ERK1 / 2 kinase inhibitors and their uses

Abstract

This invention relates to a class of ERK1 / 2 kinase inhibitors and their uses. The ERK1 / 2 kinase inhibitors have the structure shown in Formula I. In pharmacological studies, the compounds of this invention exhibit significant ERK1 / 2 kinase inhibitory activity, with some compounds showing comparable or even superior activity to existing MAPK pathway inhibitors. They also exhibit proliferative inhibitory activity against A375 and HCT116 cells, thus possessing the potential to be developed into drugs for treating diseases related to ERK1 / 2 kinases.

Description

Technical Field

[0001] This application belongs to the field of medicinal chemistry, specifically relating to a class of ERK1 / 2 kinase inhibitors and their uses. Background Technology

[0002] Extracellular signal-regulated kinase ERK is a key member of the mitogen-activated protein kinase (MAPK) signaling pathway, which plays a crucial role in biological processes such as cell proliferation, differentiation, survival, and migration. Aberrant activation of the ERK signaling pathway is closely associated with various cancers, inflammatory diseases, and fibrotic diseases; therefore, ERK inhibition has become an important therapeutic approach for these diseases. ERK inhibitors bind to ERK1 / 2 kinases, blocking their catalytic activity and thereby inhibiting the phosphorylation of downstream substrates such as p90RSK, thus blocking signal transduction. These drugs are commonly used to target tumors with aberrant activation of the RAS-RAF-MEK-ERK signaling pathway, such as melanoma, non-small cell lung cancer, and pancreatic cancer. Compared to RAF or MEK inhibitors, ERK inhibitors can act more directly on the terminal ends of the signaling pathway, especially in cases of RAF or MEK inhibitor resistance, where ERK inhibitors can provide a new therapeutic option. Currently, several ERK inhibitors have entered clinical trials, such as Ulixertinib, LY3214996, MK-8353, GDC-0994, CC-90003, ASN007, and ERAS-007. These drugs have shown good anti-tumor activity in preclinical and early clinical trials, especially in tumors carrying BRAF or KRAS mutations. However, the development and application of ERK inhibitors still face some significant challenges, including: 1) toxic side effects: Since the ERK signaling pathway also plays an important role in normal cells, inhibiting ERK may lead to serious side effects, including gastrointestinal toxicity, skin toxicity, hematologic toxicity, and hepatotoxicity; 2) complex drug resistance mechanisms: these involve multiple factors such as ERK mutations, reactivation of upstream signals, and compensatory activation of downstream signals; 3) limited clinical trial results: most drugs currently on the market are still in Phase I / II trials and have not yet obtained extensive clinical data support.

[0003] To address the above shortcomings, it is necessary to develop a novel ERK1 / 2 kinase inhibitor to tackle one or more of the challenges mentioned above for the treatment of related diseases. Summary of the Invention

[0004] Purpose of the invention

[0005] One objective of this invention is to provide a class of compounds having a pyrazolo[4,3-c]pyridine structure or pharmaceutically acceptable salts thereof, which have strong inhibitory activity against ERK1 / 2 kinases and suitable pharmaceutical properties.

[0006] Another technical object of the present invention is to provide a pharmaceutical composition comprising at least the above-described compounds.

[0007] Another technical object of the present invention is to provide the use of the above-described compound or pharmaceutical composition in the preparation of ERK1 / 2 kinase inhibitors. Summary of the Invention

[0008] On one hand, the present invention provides a class of compounds represented by Formula I, pharmaceutically acceptable salts thereof, enantiomers, diastereomers, tautomers, solvates, polymorphs, or prodrugs:

[0009]

[0010] In equation I above,

[0011] R 1 -R 3 R 1’ -R 3’ Each of them is independently H or C1-6 alkyl;

[0012] W 1 Selected from C1-6 alkyl groups, by R 4 Substituted C1-6 alkyl, -NR 5 R 5’ -C(=O)NR 6 R 6’ C6-10 aryl, 4-12 membered monocyclic or fused heterocyclic groups containing 1-3 of N, O or S as heteroatoms, 5-12 membered monocyclic or fused heteroaryl groups containing 1-3 of N, O or S as heteroatoms,

[0013] in,

[0014] R 4 Selected from hydroxyl and amino groups;

[0015] R 5 R 5’ R 6 R 6’ Each time it appears, it is independently selected from H, C1-6 alkyl, C3-8 cycloalkyl, 4-12 membered monocyclic or fused heterocyclic groups containing 1-3 of N, O or S as heteroatoms, or 5-12 membered monocyclic or fused heteroaryl groups containing 1-3 of N, O or S as heteroatoms.

[0016] The cycloalkyl, aryl, heterocyclic, and heteroaryl groups are optionally further substituted by one or more substituents selected from unsubstituted or halogenated C1-6 alkyl, halogen, hydroxyl, aldehyde, -S(=O)2R'-OS(=O)2R', where R' is selected from halogen, C1-6 alkyl, or C2-C10 alkenyl.

[0017] The cyclic carbon atom of the heterocyclic group may optionally be further oxidized;

[0018] L represents the bivalent connector shown below:

[0019]

[0020] Where * indicates the position where the nitrogen atom is attached to the pyridine ring of pyrazolo[4,3-c]pyridine, and ** indicates the position where W2 is attached.

[0021] X 1 and X 2 Each occurrence is independently -C(R) 7 )2- or -NR 8 -, provided that the two are not both -NR 8 -;

[0022] X is selected from -CH2- and -C(=O)-.

[0023] R 7 R 7’ Each of the following is independently selected from H, unsubstituted or substituted C1-6 alkyl groups, and amino groups;

[0024] R 8 R 9 Each of them is independently H or C1-6 alkyl;

[0025] n is 0 or 1;

[0026] W 2 For those not replaced or replaced by 1-3 Rs 10 Substituted C6-10 aryl groups, or 5-12 membered monocyclic or fused heteroaryl groups containing 1-3 N, O, or S as heteroatoms, each R 10 It is independently selected from halogens, unsubstituted or halogenated alkyl groups, and cycloalkyl groups.

[0027] In a specific implementation, R 1 -R 3 R 1’ -R 3’ Each of them is independently H or C1-6 alkyl;

[0028] W 1It is a C1-6 alkyl group, -(CH2) 1-6 -R 4 -NR 5 R 5’ -C(=O)NR 6 R 6’ , phenyl, naphthyl, biphenyl, heteroaryl selected from 1,2,3-triazolyl, 1,2,4-triazolyl, imidazolyl, pyrazolyl, pyridinyl and oxazolyl, or heterocyclic group selected from tetrahydropyrroleyl, tetrahydropyranyl and piperidinyl.

[0029] in,

[0030] R 4 Selected from hydroxyl and amino groups;

[0031] R 5 R 5’ R 6 R 6’ Each of the following is independently H in each occurrence, a C1-6 alkyl group, a C3-8 cycloalkyl group optionally substituted with deuterium, halogen, hydroxyl or amino, a heteroaryl group selected from imidazolyl, pyrazolyl and pyridinyl, or a heterocyclic group selected from piperidinyl, tetrahydropyrrolel, tetrahydrofuranyl and tetrahydropyranyl.

[0032] The cycloalkyl, phenyl, heterocyclic, and heteroaryl groups are optionally further substituted by one or more substituents selected from unsubstituted or halogenated C1-6 alkyl, halogen, hydroxyl, aldehyde, -S(=O)2R', -OS(=O)2R', where R' is selected from halogen, C1-6 alkyl, or C2-C10 alkenyl.

[0033] The cyclic carbon atom of the piperidinyl, tetrahydropyrrole, or tetrahydropyran may optionally be further oxidized;

[0034] L represents the bivalent connector shown below:

[0035]

[0036] Where * indicates the position where the nitrogen atom is attached to the pyridine ring of pyrazolo[4,3-c]pyridine, and ** indicates the position where W2 is attached.

[0037] X 1 and X 2 Each occurrence is independently assigned to -CR 7 R 7’ -or-NR 8 -, provided that the two are not both -NR 8 -;

[0038] X is selected from -CR 11 R 11’ -and-C(=O)-,

[0039] R 7 R 7’ Each of the following is independently selected from H, unsubstituted or substituted C1-6 alkyl groups, and amino groups;

[0040] R 8 R 9 Each of them is independently H or C1-6 alkyl;

[0041] R 11 R 11’ Each of these substances is independently H, halogen, or C1-6 alkyl;

[0042] n is 0 or 1;

[0043] W 2 For those not replaced or replaced by 1-3 Rs 10 Substituted phenyl or naphthyl, or heteroaryl selected from thiazolyl, indolyl, oxazolyl, imidazolyl, pyrazolyl, benzothiazolyl, and pyridyl, each R 10 Independently selected from halogens, unsubstituted or halogenated C1-6 alkyl groups, and C3-8 cycloalkyl groups.

[0044] Wherein, the C1-6 alkyl groups are selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, isobutyl, sec-butyl, n-pentyl, n-hexyl, or cycloheptaoctyl.

[0045] The above C3-8 cycloalkyl groups are selected from cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, or cycloheptyl.

[0046] In a specific implementation, in formula I,

[0047] R 1 R 2 and R 3 and R 1’ R 2’ and R 3’ All are H;

[0048] W 1 C(=O)NH-R 6 , where R 6 The definition is as described above;

[0049] L is Among them, X 1 and X 2 As defined above, R 9 It is H or C1-6 alkyl; and

[0050] W 2 4-tert-butylthiazolyl

[0051] * indicates the location where the group is attached.

[0052] In a specific implementation, in formula I,

[0053] R 1 R 2 and R 3 and R 1’ R 2’ and R 3’ All are H;

[0054] W 1 C(=O)NH-R 6 , where R 6 The definition is as described above;

[0055] L is as well as

[0056] W 2 As defined above.

[0057] In a specific embodiment, the compound represented by formula I can be represented by formula I-1 or I-2:

[0058]

[0059]

[0060] In formulas I-1 or I-2 above, the definitions of each substituent are as defined above.

[0061] In a specific implementation, the W 1 The substituent is selected from unsubstituted or substituted diazolyl, triazolyl, pyridyl, piperidinyl, tetrahydropyranyl, tetrahydropyrrolidinyl, and oxazolyl groups, wherein the substituent is selected from halogens, alkyl groups, haloalkyl groups, hydroxyl groups, and oxo groups.

[0062] In a specific implementation, the W 2 Selected from those that have not been replaced or have been replaced by 1-3 Rs 10 Substituted thiazolyl, indolyl, indazole, oxazolyl, imidazole, benzothiazolyl, naphthyl, and pyridyl.

[0063] In a specific embodiment, the compound of formula I is selected from the following:

[0064]

[0065]

[0066]

[0067]

[0068]

[0069]

[0070] On the other hand, the present invention provides a pharmaceutical composition comprising a therapeutically effective amount of the compound of Formula I described above, a pharmaceutically acceptable salt thereof, an enantiomer, a diastereomer, a tautomer, a solvate, a polymorph or a prodrug, and a pharmaceutically acceptable excipient.

[0071] In another aspect, the present invention provides the use of the above-mentioned compounds, pharmaceutically acceptable salts thereof, enantiomers, diastereomers, tautomers, solvates, polymorphs or prodrugs thereof, or the above-mentioned pharmaceutical compositions in the preparation of medicaments for treating or preventing diseases or symptoms related to ERK1 / 2 kinases.

[0072] In specific implementations, the diseases or symptoms associated with ERK1 / 2 kinases include cancer, asthma, stroke, and Alzheimer's disease.

[0073] In a specific implementation, the cancer is selected from neurofibroma, thyroid cancer, ovarian cancer, melanoma, colorectal cancer, non-small cell lung cancer, pancreatic adenocarcinoma, and lung cancer.

[0074] Beneficial effects

[0075] This invention provides a class of novel general formula compounds that, in pharmacological studies, have shown significant ERK1 / 2 kinase inhibitory activity. Some compounds exhibit activities comparable to or even superior to existing MAPK pathway inhibitors, and show proliferative inhibitory activity against A375 and HCT116 cells. Therefore, they have the potential to be developed into drugs for the treatment of diseases related to ERK1 / 2 kinase. Detailed Implementation

[0076] The present application will be described in detail below through specific embodiments to enable those skilled in the art to better understand the technical solutions of the present application. However, the provision of these embodiments is not intended to limit the scope of the present application.

[0077] the term

[0078] The term “C1-6 alkyl” as used herein refers to a straight-chain or branched saturated hydrocarbon group selected from those containing 1, 2, 3, 4, 5, or 6 carbon atoms, including methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, isobutyl, sec-butyl, n-pentyl, or n-hexyl.

[0079] The term "cycloalkyl" as used herein refers to a hydrocarbon group selected from saturated and partially unsaturated cyclic hydrocarbon groups, including monocyclic and polycyclic (e.g., bicyclic and tricyclic) groups. For example, a cycloalkyl group may contain 3 to 8 carbon atoms, more specifically, 3 to 6 carbon atoms, 3 to 5 carbon atoms, or 3 to 4 carbon atoms. Even further, for example, a cycloalkyl group may be selected from a monocyclic group containing 3 to 8 carbon atoms or 3 to 6 carbon atoms. A cycloalkyl group may be substituted with at least one heteroatom selected from, for example, O, S, and N.

[0080] The term "aryl" in this document refers to a group selected from the following:

[0081] A six-membered carbon ring aromatic ring, such as a phenyl group;

[0082] Bicyclic systems, such as 7- to 10-membered bicyclic systems, wherein at least one ring is a carbocyclic and aromatic, such as selected from naphthalene and indene.

[0083] In this article, the term "halogen" refers to F, Cl, Br, or I.

[0084] The term "heteroaryl" in this article refers to groups selected from the following:

[0085] A 5- to 7-membered aromatic monocyclic ring containing at least one heteroatom, for example, 1 to 3 heteroatoms, selected from N, O and S, with the remaining ring atoms being carbon;

[0086] An 8- to 12-membered bicyclic ring containing at least one heteroatom, for example, 1 to 3, or, in other embodiments, 1 or 2 heteroatoms, for example, selected from N, O and S, with the remaining ring atoms being carbon, wherein at least one ring is aromatic, and at least one heteroatom is present in the aromatic ring, and the connection point is located on any ring and on carbon or a heteroatom.

[0087] Examples of heteroaryl groups include, but are not limited to, pyrrole, furanyl, thiophene, oxazolyl, isoxazolyl, thiazolyl, imidazole, pyrazolyl, triazolyl, tetrazolyl, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, indolyl, inzolyl, benzofuranyl, benzothiophene, benzothiazolyl, benzoimidazolyl, quinolinyl, isoquinolinyl, quinazolinyl, quinoxalinyl, and pyrazolyl.

[0088] The term "heterocyclic group" or "heterocycle" herein refers to a ring selected from 4 to 12-membered monocyclic, bicyclic, and tricyclic, saturated and partially unsaturated rings, containing at least one carbon atom in addition to at least one heteroatom, for example 1 to 4 heteroatoms, further for example 1 to 3 heteroatoms, or further for example 1 or 2 heteroatoms, for example selected from O, S, and N. The bonding point of the heterocyclic group can be on the heteroatom or on the carbon. "Heterocyclic group" herein also refers to a 5 to 7-membered saturated or partially unsaturated carbocyclic ring containing at least one heteroatom (e.g., selected from N, O, and S), which is fused with a 5, 6, and / or 7-membered cycloalkyl, heterocyclic, or carbocyclic aromatic ring, provided that when the heterocyclic ring is fused with a carbocyclic aromatic ring, the bonding point is located on the heterocyclic ring, and when the heterocyclic ring is fused with a cycloalkyl group, the bonding point may be located on the cycloalkyl or heterocyclic ring. "Heterocyclic group" herein also refers to an aliphatic spirocyclic ring containing at least one heteroatom (e.g., selected from N, O, and S). The ring may be saturated or have at least one double bond (i.e., partially unsaturated). The heterocyclic group may be substituted, for example, with oxygen. The linkage may be carbon or a heteroatom. The heterocyclic group is not a heteroaryl group as defined herein.

[0089] Examples of heterocyclic compounds include, but are not limited to, oxazolidine, isoxazolidine, thiazolylidine, isothiazolidine, oxathionecyclopentane, isoxathionecyclopentane, dioxathionecyclopentane, piperidine, oxathionecyclopentane, thiazolyl, piperazine, morpholine, thiamorpholine, dithiazolyl, tetrahydropyran, tetrahydrofuran, tetrahydropyrrole, etc.

[0090] The compounds disclosed herein may contain asymmetric centers and therefore may exist in enantiomeric form. When the compounds disclosed herein have two or more asymmetric centers, they may also exist in diastereomeric form. Enantiomers and diastereomers belong to the broader category of stereoisomers. How to prepare optically active forms is well known in the art, for example by material resolution or by asymmetric synthesis. This is intended to include all possible stereoisomers, such as substantially pure resolved enantiomers, racemic mixtures thereof, and mixtures of diastereomers. This is intended to include all stereoisomers of the compounds disclosed herein and / or their pharmaceutically acceptable salts. Unless otherwise expressly stated, reference to one isomer applies to any possible isomer. When the isomer composition is not specified, all possible isomers are included.

[0091] When the compounds disclosed herein contain olefinic double bonds, such double bonds are intended to include E and Z geometric isomers unless otherwise stated.

[0092] The term "oxo" in this article refers to the combination of an oxygen atom with a carbon atom (including carbon atoms in a ring or chain) through a double bond, which can be abbreviated as "=O". The resulting group is called an "oxo group".

[0093] The terms “optional” or “optionally” indicate that the event or condition described below may or may not occur, and the description includes both the possibility that the event or condition occurs and the possibility that it does not occur. For example, “alkyl group optionally substituted with X” includes both “alkyl group not substituted with X” and “alkyl group substituted with X”. Those skilled in the art will understand that for any group containing one or more substituents, such groups are not intended to introduce any substitution or substitution pattern that is sterically impractical, synthetically infeasible, and / or inherently unstable.

[0094] "Pharmaceutically acceptable salts" include, but are not limited to, salts formed with inorganic acids, such as those selected from hydrochlorides, phosphates, diphosphates, hydrobroms, sulfates, sulfinates, and nitrates; and salts formed with organic acids, such as those selected from malates, maleates, fumarates, tartrates, succinates, citrates, lactates, methanesulfonates, p-toluenesulfonates, 2-hydroxyethylsulfonates, benzoates, salicylates, stearates, and alkylates such as acetates. Similarly, examples of pharmaceutically acceptable cations include, but are not limited to, sodium, potassium, calcium, aluminum, lithium, and ammonium.

[0095] Furthermore, if the compounds disclosed herein are obtained as acid addition salts, the free base can be obtained by alkalizing the solution of the acid addition salt. Conversely, if the product is a free base, the addition salt, such as a pharmaceutically acceptable addition salt, can be prepared by dissolving the free base in a suitable organic solvent and treating the solution with acid, which conforms to the conventional procedure for preparing acid addition salts from base compounds. Those skilled in the art will recognize various synthetic methods that can be used to prepare non-toxic, pharmaceutically acceptable addition salts without requiring excessive experimentation.

[0096] The "range" disclosed in this application is defined by a lower limit and an upper limit. A given range is defined by selecting a lower limit and an upper limit, which define the boundaries of a particular range. Ranges defined in this way can include or exclude endpoints and can be arbitrarily combined; that is, any lower limit can be combined with any upper limit to form a range. In this application, unless otherwise stated, the numerical range "ab" represents a shortened representation of any combination of real numbers from a to b, where a and b are real numbers. For example, the numerical range "0-5" indicates that all real numbers between "0-5" have been listed herein, and "0-5" is merely a shortened representation of these numerical combinations. Furthermore, when a parameter is described as an integer ≥ 2, it is equivalent to disclosing that the parameter is, for example, an integer 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, etc.

[0097] Example

[0098] Abbreviation list

[0099]

[0100]

[0101]

[0102] Example 1: Synthesis of compound WLE1

[0103]

[0104] Step 1:

[0105] Diisopropylaminolithium (2.0 M, 80.0 mL) was added to THF (125 mL), and the mixture was cooled to -70 °C. A THF (125 mL) solution of compound 1-1 (29.0 g, 145 mmol) was slowly added. The reaction mixture was stirred at -70 °C for 30 minutes, followed by the addition of a THF (50 mL) solution of compound 1-2 (23.4 g, 160 mmol, 21.8 mL). After the addition was complete, the temperature was gradually increased to 20 °C, and the reaction was allowed to proceed for 12 hours. After the reaction was monitored by LCMS to ensure complete reaction, the reaction mixture was quenched at 0 °C with saturated ammonium chloride aqueous solution (150 mL), stirred at 25 °C for 30 minutes, and the pH was adjusted to 5–6 with 1.0 M hydrochloric acid. The mixture was extracted with ethyl acetate (500 mL × 3), and the organic layers were combined. The mixture was washed with saturated sodium chloride aqueous solution (500 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain crude intermediate compound 1-3 (46.6 g), which was a brown oily compound. LCMS[M-56+H] + m / z:243.6.

[0106] Step Two:

[0107] Intermediate compounds 1-3 (41.6 g, 138 mmol) were dissolved in a mixed solution of acetic acid (60.0 mL) and ethanol (450 mL), and 4-methoxybenzylhydrazine hydrochloride (27.5 g, 145 mmol) was added. The reaction mixture was stirred at 90 °C for 3 hours. The reaction was monitored by LCMS until complete, and the solvent was removed by concentration under reduced pressure. The resulting mixture was the crude intermediate compound 1-4 (10.0 g), a brown oily compound. LCMS [M+H] + m / z:416.2.

[0108] Step 3:

[0109] Intermediate compounds 1-4 (20.0 g, 48.1 mmol) were dissolved in ethanol (50.0 mL), and MeNH₂ / EtOH (666 g, 6.44 mol, 100 mL, 30.0% purity) was added. The mixture was stirred at 90 °C for 12 hours in an autoclave. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was the crude intermediate compound 1-5 (20.0 g), a brown oily compound. LCMS [M+H] + m / z:400.7.

[0110] Step Four:

[0111] Intermediate compounds 1-5 (20.0 g, 49.9 mmol) were dissolved in dichloromethane (210 mL), and a solution of 1,4-dioxane in hydrochloric acid (4.0 M, 70.0 mL) was added. The mixture was stirred at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was the crude intermediate compound 1-6 (20.0 g, hydrochloride), a yellow solid. LCMS [M+H] + m / z:301.1.

[0112] Step 5: Synthesis of intermediate compounds 1-7

[0113] Under a nitrogen atmosphere, compound 1-7A (22.0 g, 152 mmol) was dissolved in methanol (200 mL), and palladium on carbon (3.00 g, 10% purity) was added. The suspension was degassed and purged once with hydrogen. The mixture was stirred at 25 °C for 4 hours under a hydrogen atmosphere. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was the crude intermediate compound 1-7 (22.0 g), a colorless oily compound. LCMS [M+H] + m / z:147.1.

[0114] Step Six:

[0115] Intermediate compounds 1-7 (8.70 g, 59.3 mmol) were dissolved in DMF (250 mL), and HATU (33.8 g, 89.0 mmol) and DIEA (38.3 g, 296 mmol, 51.7 mL) were added. The mixture was stirred at 25 °C for 1 hour. Then, compound 1-6 (20.0 g, 59.3 mmol) was added, and the mixture was stirred at 25 °C for 2 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution, extracted with ethyl acetate (50.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 1-8 (17.0 g, 39.6 mmol, yield 66.8%), a brown oily compound. LCMS [M+H]+ m / z:429.2.

[0116] Step Seven:

[0117] Intermediate compounds 1-8 (16.9 g, 39.4 mmol) were dissolved in a mixed solution of THF (180 mL), methanol (60.0 mL), and water (60.0 mL), and lithium hydroxide monohydrate (16.5 g, 394 mmol) was added. The mixture was stirred at 25 °C for 1 hour. The reaction was monitored by LCMS until complete. The reaction solution was diluted with water (10.0 mL), the pH was adjusted to 5-6 with 1.0 M hydrochloric acid, and the mixture was extracted with ethyl acetate (10.0 mL × 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was the crude intermediate compound 1-9 (15.0 g), a yellow solid. LCMS [M+H] + m / z:415.2.

[0118] Step 8:

[0119] Intermediate compounds 1-9 (7.00 g, 16.8 mmol) and 1-10 (7.90 g, 50.6 mmol) were dissolved in DMF (50.0 mL), and DIEA (10.9 g, 84.4 mmol, 14.7 mL) and HATU (9.60 g, 25.3 mmol) were added. The mixture was stirred at 25 °C for 2 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (50.0 mL), extracted with ethyl acetate (50.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (50.0 mL × 3), dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure to remove the solvent, and purified by silica gel column chromatography to give intermediate compound 1-11 (3.10 g, 5.39 mmol, yield 31.9%) as a white solid. LCMS [M+H] + m / z:553.0.

[0120] Step Nine:

[0121] TFA (15.3 g, 134 mmol, 10.0 mL) was added to intermediate compound 1-11 (2.90 g, 5.26 mmol), and the mixture was stirred at 60 °C for 12 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was slurried with methyl tert-butyl ether (10.0 mL) at 25 °C for 30 minutes to give the final product compound WLE1 (1.60 g, 3.66 mmol, yield 69.6%) as a white solid. 1H NMR (400MHz, DMSO-d6) δ12.08(s,1H),8.92(brs,1H),7.94-8.09(m,1H),6.68(d,J=4.4Hz,1H),4.67-4.74(m,1H),4.46(d,J=1 6.2Hz,1H),4.28(brs,1H),3.59-3.89(m,2H),2.88-2.99(m,3H),2.67-2.76(m,4H),1.24(s,9H),0.97-1.09(m,3H).LCMS[M+H] + m / z:433.3.

[0122] Example 2: Synthesis of compound WLE2

[0123]

[0124] Step 1:

[0125] Compound 2-1 (10.0 g, 44.8 mmol) was dissolved in DMF (100 mL), and NIS (12.1 g, 53.8 mmol) was added. After the addition was complete, the reaction was carried out at 60 °C for 4 hours under TLC monitoring. When the reaction was complete, saturated sodium sulfite aqueous solution (40.0 mL) and water (20.0 mL) were added to the reaction solution to quench the reaction. After stirring for 20 minutes, the mixture was filtered. The filter cake was slurried with methyl tert-butyl ether to give a yellow solid, namely intermediate compound 2-2 (11.0 g, 31.5 mmol, yield: 70.3%). 1 H NMR (400MHz, CDCl3) δ12.64-13.24(m,1H),4.12(s,2H),3.58(t,J=5.6Hz,2H),2.62(t,J=5.6Hz,2H),1.42(s,9H).LCMS(M+H) + m / z:350.0.

[0126] Step Two:

[0127] Intermediate compound 2-2 (10.5 g, 30.1 mmol) was dissolved in DMF (100 mL), and potassium carbonate (8.31 g, 60.1 mmol) and PMB-Cl (10.7 g, 68.3 mmol, 9.3 mL) were added. After the addition was complete, the reaction was carried out at 25 °C for 12 hours under TLC monitoring. Upon completion of the reaction, water (100 mL) was added to quench the reaction mixture. The mixture was extracted with ethyl acetate (100 mL × 2), and the ethyl acetate layer was washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove ethyl acetate. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 2-3 (10.0 g, 21.4 mmol, yield: 70.9%). LCMS (M+H) + m / z:470.0.

[0128] Step 3:

[0129] Intermediate compound 2-3 (10.0 g, 21.3 mmol) was dissolved in dichloromethane (90.0 mL), and a 1,4-dioxane solution of hydrochloric acid (4.0 M, 30.0 mL) was added. After the addition was complete, the reaction was carried out at 25 °C for 3 hours under TLC monitoring. Upon completion of the reaction, the organic solvent was removed by concentration under reduced pressure to obtain a white solid, which was the crude intermediate compound 2-4 (10.4 g). LCMS (M+H) + m / z:369.9.

[0130] Step Four:

[0131] Intermediate compound 1-7 (4.50 g, 30.8 mmol) was dissolved in DMF (70.0 mL). HATU (14.6 g, 38.5 mmol) and DIEA (16.6 g, 128 mmol, 22.4 mL) were added sequentially. After the additions were complete, the mixture was stirred at 25 °C for half an hour. Then, intermediate compound 2-4 (10.4 g, 25.6 mmol) was added to the reaction solution. The reaction was carried out at 25 °C for 8 hours. The reaction was monitored by TLC until completion. The reaction was quenched by adding water (100 mL), extracted with ethyl acetate (100 mL × 2), washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase, yielding crude intermediate compound 2-5 (14.0 g), a yellow liquid. LCMS (M+H) + m / z:498.4.

[0132] Step 5:

[0133] Intermediate compound 2-5 (14.0 g, 28.2 mmol) was dissolved in methanol (140 mL) and water (40.0 mL). Lithium hydroxide monohydrate (7.09 g, 169 mmol) was added. After the addition was complete, the reaction was carried out at 25 °C for 1 hour. The reaction was monitored by TLC until completion. After removing the organic phase by concentration under reduced pressure, water (10.0 mL) was added to the reaction flask, and the pH was adjusted to 5–6 with 1.0 M hydrochloric acid aqueous solution. After stirring for 10 minutes, the mixture was extracted with dichloromethane (150 mL × 2). The dichloromethane layer was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the dichloromethane, yielding a yellow oily substance, which was the crude intermediate compound 2-6 (13.5 g). LCMS (M+H) + m / z:484.1.

[0134] Step Six

[0135] Intermediate compound 2-6 (12.5 g, 25.9 mmol) was dissolved in DMF (130 mL). Then, HATU (14.8 g, 38.8 mmol), DIEA (10.0 g, 77.6 mmol, 13.5 mL), and compound 1-10 (4.85 g, 31.0 mmol) were added sequentially. After addition, the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until completion. The reaction was quenched by adding saturated sodium chloride aqueous solution (100 mL). The mixture was extracted with ethyl acetate (100 mL × 3), and the organic phase was washed with saturated sodium chloride aqueous solution and dried over anhydrous sodium sulfate. The mixture was filtered, concentrated under reduced pressure to remove the organic phase, and the resulting mixture was purified by silica gel column chromatography to give intermediate compound 2-7 (5.00 g, 8.05 mmol, yield: 31.1%) as a yellow oil. LCMS (M+H) + m / z:622.1.

[0136] Step Seven:

[0137] Under a nitrogen atmosphere, intermediate compound 2-7 (300 mg, 483 μmol) was dissolved in 1,4-dioxane (5.0 mL) and water (1.0 mL). Intermediate compound 2-8 (164 mg, 627 μmol), Pd(dppf)Cl2 (70.6 mg, 96.5 μmol), and sodium carbonate (102 mg, 965 μmol) were added sequentially. After the addition was complete, the reaction was carried out at 90 °C for 12 hours. The reaction was monitored by TLC until it ended. After the reaction solution was cooled to room temperature, water (20.0 mL) was added to quench the reaction. The mixture was extracted with ethyl acetate (30.0 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 2-9 (150 mg, 238 μmol, yield: 49.3%), which was a yellow oil. 1H NMR (400MHz, DMSO-d6) δ10.47(d,J=14.0Hz,1H),7.82-7.94(m,1H),7.43(d,J=18.0Hz,1H),7.25(br s,1H),7.14(dd,J=8.8,2.4Hz,2H),6.79-6.93(m,2H),6.49(s,1H),5.25(d,J=1 2.8Hz,2H),4.62-4.97(m,2H),4.02(d,J=15.6Hz,3H),3.82-3.97(m,1H),3.80( d,J=3.6Hz,3H),3.69-3.77(m,1H),3.34-3.49(m,1H),2.87-3.03(m,1H),2.57- 2.86(m,2H),2.45-2.56(m,1H),1.26-1.30(m,9H),1.11-1.26(m,3H).LCMS(M+H) + m / z:630.2.

[0138] Step 8:

[0139] Intermediate compound 2-9 (50.0 mg, 79.4 μmol) was dissolved in dichloromethane (1.0 mL), cooled to below 0 °C, and AlCl3 (15.9 mg, 119 μmol) was added. After the addition was complete, the mixture was stirred at 0 °C for 10 minutes, then heated to 40 °C and reacted for 12 hours, followed by a further heating to 70 °C and a 12-hour reaction. The reaction was monitored by TLC until it was complete. Water (3.0 mL) was added to the reaction solution to quench the reaction, and the mixture was extracted with dichloromethane (5.0 mL × 2). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was then subjected to high performance liquid chromatography to obtain the final product compound WLE2 (2 mg, 4.03 μmol, yield: 5.08%). 1 H NMR (400MHz, CD3OD) δ9.98-10.08(m,1H),7.76(d,J=8.0Hz,1H),7.13-7.38(m,2H),6.51-6.67(m,1H), 4.50-4.69(m,2H),3.79-4.11(m,2H),3.45-3.61(m,1H),3.00-3.17(m,1H),2.86-3.00(m,1H),2.78(br s,1H),2.48-2.63(m,1H),1.27(s,9H),1.08-1.25(m,3H).LCMS(M+H) + m / z:496.2.

[0140] Example 3: Synthesis of compound WLE3

[0141]

[0142] Step 1:

[0143] Under a nitrogen atmosphere, intermediate compound 2-7 (250 mg, 402 μmol) was dissolved in 1,4-dioxane (5.0 mL) and water (1.0 mL). Intermediate compound 3-1 (141 mg, 482 μmol), Pd(dppf)Cl2 (58.9 mg, 80.4 μmol), and potassium carbonate (111 mg, 804 μmol) were added sequentially. After addition, the reaction was carried out at 90 °C for 12 hours. The reaction was monitored by TLC until completion. After the reaction solution cooled to room temperature, water (20.0 mL) was added to quench the reaction. Extraction was performed with ethyl acetate (30.0 mL × 2), and the solution was dried over anhydrous sodium sulfate. The mixture was filtered, concentrated under reduced pressure to remove the organic phase, and the resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 3-2 (200 mg, 302 μmol, yield: 75.1%), a brown oily substance. LCMS (M+H) + m / z:662.1.

[0144] Step Two:

[0145] TFA (3.0 mL) was added to intermediate compound 3-2 (170 mg, 256 μmol). After the addition was complete, the mixture was reacted at 60 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, the TFA was removed by concentration under reduced pressure. The resulting mixture was then prepared by high performance liquid chromatography to obtain the final product compound WLE3 (5.00 mg, 9.23 μmol, yield: 3.59%). 1 H NMR (400MHz, CD3OD) δ7.79-8.03(m,4H),6.83-7.02(m,1H),6.58(s,1H),6.38-6.51(m,1H),6.15(dd,J=9.6,6.5Hz,1H),4.57-5. 01(m,3H),3.82-4.12(m,2H),3.43-3.62(m,1H),2.72-2.98(m,2H),2.43-2.66(m,1H),1.27(s,9H),1.08-1.25(m,3H).LCMS(M+H) + m / z:542.2.

[0146] Example 4: Synthesis of compound WLE4

[0147]

[0148] Step 1:

[0149] Under a nitrogen atmosphere, intermediate compound 2-7 (100 mg, 161 μmol) was dissolved in 1,4-dioxane (2.0 mL) and water (0.5 mL). 4-bromophenylboronic acid (50.0 mg, 249 μmol), Pd(dppf)Cl2 (17.7 mg, 24.1 μmol), and sodium carbonate (34.1 mg, 322 μmol) were added sequentially. After the addition was complete, the reaction was carried out at 90 °C for 12 hours. The reaction was monitored by TLC until it ended. After the reaction solution cooled to room temperature, water (5.0 mL) was added to quench the reaction. The mixture was extracted with ethyl acetate (5.0 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 4-1 (25.0 mg, 38.4 μmol, yield: 23.9%), which was a yellow oil. 1 HNMR(400MHz, CDCl3)δ9.41(brs,1H),7.55-7.66(m,2H),7.46-7.53(m,2H),7.08-7.17(m,2 H),6.86(dd,J=8.4,6.0Hz,2H),6.49(d,J=2.0Hz,1H),5.21(d,J=11.2Hz,2H),4.68-4.88(m ,1H),4.60(s,1H),3.79(d,J=4.4Hz,4H),3.64-3.72(m,1H),3.10(ddd,J=10.0,6.8,3.6Hz, 1H),2.89-3.02(m,1H),2.56-2.75(m,2H),2.38-2.54(m,1H),1.27-1.32(m,12H).LCMS(M+H) + m / z:652.2.

[0150] Step Two:

[0151] Under a nitrogen atmosphere, compound 4-1 (25 mg, 38.4 μmol) was dissolved in IPA (1.0 mL), and then 1,4-diazabicyclo[2.2.2]octane-1,4-dionyl-1,4-disulfinic acid (10.2 mg, 42.3 μmol), PdCl2(AmPhos)2 (5.44 mg, 7.68 μmol), and triethylamine (11.7 mg, 115 μmol) were added sequentially. After the addition was complete, the solution was heated to 75 mL. After reacting at ℃ for 24 hours, and allowing the reaction solution to cool to room temperature, N-fluorobisbenzenesulfonamide (18.2 mg, 57.6 μmol) was added to the reaction solution. The reaction was continued at 25℃ for 3 hours, monitored by TLC. Upon completion of the reaction, ethyl acetate was added to the reaction solution, and the mixture was filtered. The filtrate was concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 4-2 (15.0 mg, 22.9 μmol, yield: 59.7%). LCMS (M+H) + m / z:654.1.

[0152] Step 3:

[0153] Compound 4-2 (15 mg, 22.9 μmol) was dissolved in dichloromethane (1.0 mL), and TFA (0.5 mL) was added. After the addition was complete, the reaction was carried out at 70 °C for 24 hours. The reaction was monitored by TLC. When the reaction was complete, water (3.0 mL) was added to the reaction solution to quench the reaction. The mixture was extracted with dichloromethane (5.0 mL × 2), and the organic phase was dried with anhydrous sodium sulfate. The mixture was filtered and concentrated under reduced pressure to remove the organic phase. The resulting mixture was prepared by high performance liquid chromatography to obtain the final product compound WLE4 (2 mg, 3.99 μmol, yield: 17.4%). 1 H NMR (400MHz, CD3OD) δ8.08-8.19(m,2H),7.92-8.05(m,2H),6.87(s,1H),4.75-4.85(m,2H),3.86-3.99(m,2H),3.20(ddd,J=13. 6,6.8,3.6Hz,1H),3.03-3.12(m,1H),2.96(d,J=3.6Hz,1H),2.70-2.88(m,2H),1.34-1.36(m,9H),1.27-1.32(m,3H).LCMS(M+H) + m / z:534.0.

[0154] Example 5: Synthesis of compound WLE5

[0155]

[0156] Step 1:

[0157] Under a nitrogen atmosphere, compound 2-7 (300 mg, 482 μmol) was dissolved in 1,4-dioxane (5.0 mL) and water (1.0 mL). Then, 4-benzyloxyphenylboronic acid (165 mg, 724 μmol), Pd(dppf)Cl2 (71 mg, 96.5 μmol), and sodium carbonate (102 mg, 965 μmol) were added sequentially. After the additions were complete, the reaction was carried out at 90 °C for 12 hours. The reaction was monitored by TLC until completion. After the reaction solution cooled to room temperature, water (10.0 mL) was added to quench the reaction. The mixture was extracted with ethyl acetate (5.0 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 5-1 (320 mg, 472 μmol, yield 97.8%), a yellow oil. LCMS (M+H) + m / z:678.1.

[0158] Step Two:

[0159] Under a hydrogen atmosphere, intermediate compound 5-1 was dissolved in methanol (10.0 mL), and Pd / C (108 mg, 101 μmol) was added. The reaction was carried out at 40 psi and 30 °C for 12 hours. The reaction was monitored by TLC until completion. The mixture was filtered, and the filtrate was concentrated under reduced pressure to remove the organic phase. The resulting mixture was then subjected to high-performance liquid chromatography (HPLC) to prepare intermediate compound 5-2 (100 mg, 170 μmol, yield: 17.7%) as a white solid. LCMS (M+H) + m / z:588.1.

[0160] Step 3:

[0161] Intermediate compound 5-2 (75 mg, 127 μmol) was dissolved in THF (3.0 mL), and 4-(acetamido)phenyl]iminodithiodifluoride (48.1 mg, 153 μmol) and DBU (43 mg, 280 μmol) were added sequentially. The reaction was carried out at 25 °C for 10 min, monitored by TLC. After the reaction was completed, water (10.0 mL) was added to quench the reaction mixture, and the mixture was extracted with ethyl acetate (10.0 mL). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was then subjected to high performance liquid chromatography (HPLC) to prepare intermediate compound 5-3 (13.0 mg, 19.4 μmol, yield: 15.2%). LCMS (M+H) + m / z:670.0.

[0162] Step Four:

[0163] Intermediate compound 5-3 (13 mg, 19.4 μmol) was dissolved in dichloromethane (1.0 mL), and TFA (3.0 mL) was added. After the addition was complete, the reaction was carried out at 70 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, water (3.0 mL) was added to the reaction solution to quench the reaction. The mixture was extracted with dichloromethane (5.0 mL × 2), and the organic phase was dried with anhydrous sodium sulfate. The mixture was filtered and concentrated under reduced pressure to remove the organic phase. The resulting mixture was prepared by high performance liquid chromatography to obtain the final product compound WLE5 (10 mg, 18.2 μmol, yield: 93.7%). 1 H NMR (400MHz, CD3OD) δ7.74-7.87(m,2H),7.55(t,J=8.4Hz,2H),6.61-6.66(m,1H),4.90-5.00(m,1H),4.67(d,J=15.6Hz,1 H),3.88-4.12(m,2H),3.46-3.60(m,1H),2.72-3.15(m,3H),2.51-2.65(m,1H),1.28(s,9H),1.10-1.26(m,3H).LCMS(M+H) + m / z:550.0.

[0164] Example 6: Synthesis of compound WLE6

[0165]

[0166] Step 1:

[0167] Under a nitrogen atmosphere, compound 2-7 (600 mg, 965 μmol) was dissolved in THF (5.0 mL). Trimethylsilylacetylene (113 mg, 1.16 mmol), TEA (293 mg, 2.90 mmol), Pd(PPh3)2Cl2 (68 mg, 96.5 μmol), and CuI (18 mg, 96.5 μmol) were added sequentially. After addition, the reaction was carried out at 25 °C for 8 hours. The reaction was monitored by TLC until completion. Water (10.0 mL) was added to quench the reaction. The mixture was extracted with ethyl acetate (5.0 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 6-1 (450 mg, 760 μmol, yield 78.7%) as a yellow oil. LCMS (M+H) + m / z:592.1.

[0168] Step Two:

[0169] Under a nitrogen atmosphere, intermediate compound 6-1 was dissolved in DMF (9.0 mL) and methanol (1.0 mL). TMSN3 (78 mg, 675 μmol) and CuI (3 mg, 16.9 μmol) were added sequentially. The reaction was carried out at 100 °C for 12 hours. The reaction was monitored by TLC until completion. After cooling to room temperature, potassium fluoride (25 mg) was added, and the mixture was stirred at room temperature for 24 hours. The reaction was quenched by adding saturated sodium chloride aqueous solution (20.0 mL). The mixture was extracted with ethyl acetate (3 × 10.0 mL) and dried over anhydrous sodium sulfate. The mixture was filtered, concentrated under reduced pressure to remove the organic phase, and the resulting mixture was purified by silica gel column chromatography to give intermediate compound 6-2 (20 mg, 35.5 μmol, yield: 10.5%) as a white solid. LCMS (M+H) + m / z:563.2.

[0170] Step 3:

[0171] The intermediate compound 6-2 (10 mg, 17.7 μmol) was dissolved in TFA (3.0 mL) and reacted at 70 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, water (3.0 mL) was added to the reaction solution to quench the reaction. The mixture was extracted with dichloromethane (5.0 mL × 2), and the organic phase was dried with anhydrous sodium sulfate. The mixture was filtered and concentrated under reduced pressure to remove the organic phase. The resulting mixture was then subjected to high performance liquid chromatography to prepare the final product compound WLE6 (2 mg, 4.52 μmol, yield: 25.5%). 1 H NMR (400MHz, CD3OD) δ7.99-8.14(m,1H),6.59(d,J=1.6Hz,1H),3.97-4.09(m,1H),3.80-3.97(m,1H),3.48-3.61(m,2H),3.01-3.15(m,1H ),2.93(ddd,J=18.8,16.0,9.2Hz,2H),2.78(brs,1H),2.57(dt,J=15.6,5.6Hz,1H),1.27(d,J=3.2Hz,9H),1.14-1.25(m,3H).LCMS(M+H) + m / z:443.1.

[0172] Example 7: Synthesis of compound WLE7

[0173]

[0174] Step 1:

[0175] Compound 1-4 (5.00 g, 12.0 mmol) was dissolved in methanol (10.0 mL), THF (30.0 mL), and water (10.0 mL). Lithium hydroxide monohydrate (5.05 g, 120 mmol) was added. After the addition was complete, the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until completion. After removing the organic phase by concentration under reduced pressure, water (10.0 mL) was added to the reaction flask, and the pH was adjusted to 2–3 with 1.0 M hydrochloric acid aqueous solution. After stirring for 10 minutes, the mixture was filtered to obtain the filter cake, which was the crude intermediate compound 7-1 (4.66 g), a yellow solid. LCMS (M+H) + m / z:388.1.

[0176] Step Two:

[0177] Intermediate compound 7-1 (4.66 g, 12.0 mmol) was dissolved in DMF (25.0 mL), and HATU (6.86 g, 18.0 mmol), DIEA (10.9 g, 84.2 mmol, 14.7 mL), and NH4Cl (3.22 g, 60.1 mmol) were added sequentially. After the addition was complete, the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until it ended. The reaction was quenched by adding saturated sodium chloride aqueous solution (50.0 mL), extracted with ethyl acetate (30.0 mL × 3), washed with saturated ammonium chloride aqueous solution, and dried over anhydrous sodium sulfate. The mixture was filtered, concentrated under reduced pressure to remove the organic phase, and the resulting mixture was purified by silica gel column chromatography to give intermediate compound 7-2 (4.50 g, 11.6 mmol, yield: 96.8%) as a yellow solid. LCMS (M+H) + m / z:387.0.

[0178] Step 3:

[0179] Intermediate compound 7-2 (4.50 g, 11.6 mmol) was dissolved in DMF-DMA (145 g, 1.21 mol, 161 mL). After addition, the reaction was carried out at 90 °C for 2 hours. The organic phase was then removed by concentration under reduced pressure. At 25 °C, acetic acid (169 g, 2.82 mol, 161 mL) and hydrazine hydrate (7.29 g, 116 mmol, 7.06 mL) were added sequentially to the reaction flask. After addition, the reaction was carried out at 80 °C for 2 hours. The reaction was monitored by TLC. After the reaction was completed, the acetic acid was removed by concentration under reduced pressure. The pH was adjusted to neutral by adding saturated sodium bicarbonate aqueous solution to the reaction flask. The mixture was extracted with dichloromethane (20.0 mL), and the organic phase was washed with saturated sodium chloride aqueous solution (20.0 mL). The organic phase was dried over anhydrous sodium sulfate, filtered, and the organic phase was removed by concentration under reduced pressure to obtain a yellow solid, which was the crude intermediate compound 7-3 (4.50 g). LCMS (M+H) + m / z:411.1.

[0180] Step Four:

[0181] Intermediate compound 7-3 (2.50 g, 6.09 mmol) was dissolved in 1,4-dioxane (5.0 mL), and a 1,4-dioxane solution of hydrochloric acid (4.0 M, 20.0 mL) was added. After the addition was complete, the reaction was carried out at 25 °C for 2 hours under TLC monitoring. Upon completion of the reaction, the organic solvent was removed by concentration under reduced pressure to obtain a white solid, which was the crude intermediate compound 7-4 (1.89 g). LCMS (M+H) + m / z:310.6.

[0182] Step 5:

[0183] Intermediate compound 7-4 (1.89 g, 6.09 mmol) was dissolved in DMF (10.0 mL). Then, HATU (2.78 g, 7.31 mmol), DIEA (3.94 g, 30.5 mmol, 5.3 mL), and compound 1-7 (979 mg, 6.70 mmol) were added sequentially. After addition, the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until completion. The reaction was quenched by adding saturated sodium chloride aqueous solution (30.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phase was washed with saturated ammonium chloride aqueous solution and dried over anhydrous sodium sulfate. The mixture was filtered, concentrated under reduced pressure to remove the organic phase, and the resulting mixture was purified by silica gel column chromatography to give intermediate compound 7-5 (2.60 g, 5.93 mmol, yield: 97.4%) as a yellow solid. LCMS (M+H) + m / z:439.2.

[0184] Step Six:

[0185] Intermediate compound 7-5 (2.60 g, 5.93 mmol) was dissolved in methanol (5.0 mL), THF (15.0 mL), and water (5.0 mL). Lithium hydroxide monohydrate (2.49 g, 59.3 mmol) was added. After the addition was complete, the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until completion. After removing the organic phase by concentration under reduced pressure, water (10.0 mL) was added to the reaction flask, and the pH was adjusted to 5–6 with 1.0 M hydrochloric acid aqueous solution. After stirring for 10 minutes, the mixture was filtered to obtain the filter cake, which was intermediate compound 7-6 (1.25 g), as a white solid. LCMS (M+H) + m / z:425.0.

[0186] Step Seven:

[0187] Intermediate compound 7-6 (1.00 g, 2.36 mmol) was dissolved in DMF (10.0 mL). Then, HATU (1.07 g, 2.83 mmol), DIEA (1.52 g, 11.8 mmol, 2.1 mL), and compound 1-10 (1.10 g, 7.07 mmol) were added sequentially. After addition, the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until completion. The reaction was quenched by adding saturated sodium chloride aqueous solution (20.0 mL). The mixture was extracted with ethyl acetate (10.0 mL × 3), and the organic phase was washed with saturated ammonium chloride aqueous solution and dried over anhydrous sodium sulfate. The mixture was filtered, concentrated under reduced pressure to remove the organic phase, and the resulting mixture was purified by silica gel column chromatography to give intermediate compound 7-7 (1.00 g, 1.78 mmol, yield: 75.4%) as a yellow solid. LCMS (M+H) + m / z:563.1.

[0188] Step 8:

[0189] Intermediate compound 7-7 (1.00 g, 1.78 mmol) was dissolved in TFA (15.4 g, 135 mmol, 10.0 mL) and reacted at 70 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, water (3.0 mL) was added to the reaction solution to quench the reaction. The mixture was extracted with dichloromethane (10.0 mL × 2), and the organic phase was dried over anhydrous sodium sulfate. The mixture was filtered and concentrated under reduced pressure to remove the organic phase. The resulting mixture was then subjected to high performance liquid chromatography to prepare the final product compound WLE7 (430 mg, 971 μmol, yield: 54.7%). 1 HNMR(400MHz,CD3OD)δ9.53-9.57(m,1H),6.81-6.83(m,1H),4.13(dt,J=14.0,5.0Hz,1H),3.84-4.00(m,2H),3.52-3.6 0(m,1H),2.91-3.18(m,3H),2.83(t,J=5.6Hz,1H),2.67-2.74(m,1H),1.33-1.34(m,9H),1.19-1.28(m,3H).LCMS(M+H) + m / z:443.2.

[0190] Example 8: Synthesis of compound WLE8

[0191]

[0192] Step 1:

[0193] Ethyl pivaloyl ethyl acetate (1.17 g, 6.82 mmol, 1.2 mL) was dissolved in DMF (5.0 mL). Potassium carbonate (628 mg, 4.55 mmol) was added at 0 °C. After the addition was complete, the mixture was stirred at 0 °C for 30 minutes. Then, 4-bromo-1-fluoro-2-nitrobenzene (1.00 g, 4.55 mmol) was added dropwise to the reaction solution. After the addition was complete, the mixture was reacted at 25 °C for 12 hours. The reaction was monitored by TLC until it ended. Water (20.0 mL) was added to the reaction solution to quench the reaction. The mixture was filtered, and the filter cake was purified by silica gel column chromatography to obtain intermediate compound 8-1 (1.50 g, 4.03 mmol, yield: 88.7%), which was a yellow oil. 1 H NMR (400MHz, DMSO-d6) δ8.28(d,J=2.0Hz,1H),7.98(dd,J=8.4,2.0Hz,1H),7.37(d,J=8. 4Hz,1H),5.78(s,1H),4.15(q,J=7.2Hz,2H),1.15-1.18(m,3H),1.10(s,9H).LCMS(M+H) + m / z:373.8.

[0194] Step Two:

[0195] At 0 °C, intermediate compound 8-1 (1.10 g, 2.96 mmol) was dissolved in concentrated hydrochloric acid (12.0 M, 25.4 mL). After the addition was complete, the reaction was carried out at 100 °C for 2 hours under TLC monitoring. After the reaction was completed, the hydrochloric acid was removed by concentration under reduced pressure. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 8-2 (650 mg, 2.17 mmol, yield: 73.3%) as a white solid. 1 H NMR (400MHz, DMSO-d6) δ8.23(d,J=2.0Hz,1H),7.91(dd,J=8.4,2.0Hz,1H),7.43(d,J=8.4Hz,1H),4.37(s,2H),1.17(s,9H).

[0196] Step 3:

[0197] Intermediate compound 8-2 (550 mg, 1.83 mmol) was dissolved in acetic acid (11.0 mL), and iron powder (512 mg, 9.16 mmol) was added. After the addition was complete, the reaction was carried out at 115 °C for 3 hours. The reaction was monitored by TLC until it ended. The mixture was filtered, and the filtrate was concentrated under reduced pressure to remove organic matter. Water (20.0 mL) was added to the reaction flask, and the mixture was extracted with ethyl acetate (20.0 mL). The extract was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 8-3 (370 mg, 1.47 mmol, yield: 80.1%) as a yellow solid. 1 H NMR (400MHz, DMSO-d6) δ11.04(brs,1H),7.43(s,1H),7.36(d,J=8.4Hz,1H),7.04(dd,J=8.4,1.6Hz,1H),6.13(d,J=1.6Hz,1H),1.33(s,9H).

[0198] Step Four:

[0199] Under a nitrogen atmosphere, intermediate compound 8-3 (320 mg, 1.27 mmol) was dissolved in toluene (5.0 mL). TEA (193 mg, 1.90 mmol), chloro(crotonyl)(tri-tert-butylphosphine)palladium(II) (51.9 mg, 102 μmol) and ethyl methacrylate (724 mg, 6.35 mmol) were added sequentially. After the addition was complete, the mixture was reacted at 90 °C for 3 hours under TLC monitoring. After the reaction was completed, the toluene was removed by concentration under reduced pressure. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 8-4 (200 mg, 701 μmol, yield: 55.2%). 1 H NMR (400MHz, DMSO-d6) δ11.09(s,1H),7.73(s,1H),7.43-7.49(m,2H),7.08(dd,J=8.4,0.8Hz,1H),6.16(d ,J=1.6Hz,1H),4.20(q,J=7.2Hz,2H),2.15(d,J=0.8Hz,3H),1.35(s,9H),1.28(t,J=7.2Hz,3H).LCMS(M+H) + m / z:286.0.

[0200] Step 5:

[0201] Under a hydrogen atmosphere, intermediate compound 8-4 (200 mg, 703 μmol) was dissolved in methanol (2.0 mL). Pd / C (10%, 0.20 g) was added, and the reaction was carried out at 25 °C for 12 hours under TLC monitoring. After the reaction was complete, Pd / C was removed by filtration, and the filtrate was concentrated under reduced pressure to remove methanol, yielding crude intermediate compound 8-5 (160 mg), a yellow oily substance. LCMS (M+H) + m / z:288.0.

[0202] Step Six:

[0203] Intermediate compound 8-5 (160 mg, 557 μmol) was dissolved in methanol (1.0 mL), THF (3.0 mL), and water (1.0 mL). Lithium hydroxide monohydrate (234 mg, 5.57 mmol) was added. After the addition was complete, the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until completion. After removing the organic phase by concentration under reduced pressure, water (10.0 mL) was added to the reaction flask, and the pH was adjusted to 2–3 with 1.0 M hydrochloric acid aqueous solution. After stirring for 10 minutes, the mixture was extracted with dichloromethane (10.0 mL). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove dichloromethane, yielding a yellow oily substance, which was the crude intermediate compound 8-6 (140 mg). LCMS (M+H) + m / z:259.9.

[0204] Step Seven:

[0205] Intermediate compound 8-6 (140 mg, 540 μmol) was dissolved in DMF (3.0 mL). HATU (308 mg, 810 μmol) and DIEA (349 mg, 2.70 mmol) were added sequentially. After the addition was complete, the mixture was stirred at room temperature for 1 hour. Then, a DMF solution of compound 1-6 (162 mg, 540 μmol) (2.0 mL) was added dropwise. After the addition was complete, the reaction was carried out at 25 °C for 11 hours. The reaction was monitored by TLC until completion. The reaction was quenched by adding saturated sodium chloride aqueous solution (20.0 mL). The mixture was extracted with dichloromethane (5.0 mL × 3), and the organic phase was washed with saturated ammonium chloride aqueous solution and dried over anhydrous sodium sulfate. The mixture was filtered, concentrated under reduced pressure to remove the organic phase, and the resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 8-7 (40 mg, 73.8 μmol, yield: 13.7%) as a yellow solid. LCMS (M+H) + m / z:542.3.

[0206] Step 8:

[0207] Intermediate compound 8-7 (40 mg, 73.8 μmol) was dissolved in TFA (1.54 g, 13.5 mmol, 1.0 mL), and reacted at 70 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, the TFA was removed by concentration under reduced pressure. The resulting mixture was then prepared by high performance liquid chromatography to obtain the final product compound WLE8 (10 mg, 23.7 μmol, yield: 32.1%). 1 H NMR(400MHz, DMSO-d6)δ10.07(d,J=2.8Hz,1H),7.13-7.30(m,1H),7.02(brs,1H),6.72-6.82(m,1H),5.95-6.04(m,1H),4.41-4.61(m,4H) ),3.44-4.05(m,2H),3.21-3.29(m,1H),2.88(d,J=10.0Hz,3H),2.57-2.78(m,2H),1.35(d,J=12.4Hz,9H),1.14-1.23(m,3H).LCMS(M+H) + m / z:422.2.

[0208] Example 9: Synthesis of compound WLE9

[0209]

[0210] Step 1:

[0211] Boc-glycine (3.00 g, 17.1 mmol) was dissolved in dichloromethane (50.0 mL). HATU (9.77 g, 25.7 mmol), DIEA (11.1 g, 85.6 mmol, 14.9 mL), and compound 1-10 (2.68 g, 17.1 mmol) were added sequentially. After addition, the reaction was carried out at 25 °C for 11 hours. The reaction was monitored by TLC until completion. The reaction was quenched by adding saturated sodium chloride aqueous solution (50.0 mL), extracted with dichloromethane (30.0 mL × 3), and the organic phase was washed with saturated ammonium chloride aqueous solution and dried over anhydrous sodium sulfate. The mixture was filtered, concentrated under reduced pressure to remove the organic phase, and the resulting mixture was purified by silica gel column chromatography to give intermediate compound 9-1 (1.26 g, 4.02 mmol, yield: 23.5%) as a yellow solid. LCMS (M+H) + m / z:314.0.

[0212] Step Two:

[0213] Intermediate compound 9-1 (1.23 g, 3.92 mmol) was dissolved in dichloromethane (10.0 mL), and a 1,4-dioxane solution of hydrochloric acid (4.0 M, 10.0 mL) was added. After the addition was complete, the reaction was carried out at 25 °C for 12 hours under TLC monitoring. Upon completion of the reaction, the organic solvent was removed by concentration under reduced pressure to obtain a white solid, which was the crude intermediate compound 9-2 (800 mg). LCMS (M+H) + m / z:213.9.

[0214] Step 3:

[0215] Intermediate compound 9-2 (422 mg, 1.41 mmol) and TEA (285 mg, 2.81 mmol) were dissolved in dichloromethane (5.0 mL). The solution was cooled to below 0 °C, and triphosgene (138 mg, 464 μmol) was added dropwise. After the addition was complete, the mixture was stirred at 0 °C for 2 hours. Then, a dichloromethane solution (5.0 mL) of compound 1-6 (300 mg, 1.41 mmol) was slowly added dropwise to the reaction mixture. After the addition was complete, the mixture was reacted at 25 °C for 12 hours. The reaction was monitored by TLC until it ended. The reaction was quenched by adding ammonia (20.0 mL), and the mixture was extracted with dichloromethane (10.0 mL × 3). The extract was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 9-3 (450 mg, 834 μmol, yield: 59.3%) as a yellow solid. LCMS (M+H) + m / z:540.0.

[0216] Step Four:

[0217] Intermediate compound 9-3 (550 mg, 1.02 mmol) was dissolved in TFA (7.68 g, 67.3 mmol, 5.0 mL) and reacted at 60 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, DIEA was added to the reaction solution to adjust the pH to neutral. The organic phase was removed by concentration under reduced pressure. The resulting mixture was then treated by high performance liquid chromatography to obtain the final product compound WLE9 (140 mg, 333 μmol, yield: 32.9%). 1 H NMR (400MHz, DMSO-d6) δ12.93(brs,1H),11.99(s,1H),7.98(brs,1H),7.05(t,J=5.6Hz,1H),6.71(s,1H),4.51(s,2H ),3.87(d,J=5.6Hz,2H),3.60(t,J=5.6Hz,2H),2.72(d,J=4.8Hz,3H),2.67(t,J=5.2Hz,2H),1.25(s,9H).LCMS(M+H) + m / z:420.1.

[0218] Example 10: Synthesis of compound WLE10

[0219]

[0220] Step 1:

[0221] Boc-N-methylglycine (2.00 g, 10.6 mmol) was dissolved in dichloromethane (20.0 mL), and HATU (6.03 g, 15.9 mmol), DIEA (6.83 g, 52.9 mmol), and compound 1-10 (1.65 g, 10.6 mmol) were added sequentially. After the addition was complete, the reaction was carried out at 25 °C for 2 hours. The reaction was monitored by TLC until it ended. Water (50.0 mL) was added to the reaction solution to quench the reaction, and the mixture was extracted with dichloromethane (20.0 mL × 3). The extract was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 10-1 (3.20 g, 9.77 mmol, yield: 92.5%) as a white solid. HNMR (400MHz, CDCl3) δ8.95-9.48(m,1H),6.55(s,1H),4.09(brs,2H),3.01(s,3H),1.38-1.57(m,9H),1.29(s,9H).

[0222] Step Two:

[0223] Intermediate compound 10⁻¹ (2.60 g, 7.94 mmol) was dissolved in dichloromethane (24.0 mL), and a 1,4-dioxane solution of hydrochloric acid (4.0 M, 8.5 mL) was added. After the addition was complete, the reaction was carried out at 25 °C for 2 hours under TLC monitoring. Upon completion of the reaction, the organic solvent was removed by concentration under reduced pressure to obtain a white solid, which was crude intermediate compound 10⁻² (1.50 g). LCMS (M+H) + m / z:228.0.

[0224] Step 3:

[0225] Intermediate compound 10-2 (200 mg, 666 μmol) and TEA (135 mg, 1.33 mmol) were dissolved in dichloromethane (5.0 mL). The mixture was cooled to below 0 °C, and a solution of triphosgene (65 mg, 220 μmol) in dichloromethane (2.0 mL) was added dropwise. After the addition was complete, the mixture was stirred at 0 °C for 3 hours. Then, a solution of compound 1-6 (182 mg, 799 μmol) in dichloromethane (5.0 mL) was slowly added dropwise to the reaction mixture. After the addition was complete, the mixture was reacted at 25 °C for 12 hours. The reaction was monitored by TLC until it ended. The reaction was quenched by adding ammonia (20.0 mL), and the mixture was extracted with dichloromethane (10.0 mL × 3). The extract was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 10-3 (120 mg, 217 μmol, yield: 32.5%) as a yellow solid. 1 H NMR (400MHz, DMSO-d6) δ12.09(s,1H),7.96(q,J=4.4Hz,1H),7.15(d,J=8.4Hz,2H),6.90(d,J=8.4Hz,2H),6.73(s,1H),5.21(s,2H),4.34(s ,2H),3.99(s,2H),3.72(s,3H),3.38(t,J=5.2Hz,2H),2.91(s,3H),2.71(d,J=4.4Hz,3H),2.65-2.70(m,2H),1.22-1.28(m,9H).LCMS(M+H) + m / z:554.2.

[0226] Step Four:

[0227] The intermediate compound 10-3 (120 mg, 217 μmol) was dissolved in TFA (2.0 mL) and reacted at 60 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, the organic phase was removed by vacuum concentration. The resulting mixture was then prepared by high performance liquid chromatography to obtain the final product compound WLE10 (39.6 mg, 91 μmol, yield: 42.4%). 1 H NMR (400MHz, CD3OD) δ6.60-6.69(m,1H),4.57(s,2H),4.07(s,2H),3.58(J=5. 2Hz,2H),3.07(s,3H),2.88(s,3H),2.79-2.86(m,2H),1.29(s,9H).LCMS(M+H) + m / z:434.2.

[0228] Example 11: Synthesis of compound WLE11

[0229]

[0230] Step 1:

[0231] Under nitrogen protection, compound 11-1 (5.00 g, 20.9 mmol) and imidazole (2.85 g, 41.8 mmol) were dissolved in DMF (50.0 mL), cooled to below 0 °C, and TBSCl (6.30 g, 41.8 mmol) was added. After the addition was complete, the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until it ended. Water (300 mL) was added to the reaction solution, and the mixture was extracted with ethyl acetate (100 mL × 2). The organic phase was washed with saturated sodium chloride aqueous solution, filtered, and concentrated under reduced pressure to remove the organic solvent. The mixture was purified by silica gel column chromatography to obtain intermediate compound 11-2 (4.45 g, 12.6 mmol, yield: 60.2%). 1 H NMR (400MHz, DMSO-d6) δ13.55-11.63(m,1H),7.44-7.18(m,6H),5.04(d,J=2.4Hz,2H),4.20-4.08(m,1H),3.82(d,J=5.2Hz,2H),0.84(s,9H),0.06 -0.05(m,6H).LCMS(M+H) + m / z:354.0.

[0232] Step Two:

[0233] Intermediate compound 11-2 (2.00 g, 5.66 mmol) was dissolved in DMF (15.0 mL), followed by the sequential addition of HATU (3.23 g, 8.49 mmol) and DIEA (2.19 g, 17.0 mmol). After the addition was complete, the mixture was stirred at 25 °C for half an hour. Then, compound 1-10 (1.06 g, 6.79 mmol) was added to the reaction solution, and the reaction was carried out at 25 °C for 3 hours. The reaction was monitored by TLC until completion. The reaction was quenched by adding water (50.0 mL), extracted with ethyl acetate (50.0 mL), washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 11-3 (3.50 g, 8.17 mmol, yield: 91.7%) as a yellow solid. LCMS (M+H) + m / z:492.1.

[0234] Step 3:

[0235] Under a hydrogen atmosphere, intermediate compound 11-3 (930 mg, 1.89 mmol) was dissolved in methanol (20.0 mL), and Pd / C (2.01 g, 1.89 mmol) was added. After the addition was complete, the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until completion. Pd / C was removed by filtration, and the filtrate was concentrated under reduced pressure to remove the organic solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 11-4 (304 mg, 851 μmol, yield: 44.9%). LCMS (M+H) + m / z:358.0.

[0236] Step Four:

[0237] Compound 1-6 (141 mg, 419 μmol) was dissolved in dichloromethane (5.0 mL) and cooled to below 0 °C. TEA (170 mg, 1.68 mmol, 234 μL) and triphosgene (42.3 mg, 143 μmol) were added sequentially. After addition, the mixture was stirred at 0 °C for 3 hours. Then, intermediate compound 11-4 (150 mg, 419 μmol) was added to the reaction solution, and the reaction was carried out at 25 °C for 40 hours. The reaction was monitored by TLC until completion. Water (50.0 mL) was added to quench the reaction, and the mixture was extracted with dichloromethane (50.0 mL × 2). The organic phase was washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 11-5 (66 mg, 96.5 μmol, yield: 23.0%) as a white solid. LCMS (M+H) + m / z:684.1.

[0238] Step 5:

[0239] Under a nitrogen atmosphere, intermediate compound 11-5 (56 mg, 82 μmol) was dissolved in TFA (18.0 mL) and TfOH (0.3 mL), and reacted at 60 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, the organic phase was removed by vacuum concentration. The resulting mixture was then prepared by high performance liquid chromatography to obtain the final product compound WLE11 (28 mg, 61.4 μmol, yield: 75.0%). 1 HNMR (400MHz, CD3OD) δ6.65 (s, 1H), 4.70 (s, 2H), 4.54 (t, J = 5.2Hz, 1H), 3.99-3.88 ( m,2H),3.84-3.67(m,2H),2.89(s,3H),2.79(t,J=5.2Hz,2H),1.28(s,9H)LCMS(M+H) + m / z:450.2.

[0240] Example 12: Synthesis of compound WLE12

[0241]

[0242] Step 1:

[0243] Compound 1-9 (620 mg, 1.50 mmol) was dissolved in dichloromethane (5.0 mL), and oxaloyl chloride (227 mg, 1.80 mmol, 157 μL) and DMF (10.9 mg, 149 μmol) were added sequentially. After the addition was complete, the mixture was stirred at 25 °C for 1 hour, and then concentrated under reduced pressure to remove organic matter. Dichloromethane (5.00 mL) was added to the reaction flask, followed by DMAP (913 mg, 7.48 mmol) and compound 12-1 (754 mg, 4.50 μmol). A solution of 0.49 mmol / L dichloromethane (5.0 mL) was added, and the mixture was stirred at 25 °C for 1 hour. The reaction was monitored by TLC until completion. Water (20.0 mL) was added to quench the reaction mixture, followed by extraction with 20.0 mL dichloromethane. The organic phase was washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 12-2 (180 mg, 318 μmol, yield: 21.3%) as a yellow solid. LCMS (M+H) + m / z:565.0.

[0244] Step Two:

[0245] Intermediate compound 12-2 (134 mg, 237 μmol) was dissolved in TFA (12.3 g, 108 mmol, 8.0 mL), and reacted at 60 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, the organic phase was removed by vacuum concentration. The resulting mixture was then prepared by high performance liquid chromatography to obtain the final product compound WLE12 (40 mg, 90.0 μmol, yield: 38.0%). 1 H NMR (400MHz, CD3OD) δ7.58 (d, J = 0.8Hz, 1H), 4.94-5.02 (m, 1H), 4.56-4.64 (m, 1H), 3.76-4.07 (m, 2H), 3.46-3.59 (m, 1H), 2.91-3.08(m,2H),2.88(d,J=10.4Hz,3H),2.67-2.79(m,1H),2.60(dd,J=16.4,4.8Hz,1H),1.12-1.25(m,3H).LCMS(M+H) + m / z:445.4.

[0246] Example 13: Synthesis of compound WLE13

[0247]

[0248] Step 1:

[0249] Compound 1-9 (800 mg, 1.93 mmol) was dissolved in DMF (10.0 mL). Then, 13-1 (823 mg, 5.79 mmol), HATU (1.10 g, 2.90 mmol), and DIEA (1.25 g, 9.65 mmol, 1.7 mL) were added sequentially. After addition, the mixture was stirred at 25 °C for 2 hours. The reaction was monitored by TLC until complete. Water (20.0 mL) was added to quench the reaction mixture. The mixture was extracted with ethyl acetate (20.0 mL), washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 13-2 (850 mg, 1.58 mmol, yield: 81.8%), a brown liquid. LCMS (M+H) + m / z:539.2.

[0250] Step Two:

[0251] Intermediate compound 13-2 (800 mg, 1.49 mmol) was dissolved in TFA (15.3 g, 134 mmol, 10.0 mL), and reacted at 60 °C for 5 hours. The reaction was monitored by TLC. After the reaction was completed, the organic phase was removed by vacuum concentration. The resulting mixture was then prepared by high performance liquid chromatography to obtain the final product compound WLE13 (80.0 mg, 191 μmol, yield: 13%). 1 H NMR (400MHz, CD3OD) δ6.58(d,J=0.4Hz,1H),4.95(d,J=16.8Hz,1H),4.57-4.65(m,1H),3.91-4.03(m,1H),3.75-3.86(m,1H),3.45 -3.54(m,1H),2.83-3.04(m,6H),2.71-2.77(m,1H),2.56(dd,J=16.0,5.1Hz,1H),1.22-1.26(m,6H),1.13-1.21(m,3H).LCMS(M+H) + m / z:419.2.

[0252] Example 14: Synthesis of compound WLE14

[0253]

[0254] Step 1:

[0255] Compound 7-1 (2.50 g, 6.45 mmol) was dissolved in DMF (125 mL). HATU (2.94 g, 7.74 mmol) and DIEA (4.17 g, 32.3 mmol, 5.6 mL) were added sequentially. After addition, the mixture was stirred at 25 °C for half an hour. Then, compound 14-1 (789 mg, 7.10 mmol) was added, and the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until completion. Water (20.0 mL) was added to quench the reaction mixture. The mixture was filtered, and the filter cake was purified by silica gel column chromatography to obtain intermediate compound 14-2 (82.0 g, 4.16 mmol, yield: 64.5%) as a yellow solid. LCMS (M+H) + m / z:481.1.

[0256] Step Two:

[0257] Intermediate compound 14-2 (1.00 g, 2.08 mmol) was dissolved in dichloromethane (7.5 mL), and a solution of 1,4-dioxane in hydrochloric acid (4.0 M, 2.5 mL) was added. After the addition was complete, the reaction was carried out at 25 °C for 12 hours under TLC monitoring. Upon completion of the reaction, the organic solvent was removed by concentration under reduced pressure to give a yellow solid, which was the crude intermediate compound 14-3 (630 mg, 1.66 mmol, yield: 79.6%). LCMS (M+H) + m / z:381.0.

[0258] Step 3:

[0259] Intermediate compound 14-3 (630 mg, 1.66 mmol) was dissolved in DMF (125 mL). Then, HATU (756 mg, 1.99 mmol), DIEA (1.07 g, 8.28 mmol, 1.44 mL), and compound 1-7 (242 mg, 1.66 mmol) were added sequentially. After addition, the reaction was carried out at 25 °C for 2 hours. The reaction was monitored by TLC until completion. Water (20.0 mL) was added to quench the reaction mixture. The mixture was filtered, and the filter cake was purified by silica gel column chromatography to obtain intermediate compound 14-4 (630 mg, 1.24 mmol, yield: 74.8%) as a yellow solid. LCMS (M+H) + m / z:509.3.

[0260] Step Four:

[0261] Intermediate compound 14-4 (630 mg, 1.24 mmol) was dissolved in methanol (2.0 mL), THF (6.0 mL), and water (2.0 mL). Lithium hydroxide monohydrate (520 mg, 12.4 mmol) was added. After the addition was complete, the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until completion. After removing the organic phase by concentration under reduced pressure, water (10.0 mL) was added to the reaction flask, and the pH was adjusted to 5–6 with 1.0 M hydrochloric acid aqueous solution. After stirring for 10 minutes, the mixture was filtered to obtain a yellow solid, which was the crude intermediate compound 14-5 (610 mg). LCMS (M+H) + m / z:495.1.

[0262] Step 5:

[0263] Intermediate compound 14-5 (2.00 g, 5.66 mmol) was dissolved in DMF (15.0 mL). HATU (148 mg, 388 μmol) and DIEA (209 mg, 1.62 mmol) were added sequentially. After the additions were complete, the mixture was stirred at 25 °C for half an hour. Then, compound 1-10 (101 mg, 647 μmol) was added to the reaction solution, and the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until completion. Water (50.0 mL) was added to quench the reaction, and the mixture was stirred for 10 minutes. The mixture was then filtered, and the filter cake was purified by silica gel column chromatography to obtain intermediate compound 14-6 (100 mg, 158 μmol, yield: 48.6%) as a yellow solid. LCMS (M+H) + m / z:633.4.

[0264] Step Six:

[0265] Intermediate compound 14-6 (100 mg, 158 μmol) was dissolved in TFA (1.54 g, 13.5 mmol, 1.0 mL), and reacted at 60 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, the organic phase was removed by concentration under reduced pressure. The resulting mixture was then subjected to high performance liquid chromatography to obtain the final product compound WLE14 (10 mg, 19.5 μmol, yield: 12.3%). 1H NMR (400MHz, DMSO-d6) δ13.04-13.23(m,1H),11.94-12.15(m,1H),9.21-9.48(m,1H),7.32-7.51(m,1H),6.68(d,J=2.0Hz,1H),4.42-4.80(m, 2H),3.67-3.88(m,5H),2.75-2.91(m,2H),2.66(brs,1H),2.41-2.49(m ,2H),2.16(d,J=9.2Hz,3H),1.24(s,9H),0.98-1.10(m,3H).LCMS(M+H) + m / z:513.3.

[0266] Example 15: Synthesis of compound WLE15

[0267]

[0268] Step 1:

[0269] Compound 7-1 (500 mg, 1.29 mmol) was dissolved in DMF (5.0 mL), and HATU (736 mg, 1.94 mmol) and DIEA (500 mg, 3.87 mmol, 674 μL) were added sequentially. After the addition was complete, the mixture was stirred at 25 °C for half an hour. Then, 4-aminotetrahydropyran (144 mg, 1.42 mmol) was added, and the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until it was complete. The reaction was quenched by adding water (20.0 mL), extracted with ethyl acetate (30.0 mL × 2), washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove ethyl acetate. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 15-1 (376 mg, 79.9 μmol, yield: 60.1%) as a white solid. LCMS (M+H) + m / z:471.2.

[0270] Step Two:

[0271] Intermediate compound 15-1 (376 mg, 799 μmol) was dissolved in dichloromethane (3.0 mL), and a 1,4-dioxane solution of hydrochloric acid (4.0 M, 1.0 mL) was added. After the addition was complete, the reaction was carried out at 25 °C for 2 hours under TLC monitoring. Upon completion of the reaction, the organic solvent was removed by concentration under reduced pressure to obtain a yellowish-white solid, which was the crude intermediate compound 15-2 (320 mg). LCMS (M+H) + m / z:371.1.

[0272] Step 3:

[0273] Compound 1-7 (242 mg, 1.66 mmol) was dissolved in DMF (2.0 mL). HATU (429 mg, 1.13 mmol) and DIEA (486 mg, 3.76 mmol, 656 μL) were added sequentially. After addition, the mixture was stirred at 25 °C for half an hour. Then, intermediate compound 15-2 (320 mg, 786 μmol) was added. After addition, the mixture was reacted at 25 °C for 3 hours. The reaction was monitored by TLC until completion. The reaction was quenched by adding water (20.0 mL). The mixture was extracted with ethyl acetate (30.0 mL × 2), washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove ethyl acetate. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 15-3 (335 mg, 672 μmol, yield: 89.2%) as a yellow solid. LCMS (M+H) + m / z:499.4.

[0274] Step Four:

[0275] Intermediate compound 15-3 (335 mg, 672 μmol) was dissolved in methanol (3.0 mL) and water (0.5 mL). Lithium hydroxide monohydrate (141 mg, 3.36 mmol) was added. After the addition was complete, the reaction was carried out at 25 °C for 2 hours. The reaction was monitored by TLC until completion. After removing the organic phase by concentration under reduced pressure, water (10.0 mL) was added to the reaction flask, and the pH was adjusted to 5–6 with 1.0 M hydrochloric acid aqueous solution. After stirring for 10 minutes, the mixture was extracted with dichloromethane (50.0 mL × 3). The dichloromethane layer was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the dichloromethane, yielding a yellow oily substance, which was the crude intermediate compound 15-4 (230 mg). LCMS (M+H) + m / z:485.1.

[0276] Step 5:

[0277] Intermediate compound 15-4 (230 mg, 475 μmol) was dissolved in DMF (2.0 mL). HATU (271 mg, 712 μmol) and DIEA (184 mg, 1.42 mmol, 248 μL) were added sequentially. After addition, the mixture was stirred at 25 °C for half an hour. Then, compound 1-10 (111 mg, 712 μmol) was added to the reaction solution. The reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until completion. Water (50.0 mL) was added to quench the reaction, and the mixture was extracted with ethyl acetate (30.0 mL × 2). The organic phase was washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove ethyl acetate. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 15-5 (200 mg, 321 μmol, yield: 67.6%) as a yellow solid. LCMS (M+H)+ m / z:623.2.

[0278] Step Six:

[0279] Intermediate compound 15-5 (200 mg, 321 μmol) was dissolved in TFA (3.0 mL) and reacted at 60 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, the organic phase was removed by vacuum concentration. The resulting mixture was then prepared by high performance liquid chromatography to obtain the final product compound WLE15 (33 mg, 65.7 μmol, yield: 20.4%). 1 H NMR(400MHz,CD3OD)δ6.55-6.59(m,1H),4.95(d,J=16.8Hz,1H),4.91-5.00(m,1H),4 .54-4.66(m,1H),4.03-4.13(m,1H),3.91-4.02(m,3H),3.74-3.87(m,1H),3.45-3.60 (m,3H),2.84-3.10(m,2H),2.67-2.79(m,1H),2.54(dd,J=16.0,4.8Hz,1H),1.82-1.9 4(m,2H),1.65(t,J=11.2Hz,2H),1.28(d,J=2.4Hz,9H),1.13-1.23(m,3H).LCMS(M+H) + m / z:503.7.

[0280] Example 16: Synthesis of compound WLE16

[0281]

[0282] Step 1:

[0283] Under a nitrogen atmosphere, compound 2-7 (500 mg, 804 μmol) was dissolved in 1,4-dioxane (3.0 mL) and water (3.0 mL). Pyridine-4-boronic acid (198 mg, 1.61 mmol), Pd(dppf)Cl2 (118 mg, 161 μmol), and potassium carbonate (334 mg, 2.41 mmol) were added sequentially. After addition, the reaction was carried out at 90 °C for 12 hours. The reaction was monitored by TLC until completion. After the reaction solution cooled to room temperature, water (5.0 mL) was added to quench the reaction. Extraction was performed with ethyl acetate (5.0 mL × 2), and the solution was dried over anhydrous sodium sulfate. The mixture was filtered, concentrated under reduced pressure to remove the organic phase, and the resulting mixture was purified by silica gel column chromatography to give intermediate compound 16-1 (200 mg, 349 μmol, yield: 43.4%) as a yellow solid. LCMS (M+H) + m / z:573.2.

[0284] Step Two:

[0285] Intermediate compound 16-1 (210 mg, 367 μmol) was dissolved in TFA (3.07 g, 26.9 mmol, 2.0 mL) and reacted at 70 °C for 24 hours. The reaction was monitored by TLC. After the reaction was completed, water (3.0 mL) was added to the reaction solution to quench the reaction, and the mixture was extracted with dichloromethane (5.0 mL × 2). The organic phase was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was then subjected to high performance liquid chromatography to prepare the final product compound WLE16 (25 mg, 55.2 μmol, yield: 15.0%). 1 HNMR(400MHz, CD3OD)δ8.57(brs,2H),7.57-7.79(m,2H),6.58(s,1H),4.93-5.00(m,1H),4.68-4.79(m,1H),3.88 -4.10(m,2H),3.50-3.60(m,1H),2.77-3.11(m,3H),2.51-2.61(m,1H),1.27(s,9H),1.13-1.25(m,3H).LCMS(M+H) + m / z:453.2.

[0286] Example 17: Synthesis of compound WLE17

[0287]

[0288] Step 1:

[0289] Under a nitrogen atmosphere, compound 2-7 (350 mg, 563 μmol) was dissolved in 1,4-dioxane (10.0 mL) and water (1.0 mL). Then, 3,6-dihydro-2H-pyran-4-boronic acid pinacol ester (177 mg, 844 μmol), Pd(dppf)Cl2 (82.4 mg, 112 μmol), and potassium carbonate (233 mg, 1.69 mmol) were added sequentially. After the additions were complete, the reaction was carried out at 90 °C for 12 hours. The reaction was monitored by TLC until completion. After the reaction solution cooled to room temperature, water (5.0 mL) was added to quench the reaction. The mixture was extracted with dichloromethane (20.0 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 17-1 (170 mg, 294 μmol, yield: 52.3%) as a brown solid. LCMS (M+H) + m / z:578.0.

[0290] Step Two:

[0291] Under a hydrogen atmosphere, intermediate compound 17-1 (170 mg, 294 μmol) was dissolved in methanol (5.0 mL). Pd / C (500 mg, 469 μmol) was then added. After the addition was complete, the reaction was carried out at 30 °C for 12 hours. TLC analysis was performed. Upon completion of the reaction, the mixture was filtered to remove Pd / C. The filtrate was concentrated under reduced pressure to remove the organic solvent, yielding a white solid, which was the crude intermediate compound 17-2 (150 mg). LCMS (M+H) + m / z:580.1.

[0292] Step 3:

[0293] Intermediate compound 17-2 (150 mg, 258 μmol) was dissolved in TFA (14.4 g, 126 mmol, 9.4 mL) and TfOH (159 mg, 1.0 mmol). After the addition was complete, the reaction was carried out at 70 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, water (3.0 mL) was added to the reaction solution to quench the reaction. The mixture was extracted with dichloromethane (5.0 mL × 2), and the organic phase was dried over anhydrous sodium sulfate. The mixture was filtered and concentrated under reduced pressure to remove the organic phase. The resulting mixture was then subjected to high performance liquid chromatography to prepare the final product compound WLE17 (18 mg, 39.1 μmol, yield: 56.7%). 1 HNMR(400MHz,CD3OD)δ6.59(d,J=2.0Hz,1H),4.60(s,2H),3.74-4.05(m,4H),3.47-3.60(m,3H),2.75-3.00(m,3H),2. 68(t,J=5.6Hz,1H),2.55(ddd,J=16.0,8.4,5.2Hz,1H),1.75-1.89(m,4H),1.28(s,9H),1.13-1.23(m,3H).LCMS(M+H) + m / z:460.2.

[0294] Example 18: Synthesis of compound WLE18

[0295]

[0296] Step 1:

[0297] Compound 1-9 (300 mg, 723 μmol) was dissolved in dichloromethane (5.0 mL). DIEA (468 mg, 3.62 mmol, 630 μL), T4P (3.13 g, 4.34 mmol), and 2-amino-4-bromothiazole (130 mg, 724 μmol) were added sequentially. After addition, the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until completion. After the reaction solution cooled to room temperature, water (10.0 mL) was added to quench the reaction. Extraction was performed with dichloromethane (10.0 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 18-1 (160 mg, 261 μmol, yield: 36.1%) as a yellow solid. LCMS (M+H) + m / z:574.9.

[0298] Step Two:

[0299] Intermediate compound 18-1 (150 mg, 261 μmol) was dissolved in TFA (2.30 g, 20.2 mmol, 1.5 mL) and reacted at 70 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, water (3.00 mL) was added to the reaction solution to quench the reaction. The mixture was extracted with dichloromethane (5.00 mL × 2), and the organic phase was dried over anhydrous sodium sulfate. The mixture was filtered and concentrated under reduced pressure to remove the organic phase. The resulting mixture was then subjected to high performance liquid chromatography to prepare the final product compound WLE18 (20 mg, 43.9 μmol, yield: 16.8%). 1 H NMR (400MHz, DMSO-d6) δ12.97(d,J=10.8Hz,1H),12.39(s,1H),7.92-8.17(m,1H),7.24(d,J=2.8Hz,1H),4.38-4.79(m,2H),3.58-3.88(m ,2H),2.77-2.91(m,2H),2.72(dd,J=8.8,4.4Hz,3H),2.63(brs,1H),2.51-2.54(m,1H),2.45-2.49(m,1H),0.95-1.15(m,3H).LCMS(M+H) + m / z:455.0.

[0300] Example 19: Synthesis of compound WLE19

[0301]

[0302] Step 1:

[0303] Under a nitrogen atmosphere, compound 19-1 (2.00 g, 12.4 mmol) was dissolved in tert-butanol (20.0 mL), and TEA (2.51 g, 24.8 mmol, 3.5 mL) and DPPA (5.12 g, 18.6 mmol, 4.0 mL) were added sequentially. After the addition was complete, the reaction was carried out at 80 °C for 15 hours under TLC monitoring. When the reaction was complete, water (50.0 mL) was added to the reaction solution to quench the reaction, and the mixture was extracted with ethyl acetate (100 mL × 2). The organic layer was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove ethyl acetate. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 19-2 (920 mg, 3.96 mmol, yield: 31.9%), which was a yellow oil. 1 H NMR (400MHz, CDCl3) δ10.64(brs,1H),9.97(brs,1H),7.26-7.32(m,2H),6.84-6.94(m,2H),5.88(d,J=1.6Hz,1H),1.50(s,9H).LCMS(M-56+H) + m / z:177.1.

[0304] Step Two:

[0305] Intermediate compound 19-2 (500 mg, 2.15 mmol) was dissolved in 1,4-dioxane hydrochloric acid (4.0 M, 3.0 mL), and reacted at 25 °C for 12 hours. The reaction was monitored by TLC until it ended. The organic solvent was removed by concentration under reduced pressure to obtain crude intermediate compound 19-3 (350 mg), which was a yellow oil. 1 H NMR(400MHz,DMSO-d6)δ12.30(brs,1H),9.64-10.18(m,1H),7.39-7.52(m,2H),7.28-7.37(m,3H),7.05-7.23(m,2H).LCMS(M+H) + m / z:133.0.

[0306] Step 3:

[0307] Compound 1-9 (300 mg, 724 μmol) was dissolved in DMF (3.0 mL). DIEA (468 mg, 3.62 mmol, 630 μL) and HATU (413 mg, 1.09 mmol) were added sequentially. After addition, the mixture was stirred at 25 °C for half an hour. Intermediate compound 19-3 (200 mg, 1.19 mmol) was then added. After addition, the mixture was reacted at 25 °C for 2 hours. The reaction was monitored by TLC until completion. After the reaction solution cooled to room temperature, water (20.0 mL) was added to quench the reaction. The mixture was extracted with dichloromethane (50.0 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic solvent. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 19-4 (300 mg, 568 μmol, yield: 78.5%), a yellow oil. LCMS (M+H) + m / z:529.1.

[0308] Step Four:

[0309] Intermediate compound 19-4 (150 mg, 284 μmol) was dissolved in TFA (3.0 mL) and reacted at 70 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, water (3.0 mL) was added to the reaction solution to quench the reaction. The mixture was extracted with dichloromethane (5.0 mL × 2), and the organic phase was dried with anhydrous sodium sulfate. The mixture was filtered and concentrated under reduced pressure to remove the organic solvent. The resulting mixture was then subjected to high performance liquid chromatography to prepare the final product compound WLE19 (2 mg, 5.75 μmol, yield: 2.02%). 1 H NMR (400MHz, CD3OD) δ7.18-7.38(m,2H),7.00-6.92(m,2H),5.93(d,J=12.4Hz,1H),4.93-5.03(m,2H),4.56-4.66(m,1H),3.76-4. 02(m,2H),3.44-3.62(m,1H),2.83-2.91(m,4H),2.74(d,J=4.4Hz,1H),2.52(dd,J=15.6,5.2Hz,1H),1.14-1.25(m,3H).LCMS(M+H) + m / z:409.2.

[0310] Example 20: Synthesis of compound WLE20

[0311]

[0312] Step 1:

[0313] Compounds 1-9 (250 mg, 603 μmol) were dissolved in dichloromethane (5.0 mL), and HATU (344 mg, 905 μmol) and DIEA (390 mg, 3.02 mmol, 525 μL) were added. The mixture was stirred at 25 °C for 1 hour. Then, compound 20-1 (84.6 mg, 603 μmol) was added, and the mixture was stirred at 25 °C for 3 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10 mL × 3), and the organic phases were combined. The mixture was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 20-2 (200 mg, 373 μmol, yield 61.8%), a yellow oily compound. LCMS [M+H] + m / z:537.3.

[0314] Step Two:

[0315] TFA (3.0 mL) was added to intermediate compound 20-2 (130 mg, 242 μmol), and the mixture was stirred at 70 °C for 12 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was prepared by high performance liquid chromatography to obtain the final product compound WLE20 (10 mg, 24.3 μmol, yield 10.0%), which was a white solid. 1 H NMR(400MHz,CD3OD)δ7.25(s,1H),4.96-5.07(m,1H),4.51-4.63(m,2H),3.87-4.08(m,1H),3.71-3.86(m,1H),3.42-3.56(m,1H),2 .96-3.12(m,1H),2.86-2.91(m,3H),2.75(q,J=5.6Hz,1H),2.47-2.63(m,1H),1.22(d,J=1.6Hz,9H),1.12-1.17(m,3H).LCMS[M+H] + m / z:417.2.

[0316] Example 21: Synthesis of compound WLE21

[0317]

[0318] Step 1:

[0319] Under a nitrogen atmosphere, compound 1-bromopinazone (2.00 g, 12.6 mmol) was dissolved in DMF (10.0 mL), and potassium iodide (209 mg, 1.26 mmol) and compound 21-1 (2.25 g, 12.6 mmol, 1.69 mL) were added sequentially. After the addition was complete, the reaction was carried out at 25 °C for 60 hours under TLC monitoring. When the reaction was complete, water (100 mL) was added to the reaction solution to quench the reaction, and the mixture was extracted with ethyl acetate (150 mL × 2). The organic layer was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove ethyl acetate. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 21-2 (630 mg, 2.64 mmol, yield: 20.9%), which was a yellow oil. 1 HNMR(400MHz,DMSO-d6)δ6.42-6.40(m,1H),6.38(brs,2H),1.53(s,9H),1.12(s,9H).LCMS(M-56+H) + m / z:184.2.

[0320] Step Two:

[0321] Intermediate compound 21-2 (100 mg, 418 μmol) was dissolved in 1,4-dioxane hydrochloric acid (4.0 M, 1.0 mL), and the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until it ended. The organic solvent was removed by concentration under reduced pressure to obtain crude intermediate compound 21-3 (73.0 mg), which was a yellow oil. 1 H NMR(400MHz,DMSO-d6)δ12.25(brs,1H),11.63(brs,1H),7.14(s,2H),6.49(s,1H),1.20(s,9H).LCMS(M+H) + m / z:140.2.

[0322] Step 3:

[0323] Compound 1-9 (130 mg, 314 μmol) was dissolved in 3.0 mL of dichloromethane. Then, DIEA (203 mg, 1.57 mmol, 273 μL) and HATU (179 mg, 471 μmol) were added sequentially. After the addition was complete, the mixture was stirred at 25 °C for half an hour. Then, intermediate compound 21-3 (55.1 mg, 314 μmol) was added. After the addition was complete, the mixture was reacted at 25 °C for 2 hours. The reaction was monitored by TLC until completion. After the reaction solution cooled to room temperature, water (20.0 mL) was added to quench the reaction. The mixture was extracted with dichloromethane (30.0 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 21-4 (160 mg, 299 μmol, yield: 95.2%), which was a yellow oil. LCMS (M+H) + m / z:536.5.

[0324] Step Four:

[0325] Intermediate compound 21-4 (60.0 mg, 112 μmol) was dissolved in TFA (2.0 mL) and TfOH (0.1 mL), and reacted at 25 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, water (3.0 mL) was added to the reaction solution to quench the reaction, and the mixture was extracted with dichloromethane (5.0 mL × 2). The organic phase was dried with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was then subjected to high performance liquid chromatography to prepare the final product compound WLE21 (5 mg, 12.0 μmol, yield: 10.5%). 1 H NMR (400MHz, CD3OD) δ6.81(d,J=8.0Hz,1H),5.05(d,J=16.4Hz,1H),4.52(d,J=16.4Hz,1H),3.99-4.12(m,1H),3.74-3.97(m,2H),3.42-3.54(m, 1H),2.96-3.03(m,1H),2.87-2.90(m,3H),2.75(t,J=5.2Hz,1H),2.65( dd,J=16.8,4.0Hz,1H),1.27-1.35(m,9H),1.09-1.23(m,3H).LCMS(M+H) + m / z:416.3.

[0326] Example 22: Synthesis of compound WLE22

[0327]

[0328] Step 1:

[0329] Compound 1-9 (150 mg, 362 μmol) was dissolved in DMF (2.0 mL). DIEA (234 mg, 1.81 mmol, 315 μL) and HATU (206 mg, 543 μmol) were added sequentially. After the additions were complete, the mixture was stirred at 25 °C for half an hour. Then, 2-aminobenzothiazole (59.8 mg, 398 μmol) was added. After the additions were complete, the mixture was reacted at 25 °C for 2 hours. The reaction was monitored by TLC until completion. Water (20.0 mL) was added to quench the reaction. The mixture was filtered, and the resulting filter cake was the crude intermediate compound 22-1 (200 mg), a yellow solid. LCMS (M+H) + m / z:547.1.

[0330] Step Two:

[0331] Intermediate compound 22-1 (150 mg, 274 μmol) was dissolved in TFA (3.07 g, 26.9 mmol, 2.0 mL) and reacted at 70 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, water (3.0 mL) was added to the reaction solution to quench the reaction. The mixture was extracted with dichloromethane (5.0 mL × 2), and the organic phase was dried over anhydrous sodium sulfate. The mixture was filtered and concentrated under reduced pressure to remove the organic phase. The resulting mixture was subjected to high performance liquid chromatography to obtain the final product compound WLE22 (10 mg, 5.93 μmol, yield: 8.55%). 1 H NMR(400MHz,DMSO-d6)δ12.99(d,J=9.6Hz,1H),12.34(br s,1H),7.95-8.09(m,1H),7.93(d,J=8.0Hz,1H),7.72(d,J=8.0Hz,1H),7.42(t,J=7.6Hz,1H),7.25-7.32(m,1H),4.41-4.78(m,2H),3.64-3. 89(m,2H),2.80-2.97(m,2H),2.72(dd,J=12.4,4.4Hz,3H),2.64(s,1H),2.53-2.60(m,1H),2.28-2.41(m,1H),1.02-1.12(m,3H).LCMS(M+H) + m / z:427.1.

[0332] Example 23: Synthesis of compound WLE23

[0333]

[0334] Step 1:

[0335] Compound 1-9 (300 mg, 724 μmol) was dissolved in DMF (5.0 mL). DIEA (468 mg, 3.62 mmol, 630 μL) and HATU (413 mg, 1.09 mmol) were added sequentially. After the addition was complete, the mixture was stirred at 25 °C for half an hour. Then, 2-naphthylamine (114 mg, 796 μmol) was added. After the addition was complete, the mixture was reacted at 25 °C for 2 hours. The reaction was monitored by TLC until it ended. Water (20.0 mL) was added to quench the reaction. The mixture was filtered, and the filter cake was purified by silica gel column chromatography to obtain intermediate compound 23-1 (370 mg, 686 μmol, yield: 94.7%), which was a yellow solid. 1 H NMR (400MHz, DMSO-d6) δ8.50(brs,1H),8.16(s,1H),7.72(brs,4H),7.38-7.46(m,3H),7.03(d,J=8.8Hz,2H),6.81(d,J=8.8Hz,2H),5. 02-5.09(m,2H),4.81-4.92(m,2H),3.75(s,3H),2.95-2.98(m,4H),2.51-2.58(m,3H),1.68(brs,1H),1.23(d,J=7.2Hz,3H).LCMS(M+H) + m / z:540.1.

[0336] Step Two:

[0337] Intermediate compound 23-1 (300 mg, 556 μmol) was dissolved in TFA (6.14 g, 53.9 mmol, 4.0 mL) and TfOH (339 mg, 2.26 mmol, 200 μL). The reaction was carried out at 70 °C for 12 hours under TLC monitoring. After the reaction was completed, water (3.00 mL) was added to the reaction solution to quench the reaction. The mixture was extracted with dichloromethane (5.00 mL × 2), and the organic phase was dried over anhydrous sodium sulfate. The mixture was filtered and concentrated under reduced pressure to remove the organic phase. The resulting mixture was then subjected to high performance liquid chromatography to prepare the final product compound WLE23 (15 mg, 35.8 μmol, yield: 6.43%). 1 H NMR (400MHz, DMSO-d6) δ8.06-8.12(m,1H),7.71-7.78(m,3H),7.35-7.49(m,3H),4.89-5.01(m,2H),3.85-4.12(m,2H),3.49-3. 57(m,1H),2.90-3.05(m,1H),2.84-2.89(m,4H),2.72-2.77(m,1H),2.54(dd,J=15.2,4.8Hz,1H),1.17-1.25(m,3H).LCMS(M+H)+ m / z:420.2.

[0338] Example 24: Synthesis of compounds WLE24 and WLE25

[0339]

[0340] Step 1:

[0341] Under a nitrogen atmosphere, compound 24-1 (600 mg, 4.11 mmol) was dissolved in dichloromethane (10.0 mL), cooled to 0 °C, and trifluoroacetic anhydride (1.03 g, 4.93 mmol, 684 μL) was added. The reaction was carried out at 0 °C for 2 hours. TLC monitoring showed the reaction was complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was the crude intermediate compound 24-2 (520 mg), a white solid.

[0342] Step Two:

[0343] Under a nitrogen atmosphere, intermediate compound 24-2 (456 mg, 3.56 mmol), TEA (721 mg, 7.13 mmol, 992 μL), and DMAP (43.5 mg, 356 μmol) were dissolved in DMF (10.0 mL). Compound 1-6 (1.20 g, 3.56 mmol) and TEA (721 mg, 7.13 mmol, 992 μL) were added, and the mixture was stirred at 25 °C for 12 hours. TLC monitoring showed the reaction was complete. The reaction was quenched by adding water (20.0 mL), extracted with ethyl acetate (50.0 mL × 2), washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 24-3 (700 mg, 1.64 mmol, yield: 46%) as a yellow solid. LCMS [M+H] + m / z:429.1.

[0344] Step 3:

[0345] Intermediate compound 24-3 (700 mg, 1.64 mmol) was dissolved in DMF (10.0 mL), and HATU (932 mg, 2.45 mmol) and DIEA (845 mg, 6.53 mmol, 1.14 mL) were added. The mixture was stirred at 25 °C for 1 hour. Then, compound 1-10 (306 mg, 1.96 mmol) was added, and the mixture was stirred at 25 °C for 5 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (50.0 mL), extracted with ethyl acetate (50.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (50.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 24-4 (836 mg, 1.48 mmol, yield: 90%) as a yellow solid. LCMS [M+H] + m / z:567.2.

[0346] Step Four:

[0347] TFA (5.0 mL) was added to intermediate compound 24-4 (400 mg, 705 μmol), and the reaction was carried out at 60 °C for 12 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was subjected to SFC to prepare final product compounds WLE24 (7 mg, 15.7 μmol, yield: 2%) and WLE25 (11 mg, 24.7 μmol, yield: 3.5%). WLE24 was a white solid. 1 H NMR (400MHz, CD3OD) δ6.65 (d, J = 8.4Hz, 1H), 4.75-4.52 (m, 1H), 4.05-3.63 (m, 3H), 3.29-3.21 ( m,1H),2.94-2.75(m,5H),2.23-1.60(m,1H),1.32-1.23(m,9H),1.17-0.86(m,6H).LCMS[M+H] + m / z: 447.2. WLE25 is a white solid. 1 HNMR (400MHz, CD3OD) δ6.55 (d, J = 8.4Hz, 1H), 4.56-4.58 (m, 1H), 3.92-3.53 (m, 3H), 3. 20-3.24(m,1H),2.93-2.65(m,5H),2.24-1.60(m,1H),1.31-0.71(m,15H).LCMS[M+H] + m / z:447.2.

[0348] Example 25: Synthesis of compound WLE26

[0349]

[0350] Step 1:

[0351] Under a nitrogen atmosphere, compound 2-7 (150 mg, 241 μmol) was dissolved in DMF (4.0 mL). CuI (9.19 mg, 48.3 μmol), L-proline (11 mg, 96.5 μmol), potassium carbonate (67 mg, 483 μmol), and 4-aminotetrahydropyran (292 mg, 289 μmol) were added sequentially. After addition, the reaction was carried out at 95 °C for 12 hours. The reaction was monitored by TLC until completion. After the reaction solution cooled to room temperature, water (20.0 mL) was added to quench the reaction. Extraction was performed with ethyl acetate (30.0 mL × 2), and the solution was dried over anhydrous sodium sulfate. The mixture was filtered, concentrated under reduced pressure to remove the organic phase, and the resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 26-1 (35 mg, 58.9 μmol, yield: 24.4%) as a yellow solid. LCMS (M+H) + m / z:595.3.

[0352] Step Two:

[0353] Compound 26-1 (25 mg, 42.0 μmol) was dissolved in TFA (5 mg, 42.0 μmol) and reacted at 70 °C for 48 hours. The reaction was monitored by TLC. After the reaction was completed, the organic phase was removed by vacuum concentration. The resulting mixture was then prepared by high performance liquid chromatography to obtain the final product compound WLE26 (3 mg, 6.52 μmol, yield: 15.6%). 1 H NMR (400MHz, CD3OD) δ6.60 (d, J = 5.2Hz, 1H), 4.62 (brs, 1H), 4.44-4.54 (m, 1H), 3.90-4.01 (m, 3H), 3.86-3.90 (m, 0.5H), 3.60-3.73 (m, 0.5H), 3.42- 3.58(m,4H),2.82-3.00(m,2H),2.49-2.68(m,2H),1.99(t,J=13.6Hz,2H ),1.40-1.55(m,2H),1.28(d,J=2.0Hz,9H),1.10-1.24(m,3H).LCMS(M+H) + m / z:475.2.

[0354] Example 26: Synthesis of compound WLE27

[0355]

[0356] Step 1:

[0357] Compound 2-7 (100 mg, 161 μmol), 4-aminopyridine (23 mg, 241 μmol, 41 μL), potassium phosphate (51 mg, 241 μmol), CuI (31 mg, 160 μmol), and compound 27-1 (11.4 mg, 80.5 μmol) were dissolved in tert-amyl alcohol (5.0 mL) under a nitrogen atmosphere and reacted at 90 °C for 12 h. After the reaction was complete as monitored by TLC, the reaction was quenched with water (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was then subjected to high-performance liquid chromatography (HPLC) to prepare intermediate compound 27-2 (25 mg, 42.6 μmol, yield 26.5%) as a white solid. LCMS [M+H] + m / z:588.3.

[0358] Step Two:

[0359] TFA (3.0 mL) was added to intermediate compound 27-2 (18 mg, 30.6 μmol), and the mixture was stirred at 70 °C for 2 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was used to prepare the final product compound WLE27 (5 mg, 10.6 μmol, yield 34.6%) as a white solid by high performance liquid chromatography. 1 H NMR (400MHz, CD3OD) δ8.16-8.30(m,2H),7.40-7.69(m,2H),6.79(s,1H),4.62-4.75(m,2H),4.35-4.00-4.12(m,1H),3.82- 3.93(m,1H),3.59-3.59(m,1H),2.84-3.11(m,2H),2.62-2.79(m,2H),1.32(d,J=3.2Hz,9H),1.16-1.26(m,3H).LCMS[M+H] + m / z:468.2.

[0360] Example 27: Synthesis of compound WLE28

[0361]

[0362] Step 1:

[0363] Compound 28-1 (7.64 g, 46.8 mmol) was dissolved in DMF (50.0 mL), and potassium carbonate (13.0 g, 93.7 mmol) and PMB-Cl (14.7 g, 93.7 mmol, 12.7 mL) were added. The reaction mixture was reacted at 25 °C for 12 hours. After the reaction was complete as monitored by TLC, the solvent was removed by vacuum concentration, water was added for dilution, and the mixture was concentrated by vacuum filtration. The filter cake was the crude intermediate compound 28-2 (18.0 g), a yellow solid. LCMS [M+H] + m / z:404.0.

[0364] Step Two:

[0365] Under a nitrogen atmosphere, intermediate compound 28-2 (18.0 g, 44.6 mmol) was dissolved in a mixed solution of THF (20.0 mL), methanol (100 mL), and water (20.0 mL). Lithium hydroxide monohydrate (9.36 g, 223 mmol) was added, and the mixture was stirred at 25 °C for 1 hour. The reaction was monitored by TLC until complete. The reaction solution was diluted with water (100 mL), the pH was adjusted to 3 with 1.0 M hydrochloric acid, and the mixture was extracted with ethyl acetate (100 mL × 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 28-3 (7.97 g, 28.2 mmol, yield 63.1%) as a yellow solid. LCMS [M+H] + m / z:283.9.

[0366] Step 3:

[0367] Intermediate compound 28-3 (4.50 g, 15.9 mmol) was dissolved in dichloromethane (45.0 mL), and oxalyl chloride (4.03 g, 31.8 mmol, 2.78 mL) and DMF (10 μL) were added. The reaction was carried out at 25 °C for 12 hours. The reaction was monitored by LCMS until completion. The solvent was removed by concentration under reduced pressure, and the resulting mixture was the crude intermediate compound 28-4 (4.79 g), a brown solid. LCMS: [M-Cl + MeO + H] + m / z:297.9.

[0368] Step 4: Synthesis of intermediate compound 28-5

[0369] Compound 28-5-1 (4.00 g, 27.4 mmol, 3.64 mL) was dissolved in THF (456 mL) and water (456 mL). A 0.25 M, 131 mL aqueous solution of potassium hydroxide was added at 0 °C, and the reaction was carried out at 0 °C for 3 hours. The reaction was monitored by TLC until completion. Water (100 mL) was added to quench the reaction, and the pH was adjusted to 3. Extraction was performed with ethyl acetate (100 mL × 3), followed by washing with a saturated sodium chloride solution (100 mL × 3). The mixture was dried over anhydrous sodium sulfate, and the solvent was removed by concentration under reduced pressure. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 28-5 (3.60 g, 27.3 mmol, yield 99.6%), a colorless liquid. 1 H NMR: (400MHz, DMSO-d6) δ13.07-12.33(m,1H),3.64(s,3H),1.25(d,J=7.2Hz,3H),1.17(t,J=7.2Hz,1H).

[0370] Step 5:

[0371] Under a nitrogen atmosphere, intermediate compound 28-5 (3.15 g, 23.8 mmol) was dissolved in THF, and isopropyl magnesium chloride (2.0 M, 23.8 mL) was added at 0 °C, and the reaction was carried out at 0 °C for 30 min. Then, intermediate compound 28-4 (4.79 g, 15.9 mmol) dissolved in THF (15.0 mL) was added at 0 °C, and the reaction was carried out at 25 °C for 5 h. After TLC monitoring showed the reaction was complete, the reaction was quenched by adding saturated ammonium chloride aqueous solution (400 mL) at 0 °C, stirred at room temperature for 30 min, extracted with ethyl acetate (200 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (100 mL × 3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 28-6 (2.52 g, 7.14 mmol, yield 44.9%) as a colorless oil. LCMS [M+H] + m / z:354.0.

[0372] Step Six:

[0373] Under a nitrogen atmosphere, NaH (570 mg, 14.3 mmol, 60% purity) was dissolved in DMF (20.0 mL). Intermediate compound 28-6 (2.52 g, 7.14 mmol) was added at 0 °C, and the mixture was stirred at 0 °C for 30 minutes. Then, compound 28-7 (2.18 g, 14.3 mmol, 1.35 mL) dissolved in DMF (10.0 mL) was slowly added dropwise to the reaction mixture, and the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until completion. The mixture was quenched at 0 °C with saturated ammonium chloride solution (100 mL), extracted with ethyl acetate (250 mL × 2), and the organic phases were combined. The mixture was dried over anhydrous sodium sulfate, concentrated under reduced pressure to remove the solvent, and the resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 28-8 (1.29 g, 3.04 mmol, yield 42.7%) as a colorless oil. LCMS [M+H] + m / z:425.9.

[0374] Step Seven:

[0375] Under a nitrogen atmosphere, intermediate compound 28-8 (890 mg, 2.09 mmol) was dissolved in DMF (5.0 mL), and lithium bromide (363 mg, 4.18 mmol, 105 μL) and water (75 mg, 4.18 mmol, 75 μL) were added. The mixture was stirred at 120 °C for 12 hours. The reaction was monitored by LCMS until completion. After cooling to room temperature, the solvent was removed by concentration under reduced pressure. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 28-9 (550 mg, 1.50 mmol, yield 71.7%) as a yellow oil. LCMS [M+H] + m / z:368.3.

[0376] Step 8:

[0377] TFA (30.0 mL) was added to intermediate compound 28-9 (2.78 g, 7.57 mmol), and the mixture was stirred at 70 °C for 12 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was subjected to high-performance liquid chromatography (HPLC) to prepare intermediate compound 28-10 (1.10 g, 4.45 mmol, yield 58.8%) as a white solid. LCMS [M+H] + m / z:247.9.

[0378] Step Nine:

[0379] Intermediate compound 28-10 (1.00 g, 4.04 mmol) was dissolved in DMF (10.0 mL), and NIS (1.82 g, 8.09 mmol) was added. The reaction was carried out at 25 °C for 2 hours. The reaction was monitored by LCMS until completion. The reaction was quenched with water (30.0 mL), extracted with ethyl acetate (30.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride solution (30.0 mL × 3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 28-11 (1.06 g, 2.84 mmol, yield 70.2%) as a yellow solid. LCMS [M+H] + m / z:373.9.

[0380] Step 10:

[0381] Intermediate compound 28-11 (1.10 g, 2.95 mmol) was dissolved in DMF (11.0 mL), and cesium carbonate (1.06 g, 3.24 mmol) and SEMCl (541 mg, 3.24 mmol, 574 μL) were added. The reaction was carried out at 25 °C for 2 hours. After TLC monitoring, the reaction was quenched with water (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride solution (10.0 mL × 3), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 28-12 (850 mg, 1.69 mmol, yield 57.3%) as a colorless oil. LCMS [M+H] + m / z:504.1.

[0382] Step Eleven:

[0383] Intermediate compound 28-12 (850 mg, 1.69 mmol) was dissolved in a mixed solution of THF (6.0 mL), methanol (2.0 mL), and water (2.0 mL). Lithium hydroxide monohydrate (708 mg, 16.9 mmol) was added, and the mixture was stirred at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The reaction solution was diluted with water (10.0 mL), and the pH was adjusted to 3–4 with 1.0 M hydrochloric acid. The mixture was then extracted with ethyl acetate (10.0 mL × 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was the crude intermediate compound 28-13 (766 mg), a colorless oil. LCMS [M+H] + m / z:490.1.

[0384] Step Twelve:

[0385] Compound 28-13 (200 mg, 408 μmol) was dissolved in DMF (3.0 mL), and HATU (233 mg, 613 μmol), DIEA (264 mg, 2.04 mmol, 355 μL), and compound 1-10 (255 mg, 1.63 mmol) were added. The mixture was stirred at 25 °C for 2 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 28-14 (200 mg, 318 μmol, yield 77.9%) as a brown solid. LCMS [M+H] + m / z:628.5.

[0386] Step Thirteen:

[0387] Intermediate compound 28-14 (200 mg, 318 μmol), Pd(dppf)Cl2 (47 mg, 63.7 μmol), and TEA (97 mg, 956 μmol, 133 μL) were dissolved in a mixed solvent of DMF (8.0 mL) and water (2.0 mL). After degassing, the mixture was purged once with CO, and then reacted at 80 °C for 12 hours under a CO atmosphere. The reaction was monitored by LCMS until completion. The solvent was removed by concentration under reduced pressure, and the resulting mixture was used to prepare intermediate compound 28-15 (90 mg, 165 μmol, yield 51.9%) as a brown solid by high performance liquid chromatography. 1 HNMR (400MHz, CDCl3) δ9.27 (s, 2H), 6.53 (s, 1H), 5.99 (d, J = 4.0Hz, 2H), 4.06-4.32 (m, 1H), 3.64-3.80 (m, 3H), 3.34 ( dd,J=16.4,7.5Hz,2H),2.96(dd,J=16.4,6.4Hz,2H),1.34-1.48(m,9H),0.87-1.03(m,3H),-0.03(s,9H).LCMS[M+H] + m / z:546.7.

[0388] Step Fourteen:

[0389] Compound 28-15 (70 mg, 128 μmol) was dissolved in DMF (5.0 mL), and HATU (73 mg, 192 μmol), DIEA (83 mg, 641 μmol, 111 μL), and methylamine hydrochloride (43 mg, 641 μmol) were added. The mixture was stirred at 25 °C for 2 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 28-16 (40 mg, 72 μmol, yield 56.0%) as a brown solid. LCMS [M+H] + m / z:559.1.

[0390] Step Fifteen:

[0391] Intermediate compound 28-16 (30 mg, 53.7 μmol) was dissolved in dichloromethane (3.0 mL), and TFA (1.0 mL) was added. The reaction was carried out at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure. The resulting mixture was then subjected to high performance liquid chromatography to prepare the final product compound WLE28 (16 mg, 37.3 μmol, yield 69.5%) as a white solid. 1 H NMR (400MHz, CD3OD) δ9.16-9.33(m,2H),6.60(s,1H),4.62(s,3H),4.17-4.27(m,1H),3.13(dd, J=16.0,9.1Hz,1H),2.74(dd,J=16.0,5.1Hz,1H),1.33(d,J=7.2Hz,3H),1.30(s,9H).LCMS[M+H] + m / z:429.3.

[0392] Example 28: Synthesis of compound WLE29

[0393]

[0394] Step 1:

[0395] At 0°C, sodium hydroxide (1.34 g, 33.5 mmol) was added to water (33.00 mL). After stirring for half an hour, 1-bromopinazone (2.00 g, 11.2 mmol, 1.50 mL) was slowly added dropwise to the reaction solution. After the addition was complete, the reaction was carried out at 0°C for 2 hours. The reaction was monitored by TLC. When the reaction was complete, the mixture was extracted with ethyl acetate (50 mL × 2). The ethyl acetate layer was washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove ethyl acetate, yielding a colorless liquid, which is the crude intermediate compound 29-5-1 (980 mg). 1 H NMR(400MHz,DMSO-d6)δ4.61(t,J=5.6Hz,1H),4.33(d,J=5.6Hz,2H),1.08(s,9H)

[0396] Step Two:

[0397] Under a nitrogen atmosphere, intermediate compound 29-5-1 (975 mg, 8.39 mmol), cyanamide (706 mg, 8.39 mmol, 706 μL), tetrabutylammonium hydroxide (0.4 M, 878 μL), sodium hydroxide (1.0 M, 353 μL), and sodium acetate (1.00 M, 2.0 mL) were dissolved in tetrahydrofuran (2.5 mL). The reaction was carried out at 25 °C for 48 hours under TLC monitoring. After the reaction was completed, water (10.0 mL) was added to the reaction solution, and the mixture was extracted with ethyl acetate (50.0 mL × 2). The extract was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove ethyl acetate. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 29-5 (176 mg, 1.26 mmol, yield: 14.9%).

[0398] Step 3:

[0399] Under a nitrogen atmosphere, compound 2-3 (5.00 g, 10.7 mmol) was dissolved in 1,4-dioxane (50.0 mL) and water (10.0 mL). Then, 4-pyridineboronic acid (2.62 g, 21.3 mmol), Pd(dppf)Cl2 (780 mg, 1.07 mmol), and sodium carbonate (3.68 g, 26.6 mmol) were added sequentially. After the additions were complete, the mixture was reacted at 90 °C for 12 hours. The reaction was monitored by TLC until completion. After the reaction solution cooled to room temperature, water (70.0 mL) was added to quench the reaction. The mixture was extracted with ethyl acetate (30.0 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 29-1 (3.60 g, 8.56 mmol, yield: 80.4%) as a yellow solid. LCMS (M+H) + m / z:421.2.

[0400] Step Four:

[0401] Intermediate compound 29-1 (2.3 g, 5.47 mmol) was dissolved in dichloromethane (10.0 mL), and a 1,4-dioxane solution of hydrochloric acid (4.0 M, 10.0 mL) was added. After the addition was complete, the reaction was carried out at 25 °C for 12 hours under TLC monitoring. Upon completion of the reaction, the organic solvent was removed by concentration under reduced pressure to obtain a yellow oily substance, which was the crude intermediate compound 29-2 (1.95 g). LCMS (M+H) + m / z:321.2.

[0402] Step 5:

[0403] Compound 1-7 (700 mg, 4.79 mmol) was dissolved in dichloromethane (10.0 mL). HATU (2.73 g, 7.18 mmol) and DIEA (3.10 g, 23.9 mmol, 4.2 mL) were added sequentially. After the addition was complete, the mixture was stirred at 25 °C for half an hour. Then, intermediate compound 29-2 (1.20 g, 3.35 mmol) was added. After the addition was complete, the mixture was reacted at 25 °C for 12 hours. The reaction was monitored by TLC until completion. The reaction was quenched by adding water (20.0 mL). The mixture was extracted with ethyl acetate (30.0 mL × 2), washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove ethyl acetate. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 29-3 (2.00 g, 4.45 mmol, yield: 93.0%) as a yellow solid. LCMS (M+H) + m / z:449.3.

[0404] Step Six:

[0405] Intermediate compound 29-3 (2.80 g, 6.24 mmol) was dissolved in methanol (20.0 mL), THF (10.0 mL), and water (10.0 mL). Lithium hydroxide monohydrate (1.31 g, 31.2 mmol) was added to the solution. After the addition was complete, the reaction was carried out at 25 °C for 2 hours. The reaction was monitored by TLC until it ended. After removing the organic phase by concentration under reduced pressure, water (10.0 mL) was added to the reaction flask, and the pH was adjusted to 5-6 with 1.0 M hydrochloric acid aqueous solution. After stirring for 10 minutes, the mixture was filtered to obtain a yellow solid, which was the crude intermediate compound 29-4 (1.16 g). 1H NMR(400MHz,DMSO-d6)δ12.01(brs,1H),8.60(d,J=6.0Hz,2H),7.53-7.66(m, 2H),7.19(d,J=8.4Hz,2H),6.91(d,J=8.4Hz,2H),5.27(brs,2H),4.79-4.92( m,1H),4.58(d,J=15.6Hz,1H),3.68-3.98(m,5H),3.17-3.28(m,1H),2.75-2. 85(m,1H),2.56-2.69(m,2H),2.17-2.35(m,1H),0.85-1.08(m,3H).LCMS(M+H) + m / z:435.2.

[0406] Step Seven:

[0407] Intermediate compound 29-4 (100 mg, 230 μmol) was dissolved in dichloromethane (10.0 mL). HATU (131 mg, 345 μmol) and DIEA (149 mg, 1.15 mmol, 200 μL) were added sequentially. After the additions were complete, the mixture was stirred at 25 °C for half an hour. Then, compound 1-10 (41 mg, 290 μmol) was added. After the additions were complete, the mixture was reacted at 25 °C for 12 hours. The reaction was monitored by TLC until completion. The reaction was quenched by adding water (20.0 mL). The mixture was extracted with ethyl acetate (10.0 mL × 2), washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove ethyl acetate. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 29-5 (20 mg, 35.9 μmol, yield: 15.6%) as a yellow solid. LCMS (M+H) + m / z:557.4.

[0408] Step 8:

[0409] Intermediate compound 29-5 (15 mg, 27.0 μmol) was dissolved in TFA (3 mg, 27.0 μmol, 2 μL) and reacted at 70 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, the organic phase was removed by vacuum concentration. The resulting mixture was then prepared by high performance liquid chromatography to obtain the final product compound WLE29 (1 mg, 2.84 μmol, yield: 10.5%). 1H NMR(400MHz,CD3OD)δ8.52-8.76(m,2H),7.62-7.79(m,2H),7.16-7.32(m,2H),4.59-4.66(m,2H),3.85 -4.14(m,2H),3.51-3.58(m,1H),2.88-3.19(m,3H),2.45-2.65(m,1H),1.09-1.35(m,12H).LCMS(M+H) + m / z:437.1.

[0410] Example 29: Synthesis of WLE30

[0411]

[0412] Step 1:

[0413] Compound 29-4 (200 mg, 460 μmol) was dissolved in dichloromethane (3.0 mL), and HATU (263 mg, 690 μmol) and DIEA (298 mg, 2.30 mmol, 401 μL) were added. The mixture was stirred at 25 °C for 30 min. Then, compound 30-1 (112 mg, 598 μmol) was added, and the mixture was stirred at 25 °C for 12 h. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 30-2 (120 mg, 212 μmol, yield 46.0%) as a yellow solid. LCMS [M+H] + m / z:567.3.

[0414] Step Two:

[0415] TFA (3.0 mL) was added to intermediate compound 30-2 (100 mg, 176 μmol), and the mixture was stirred at 70 °C for 12 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was prepared by high performance liquid chromatography to obtain the final product compound WLE30 (11 mg, 24.9 μmol, yield 14.1%), which was a white solid. 1H NMR (400MHz, CD3OD) δ8.58(brs,2H),7.50-7.84(m,4H),7.08(d,J=7.6Hz,1H),4.90-5.10(m,1H),4.67-4.79(m,1H),3.82-4.14(m ,2H),3.45-3.61(m,1H),3.04-3.17(m,1H),2.73-3.02(m,2H),2.45-2.68(m,1H),1.28-1.33(m,9H),1.13-1.26(m,3H).LCMS[M+H] + m / z:447.1.

[0416] Example 30: Synthesis of compound WLE31

[0417]

[0418] Step 1:

[0419] N-Boc-D-alanine (1.0 g, 5.29 mmol) was dissolved in DMF (10.0 mL), and HATU (3.01 g, 7.93 mmol), DIEA (3.42 g, 26.4 mmol, 4.60 mL), and 2-amino-4-tert-butylthiazole (826 mg, 5.29 mmol) were added sequentially. After the addition was complete, the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until it ended. Water (20.0 mL) was added to the reaction solution to quench the reaction, and the mixture was extracted with ethyl acetate (30.0 mL × 2). The organic phase was washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove ethyl acetate. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 31-1 (1.20 g, 3.66 mmol, yield: 69.3%) as a yellow solid. LCMS (M+H) + m / z:328.2.

[0420] Step Two:

[0421] Intermediate compound 31-1 (1.00 g, 3.05 mmol) was dissolved in a 1,4-dioxane solution of hydrochloric acid (4.0 M, 5.0 mL). The reaction was carried out at 25 °C for 2 hours under TLC monitoring. After the reaction was completed, the organic solvent was removed by concentration under reduced pressure to obtain a yellow oily substance, namely the crude intermediate compound 31-2 (680 mg). LCMS (M+H) + m / z:228.2.

[0422] Step 3:

[0423] At 0 °C, intermediate compound 31-2 (300 mg, 1.32 mmol) was dissolved in dichloromethane (10.0 mL). TEA (1267 mg, 2.64 mmol, 367 μL) and triphosgene (129 mg, 436 μmol) were added dropwise. After the addition was complete, the reaction was carried out at 0 °C for 2 hours. Then, compound 1-6 (793 mg, 2.64 mmol) was added. After the addition was complete, the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until it ended. Water (20.0 mL) was added to the reaction solution to quench the reaction. The mixture was extracted with ethyl acetate (10.0 mL × 2). The organic phase was washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove ethyl acetate. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 31-3 (280 mg, 506 μmol, yield: 58.3%) as a yellow solid. H NMR(400MHz,DMSO-d6)δ10.49(br s,1H),7.92-8.12(m,1H),7.14(dd,J=8.4,3.1Hz,2H),6.89(d,J=8.4Hz,2H),6.53(s,1H),5.22(s,2H),4 .66-4.84(m,2H),4.43-4.54(m,1H),3.70-4.06(m,5H),2.61-2.80(m,5H),1.19-1.25(m,12H).LCMS(M+H) + m / z:554.3.

[0424] Step Four:

[0425] Compound 31-3 (80 mg, 144 μmol) was dissolved in TFA (3.07 g, 26.9 mmol, 2.0 mL) and TfOH (170 mg, 1.13 mmol, 0.1 mL). The reaction was carried out at 25 °C for 12 hours under TLC monitoring. After the reaction was completed, the organic phase was removed by vacuum concentration. The resulting mixture was then prepared by high performance liquid chromatography to obtain the final product compound WLE31 (35 mg, 80.7 μmol, yield: 55.8%). 1 H NMR (400MHz, DMSO-d6) δ12.93-13.11(m,1H),10.45-10.79(m,1H),7.90-8.25(m,1H),6.85-7.30(m,1H) ),6.54(s,1H),4.48-4.85(m,3H),3.66-3.84(m,2H),2.66-2.81(m,5H),1.21-1.28(m,12H).LCMS(M+H) + m / z:434.2.

[0426] Example 31: Synthesis of compound WLE32

[0427]

[0428] Step 1:

[0429] At 0 °C, compound 1-methylcyclopropyl methyl ketone (2.00 g, 20.4 mmol) was dissolved in methanol (20.0 mL). Bromine (3.26 g, 20.4 mmol, 1.05 mL) was slowly added dropwise to the reaction solution. After the addition was complete, the reaction was carried out at 25 °C for 1 hour. When the reaction solution became a colorless and transparent solution, the reaction was terminated. Water (50.0 mL) was added to the reaction solution to quench the reaction. The mixture was extracted with petroleum ether (50 mL × 2). The organic phase was washed with saturated sodium sulfite aqueous solution (10.0 mL) and saturated sodium chloride aqueous solution (20.0 mL). The mixture was concentrated under reduced pressure to remove petroleum ether, yielding a yellow oily compound 32-1-1 (3.15 g, 17.3 mmol, yield: 87.3%).

[0430] Step Two:

[0431] Compound 32-1-1 (3.00 g, 17.0 mmol) and thiourea (1.29 g, 17.0 mmol) were dissolved in ethanol (50.0 mL) and reacted at 80 °C for 2 hours under TLC monitoring. After the reaction was completed, saturated sodium bicarbonate aqueous solution (50.0 mL) and water (200 mL) were added to the reaction solution. The mixture was extracted with ethyl acetate (200 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove organic solvent. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 32-1 (1.64 g, 10.6 mmol, yield: 62.7%).

[0432] Step 3:

[0433] Compound 1-9 (700 mg, 1.69 mmol) was dissolved in DMF (5.0 mL). HATU (963 mg, 2.53 mmol), DIEA (1.09 g, 8.44 mmol, 1.5 mL), and intermediate compound 32-1 (313 mg, 2.03 mmol) were added sequentially. After addition, the reaction was carried out at 25 °C for 2 hours. The reaction was monitored by TLC until completion. Water (10.0 mL) was added to quench the reaction mixture. Extraction was performed with ethyl acetate (10.0 mL × 2). The organic phase was washed with saturated sodium chloride aqueous solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove ethyl acetate. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 32-2 (210 mg, 381 μmol, yield: 22.6%) as a yellow solid. LCMS (M+H) + m / z:551.0.

[0434] Step Four:

[0435] Intermediate compound 32-2 (200 mg, 363 μmol) was dissolved in TFA (7.68 g, 67.3 mmol, 5.0 mL) and TfOH (848 mg, 5.65 mmol, 0.5 mL). The reaction was carried out at 25 °C for 2 hours under TLC monitoring. After the reaction was completed, the organic phase was removed by vacuum concentration. The resulting mixture was then prepared by high performance liquid chromatography to obtain the final product compound WLE32 (70 mg, 163 μmol, yield: 44.7%). 1 HNMR(400MHz,CD3OD)δ6.56(s,1H),4.92(brs,2H),4.52-4.65(m,1H),3.90-4.04(m,1H),3.73-3.87(m,1H),2.93-3.05(m,1H),2.82-2.9 2(m,4H),2.70-2.76(m,1H),2.53(dd,J=16.0,5.0Hz,1H),1.41(s,3H),1.11-1.22(m,3H),1.06(d,J=2.8Hz,2H),0.66(brs,2H)LCMS(M+H) + m / z:431.2.

[0436] Example 32: Synthesis of compound WLE33

[0437]

[0438] Step 1:

[0439] At 0 °C, compound WLE1 (30 mg, 69.4 μmol) was dissolved in DMF (1.0 mL), and 2,4,6-trimethylpyridine (8 mg, 69.4 μmol) and 1-chloromethyl-4-fluoro-1,4-diazabicyclo[2.2.2]octane di(tetrafluoroborate) salt (37 mg, 104 μmol) were added sequentially. After the addition was complete, the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until it ended. Water (10.0 mL) was added to the reaction solution to quench the reaction, and the mixture was extracted with ethyl acetate (10.0 mL × 2). The organic phase was washed with saturated sodium chloride aqueous solution, dried with anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove ethyl acetate. The resulting mixture was prepared by high performance liquid chromatography to obtain the final product compound WLE33 (2 mg, 4.44 μmol, yield: 6.40%). 1H NMR(400MHz,CD3OD)δ12.97(d,J=10.4Hz,1H),12.08(brs,1H),7.85-8.17(m,1H),4.40-4.77(m,2H),3.58-3.91(m,2H),2.7 6-2.92(m,2H),2.72(dd,J=8.0,4.6Hz,3H),2.61-2.68(m,1H),2.38-2.47(m,2H),1.28(s,9H),0.96-1.09(m,3H).LCMS(M+H) + m / z:451.2.

[0440] Example 33: Synthesis of compound WLE34:

[0441]

[0442] Step 1:

[0443] Compound 1-10 (500 mg, 3.20 mmol) was dissolved in acetonitrile (10.0 mL), and NCS (512 mg, 3.84 mmol) was added. The reaction was carried out at 25 °C for 1 hour. After the reaction was monitored by TLC, sodium sulfite solution (10.0 mL) was added to quench the reaction, and the mixture was extracted with ethyl acetate (10.0 mL × 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The obtained compound was purified by silica gel column chromatography to give intermediate compound 34-1 (600 mg, 3.16 mmol, yield 98.7%), a brown oily compound. LCMS [M+H] + m / z:191.1.

[0444] Step Two:

[0445] Compounds 1-9 (434 mg, 1.05 mmol) were dissolved in DMF (10.0 mL), and HATU (598 mg, 1.57 mmol) and DIEA (677 mg, 5.24 mmol, 913 μL) were added. The mixture was stirred at 25 °C for 2 hours. Then, compound 34-1 (200 mg, 1.05 mmol) was added, and the mixture was stirred at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 34-2 (60 mg, 104 μmol, yield 9.9%) as a yellow solid. LCMS [M+H] + m / z:587.1.

[0446] Step 3:

[0447] TFA (1.0 mL) and TfOH (0.1 mL) were added to intermediate compound 34-2 (59 mg, 100 μmol), and the mixture was stirred at 25 °C for 1 hour. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to give the final product compound WLE34 (20 mg, 42.9 μmol, yield 42.9%) as a white solid. 1 H NMR (400MHz, CD3OD) δ4.91-5.05(m,2H),3.77-4.06(m,2H),3.42-3.55(m,1H),2.94-3.09(m,1H),2.83-2.93 (m,4H),2.69-2.80(m,1H),2.53(dd,J=16.0,5.0Hz,1H),1.38(d,J=2.8Hz,9H),1.09-1.24(m,3H).LCMS[M+H] + m / z:467.2.

[0448] Example 34: Synthesis of compounds WLE35 and WLE36:

[0449]

[0450] Step 1:

[0451] Compound 2-7 (250 mg, 402 μmol), Pd(dppf)Cl2 (29 mg, 40.2 μmol), and TEA (122 mg, 1.21 mmol, 167 μL) were dissolved in methanol (10.0 mL) under a nitrogen atmosphere. After degassing, the mixture was reacted at 80 °C for 24 hours under a CO atmosphere. The reaction was monitored by TLC until complete. The solvent was removed by concentration under reduced pressure. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 35-1 (198 mg, 358 μmol, yield 89.1%) as a yellow solid. LCMS [M+H] + m / z:554.2.

[0452] Step Two:

[0453] Under a nitrogen atmosphere, intermediate compound 35-1 (198 mg, 358 μmol) was dissolved in THF (10.0 mL), and LAH (2.5 M, 140 μL) was added dropwise at 0 °C. The reaction was carried out at 0 °C for 13 hours. The reaction was monitored by TLC until complete. The reaction was quenched by adding water (0.40 mL), 15% sodium hydroxide (1.20 mL), and water (1.2 mL). The solvent was removed by concentration under reduced pressure. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 35-2 (100 mg, 190 μmol, yield 53.2%) as a yellow solid. LC-MS [M+H] + m / z: 526.3. An intermediate compound 36-2 (20.0 mg, 39.1 μmol, yield 10.9%) was also isolated as a yellow solid. LCMS [M+H] + m / z:512.3.

[0454] Step 3:

[0455] TFA (2.0 mL) and TfOH (0.2 mL) were added to intermediate compound 35-2 (50 mg, 95.1 μmol) and intermediate compound 36-2 (10.0 mg, 19.5 μmol), respectively, and the mixture was stirred at 70 °C for 8 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to give the final product compound WLE35 (5 mg, 12.3 μmol, yield 13.0%) as a white solid. 1 H NMR (400MHz, CD3OD) δ6.60 (d, J = 3.2Hz, 1H), 4.74-4.80 (m, 2H), 4.47-4.65 (m, 3H), 3.71-4.02 (m, 2H), 3.45-3.63 (m, 1H), 2.79-3.03(m,2H),2.68-2.74(m,1H),2.55(dt,J=15.6,4.8Hz,1H),1.29(d,J=2.8Hz,9H),1.14-1.24(m,3H).LCMS[M+H] + m / z: 406.2. The final product compound WLE36 (1.00 mg, 2.56 μmol, yield 13%) was further isolated and purified. 1HNMR(400MHz,CD3OD)δ6.57(s,1H),4.53(s,2H),3.49(td,J=3.2,1.6Hz,2H),2.86(br t,J=6.0Hz,1H),2.66-2.77(m,3H),2.38-2.57(m,4H),2.27-2.37(m,1H),1.26-1.29(m,9H),1.03(d,J=6.0Hz,3H)LCMS[M+H] + m / z:392.3.

[0456] Example 35: Synthesis of compound WLE37: Step 1:

[0457] Compound 35-1 (300 mg) was dissolved in a mixed solvent of THF (5.0 mL) and methanol (5.0 mL), and 2.0 M sodium hydroxide solution (4.0 mL) was added. The reaction was carried out at 25 °C for 3 hours. The reaction was monitored by LCMS until completion. The pH was adjusted to neutral with 1.0 M hydrochloric acid, and the solvent was removed by concentration under reduced pressure. The resulting mixture was the crude intermediate compound 37-1 (200 mg), a yellow oily compound. LCMS [M+H] + m / z:540.2.

[0458] Step Two:

[0459] TFA (2.0 mL) and TfOH (0.2 mL) were added to intermediate compound 37-1 (200 mg, 371 μmol). The mixture was stirred at 25 °C for 12 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was further analyzed by high-performance liquid chromatography (HPLC) to obtain intermediate compound 37-2 (100 mg, 238 μmol, yield 64.3%) as a white solid. LCMS [M+H] + m / z:420.2.

[0460] Step 3:

[0461] Intermediate compound 37-2 (30 mg, 71.5 μmol) was dissolved in DMF (1.0 mL), and T4P (155 mg, 215 μmol, 50% purity), DIEA (46 mg, 358 μmol, 62.3 μL), and ethylamine (2.0 M, 108 μL) were added. The reaction was carried out at 25 °C for 2 hours. After the reaction was monitored by LCMS, water (10.0 mL) and dichloromethane (10.0 mL) were added for extraction, the organic phase was separated, washed with water (10.0 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to remove the solvent. The resulting mixture was then subjected to high performance liquid chromatography to prepare the final product compound WLE37 (10 mg, 22.4 μmol, yield 31.4%) as a white solid. 1 H NMR (400MHz, DMSO-d6) δ12.78-13.15(m,1H),12.06(brs,1H),7.95-8.16(m,1H),6.68(d,J=4.4Hz,1H),4.42-4.75(m,2H),3.64-3 .84(m,2H),3.18-3.27(m,4H),2.76-2.87(m,2H),2.59-2.69(m,1H),2.37-2.48(m,1H),1.24(s,9H),0.98-1.11(m,6H).LCMS[M+H] + m / z:447.2.

[0462] Example 36: Synthesis of compound WLE38:

[0463]

[0464] Step 1:

[0465] Compound 37-2 (60 mg, 143 μmol) was dissolved in DMF (2.0 mL), and T4P (309 mg, 429 μmol, 50% purity), DIEA (92.4 mg, 715 μmol, 125 μL), and methyl-D3-amine (30 mg, 429 μmol) were added. The reaction was carried out at 25 °C for 2 hours. The reaction was monitored by LCMS until completion. Water (10.0 mL) and dichloromethane (10.0 mL) were added for extraction, the organic phase was separated, washed with water (10.0 mL), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to remove the solvent. The resulting mixture was subjected to high performance liquid chromatography to prepare the final product compound WLE38 (15.0 mg, 34.5 μmol, yield 24.1%) as a white solid. 1H NMR (400MHz, DMSO-d6) δ12.97(d,J=12.4Hz,1H),12.06(brs,1H),7.88-8.15(m,1H),6.68(d,J=4.0Hz,1H),4.44-4.77(m,2 H),3.60-3.86(m,2H),2.76-2.88(m,2H),2.60-2.68(m,1H),2.30-2.48(m,2H),1.24(s,9H),0.98-1.09(m,3H).LCMS[M+H] + m / z:436.2.

[0466] Example 37: Synthesis of compound WLE39:

[0467]

[0468] Step 1:

[0469] Compound 37-1 (80 mg, 148 μmol) was dissolved in DMF (3.0 mL), and HATU (85 mg, 222 μmol) and DIEA (96 mg, 741 μmol, 129 μL) were added. The mixture was stirred at 25 °C for 2 hours. Then, cyclopropylamine (17 mg, 296 μmol, 20.5 μL) was added, and the mixture was stirred at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 39-1 (80 mg, 138 μmol, yield 93.3%) as a white solid. LCMS [M+H] + m / z:579.1.

[0470] Step Two:

[0471] TFA (1.0 mL) and TfOH (0.1 mL) were added to intermediate compound 39-1 (80 mg, 138 μmol), and the mixture was stirred at 25 °C for 1 hour. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was subjected to high performance liquid chromatography to prepare the final product compound WLE39 (11 mg, 24.0 μmol, yield 17.4%) as a white solid. 1HNMR(400MHz,CD3OD)δ6.82(s,1H),4.98-4.59(m,1H),3.89-4.10(m,1H),3.73-3.86(m,1H),3.44-3.59(m,1H),2.92-3.1 6(m,2H),2.59-2.90(m,4H),1.34(d,J=2.0Hz,9H),1.15-1.27(m,3H),0.79(t,J=6.8Hz,2H),0.57-0.65(m,2H).LCMS[M+H] + m / z:459.2.

[0472] Example 38: Synthesis of compound WLE40:

[0473]

[0474] Step 1:

[0475] Compound 37-1 (80 mg, 148 μmol) was dissolved in DMF (5.0 mL), and HATU (85 mg, 222 μmol) and DIEA (96 mg, 741 μmol, 129 μL) were added. The mixture was stirred at 25 °C for 2 hours. Then, 3,3-difluorocyclobutylamine (32 mg, 296 μmol) was added, and the mixture was stirred at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 40-1 (80 mg, 127 μmol, yield 86.1%) as a white solid. LCMS [M+H] + m / z:629.3.

[0476] Step Two:

[0477] TFA (1.0 mL) and TfOH (0.1 mL) were added to intermediate compound 40-1 (70 mg, 111 μmol). The mixture was stirred at 25 °C for 1 hour. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was subjected to high performance liquid chromatography to prepare the final product compound WLE40 (15 mg, 29.5 μmol, yield 26.6%) as a white solid. 1HNMR(400MHz,CD3OD)δ6.58(s,1H),5.02-5.12(m,1H),4.53-4.62(m,2H),4.22-4.38(m,1H),3.76-4.08(m,2H),3.48(dd,J=6.4,3 .2Hz,1H),2.80-3.02(m,4H),2.64-2.78(m,3H),2.54(dd,J=16.0,5.1Hz,1H),1.27(d,J=2.8Hz,9H),1.12-1.23(m,3H).LCMS[M+H] + m / z:509.2.

[0478] Example 39: Synthesis of compound WLE41:

[0479]

[0480] Step 1:

[0481] Compound 37-1 (40 mg, 74.1 μmol) was dissolved in DMF (3.0 mL), and HATU (42 mg, 112 μmol) and DIEA (48 mg, 370 μmol, 65 μL) were added. The mixture was stirred at 25 °C for 2 hours. Then, 2,2-difluoroethylamine (6 mg, 74.1 μmol) was added, and the mixture was stirred at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 41-1 (15 mg, 24.9 μmol, yield 33.6%) as a white solid. LCMS [M+H] + m / z:603.3.

[0482] Step Two:

[0483] TFA (1.0 mL) and TfOH (0.1 mL) were added to intermediate compound 41-1 (15 mg, 24.9 μmol), and the mixture was stirred at 25 °C for 1 hour. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to give the final product compound WLE41 (3 mg, 6.22 μmol, yield 24.9%) as a white solid. 1H NMR(400MHz,CD3OD)δ6.80(s,1H),5.76-6.15(m,1H),4.96-5.04(m,1H),4.58(d,J=16.8Hz,1H),3.87-4.07(m,1H),3.63- 3.86(m,3H),3.43-3.58(m,1H),2.82-3.11(m,2H),2.58-2.77(m,2H),1.33(d,J=2.4Hz,9H),1.15-1.26(m,3H).LCMS[M+H] + m / z:483.2.

[0484] Example 40: Synthesis of compound WLE42:

[0485]

[0486] Step 1:

[0487] Compound 37-1 (70 mg, 129 μmol) was dissolved in DMF (3.0 mL), and HATU (74 mg, 194 μmol) and DIEA (84 mg, 648 μmol, 112 μL) were added. The mixture was stirred at 25 °C for 2 hours. Then, compound 3-aminotetrahydrofuran (23 mg, 259 μmol) was added, and the mixture was stirred at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 42-1 (60 mg, 98.7 μmol, yield 76.5%) as a yellow solid. LCMS [M+H] + m / z:609.2.

[0488] Step Two:

[0489] TFA (1.0 mL) and TfOH (0.1 mL) were added to intermediate compound 42-1 (50 mg, 82.1 μmol), and the mixture was stirred at 25 °C for 1 hour. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was subjected to high performance liquid chromatography to prepare the final product compound WLE42 (12 mg, 24.6 μmol, yield 30.0%) as a white solid. 1HNMR(400MHz,CD3OD)δ6.59(s,1H),4.59-4.61(m,2H),4.51-4.57(m,1H),4.3 3-4.44(m,1H),4.07-4.21(m,1H),3.81-4.06(m,2H),3.75(q,J=6.8Hz,2H),3. 44-3.55(m,1H),2.83-3.08(m,2H),2.71-2.79(m,1H),2.55(dt,J=16.0,4.7H z,1H),1.71-1.95(m,2H),1.28(d,J=2.8Hz,9H),1.13-1.24(m,3H).LCMS[M+H] + m / z:489.2.

[0490] Example 41: Synthesis of compound WLE43:

[0491]

[0492] Step 1:

[0493] Compounds 1-6 (500 mg, 1.48 mmol) were dissolved in DMF (5.0 mL), and potassium carbonate (410 mg, 2.97 mmol) and succinic anhydride (163 mg, 1.63 mmol) were added. The mixture was stirred at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was the crude intermediate compound 43-1 (600 mg), a yellow oily compound. LCMS [M+H] + m / z:401.2.

[0494] Step Two:

[0495] Intermediate compound 43-1 (500 mg, 1.25 mmol) and compound 1-10 (390 mg, 2.50 mmol) were dissolved in DMF (5.0 mL), and DIEA (807 mg, 6.24 mmol, 1.1 mL) and HATU (712 mg, 1.87 mmol) were added. The reaction was carried out at 25 °C for 2 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure to remove the solvent, and the resulting mixture was purified by silica gel column chromatography to give intermediate compound 43-2 (300 mg, 557 μmol, yield 44.6%), a yellow oily compound. LCMS [M+H] + m / z:539.2.

[0496] Step 3:

[0497] TFA (1.0 mL) and TfOH (0.1 mL) were added to intermediate compound 43-2 (60 mg, 111 μmol), and the reaction was carried out at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to obtain the final product compound WLE43 (35 mg, 83.7 μmol, yield 75.4%) as a white solid. 1 H NMR (400MHz, DMSO-d6) δ12.57-13.27(m,1H),12.04(s,1H),7.88-8.14(m,1H),6.68(s,1H),4.61(d,J=20.0Hz ,2H),3.68-3.75(m,2H),2.77(d,J=6.0Hz,2H),2.70-2.75(m,4H),2.62-2.68(m,3H),1.25(s,9H).LCMS[M+H] + m / z:419.2.

[0498] Example 42: Synthesis of compound WLE44:

[0499]

[0500] Step 1:

[0501] Compounds 44-1 (4.00 g, 22.3 mmol, 3.0 mL), 44-2 (10.7 g, 67.0 mmol), and sodium iodide (334 mg, 2.23 mmol) were dissolved in DMF (100 mL) under a nitrogen atmosphere and reacted at 25 °C for 96 hours. After the reaction was complete as monitored by TLC, the mixture was quenched with water (100 mL), extracted with ethyl acetate (100 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (100 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 44-3 (2.30 g, 9.58 mmol, 43.0%) as a white solid. 1 H NMR(400MHz, CDCl3)δ6.45(s,1H),5.86(brs,2H),1.59(s,9H),1.21(s,9H).LCMS[M+H] + m / z:240.1.

[0502] Step Two:

[0503] Intermediate compound 44-3 (500 mg, 2.09 mmol) was dissolved in dichloromethane (10.0 mL), and a 1,4-dioxane solution of hydrochloric acid (2.0 M, 5.0 mL) was added. The reaction was carried out at 25 °C for 8 hours. The reaction was monitored by LCMS until completion. The solvent was removed by concentration under reduced pressure, and the resulting mixture was the crude intermediate compound 44-4 (500 mg), a white solid. LCMS [M+H] + m / z:140.2.

[0504] Step 3:

[0505] Intermediate compound 44-4 (178 mg, 1.01 mmol) and compound 1-9 (280 mg, 675 μmol) were dissolved in DMF (10.0 mL), and HATU (385 mg, 1.01 mmol) and DIEA (436 mg, 3.38 mmol, 588 μL) were added. The mixture was stirred at 25 °C for 3 hours. The reaction was monitored by LCMS until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 44-5 (100 mg, 187 μmol, yield 27.7%) as a yellow solid. LCMS [M+H] + m / z:536.1.

[0506] Step Four:

[0507] TFA (3.0 mL) and TfOH (0.3 mL) were added to intermediate compound 44-5 (90 mg, 168 μmol), and the reaction was carried out at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to obtain the final product compound WLE44 (5 mg, 12.0 μmol, 7.2%) as a white solid. 1 H NMR (400MHz, DMSO-d6) δ12.82-13.13(m,1H),7.86-8.13(m,1H),6.76(d,J=3.6Hz,1H),6.49-6.69(m,1H),5.74(s,1H),4.64-4.81(m,2H),3. 56-3.63(m,2H),2.78-2.87(m,1H),2.69-2.75(m,4H),2.61-2.68(m,1 H),1.76-1.72(m,2H),1.17-1.20(m,3H),1.13-1.16(m,9H).LCMS[M+H] + m / z:416.3.

[0508] Example 43: Synthesis of compound WLE45:

[0509]

[0510] Step 1:

[0511] Compound 45-1 (106 mg, 483 μmol), compound 2-7 (100 mg, 160 μmol), Pd(PPh3)2Cl2 (23 mg, 32.2 μmol), and potassium carbonate (67 mg, 483 μmol) were dissolved in 1,4-dioxane (2.0 mL) and water (0.2 mL) under a nitrogen atmosphere and reacted at 90 °C for 12 h. After the reaction was complete as monitored by TLC, the reaction was quenched with water (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was then subjected to high performance liquid chromatography (HPLC) to prepare intermediate compound 45-2 (70 mg, 119 μmol, yield 74.2%) as a yellow solid. LCMS [M+H] + m / z:587.1.

[0512] Step Two:

[0513] TFA (2.0 mL) and TfOH (0.2 mL) were added to intermediate compound 45-2 (70 mg, 119 μmol), and the reaction was carried out at 40 °C for 5 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to obtain the final product compound WLE45 (30 mg, 64.3 μmol, 54.1%) as a white solid. 1 H NMR (400MHz, DMSO-d6) δ12.84-13.60(m,1H),12.06(s,1H),8.49(d,J=5.2Hz,1H),7.33-7.64(m,2H),6.67(s,1H),4.62-4.87(m,2H),3.76-3.93 (m,2H),3.60-3.70(m,1H),2.77-2.89(m,2H),2.66(d,J=4.4Hz,1H),2. 52(s,3H),2.39-2.47(m,1H),1.23(s,9H),0.96-1.11(m,3H).LCMS[M+H] + m / z:467.2.

[0514] Example 44: Synthesis of compound WLE46:

[0515]

[0516] Step 1:

[0517] Under a nitrogen atmosphere, compound 46-1 (171 mg, 483 μmol), compound 2-7 (100 mg, 160 μmol), Pd(PPh3)2Cl2 (22.6 mg, 32.2 μmol), and potassium carbonate (66.7 mg, 483 μmol) were dissolved in 1,4-dioxane (2.0 mL) and water (0.2 mL), and reacted at 90 °C for 12 h. After the reaction was complete as monitored by TLC, the reaction was quenched with water (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was then subjected to high-performance liquid chromatography (HPLC) to prepare intermediate compound 46-2 (80 mg, 100 μmol, yield 61.8%), which was a yellow oily compound. LCMS [M+H] + m / z:804.2.

[0518] Step Two:

[0519] TFA (2.0 mL) and TfOH (0.2 mL) were added to intermediate compound 46-2 (80 mg, 100 μmol). The reaction was carried out at 40 °C for 5 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to obtain the final product compound WLE46 (10 mg, 22.7 μmol, yield 22.7%) as a white solid. 1 H NMR (400MHz, DMSO-d6) δ12.05(brs,1H),7.87(brs,1H),7.31-7.49(m,1H),6.67(d,J=3.6Hz,1H),4.48-4.79(m,2H),3.75-3.88( m,2H),3.59-3.64(m,1H),2.76-2.87(m,2H),2.60(t,J=5.2Hz,1H),2.40-2.47(m,1H),1.24(s,9H),0.98-1.10(m,3H).LCMS[M+H] + m / z:442.2.

[0520] Example 45: Synthesis of compound WLE47:

[0521]

[0522] Synthesis of compound 47-2:

[0523] Compound 47-2A (3.00 g, 11.4 mmol) was dissolved in dichloromethane (30.0 mL), and oxalyl chloride (1.74 g, 13.7 mmol, 1.2 mL) and DMF (33.3 mg, 456 μmol, 35 μL) were added. The reaction was carried out at 25 °C for 1 hour. The reaction was monitored by LCMS until completion. The solvent was removed by concentration under reduced pressure, and the resulting mixture was the crude intermediate compound 47-2 (3.50 g), a yellow oily compound. LCMS: [M-Cl + OMe + H] + m / z:278.2.

[0524] Synthesis of compound WLE47

[0525] Step 1:

[0526] Under a nitrogen atmosphere, compound 47-1 (2.80 g, 11.1 mmol) was dissolved in dichloromethane (30.0 mL), and TEA (1.68 g, 16.6 mmol, 2.3 mL) was added. After cooling to 0 °C, intermediate compound 47-2 (3.13 g, 11.1 mmol) was added dropwise. After the addition was complete, the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until completion. The pH was adjusted to 3 with 1.0 M hydrochloric acid, and the mixture was stirred at 25 °C for 30 minutes. The organic phase was separated, and the aqueous phase was extracted with ethyl acetate (50.0 mL × 3). The combined organic phases were washed with saturated sodium chloride solution (50.0 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 47-3 (1.50 g, 3.37 mmol, yield 30.4%), a yellow oily compound. LCMS [M + Na] + m / z:467.3.

[0527] Step Two:

[0528] Intermediate compound 47-3 (1.60 g, 3.60 mmol) was dissolved in acetic acid (20.0 mL), and 4-methoxybenzylhydrazine hydrochloride (548 mg, 3.60 mmol) was added. The reaction was carried out at 80 °C for 12 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to give intermediate compound 47-4 (800 mg, 1.74 mmol, yield 48.26%), a yellow oily compound. LCMS [M+H] + m / z:461.2.

[0529] Step 3:

[0530] Compound 1-7 (508 mg, 3.47 mmol) was dissolved in DMF (5.0 mL), and HATU (991 mg, 2.61 mmol) and DIEA (1.12 g, 8.68 mmol, 1.51 mL) were added. The mixture was stirred at 25 °C for 1 hour. Then, intermediate compound 47-4 (800 mg, 1.74 mmol) was added, and the mixture was stirred at 25 °C for 12 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (20.0 mL), extracted with ethyl acetate (20.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (20.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 47-5 (410 mg, 696 μmol, yield 40.1%), which was a yellow oily compound. LCMS [M+H] + m / z:589.4.

[0531] Step Four:

[0532] Intermediate compound 47-5 (410 mg, 696 μmol) was dissolved in a mixed solution of THF (3.0 mL), methanol (1.0 mL), and water (1.0 mL). Lithium hydroxide monohydrate (292 mg, 6.96 mmol) was added, and the mixture was stirred at 25 °C for 12 hours. The reaction was monitored by LCMS until complete. The reaction solution was diluted with water (5.0 mL), and the pH was adjusted to 2–3 with 1.0 M hydrochloric acid. The solution was then extracted with ethyl acetate (10.0 mL × 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was the crude intermediate compound 47-6 (410 mg), a yellow solid. LCMS [M+H] + m / z:575.1.

[0533] Step 5:

[0534] Intermediate compound 47-6 (400 mg, 696 μmol) and compound 1-10 (218 mg, 1.39 mmol) were dissolved in DMF (5.00 mL), and DIEA (450 mg, 3.48 mmol, 606 μL) and HATU (397 mg, 1.04 mmol) were added. The mixture was stirred at 25 °C for 2 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 47-7 (210 mg, 295 μmol, yield 42.3%) as a yellow solid. LCMS [M+H] + m / z:713.2.

[0535] Step Six:

[0536] TFA (2.0 mL) and TfOH (0.2 mL) were added to intermediate compound 47-7 (100 mg, 140 μmol). The reaction was carried out at 65 °C for 12 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to obtain the final product compound WLE47 (19.0 mg, 40.3 μmol, yield 28.8%) as a white solid. 1H NMR(400MHz,DMSO-d6)δ12.06(brs,1H),7.49-7.65(m,1H),6.68(s,1H),4.46-4.70(m,2H),4.32-4.45(m,1H),3.19-3.32(m,4H),3.09-3.16(m ,1H),3.01-3.08(m,1H),2.73-2.84(m,2H),2.37-2.48(m,2H),2.23-2. 33(m,2H),1.84-1.98(m,2H),1.24(s,9H),0.97-1.08(m,3H).LCMS[M+H] + m / z:473.2.

[0537] Example 46: Synthesis of compound WLE48:

[0538]

[0539] Synthesis of Compound 48-2

[0540] Step 1:

[0541] Compound 48-2-1 (10.0 g, 63.2 mmol, 8.9 mL) was dissolved in methanol (100 mL), and 4-methoxybenzylamine (8.67 g, 63.2 mmol, 8.2 mL) was added. The mixture was stirred at 25 °C for 13 hours, then reacted at 80 °C for 4 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, yielding the crude intermediate 48-2-2 (18.0 g), a colorless oily compound. LCMS [M+H] + m / z:263.9.

[0542] Step Two:

[0543] Compound 48-2-2 (16.6 g, 63.1 mmol) was dissolved in 1,4-dioxane (330 mL), and sodium hydroxide aqueous solution (2.5 M, 75.7 mL) was added at 0 °C. The reaction was carried out at 80 °C for 12 hours. The reaction was monitored by TLC until complete. The solvent was removed by concentration under reduced pressure. The pH was adjusted to 2–3 with 1.0 M potassium hydrogen sulfate, diluted with dichloromethane (200 mL), and extracted with water (200 mL × 2). The solvent was removed by concentration under reduced pressure. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 48-2-3 (13.0 g, 52.2 mmol, yield 82.7%) as a yellow solid. LCMS [M+H] + m / z:250.0.

[0544] Step 3:

[0545] Intermediate compound 48-2-3 (3.00 g, 12.0 mmol) was dissolved in dichloromethane (30.0 mL), and oxalyl chloride (1.83 g, 14.4 mmol, 1.26 mL) and DMF (35.2 mg, 481 μmol, 37 μL) were added. The reaction was carried out at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was the crude intermediate compound 48-2 (3.50 g), a yellow oily compound. LCMS [M-Cl+MeO] + m / z:264.2.

[0546] Synthesis of compound WLE48

[0547] Step 1:

[0548] Under a nitrogen atmosphere, compound 48-1 (3.00 g, 11.9 mmol) was dissolved in dichloromethane (30.0 mL), and TEA (1.80 g, 17.8 mmol, 2.5 mL) was added. Then, intermediate compound 48-2 (3.18 g, 11.9 mmol) was added dropwise at 0 °C, and the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until complete. The pH was adjusted to 3 with 1.0 M hydrochloric acid, and the mixture was stirred at 25 °C for 30 minutes. The dichloromethane layer was separated, and the aqueous phase was extracted with ethyl acetate (50.0 mL), washed with sodium chloride aqueous solution (20.0 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 48-3 (950 mg, 2.21 mmol, yield 18.6%), a yellow oily compound. LCMS [M-56+H] + m / z:375.0.

[0549] Step Two:

[0550] Intermediate compound 48-3 (950 mg, 2.21 mmol) was dissolved in acetic acid (10.0 mL), and 4-methoxybenzylhydrazine hydrochloride (335 mg, 2.21 mmol) was added. The mixture was stirred at 80 °C for 12 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to give intermediate compound 48-4 (890 mg, 1.99 mmol, yield 90.3%), a yellow oily compound. LCMS [M+H] + m / z:447.1.

[0551] Step 3:

[0552] Compounds 1-7 (583 mg, 3.99 mmol) were dissolved in DMF (10.0 mL), and HATU (1.14 g, 2.99 mmol) and DIEA (1.29 g, 9.97 mmol, 1.7 mL) were added. The mixture was stirred at 25 °C for 30 min. Then, intermediate compound 48-4 (890 mg, 1.99 mmol) was added, and the mixture was stirred at 25 °C for 12 h. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 48-5 (800 mg, 1.39 mmol, yield 69.8%), a yellow oily compound. LCMS [M+H] + m / z:575.2.

[0553] Step Four:

[0554] Intermediate compound 48-5 (800 mg, 1.39 mmol) was dissolved in a mixed solution of THF (6.0 mL), methanol (2.0 mL), and water (2.0 mL). Lithium hydroxide monohydrate (584 mg, 13.9 mmol) was added, and the mixture was stirred at 25 °C for 12 hours. The reaction was monitored by LC-MS until complete. The solvent was removed by concentration under reduced pressure. The reaction solution was diluted with water (20.0 mL), extracted with dichloromethane (20.0 mL × 3), and the organic phases were combined. The mixture was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was intermediate compound 48-6 (760 mg, 1.36 mmol, yield 97.6%), a yellow solid. LC-MS [M+H] + m / z:561.2.

[0555] Step 5:

[0556] Intermediate compound 48-6 (360 mg, 642 μmol) was dissolved in DMF (4.0 mL), and HATU (293 mg, 771 μmol), DIEA (415 mg, 3.21 mmol, 559 μL), and compound 1-10 (200 mg, 1.28 mmol) were added. The mixture was stirred at 25 °C for 12 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 48-7 (300 mg, 429 μmol, yield 66.9%) as a yellow solid. LCMS [M+H] +m / z:699.2.

[0557] Step Six:

[0558] TFA (2.0 mL) and TfOH (0.2 mL) were added to intermediate compound 48-7 (100 mg, 143 μmol), and the mixture was stirred at 60 °C for 12 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to give the final product compound WLE48 (17 mg, 36.5 μmol, yield 25.5%) as a white solid. 1 H NMR (400MHz, CDCl3) δ6.49(s,1H),4.46-4.73(m,2H),3.79-3.93(m,2H),3.69-3.79(m,2H),3.43-3.65(m ,2H),2.90-3.12(m,2H),2.65-2.88(m,3H),2.52-2.64(m,2H),1.29(s,9H),1.16-1.27(m,3H).LCMS[M+H] + m / z:459.3.

[0559] Example 47: Synthesis of compound WLE49:

[0560]

[0561] Step 1:

[0562] Compound 49-1 (10.0 g, 50.2 mmol) was dissolved in toluene (100 mL), and tetrahydropyrrole (3.57 g, 50.2 mmol, 4.2 mL) and p-toluenesulfonic acid (43.2 mg, 251 μmol) were added. The reaction was carried out at 140 °C for 20 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was the crude intermediate compound 49-2 (13.0 g), which was a yellow oily compound. 1 H NMR(400MHz,DMSO-d6)δ4.02(brs,1H),3.83(brs,2H),3.41(t,J=5.6Hz,2H),2.95 (t,J=6.4Hz,4H),2.19(t,J=5.6Hz,2H),1.78(dt,J=6.4,3.3Hz,4H),1.39(s,9H).

[0563] Step 2: Synthesis of intermediate compound 49-3

[0564] Under a nitrogen atmosphere, compound 49-3-1 (2.00 g, 17.7 mmol), oxalyl chloride (2.69 g, 21.2 mmol, 1.86 mL), and DMF (38.8 mg, 531 μmol, 40.8 μL) were dissolved in 1,4-dioxane (20.0 mL) and reacted at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, yielding the crude intermediate compound 49-3 (2.50 g), which was a yellow oily compound. LCMS [M-Cl+CH3O] + m / z:128.0.

[0565] Step 3:

[0566] Under a nitrogen atmosphere, intermediates 49-2 (3.50 g, 13.9 mmol), 49-3 (2.19 g, 16.6 mmol), and DIEA (2.15 g, 16.6 mmol, 2.90 mL) were dissolved in 1,4-dioxane (15.0 mL) and reacted at 25 °C for 16 h. Then, water (15.0 mL) was added dropwise, and the reaction was continued at 100 °C for 1 h. The reaction was monitored by TLC until completion. After cooling to room temperature, the reaction was quenched with water (30.0 mL), extracted with ethyl acetate (30.0 mL × 3), and the organic phases were combined. The mixture was washed with a saturated aqueous solution of sodium chloride (30.0 mL), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate 49-4 (700 mg, 2.21 mmol, yield 15.9%) as a yellow solid. LCMS [M-56+H] + m / z:238.9.

[0567] Step Four:

[0568] Intermediate compound 49-4 (600 mg, 2.04 mmol) was dissolved in acetic acid (6.0 mL), and 4-methoxybenzylhydrazine hydrochloride (310 mg, 2.04 mmol) was added. The mixture was stirred at 80 °C for 2 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was the crude intermediate compound 49-5 (850 mg), a colorless oily compound. LCMS [M-100+H] + m / z:311.3.

[0569] Step 5:

[0570] Intermediate compound 49-5 (800 mg, 1.95 mmol) was added to a 1,4-dioxane solution (2.0 M, 8.0 mL) in hydrochloric acid and stirred at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to give intermediate compound 49-6 (350 mg, 1.13 mmol, yield 57.9%) as a yellow solid. LCMS [M+H] + m / z:310.9.

[0571] Step Six:

[0572] Compound 1-7 (329 mg, 2.26 mmol) was dissolved in DMF (4.0 mL), and HATU (472 mg, 1.24 mmol) and DIEA (729 mg, 5.64 mmol, 982 μL) were added. The mixture was stirred at 25 °C for 30 min. Then, intermediate compound 49-6 (350 mg, 1.13 mmol) was added, and the mixture was stirred at 25 °C for 2 h. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 49-7 (330 mg, 753 μmol, yield 66.7%), which was a yellow oily compound. 1 H NMR(400MHz,DMSO-d6)δ8.51-8.66(m,1H),8.34-8.48(m,1H),7.03-7.18(m,2H) ,6.80-6.94(m,2H),5.18-5.57(m,2H),4.35-4.77(m,2H),3.73-3.96(m,2H),3.6 8-3.72(m,3H),3.43-3.52(m,3H),3.27(dd,J=6.4,2.4Hz,2H),2.75-2.84(m,1H ),2.57-2.63(m,1H),2.36(dt,J=16.8,4.8Hz,1H),0.93-1.09(m,3H).LCMS[M+H] + m / z:439.0.

[0573] Step Seven:

[0574] Intermediate compound 49-7 (350 mg, 798 μmol) was dissolved in a mixed solution of THF (3.0 mL), methanol (1.0 mL), and water (1.0 mL). Lithium hydroxide monohydrate (335 mg, 7.98 mmol) was added, and the mixture was stirred at 25 °C for 12 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the reaction was diluted with water (10.0 mL). Extraction was performed with dichloromethane (10.0 mL × 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was the crude intermediate compound 49-8 (230 mg), a yellow oily compound. LCMS [M+H] + m / z:425.2.

[0575] Step 8:

[0576] Intermediate compound 49-8 (180 mg, 424 μmol) and compound 1-10 (199 mg, 1.27 mmol) were dissolved in DMF (2.0 mL), and DIEA (274 mg, 2.12 mmol, 369 μL) and HATU (194 mg, 509 μmol) were added. The mixture was stirred at 25 °C for 2 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 49-9 (120 mg, 213 μmol, yield 50.3%) as a yellow solid. LCMS [M+H] + m / z:563.2.

[0577] Step Nine:

[0578] TFA (2.0 mL) and TfOH (159 mg, 1.06 mmol, 94 μL) were added to intermediate compound 49-9 (150 mg, 267 μmol), and the mixture was stirred at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to give the final product compound WLE49 (15 mg, 33.9 μmol, yield 12.7%) as a white solid. 1H NMR(400MHz,DMSO-d6)δ12.65-13.10(m,1H),12.06(s,1H),8.24-8.64(m,2H),6.64-6.74(m,1H),4.50-4.79(m,2H),3.60-3.90 (m,2H),3.35-3.43(m,1H),2.70-2.88(m,2H),2.58-2.68(m,1H),2.41-2.47(m,1H),1.24(s,9H),0.98-1.10(m,3H).LCMS[M+H] + m / z:443.2.

[0579] Example 48: Synthesis of compound WLE50:

[0580]

[0581] Step 1:

[0582] Under a nitrogen atmosphere, N-tert-butoxycarbonyl-1,2-ethylenediamine (77 mg, 483 μmol, 76.1 μL), compound 2-7 (100 mg, 160 μmol), potassium carbonate (45 mg, 321 μmol), CuI (6 mg, 32.1 μmol), and L-proline were dissolved in DMF (5.0 mL) and reacted at 80 °C for 12 h. The reaction was monitored by TLC until complete. The reaction was quenched with water (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was then subjected to high-performance liquid chromatography (HPLC) to prepare intermediate compound 50-1 (100 mg, 152 μmol, yield 95.1%) as a brown solid. LCMS [M+H] + m / z:654.3.

[0583] Step Two:

[0584] TFA (3.0 mL) and TfOH (508 mg, 3.39 mmol, 0.30 mL) were added to intermediate compound 50-1 (90 mg, 137 μmol), and the mixture was stirred at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was used to prepare WLE50 (3 mg, 6.91 μmol, yield 5.05%) as a white solid by high performance liquid chromatography. 1H NMR(400MHz,CD3OD)δ6.60(d,J=7.2Hz,1H),4.49-4.57(m,2H),3.70-4.00(m,2H),3.37-3.57(m,3H),3.0 6-3.21(m,2H),2.86-2.99(m,2H),2.50-2.79(m,2H),1.28(d,J=2.4Hz,9H),1.13-1.23(m,3H).LCMS[M+H] + m / z:434.2.

[0585] Example 49: Synthesis of compound WLE51:

[0586]

[0587] Step 1:

[0588] Compound 37-1 (100 mg, 185.3 μmol) was dissolved in DMF (2.0 mL), and HATU (106 mg, 278 μmol), DIEA (120 mg, 927 μmol, 161 μL), and ammonium chloride (50 mg, 927 μmol) were added. The mixture was stirred at 25 °C for 12 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 51-1 (80 mg, 149 μmol, yield 80.2%) as a yellow solid. LCMS [M+H] + m / z:539.1.

[0589] Step Two:

[0590] TFA (1.0 mL) and TfOH (0.1 mL) were added to intermediate compound 51-1 (80 mg, 149 μmol), and the mixture was stirred at 40 °C for 2 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to obtain the final product compound WLE51 (40 mg, 95.58 μmol, yield 64.3%) as a white solid. 1H NMR (400MHz, DMSO-d6) δ12.97(d,J=12.8Hz,1H),12.07(s,1H),7.07-7.51(m,2H),6.68(d,J=4.0Hz,1H),4.41-4.75(m,2H),3.59 -3.86(m,2H),2.76-2.87(m,2H),2.61-2.68(m,1H),2.52(brs,1H),2.41-2.47(m,1H),1.24(s,9H),0.98-1.08(m,3H).LCMS[M+H] + m / z:419.2.

[0591] Example 50: Synthesis of compound WLE52:

[0592]

[0593] Step 1:

[0594] Compound 52-1 (76 mg, 320 μmol), compound 2-7 (100 mg, 160 μmol), cataCXium A-Pd-G2 (16 mg, 24.0 μmol), silver carbonate (7 mg, 24.0 μmol), and cesium carbonate (75 mg, 480 μmol) were dissolved in 1,4-dioxane (5.0 mL) and water (1.0 mL) under a nitrogen atmosphere and reacted at 100 °C for 12 h. After the reaction was complete as monitored by TLC, the reaction was quenched with water (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), the organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was then subjected to high-performance liquid chromatography (HPLC) to prepare intermediate compound 52-2 (50 mg, 80 μmol, yield 50.0%), a colorless oily compound. LCMS [M+H] + m / z:625.2.

[0595] Step Two:

[0596] TFA (1.0 mL) and TfOH (0.10 mL) were added to intermediate compound 52-2 (50 mg, 80 μmol), and the reaction was carried out at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to obtain the final product compound WLE52 (22 mg, 54.4 μmol, yield 68.1%) as a white solid. 1H NMR (400MHz, DMSO-d6) δ8.33 (s, 1H), 6.68 (d, J = 3.0Hz, 1H), 4.32-4.63 (m, 2H), 3.81-3.84 (brs, 1H), 3.61-3.78 (m, 2H), 3.35-3.37(m,2H),2.80-2.84(m,2H),2.65-2.68(m,1H),2.40-2.56(m,2H),1.25(s,9H),0.98-1.08(m,3H).LCMS[M+H] + m / z:405.2.

[0597] Example 51: Synthesis of compound WLE53:

[0598]

[0599] Step 1:

[0600] Diisopropylaminolithium (2.0 M, 2.58 mL) was added to THF (5.0 mL), and the mixture was cooled to -70 °C. A THF (5.0 mL) solution of compound 53-1 (1.00 g, 4.69 mmol) was slowly added. After stirring the reaction mixture at -70 °C for 1 hour, a THF (2.0 mL) solution of compound 1-2 (754 mg, 5.16 mmol, 704 μL) was added. After the addition was complete, the temperature was gradually increased to 25 °C, and the reaction was allowed to proceed for 12 hours. The reaction was monitored by LCMS until complete. The mixture was quenched at 0 °C with saturated ammonium chloride aqueous solution (20.0 mL). The mixture was stirred at 25 °C for 30 minutes. The pH was adjusted to 5–6 with 1.0 M hydrochloric acid. The mixture was extracted with ethyl acetate (20.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (20.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent, yielding a mixture, i.e., crude intermediate compound 53-2 (1.5 g), which was a yellow oily compound. LCMS[M-56+H] + m / z:257.9.

[0601] Step Two:

[0602] Intermediate compound 53-2 (1.47 g, 4.69 mmol) was dissolved in ethanol (17.0 mL), and 4-methoxybenzylhydrazine hydrochloride (750 mg, 4.93 mmol) and acetic acid (338 mg, 5.63 mmol, 322 μL) were added. The mixture was stirred at 90 °C for 3 hours. The reaction was monitored by TLC until complete. The solvent was removed by concentration under reduced pressure. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 53-3 (600 mg, 1.40 mmol, yield 29.8%) as a yellow solid. LCMS [M+H] + m / z:430.2.

[0603] Step 3:

[0604] MeNH₂ / EtOH (5.78 g, 55.9 mmol, 6.0 mL) was added to compound 53-3 (600 mg, 1.40 mmol), and the mixture was stirred at 90 °C for 12 hours in an autoclave. The reaction was monitored by LCMS until complete, and the solvent was removed by concentration under reduced pressure. The resulting mixture, intermediate compound 53-4 (570 mg, 1.38 mmol, 98.4%), was a yellow oily compound. LCMS [M+H] + m / z:415.0.

[0605] Step Four:

[0606] In intermediate compound 53-4 (570 mg, 1.38 mmol), a solution of 1,4-dioxane hydrochloric acid (2.0 M, 6.0 mL) was added, and the mixture was stirred at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was the crude intermediate compound 53-5 (500 mg), a yellow oily compound. LCMS [M+H] + m / z:315.2.

[0607] Step 5:

[0608] Compounds 1-7 (400 mg, 2.74 mmol) were dissolved in DMF (5.0 mL), and HATU (780 mg, 2.05 mmol) and DIEA (884 mg, 6.84 mmol, 1.2 mL) were added. The mixture was stirred at 25 °C for 1 hour. Then, intermediate compound 53-5 (430 mg, 1.37 mmol) was added, and the mixture was stirred at 25 °C for 12 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (20.0 mL), extracted with ethyl acetate (20.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (20.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 53-6 (550 mg, 1.24 mmol, yield 90.5%), a yellow oily compound. LCMS [M+H] + m / z:443.1.

[0609] Step Six:

[0610] Intermediate compound 53-6 (550 mg, 1.24 mmol) was dissolved in a mixed solution of THF (3.0 mL), methanol (1.0 mL), and water (1.0 mL). Lithium hydroxide monohydrate (522 mg, 12.4 mmol) was added, and the mixture was stirred at 25 °C for 12 hours. The reaction was monitored by LCMS until complete. The reaction solution was diluted with water (5.0 mL), and the pH was adjusted to 2–3 with 1.0 M hydrochloric acid. The solution was then extracted with ethyl acetate (10.0 mL × 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was the crude intermediate compound 53-7 (550 mg), a yellow solid. LCMS [M+H] + m / z:429.3.

[0611] Step Seven:

[0612] Intermediate compound 53-7 (500 mg, 1.17 mmol) and compound 1-10 (364 mg, 2.33 mmol) were dissolved in DMF (5.0 mL), and DIEA (754 mg, 5.83 mmol, 1.0 mL) and HATU (665 mg, 1.75 mmol) were added. The mixture was stirred at 25 °C for 12 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure to remove the solvent, and the resulting mixture was purified by silica gel column chromatography to give intermediate compound 53-8 (350 mg, 618 μmol, yield 52.9%) as a yellow solid. LCMS [M+H] + m / z:567.1.

[0613] Step 8:

[0614] TFA (4.0 mL) was added to intermediate compound 53-8 (350 mg, 618 μmol), and the reaction was carried out at 65 °C for 12 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to obtain the final product compound WLE53 (70 mg, 152 μmol, yield 24.6%) as a white solid. 1H NMR(400MHz,DMSO-d6)δ12.98(d,J=17.6Hz,1H),11.90-12.19(m,1H),7.92-8.17(m, 1H),6.67(d,J=13.6Hz,1H),4.64-5.42(m,2H),3.71-4.48(m,1H),3.19-3.29(m,1H) ,2.79-3.05(m,2H),2.72(dd,J=10.8,4.4Hz,3H),2.55-2.66(m,1H),2.39-2.48(m,1 H),1.24(d,J=7.6Hz,9H),1.07-1.17(m,3H),0.96(dd,J=6.8,3.2Hz,3H).LCMS[M+H] + m / z:447.2.

[0615] Example 52: Synthesis of compound WLE54

[0616]

[0617] By replacing compound 53-1 with compound 54-1 and following the same synthetic method as compound WLE53, compound WLE54 was prepared, which is a white solid. 1 H NMR (400MHz, DMSO-d6) δ13.06(d,J=11.0Hz,1H),12.05(s,1H),7.96-8.08(m,1H),6.67(d,J=3.5Hz,1H),4.20-4.86(m,2H),3.35-3.5 0(m,3H),2.68-2.79(m,1H),2.41-2.50(m,4H),1.30-1.32(m,3H),1.23-1.25(m,9H),1.14-1.17(m,3H),1.01-1.09(m,3H).LCMS[M+H] + m / z:461.2.

[0618] Example 53: Synthesis of compound WLE55

[0619]

[0620] Step 1:

[0621] Diisopropylaminolithium (2.0 M, 9.9 mL) was added to THF (30.0 mL), and the mixture was cooled to -78 °C. Compound 55-2 (1.87 g, 18.36 mmol, 2.1 mL) was slowly added. The reaction mixture was stirred at -70 °C for 30 min, and then compound 55-1 (3.20 g, 15.3 mmol) was added. The reaction was allowed to proceed for 2 h. The reaction was monitored by LCMS until complete. The mixture was quenched at 0 °C with saturated aqueous ammonium chloride solution (50.0 mL), stirred at 25 °C for 30 min, and the pH was adjusted to 5–6 with 1.0 M hydrochloric acid. The mixture was extracted with ethyl acetate (50.0 mL × 3), and the organic layers were combined. The mixture was washed with saturated aqueous sodium chloride solution (50.0 mL × 3), dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 55-3 (1.90 g, 8.26 mmol, yield 54.0%), which was a colorless oily compound. 1 H NMR (400MHz, CDCl3) δ3.69 (s, 3H), 2.84 (q, J = 7.2Hz, 1H), 1.43 (s, 9H), 1.20 (s, 3H), 1.19 (s, 3H), 1.10 (d, J = 7.6Hz, 3H).

[0622] Step Two:

[0623] Intermediate compound 55-3 (1.90 g, 8.26 mmol) was dissolved in dichloromethane (10.0 mL), and TFA (5.0 mL) was added. The mixture was stirred at 25 °C for 2 hours. The reaction was monitored by TLC until complete. The solvent was removed by concentration under reduced pressure. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 55-4 (1.43 g, 8.24 mmol, yield 99.8%) as a yellow liquid. 1 HNMR (400MHz, CDCl3) δ3.71 (s, 3H), 2.98 (q, J = 7.2Hz, 1H), 1.27 (s, 3H), 1.24 (s, 3H), 1.20 (d, J = 7.2Hz, 3H).

[0624] Step 3:

[0625] Intermediate compound 55-4 (500 mg, 2.87 mmol) was dissolved in DMF (15.0 mL), and HATU (1.64 g, 4.31 mmol) and DIEA (1.85 g, 14.3 mmol, 2.50 mL) were added. The mixture was stirred at 25 °C for 1 hour. Then, compound 1-6 (966 mg, 2.87 mmol) was added, and the mixture was stirred at 25 °C for 2 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (20.0 mL), extracted with ethyl acetate (20.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (20.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 55-5 (1.30 g, 2.86 mmol, 99.5% yield), which was a yellow oily compound. LCMS [M+H] + m / z:457.4.

[0626] Step Four:

[0627] Intermediate compound 55-5 (1.30 g, 2.86 mmol) was dissolved in a mixture of methanol (21.0 mL) and water (7.0 mL), and lithium hydroxide monohydrate (1.20 g, 28.6 mmol) was added. The mixture was stirred at 50 °C for 12 hours. The reaction was monitored by LCMS until complete. The reaction solution was diluted with water (10.0 mL), the pH was adjusted to 2–3 with 1.0 M hydrochloric acid, and the mixture was extracted with ethyl acetate (10.0 mL × 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 55-6 (261 mg, 590 μmol, yield 20.6%), a colorless oily compound. LCMS [M+H] + m / z:443.4.

[0628] Step 5:

[0629] Intermediate compound 55-6 (241 mg, 544 μmol) and compound 1-10 (170 mg, 1.09 mmol) were dissolved in DMF (5.0 mL), and DIEA (351 mg, 2.72 mmol, 474 μL) and HATU (414 mg, 1.09 mmol) were added. The reaction was carried out at 80 °C for 40 min. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 55-7 (46 mg, 79.2 μmol, yield 14.6%) as a yellow solid. LCMS [M+H]+ m / z:581.4.

[0630] Step Six:

[0631] TFA (2.0 mL) was added to intermediate compound 55-7 (46 mg, 79.2 μmol), and the reaction was carried out at 40 °C for 1 hour. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to obtain the final product compound WLE55 (1 mg, 3.2 μmol, yield 4.1%) as a white solid. 1 H NMR(400MHz,CH3OD)δ6.61(s,1H),4.93(t,J=16.8Hz,1H),4.80-4.56(m,1H),3.98-3.77(m,2H ),3.35(q,J=7.2Hz,1H),2.98-2.69(m,5H),1.42-1.29(m,15H),1.26-1.15(m,3H).LCMS[M+H] + m / z:461.2.

[0632] Example 54: Synthesis of compound WLE56

[0633]

[0634] Step 1:

[0635] Compound 56-1 (300 mg, 1.87 mmol) was dissolved in DMF (3.0 mL), and HATU (1.07 g, 2.81 mmol) and DIEA (1.21 g, 9.37 mmol, 1.6 mL) were added. The mixture was stirred at 25 °C for 1 hour. Then, compound 1-10 (878 mg, 5.62 mmol) was added, and the mixture was stirred at 25 °C for 12 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 56-2 (500 mg, 1.68 mmol, yield 89.5%), a yellow oily compound. LCMS [M+H] + m / z:299.0.

[0636] Step Two:

[0637] Compound 56-2 (500 mg, 1.68 mmol) was dissolved in a mixed solution of THF (3.0 mL), methanol (1.0 mL), and water (1.0 mL). Lithium hydroxide monohydrate (703 mg, 16.7 mmol) was added, and the mixture was stirred at 25 °C for 12 hours. The reaction was monitored by LCMS until complete. The reaction solution was diluted with water (10.0 mL), and the pH was adjusted to 2–3 with 1.0 M hydrochloric acid. Extraction was performed with ethyl acetate (10.0 mL × 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 56-3 (400 mg, 1.41 mmol, yield 83.9%) as a white solid. LCMS [M+H] + m / z:285.2.

[0638] Step 3:

[0639] Intermediate compound 56-3 (400 mg, 1.41 mmol) and compound 1-6 (422 mg, 1.41 mmol) were dissolved in DMF (5.0 mL), and DIEA (909 mg, 7.03 mmol, 1.2 mL) and HATU (802 mg, 2.11 mmol) were added. The reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure to remove the solvent, and purified by silica gel column chromatography to obtain intermediate compound 56-4 (30 mg, 52.9 μmol, yield 3.8%), a yellow oily compound. LCMS [M+H] + m / z:567.1.

[0640] Step Four:

[0641] TFA (1.0 mL) and TfOH (0.1 mL) were added to intermediate compound 56-4 (25 mg, 44.1 μmol), and the reaction was carried out at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to obtain the final product compound WLE56 (10 mg, 22.4 μmol, yield 50.8%) as a white solid. 1H NMR(400MHz, DMSO-d6)δ6.61(s,1H),4.93(t,J=16.8Hz,1H),4.80-4.56(m,1H),3.98-3.77(m,2 H),3.35(q,J=7.2Hz,1H),2.98-2.69(m,5H),1.42-1.29(m,15H),1.26-1.15(m,3H).LCMS[M+H] + m / z:447.2.

[0642] Example 55: Synthesis of compound WLE57

[0643]

[0644] Step 1:

[0645] Compound 57-1 (500 mg, 2.02 mmol) was dissolved in DMF (5.0 mL), and HATU (1.15 g, 3.03 mmol) and DIEA (1.31 g, 10.1 mmol, 1.8 mL) were added. The mixture was stirred at 25 °C for 1 hour. Then, compound 1-6 (607 mg, 2.02 mmol) was added, and the mixture was stirred at 25 °C for 2 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (10.0 mL), extracted with ethyl acetate (10.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (10.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 57-2 (750 mg, 1.42 mmol, 70.0%) as a yellow solid. LCMS [M+H] + m / z:530.4.

[0646] Step Two:

[0647] Intermediate compound 57-2 (750 mg, 1.42 mmol) was dissolved in a mixed solution of THF (6.0 mL), methanol (2.0 mL), and water (2.0 mL). Lithium hydroxide monohydrate (594 mg, 14.2 mmol) was added, and the mixture was stirred at 25 °C for 12 hours. The reaction was monitored by LCMS until complete. The reaction solution was diluted with water (10.0 mL), and the pH was adjusted to 2–3 with 1.0 M hydrochloric acid. Extraction was performed with ethyl acetate (10.0 mL × 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 57-3 (640 mg, 1.24 mmol, yield 87.7%) as a yellow solid. LCMS [M-56+H] + m / z:460.0.

[0648] Step 3:

[0649] Intermediate compound 57-3 (600 mg, 1.16 mmol) and compound 1-10 (422 mg, 1.41 mmol) were dissolved in DMF (7.0 mL), and DIEA (752 mg, 5.82 mmol, 1.0 mL) and HATU (664 mg, 1.75 mmol) were added. The reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (20.0 mL), extracted with ethyl acetate (20.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (20.0 mL × 3), dried over anhydrous sodium sulfate, filtered, concentrated under reduced pressure to remove the solvent, and the resulting mixture was purified by silica gel column chromatography to give intermediate compound 57-4 (650 mg, 994 μmol, yield 85.4%) as a yellow solid. LCMS [M+H] + m / z:654.4.

[0650] Step Four:

[0651] TFA (1.0 mL) and TfOH (0.1 mL) were added to intermediate compound 57-4 (100 mg, 153 μmol), and the reaction was carried out at 25 °C for 2 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by silica gel column chromatography to obtain the final product compound WLE57 (40 mg, 91.4 μmol, yield 59.8%) as a white solid. 1 H NMR(400MHz,DMSO-d6)δ8.20(s,1H),7.95-8.05(m,1H),6.71(s,1H),4.51-4.85(m,2H),4.17-4.30(m,1H ),3.62-3.86(m,2H),2.74-2.90(m,2H),2.72(d,J=4.4Hz,3H),2.54-2.67(m,2H),1.25(s,9H).LCMS[M+H] + m / z:434.2.

[0652] Example 56: Synthesis of compound WLE58:

[0653]

[0654] Step 1:

[0655] Compound 58-1 (2.00 g, 6.19 mmol) was dissolved in DMF (20.0 mL), and potassium carbonate (1.28 g, 9.28 mmol) and methyl iodide (1.76 g, 12.4 mmol, 770 μL) were added. The reaction was carried out at 20 °C for 2 hours. After the reaction was monitored by TLC, water (20.0 mL) was added to quench the reaction, and the mixture was extracted with ethyl acetate (20.0 mL × 2). The mixture was washed with lithium chloride solution (5%, 20.0 mL) and saturated sodium chloride aqueous solution (20.0 mL), dried over anhydrous sodium sulfate, filtered, and the solvent was removed by rotary evaporation under reduced pressure. The resulting mixture was intermediate compound 58-2 (2.01 g, 5.97 mmol, yield 96.5%), which was a yellow oily compound. 1 H NMR (400MHz, CDCl3) δ7.39-7.34(m,5H),5.49(d,J=8.0Hz,1H),5.17-5.10(m,2H),4.60-4.58(m,1H),3. 71(s,3H),3.06(dd,J=16.8Hz,J=4.0Hz,1H),2.88(dd,J=16.8Hz,J=4.8Hz,1H),1.45(s,9H).LCMS[M+Na] + m / z:360.0.

[0656] Step Two:

[0657] Under a nitrogen atmosphere, intermediate compound 58-2 (2.09 g, 6.20 mmol) was dissolved in anhydrous THF (20.0 mL), cooled to -78 °C, and LiHMDS (1.0 M, 13.9 mL) was added dropwise. The mixture was stirred for 30 minutes, and iodomethane (1.06 g, 7.43 mmol, 462 μL) was added. The reaction was allowed to proceed for 2.5 hours. The reaction was monitored by TLC until completion. The mixture was quenched with saturated ammonium chloride aqueous solution (20.0 mL), extracted with ethyl acetate (50.0 mL × 2), washed with saturated sodium chloride aqueous solution (50.0 mL × 2), dried over anhydrous sodium sulfate, filtered, and the solvent was removed by rotary evaporation under reduced pressure. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 58-3 (1.18 g, 3.36 mmol, yield 54.2%).

[0658] Step 3:

[0659] Under a nitrogen atmosphere, intermediate compound 58-3 (1.00 g, 2.85 mmol) was dissolved in methanol (20.0 mL), and Pd(OH)₂ / C (399 mg, 569 μmol) was added. After degassing, the mixture was reacted at 25 °C for 14 hours under a hydrogen atmosphere. The reaction was monitored by LCMS until completion. The solvent was removed by concentration under reduced pressure, and the resulting mixture, intermediate compound 58-4 (750 mg, 2.67 mmol, yield 93.8%), was a colorless oily compound. 5.49(minor,d,J=9.6Hz,1H),5.39(major,d,J=9.2Hz,1H),4.67(major,br,1H),4.52(minor,dd,J=9.4Hz,J=3.4Hz,1H),3.77(major,s,3H),3.75(minor,s,3H),3 .32-3.26(minor,m,1H),3.03-2.97(major,m,1H),1.47(minor,s,9H),1.45(major,s,9H),1.29(major,d,J=6.4Hz,3H),1.24(minor,d,J=7.2Hz,3H).LCMS[M+Na] + m / z:284.2.

[0660] Step Four:

[0661] Intermediate compound 58-4 (750 mg, 2.87 mmol) was dissolved in DMF (15.0 mL), and HATU (1.64 g, 4.31 mmol) and DIEA (1.86 g, 14.4 mmol, 2.50 mL) were added. The mixture was stirred at 25 °C for 1 hour. Then, compound 1-6 (966 mg, 2.87 mmol) was added, and the mixture was stirred at 25 °C for 2 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (30.0 mL), extracted with ethyl acetate (30.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (30.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 58-5 (1.50 g, 2.77 mmol, yield 96.4%) as a yellow solid. LCMS [M+H] + m / z:544.2.

[0662] Step 5:

[0663] Intermediate compound 58-5 (1.50 g, 2.77 mmol) was dissolved in a mixture of methanol (36.0 mL) and water (12.0 mL), and lithium hydroxide monohydrate (1.16 g, 27.7 mmol) was added. The mixture was stirred at 25 °C for 12 hours. The reaction was monitored by LCMS until complete. The reaction solution was diluted with water (10.0 mL), and the pH was adjusted to 2–3 with 1.0 M hydrochloric acid. The mixture was extracted with ethyl acetate (10.0 mL × 3). The organic phases were combined, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 58-6 (1.45 g, 2.74 mmol, yield 98.9%) as a white solid. 1 H NMR(400MHz, CDCl3)δδ12.32(br,1H),8.05-7.94(m,1H),7.14(dd,J=8.8Hz,J=2.8Hz,2H),6.90(d,J= 8.0Hz,2H),4.73(minor,dd,J=16.4Hz,J=4.8Hz,1H),4.40(major,dd,J=16.4Hz,J=4.4Hz,1H),4.23(m ajor,t,J=9.4Hz,1H),4.17-4.12(minor,m,1H),3.72(s,3H),3.67-3.61(m,1H),3.25-3.19(m,1H),2 .73-2.70(m,3H),2.69(s,3H),2.59(br,1H),1.99(s,1H),1.37(br,9H),1.06-0.89(m,3H).LCMS[M+H] + m / z:530.4.

[0664] Step Six:

[0665] Intermediate compound 58-6 (500 mg, 944 μmol) and compound 1-10 (221 mg, 1.42 mmol) were dissolved in DMF (10.0 mL), and DIEA (610 mg, 4.72 mmol, 822 μL) and HATU (538 mg, 1.42 mmol) were added. The reaction was carried out at 25 °C for 2 hours. The reaction was monitored by TLC until complete. The reaction was quenched with saturated ammonium chloride aqueous solution (20.0 mL), extracted with ethyl acetate (20.0 mL × 3), and the organic phases were combined. The mixture was washed with saturated sodium chloride aqueous solution (20.0 mL × 3), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the solvent. The resulting mixture was purified by silica gel column chromatography to give intermediate compound 58-7 (579 mg, 868 μmol, yield 91.9%) as a yellow solid. LCMS [M+H] + m / z:668.3.

[0666] Step Seven:

[0667] TFA (10.0 mL) and TfOH (1.00 mL) were added to intermediate compound 58-7 (579 mg, 868 μmol), and the reaction was carried out at 40 °C for 3 hours. The reaction was monitored by LCMS until complete. The solvent was removed by concentration under reduced pressure, and the resulting mixture was purified by prep-HPLC to obtain the final product compound WLE58 (143 mg, 319 μmol, yield 36.8%) as a white solid. 1 H NMR (400MHz, CDCl3) δ11.73 (brs, 1H), 7.70 (d, J = 5.2Hz, 1H), 6.48 (s, 1H), 5.0 0(d,J=16.0Hz,1H),4.60(d,J=16.4Hz,1H),4.46(dt,J=12.4Hz,3.6Hz,1H),3 .73-3.66(m,1H),3.45(d,J=3.6Hz,1H),3.14-3.07(m,1H),2.89(d,J=5.2Hz, 3H),2.73(d,J=15.6Hz,1H),2.59-2.51(m,1H),1.29-1.24(m,12H).LCMS[M+H] + m / z:448.1.

[0668] Example 57: Synthesis of compound WLE59

[0669]

[0670] Step 1:

[0671] Under a nitrogen atmosphere, intermediate compound 2-7 (100 mg, 161 μmol) was dissolved in 1,4-dioxane (1.0 mL) and water (0.2 mL). Intermediate compound 59-1 (61 mg, 402 μmol), Pd(dppf)Cl2 (13 mg, 16.1 μmol), and potassium carbonate (89 mg, 644 μmol) were added sequentially. After addition, the mixture was reacted at 100 °C for 12 hours. The reaction was monitored by TLC until completion. After cooling to room temperature, water (10.0 mL) was added to quench the reaction. The mixture was extracted with ethyl acetate (10.0 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 59-2 (40 mg, 66.6 μmol, yield: 41.4%) as a yellow solid. LCMS (M+H) + m / z:601.4.

[0672] Step Two:

[0673] TFA (1.0 mL) and TfOH (0.1 mL) were added to compound 59-2 (40 mg, 66.6 μmol). After the addition was complete, the mixture was reacted at 40 °C for 2 hours. The reaction was monitored by TLC. After the reaction was completed, TFA and TfOH were removed by concentration under reduced pressure. The resulting mixture was then prepared by high performance liquid chromatography to obtain the final product compound WLE59 (20 mg, 41.6 μmol, yield: 62.5%). 1 H NMR (400MHz, DMSO-d6) δ12.59-13.64(m,1H),12.06(s,1H),7.16-7.39(m,2H),6.59-6.74(m,1H),4.62-4.84(m,2H),3.76-3.96 (m,2H),3.05-3.20(m,1H),2.74-3.04(m,3H),2.58-2.71(m,1H),2.45-2.48(m,6H),1.24(s,9H),0.95-1.11(m,3H).LCMS(M+H) + m / z:481.3.

[0674] Example 58: Synthesis of compound WLE60

[0675]

[0676] Step 1:

[0677] Under a nitrogen atmosphere, intermediate compound 2-7 (100 mg, 161 μmol) was dissolved in 1,4-dioxane (5.0 mL) and water (1.0 mL). Intermediate compound 60-1 (61 mg, 321 μmol), Pd(dppf)Cl2 (12 mg, 16.1 μmol), and potassium carbonate (67 mg, 482 μmol) were added sequentially. After addition, the reaction was carried out at 90 °C for 12 hours. The reaction was monitored by TLC until completion. After the reaction solution cooled to room temperature, water (10.0 mL) was added to quench the reaction. Extraction was performed with ethyl acetate (10.0 mL × 2), and the solution was dried over anhydrous sodium sulfate. The mixture was filtered, concentrated under reduced pressure to remove the organic phase, and the resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 60-2 (85 mg, 133 μmol, yield: 82%) as a yellow solid. LCMS (M+H) + m / z:641.1.

[0678] Step Two:

[0679] TFA (3.0 mL) and TfOH (0.3 mL) were added to compound 60-2 (75.0 mg, 117 μmol). After the addition was complete, the mixture was reacted at 70 °C for 8 hours. The reaction was monitored by TLC. After the reaction was completed, TFA and TfOH were removed by concentration under reduced pressure. The resulting mixture was then prepared by high performance liquid chromatography to obtain the final product compound WLE60 (20 mg, 38.5 μmol, yield: 33%). 1 H NMR(400MHz,CD3OD)δ8.64-8.82(m,1H),8.07-8.19(m,1H),7.79-7.93(m,1H ),6.51-6.62(m,1H),4.95-5.08(m,1H),4.75(d,J=16.0Hz,1H),4.04-4.16(m ,1H),3.88-3.99(m,1H),3.52-3.63(m,1H),3.06-3.18(m,1H),2.77-3.02(m ,2H),2.53-2.64(m,1H),1.23-1.31(m,9H),1.17(d,J=6.8Hz,3H).LCMS(M+H) + m / z:521.6.

[0680] Example 59: Synthesis of compound WLE61

[0681]

[0682] Step 1:

[0683] Under a nitrogen atmosphere, intermediate compound 2-7 (100 mg, 161 μmol) was dissolved in 1,4-dioxane (1.0 mL) and water (0.2 mL). Intermediate compound 61-1 (63 mg, 402 μmol), Pd(dppf)Cl2 (13 mg, 16.1 μmol), and potassium carbonate (89 mg, 644 μmol) were added sequentially. After addition, the reaction was carried out at 70 °C for 2 hours. The reaction was monitored by TLC until completion. After the reaction solution cooled to room temperature, water (10.0 mL) was added to quench the reaction. The mixture was extracted with ethyl acetate (10.0 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 61-2 (44 mg, 72.5 μmol, yield: 45.0%) as a white solid. LCMS (M+H) + m / z:607.3.

[0684] Step Two:

[0685] TFA (1.0 mL) and TfOH (0.1 mL) were added to intermediate compound 61-2 (44 mg, 72.5 μmol). After the addition was complete, the reaction was carried out at 25 °C for 2 hours. The reaction was monitored by TLC. After the reaction was completed, TFA and TfOH were removed by concentration under reduced pressure. The resulting mixture was then prepared by high performance liquid chromatography to obtain the final product compound WLE61 (20 mg, 41.2 μmol, yield: 57%). 1 H NMR (400MHz, DMSO-d6) δ13.00-13.44(m,1H),12.06(brs,1H),8.44(d,J=4.4Hz,1H),7.55-7.72(m,2H),6.67(s,1H),4.61-4.88( m,2H),3.67-3.90(m,2H),2.71-2.99(m,3H),2.66(d,J=3.2Hz,1H),2.40-2.47(m,1H),1.24(s,9H),0.98-1.11(m,3H).LCMS(M+H) + m / z:487.2.

[0686] Example 60: Synthesis of compound WLE62

[0687]

[0688] Step 1:

[0689] Under a nitrogen atmosphere, intermediate compound 2-7 (100 mg, 161 μmol) was dissolved in 1,4-dioxane (1.0 mL) and water (0.2 mL). Intermediate compound 62-1 (62 mg, 402 μmol), Pd(dppf)Cl2 (13 mg, 16.1 μmol), and potassium carbonate (89 mg, 644 μmol) were added sequentially. After addition, the reaction was carried out at 70 °C for 2 hours. The reaction was monitored by TLC until completion. After the reaction solution cooled to room temperature, water (10.0 mL) was added to quench the reaction. The mixture was extracted with ethyl acetate (10.0 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 62-2 (53 mg, 88 μmol, yield: 55%) as a yellow solid. LCMS (M+H) + m / z:603.3.

[0690] Step Two:

[0691] TFA (1.0 mL) and TfOH (0.1 mL) were added to intermediate compound 62-2 (53 mg, 88 μmol). After addition, the reaction was carried out at 25 °C for 12 hours, monitored by TLC. Upon completion of the reaction, the mixture was concentrated under reduced pressure to remove TFA and TfOH. The resulting mixture was then subjected to high-performance liquid chromatography (HPLC) to prepare intermediate compound 62-3 (20 mg, 41.5 μmol, yield: 47%), which was a yellow oil. LCMS (M+H) + m / z:483.2.

[0692] Step 3:

[0693] Intermediate compound 62-3 (20 mg, 41.5 μmol) was dissolved in DCE (1.0 mL), cooled to below 0 °C, and boron tribromide (31 mg, 125 μmol) was slowly added dropwise. After the addition was complete, the reaction was carried out at 70 °C for 16 hours. The reaction was monitored by TLC until it ended. After the reaction solution cooled to room temperature, water (10.0 mL) was added to quench the reaction, and the mixture was extracted with ethyl acetate (10.0 mL × 2). The extract was dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was then subjected to high performance liquid chromatography to prepare the final product compound WLE62 (5 mg, 10.7 μmol, yield: 26%), which was a white solid. 1 H NMR(400MHz,CD3OD)δ7.47(brs,1H),6.82-6.89(m,1H),6.68-6.80(m,1H),6.55-6.59(m,1H),4.92-4.99(m,1H),4. 63-4.69(m,1H),3.88-4.01(m,2H),3.51-3.54(m,1H),2.52-2.98(m,4H),1.24(s,9H),0.95-1.11(m,3H).LCMS(M+H) + m / z: 481.3. LCMS(M+H) + m / z:469.3.

[0694] Example 61: Synthesis of compound WLE63

[0695]

[0696] Step 1:

[0697] Under a nitrogen atmosphere, intermediate compound 2-7 (200 mg, 321 μmol) was dissolved in 1,4-dioxane (5.0 mL) and water (0.5 mL). Intermediate compound 63-1 (146 mg, 643 μmol), Pd(dppf)Cl2 (24 mg, 32.2 μmol), and potassium carbonate (133 mg, 965 μmol) were added sequentially. After the additions were complete, the reaction was carried out at 25 °C for 12 hours. The reaction was monitored by TLC until completion. After the reaction solution cooled to room temperature, water (10.0 mL) was added to quench the reaction. The mixture was extracted with ethyl acetate (10.0 mL × 2), dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to remove the organic phase. The resulting mixture was purified by silica gel column chromatography to obtain intermediate compound 63-2 (135 mg, 200 μmol, yield: 68%) as a white solid. LCMS (M+H) + m / z:677.3.

[0698] Step Two:

[0699] Under a hydrogen atmosphere, intermediate compound 63-2 (135 mg, 200 μmol) was dissolved in methanol (10.0 mL), and Pd / C (1.00 g, 939 μmol) was added. The reaction was carried out at 25 °C for 3 hours. The reaction was monitored by TLC until completion. The mixture was filtered, and the filtrate was concentrated under reduced pressure to remove the organic phase. The resulting mixture was the crude intermediate compound 63-3 (110 mg), a yellow solid. LCMS (M+H) + m / z:679.6.

[0700] Step 3:

[0701] TFA (3.0 mL) and TfOH (0.3 mL) were added to compound 63-3 (50 mg, 73.6 μmol). After the addition was complete, the mixture was reacted at 70 °C for 12 hours. The reaction was monitored by TLC. After the reaction was completed, TFA and TfOH were removed by concentration under reduced pressure. The resulting mixture was then prepared by high performance liquid chromatography to obtain the final product compound WLE63 (10 mg, 21.8 μmol, yield: 30%). 1 H NMR (400MHz, CD3OD) δ8.54(brs,1H),6.59(s,1H),4.43-4.62(m,2H),3.80-4.03(m,2H),3.47(t,J=13.6Hz,3H),3.05-3.17(m,2H),2.77-3 .03(m,3H),2.69(t,J=5.2Hz,1H),2.49-2.60(m,1H),2.10(d,J=12.4Hz,2H),1.89-2.03(m,2H),1.28(s,9H),1.13-1.24(m,3H).LCMS(M+H) +m / z:459.2.

[0702] Test Implementation Examples

[0703] Biological assay

[0704] Test Example 1: In vitro ERK1 kinase inhibitory activity assay

[0705] Reaction system: Substrate (FFKNIVTPRTPPPSQGK, SEQ ID No.:1) MW: 1914Da; Composition: Activated MEK-ΔN3-SESD (MEK-Del32-51--S218E, S222D), unphosphorylated ERK1, Promega kinase-Glo (Cat: V3772; Promega);

[0706] instrument: NBS TM 384-well microplate (Cat: 3574); Multifunctional microplate testing instrument (Sparktecan); Microplate thermostatic oscillator (Mio).

[0707] 1) Preparation of kinase reaction solution: HEPES (pH=7.4) 50mM, NaCl 150mM, BSA 0.1mg / mL, DTT 2mM, MgCl2 10mM, Triton X-100 0.01%.

[0708] 2) Prepare solutions of the test compounds at different concentrations using DMSO. Eight solutions of the corresponding test compounds at different concentrations were prepared by equal-fold (3-fold) dilution with DMSO. The concentrations were 100, 33.33, 11.11, 3.70, 1.23, 0.41, 0.14, and 0.05 μM.

[0709] 3) After dilution, take 1 μL of DMSO to prepare different concentrations of the test compound solution and add 24 μL of kinase reaction solution to each solution. Mix thoroughly to obtain kinase reaction solutions with test compound concentrations of 4000, 1333.33, 444.44, 148.15, 49.38, 16.46, 5.49 and 1.83 nM. Prepare a DMSO control solution.

[0710] 4) Preparation of other components of the system: Prepare 800 μM substrate, 120 nM unphosphorylated ERK1, 400 μM ATP and 400 nM MEKΔN-SESD using kinase reaction solution.

[0711] 5) In a horizontally arranged 8-tube array, add 6.5 μL of unphosphorylated ERK1 to each tube, then add 6.5 μL of the kinase reaction solution containing different concentrations of the test compound prepared in step 3) above to the pipette and mix well. After sealing with sealing film, incubate at 25°C and 400 rpm in a microplate shaker for 1 hour.

[0712] 6) After shaking for 1 hour, add 6.5 μL of substrate to the octet and mix by rotation. Continue adding 6.5 μL of ATP and the activated MEKΔN-SESD to the octet and mix by rotation. The final reaction system (containing 0.1% DMSO) yielded test compound concentrations of 1000, 333.33, 111.11, 37.04, 12.35, 4.12, 1.37, and 0.46 nM. Control group Lut setting: 6.5 μL kinase reaction solution + 6.5 μL test compound + 6.5 μL substrate + 6.5 μL (ATP + MEKΔN-SESD) and control group Luc setting b: 6.5 μL unphosphorylated ERK1 + 6.5 μL kinase reaction solution containing DMSO + 6.5 μL substrate + 6.5 μL (ATP + MEKΔN-SESD).

[0713] 7) Incubate the reaction system in a microplate shaker at 25°C, 400 rpm for 30 min. Meanwhile, remove the pre-prepared Kinase-Glo solution and allow it to thaw at room temperature for 30 min.

[0714] 8) After incubation, transfer 10 μL of the reaction mixture to a 384-well plate, repeating this process once more as a replicate. Then, transfer 10 μL of Kinase-Glo solution into each well. Place the 384-well plate containing the transferred reaction mixture in a microplate shaker at 25°C, 400 rpm, for 10 min.

[0715] 9) Place the incubated 384-well plate into a microplate reader, select the luciferase assay program, and record the obtained chemiluminescence readings (Luc). Simultaneously, collect the chemiluminescence readings (Lut and Luc) of the control group (Lut and Luc, respectively).

[0716] 10) Inhibition rate calculation: Inhibition rate = [1 - ((Lut - lu)) / ((Lut - Luc))] * 100%

[0717] 11) Substitute the data into Graphpad Prism and calculate the IC using log(inhibitor) vs. response -- Variable slope (four parameters) analysis. 50 The values ​​are shown in Table 1 below.

[0718] Table 1. Inhibitory activity of compounds against ERK1 kinase

[0719]

[0720] Test Example 2: In vitro ERK2 kinase inhibitory activity assay

[0721] Except that unphosphorylated ERK1 was replaced with unphosphorylated ERK2, the inhibitory activity of the compounds against ERK2 kinase was tested using the same method as in Test Example 1, and the results are shown in Table 2 below.

[0722] Table 2. Inhibitory activity of compounds against ERK2 kinase

[0723]

[0724]

[0725] Test Example 3: Detection of Thr359 phosphorylation of p90RSK protein in A375 cells

[0726] Reagents and Instruments: Culture media: 10% FBS (Cat: SA211.02; CellMax) + 89% DMEM (Cat: C11995500BT; Thermo) + 1% Pen Strep (Cat: 15140; Gibco); Hoechst 33342 (Cat: C1022; Beyotime); 37% formaldehyde solution (Cat: R010480-500mL; Ron Reagent); Multifunctional microplate reader (Spark Tecan); Corning 96well black (Cat: 3916); Phospho-p90RSK (Thr359)(D1E9) Rabbit mAb (Cat: 8753; CST); Alex fluoride 488 Sheep Anti-Rabbit (Cat: A0423; Beyotime); Tween-20 (Cat: T818927-500ml; McLean).

[0727] 1) A375 cells were prepared at a rate of 2 × 10⁶ cells per well. 4 Cells were seeded in 80 μL of medium in 96-well plates and cultured overnight. (The outermost wells of the 96-well plates were filled with 100 μL of PBS except for the rightmost well; the rightmost well was filled with 100 μL of medium as the Blank group; 80 μL of sample was added using a pipette.)

[0728] 2) Prepare solutions of the test compounds at different concentrations using DMSO. Nine solutions of the corresponding test compounds at different concentrations were prepared by equal-fold (4-fold) dilution with DMSO. The concentrations of the test compound solutions were 10000, 2500, 625, 156.25, 39.06, 9.77, 2.44, 0.61, and 0.15 μM.

[0729] 3) After dilution, take 3 μL of DMSO to prepare different concentrations of the test compound solution and add 198 μL of the corresponding culture medium, then mix thoroughly. Next, add 20 μL / well of the above-mentioned culture medium containing different concentrations of the test compound to each of the 96 wells already seeded with cells. Finally, culture media (containing 0.3% DMSO) were prepared with final concentrations of the test compounds of 30000, 7500, 1875, 468.8, 117.2, 29.3, 7.3, 1.8, and 0.5 nM, and a 0.3% DMSO control solution was also prepared.

[0730] 4) Place the test compound and DMSO-treated cells back into a 37°C incubator for 2 hours.

[0731] 5) After incubation, remove almost all the culture medium by decantation and invert the 96-well plate onto absorbent paper. Prepare a 3.7% formaldehyde solution using 37% formaldehyde solution at a 10× concentration. Add 50 μL of the 3.7% formaldehyde solution to each well using a multi-channel pipette. Incubate on a horizontal shaker at 200 rpm for 20 min at room temperature. After incubation, discard the formaldehyde solution by decantation and gently add 100 μL of PBS to the multi-channel pipette for washing. Incubate on a horizontal shaker at 200 rpm for 5 min at room temperature. Discard almost all of the PBS by decantation, shake dry thoroughly, and then blot dry with paper.

[0732] 6) Prepare 0.1% Triton X-100 using the 10% Triton X-100 method as per 100X. Add 0.1% Triton X-100 to each well at a rate of 50 μL. Incubate on a horizontal shaker at 200 rpm for 20 min at room temperature. Repeat step 5), using a pump to remove virtually all of the PBS with minimal suction.

[0733] 7) Prepare an antibody dilution buffer by dissolving 0.05% Tween-20 and 5% BSA in PBS (store at -20°C). Dilute the primary antibody (Phospho-p90RSK(Thr359)(D1E9)Rabbit mAb) at a ratio of 1:1000 to obtain the antibody solution. Add 25 μL of the solution to each well. Gently shake to cover the bottom of the 96-well plate and incubate overnight at 4°C on a shaker.

[0734] 8) After removing unbound antibodies by the decantation method, gently add 100 μL of PBST containing 0.05% Tween-20 to the micropipe for washing. Run on a horizontal shaker at 200 rpm for 5 min, then at rest (RT), repeating 3 times. Finally, use a pump with minimal suction to remove almost all PBST. Dilute Alex fluor 488 goat anti-rabbit fluorescent secondary antibody 1:500 using antibody dilution buffer, and simultaneously add Hoechst 33342 to a final concentration of 10 μg / ml. Add the diluted secondary antibody and Hoechst 33342 mixture to a minimum of 25 μL per well using a micropipe. Gently shake to cover the bottom of the 96-well plate, then run on a horizontal shaker at 200 rpm in the dark for 90 min, then at RT. Remove the secondary antibody by the decantation method, repeating the washing process 3 times, while protecting the plate from light during shaking.

[0735] 9) Place the 96-well plate, which contains virtually no solution, into an ELISA reader to detect fluorescence. Use an excitation light of 485 nm (20) to detect the emission light at 535 nm (25); and use an excitation light of 320 nm (25) to detect the emission light at 460 nm (20).

[0736] 10) Calculation of P90RSK phosphorylation level:

[0737] Calculation formula = [(

RFU_compound535-RFU_blank535

RFU_DMSO535-RFU_blank535

RFU_compound460-RFU_blank460

RFU_DMSO460-RFU_blank460

[0738] RFU_compound535 is the average emission reading at 535 nm for the analyte processing well; RFU_DMSO535 is the average emission reading at 535 nm for the DMSO processing well; RFU_blank535 is the average reading at 535 nm for the blank well; RFU_compound460 is the average emission reading at 460 nm for the analyte processing well; RFU_DMSO460 is the average emission reading at 460 nm for the DMSO processing well; RFU_blank460 is the average reading at 460 nm for the blank well.

[0739] 11) Substitute the data into Graphpad Prism and calculate EC using log(inhibitor) vs. response -- Variable slope (four parameters) analysis. 50 The values ​​are shown in Table 3 below.

[0740] Table 3. Inhibitory activity of compounds against phosphorylation of P90RSK protein at Thr359 site in A375 cells.

[0741]

[0742]

[0743] Test Example 4: Detection of Thr359 phosphorylation of P90RSK protein in HCT116 cells

[0744] In addition to using a culture medium of 10% FBS (Cat: SA211.02; CellMax) + 89% McCoy's 5A (Cat: 16600082; Thermo) + 1% Pen Strep (Cat: 15140; Gibco), and HCT116 cell seeding density of 4 × 10⁶ cells per well in step 1), the following conditions must be met. 4 In addition, the inhibitory activity of the compound on phosphorylation of p90RSK protein at Thr359 site in HCT116 cells was determined using the same method as in Test Example 3, and the results are shown in Table 4 below.

[0745] Table 4. Inhibitory activity of compounds against phosphorylation of p90RSK protein at Thr359 site in HCT116 cells.

[0746]

[0747] Test Example 5: Detection of A375 cell proliferation capacity

[0748] Reagents and instruments: Culture media: 10% FBS (Cat: SA211.02; CellMax) + 89% DMEM (Cat: C11995500BT; Thermo) + 1% Pen Strep (Cat: 15140; Gibco); Hoechst 33342 (Cat: C1022; Beyotime); 37% formaldehyde solution (Cat: R010480-500mL; Ron Reagent); Multifunctional microplate reader (Spark Tecan); Corning 96well black (Cat: 3916).

[0749] 1) A375 cells were grown at a rate of 6 × 10⁶ cells per well. 3 Cells were seeded in 80 μL of culture medium in 96-well plates and cultured overnight. (The outermost wells of the 96-well plates were filled with 100 μL of PBS except for the rightmost well; the rightmost well was filled with 100 μL of culture medium as a control group; 80 μL of sample was added using a pipette).

[0750] 2) Prepare test compound solutions of different concentrations using DMSO and further dilute with culture medium. Prepare nine different concentrations of the corresponding test compound solutions by isotropic (4-fold) dilution with DMSO. Then, add 198 μL of the corresponding culture medium to each of the different concentrations prepared with 3 μL of DMSO and mix thoroughly (pipe-pipette at least 20 times). Add 20 μL / well of the above-mentioned culture medium containing different concentrations of the test compound to each of the 96 wells containing cells. The final test compound concentrations were obtained as follows: 7.5, 1.88, 0.47, 0.12, 0.029, 0.0073, 0.0018, 0.00045, and 0.00011 μM culture medium (containing 0.3% DMSO), and a 0.3% DMSO control solution was also prepared. Each concentration of the test compound was tested in duplicate.

[0751] 3) The 96-well plates containing the test compound and DMSO-treated cells were then placed back into a 37°C incubator for three days.

[0752] 4) On the fifth day, harvest the cells. Use the inverted plate method to remove almost all the culture medium from the 96-well plate after culture, and invert the 96-well plate onto absorbent paper.

[0753] 5) Preparation of fixation and staining solution: Prepare 3.7% formaldehyde solution by using 37% formaldehyde water at a 10× concentration. Dilute Hoechst 33342 in the 3.7% formaldehyde solution to obtain a fixation and staining solution with a final concentration of Hoechst 33342 of 10 μg / mL.

[0754] 6) Add 50 μL of fixative staining solution to each well of a 96-well plate after removing the culture medium using a pipette. Incubate on a horizontal shaker at 200 rpm for 30 min at room temperature, protected from light.

[0755] 7) After fixation and staining, remove almost all the fixative solution from the 96-well plate using the decantation method. Gently add 100 μL of PBS to wash the plate. Incubate on a horizontal shaker at 200 rpm for 5 min at room temperature, protected from light. Repeat twice.

[0756] 8) Data detection: After cleaning and removing all liquid from the 96-well plate, fluorescence was detected using a microplate reader. Microplate reader parameters: excitation light 320nm (25), emission light 460nm (20).

[0757] 9) Data calculation: Cell viability = 100 * (drug-treated group - blank group / DMSO group - blank group). The values ​​for the drug-treated group are the average 460nm readings of the wells treated with the test compound; the values ​​for the DMSO group are the average 460nm readings of the wells treated with DMSO; and the values ​​for the blank group are the average 460nm readings of the culture medium.

[0758] 10) Substitute the data into Graphpad Prism and calculate EC using log(inhibitor) vs. response -- Variable slope (four parameters) analysis. 50 The values ​​are shown in Table 5 below.

[0759] Table 5. Inhibitory activity of compounds on the proliferation of A375 cells

[0760]

[0761]

[0762] Test Example 6: HCT116 Cell Proliferation Capacity Detection

[0763] In addition to using 10% FBS (Cat: SA211.02; CellMax) + 89% McCoy's 5A (Cat: 16600082; Thermo) + 1% Pen Strep (Cat: 15140; Gibco) as the culture medium, HCT116 cells were cultured at a rate of 5 × 10⁻⁶ cells / year. 3 The cells were seeded in 80 μL of culture medium in 96-well plates, and the remaining procedures were the same as in test example 5. The results are shown in Table 6 below.

[0764] Table 6. Inhibitory activity of compounds on the proliferation of HCT116 cells.

[0765]

[0766] The results of the above test examples show that the compounds in this application generally exhibit significant inhibitory activity against ERK1 / 2. Some compounds show comparable or even superior activity to existing MAPK pathway inhibitors and exhibit proliferative inhibitory activity against A375 cells and HCT116 cells. Therefore, they have the potential to be developed into drugs for the treatment of diseases related to ERK1 / 2 kinases.

Claims

1. A class of compounds represented by Formula I, their pharmaceutically acceptable salts, enantiomers, diastereomers, tautomers, solvates, polymorphs, or prodrugs: In equation I above, R 1 -R 3 R 1’ -R 3’ Each of them is independently H or C1-6 alkyl; W 1 Selected from C1-6 alkyl groups, by R 4 Substituted C1-6 alkyl, -NR 5 R 5’ -C(=O)NR 6 R 6’ C6-10 aryl, 4-12 membered monocyclic or fused heterocyclic groups containing 1-3 heteroatoms of N, O or S, and 5-12 membered monocyclic or fused heteroaryl groups containing 1-3 heteroatoms of N, O or S. in, R 4 Selected from hydroxyl and amino groups; R 5 R 5’ R 6 R 6’ Each time it appears, it is independently selected from H, C1-6 alkyl, C3-8 cycloalkyl, 4-12 membered monocyclic or fused heterocyclic groups containing 1-3 of N, O or S as heteroatoms, or 5-12 membered monocyclic or fused heteroaryl groups containing 1-3 of N, O or S as heteroatoms. The cycloalkyl, aryl, heterocyclic, and heteroaryl groups are optionally further substituted by one or more substituents selected from unsubstituted or halogenated C1-6 alkyl, halogen, hydroxyl, aldehyde, -S(=O)2R', and -OS(=O)2R', where R' is selected from halogen, C1-6 alkyl, or C2-C10 alkenyl. The cyclic carbon atom of the heterocyclic group may optionally be further oxidized; L represents the following bivalent connector: Where * indicates the position where the nitrogen atom is attached to the pyridine ring of pyrazolo[4,3-c]pyridine, and ** indicates the position where W2 is attached. X 1 and X 2 Each occurrence is independently assigned to -CR 7 R 7’ -or-NR 8 -, provided that the two are not both -NR 8 -; X is selected from -CR 11 R 11’ -and-C(=O)-, R 7 R 7’ Each of the following is independently selected from H, unsubstituted or substituted C1-6 alkyl groups, and amino groups; R 8 R 9 Each of them is independently H or C1-6 alkyl; R 11 R 11’ Each of these substances is independently H, halogen, or C1-6 alkyl; n is 0 or 1; W 2 For those not replaced or replaced by 1-3 Rs 10 Substituted C6-10 aryl groups, or 5-12 membered monocyclic or fused heteroaryl groups containing 1-3 N, O, or S as heteroatoms, each R 10 It is independently selected from halogens, unsubstituted or halogenated C1-6 alkyl groups, and C3-8 cycloalkyl groups.

2. The compound according to claim 1, its pharmaceutically acceptable salt, enantiomer, diastereomer, tautomer, solvate, polymorph, or prodrug, wherein, R 1 -R 3 R 1’ -R 3’ Each of them is independently H or C1-6 alkyl; W 1 It is a C1-6 alkyl group, -(CH2) 1-6 -R 4 -NR 5 R 5’ -C(=O)NR 6 R 6’ , phenyl, naphthyl, biphenyl, heteroaryl selected from 1,2,3-triazolyl, 1,2,4-triazolyl, imidazolyl, pyrazolyl, pyridinyl and oxazolyl, or heterocyclic group selected from tetrahydropyrroleyl, tetrahydropyranyl and piperidinyl. in, R 4 Selected from hydroxyl and amino groups; R 5 R 5’ R 6 R 6’ Each of the following is independently H in each occurrence, a C1-6 alkyl group, a C3-8 cycloalkyl group optionally substituted with deuterium, halogen, hydroxyl or amino, a heteroaryl group selected from imidazolyl, pyrazolyl and pyridinyl, or a heterocyclic group selected from piperidinyl, tetrahydropyrrolel, tetrahydrofuranyl and tetrahydropyranyl. The cycloalkyl, phenyl, heterocyclic, and heteroaryl groups are optionally further substituted by one or more substituents selected from unsubstituted or halogenated C1-6 alkyl, halogen, hydroxyl, aldehyde, -S(=O)2R', -OS(=O)2R', where R' is selected from halogen, C1-6 alkyl, or C2-C10 alkenyl. The cyclic carbon atom of the piperidinyl, tetrahydropyrrole, or tetrahydropyran may optionally be further oxidized; L represents the following bivalent connector: Where * indicates the position where the nitrogen atom is attached to the pyridine ring of pyrazolo[4,3-c]pyridine, and ** indicates the position where W2 is attached. X 1 and X 2 Each occurrence is independently assigned to -CR 7 R 7’ -or-NR 8 -, provided that the two are not both -NR 8 -; X is selected from -CH2- and -C(=O)-. R 7 R 7’ Each of the following is independently selected from H, unsubstituted or substituted C1-6 alkyl groups, and amino groups; R 8 R 9 Each of them is independently H or C1-6 alkyl; n is 0 or 1; W 2 For those not replaced or replaced by 1-3 Rs 10 Substituted phenyl or naphthyl, or heteroaryl selected from thiazolyl, indolyl, oxazolyl, imidazolyl, pyrazolyl, benzothiazolyl, and pyridyl, each R 10 Independently selected from halogens, unsubstituted or halogenated C1-6 alkyl groups, and C3-8 cycloalkyl groups. Wherein, the above C1-6 alkyl groups are selected from methyl, ethyl, n-propyl, isopropyl, n-butyl, tert-butyl, isobutyl, sec-butyl, n-pentyl, or n-hexyl. The above C3-8 cycloalkyl groups are selected from cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, or cyclooctyl.

3. The compound according to claim 1, its pharmaceutically acceptable salt, enantiomer, diastereomer, tautomer, solvate, polymorph, or prodrug, wherein, In equation I, R 1 R 2 and R 3 and R 1’ R 2’ and R 3’ All are H; W 1 -C(=O)NH-R 6 , where R 6 As defined in claim 1; L is Among them, X 1 and X 2 As defined in claim 1, R 9 It is H or C1-6 alkyl; and W 2 4-tert-butylthiazolyl * indicates the location of the group connection; or, In equation I, R 1 R 2 and R 3 and R 1’ R 2’ and R 3’ All are H; W 1 -C(=O)NH-R 6 , where R 6 As defined in claim 1; L is as well as W 2 As defined in claim 1.

4. The compound according to claim 1, its pharmaceutically acceptable salt, enantiomer, diastereomer, tautomer, solvate, polymorph, or prodrug, wherein, The compound represented by Formula I is represented by Formula I-1 or I-2: In formula I-1 or I-2 above, the definitions of each substituent are as defined in claim 1 or 2, respectively.

5. The compound according to claim 1, its pharmaceutically acceptable salt, enantiomer, diastereomer, tautomer, solvate, polymorph, or prodrug, wherein, The W 1 The substituent is selected from unsubstituted or substituted diazolyl, triazolyl, pyridyl, piperidinyl, tetrahydropyranyl, tetrahydropyrrolidinyl, and oxazolyl groups, wherein the substituent is selected from halogens, alkyl groups, haloalkyl groups, hydroxyl groups, and oxo groups; and / or The W 2 Selected from those that have not been replaced or have been replaced by 1-3 Rs 10 Substituted thiazolyl, indolyl, indazole, oxazolyl, imidazole, benzothiazolyl, naphthyl, and pyridyl.

6. The compound according to claim 1, its pharmaceutically acceptable salt, enantiomer, diastereomer, tautomer, solvate, polymorph, or prodrug, wherein, The compounds of Formula I are selected from the following:

7. A pharmaceutical composition comprising a therapeutically effective amount of the compound of formula I as claimed in any one of claims 1-6, a pharmaceutically acceptable salt thereof, an enantiomer, a diastereomer, a tautomer, a solvate, a polymorph or a prodrug, and a pharmaceutically acceptable excipient.

8. Use of the compound of any one of claims 1-6, its pharmaceutically acceptable salt, enantiomer, diastereomer, tautomer, solvate, polymorph or prodrug, or the pharmaceutical composition of claim 7 in the preparation of a medicament for treating or preventing diseases or symptoms related to ERK1 / 2 kinase.

9. The use according to claim 8, wherein, Diseases or symptoms associated with ERK1 / 2 kinases include cancer, asthma, stroke, and Alzheimer's disease.

10. The use according to claim 9, wherein, The cancer is selected from neurofibroma, thyroid cancer, ovarian cancer, melanoma, colorectal cancer, pancreatic adenocarcinoma, and lung cancer, wherein the lung cancer can be non-small cell lung cancer.

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