WO2023050975A1

WO2023050975A1 - α-FLUOROACYL PIPERAZINE DERIVATIVE, AND PREPARATION AND APPLICATION THEREOF

Info

Publication number: WO2023050975A1
Application number: PCT/CN2022/105698
Authority: WO
Inventors: 沈征武; 邓斌; 江亮; 查雨峰; 张梦麒; 边泓竹
Original assignee: 云白药征武科技(上海)有限公司
Priority date: 2021-09-28
Filing date: 2022-07-14
Publication date: 2023-04-06
Also published as: CN113754680B; CN113754680A

Abstract

Disclosed in the present invention is an α-fluoroacyl piperazine derivative, comprising an α-fluoroacyl piperazine derivative of general formula (I) or an isomer thereof, or a pharmaceutically acceptable salt thereof, or a prodrug molecule thereof, and having a chemical structure as shown in general formula (I). The present invention also provides a preparation method for the α-fluoroacyl piperazine derivative. The present invention further provides an application of the α-fluoroacyl piperazine derivative or the isomer thereof, the pharmaceutically acceptable salt thereof, or the prodrug molecule thereof in the preparation of a drug for treating cancer. According to the α-fluoroacyl piperazine derivative of the present invention, a fluorine atom having a strong electron-withdrawing capability is introduced into an ortho-position of an acyl group, so that the electron cloud distribution of acyl piperazine is changed. Therefore, the α-fluoroacyl piperazine derivative has extremely strong PI3K kinase inhibitory activity and extremely strong antitumor activity, and has broad application prospects.

Description

A kind of alpha fluoroacylpiperazine derivative and its preparation and application

technical field

The invention belongs to the field of medicinal chemistry, and in particular relates to an α-fluoroacylpiperazine derivative and its preparation and application.

Background technique

PI3K is closely related to intracellular signal transduction, and plays a regulatory role in many important cellular processes, such as cell growth, differentiation, proliferation, apoptosis, and intracellular transport. The PI3K-Akt-mTOR signaling pathway plays an important role in cell growth, differentiation, and apoptosis. Many member molecules of signal transduction are key drug targets in the processes of cancer, immunity, and thrombus control. . When this signaling pathway is abnormally activated in the human body, it often leads to the occurrence of cancer.

PI3K kinases are divided into three types: I, II, and III. Among them, type I PI3K is the most deeply studied, and type I PI3K is a heterodimer composed of a regulatory subunit and a catalytic subunit. There are four catalytic subunits, namely p110α, β, δ, γ. Today, cancer and the target-pathological role of PI3K have been identified.

Each of the four catalytic subunits of class I PI3K kinases preferentially regulates specific signaling roles, depending on the type of malignancy and the genetic or epigenetic alterations that occur. For example, p110α is critical for the growth of tumor cells driven by PIK3CA mutations or oncogene RAS and receptor tyrosine kinases; p110β mediates PTEN-deficient tumorigenesis; and p110δ is highly expressed in leukocytes, thereby making It becomes an ideal target for the treatment of hematological malignancies. p110γ may play an important role in the pathogenesis of IPF (idiopathic pulmonary fibrosis), and may be a specific pharmacological target of IPF.

Due to the great potential of PI3K inhibitors in the treatment of tumors and various diseases, many pharmaceutical companies and research institutions have invested a lot of resources in the development of such drugs. PI3K inhibitors currently approved by the FDA include: Idelalisib, a PI3Kδ selective inhibitor, for the treatment of lymphoma; Copanlisib, a PI3K-α/δ inhibitor, for the treatment of recurrent follicular lymphoma; Duvelisib, a PI3Kδ/γ inhibitor, Treatment of lymphoma; Alpelisib, PI3K-α inhibitor, for the treatment of advanced metastatic breast cancer; Umbralisib, PI3Kδ and CK1ε inhibitor, for the treatment of lymphoma.

The PI3K inhibitor (Apitolisib, GDC-0980, RG7422, CAS: 1032754-93-0), developed by Genentech, is in the second phase of clinical trials. Kidney tumors;

Contents of the invention

The object of the present invention is to provide a kind of α-fluoroacylpiperazine derivative and its preparation and application, and the described α-fluoroacylpiperazine derivative and its preparation and application will solve the problems of drugs in the prior art for the treatment of Cancer's ineffective technical problems.

The present invention provides an α-fluoroacylpiperazine derivative, the α-fluoroacylpiperazine derivative having the general formula (I) or its isomer, or its pharmaceutically acceptable salt, or its prodrug molecule, which General formula (I) chemical structure is:

In the general formula (I), X=O or S; wherein, the acyl α position of the acylpiperazine contains at least one fluorine atom;

In the general formula (I), _R is H, halogen, cyano, alkyl, alkenyl, alkynyl or derivatives thereof of 1-15 carbons; monocyclic or condensed aromatic rings containing 5-22 carbon atoms Group or its derivatives; 5- to 8-membered heterocycle or heterocycle or its derivatives containing 1-4 heteroatoms; carboxyl or its derivatives; hydroxyl or its derivatives; amino or its derivatives; mercapto or derivatives thereof; sulfone or sulfoxide derivatives; sulfonates or sulfonates; phosphates or phosphates;

In the general formula (I), _R2 is H, halogen, cyano, alkyl, alkenyl, alkynyl or derivatives thereof of 1-15 carbons; monocyclic or condensed aromatic rings containing 5-22 carbon atoms Group or its derivatives; 5- to 8-membered heterocycle or heterocycle or its derivatives containing 1-4 heteroatoms; carboxyl or its derivatives; hydroxyl or its derivatives; amino or its derivatives; mercapto or derivatives thereof; sulfone or sulfoxide derivatives; sulfonate or sulfonate; phosphate or phosphate.

Further, in the general formula (I), the acyl α position of the acylpiperazine is a single optical configuration or a racemate; R ₁ and R ₂ can be the same or different; in the general formula (I), R ₁ and _R2 can be connected to form a ring structure.

Further, the preferred structure of the α-fluoroacylpiperazine derivative is as follows:

The present invention also provides a pharmaceutical composition, which contains a therapeutically effective amount of the above-mentioned α-fluoroacylpiperazine derivative or its isomer, or its pharmaceutically acceptable salt, or its prodrug molecule and one or more A pharmaceutically acceptable carrier, diluent or excipient.

The present invention also provides a method for preparing the above-mentioned α-fluoroacylpiperazine derivative, the reaction equation of which is as follows:

or as follows:

In the first reaction formula, P ₁ is H or an amino protecting group, P ₂ is H or an amino protecting group; P ₁ and P ₂ are not H at the same time;

Amino protecting groups include formyl, acetyl, trifluoroacetyl, benzoyl, tert-butoxycarbonyl, benzyloxycarbonyl, 9-fluorenylmethoxycarbonyl, phthaloyl, cyclosuccinyl, 2- Biphenyl-2-propoxycarbonyl, p-toluenesulfonyl, trityl, etc.;

The reductive amination reaction solvent can be anhydrous or water-containing, including dichloromethane, tetrahydrofuran, methanol, ethanol, isopropanol, 1,2-dichloroethane, ethylene glycol dimethyl ether, bis(ethylene glycol) Dimethyl ether, etc.;

The reducing reagents used in reductive amination include NaBH ₄ , KBH ₄ , LiBH ₄ , Zn(BH ₄ ) ₂ , NaBH ₃ CN, NaBH(OAc) ₃ , borane complex, Bu ₃ SnH, PhSiH _{4 ,} etc.;

Catalysts used in reductive amination include protic acid and Lewis acid, such as hydrochloric acid, sulfuric acid, phosphoric acid, formic acid, acetic acid, trifluoroacetic acid, BF ₃ , SnCl ₂ , SnCl ₄ , Ti(O ⁱ Pr) ₄ , SiO ₂ , BuSnCl ₂ etc. ;

Acids used for deprotection include protic acids and Lewis acids, such as hydrochloric acid, sulfuric acid, phosphoric acid, formic acid, acetic acid, p-toluenesulfonic acid, trifluoromethanesulfonic acid, trifluoroacetic acid, BF ₃ , SnCl ₂ , SnCl ₄ , Ti(O ⁱ Pr) ₄ , SiO ₂ , BuSnCl ₂ , AlCl ₃ etc.;

The bases used for deprotection include inorganic bases and organic bases. Inorganic bases include: sodium hydroxide, potassium hydroxide, sodium hydride, calcium hydride, calcium fluoride, cesium fluoride, sodium carbonate, potassium carbonate, cesium carbonate, potassium phosphate, etc.; Organic bases include: hydrazine hydrate, lithium diisopropylamide, butyllithium, lithium bis(trimethylsilyl)amide, triethylamine, diisopropylethylamine, pyridine, pyrrole, piperidine, morpholine, N-methylmorpholine, 1,8-diazabicycloundec-7-ene, etc.;

The hydrogen source used in the catalytic reduction deprotection reaction includes hydrogen, formic acid, ammonium formate, etc.; the catalyst includes metal catalysts such as Pd, Pt, Ni, and Cu;

The solvent used in the deprotection reaction may be a protic solvent, an aprotic solvent or a mixed solvent. Preferably dichloromethane, dichloroethane, tetrahydrofuran, acetonitrile, methanol, ethanol, water, ethylene glycol dimethyl ether, N,N-dimethylformamide, dimethyl sulfoxide, etc.;

The thio reagents are P ₂ S ₅ , 2,4-bis(methylthio)-1,3,2,4-dithiadiphosphatidine-2,4-disulfide, 2,4 -Bis(phenylthio)-1,3-dithio-2,4-diphosphetane-2,4 disulfide, or 2,4-bis(4-phenoxyphenyl)- 1,3,2,4-dithiodiphosphetane-2,4-disulfide; the solvent used in the reaction is either a protic solvent or an aprotic solvent or a mixture of both;

In the second reaction formula, the fluorinated reagents used include HF and its salts, SF ₄ , diethylaminosulfur trifluoride, bis(2-methoxyethyl)aminosulfur trifluoride, 4-tert-butyl Base-2,6-dimethylphenylsulfur trifluoride, pyridine-2-sulfonyl fluoride, bis(2-methoxyethyl)aminosulfur trifluoride, diethylamino)difluorosulfonium tetrafluoro Borate, difluoro(4-morpholino)sulfonium tetrafluoroborate, 1,3-bis(2,6-diisopropylphenyl)-2,2-difluoroimidazoline, 4-chloro -N-[(4-methylphenyl)sulfonyl]-benzenesulfonyl fluoride; the solvent used in the reaction is an aprotic solvent or a mixed solvent. Preferred are dichloromethane, dichloroethane, tetrahydrofuran, acetonitrile, ethylene glycol dimethyl ether, 1,4-dioxane and the like.

The reaction temperature is -78 to 180 degrees Celsius;

The crude product of the final product in the above-mentioned reaction can use solvent extraction method, precipitation method, crystallization method to further purify, also can use column chromatography to carry out purification, filler uses silica gel, gel, macroporous resin or aluminum oxide, and eluent can be Mix petroleum ether-acetone, petroleum ether-ethyl acetate, petroleum ether-dichloromethane, etc. in different proportions.

The present invention also provides an application of the above-mentioned α-fluoroacylpiperazine derivative or its isomer, or its pharmaceutically acceptable salt, or its prodrug molecule in the preparation of a drug for treating cancer.

Further, the cancer is brain cancer, glioma, endometrial cancer, ovarian cancer, cervical cancer, breast cancer, colon cancer, lung cancer, prostate cancer, liver cancer, leukemia, lymphoma, skin cancer, basal cell tumor , hemangioma, uterine cancer, laryngeal cancer, stomach cancer, lip cancer, esophagus cancer, nasopharyngeal cancer, gallbladder cancer, pancreatic cancer, kidney cancer, tongue cancer, bladder cancer, melanoma, lipoma, thyroid cancer, thymus cancer or bone cancer.

The present invention also provides the above-mentioned α-fluoroacylpiperazine derivative or its isomer, or its pharmaceutically acceptable salt, or its prodrug molecule combined with at least one other anti-cancer agent in the preparation of the treatment of cancer application in medicines.

Further, the other anticancer agents are doxorubicin, bleomycin, vinblastine, taxanes, etoposide, 5-fluorouracil, cyclophosphamide, methotrexate, cisplatin, Retinoic acid, temozolomide, actinomycin, imatinib, gefitinib, sorafenib, erlotinib, sunitinib, afatinib, cabozantinib, ostinib, Any one or both of rituximab, cetuximab, trastuzumab, nivolumab, panlizumab, atezolizumab, durvalumab, or avelumab combination of the above.

The α-fluorinated amide piperazine derivatives of the present invention introduce a fluorine atom with strong electron-withdrawing ability at the ortho-position of the amide group, which changes the electron cloud distribution of amide piperazine, not only enhances the PI3K kinase inhibitory activity of this type of molecule, but also On the one hand, the lipid-water distribution coefficient of the compound is also optimized, making it easier for this type of drug to permeate the cell membrane, so it has a strong anti-tumor activity and has a very broad application prospect.

Detailed ways

The following clearly and completely describes the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only some of the embodiments of the present invention, but not all of them. Based on the embodiments of the present invention, all other embodiments obtained by persons of ordinary skill in the art without creative efforts fall within the protection scope of the present invention.

Embodiment one: the preparation of compound 1:

(1), preparation of compound 1a: under nitrogen protection, 2-chloro-7-methyl-4-morpholin-4-ylthieno[3,2-d]pyrimidine-6-carbaldehyde (CAS: 955979- 02-9, 190 g, 0.640 mol), 2-(N,N-di-tert-butoxycarbonyl)aminopyrimidine-5-boronic acid pinacol ester (350 g, 0.831 mol), Pd(dppf)Cl ₂ (4.7 g, 0.0064mol), K ₂ CO ₃ (180 grams, 1.28mol) were added to the mixed solution of 1,4-dioxane (2 liters) and H ₂ O (0.2 liters), then heated to 80°C and stirred for 2 Hour. Thin-layer silica gel chromatography showed that the reaction was complete. The temperature of the reaction solution was lowered to room temperature, and saturated ammonium chloride solution (2 L) and ethyl acetate (2 L) were added for extraction. The organic phase was dried over anhydrous sodium sulfate and concentrated under reduced pressure, and the resulting crude product was subjected to silica gel column chromatography separation and purification (eluent: petroleum ether, ethyl acetate/petroleum ether (1/1), ethyl acetate) to obtain a yellow solid 160g, yield 45%.

MS: [M+1] ⁺ = 557.2

¹ H NMR (400MHz, DMSO-d ₆ ) δ: 10.4(s,1H), 9.67(s,2H), 3.98-4.03(m,4H), 3.77-3.81(m,4H), 2.73(s,3H ),1.43(s,18H).

(2), preparation of compound 1b: Dissolve compound 1a (500 mg, 0.9 mmol) in dichloromethane (5 ml), then add 1-(α-fluoropropionyl) piperazine (1.44 g, 9 mmol) and acetic acid (54 mg). The reaction solution was stirred at room temperature for half an hour and then sodium cyanoborohydride (226 mg, 3.6 mmol) was added. The reaction was continued to stir at room temperature for 12 hours. 5 ml of saturated aqueous sodium bicarbonate solution was added, and the aqueous phase was extracted twice with 5 ml of ethyl acetate. The organic phases were combined, dried over anhydrous Na ₂ SO ₄ and concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: petroleum ether/ethyl acetate=2/1) to obtain 300 mg of a white solid product, Yield 47%.

MS: [M+1] ⁺ = 701.3

¹ H NMR (400MHz, DMSO-d ₆ ) δ: 9.73(s, 2H), 5.21 and 5.33(dq, J=48.2, 6.6Hz, 1H), 4.09-4.01(m, 4H), 3.94-3.87(m ,4H),3.84(s,2H),3.60-3.80(m,4H),2.50-2.65(m,4H),2.44(s,3H),1.55and 1.62(dd,J＝27.0,6.6Hz,3H ),1.47(s,18H).

(3) Preparation of Compound 1: Compound 1b (300 mg, 0.43 mmol) was dissolved in methanol (3 mL), and then hydrochloric acid (37%, 1 mL) was added at 0°C. The reaction was stirred overnight at room temperature. When thin-layer silica gel chromatography showed that the reaction was complete, cold saturated aqueous sodium bicarbonate (20 mL) was added dropwise to the reaction solution and extracted three times with 10 mL of ethyl acetate. The combined organic phases were dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by preparative liquid chromatography and lyophilized to obtain 60 mg of a white solid product with a yield of 30%.

MS: [M+1] ⁺ = 501.2

¹ H NMR (400MHz, CDCl ₃ )δ:9.33(s,2H),5.51(s,2H),5.30and 5.20(dq,J=6.6,48.4Hz,1H),4.00-4.04(m,4H), 3.80-3.95(m,6H),3.60-3.80(m,4H),2.55-2.65(m,4H),2.42(s,3H),1.60 and 1.54(dd,J＝6.6,24.6Hz,3H).

Embodiment two: the preparation of compound 2:

Under argon protection, starting material 13 (50 mg, 0.1 mmol) was dissolved in dichloromethane (1 mL), followed by dropwise addition of 4 drops of DAST. After the reaction solution was stirred at room temperature for 5 hours, the reaction solution was poured into 10 ml of saturated aqueous sodium bicarbonate solution and extracted twice with 20 ml of ethyl acetate. The combined organic phases were dried over anhydrous Na ₂ SO ₄ and concentrated under reduced pressure. The residue was purified by preparative liquid chromatography to obtain 31 mg of white solid.

MS: [M+1] ⁺ = 501.2

¹ H NMR (400MHz,DMSO-d ₆ )δ:9.16(s,2H),7.10(s,2H),5.57and 5.55(dq,J＝6.6,48.4Hz,1H),3.90-4.00(m,4H ),3.86(s,2H),3.70-3.80(m,4H),3.30-3.60(m,8H),2,35(s,3H),1.42and 1.36(dd,J＝6.6,24.8Hz,3H ).

Embodiment three: the preparation of compound 3

Under argon protection, compound 1 (200 mg, 0.4 mmol) and Lawson's reagent (170 mg, 0.42 mmol) were dissolved in anhydrous THF (12 mL) and refluxed overnight. The reaction solution was concentrated under reduced pressure, and the residue was purified by preparative liquid chromatography and freeze-dried to obtain 80 mg of a white product with a yield of 39%.

MS: [M+1] ⁺ = 517.2

¹ H NMR (400MHz, CDCl ₃ ) δ: 9.34(s, 2H), 5.80 and 5.67(dq, J=6.8, 49.9Hz, 1H), 5.49(s, 2H), 4.25-4.50(m, 2H), 3.95-4.15(m,5H),3.75-3.95(m,7H),2.60-2.80(m,4H),2.42(s,3H),1.72and 1.66(dd,J＝6.8,24.7Hz,3H).

Embodiment four: the preparation of compound 4

(1), Preparation of compound 4a: Dissolve compound 1a (150mg, 0.27mmol) in dichloromethane (3ml), then add 1-(2-fluoro-2-methylpropionyl)piperazine (469.48mg , 2.69mmol) and acetic acid (16mg). The reaction solution was stirred at room temperature for half an hour and then sodium cyanoborohydride (68 mg, 1.08 mmol) was added. The reaction was continued to stir at room temperature for 12 hours. 10 ml of saturated aqueous sodium bicarbonate solution was added, and the aqueous phase was extracted twice with 10 ml of ethyl acetate. The organic phases were combined, dried over anhydrous Na ₂ SO ₄ and concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: petroleum ether/ethyl acetate=2/1) to obtain 100 mg of a white solid product, Yield 52%.

MS: [M+1] ⁺ = 715.3

¹ H NMR (400MHz, CDCl ₃ )δ:9.72(s,2H),3.90–4.00(m,4H),3.75–3.90(m,8H),3.70(s,2H),2.55-2.70(m,4H ), 2.43(s,3H), 1.59 and 1.64(d,J=21.8Hz,6H),1.46(s,18H).

(2) Preparation of Compound 4: Compound 4a (100 mg, 0.14 mmol) was dissolved in methanol (3 mL), and then hydrochloric acid (37%, 1 mL) was added at 0°C. The reaction was stirred overnight at room temperature. When thin-layer silica gel chromatography showed that the reaction was complete, cold saturated aqueous sodium bicarbonate (20 mL) was added dropwise to the reaction solution and extracted three times with 10 mL of ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by preparative liquid chromatography and lyophilized to obtain 53 mg of a white solid product with a yield of 74%.

MS: [M+1] ⁺ = 515.2

¹ H NMR (400MHz, CDCl ₃ )δ:9.33(s,2H),5.49(brs,2H),4.03(m,4H),3.88(m,4H),3.82(s,2H),3.60-3.75( m,4H),2.55-2.65(m,4H),2.42(s,3H),1.64and 1.59(d,J＝21.8Hz,6H).

Embodiment five: the preparation of compound 5

(1), Preparation of Compound 5a: Dissolve Compound 1a (150mg, 0.27mmol) in dichloromethane (3ml), then add 1-(2,2-difluoropropionyl)piperazine (479mg, 2.69mmol ) and acetic acid (16mg). The reaction solution was stirred at room temperature for half an hour and then sodium cyanoborohydride (68 mg, 1.08 mmol) was added. The reaction was continued to stir at room temperature for 12 hours. 10 ml of saturated aqueous sodium bicarbonate solution was added, and the aqueous phase was extracted twice with 10 ml of ethyl acetate. The organic phases were combined, dried over anhydrous Na ₂ SO ₄ and concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: petroleum ether/ethyl acetate=2/1) to obtain 105 mg of a white solid product, Yield 54%.

MS: [M+1] ⁺ = 719.3

¹ H NMR (400MHz, CDCl ₃ )δ:9.73(s,2H),4.00-4.10(m,4H),3.75-3.95(m,8H),3.72(s,2H),2.55-2.70(m,4H ),2.44(s,3H),1.84(t,J=19.9Hz,3H),1.47(s,18H).

(2) Preparation of Compound 5: Compound 5a (105 mg, 0.15 mmol) was dissolved in methanol (3 mL), and then hydrochloric acid (37%, 1 mL) was added at 0°C. The reaction was stirred overnight at room temperature. When thin-layer silica gel chromatography showed that the reaction was complete, cold saturated aqueous sodium bicarbonate (20 mL) was added dropwise to the reaction solution and extracted three times with 10 mL of ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by preparative liquid chromatography and lyophilized to obtain 57 mg of a white solid product with a yield of 75%.

MS: [M+1] ⁺ = 519.2

¹ H NMR (400MHz, CDCl ₃ )δ:9.33(s,2H),5.29(brs,2H),3.90-4.10(m,4H),3.70-3.90(m,4H),3.82(s,2H), 3.60-3.80(m,4H),2.58-2.64(m,4H),2.42(s,3H),1.84(t,J＝19.9Hz,3H).

Embodiment six: the preparation of compound 6

(1), Preparation of Compound 6a: Dissolve Compound 1a (150mg, 0.27mmol) in dichloromethane (3ml), then add 1-(2,2,2-trifluoroacetyl)piperazine (489mg, 2.69mmol) and acetic acid (16mg). The reaction solution was stirred at room temperature for half an hour and then sodium cyanoborohydride (68 mg, 1.08 mmol) was added. The reaction was continued to stir at room temperature for 12 hours. 10 ml of saturated aqueous sodium bicarbonate solution was added, and the aqueous phase was extracted twice with 10 ml of ethyl acetate. The organic phases were combined, dried over anhydrous Na ₂ SO ₄ and concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: petroleum ether/ethyl acetate=3/1) to obtain 100 mg of a white solid product, Yield 51%.

MS: [M+1] ⁺ = 723.3

¹ H NMR (400MHz, CDCl ₃ )δ:9.73(s,2H),4.00-4.10(m,4H),3.80-3.95(m,6H),3.76(s,2H),3.60-3.70(m,2H ),2.60-2.70(m,4H),2.44(s,3H),1.48(s,18H).

(2) Preparation of Compound 6: Compound 6a (100 mg, 0.14 mmol) was dissolved in methanol (3 mL), and then hydrochloric acid (37%, 1 mL) was added at 0°C. The reaction was stirred overnight at room temperature. When thin-layer silica gel chromatography showed that the reaction was complete, cold saturated aqueous sodium bicarbonate solution (20 ml) was added dropwise to the reaction solution and extracted three times with 10 ml of ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by preparative liquid chromatography and lyophilized to obtain 65 mg of a white solid product with a yield of 86%.

MS: [M+1] ⁺ = 523.2

¹ H NMR (400MHz, CDCl ₃ )δ:9.32(s,2H),5.41(brs,2H),3.90-4.10(m,4H),3.75-3.90(m,4H),3.84(s,2H), 3.60-3.80(m,4H),2.60-2.70(m,4H),2.42(s,3H).

Embodiment seven: the preparation of compound 7

(1), the preparation of compound 7a: 1-Boc-piperazine (9.01g, 48.35mmol) and triethylamine (9.79g, 96.71mmol) were dissolved in dichloromethane (30mL) and cooled to 0 ℃, then drop Add a solution of 2,2-difluorobutyryl chloride (3.4 g, 23.8 mmol) in dichloromethane (20 mL). The reaction was warmed to room temperature and stirring was continued for 1.5 hours. 30 mL of water was added dropwise to the reaction solution, the organic phase was separated, and the aqueous phase was extracted twice with 20 mL of dichloromethane. The organic phases were combined, dried over anhydrous Na ₂ SO ₄ and concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: ethyl acetate/petroleum ether=1/10) to obtain 4.9 g of a white solid product, Yield 69%.

¹ H NMR (400MHz, CDCl ₃ )δ:3.67-3.72(m,2H),3.58-3.63(m,2H),3.43-3.50(m,4H),2.08-2.24(m,2H),1.47(s ,9H),1.07(t,J=7.44Hz,3H).

(2) Preparation of compound 7b: Compound 7a (3 g, 10.3 mmol) was dissolved in dichloromethane (10 mL). Trifluoroacetic acid (10 mL) was added dropwise at 0° C., and the reaction solution was stirred at room temperature for 1.5 hours after the drop was completed. When thin-layer silica gel chromatography showed that the reaction was complete, 10 mL of saturated cold NaHCO ₃ aqueous solution was added to the reaction solution, and the aqueous phase was extracted 3 times with 10 mL of dichloromethane. The organic phases were combined, washed once with 20 mL of saturated brine, dried over anhydrous Na ₂ SO ₄ , concentrated under reduced pressure, and dried in vacuo. The obtained yellow oil was directly used in the next reaction without further purification. .

MS: [M+1] ⁺ = 193.1

(3) Preparation of compound 7c: Compound 1a (150 mg, 0.27 mmol) was dissolved in dichloromethane (3 ml), then 7b (516 mg, 2.69 mmol) and acetic acid (16 mg) were added. The reaction solution was stirred at room temperature for half an hour and then sodium cyanoborohydride (68 mg, 1.08 mmol) was added. The reaction was continued to stir at room temperature for 12 hours. 10 ml of saturated aqueous sodium bicarbonate solution was added, and the aqueous phase was extracted twice with 10 ml of ethyl acetate. The organic phases were combined, dried over anhydrous Na ₂ SO ₄ and concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: petroleum ether/ethyl acetate=3/1) to obtain 100 mg of a white solid product, Yield 50%.

MS: [M+1] ⁺ = 733.3

¹ H NMR (400MHz, CDCl ₃ )δ:9.72(s,2H),4.07-4.03(m,4H),3.90-3.86(m,4H),3.83(s,2H),3.77(m,2H), 3.70(m,2H),2.60(m,4H),2.43(s,3H),2.05-2.30(m,2H),1.46(s,18H),1.07(t,J＝7.4Hz,3H).

(4) Preparation of compound 7: Compound 7c (100 mg, 0.14 mmol) was dissolved in methanol (3 mL), and then hydrochloric acid (37%, 1 mL) was added at 0°C. The reaction was stirred overnight at room temperature. When thin-layer silica gel chromatography showed that the reaction was complete, cold saturated aqueous sodium bicarbonate (20 mL) was added dropwise to the reaction solution and extracted three times with 10 mL of ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by preparative liquid chromatography and lyophilized to obtain 54 mg of a white solid product with a yield of 74%.

MS: [M+1] ⁺ = 533.2

¹ H NMR (400MHz, CDCl ₃ )δ:9.32(s,2H),5.27(brs,2H),3.95-4.05(m,4H),3.83-3.90(m,4H),3.82(s,2H), 3.65-3.81(m,4H),2.58-2.61(m,4H),2.41(s,3H),2.05-2.35(m,2H),1.07(t,J＝7.4Hz,3H).

Embodiment eight: the preparation of compound 8

(1), preparation of compound 8a: 1-Boc-piperazine (10.74g, 57.65mmol) and triethylamine (11.67g, 115.30mmol) were dissolved in dichloromethane (30mL) and cooled to 0°C, then drop Add a solution of 1-fluorocyclopropanoyl chloride (3.5 g, 28.8 mmol) in dichloromethane (20 mL). The reaction was warmed to room temperature and stirring was continued for 1.5 hours. 30 mL of water was added dropwise to the reaction solution, the organic phase was separated, and the aqueous phase was extracted twice with 20 mL of dichloromethane. The organic phases were combined, dried over anhydrous Na ₂ SO ₄ and concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: ethyl acetate/petroleum ether=1/6) to obtain 4.2 g of a white solid product, Yield 54%.

¹ H NMR (400MHz, CDCl ₃ ) δ: 3.50-3.75(m,4H), 3.30-3.50(m,4H), 1.40(s,9H), 1.10–1.30(m,4H).

(2) Preparation of compound 8b: Compound 8a (3 g, 11.2 mmol) was dissolved in dichloromethane (10 mL). Trifluoroacetic acid (10 mL) was added dropwise at 0° C., and the reaction solution was stirred at room temperature for 1.5 hours after the drop was completed. When thin-layer silica gel chromatography showed that the reaction was complete, 20 mL of saturated cold NaHCO ₃ aqueous solution was added to the reaction solution, and the aqueous phase was extracted 3 times with 10 mL of dichloromethane. The organic phases were combined, washed once with 20 mL of saturated brine, dried over anhydrous Na ₂ SO ₄ , concentrated under reduced pressure, and dried in vacuo. The obtained yellow oil was directly used in the next reaction without further purification.

MS: [M+1] ⁺ = 173.1

(3) Preparation of compound 8c: Compound 1a (150mg, 0.27mmol) was dissolved in dichloromethane (3ml), then 8b (462mg, 2.69mmol) and acetic acid (16mg) were added. The reaction solution was stirred at room temperature for half an hour and then sodium cyanoborohydride (68 mg, 1.08 mmol) was added. The reaction was continued to stir at room temperature for 12 hours. 10 ml of saturated aqueous sodium bicarbonate solution was added, and the aqueous phase was extracted twice with 10 ml of ethyl acetate. The organic phases were combined, dried over anhydrous Na ₂ SO ₄ and concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: petroleum ether/ethyl acetate=2/1) to obtain 98 mg of a white solid product, Yield 51%.

MS: [M+1] ⁺ = 713.3

¹ H NMR (400MHz, CDCl ₃ )δ:9.73(s,2H),4.00-4.10(m,4H),3.60-3.90(m,10H),2.58-2.66(m,4H),2.45(s,3H ),1.47(s,18H),1.15-1.34(m,4H).

(4) Preparation of Compound 8: Compound 8c (98 mg, 0.14 mmol) was dissolved in methanol (3 mL), and then hydrochloric acid (37%, 1 mL) was added at 0°C. The reaction was stirred overnight at room temperature. When thin-layer silica gel chromatography showed that the reaction was complete, cold saturated aqueous sodium bicarbonate (20 mL) was added dropwise to the reaction solution and extracted three times with 10 mL of ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by preparative liquid chromatography and lyophilized to obtain 53 mg of a white solid product with a yield of 75%.

MS: [M+1] ⁺ = 513.2

¹ H NMR (400MHz, CDCl ₃ )δ: 9.33(s,2H),5.53(brs,2H), 4.00-4.10(m,4H),3.85-3.90(m,4H),3.84(s,2H), 3.65-3.80(m,4H),2.60-2.65(m,4H),2.42(s,3H),1.15-1.40(m,4H).

Embodiment nine: the preparation of compound 9

(1), Preparation of compound 9a: 1-fluorocyclohexylcarboxylic acid (1g, 6.84mmol), 1-Boc-piperazine (2.55g, 13.68mmol), EDCI hydrochloride (1.57g, 8.21mmol), DMAP (1.67g, 13.68mmol) was dissolved in dichloromethane (50mL), and the reaction solution was stirred at room temperature for 10 hours. The reaction solution was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: ethyl acetate/petroleum ether=1/10) to obtain 1.3 g of a white solid product with a yield of 60%.

¹ H NMR (400MHz, CDCl ₃ )δ:3.7-3.80(m,2H),3.50-3.70(m,2H),3.4-0-3.49(m,4H),1.70-2.00(m,4H),1.55 -1.70(m,5H),1.47(s,9H),1.20-1.40(m,1H).

(2) Preparation of compound 9b: Compound 9a (1.3 g, 4.3 mmol) was dissolved in dichloromethane (6 mL). Trifluoroacetic acid (6 mL) was added dropwise at 0° C., and the reaction solution was stirred at room temperature for 1.5 hours after the drop was completed. When thin-layer silica gel chromatography showed that the reaction was complete, 10 mL of saturated cold NaHCO ₃ aqueous solution was added to the reaction solution, and the aqueous phase was extracted 3 times with 10 mL of dichloromethane. The organic phases were combined, washed once with 20 mL of saturated brine, dried over anhydrous Na ₂ SO ₄ , concentrated under reduced pressure, and dried in vacuo. The obtained yellow oil was directly used in the next reaction without further purification.

MS: [M+1] ⁺ = 215.2

(3) Preparation of compound 9c: Compound 1a (150mg, 0.27mmol) was dissolved in dichloromethane (3ml), then 9b (577mg, 2.69mmol) and acetic acid (16mg) were added. The reaction solution was stirred at room temperature for half an hour and then sodium cyanoborohydride (68 mg, 1.08 mmol) was added. The reaction was continued to stir at room temperature for 12 hours. 10 ml of saturated aqueous sodium bicarbonate solution was added, and the aqueous phase was extracted twice with 10 ml of ethyl acetate. The organic phases were combined, dried over anhydrous Na ₂ SO ₄ and concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: petroleum ether/ethyl acetate=2/1) to obtain 110 mg of a white solid product, Yield 54%.

MS: [M+1] ⁺ = 755.4

¹ H NMR (400MHz, CDCl ₃ )δ:9.73(s,2H),4.00-4.10(m,4H),3.87-3.91(m,4H),3.83(s,2H),3.55-3.80(m,4H ),2.50-2.63(m,4H),2.44(s,3H),1.80-2.01(m,4H),1.55-1.80(m,5H),1.47(s,18H),1.20-1.40(m,1H ).

(4) Preparation of Compound 9: Compound 9c (110 mg, 0.15 mmol) was dissolved in methanol (3 mL), and then hydrochloric acid (37%, 1 mL) was added at 0°C. The reaction was stirred overnight at room temperature. When thin-layer silica gel chromatography showed that the reaction was complete, cold saturated aqueous sodium bicarbonate (20 mL) was added dropwise to the reaction solution and extracted three times with 10 mL of ethyl acetate. The combined organic phases were dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by preparative liquid chromatography and lyophilized to obtain 43 mg of a white solid product with a yield of 39%.

MS: [M+1] ⁺ = 555.2

¹ H NMR (400MHz, CDCl ₃ )δ:9.33(s,2H),5.38(brs,2H),4.00-4.08(m,4H),3.82-3.90(m,6H),3.82(s,2H), 3.60-3.75(m,2H),2.55-2.65(m,4H),2.42(s,3H),1.80-2.00(m,4H),1.50-1.75(m,5H),1.20-1.35(m,1H ).

Embodiment 10: Preparation of compound 10

(1), preparation of compound 10a: 4-fluoropiperidine-4-carboxylate hydrochloride (2.00g, 10.89mmol) was dissolved in tetrahydrofuran (27mL) and water (27mL), then CbzCl (2.23g, 13.07mmol) and K ₂ CO ₃ (3.76g, 27.23), the reaction solution was stirred at room temperature for 3 hours. When thin-layer silica gel chromatography showed that the reaction was complete, the pH was adjusted to 5 with 1M dilute hydrochloric acid, and then extracted three times with 20 ml of ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and dried in vacuo. The obtained colorless oil was directly used in the next reaction without purification.

MS: [M-1] ^- = 280.1

(2), the preparation of compound 10b: 10a (3g, 10.89mmol), 1-Boc-piperazine (4.07g, 21.83mmol), EDCI hydrochloride (2.51g, 13.10mmol), DMAP (2.67g, 21.83 mmol) was dissolved in dichloromethane (50 mL), and the reaction solution was stirred at room temperature for 10 hours. The reaction solution was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: ethyl acetate/petroleum ether=1/10) to obtain 2.7 g of a white solid product with a yield of 55%.

¹ H NMR (400MHz, CDCl ₃ )δ:7.28-7.45(m,5H),5.17(s,2H),3.95-4.20(m,2H),3.55-3.60(m,4H),3.40-3.50(m ,4H),3.15-3.30(m,2H),1.80-2.30(m,4H),1.47(s,9H).

(3) Preparation of compound 10c: Compound 10b (2.7 g, 6.01 mmol) was dissolved in dichloromethane (10 mL). Trifluoroacetic acid (10 mL) was added dropwise at 0° C., and the reaction solution was stirred at room temperature for 1.5 hours after the drop was completed. When thin-layer silica gel chromatography showed that the reaction was complete, 20 mL of saturated NaHCO ₃ aqueous solution was added to the reaction solution, and the aqueous phase was extracted three times with 20 mL of dichloromethane. The organic phases were combined, washed once with 30 mL of saturated brine, dried over anhydrous Na ₂ SO ₄ , concentrated under reduced pressure, and dried in vacuo. The obtained yellow oil was directly used in the next reaction without further purification.

MS: [M+1] ⁺ = 350.2

(4) Preparation of compound 10d: Compound 1a (270.00 mg, 0.49 mmol) was dissolved in dichloromethane (5 ml), and then 10c (1.6 g, 4.85 mmol) and acetic acid (29 mg) were added. After the reaction solution was stirred at room temperature for half an hour, sodium cyanoborohydride (122 mg, 1.96 mmol) was added. The reaction was continued to stir at room temperature for 12 hours. 20 ml of saturated aqueous sodium bicarbonate solution was added, and the aqueous phase was extracted twice with 20 ml of ethyl acetate. The organic phases were combined, dried over anhydrous Na ₂ SO ₄ and concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: petroleum ether/ethyl acetate=2/1) to obtain 200 mg of a white solid product, Yield 46%.

MS: [M+1] ⁺ = 890.4

¹ H NMR (400MHz, CDCl ₃ )δ:9.73(s,2H),7.28-7.45(m,5H),5.14(s,2H),4.00-4.20(m,6H),3.60-3.90(m,10H ), 3.10-3.25(m,2H), 2.55-2.65(m,4H), 2.44(s,3H), 2.05-2.40(m,4H), 1.47(s,18H).(5), compound 10e Preparation: Compound 10d (200 mg, 0.22 mmol) was dissolved in methanol (3 mL), followed by addition of hydrochloric acid (37%, 1 mL) at 0°C. The reaction was stirred overnight at room temperature. When thin-layer silica gel chromatography showed that the reaction was complete, cold saturated aqueous sodium bicarbonate (20 mL) was added dropwise to the reaction solution and extracted three times with 10 mL of ethyl acetate. The organic phases were combined, washed once with 20 ml of saturated brine, dried over anhydrous sodium sulfate, concentrated under reduced pressure, and dried in vacuo to obtain 170 mg of a white crude product that was directly used in the next reaction without purification.

MS: [M+1] ⁺ = 690.3

(6) Preparation of Compound 10: Compound 10e (170 mg, 0.22 mmol) was dissolved in methanol (5 mL), 15% Pd/C (34 mg) was added, and hydrogenation was carried out at room temperature for 12 hours. The reaction solution was filtered with diatomaceous earth, the filtrate was concentrated under reduced pressure, and the residue was purified by preparative liquid chromatography and freeze-dried to obtain 20 mg of a white solid product with a yield of 16%.

MS: [M+1] ⁺ = 556.2

¹ H NMR (400MHz, CDCl ₃ )δ:9.32(s,2H),5.33(brs,2H),3.95-4.15(m,4H),3.75-3.90(m,8H),3.60-3.75(m,2H ),2.90-3.10(m,3H),2.50-2.65(m,4H),2.41(s,3H),1.80-2.00(m,4H),1.70-2.30(m,6H).

Embodiment 11: Preparation of compound 11

(1), Preparation of Compound 11a: Dissolve 2-fluoro-2-phenylacetic acid (1g, 6.49mmol) in dichloromethane (10mL) and cool to 0°C, then add dropwise oxalyl chloride (894.58mg, 7.79 mmol). After the oxalyl chloride was dropped, 1 drop of DMF was added. The temperature of the reaction solution was raised to room temperature and stirred for 1.5 hours to obtain a 2-fluoro-2-phenylacetyl chloride solution, which was directly used in the next reaction without any treatment.

1-Boc-piperazine (2.42g, 12.98mmol) and triethylamine (2.63g, 25.95mmol) were dissolved in dichloromethane (10mL) and cooled to 0°C, then the above 2-fluoro-2-benzene A dichloromethane solution of acetyl chloride (10 ml) was added dropwise to the reaction solution. After the dropwise addition, the reaction solution was stirred at room temperature for 1.5 hours. 30 ml of water was added, and the aqueous phase was further extracted three times with 10 ml of dichloromethane. The organic phases were combined, dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: ethyl acetate/petroleum ether=1/7) to obtain 1.0 g of a white solid product with a yield of 48 %.

MS: [M+1] ⁺ = 323.2

¹ H NMR (400MHz, CDCl ₃ ) δ: 7.37-7.47(m, 5H), 6.09(d, J=49.4 Hz, 1H), 3.00-3.80(m, 8H), 1.43(s, 9H).

(2) Preparation of compound 11b: Compound 11a (1 g, 3.1 mmol) was dissolved in dichloromethane (6 mL). Trifluoroacetic acid (6 mL) was added dropwise at 0° C., and the reaction solution was stirred at room temperature for 1.5 hours after the drop was completed. When thin-layer silica gel chromatography showed that the reaction was complete, 10 mL of saturated cold NaHCO ₃ aqueous solution was added to the reaction solution, and the aqueous phase was extracted 3 times with 10 mL of dichloromethane. The organic phases were combined, washed once with 20 mL of saturated brine, dried over anhydrous Na ₂ SO ₄ , concentrated under reduced pressure, and dried in vacuo. The obtained yellow oil was directly used in the next reaction without further purification.

MS: [M+1] ⁺ = 223.1

(3) Preparation of compound 11c: Compound 1a (150mg, 0.27mmol) was dissolved in dichloromethane (3ml), then 11b (599mg, 2.69mmol) and acetic acid (16mg) were added. The reaction solution was stirred at room temperature for half an hour and then sodium cyanoborohydride (68 mg, 1.08 mmol) was added. The reaction was continued to stir at room temperature for 12 hours. 10 ml of saturated aqueous sodium bicarbonate solution was added, and the aqueous phase was extracted twice with 10 ml of ethyl acetate. The organic phases were combined, dried over anhydrous Na ₂ SO ₄ and concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: petroleum ether/ethyl acetate=2/1) to obtain 118 mg of a white solid product, Yield 57%.

MS: [M+1] ⁺ = 763.3

¹ H NMR (400MHz, CDCl ₃ )δ: 9.72(s, 2H), 7.34-7.49(m, 5H), 6.08(d, J=49.4Hz, 1H), 4.00-4.09(m, 4H), 3.84- 3.93(m,4H),3.76(s,2H),3.70-3.76(m,2H),3.40-3.50(m,2H),2.50-2.60(m,2H),2.39(s,3H),2.25- 2.35(m,2H),1.47(s,18H).

(4) Preparation of compound 11: Compound 11c (118 mg, 0.15 mmol) was dissolved in methanol (3 mL), and then hydrochloric acid (37%, 1 mL) was added at 0°C. The reaction was stirred overnight at room temperature. When thin-layer silica gel chromatography showed that the reaction was complete, cold saturated aqueous sodium bicarbonate (20 mL) was added dropwise to the reaction solution and extracted three times with 10 mL of ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was purified by preparative liquid chromatography and lyophilized to obtain 50 mg of a white solid product with a yield of 57%.

MS: [M+1] ⁺ = 563.2

¹ H NMR (400MHz, CDCl ₃ ) δ: 9.31 (s, 2H), 7.30-7.50 (m, 5H), 6.07 (d, J=49.4Hz, 1H), 5.31 (brs, 2H), 3.95-4.05 ( m,4H),3.80-3.90(m,4H),3.60-3.80(m,4H),3.35-3.50(m,2H),2.50-2.69(m,2H),2.37(s,3H),2.20- 2.37(m,2H).

Embodiment 12: Preparation of compound 12

(1), Preparation of compound 12a: 2,2-difluoro-2-(2-pyridyl)acetic acid (1g, 5.78mmol), 1-Boc-piperazine (2.15g, 11.55mmol), EDCI hydrochloride (1.33g, 6.93mmol), DMAP (1.41g, 11.55mmol) were dissolved in dichloromethane (30mL), and the reaction solution was stirred at room temperature for 10 hours. The reaction solution was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: ethyl acetate/petroleum ether=1/10) to obtain 1.0 g of a white solid product with a yield of 51%.

¹ H NMR (400MHz, CDCl ₃ )δ: 8.60(d, J=4.4Hz, 1H), 7.88(m, 1H), 7.74(d, J=7.9Hz, 1H), 7.44(m, 1H), 3.65 -3.72(m,2H),3.55-3.62(m,2H),3.45-3.51(m,2H),3.32-3.38(m,2H),1.46(s,9H).

(2) Preparation of compound 12b: Compound 12a (1.0 g, 2.93 mmol) was dissolved in dichloromethane (5 mL). Trifluoroacetic acid (5 mL) was added dropwise at 0° C., and the reaction solution was stirred at room temperature for 1.5 hours after the drop was completed. When thin-layer silica gel chromatography showed that the reaction was complete, 20 mL of saturated cold NaHCO ₃ aqueous solution was added to the reaction liquid, and extracted 3 times with 20 mL of dichloromethane. The organic phases were combined, washed once with 30 mL of saturated brine, dried over anhydrous Na ₂ SO ₄ , concentrated under reduced pressure, and dried in vacuo. The obtained yellow oil was directly used in the next reaction without further purification.

MS: [M+1] ⁺ = 242.1

(3) Preparation of compound 12c: Compound 1a (150 mg, 0.27 mmol) was dissolved in dichloromethane (3 mL), followed by addition of 12b (325 mg, 1.35 mmol) and acetic acid (16 mg). The reaction solution was stirred at room temperature for half an hour and then sodium cyanoborohydride (68 mg, 1.08 mmol) was added. The reaction was continued to stir at room temperature for 12 hours. 10 ml of saturated aqueous sodium bicarbonate solution was added, and the aqueous phase was extracted twice with 10 ml of ethyl acetate. The organic phases were combined, dried over anhydrous Na ₂ SO ₄ and concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: petroleum ether/ethyl acetate=2/1) to obtain 50 mg of a white solid product, Yield 26%.

MS: [M+1] ⁺ = 782.3

¹ H NMR (400MHz, CDCl ₃ ) δ: 9.72(s, 2H), 8.66(d, J=4.7Hz, 1H), 7.87(m, 1H), 7.72(d, J=7.9Hz, 1H), 7.43 (m,1H),4.02-4.08 (m,4H),3.86-3.91(m,4H),3.81(s,2H),3.70-3.80(m,2H),3.60-3.66(m,2H),2.61 (m, 2H), 2.48 (m, 2H), 2.42 (s, 3H), 1.47 (s, 18H). (4), preparation of compound 12: compound 12c (50 mg, 0.06 mmol) was dissolved in methanol ( 3 mL), then hydrochloric acid (37%, 1 mL) was added at 0°C. The reaction was stirred overnight at room temperature. When thin-layer silica gel chromatography showed that the reaction was complete, cold saturated aqueous sodium bicarbonate (20 mL) was added dropwise to the reaction solution and extracted three times with 10 mL of ethyl acetate. The organic phases were combined, dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by preparative liquid chromatography and lyophilized to obtain 20 mg of a white solid product with a yield of 54%.

MS: [M+1] ⁺ = 582.2

¹ H NMR (400MHz, CDCl ₃ ) δ: 9.32(s, 2H), 8.66(d, J=4.4Hz, 1H), 7.80-7.90(m, 1H), 7.71(d, J=7.9Hz, 1H) ,7.43(m,1H),5.36(brs,2H),3.95-4.10(m,4H),3.80-3.90(m,4H),3.70-3.80(m,4H),3.55-3.65(m,2H) ,2.55-2.60(m,2H),2.40-2.50(m,2H),2.40(s,3H).

Example 13: Calculation of compound clogP

The clogP value refers to the logarithmic value of the ratio of the partition coefficient of a substance in n-octanol (oil) and water. It reflects the distribution of substances in the oil-water two-phase. The larger the clogP value, the more lipophilic the substance is, on the contrary, the smaller the value, the more hydrophilic, that is, the better the water solubility. The dissolution, absorption, distribution, and transport of drugs in the body are related to the water solubility and fat solubility of the drug, that is, the oil-water partition coefficient clogP. Using the online tool provided by VCCLAB (http://www.vcclab.org/), the calculation results are shown in Table 1:

Table 1. clogP of compounds 1-12

The data obtained from the calculation results show that the clogP corresponding to the compounds 1-12 is between 1-5, and has a good ester-water distribution coefficient.

Example 14: Determination of Compound PI3K Kinase Activity

The purpose of the experiment: to test the inhibitory activity of the compound on PI ₃ Kα/β/δ/γ kinase by ADP-Glo luminescence method

Experimental method: The highest concentration of the compound is set to 10uM, diluted 4 times downward with DMSO, a total of 10 concentrations: 10uM, 2.5uM, 0.625uM, 0.156uM, 0.039uM, 0.0098uM, 0.0024uM, 0.0006uM, 0.00015uM, 0.000038 uM. Add the corresponding PI3Kα/β/δ/γ kinase solution to each well of the test plate containing the compound DMSO solution, mix well; add the PIP2 solution and ATP solution to each well to start the kinase reaction, and incubate at room temperature for 1h . Add the ADPGlo reagent equilibrated to room temperature into the wells to terminate the kinase reaction, centrifuge and mix slowly on a shaker, and equilibrate for 120 minutes. Add kinase test reagent to each well, mix well and equilibrate for 30 minutes, record the reading. Data curves were plotted using XLFit and _IC50 values were calculated. The results are shown in Table 2.

The experimental results are shown in the table below:

Table 2. IC ₅₀ data of four isoforms of PI3K for compounds 1-12 and other compounds

The results showed that for the tested PI3Kα/β/δ/γ kinase inhibitory activities, compounds 1-12 showed good activity on the four subtypes of PI3K and were stronger than apelixed. It is particularly worth mentioning that compound 12 is the strongest inhibitor of PI3Kα found so far. Compared with apelixed, the inhibitory activity of compound 12 on PI3Kα is 31 times that of apelixed, and its inhibitory activity on PI3Kβ It is 29 times that of Apirised; the inhibitory activity on PI3Kδ is 8 times that of Apirised,

Example 15: Compounds inhibit the proliferation of tumor cells:

The purpose of the experiment: to use the MTT method to verify the effect of the compound on MCF-7 cells (human breast cancer cells), NCI-H460 cells (human large cell lung cancer cells), HCT116 cells (human colon cancer cells), PC-3 cells (human prostate cancer cells) , HeLa cell (human cervical cancer cell) proliferation inhibitory toxicity.

Experimental method: The initial concentration of the sample is 5μM, and it is diluted 4 times downwards sequentially to obtain 10 concentrations: 5000nM, 1250nM, 312nM, 78nM, 19.5nM, 4.9nM, 1.2nM, 0.30nM, 0.076nM, 0.019nM. Take the cancer cells in the logarithmic growth phase, count them under a microscope, adjust the cell concentration to 7× ¹⁰⁴ /mL with the corresponding complete culture solution, plant them in a 96-well culture plate, 100 μL/well, and place at 37°C, 5% CO ₂ incubator culture 24h. Absorb the culture medium with a syringe (suspended cells are centrifuged and then sucked out the culture medium, and the adherent cells are sucked directly), and then add 200 μL/well of culture medium containing different concentrations of test samples, and directly add 200 μL of complete culture medium to the blank control well , with 3 replicate wells for each concentration. After adding the drug, the cells were incubated for 72 hours at 37°C in a 5% CO ₂ incubator. The culture medium was sucked off with a syringe (suspended cells were centrifuged and the culture medium was sucked off, and the adherent cells were sucked directly), and then 200 μL of freshly prepared complete culture medium containing 10% MTT was added, and the culture was continued for 4 hours. Aspirate the upper layer culture solution, add 150 μL DMSO to each well, shake in the dark for 10 min, and measure the OD value of each well at a wavelength of 490 nm. _IC50 values were calculated using SPSS17.0 software.

The experimental results are shown in Table 3:

Table 3. IC ₅₀ data of compounds 1-12 and other compounds on various tumor cell proliferation inhibition

The results showed that for the 5 tested tumor cells, the IC ₅₀ of the in vitro tumor cell proliferation inhibitory activity of compounds 1-12 were all lower than 0.2 μM, and the tumor cell proliferation inhibitory activity was stronger than that of apelixed. Among them, the IC ₅₀ of compound 12 on the five tested tumor cells was all lower than 50nM, which was 9-20 times of the activity of apelixed. It has a very high application prospect.

The above-mentioned embodiments only express the implementation manner of the present invention, and the description thereof is relatively specific and detailed, but should not be construed as limiting the patent scope of the present invention. It should be noted that, for those skilled in the art, several modifications and improvements can be made without departing from the concept of the present invention, and these all belong to the protection scope of the present invention. Therefore, the protection scope of the patent for the present invention should be based on the appended claims.

Claims

An α-fluoroacylpiperazine derivative, characterized in that: the α-fluoroacylpiperazine derivative with general formula (I) or its isomer, or its pharmaceutically acceptable salt, or its prodrug molecule, its General formula (I) chemical structure is:

In the general formula (I), X=O or S; wherein, the acyl α position of the acylpiperazine contains at least one fluorine atom;

In the general formula (I), R is H, halogen, cyano, 1-15 carbon alkyl, alkenyl, alkynyl or derivatives thereof; or a monocyclic or condensed ring containing 5-22 carbon atoms Aryl or its derivatives; or a 5-8 membered heterocycle or a heterocycle or its derivatives containing 1-4 heteroatoms; or carboxyl or its derivatives; or hydroxyl or its derivatives; or amino or its derivatives; or mercapto or its derivatives; or sulfone or sulfoxide derivatives; or sulfonate or sulfonate; or phosphate or phosphate;

In the general formula (I), R is H, halogen, cyano, 1-15 carbon alkyl, alkenyl, alkynyl or derivatives thereof; or a monocyclic or condensed ring containing 5-22 carbon atoms Aryl or its derivatives; or a 5-8 membered heterocycle or a heterocycle or its derivatives containing 1-4 heteroatoms; or carboxyl or its derivatives; or hydroxyl or its derivatives; or amino or or a mercapto group or a derivative thereof; or a sulfone or sulfoxide derivative; or a sulfonate or a sulfonate; or a phosphate or a phosphate.
A kind of α-fluorinated acylpiperazine derivative according to claim 1, characterized in that: in the general formula (I), the α-position of the acyl group of acylpiperazine is a single optical configuration or a racemate; R 1 and R 2 may be the same or different; in general formula (I), R 1 and R 2 may be connected to form a ring structure.
A kind of α-fluoroacylpiperazine derivative according to claim 1, characterized in that: its preferred structure is as follows:
A pharmaceutical composition, characterized in that it contains a therapeutically effective amount of the α-fluoroacylpiperazine derivative or its isomer, or its pharmaceutically acceptable salt, or its prodrug molecule as claimed in claim 1 and a or multiple pharmaceutically acceptable carriers, diluents or excipients.
The preparation method of a kind of α-fluorinated acylpiperazine derivative described in claim 1, is characterized in that: its reaction equation is as follows:

P 1 is H or an amino protecting group, P 2 is H or an amino protecting group; P 1 and P 2 are not H at the same time;

The amino protecting group is formyl, acetyl, trifluoroacetyl, benzoyl, tert-butoxycarbonyl, benzyloxycarbonyl, 9-fluorenylmethoxycarbonyl, phthaloyl, cyclosuccinyl , 2-biphenyl-2-propoxycarbonyl, p-toluenesulfonyl or trityl;

The reducing agent used in the reductive amination is NaBH 4 , KBH 4 , LiBH 4 , Zn(BH 4 ) 2 , NaBH 3 CN, NaBH(OAc) 3 , borane complex, Bu 3 SnH or PhSiH 4 ;

The catalyst used in the reductive amination is protonic acid or Lewis acid;

The reductive amination reaction solvent is dichloromethane, tetrahydrofuran, methanol, ethanol, isopropanol, 1,2-dichloroethane, ethylene glycol dimethyl ether or bis(ethylene glycol) dimethyl ether;

The acid used for the deprotection is a protonic acid or a Lewis acid;

The base used for the deprotection is an inorganic base or an organic base;

The hydrogen source used in the catalytic reduction deprotection reaction is hydrogen, formic acid or ammonium formate; the catalyst is a metal catalyst;

The solvent used in the deprotection reaction is any one of a protic solvent or an aprotic solvent or a mixture of both;

The thio reagent is P 2 S 5 , Lawson's reagent, 2,4-di(methylthio)-1,3,2,4-dithiadiphosphatidine-2,4-disulfide , 2,4-bis(phenylthio)-1,3-dithio-2,4-diphosphetane-2,4 disulfide, or 2,4-bis(4-phenoxy Phenyl)-1,3,2,4-dithiodiphosphetane-2,4-disulfide; the solvent used in the reaction is any one or both of protic solvents or aprotic solvents the mix of;

The reaction temperature is between -78°C and 180°C.
The preparation method of a kind of α-fluorinated acylpiperazine derivative described in claim 1, is characterized in that: its reaction equation is as follows:

The fluorinated reagent is HF or its salt, SF 4 , diethylaminosulfur trifluoride, bis(2-methoxyethyl)aminosulfur trifluoride, 4-tert-butyl-2,6- Dimethylphenylsulfur trifluoride, pyridine-2-sulfonyl fluoride, bis(2-methoxyethyl)aminosulfur trifluoride, diethylamino)difluorosulfonium tetrafluoroborate, difluoro (4-morpholino)sulfonium tetrafluoroborate, 1,3-bis(2,6-diisopropylphenyl)-2,2-difluoroimidazoline, 4-chloro-N-[(4 -methylphenyl)sulfonyl]-benzenesulfonyl fluoride; the solvent used in the reaction is an aprotic solvent or a mixed solvent, and the reaction temperature is at -78 to 180°C in Celsius.

The solvent used in the reaction is dichloromethane, dichloroethane, tetrahydrofuran, acetonitrile, ethylene glycol dimethyl ether or 1,4-dioxane.
The application of an α-fluoroacylpiperazine derivative or its isomer, or its pharmaceutically acceptable salt, or its prodrug molecule described in claim 1 in the preparation of a drug for treating cancer.
The application according to claim 7, characterized in that: said cancer is brain cancer, glioma, endometrial cancer, ovarian cancer, cervical cancer, breast cancer, colon cancer, lung cancer, prostate cancer, liver cancer, Leukemia, lymphoma, skin cancer, basal cell tumor, hemangioma, uterine cancer, laryngeal cancer, stomach cancer, lip cancer, esophagus cancer, nasopharyngeal cancer, gallbladder cancer, pancreatic cancer, kidney cancer, tongue cancer, bladder cancer, melanoma tumor, lipoma, thyroid cancer, thymus cancer, or bone cancer.
An α-fluoroacylpiperazine derivative or its isomer, or a pharmaceutically acceptable salt thereof, or a prodrug molecule thereof as claimed in claim 1 is used in combination with at least one other anticancer agent in the preparation of a drug for treating cancer application in medicine.
The application according to claim 9, characterized in that: the other anticancer agents are doxorubicin, bleomycin, vinblastine, taxanes, etoposide, 5-fluorouracil, cyclophosphine Amide, methotrexate, cisplatin, tretinoin, temozolomide, actinomycin, imatinib, gefitinib, sorafenib, erlotinib, sunitinib, afatinib, Cabozantinib, Ostinib, Rituximab, Cetuximab, Trastuzumab, Nivolumab, Palivizumab, Atezolizumab, Durvalumab, or Avi Any one of monoclonal antibodies or a combination of two or more.