceptable salt thereof as described in any one of claims 1 to 14 in the preparation of a medicament for treating at least one of the following diseases: breast cancer, prostate cancer, lung cancer, colorectal cancer, liver cancer, pancreatic cancer, gastric cancer, esophageal cancer, melanoma, multiple myeloma, leukemia, and lymphoma.

21. A pharmaceutical combination comprising a pyrazoloquinazoline compound or a pharmaceutically acceptable salt thereof as described in any one of claims 1 to 14, and one or more chemotherapeutic agents, said one or more chemotherapeutic agents being used simultaneously, separately or sequentially with said compound or a pharmaceutically acceptable salt thereof.