Method for preparing camptothecin conjugate

Abstract

Provided is a method for preparing a camptothecin conjugate. The new method for preparing a camptothecin conjugate exhibits one or more effects selected from the group consisting of a high yield, a simple purification process, low impurities, suitability for industrial production, and high stability.

Description

Preparation method of a class of camptothecin conjugates Technical Field

[0001] This invention relates to the field of chemical medicine, specifically to a method for preparing a class of camptothecin conjugates. Background Technology

[0002] Camptothecin (CPT) is a cytotoxic alkaloid isolated from the camptotheca tree (Campanula stenoptera), a plant in the Davidiaceae family. It forms a ternary complex with cellular DNA topoisomerase I, thereby inhibiting DNA unwinding, leading to DNA replication arrest and ultimately cell death (Cancer Res. 1989, 49, 6365), exhibiting broad-spectrum antiproliferative activity. However, its low solubility, instability, acquired tumor cell resistance, and significant toxicity make it unsuitable for clinical development. Camptothecin derivatives can increase water solubility and improve drugability by introducing water-soluble groups or preparing prodrugs. Several camptothecin derivatives with significantly improved solubility have been approved for marketing (Med. Res. Rev. 2015, 35, 753), such as topotecan, irinotecan, and beloteccan, for the treatment of various types of cancer.

[0003] Camptothecin derivatives are also used in antibody conjugation as small molecule toxins in antibody-drug conjugates (ADCs), also known as payloads. ADCs combine the high potency of cytotoxic small molecules with the high selectivity of antibodies for specific tumor cells. Compared to traditional chemotherapy drugs, ADCs can more precisely kill tumor cells and reduce the impact on normal cells. In recent years, ADCs using camptothecin derivatives as small molecule toxins have made significant progress. Two camptothecin-based ADCs have been approved for cancer treatment: DS-8101a, where the camptothecin analog dxd is conjugated to the anti-HER2 antibody trastuzumab via a cleavable tetrapeptide-based linker; and Immu-132, where the camptothecin analog SN-38 is conjugated to the anti-Trop-2 antibody cetuzumab via a hydrolyzable pH-sensitive linker.

[0004] However, ADCs using camptothecin-based drugs or derivatives as toxins generally have a high drug-to-antibody ratio (DAR), making their production process difficult and prone to instability. Furthermore, camptothecin compounds often exhibit bone marrow suppression leading to hematologic toxicity, such as neutropenia, leukopenia, thrombocytopenia, and anemia, as well as gastrointestinal side effects such as nausea, vomiting, and diarrhea.

[0005] Therefore, it is crucial to develop novel camptothecin compounds and their conjugates, and to develop stable, high-yield, and scale-up production processes. Summary of the Invention

[0006] The purpose of this disclosure is to provide a novel, industrially applicable method for preparing camptothecin conjugates. The preparation method described above has one or more effects selected from the group consisting of:

[0007] (1) High yield;

[0008] (2) The purification process is simple and has low impurities;

[0009] (3) Suitable for industrial production;

[0010] (4) High stability.

[0011] This disclosure also provides a method for preparing a compound of formula I, comprising the step of reacting a compound of formula Ii with an intermediate compound a via a nucleophilic substitution reaction to obtain the compound of formula I.

[0012] in:

[0013] L2 is -(C(R) L21 )2) n -,

[0014] Any unit CHR in L2 L21 Each of these can be independently replaced by the following structural units: -Cy-, -C(O)-, NR L22 , -O-, -S-, -SO-, SO2, -P(R L22 )-、-(P=O)R L22 -C(=S)-, C(=NR) L22 -N=N-, -C=N-,

[0015] n is a natural number from 0 to 50;

[0016] L3 is any combination of amino acid residues and short peptides consisting of 2-10 amino acid residues, wherein the amino acid residues are natural amino acid residues or non-natural amino acid residues.

[0017] Tr is Or any combination of the above groups;

[0018] "*" indicates a symbol related to L. 3 connect;

[0019] -Cy- is selected from phenylene, 5- to 8-membered heteroaryl, 3- to 10-membered heterocyclic, or 3- to 10-membered cycloalkylene, wherein -Cy- is unsubstituted or independently formed by one or more R- atoms. x replace,

[0020] RL21 R L22 R cx Each is independently selected from hydrogen, deuterium, halogens, -NO2, -CN, and -OR. L2a -SR L2a -N(R) L2a )2、-N + (R L2a 3、-CH2C(O)R L2a --OC(O)R L2a -N(R) L2a SO2R L2b -N(R) L2a )COR L2b --(N(Me)CH2C(O)) m -OR L2a --(N(Me)CH2C(O)) m -NHR L2a -(N(Me)CH2C(O) m -N*(R L2a 3、-(CH2) y -NHCOCH2(OCH2CH2)OR L2a -(CH2) y -NH(COCH2(N(Me) m -R L2a -NH-(CH2CH2O) m -R L2a -(CH2CH2O) m -R L2a -(CH2N(Me)) m -R L2a -CH2(OCH2CH2) m -OR L2a -(CH2CH2O) m -R L2a -NH-(CH2CH2O) m -R L2a or R L2a Optional substitution of -C 1-6 Alkyl, -C 1-6 alkenyl, -C 1-6 Alkyne, 3-8 membered cycloalkyl, 4-10 membered heterocycloalkyl, 6-10 membered aryl or 3-10 membered heteroaryl,

[0021] m and y are each natural numbers from 0 to 50.

[0022] R L2a R L2bEach is independently selected from hydrogen, deuterium, halogens, -NO2, -CN, -OH, -SH, -NH2, -N(Me)2, -CO2H, -S(O)2Me, -S(O)2OH, -C(O)NH2, -SO2NH2, -C 1-6 Alkyl, -C 1-6 alkenyl, -C 1-6 Alkynyl, 3-8 membered cycloalkyl, 4-10 membered heterocycloalkyl, 6-10 membered aryl or 3-10 membered heteroaryl;

[0023] R1 and R2 are each independently selected from H, -OH, -CN, halogen, and C. 1-3 Alkyl, C 1-3 Alkoxy;

[0024] Rx is selected from H, halogen, or C. 1-3 alkyl;

[0025] p can be 0, 1, or 2.

[0026] In some implementations, L3 is selected from Val, D-Val, Phe, Lys, Leu, Ile, Gly, Ala, Cit, Asp, Asn, Glu, Gln, Val-Cit, Val-Ala, Val-Lys, Val-Lys(Ac), Phe-Lys, Phe-Lys(Ac), Leu-Lys, Leu-Lys(Ac), Ala-Ala, Ala-Lys, D-Ala-Ala, Gly-Glu, Gly-Asp, Gly-Asn, Val-Glu, Val-Asp, Asn-Asn, Asp-Glu, Gly-Gly-Glu, Gly-Gly-Asp, Gly-Gly-Asn, Gly-Ala-Ala, Gly-Val-Ala, Gly-Val-Cit, Glu-Val-Cit, Al a-Ala-Ala、Ala-(D-Ala)-Ala、Ala-Ala-Asn、Ala-(D-Ala)-Asn、Ala-Ala-Asp.Val-Lys-Gly , D-Val-Leu-Lys, Gly-Gly-Arg, Gly-Gly-Gly, Lys-Ala-Asn, Gly-Phe-Gly, Gly-Gly-Phe, Asn -Pro-Val, Ala-Lys-Gly, Gly-Lys-Gly, Gly-Gly-Gly-Gly, Gly-Gly-Phe-Gly, Gly-Gly-Glu-Gly, Lys-Ala-Ala-Asn, Lys-Ala-Ala-Asp, Ala-Ala-Pro-Val, Ala-Ala-Pro-Nva, or any combination of the above fragments.

[0027] In some implementations, L3 is selected from Lys, Gly, Asp, Asn, Glu, Gln, Val-Cit, Val-Ala, Ala-Ala, Gly-Glu, Gly-Asp, Gly-Asn, Asn-Asn, Asp-Glu, Gly-Glu-Gly, Gly-Gly-Phe-Gly, or any combination of the above fragments; preferably, L3 is selected from Lys, Gly, Val-Ala, Ala-Ala, Gly-Glu, Gly-Asp, Gly-Asn, Asn-Asn, Asp-Glu, Gly-Gly-Phe-Gly, and more preferably from Val-Ala, Ala-Ala, Gly-Glu, Gly-Asp, Gly-Asn, Asn-Asn, Asp-Glu, Gly-Gly-Phe-Gly.

[0028] In some implementations, L2 is -(C(R) L21 )2) n -,

[0029] Any unit CHR in L2 L21 Each of these can be independently replaced by the following structural units: -Cy-, -C(O)-, NR L22 -O-;

[0030] n is a natural number from 0 to 50;

[0031] -Cy- is selected from phenylene, 5- to 6-membered heteroaryl, 4- to 10-membered heterocyclic, or 3- to 6-membered cycloalkylene, wherein -Cy- is unsubstituted or independently represented by one or more R- atoms. x replace,

[0032] R L21 R L22 R cx Each is independently selected from hydrogen, halogen, -OR L2a -SR L2a -N(R) L2a )2、-N(R L2a SO2R L2b -N(R) L2a )COR L2b -(N(Me)CH2C(O) m -OR L2a --(N(Me)CH2C(O)) m -NHR L2a -(N(Me)CH2C(O) m -N*(R L2a3. -NH-(CH2CH2O) m -R L2a -(CH2CH2O) m -R L2a -(CH2N(Me)) m -R L2a -CH2(OCH2CH2) m -OR L2a -(CH2CH2O) m -R L2a -NH-(CH2CH2O) m -R L2a or R L2a Optional substitution of -C 1-6 Alkyl, -C 1-6 alkenyl, -C 1-6 Alkyne, 3-8 membered cycloalkyl, 4-10 membered heterocycloalkyl, 6-10 membered aryl or 3-10 membered heteroaryl,

[0033] m is a natural integer from 0 to 8.

[0034] y is 0, 1, 2, 3 or 4.

[0035] R L2a R L2b Each is independently selected from hydrogen, halogen, -CN, -OH, -NH2, -N(Me)2, -CO2H, -C(O)NH2, -C 1-6 alkyl.

[0036] In some implementations, L2 is -(CH2). n -,

[0037] In L2, any CH2 unit can be independently replaced by the following structural units: 4-6 membered heterocyclic group, 3-6 membered cycloalkyl group, -C(O)-, NR L22 -O-;

[0038] Each R L22 Each is independently selected from hydrogen, -OR L2a or R L2a Optional substitution of -C 1-6 alkyl,

[0039] Each R L2a Each is independently selected from hydrogen, halogen, -CN, -OH, -NH2, -N(Me)2, -CO2H, -C(O)NH2, -C 1-6 alkyl.

[0040] In some implementations, L2 is

[0041] In some embodiments, the compound Ii is

[0042] In some implementations, compound a is

[0043] In some embodiments, compound a is typically present in the form of a salt; preferably, when the salt of compound a undergoes the nucleophilic substitution reaction, a base is typically added first to obtain the free state of compound a; more preferably, the salt of compound a is a methanesulfonate of compound a, and sodium bicarbonate is typically added first to obtain the free state of compound a during the nucleophilic substitution reaction.

[0044] In some embodiments, the reactants of the nucleophilic substitution reaction further include a base, which is a conventional base used in the art for this type of reaction, such as one or more of DIPEA, N-methylmorpholine, and 2,6-dimethylpyridine, preferably N-methylmorpholine.

[0045] In some embodiments, the nucleophilic substitution reaction is carried out in a solvent, which is a conventional solvent used in the art for this type of reaction, such as an organic solvent, or one or more of DMF, DMAc, DMSO, and 1,4-dioxane, preferably DMSO or DMF.

[0046] In some preferred embodiments, the molar volume ratio of the compound Ii to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.08 to 0.4 mol / L, or 0.1 mol / L.

[0047] In some preferred embodiments, the molar volume ratio of compound a to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.08 to 0.18 mol / L, or, for example, 0.12 mol / L.

[0048] In some embodiments, the molar ratio of compound Ii to compound a is a conventional molar ratio used in the art for this type of reaction, such as 1:1 to 1:1.5, or, for example, 1:1.2.

[0049] In some embodiments, the molar ratio of the compound Ii to the base is a conventional molar ratio used in the art for this type of reaction, such as 1:1 to 1:4, or, for example, 1:2.

[0050] In some embodiments, the nucleophilic reaction is carried out at a temperature that is conventional for this type of reaction in the art, such as room temperature, or 15–25°C, or 20°C.

[0051] In some preferred embodiments, the reaction solution A is treated in accordance with conventional treatment methods for this type of reaction solution in the art. The reaction solution A needs to be further filtered, washed and dried. For example, the filtration is vacuum filtration; the washing is rinsing with water; the drying is vacuum drying, preferably drying at 35-45°C for 18-24 hours.

[0052] In some embodiments, the nucleophilic substitution reaction is carried out in a conventional order in the art, for example, by dissolving compound Ii in the solvent, then adding a base, stirring, and then adding the reaction solution A.

[0053] In some embodiments, the reaction time of the nucleophilic substitution reaction is the conventional reaction time used in the art for this type of reaction, such as 8-24 hours, or 12-18 hours, or 16 hours.

[0054] In some embodiments, the nucleophilic substitution reaction further includes a post-processing step, which is a conventional post-processing step in the art, such as extraction, washing, separation and purification.

[0055] The organic phase used for extraction is dichloromethane, and the aqueous phase is water. The liquid used for washing is saturated saline solution. The chromatographic column used for separation and purification is a 1010-C18-BS column, with mobile phase A being 0.1% aqueous phosphoric acid solution and mobile phase B being methanol, the ratio of mobile phase A to mobile phase B being 50%:50%, and the flow rate for separation and purification being 1120 mL / min.

[0056] In some embodiments, the method for preparing the compound of formula I further includes reacting the compound of formula I-ii with compound b via a substitution reaction to prepare the compound of formula Ii, or

[0057] In some embodiments, in the method for preparing the compound represented by Formula I, the compound of Formula Ii is prepared by a method comprising a substitution reaction of the compounds represented by Formulas I-ii with compound b, or

[0058] In some embodiments, the preparation method of the compound of formula Ii in the preparation method of the compound of formula I includes a substitution reaction between the compound of formula I-ii and compound b.

[0059] Where: Tx is -OH or

[0060] X is a halogen;

[0061] The definitions of L2, L3, and Tr are as described above.

[0062] In some preferred embodiments, X is F, Cl, Br or I.

[0063] In some preferred embodiments, the reactants of the substitution reaction further include a base, which is a conventional base used in the art for this type of reaction, such as one or more of sodium bicarbonate, DIPEA, triethylamine, N-methylmorpholine, imidazole, pyridine, DBU, 2,6-dimethylpyridine, and diethylamine, for example, N-methylmorpholine.

[0064] In some preferred embodiments, the substitution reaction is carried out in an organic solvent, which is a conventional organic solvent used in the art for this type of reaction, such as one or more of THF, DMF, EA, DCM, 1,4-dioxane, and DMAc, or THF, DMF, DMAc, and DCM, or DCM.

[0065] In some preferred embodiments, the molar volume ratio of the compound of formula I-ii to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.1 to 0.16 mol / L, or for example, 0.133 mol / L.

[0066] In some preferred embodiments, the molar volume ratio of compound b to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.2 to 0.8 mol / L, or 0.4 mol / L.

[0067] In some preferred embodiments, the molar ratio of the compound of formula I-ii to the compound b is a conventional molar ratio used in the art for this type of reaction, for example, 1:2 to 1:5, or even 1:3.

[0068] In some preferred embodiments, the molar ratio of the compound of formula I-ii to the base is a conventional molar ratio used in the art for this type of reaction, for example, 1:2 to 1:5, or, for example, 1:3.

[0069] In some preferred embodiments, the reaction temperature of the substitution reaction is a conventional temperature used in the art for this type of reaction, such as room temperature, or 20–30°C, or 25°C.

[0070] In some preferred embodiments, the substitution reaction further includes a pretreatment of the reactants prior to the reaction. The pretreatment is a conventional pretreatment used in the art for this type of reaction, such as dissolving the compound of formula I-ii and the base in a solvent, or, for example, the pretreatment temperature is -5 to 5°C, preferably 0°C.

[0071] In some preferred embodiments, the reaction time of the substitution reaction is the conventional reaction time for this type of reaction in the art, such as 4-12 hours, or for example, 8 hours.

[0072] In some preferred embodiments, the substitution reaction further includes a post-processing step, which is a conventional post-processing step in the art, such as concentration, crystallization, or extraction.

[0073] The concentration is carried out in a water bath at 30-40°C, for example, at 35°C. The crystallization is performed by adding dichloromethane to the solution and allowing it to stand in layers. The extraction includes extraction with n-hexane and ethyl acetate, followed by vacuum concentration, which is carried out in a water bath at 30-40°C, for example, at 35°C.

[0074] In some embodiments, the method for preparing the compound of formula I further includes a condensation reaction of the compounds of formulas I-iii with compound c to prepare the compounds of formulas I-ii, or

[0075] In some embodiments, in the method for preparing the compound shown in Formula I, the compound of Formula I-ii is prepared by a method comprising a condensation reaction of the compound shown in Formulas I-iii with compound c, or

[0076] In some embodiments, the preparation method of the compounds of formula I-ii in the preparation method of the compounds shown in formula I includes a condensation reaction of the compounds shown in formulas I-iii with compound c.

[0077] Wherein: compound c is NH-L3-Tx, and L2, L3, and Tx are as described above.

[0078] In some preferred embodiments, the reactants of the condensation reaction further include a condensing agent, which is a conventional condensing agent used in this type of reaction in the art, such as one or more of DMTMM, CDMT, NMM, HATU, and EDCl, or HATU, DIPEA, CDMT, NMM, or DMTMM, or DMTMM, or DMTMM.

[0079] In some preferred embodiments, the condensation reaction is carried out in an organic solvent, which is a conventional organic solvent used in the art for this type of reaction, such as one or more of THF, DMAc, and DCM, for example, THF and DMAc.

[0080] In some preferred embodiments, the molar volume ratio of the compounds of formulas I-iii to the organic solvent is a commonly used molar volume ratio in the art for this type of reaction, for example, 0.3-0.7 mol / L, or 0.54 mol / L.

[0081] In some preferred embodiments, the molar volume ratio of compound c to the organic solvent is a commonly used molar volume ratio in the art for this type of reaction, for example 0.3-0.7 mol / L, or 0.52 mol / L.

[0082] In some preferred embodiments, the molar ratio of the compounds of formulas I-iii to compound c is a conventional molar ratio used in the art for this type of reaction, such as 1:0.6 to 1:1.5, or for example 1:0.67, 1:0.83, 1:1.1 or 1:1.2.

[0083] In some preferred embodiments, the molar ratio of the compound of formula I-iii to the condensing agent is a conventional molar ratio for this type of reaction in the art, for example, 1:1.2 to 1:1.8, or even 1:1.5.

[0084] In some preferred embodiments, the reaction temperature of the condensation reaction is a conventional temperature used in the art for this type of reaction, such as 5-30°C, or 20-30°C, or 25°C.

[0085] In some preferred embodiments, the reaction time of the condensation reaction is the conventional reaction time used in the art for this type of reaction, such as 12-24 hours, or for example, 18 hours.

[0086] In some preferred embodiments, the condensation reaction further includes a post-processing step, which is a conventional post-processing step in the art, such as filtration, extraction, or drying.

[0087] The filtration is vacuum filtration to remove filter residue. Preferably, the post-filtration step further includes concentration, which is carried out in a water bath at 30-40°C, for example, at 35°C. The extraction includes extraction with n-hexane and ethyl acetate. The extraction is further followed by drying, using anhydrous sodium sulfate as the drying agent. The drying is further followed by vacuum concentration, which is carried out in a water bath at 30-40°C, for example, at 35°C.

[0088] In some preferred embodiments, in the preparation method of the compound shown in Formula I, the preparation method of the compound shown in Formula IIi is as follows: the compound shown in Formulas I-iii undergoes a condensation reaction with compound c, and the resulting reaction solution containing the compound shown in Formulas I-ii directly undergoes the substitution reaction with compound b.

[0089] In some embodiments, the method for preparing the compound represented by Formula I further includes hydrolyzing the compounds represented by Formulas I-iv with a strong acid to prepare the compounds represented by Formulas I-iii, or

[0090] In some embodiments, in the method for preparing the compound represented by Formula I, the compounds of Formulas I-iii are prepared by a method comprising hydrolyzing the compounds of Formulas I-iv with a strong acid, or

[0091] In some embodiments, the preparation method of the compounds of formulas I-iii in the method for preparing the compounds shown in formula I includes a hydrolysis reaction of the compounds shown in formulas I-iv with a strong acid.

[0092] L2 is as described above.

[0093] In some preferred embodiments, I-iv is:

[0094] In some preferred embodiments, I-iii are:

[0095] In some preferred embodiments, the strong acid is a conventional strong acid used in this type of reaction, such as trifluoroacetic acid.

[0096] In some preferred embodiments, the hydrolysis reaction is carried out in an organic solvent, which is a conventional organic solvent used in the art for this type of reaction, such as one or more of THF, DMAc, and DCM, for example, DCM.

[0097] In some preferred embodiments, the molar volume ratio of compounds I-iv to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.1-0.4 mol / L, or 0.2 mol / L.

[0098] In some preferred embodiments, the molar volume ratio of the compounds I-iv to the strong acid is a conventional molar volume ratio for this type of reaction in the art, for example, 0.2 to 0.7 mol / L, or, for example, 0.4 mol / L.

[0099] In some preferred embodiments, the hydrolysis reaction is carried out at a temperature that is conventional for this type of reaction in the art, such as 15–25°C, or 20°C.

[0100] In some preferred embodiments, the reaction time of the hydrolysis reaction is the conventional reaction time used in the art for this type of reaction, such as 1-5 hours, or for example, 2 hours.

[0101] In some preferred embodiments, the hydrolysis reaction further includes a post-processing step, which is a conventional post-processing step in the art, such as concentration, extraction, or drying.

[0102] The concentration is carried out in a water bath at 30-40°C, for example, in a water bath at 35°C. The extraction includes extraction with dichloromethane and methyl tert-butyl ether, and the extraction is further followed by drying, the drying agent being anhydrous sodium sulfate, and the drying is further followed by vacuum concentration, which is carried out in a water bath at 30-40°C, for example, in a water bath at 35°C.

[0103] In some embodiments, the method for preparing the compound of formula I further includes an amide condensation reaction of the compound of formula Iv with compound 1 to prepare the compound of formula I-iv, or

[0104] In some embodiments, in the method for preparing the compound of formula I, the compound of formula I-iv is prepared by a method comprising an amide condensation reaction of the compound of formula Iv with compound 1, or

[0105] In some embodiments, the preparation of compounds of formula I-iv in the method for preparing the compound shown in formula I includes an amide condensation reaction of the compound shown in formula Iv with compound 1.

[0106] L2 is as described above.

[0107] In some preferred embodiments, I-iv is:

[0108] In some preferred embodiments, the reactants of the amide condensation reaction further include a condensing agent, which is a conventional condensing agent used in this type of reaction in the art, such as one or more of DMTMM, CDMT, HATU, and EDCl, for example, DMTMM.

[0109] In some preferred embodiments, the amide condensation reaction is carried out in an organic solvent, which is a conventional organic solvent used in the art for this type of reaction, such as one or more of THF, DMAc, DCM, and DMF, for example, DMF, DMAc, or THF. In some embodiments, the organic solvent is DMF or DMAc.

[0110] In some preferred embodiments, the molar volume ratio of compound IV to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.2-0.6 mol / L, or 0.46 mol / L.

[0111] In some preferred embodiments, the molar volume ratio of compound 1 to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.4-0.7 mol / L, or 0.56 mol / L.

[0112] In some preferred embodiments, the molar ratio of compound IV to compound 1 is a conventional molar ratio used in the art for this type of reaction, such as 1:0.8 to 1:1.5, or even 1:1.1 or 1:1.2.

[0113] In some preferred embodiments, the molar ratio of the compound IV to the condensing agent is a conventional molar ratio used in the art for this type of reaction, such as 1:1.2 to 1:1.8, or even 1:1.5.

[0114] In some preferred embodiments, the reaction temperature of the amide condensation reaction is a conventional temperature used in the art for this type of reaction, such as 10–30°C, or 20–30°C, or 25°C.

[0115] In some preferred embodiments, the reaction time of the amide condensation reaction is the conventional reaction time used in the art for this type of reaction, such as 18-36 hours, or for example, 24 hours.

[0116] In some preferred embodiments, the amide condensation reaction further includes a post-processing step, which is a conventional post-processing step in the art, such as extraction, drying, or filtration.

[0117] The extraction requires prior concentration, which is carried out in a water bath at 30-40°C, for example, at 35°C. The extraction includes extraction with methyl tert-butyl ether, followed by drying using anhydrous sodium sulfate as a drying agent. The drying process further includes vacuum concentration, which is carried out in a water bath at 30-40°C, for example, at 35°C.

[0118] In another aspect, this disclosure provides a method for preparing a compound as shown in formula Ii, comprising reacting a compound of formulas I-ii with compound b in a substitution reaction.

[0119] Where Tx is -OH or X is a halogen; L2, L3, and Tr are defined as described above, and the preferred reaction conditions for the substitution reaction are as described above.

[0120] In some embodiments, the preparation method of the compound represented by formula Ii further includes a condensation reaction of the compounds represented by formulas I-iii with compound c to prepare the compounds represented by formulas I-ii, or

[0121] In some embodiments, in the method for preparing the compound represented by formula Ii, the compound represented by formulas I-ii is prepared by a method comprising a condensation reaction of the compound represented by formulas I-iii with compound c, or

[0122] In some embodiments, the preparation method of the compounds of formulas I-ii in the preparation method of the compounds shown in formula Ii includes a condensation reaction of the compounds shown in formulas I-iii with compound c.

[0123] Wherein: compound c is H-L3-Tx, and L2, L3, and Tx are as described above, and the reaction conditions for the condensation reaction are preferably as described above.

[0124] In some embodiments, the preparation method of the compound represented by formula Ii further includes hydrolyzing the compounds represented by formulas I-iv with a strong acid to prepare the compounds represented by formulas I-iii, or

[0125] In some embodiments, in the method for preparing the compound represented by formula Ii, the compounds of formulas I-iii are prepared by a method comprising hydrolyzing the compounds represented by formulas I-iv with a strong acid, or

[0126] In some embodiments, the preparation of compounds I-iii in the method for preparing the compound represented by formula Ii includes a hydrolysis reaction of the compound represented by formulas I-iv with a strong acid.

[0127] Wherein, L2 is as described above, and the reaction conditions for the hydrolysis reaction are preferably as described above.

[0128] In some embodiments, the method for preparing the compound of formula Ii further includes reacting the compound of formula Iv with compound 1 via an amide condensation reaction to prepare the compounds of formulas I-iv, or

[0129] In some embodiments, in the method for preparing the compound represented by formula Ii, the compounds of I-iv are prepared by a method comprising an amide condensation reaction of the compound represented by formula Iv with compound 1, or

[0130] In some embodiments, the preparation method of the compounds of formula I-iv in the preparation method of the compound shown in formula Ii includes an amide condensation reaction between the compound shown in formula Iv and compound 1.

[0131] Wherein, L2 is as described above, and the reaction conditions for the amide condensation reaction are preferably as described above.

[0132] In another aspect, this disclosure provides a method for preparing compounds of formulas I-ii, comprising a condensation reaction of compounds of formulas I-iii with compound c.

[0133] Wherein compound c is H-L3-Tx, and L2, L3, and Tx are as described above, and the reaction conditions for the condensation reaction are preferably as described above.

[0134] In some embodiments, the preparation method of the compounds represented by formulas I-ii further includes hydrolyzing the compounds represented by formulas I-iv with a strong acid to prepare the compounds represented by formulas I-iii, or

[0135] In some embodiments, in the preparation methods of the compounds represented by formulas I-ii, the compounds of formulas I-iii are prepared by a method comprising hydrolyzing the compounds represented by formulas I-iv with a strong acid, or

[0136] In some embodiments, the preparation of compounds I-iii in the methods for preparing compounds of formulas I-ii includes a hydrolysis reaction of compounds of formulas I-iv with a strong acid.

[0137] Wherein, L2 is as described above, and the reaction conditions for the hydrolysis reaction are preferably as described above.

[0138] In some embodiments, the preparation method of the compounds represented by formulas I-ii further includes an amide condensation reaction of the compound represented by formula Iv with compound 1 to prepare the compounds represented by formulas I-iv, or

[0139] In some embodiments, in the preparation methods of the compounds represented by formulas I-ii, the compounds I-iv are prepared by a method comprising an amide condensation reaction of the compound represented by formula Iv with compound 1, or

[0140] In some embodiments, the preparation of compounds I-iv in the methods for preparing compounds of formulas I-ii includes an amide condensation reaction of the compound of formula Iv with compound 1.

[0141] Wherein, L2 is as described above, and the reaction conditions for the amide condensation reaction are preferably as described above.

[0142] In another aspect, this disclosure provides a method for preparing compounds of formulas I-iii, comprising hydrolyzing the compounds of formulas I-iv with a strong acid.

[0143] Wherein, L2 is as described above, and the reaction conditions for the hydrolysis reaction are preferably as described above.

[0144] In some embodiments, the preparation method of the compounds represented by formulas I-iii further includes an amide condensation reaction of the compound represented by formula Iv with compound 1 to prepare the compounds represented by formulas I-iv, or

[0145] In some embodiments, in the preparation methods of the compounds represented by formulas I-iii, the compounds of formulas I-iv are prepared by a method comprising an amide condensation reaction of the compound represented by formula Iv with compound 1, or

[0146] In some embodiments, the preparation of compounds I-iv in the methods for preparing compounds of formulas I-iii includes an amide condensation reaction of the compound of formula Iv with compound 1.

[0147] Wherein, L2 is as described above, and the reaction conditions for the amide condensation reaction are preferably as described above.

[0148] In another aspect, this disclosure provides a method for preparing compounds as shown in formulas I-iv, comprising an amide condensation reaction of a compound shown in formula Iv with compound 1.

[0149] Wherein, L2 is as described above, and the reaction conditions for the amide condensation reaction are preferably as described above.

[0150] In another aspect, this disclosure provides a second method for preparing compounds as shown in Formula I, comprising dehydration condensation of compounds shown in Formulas I-iii with compound a-1.

[0151] The definitions of groups L2, L3, Tr, Rx, p, R1, and R2 are as described above; the preparation methods of the compounds of formulas I-iii are as described above.

[0152] In some preferred embodiments, compound a-1 is:

[0153] In some preferred embodiments, the dehydration condensation of the compounds of formulas I-iii with compound a-1 further comprises a condensing agent, which is HBTU, TBTU, EEDQ, EDCl, T4P, DMTMM, TSTU or HATU, preferably HBTU, TBTU, or HATU, and more preferably HATU.

[0154] In some preferred embodiments, the dehydration condensation of the compounds of formulas I-iii with compound a-1 further comprises a base, which is a conventional base used in the art for this type of reaction, preferably DIPEA or TEA, more preferably DIPEA.

[0155] In some preferred embodiments, the compounds of formulas I-iii undergo dehydration condensation with compound a-1 in an organic solvent, wherein the organic solvent is DMAc, DMSO, MeCN, DMF, NMP or HMPA, preferably DMAc, DMSO, MeCN, DMF or NMP, and more preferably DMSO.

[0156] In some preferred embodiments, the molar volume ratio of compound a-1 to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.07-0.12 mol / L, or 0.1 mol / L.

[0157] In some preferred embodiments, the molar volume ratio of the compound of formulas I-iii to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.07-0.12 mol / L, or 0.1 mol / L.

[0158] In some preferred embodiments, the molar ratio of compound a-1 to the compounds of formula I-iii is a conventional molar ratio used in the art for this type of reaction, for example, 1:0.8 to 1:1.5, or even 1:1.

[0159] In some preferred embodiments, the molar ratio of compound a-1 to the condensing agent is a conventional molar ratio used in the art for this type of reaction, for example, 1:1.1 to 1:1.8, or, for example, 1:1.2.

[0160] In some preferred embodiments, the molar ratio of the base to the compound a-1 is a conventional molar ratio used in the art for this type of reaction, for example, 1:1 to 3:1, or, for example, 2:1.

[0161] In some preferred embodiments, the reaction temperature for the dehydration condensation of the compounds of formulas I-iii with compound a-1 is a conventional temperature used in the art for this type of reaction, for example -30 to -10°C, or -25 to -15°C, or -20°C.

[0162] In some preferred embodiments, the reaction time for the dehydration condensation of the compounds of formulas I-iii with compound a-1 is the conventional reaction time used in the art for this type of reaction, for example 0.2-1 hours, or for example 0.5 hours.

[0163] In some preferred embodiments, the dehydration condensation of the compounds of formulas I-iii with compound a-1 further includes a post-processing step, which is a conventional post-processing step in the art, such as extraction or HPLC separation.

[0164] The post-processing step involves adding methyl tert-butyl ether to the reaction solution, extracting, and then performing HPLC preparation. The preferred HPLC conditions are mobile phase A (0.1% aqueous phosphoric acid): mobile phase B (methanol) = 50%: 50%, flow rate 1120 mL / min, and elution time 5 min.

[0165] In some preferred embodiments, the dehydration condensation reaction step of the compound of formula I-iii with compound a-1 is as follows: placing the compound of formula I-iii in a container, adding the solvent, the base and the condensing agent at the reaction temperature, stirring, and then adding compound a-1 to react.

[0166] In some embodiments, the second preparation method of the compound of formula I further includes performing a deprotection reaction of compound ai to prepare compound a-1, or

[0167] In some embodiments, in the second preparation method of the compound of formula I, compound a-1 is prepared by a deprotection reaction including compound ai, or

[0168] In some embodiments, the preparation method of compound a-1 in the second preparation method for the compound of formula I includes carrying out a deprotection reaction of compound ai.

[0169] The protecting group is a -Boc group.

[0170] The definitions of groups L3, Tr, Rx, p, R1, and R2 are as described above.

[0171] In some preferred embodiments, the compound ai is preferably:

[0172] In some preferred embodiments, the deprotection reaction of the compound ai further comprises an organic acid, such as hydrochloric acid, zinc bromide, TMSOTf, or trifluoroacetic acid, for example hydrochloric acid or trifluoroacetic acid, or trifluoroacetic acid for example.

[0173] In some preferred embodiments, the deprotection reaction of compound ai is carried out in an organic solvent, which is a conventional organic solvent used in the art for this type of reaction, preferably methanol, dichloromethane, 1,4-dioxane, ethyl acetate, more preferably methanol, dichloromethane, ethyl acetate, and even more preferably dichloromethane.

[0174] In some preferred embodiments, the molar volume ratio of the compound ai to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.1-0.4 mol / L, or 0.23 mol / L.

[0175] In some preferred embodiments, the volume ratio of the organic acid to the organic solvent is a conventional volume ratio used in the art for this type of reaction, such as 1:2 to 1:5, or, for example, 1:2.

[0176] In some preferred embodiments, the volume molar ratio of the organic acid to the compound ai is 3:1 to 0.8:1, for example 2.5:1 to 1:1, or 2:1 to 2.5:1, or even 2.5:1, 2.1:1 or 2:1.

[0177] In some preferred embodiments, the deprotection reaction of the compound ai is carried out at a temperature that is conventional for this type of reaction in the art, such as -20 to 20°C, or -20 to -10°C, or 0°C.

[0178] In some preferred embodiments, the deprotection reaction of the compound ai takes place over a time that is conventional for this type of reaction in the art, for example, 0.5-4 hours, or for example, 1 hour.

[0179] In some preferred embodiments, the deprotection reaction of the compound ai further includes a post-processing step, which is a conventional post-processing step in the art, such as distillation, extraction, or HPLC separation.

[0180] In some preferred embodiments, the post-treatment step of the compound is to remove the organic solvent from the reaction solution of the deprotection reaction of compound ai under reduced pressure, extract with methyl tert-butyl ether, and finally separate by HPLC, wherein the HPLC separation conditions are: mobile phase A (0.1% aqueous phosphoric acid solution) and mobile phase B (methanol).

[0181] In some embodiments, the second preparation method of the compound of formula I further includes an addition reaction of compound a with compound f to prepare compound ai, or

[0182] In some embodiments, in the second preparation method of the compound of formula I, compound ai is prepared by a method comprising an addition reaction of compound a and compound f, or

[0183] In some embodiments, the preparation method of compound ai in the method for preparing the compound of formula I includes an addition reaction between compound a and compound f.

[0184] The definitions of groups L3, Tr, Rx, p, R1, and R2 are as described above.

[0185] In some preferred embodiments, compound a is:

[0186] In some preferred embodiments, the addition reaction further comprises an activator, which is DMAP, HOAt, HOSu or HOBt, preferably DMAP, HOSu or HOBt, more preferably HOSu.

[0187] In some preferred embodiments, the addition reaction further comprises a base, which is DIPEA or NMM, preferably DIPEA.

[0188] In some preferred embodiments, the addition reaction is carried out in an organic solvent, which is a conventional organic solvent used in the art for this type of reaction, such as DMAc, DMSO, MeCN, DMF, NMP or DIPEA, more preferably DMF.

[0189] In some preferred embodiments, the molar volume ratio of compound a to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.07-0.2 mol / L, or 0.18 mol / L.

[0190] In some preferred embodiments, the molar volume ratio of compound f to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example 0.07-0.2 mol / L, or 0.18 mol / L.

[0191] In some preferred embodiments, the molar ratio of compound f to compound a is 1:0.8 to 1:1.5, for example 1:0.9 to 1:1, or even 1:1.

[0192] In some preferred embodiments, the molar ratio of the activator to the compound a is a conventional molar ratio used in the art for this type of reaction, for example, 0.2:1 to 1.5:1, or 0.5:1 to 1.5:1, or 0.5:1, 1:1, or 1.5:1.

[0193] In some preferred embodiments, the molar ratio of the base to the compound a-1 is a conventional molar ratio used in the art for this type of reaction, for example, 1:1 to 3:1, or, for example, 2:1.

[0194] In some preferred embodiments, the reaction temperature of the addition reaction is a conventional temperature used in the art for this type of reaction, such as 10-15°C.

[0195] In some preferred embodiments, the reaction time of the addition reaction is the conventional reaction time used in the art for this type of reaction, such as 2-5 hours, or for example, 3 hours.

[0196] In some preferred embodiments, the addition reaction further includes a post-processing step, which is a conventional post-processing step in the art, such as extraction or filtration.

[0197] In some preferred embodiments, the post-treatment step of the addition reaction involves the addition of ethanol, followed by stirring and the addition of water, filtration, and subsequent rinsing with a mixture of ethanol and water.

[0198] In another aspect, this disclosure provides a method for preparing compound a-1, comprising a deprotection reaction of compound ai, wherein the protecting group is a -Boc group.

[0199] The definitions of groups L3, Tr, Rx, p, R1, and R2 are as described above, and the preparation conditions of compound a-1 are as described above.

[0200] In some preferred embodiments, the preparation method of compound a-1 further includes an addition reaction between compound a and compound f to prepare compound ai, or

[0201] In some preferred embodiments, in the method for preparing compound a-1, compound ai is prepared by a method involving an addition reaction between compound a and compound f, or

[0202] In some preferred embodiments, the preparation of compound ai in the method for preparing compound a-1 includes an addition reaction between compound a and compound f.

[0203] The definitions of groups L3, Tr, Rx, p, R1, and R2 are as described above, and the preparation method of compound ai is as described above.

[0204] In another aspect, this disclosure provides a method for preparing compound ai, comprising reacting compound a with compound f in an addition reaction to obtain said compound ai.

[0205] The definitions of groups L3, Tr, Rx, p, R1, and R2 are as described above, and the preparation method of compound ai is as described above.

[0206] In another aspect, this disclosure provides a third method for preparing compounds as shown in Formula I, comprising carrying out substitution reactions of compounds shown in Formulas I-VI.

[0207] The definitions of groups L2, L3, Tr, Rx, p, R1, and R2 are as described above.

[0208] In some preferred embodiments, the substitution reaction further comprises an oxidizing agent, which is a conventional oxidizing agent used in this type of reaction in the art, preferably m-chloroperoxybenzoic acid.

[0209] In some preferred embodiments, the substitution reaction is carried out in an organic solvent, which is a conventional organic solvent used in the art for this type of reaction, such as DCM, DMAc, DMSO, MeCN, DMF, NMP, or DIPEA, more preferably DCM.

[0210] In some preferred embodiments, the molar volume ratio of the compounds of formulas I-VI to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.01-0.2 mol / L, or 0.02 mol / L.

[0211] In some preferred embodiments, the molar volume ratio of the oxidant to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.01-0.2 mol / L, or 0.07-0.2 mol / L, or 0.024 mol / L or 0.18 mol / L.

[0212] In some preferred embodiments, the molar ratio of the compounds of formulas I-VI to the oxidant is a conventional molar ratio for this type of reaction in the art, for example, 1:0.8-1:2, or, for example, 1:1.2.

[0213] In some preferred embodiments, the reaction time of the addition reaction is the conventional reaction time used in the art for this type of reaction, such as 6-18 hours, or for example, 12 hours.

[0214] In some preferred embodiments, the substitution reaction includes a post-treatment process, which includes extraction and HPLC separation. Preferably, the post-treatment process includes extraction of the reaction solution of the substitution reaction with methyl tert-butyl ether, followed by HPLC separation. The HPLC column is 1010-C18-BS, and the mobile phase A (0.1% aqueous phosphoric acid): mobile phase B (methanol) = 50%:50% is used at a flow rate of 1120 mL / min.

[0215] In some preferred embodiments, the oxidation reaction is carried out at a temperature that is conventional for this type of reaction in the art, such as -30 to -10°C, or -20°C.

[0216] In some embodiments, the third method for preparing the compound of formula I further includes a condensation reaction of the compounds of formulas I-VII with compound e to prepare the compounds of formulas I-VI, or

[0217] In some embodiments, in the third preparation method of the compounds of formula I, the compounds of formulas I-VI are prepared by a method comprising a condensation reaction of the compounds shown in formulas I-VII with compound e, or

[0218] In some embodiments, the preparation of compounds I-VI in the third preparation method for compounds of formula I includes a condensation reaction of compounds of formulas I-VII with compound e.

[0219] In some preferred embodiments, the condensation reaction further comprises a condensing agent, which is a conventional condensing agent used in this type of reaction in the art, preferably NMM.

[0220] In some preferred embodiments, the condensation reaction further comprises a coupling agent, which is a conventional coupling agent used in this type of reaction in the art, preferably CDMT.

[0221] In some preferred embodiments, the condensation reaction is carried out in an organic solvent, which is a conventional organic solvent used in the art for this type of reaction, such as DMAc, DMSO, MeCN, DMF, NMP or DIPEA, more preferably DMF.

[0222] In some preferred embodiments, the molar volume ratio of compound e to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.01-0.2 mol / L, or 0.05 mol / L.

[0223] In some preferred embodiments, the molar volume ratio of the compounds of formulas I-VII to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.01-0.2 mol / L, or, for example, 0.05 mol / L.

[0224] In some preferred embodiments, the molar ratio of compound e to the compounds of formulas I-VII is a conventional molar ratio used in the art for this type of reaction, for example, 1:0.8 to 1:1.5, or even 1:1.

[0225] In some preferred embodiments, the molar ratio of the coupling agent to the compounds of formulas I-VII is a conventional molar ratio used in the art for this type of reaction, for example, 1:1 to 2:1, or 2:1.

[0226] In some preferred embodiments, the molar ratio of the condensing agent to the compounds of formulas I-VII is a conventional molar ratio used in the art for this type of reaction, for example, 1:1 to 2:1, or 2:1.

[0227] In some preferred embodiments, the reaction temperature of the addition reaction is a conventional temperature used in the art for this type of reaction, such as 15-30°C, or for example, 25°C or 20°C.

[0228] In some preferred embodiments, the reaction time of the addition reaction is the conventional reaction time used in the art for this type of reaction, such as 6-14 hours, or for example, 8 hours.

[0229] In some embodiments, the third preparation method of the compound of formula I further includes a deprotection reaction of the compound of formula IX with piperidine to prepare the compounds of formulas I-VII, or

[0230] In some embodiments, in the third preparation method of the compounds of formula I, the compounds of formulas I-VII are prepared by a method comprising a deprotection reaction of the compound of formula IX with piperidine.

[0231] In some embodiments, the preparation methods of compounds I-VII in the third preparation method for the compound of formula I include a deprotection reaction of the compound of formula IX with piperidine.

[0232] In some preferred embodiments, the deprotection reaction further comprises a deprotecting agent, which is a conventional deprotecting agent used in this type of reaction in the art, preferably piperidine.

[0233] In some preferred embodiments, the deprotection reaction is carried out in an organic solvent, which is a conventional organic solvent used in the art for this type of reaction, such as DMAc, DMSO, MeCN, DMF, NMP or DIPEA, more preferably DMF.

[0234] In some preferred embodiments, the molar volume ratio of the compound of formula IX to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.01-0.2 mol / L, or 0.1 mol / L.

[0235] In some preferred embodiments, the molar volume ratio of the deprotecting agent to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.5-2 mol / L, or, for example, 1 mol / L.

[0236] In some preferred embodiments, the molar ratio of the deprotecting agent to the compound of formula IX is a conventional molar ratio used in the art for this type of reaction, for example, 1:8 to 1:15, or even 1:10.

[0237] In some preferred embodiments, the deprotection reaction is carried out at a temperature that is conventional for this type of reaction in the art, such as 15-30°C, or 20°C.

[0238] In some preferred embodiments, the deprotection reaction time is the conventional reaction time used in the art for this type of reaction, for example, 0.5-4 hours, or for example, 2 hours.

[0239] In some embodiments, the third method for preparing the compound of formula I further includes a condensation reaction of compound a-1 and compound a-2 to prepare the compound of formula IX, or

[0240] In some embodiments, in the third preparation method of the compound of formula I, the compound of formula IX is prepared by a method comprising a condensation reaction of compound a-1 and compound a-2, or

[0241] In some embodiments, the preparation method of the compound of formula I in the third preparation method includes a condensation reaction of compound a-1 and compound a-2.

[0242] In some preferred embodiments, the condensation reaction further comprises a base, which is a conventional base used in the art for this type of reaction, preferably DIPEA.

[0243] In some preferred embodiments, the condensation reaction further comprises a condensing agent, which is a conventional condensing agent used in this type of reaction in the art, preferably HATU.

[0244] In some preferred embodiments, the condensation reaction is carried out in an organic solvent, which is a conventional organic solvent used in the art for this type of reaction, such as DMAc, DMSO, MeCN, DMF, NMP or DIPEA, more preferably DMF.

[0245] In some preferred embodiments, the molar volume ratio of compound a-1 to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.01-0.2 mol / L, or 0.1 mol / L.

[0246] In some preferred embodiments, the molar volume ratio of the base to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.2-2 mol / L, or 0.5 mol / L.

[0247] In some preferred embodiments, the molar volume ratio of the condensing agent to the organic solvent is a conventional molar volume ratio used in the art for this type of reaction, for example, 0.05-0.5 mol / L, or, for example, 0.12 mol / L.

[0248] In some preferred embodiments, the molar ratio of the base to the compound a-1 is a conventional molar ratio used in the art for this type of reaction, such as 1:1 to 3:1, or even 2:1.

[0249] In some preferred embodiments, the molar ratio of compound a-2 and compound a-1 is a conventional molar ratio used in the art for this type of reaction, for example, 0.8:1 to 1.5:1, or even 1:1.

[0250] In some preferred embodiments, the molar ratio of the condensing agent to the compound a-1 is a conventional molar ratio used in the art for this type of reaction, for example, 0.8:1 to 1.5:1, or, for example, 1.2:1.

[0251] In some preferred embodiments, the condensation reaction is carried out at a temperature that is conventional for this type of reaction in the art, such as -5 to 100°C, or 0°C.

[0252] In some preferred embodiments, the reaction time of the condensation reaction is the conventional reaction time used in the art for this type of reaction, such as 2-6 hours, or for example, 4 hours.

[0253] In some embodiments, the third method for preparing the compound of formula I further includes reacting compounds I-VIII with piperidine to prepare compound a-1, or

[0254] In some embodiments, in the third preparation method of the compound of formula I, compound a-1 is prepared by a method comprising a substitution reaction of compounds I-VIII with piperidine, or

[0255] In some embodiments, the preparation of compound a-1 in the third preparation method for compounds of formula I includes a substitution reaction of compounds I-VIII with piperidine.

[0256] Preferably, the preparation method of compound a-1 is as described above.

[0257] In some embodiments, the third method for preparing the compounds of formula I further includes a substitution reaction of compound a with compound a-3 to prepare the compounds of formulas I-VII, or

[0258] In some embodiments, in the third preparation method of the compounds of formula I, the compounds of formulas I-VII are prepared by a method involving a substitution reaction between compound a and compound a-3, or

[0259] In some embodiments, the preparation method of compounds of formulas I-VII in the third preparation method for compounds of formula I includes a substitution reaction between compound a and compound a-3.

[0260] Preferably, the preparation method of the compounds of formulas I-VII is as described above.

[0261] In another aspect, the present invention provides an intermediate for preparing the compound of formula I:

[0262] In another aspect, the present invention provides a compound of formula I obtained by the above preparation method, wherein the compound of formula I and the various features of the preparation method are as described above.

[0263] In any embodiment according to the foregoing aspects, the compound of formula I is:

[0264] Terminology Definition

[0265] In this application, unless otherwise stated, the scientific and technical terms used herein have the meanings commonly understood by those skilled in the art. Furthermore, the laboratory procedures for cell culture, molecular genetics, nucleic acid chemistry, and immunology used herein are all standard procedures widely used in their respective fields. To better understand this disclosure, definitions and explanations of relevant terms are provided below.

[0266] When the lower and upper limits of a numerical range are disclosed, any numerical value falling within that range and any included range are specifically disclosed. In particular, each range of values ​​disclosed herein (in the form of “about a to b”, or equivalently, “approximately a to b”, or equivalently, “about ab”) should be understood to represent each numerical value and range encompassed within a wider range;

[0267] For example, the expression "C1-6" should be understood as encompassing any subrange and each point value within it, such as C2-5, C3-4, C1-2, C1-3, C1-4, C1-5, etc., as well as C, C2, C3, C4, C5, C6, etc. Similarly, the expression "C3-10" should be understood in a similar way, for example, it can encompass any subrange and point value contained within it, such as C3-9, C6-9, C6-8, C6-7, C7-10, C7-9, C7-8, C8-9, etc., as well as C3, C4, C5, C6, C7, C8, C9, C10, etc. For example, the expression "3-10 yuan" should be understood as encompassing any sub-range and each point value within it, such as 3-4 yuan, 3-5 yuan, 3-6 yuan, 3-7 yuan, 3-8 yuan, 3-9 yuan, 4-5 yuan, 4-6 yuan, 4-7 yuan, 4-8 yuan, 5-7 yuan, 5-8 yuan, 6-7 yuan, etc., as well as 3, 4, 5, 6, 7, 8, 9, 10 yuan, etc. Similarly, the expression "5-10 yuan" should be understood in a similar way, for example, it can encompass any sub-range and point value included within it, such as 5-6 yuan, 5-7 yuan, 5-8 yuan, 5-9 yuan, 5-10 yuan, 6-7 yuan, 6-8 yuan, 6-9 yuan, 6-10 yuan, 7-8 yuan, etc., as well as 5, 6, 7, 8, 9, 10 yuan, etc.

[0268] In this application, the term "alkyl" refers to a saturated straight-chain or branched hydrocarbon group. As used herein, the term "C1-6 alkyl" refers to a saturated straight-chain or branched hydrocarbon group having 1 to 6 carbon atoms (e.g., 1, 2, 3, 4, 5, or 6 carbon atoms). "C1-6 alkyl" includes, for example, methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, n-pentyl, isopentyl, neopentyl, or n-hexyl. Alkyl groups in this invention are optionally substituted with one or more substituents described in this invention. In this application, the term "alkylene" refers to a saturated straight-chain or branched divalent hydrocarbon group. As used herein, the term "C1-6 alkylene" refers to a saturated straight-chain or branched divalent hydrocarbon group having 1 to 6 carbon atoms. "C1-6 alkylene" includes, for example, but not limited to, methylene, ethylene, propylene, or butylene. Alkylene groups in this invention are optionally substituted with one or more substituents described in this invention.

[0269] In this application, the term "aryl" refers to an aromatic hydrocarbon group having a monocyclic or fused ring with a conjugated π-electron system. For example, the term "6-10 aryl" as used herein refers to an aryl group (such as phenyl, naphthyl, etc.) having 6-10 carbon atoms, which is optionally substituted with one or more substituents described herein (such as halogen substitution, etc.).

[0270] In this application, the term "arylene" refers to a monocyclic or fused-ring divalent aromatic hydrocarbon group having a conjugated π-electron system. For example, the term "C6-10 arylene" as used herein refers to an arylene having 6-10 carbon atoms, which is optionally substituted with one or more substituents described herein (such as halogen substitution, etc.).

[0271] In this application, the term "heteroaryl" or "heteroary ring" refers to a monocyclic and fused heterocyclic system having one or more conjugated π-electron systems, wherein one or more (e.g., 1, 2, or 3) ring atoms are heteroatoms selected from N, O, P, and S, and the remaining ring atoms are C. Heteroaryl or heteroary rings can be characterized by the number of ring atoms. For example, a 5-12 membered heteroaryl may contain 5-12 (e.g., 5, 6, 7, 8, 9, 10, 11, or 12) ring atoms, particularly 5, 6, 9, or 10 ring atoms. Examples of heteroaryl groups include thiophene, furanyl, pyrrole, oxazolyl, thiazolyl, imidazolyl, pyrazolyl, pyridinyl, pyrazinyl, isoxazolyl, isothiazolyl, oxadiazolyl, triazolyl, thiadiazolyl, indolyl, etc., optionally substituted by one or more substituents described herein.

[0272] In this application, the term "hybrid aryl" refers to a monocyclic or fused-ring divalent aromatic group having a conjugated π-electron system, the ring having one or more carbon atoms (e.g., 1, 2, 3, 4, 5, 6, 9, or 10 carbon atoms) and one or more (e.g., 1, 2, 3, or 4) heteroatoms, each independently selected from N, O, P, and S, for example having a total of 5-12 (preferably 5-10, more preferably 5, 6, 9, or 10) ring atoms. For example, the term "5-12-membered heteroaryl" as used herein refers to a heteroaryl having 5-12 ring atoms, optionally substituted with one or more substituents described herein.

[0273] In this application, the terms “cycloalkyl” or “carbocyclic” refer to a saturated or partially saturated, monocyclic or polycyclic (such as bicyclic) non-aromatic hydrocarbon group. For example, “C3-12 cycloalkyl” or “3-12 membered cycloalkyl” refers to a cycloalkyl group having 3-12 ring carbon atoms (such as 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12). Common cycloalkyl groups include (but are not limited to) monocyclic cycloalkyl groups, such as cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclobutene, cyclopentene, cyclohexene, etc.; or bicyclic cycloalkyl groups, including fused rings, bridged rings, or spirocyclic groups, such as bicyclo[1.1.1]pentyl, bicyclo[2.2.1]heptyl, bicyclo[3.2.1]octyl, bicyclo[5.2.0]nonyl, decahydronaphthyl, etc., which are optionally substituted by one or more substituents described herein.

[0274] In this application, the term "cycloalkylene" refers to a saturated or partially saturated, monocyclic or polycyclic (such as bicyclic) non-aromatic divalent cyclic group. For example, "C3-12 cycloalkylene" or "3-12 membered cycloalkylene" refers to a cycloalkylene group having 3-12 ring carbon atoms (such as 3, 4, 5, 6, 7, 8, 9, 10, 11 or 12). Common cycloalkylene compounds include (but are not limited to) monocyclic cycloalkylene compounds, such as cyclopropylene, cyclobutylene, cyclopentylene, cyclohexylene, cycloheptylene, cyclobutene, cyclopentene, cyclohexene, etc.; or bicyclic cycloalkylene compounds, including fused rings, bridged rings, or spiro rings, such as bicyclic[1.1.1]pentyl, bicyclic[2.2.1]heptyl, bicyclic[3.2.1]octyl, bicyclic[5.2.0]nonyl, decahydronaphthylene, etc., which are optionally substituted by one or more substituents described herein.

[0275] The term "heterocyclic alkyl" or "heterocycle" refers to a saturated or partially saturated non-aromatic cyclic group containing at least one heteroatom selected from N, O, P, and S as a ring member, preferably one, two, three, or four. Examples include 3-8-membered and 3-6-membered heterocyclic alkyl groups. Specific examples include, but are not limited to, ethylene oxide, oxocyclobutane, pyrrolidinyl, tetrahydrofuranyl, piperidinyl, piperazinyl, tetrahydropyranyl, homopiperazinyl, etc., optionally substituted with one or more oxo groups or substituents described herein.

[0276] The term "heterocyclic alkylene" refers to a saturated or partially saturated, non-aromatic divalent cyclic group containing at least one heteroatom selected from N, O, P, and S as a ring member, preferably 1, 2, 3, or 4 heteroatoms. Examples include 3-8-membered and 3-6-membered heterocyclic alkylenes. Specific examples include, but are not limited to, ethylene oxide, cyclobutane, pyrrolidinyl, tetrahydrofuranyl, piperidinyl, piperazineyl, tetrahydropyranyl, homopiperazineyl, etc., optionally substituted with one or more oxo groups or substituents described herein.

[0277] The term "fused ring (fused ring system)" refers to a polycyclic structure formed by two or more (e.g., 3, 4, or 5) carbon rings or heterocycles sharing a common ring edge, wherein the carbon rings include cycloalkyl and aryl groups, and the heterocycles include heteroaromatic and heterocyclic alkyl groups. The fused ring systems include, but are not limited to: fused ring systems formed by cycloalkyl groups with cycloalkyl groups, fused ring systems formed by cycloalkyl groups with heterocyclic alkyl groups, fused ring systems formed by cycloalkyl groups with aromatic rings, fused ring systems formed by cycloalkyl groups with heteroaromatic rings, fused ring systems formed by heterocyclic alkyl groups with aromatic rings, fused ring systems formed by heteroaromatic rings with heteroaromatic rings, and fused ring systems formed by heteroaromatic rings with aromatic rings.

[0278] In this application, the term "halogen" generally refers to fluorine, chlorine, bromine, or iodine, for example, fluorine or chlorine.

[0279] In this application, the term "each independently" means that at least two groups (or segments) in the structure with the same or similar value ranges can have the same or different meanings under specific circumstances. For example, if substituent X and substituent Y are each independently hydrogen, halogen, hydroxyl, cyano, alkyl, or aryl, then when substituent X is hydrogen, substituent Y can be hydrogen, halogen, hydroxyl, cyano, alkyl, or aryl; similarly, when substituent Y is hydrogen, substituent X can be hydrogen, halogen, hydroxyl, cyano, alkyl, or aryl.

[0280] In this application, the terms “optional” or “optionally” generally mean that the event or environment described below may but does not have to occur, and the description includes situations in which the event or environment occurs or does not occur. For example, “optionally alkyl-substituted heterocyclic group” means that an alkyl group may but does not have to be present, and the description can include cases where the heterocyclic group is substituted with an alkyl group and cases where the heterocyclic group is not substituted with an alkyl group.

[0281] In this application, the term "substitution" and its other variant forms herein refer to the replacement of one or more (e.g., 1, 2, 3, or 4) atoms or groups of atoms (e.g., hydrogen atoms) on a specified atom with other equivalents, provided that the replacement does not exceed the normal valence of the specified atom or group of atoms in the present case and is capable of forming a stable compound. If an atom or group of atoms is described as "optionally substituted," it may or may not be substituted. Unless otherwise stated, the linking site of a substituent herein may be derived from any suitable position of the substituent. When the linking bond in a substituent is shown as a chemical bond between two atoms connected to each other in a ring system, it indicates that the substituent may be linked to any one of the cyclic atoms in the ring system.

[0282] In this application, the term "replaced" for zero or more (e.g., zero or more than one, zero or one, zero) methylene units generally means that when the structure contains one or more methylene units, the one or more methylene units may not be replaced, or may be replaced by one or more groups described herein (e.g., -NHC(O)-, -C(O)NH-, -C(O)-, -OC(O)-, -C(O)O-, -NH-, -O-, -S-, -SO-, -SO2-, -PH-, -P(=O)H-, -NHSO2-, -SO2NH-, -C(=S)-, -C(=NH)-, -N=N-, -C=N-, -N=C- or -C(=N2)- etc.).

[0283] Without violating common sense in the field, the above-mentioned preferred conditions can be combined arbitrarily to obtain various preferred embodiments of the present invention.

[0284] The reagents and raw materials used in this invention are all commercially available.

[0285] The preparation method of the present invention has one or more effects selected from the group consisting of:

[0286] (1) High yield;

[0287] (2) The purification process is simple and has low impurities;

[0288] (3) Suitable for industrial production;

[0289] (4) High stability. Example

[0290] This invention includes all combinations of the specific embodiments described. Further embodiments of the invention and the full scope of its applicability will become apparent from the detailed description provided below. However, it should be understood that although the detailed description and specific embodiments indicate preferred embodiments of the invention, these descriptions and embodiments are provided by way of illustration only, as various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description. For all purposes, all disclosures, patents, and patent applications cited herein, including in quotation marks, are incorporated herein by reference in their entirety. The invention is further illustrated below by way of examples, but this does not limit the invention to the scope of the examples described. Experimental methods in the following examples, unless specific conditions are specified, are performed according to conventional methods and conditions, or as selected according to the trade specification.

[0291] Mass spectrometry (MS) measurements were performed using an Agilent (ESI) mass spectrometer, manufacturer: Agilent, model: Agilent 6200 series TOF / 6500 series Q-TOF.

[0292] Preparative high performance liquid chromatography (HPLC) was performed using a rapid preparative liquid chromatograph (Unisil 100-8DAC50 column).

[0293] Thin-layer chromatography purification was performed using GF 254 (0.4–0.5 nm) silica gel plates produced in Yantai.

[0294] The reaction was monitored using thin-layer chromatography (TLC) or liquid chromatography-mass spectrometry (LC-MS). The developing solvent systems used included, but were not limited to, dichloromethane and methanol systems, n-hexane and ethyl acetate systems, and petroleum ether and ethyl acetate systems. The volume ratio of the solvent was adjusted according to the polarity of the compound, or by adding triethylamine, etc.

[0295] Unless otherwise specified in the examples, the reaction temperature is room temperature (20℃~30℃).

[0296] Unless otherwise specified, the reagents used in the examples were purchased from Aladdin Biochemical Technology Co., Ltd., Shanghai Haoyuan Pharmaceutical Co., Ltd., Shanghai Bide Pharmaceutical Technology Co., Ltd., Anhui Zesheng Technology Co., Ltd., and other companies.

[0297] The above embodiments do not limit the scope of this application in any way. In addition to those described herein, various modifications to the invention will be apparent to those skilled in the art based on the foregoing description. Such modifications are also intended to fall within the scope of the appended claims. All references cited in this application (including all patents, patent applications, journal articles, books, and any other disclosures) are incorporated herein by reference in their entirety.

[0298] In this document, the meanings of the abbreviations included in conventional synthesis methods, preparation examples, and examples and intermediate synthesis examples are shown in the table below.

[0299] Example 1: Synthesis of Compound I

[0300] Step 1: Preparation of compound V

[0301] Compound 1 (1.13 g, 5.6 mmol) and compound 2 (2.31 g, 4.6 mmol) were added to tetrahydrofuran (10 mL), stirred until dissolved, and then DMTMM (1.93 g, 6.6 mmol) was added. The mixture was reacted at (25 ± 5) °C for 24 hours, then filtered under reduced pressure and the residue was discarded to obtain the organic phase. The organic phase was concentrated to dryness to give compound V (2.68 g, 85%). HRMS (ESI) calculated value C 29 H 52 N3O 13 S[M+H] + :682.3215, measured value 682.3223.

[0302] Table 1. Effects of different condensing agents, solvents, and condensation temperatures on yield

[0303] Step 2: Preparation of Compound IV

[0304] Compound V (2.68 g, 4.0 mmol) was diluted with dichloromethane (20 mL), and trifluoroacetic acid (10 mL) was slowly added at (20 ± 5) °C, with stirring for 2 hours. The organic solvent was then removed under reduced pressure, and the residue was dissolved in dichloromethane. Methyl tert-butyl ether was added and the mixture was stirred vigorously. After standing, the upper organic phase was discarded. The lower organic phase was then diluted with dichloromethane, dried thoroughly with anhydrous sodium sulfate, filtered, and concentrated to obtain the crude product. The crude product was then subjected to preparative HPLC separation to obtain compound IV (2.38 g, 95%). HRMS (ESI) calculated value C 28 H 56 N3O 13 S[M+H] +:674.3528, measured value 674.3524.

[0305] Step 3: Preparation of Compound III

[0306] Compound IV and compound 3 undergo an amide condensation reaction to generate compound III:

[0307] Compound IV (2.38 g, 5.4 mmol) and compound 3 (2.38 g, 5.2 mmol) were added to tetrahydrofuran (10 mL), stirred until dissolved, and then DMTMM (1.79 g, 6.1 mmol) was added. The mixture was reacted at (25 ± 5) °C for 18 hours, and then filtered under reduced pressure to obtain the organic phase. The organic phase was concentrated to dryness and dissolved in dichloromethane. Hexane and ethyl acetate were added and the mixture was stirred vigorously. After standing, the upper organic phase was discarded. The lower organic phase was diluted with dichloromethane, dried thoroughly with anhydrous sodium sulfate, filtered, and then concentrated to obtain the crude product. The crude product was separated by preparative HPLC to obtain compound III (3.28 g, 70%). HRMS (ESI) calculated value C 40 H 64 N6NaO 15 S[M+Na] + :923.4043, measured value 923.4060.

[0308] Table 2. Effects of different condensing agents, solvents, feed ratios, and reaction temperatures on yield.

[0309] Step 4: Preparation of Compound II

[0310] Compound III undergoes a nucleophilic substitution reaction to generate compound II:

[0311] Compound III (12.0 g, 13.3 mmol) was added to a 250 mL single-necked round-bottom flask, followed by the addition of dichloromethane (100 mL) and NMM (4.0 g, 40 mmol) and stirring until dissolved. A dichloromethane solution of phenyl p-nitrochloroformate (8.0 g, 40 mmol) was added to the solution, and the reaction was carried out under controlled temperature. The reaction was monitored by TLC until compound III was completely eliminated. After the reaction was complete, the solution was evaporated to dryness, dissolved in 1 V dichloromethane, and added dropwise to 5 V ethyl acetate. The mixture was stirred for 20 minutes and then filtered. The filtrate was evaporated to dryness, and n-hexane and ethyl acetate were added. After vigorous stirring, the mixture was separated to obtain crude compound II. The crude product was further purified by preparative HPLC to obtain compound II (8.6 g, 61%). 1H NMR (500MHz, DMSO-d6): δ10.0(s,1H),9.37(s,2H),9.11(t,J=5.5Hz,1H),8.32-8.29(m,2H),8.19(d,J=6.9 Hz,1H),7.88(d,J=8.6Hz,1H),7.65(d,J=8.6Hz,2H),7.58-7.55(m,2H),7.41(d,J=8.7Hz,2H),5.24(s,2H), 4.40(p,J=7.1Hz,1H),4.21(dd,J1=8.6Hz,J2=6.7Hz,1H),3.62-3.48(m,34H),3.45(s,3H),2.49-2.45(m,1H ),2.41-2.36(m,1H),2.02-1.92(m,1H),1.32(d,J=7.1Hz,3H),0.88(d,J=6.8Hz,3H),0.84(d,J=6.8Hz,3H). 13 C NMR (125MHz, DMSO-d6): δ171.6,171.3,170.8,166.8,162.4,158.2,155.7,152.4,145.6,139.9,130.2,130.0,129.6,125.8 ,123.0,119.4,70.7,70.16,70.08,69.8,69.0,67.3,57.9,49.8,39.9,39.5,36.3,31.0,19.5,18.5,18.2.HRMS(ESI)Calcd for C 47 H 67 N7O 19 SNa[M+Na] + :1088.4105,Found:1088.4147.

[0312] Table 3. Effects of different types of alkalis and solvents on yield

[0313] Step 5: Preparation of Compound I

[0314] Compound 4 (10.0 g) was added to a 250 mL single-necked round-bottom flask, and 100 mL of water was added and stirred to dissolve. The temperature was controlled at 25 ± 5 °C, and 100 mL of sodium bicarbonate aqueous solution (0.06 N) was slowly added dropwise to the solution, taking care not to generate a large number of bubbles. After reacting for 4 hours, the mixture was filtered, the filter cake was rinsed with 100 mL of water, and dried to obtain free eczema (8.3 g, 101%).

[0315] Free eczema (1.0 g, 2.4 mmol) was added to a 50 mL single-necked round-bottom flask A, followed by the addition of DMF (20 mL) and NMM (4.0 g, 4.0 mmol) and stirring until dissolved, yielding solution 1. Compound II (2.1 g, 2.0 mmol) was added to a 50 mL single-necked round-bottom flask B, followed by the addition of DMF (20 mL) and stirring until dissolved, yielding solution 2. Solution 1 was then added to solution 2, and the reaction was carried out at (0 ± 5) °C for 15 hours. The reaction was monitored by TLC (developing solvent: dichloromethane:methanol = 10:1 (V:V), with B00 (compound II) as a control). The reaction was stopped when the spot for compound II disappeared. If the spot for compound II was present, samples were taken every hour until compound II disappeared. Then, 10% sodium chloride aqueous solution and dichloromethane were added to the reaction solution, and the mixture was extracted and separated. The organic phase was washed three times with saturated brine, dried over anhydrous sodium sulfate, and concentrated to obtain crude compound I. The crude product was then subjected to preparative HPLC separation to obtain compound I (2.1 g, 77%). 1 H NMR (500MHz, DMSO-d6): δ9.91(s,1H),9.37(s,2H),9.10(t,J=5.5Hz,1H),8.16(d,J=7.0Hz,1H),8.04(d,J=8.7Hz,1H),7.87(d,J=8.6Hz,1H),7.7 5(d,J=10.9Hz,1H),7.60(d,J=8.5Hz,2H),7.37(d,J=8.4Hz,2H),7.30(s ,1H),6.50(s,1H),5.44(s,2H),5.31-5.21(m,3H),5.08(s,2H),4.42-4. 36(m,1H),4.21(dd,J1=8.6Hz,J2=6.8Hz,1H),3.62-3.48(m,34H),3.45( s,3H),3.26-3.22(m,1H),3.12-3.07(m,1H),2.49-2.44(m,1H),2.41-2. 37(m,1H),2.36(s,3H),2.44-2.14(m,2H),2.00-1.93(m,1H),1.92-1.81 (m,2H),1.31(d,J=7.1Hz,3H),0.89-0.86(m,6H),0.84(d,J=6.8Hz,3H). 13CNMR (125MHz, DMSO-d6): δ172.5,171.2,170.9,170.4,166.5,162.0,160.9,157.9,156.8,156.1,152.6, 150.0,148.1,147.9,145.2,141.0,138.8,136.5,136.4,131.6,129.9,128.7,125.4,123.8,123.6,121.6 ,119.2,119.1,110.0,109.8,96.7,72.4,69.83,69.79,69.75,69.5,68.7,67.0,65.8,65.3,57.5,49.9,4 9.1,47.5,39.6,39.2,36.0,30.6,30.4,28.3,23.9,19.2,18.2,18.0,11.04,11.00,7.8.HRMS(ESI)Calcd for C 65 H 84 FN9O 20 SNa[M+Na] + :1384.5430,Found:1384.5437.

[0316] Separation and purification:

[0317] Column packing: Pack 500g of 1010-C18-BS packing material into the chromatography system.

[0318] Equilibration: Mobile phase A (0.1% aqueous phosphoric acid solution): Mobile phase B (methanol) = 50%: 50%, 220nm UV detection, flow rate set at 1120mL / min, the detection baseline after equilibration should be flat and without significant fluctuations for more than 5 minutes.

[0319] Sample loading: Add 40% acetonitrile / phosphoric acid aqueous solution to the crude compound I, stir to dissolve, and filter to obtain the sample loading solution. Set the flow rate to 100-500 ml / min, and load the sample loading solution onto the column into the chromatography system. Wash with 200-1000 ml of equilibration buffer.

[0320] Elution: Gradient elution with mobile phase A and mobile phase B, detected by 220 nm UV.

[0321] Post-processing:

[0322] The collected liquid was added to a reaction vessel, and dichloromethane was added and stirred. The mixture was extracted three times by separation, and then washed three times with purified water and saturated sodium chloride solution, respectively. After drying with anhydrous sodium sulfate, the mixture was filtered to obtain the organic phase. The organic phase was concentrated to a certain volume and then slowly added to cyclohexane and ethyl acetate. After stirring and crystallizing for 1-1.5 hours, the mixture was filtered to obtain compound I.

[0323] Table 4 shows the yield of compound I prepared using the preparation method in step 5, with only the types of base and solvent changed while other conditions remained the same.

[0324] Table 4. Yields of Compound I prepared with different bases and solvents

[0325] Example 2

[0326] Preparation of compound 3

[0327] Compound 1 (15.0 g, 26.9 mmol) was added to a 200 mL single-necked round-bottom flask, followed by the addition of DMF (150 mL) and DIPEA (6.94 g, 53.7 mmol) and stirring until dissolved. Then, compound 2 (14.25 g, 26.9 mmol) and HOSu (1.55 g, 13.4 mmol) were added to the round-bottom flask. The mixture was reacted at (10-15) °C for 3 hours, and samples were taken. The reaction was monitored by TLC (developing solvent: dichloromethane:methanol = 10:1 (V:V), with compound 1 as a control). The reaction was stopped when the spot for compound 1 disappeared. If the spot for compound 1 remained, samples were taken every hour until compound 1 disappeared. After the reaction was complete, the reaction solution was slowly added to water, stirred, and filtered to obtain crude compound 3. The crude compound 3 was added to a reaction flask with ethanol and stirred for 30 minutes. Then purified water was added, and the mixture was stirred for 1 hour. The mixture was then filtered, and the solid was washed with a mixture of ethanol and water to obtain compound 3 (21.6 g, 94%). HRMS (ESI) calculated value C 45 H 51 FN6NaO 10 [M+Na] + :877.3543, measured value 877.3531.

[0328] Table 5. Yields of compound 3 prepared with different activators, feed ratios, and bases

[0329] Preparation of compound 4

[0330] Compound 3 (15.0 g, 17.5 mmol) was added to a 200 mL single-necked round-bottom flask, followed by the addition of dichloromethane (75 mL) and stirring until dissolved. Trifluoroacetic acid (37.5 mL) was slowly added while maintaining the temperature at 0 °C. The reaction was stopped after stirring at 0 °C for 1 hour. After the reaction was complete, the organic solvent was removed under reduced pressure. The remaining solution was added to methyl tert-butyl ether and stirred vigorously for 1 hour, resulting in precipitation of crude compound 4. The crude compound 4 (14.1 g, 93%) was obtained by preparative HPLC separation. HRMS (ESI) calculated value C 40 H 44 FN6O8[M-CF3CO2H+H] + 755.3199, measured value 755.3199.

[0331] HPLC separation and elution gradient:

[0332] Gradient elution was performed using mobile phase A (0.1% aqueous phosphoric acid solution) and mobile phase B (methanol), with detection at 220 nm UV.

[0333] Table 6. Yields of compound 4 prepared with different acids, feed ratios, and reaction temperatures

[0334] Preparation of Compound I

[0335] Compound IV (625.7 mg, 1.0 mmol) was added to a 25 mL single-necked round-bottom flask. Then, DMSO (10 mL), DIPEA (258.5 mg, 2.0 mmol), and HATU (456.3 mg, 1.2 mmol) were added at -20 °C, and the reaction was continued for 30 min. Next, compound 4 (868.8 mg, 1.0 mmol) was added, and a sample was taken after 0.5 hours of reaction. HPLC monitoring showed that compound 4 had completely reacted, and the reaction was stopped. The reaction solution was then added to methyl tert-butyl ether (5V), stirred under a nitrogen atmosphere for 0.5 h, and then allowed to stand. The upper organic phase was discarded to obtain crude compound I. The crude product was separated by preparative HPLC (separation conditions as described in step 5 of Example 1) to obtain compound I (1.13 g, 83%).

[0336] Table 7. Yields of Compound I prepared with different condensing agents and solvents

[0337] Example 3

[0338] Preparation of compound IX

[0339] Compound a-1 (754.8 mg, 1.0 mmol) was added to a 25 mL single-necked round-bottom flask. DMF (10 mL), DIPEA (258.5 mg, 2.0 mmol), and HATU (456.3 mg, 1.2 mmol) were then added at 0 °C, and the reaction was continued for 30 min. Compound 4 (compound a-2) (663.8 mg, 1.0 mmol) was then added. After 4 hours of reaction, a sample was taken, and HPLC monitoring showed complete reaction of compound a-1. The reaction was then stopped. Acetonitrile (50 mL) and 0.1% phosphoric acid aqueous solution (50 mL) were added to the reaction solution to obtain a preparative loading solution. This solution was then separated by preparative HPLC (under the same conditions as described in step 5 of Example 1) to obtain compound IX (700.6 mg, 50%). HRMS (ESI) calculated value C 74 H 91 FN7O 19 [M+H] + :1400.6348, measured value 1400.6329.

[0340] Preparation of compounds I-VII

[0341] Compound IX (1.4 g, 1 mmol) was added to a 50 mL single-necked round-bottom flask. DMF (10 mL) and piperidine (851.6 mg, 10 mmol) were then added at 20 °C. After reacting for 2 hours, a sample was taken, and HPLC monitoring showed complete reaction of compound IX. The reaction was then stopped. Acetonitrile (50 mL) and 0.1% phosphoric acid aqueous solution (50 mL) were then added to the reaction solution to obtain a preparative loading solution. This solution was then separated by preparative HPLC (under the same conditions as described in step 5 of Example 1) to obtain compounds I-VII (801.3 mg, 68%). HRMS (ESI) calculated value C 59 H 81 FN7O 17 [M+H] + :1178.5667, measured value 1178.5680.

[0342] Preparation of compounds I-VI

[0343] Compound e (168.2 mg, 1.0 mmol) was added to a 50 mL single-necked round-bottom flask. Then, DMF (20 mL), NMM (202.3 mg, 2.0 mmol), CDMT (351.1 mg, 2.0 mmol), and compounds I-VII (1178.5 mg, 1.0 mmol) were added at 20 °C. After reacting for 8 hours, a sample was taken, and HPLC monitoring showed complete reaction of compound e. The reaction was then stopped. Acetonitrile (100 mL) and 0.1% phosphoric acid aqueous solution (100 mL) were added to the reaction solution to obtain a preparative loading solution. This solution was then separated by preparative HPLC (under the same conditions as described in step 5 of Example 1) to obtain compounds I-VI (585.4 mg, 44%). HRMS (ESI) calculated value C 65 H 85 FN9O 18 [M+H] + :1330.5712, measured value 1330.5704.

[0344] Preparation of Compound I

[0345] Compound I-VI (13.3 g, 10 mmol) was added to a 1000 mL single-necked round-bottom flask. Then, DCM (500 mL) and m-chloroperoxybenzoic acid (2.07 g, 12 mmol) were added at -20 °C. After reacting for 12 hours, a sample was taken, and HPLC monitoring showed complete reaction of compound I-VI. The reaction was then stopped. The reaction solution was then added to methyl tert-butyl ether (5V), stirred for 0.5 h under a nitrogen atmosphere, and allowed to stand. The upper organic phase was discarded to obtain crude compound I. The crude product was then separated by preparative HPLC (under the same conditions as described in step 5 of Example 1) to obtain compound I (5.45 g, 40%).

Claims

1. A process for the preparation of a compound of formula I, comprising the step of a nucleophilic substitution reaction of a compound of formula I-i with compound a to give a compound of formula I, ###0001### I I-i a wherein: L2is -(C(R L21 )2) n -, any of the units CHR in L2 L21 may each independently be replaced by the following structural units: -Cy-, -C(O)-, NR L22 , -O-, -S-, -SO-, SO2, -P(R L22 )-, -(P=O)R L22 , -C(=S)-, C(=NR L22 ), -N=N-, -C=N-, n is a natural number from 0 to 50; L3 is any combination of amino acid residues, short peptides consisting of 2 to 10 amino acid residues, said amino acid residues being natural or unnatural amino acid residues; Tr is or any combination of the above groups; " " indicates a bond to L 3 Connection; - Cy- is selected from phenylene, 5- to 8-membered heteroarylene, 3- to 10-membered heterocyclylene, or 3- to 10-membered cycloalkylene, wherein said -Cy- is unsubstituted or independently substituted with 1 or more R x substituents, R L21 R L22 R cx Each is independently selected from hydrogen, deuterium, halogens, -NO2, -CN, and -OR. L2a -SR L2a -N(R) L2a )2、-N + (R L2a 3、-CH2C(O)R L2a --OC(O)R L2a -N(R) L2a SO2R L2b -N(R) L2a COR L2b --(N(Me)CH2C(O)) m -OR L2a --(N(Me)CH2C(O)) m -NHR L2a -(N(Me)CH2C(O) m -N*(R L2a 3、-(CH2) y -NHCOCH2(OCH2CH2)OR L2a -(CH2) y -NH(COCH2(N(Me) m -R L2a -NH-(CH2CH2O) m -R L2a -(CH2CH2O) m -R L2a -(CH2N(Me)) m -R L2a -CH2(OCH2CH2) m -OR L2a -(CH2CH2O) m -R L2a -NH-(CH2CH2O) m -R L2a or R L2a Optional substitution of -C 1-6 Alkyl, -C 1-6 alkenyl, -C 1-6 Alkyne, 3-8 membered cycloalkyl, 4-10 membered heterocycloalkyl, 6-10 membered aryl or 3-10 membered heteroaryl, m, y are each a natural number from 0 to 50, R L2a , R L2b are each independently selected from hydrogen, deuterium, halogen, -N02, -CN, -OH, -SH, -NH2, -N(Me)2, -C02H, -S(0)2Me, -S(0)2OH, -C(0)NH2, -SO2NH2, -C 1-6 alkyl, -C 1-6 alkenyl, -C 1-6 alkynyl, 3-8 membered cycloalkyl, 4-10 membered heterocycloalkyl, 6-10 membered aryl, or 3-10 membered heteroaryl; R1, R2are each independently selected from the group consisting of H, -OH, -CN, halogen, C 1-3 alkyl, C 1-3 alkoxy; Rx is selected from H, halo, or C 1-3 alkyl; p is 0, 1 or 2.

2. The production method according to claim 1, wherein L3 is selected from Val, D-Val, Phe, Lys, Leu, lie, Gly, Ala, Cit, Asp, Asn, Glu, Gin, Val-Cit, Val-Ala, Val-Lys, Val-Lys(Ac), Phe-Lys, Phe-Lys(Ac), Leu-Lys, Leu-Lys(Ac), Ala-Ala, Ala-Lys, D-Ala-Ala, Gly-Glu, Gly-Asp, Gly-Asn, Val-Glu, Val-Asp, Asn-Asn, Asp-Glu, Gly-Gly-Glu, Gly-Gly-Asp, Gly-Gly-Asn, Gly-Ala-Ala, Gly-Val-Ala, Gly-Val-Cit, Glu-Val-Cit, Ala-Ala-Ala, Ala-(D-Ala)-Ala, Ala-Ala-Asn, Ala-(D-Ala)-Asn, Ala-Ala-Asp, Val-Lys-Gly, D-Val-Leu-Lys, Gly-Gly-Arg, Gly-Gly-Gly, Lys-Ala-Asn, Gly-Phe-Gly, Gly-Gly-Phe, Asn-Pro-Val, Ala-Lys-Gly, Gly-Lys-Gly, Gly-Gly-Gly-Gly, Gly-Gly-Phe-Gly, Gly-Gly-Glu-Gly, Lys-Ala-Ala-Asn, Lys-Ala-Ala-Asp, Ala-Ala-Pro-Val, Ala-Ala-Pro-Nva, or any combination of the above fragments; Preferably, L3 is selected from Lys, Gly, Asp, Asn, Glu, Gin, Val-Cit, Val-Ala, Ala-Ala, Gly-Glu, Gly-Asp, Gly-Asn, Asn-Asn, Asp-Glu, Gly-Glu-Gly, Gly-Gly-Phe-Gly, or any combination of the above fragments; More preferably, L3 is selected from Lys, Gly, Val-Ala, Ala-Ala, Gly-Glu, Gly-Asp, Gly -Asn, Asn-Asn, Asp-Glu, Gly-Gly-Phe-Gly. More preferably, L3 is selected from Val-Ala, Ala-Ala, Gly-Glu, Gly-Asp, Gly-Asn, Asn-Asn, Asp-Glu, Gly-Gly-Phe-Gly.

3. The production method according to claim 1 or 2, characterized by, said L2is -(C(R L21 )2) n -, any of the units CHR in L2 L21 may each independently be replaced by the following structural units: -Cy-, -C(O)-, NR L22 , -O-; n is a natural number from 0 to 50; - Cy- is selected from phenylene, 5- to 6-membered heteroarylene, 4- to 10-membered heterocyclylene, or 3- to 6-membered cycloalkylenyl, wherein said -Cy- is unsubstituted or independently substituted with 1 or more R x substituents, R L21 R L22 R cx Each is independently selected from hydrogen, halogen, and -OR L2a -SR L2a -N(R) L2a )2、-N(R L2a SO2R L2b -N(R) L2a COR L2b -(N(Me)CH2C(O) m -OR L2a --(N(Me)CH2C(O)) m -NHR L2a -(N(Me)CH2C(O) m -N*(R L2a 3. -NH-(CH2CH2O) m -R L2a -(CH2CH2O) m -R L2a -(CH2N(Me)) m -R L2a -CH2(OCH2CH2) m -OR L2a -(CH2CH2O) m -R L2a -NH-(CH2CH2O) m -R L2a or R L2a Optional substitution of -C 1-6 Alkyl, -C 1-6 alkenyl, -C 1-6 Alkyne, 3-8 membered cycloalkyl, 4-10 membered heterocycloalkyl, 6-10 membered aryl or 3-10 membered heteroaryl, m is a natural integer from 0 to 8, y is 0, 1, 2, 3 or 4, R L2a R L2b Each is independently selected from hydrogen, halogen, -CN, -OH, -NH2, -N(Me)2, -CO2H, -C(O)NH2, -C 1-6 alkyl.

4. The production method according to claim 3, wherein said L2is -(CH2) n - and Any of the units CH2in L2may each independently be replaced with a structural unit selected from the group consisting of 4-6 membered heterocyclylene, 3-6 membered cycloalkylen, -C(O)-, NR L22 , -O-; each R is independently selected from hydrogen, -OR L22 hydrogen, -OR L2a , or -NR L2a optionally substituted -C 1-6 alkyl, Each R L2a Independently selected from hydrogen, halogens, -CN, -OH, -NH2, -N(Me)2, -CO2H, -C(O)NH2, -C 1-6 alkyl.

5. The production method according to any one of claims 1 to 4, wherein L2 is: The compounds I-i are: The compound a is 6. The production method according to any one of claims 1 to 5, wherein It comprises any of the following characteristics: (1) the compound a exists in the form of a salt, Preferably, the salt of the compound a is first added with a base to obtain the free form of the compound a when the nucleophilic substitution reaction is carried out, Further preferably, the salt of the compound a is the methanesulfonic acid salt of the compound a, and sodium bicarbonate is first added to obtain the free form of the compound a when the nucleophilic substitution reaction is carried out; (2) the reactants of the nucleophilic substitution reaction further comprise a base, and the base is one or more of DIPEA, N-methylmorpholine, 2,6-lutidine, preferably N-methylmorpholine; (3) the nucleophilic substitution reaction is carried out in a solvent, and the solvent is an organic solvent, preferably one or more of DMF, DMAc, DMSO, 1,4-dioxane, more preferably DMSO or DMF; (4) the molar volume ratio of the compound I-i to the organic solvent is 0.08-0.4 mol / L, preferably 0.1 mol / L; (5) the molar volume ratio of the compound a to the organic solvent is 0.08-0.18 mol / L, preferably 0.12 mol / L; (6) the molar ratio of the compound I-i to the compound a is 1:1-1:1.5, preferably 1:1.2; (7) the molar ratio of the compound I-i to the base is 1:1-1:4, preferably 1:2; (8) the reaction temperature of the nucleophilic reaction is room temperature, or 15-25°C, preferably 20°C; (9) the feeding sequence of the nucleophilic substitution reaction is: dissolving compound I-i in the solvent, then adding a base, and then adding the reaction solution A after stirring; (10) the reaction time of the nucleophilic substitution reaction is 8-24 hours, 12-18 hours or 16 hours; (11) the nucleophilic substitution reaction further comprises a post-treatment step, and the post-treatment step is extraction, washing, separation and purification; Preferably, the organic phase used for the extraction is dichloromethane, and the aqueous phase is water; and / or the liquid used for the washing is saturated brine; and / or the chromatographic column used for the separation and purification is 1010-C18-BS, the mobile phase A of the chromatographic column is 0.1% phosphoric acid aqueous solution, the mobile phase B is methanol, the ratio of the mobile phase A to the mobile phase B is 50%:50%, and the flow rate of the separation and purification is 1120 mL / min.

7. The preparation method of any one of claims 1-6, wherein, the preparation method further comprises a substitution reaction of a compound of formula I-ii with compound b to prepare the compound of formula I-i, or In the process for the preparation of the compound of formula I, the compound of formula I-i is prepared by a process comprising a substitution reaction of a compound of formula I-ii with a compound b, wherein, Tx is -OH or X is halogen; preferably, X is F, Cl, Br or I.

8. The production method according to claim 7, wherein It comprises any of the following characteristics: (1) the substituting reaction further comprises a base, and the base is one or more of sodium bicarbonate, DIPEA, triethylamine, N-methylmorpholine, imidazole, pyridine, DBU, 2,6-lutidine, diethylamine, preferably N-methylmorpholine; (2) the substituting reaction is carried out in an organic solvent, and the organic solvent is one or more of THF, DMF, EA, DCM, 1,4-dioxane, DMA, preferably THF, DMF, DMA, DCM, more preferably DCM; (3) the molar volume ratio of the compound of formula I-ii to the organic solvent is 0.1-0.16 mol / L, preferably 0.133 mol / L; (4) the molar volume ratio of the compound b to the organic solvent is 0.2-0.8 mol / L, preferably 0.4 mol / L; (5) the molar ratio of the compound of formula I-ii to the compound b is 1:2-1:5, preferably 1:3; (6) the molar ratio of the compound of formula I-ii to the base is 1:2-1:5, preferably 1:3; (7) the substituting reaction is carried out at room temperature, or at 20-30°C, preferably at 25°C; (8) the substituting reaction further comprises a pretreatment of the reactants before the reaction, and the pretreatment is dissolving the compound of formula I-ii and the base in a solvent; preferably, the temperature of the pretreatment is -5-5°C, preferably 0°C; (9) the substituting reaction is carried out for 4-12 hours, or for 8 hours; (10) the substituting reaction further comprises a post-treatment step, and the post-treatment step is concentration, crystallization, extraction; wherein: preferably, the concentration is carried out in a 30-40°C water bath, preferably in a 35°C water bath; and / or, the crystallization is carried out by adding dichloromethane to the solution, and the layers are allowed to stand; and / or, the extraction comprises extraction with n-hexane and ethyl acetate, and the extraction is further followed by reduced pressure concentration, and the reduced pressure concentration is carried out in a 30-40°C water bath, preferably in a 35 °C water bath.

9. The preparation method of claim 7 or 8, wherein the preparation method further comprises condensation reaction of a compound of formula I-iii with a compound c to prepare the compound of formula I-ii, or wherein the compound c is H-L3-Tx. which comprises any one of the following features: In the method for preparing the compound of formula I, the compound of formula I-ii is prepared by a method comprising condensation of a compound of formula I-iii with a compound c, (1) the condensation reaction further comprises a condensing agent, and the condensing agent is one or more of DMTMM, CDMT, NMM, HATU, EDCI, preferably HATU, DIPEA, or CDMT, NMM, or DMTMM, more preferably DMTMM; 10. The production method according to claim 9, wherein (2) the condensation reaction is carried out in an organic solvent, and the organic solvent is one or more of THP, DMAc, DCM, preferably THF, DMAc; (3) the molar volume ratio of the compound of formula I-iii to the organic solvent is 0.3-0.7 mol / L, preferably 0.54 mol / L; ​ ​ (4) the molar volume ratio of the compound c to the organic solvent is 0.3-0.7 mol / L, preferably 0.52 mol / L; (5) the molar ratio of the compound of formula I-iii to the compound c is 1:0.6-1:1.5, preferably 1:0.67, 1:0.83, 1:1.1 or 1:1.2; (6) the molar ratio of the compound of formula I-iii to the condensing agent is 1:1.2-1:1.8, preferably 1:1.5; (7) the reaction temperature of the condensation reaction is 5-30°C, preferably 20-30°C, more preferably 25°C; (8) the reaction time of the condensation reaction is 12-24 hours, or 18 hours; (9) the condensation reaction further comprises a post-treatment step, and the post-treatment step is filtration, extraction, drying; wherein: preferably, the filtration is filtration under reduced pressure, and the residue is removed, and the step after the filtration preferably further comprises concentration, and the concentration is performed in a water bath at 30-40°C, preferably in a water bath at 35°C; and / or, the extraction comprises extraction with n-hexane and ethyl acetate, and the extraction is further followed by drying, and the drying agent used for the drying is anhydrous sodium sulfate, and the drying is further followed by concentration under reduced pressure, and the concentration under reduced pressure is performed in a water bath at 30-40°C, preferably in a water bath at (10) in the preparation method of the compound of formula I, the preparation method of the compound of formula I-i is that the compound of formula I-iii is subjected to condensation reaction with compound c, and the reaction solution containing the compound of formula I-ii obtained is directly subjected to the substitution reaction with the compound b.

11. The preparation method of claim 9 or 10, wherein the preparation method further comprises that the compound of formula I-iv is subjected to hydrolysis reaction with a strong acid to prepare the compound of formula I-iii, or the preparation method comprises any one of the following characteristics: (1) the strong acid is trifluoroacetic acid; In the method for preparing the compound of formula I, the compound of formula I-iii is prepared by a method comprising hydrolysis of a compound of formula I-iv with a strong acid, Preferably, said I-iv is: Preferably, said I-iii is:

12. The production method according to claim 11, wherein (2) the hydrolysis reaction is performed in an organic solvent, and the organic solvent is one or more of THF, DMAc, DCM, preferably DCM; (3) the molar volume ratio of the compound of formula I-iii to the organic solvent is 0.3-0.7 mol / L, preferably 0,54 mol / L; (4) the molar volume ratio of the compound I-iv to the organic solvent is 0.1-0.4 mol / L, preferably 0.2 mol / L; (5) the molar volume ratio of the compound I-iv to the strong acid is 0.2-0.7 mol / L, preferably 0.4 mol / L; (6) the reaction temperature of the hydrolysis reaction is 15-25°C, preferably 20°C; (7) the reaction time of the hydrolysis reaction is 1-5 hours, or 2 hours; (8) the hydrolysis reaction further comprises a post-treatment step, and the post-treatment step is concentration, extraction, drying; ​ ​ Preferably, the concentration is performed in a water bath at 30-40°C, preferably in a water bath at 35°C; and / or, the extraction comprises extraction with dichloromethane and methyl tert-butyl ether, the extraction further comprises drying after the extraction, the drying agent used for the drying is anhydrous sodium sulfate, the drying further comprises concentration under reduced pressure after the drying, the concentration under reduced pressure is performed in a water bath at 30-40°C, preferably in a water bath at35°C.

13. The preparation method of claim 11 or 12, wherein, the preparation method further comprises an amide condensation reaction of the compound of formula I-v with compound 1 to prepare the compound of formula I-iv, or In the method for preparing the compound of formula I, the compound of formula I-iv is prepared by a method comprising an amide condensation reaction of a compound of formula I-v with compound 1, Preferably, said I-iv is:

14. The production method according to claim 13, wherein which comprises any one of the following characteristics: (1) the reactants of the amide condensation reaction further comprise a condensing agent, the condensing agent is one or more of DMTMM, CDMT, HATU, EDCl, preferably DMTMM; (2) the amide condensation reaction is performed in an organic solvent, the organic solvent is one or more of THF, DMAc, DCM, DMF, preferably DMF, DMAc or THF; (3) the molar volume ratio of the compound I-v to the organic solvent is 0.2-0.6 mol / L, preferably 0.46 mol / L; (4) the molar volume ratio of the compound 1 to the organic solvent is 0.4-0.7 mol / L, preferably 0.56 mol / L; (5) the molar ratio of the compound I-v to the compound 1 is 1:0.8-1:1.5, preferably 1:1.1 or 1:1.2; (6) the molar ratio of the compound I-v to the condensing agent is 1:1.2-1:1.8, preferably 1:1.5; (7) the reaction temperature of the amide condensation reaction is 10-30°C, preferably 20-30°C, more preferably 25°C; (8) the reaction time of the amide condensation reaction is 18-36 hours, or 24 hours; (9) the amide condensation reaction further comprises a post-treatment step, the post-treatment step is extraction, drying, filtration; Preferably, the extraction is preceded by concentration, the concentration is performed in a water bath at 30-40°C, preferably in a 35°C water bath; and / or, the extraction comprises extraction with methyl tert-butyl ether, the extraction further comprises drying after the extraction, the drying agent used in the drying is anhydrous sodium sulfate, the drying further comprises concentration under reduced pressure after drying, the concentration under reduced pressure is performed in a water bath at 30-40°C, preferably in a water bath at 35°C.

15. A method of preparing a compound of Formula I-i comprising a substitution reaction of a compound of Formula I-ii with compound b, ###00033### I-i I-ii b wherein: Tx is -OH or -CH2OH X is halogen; L2, L3, Tr are as defined in any one of claims 1-4; the reaction conditions of the substitution reaction are preferably as described in claim 8; the compound of formula I-ii can be prepared from a condensation reaction of a compound of formula I-iii with compound c as described in claim 9; the reaction conditions of the condensation reaction are preferably as described in claim 10; the compound of formula I-iii can be prepared from a hydrolysis reaction of a compound of formula I-iv with a strong acid as described in claim 11; the reaction conditions of the hydrolysis reaction are preferably as described in claim 12; The compound of formula I-iv can be prepared by an amide condensation reaction of a compound of formula I-v with compound 1, as described in claim 13; The reaction conditions of the amide condensation reaction are preferably as described in claim 14.

16. A method of preparing a compound of Formula I-ii comprising condensing a compound of Formula I-iii, ###00023### I-iii with compound c, ###00024### c wherein: compound c is H-L3-Tx; Tx is -OH or -NHRx2; and L2, L3 are as defined in any one of claims 1 to 4; The reaction conditions of the condensation reaction are preferably as described in claim 10; The compound of formula I-iii can be prepared by a hydrolysis reaction of a compound of formula I-iv with a strong acid, as described in claim 11; The reaction conditions of the hydrolysis reaction are preferably as described in claim 12; The compound of formula I-iv can be prepared by an amide condensation reaction of a The reaction conditions of the amide condensation reaction are preferably as described in claim 14.

17. A method of preparing a compound of Formula I-iii, comprising hydrolyzing a compound of Formula I-iv, ###00019### I-iii I-iv with a strong acid. L2 is as defined in any one of claims 1 to 4; The reaction conditions of the hydrolysis reaction are preferably as described in claim 12; compound 1, as described in claim 13; The reaction conditions of the amide condensation are preferably as described in claim 14. L2 is as defined in any one of claims l to 4; 18. A method of preparing a compound of Formula I-iv comprising amide condensation of a compound of Formula I-v with Compound 1, The reaction conditions of the amide condensation reaction are preferably as described in claim 14. wherein: L2, L3, Tr, Rx, p, R1, R2 are as defined in any one of claims 1 to 4; Preferably, the method for preparing a compound of formula I-iii is as described in claim 17; 19. A method of preparing a compound of Formula I, comprising dehydrating condensing a compound of Formula I-iii with compound a-1, L2, L3, Tr, Rx, p, R1, R2 are as described in any one of claims 1 to 4. It comprises any one of the following features: (1) The dehydrative condensation of the compound of formula I-iii with compound a-1 further comprises a condensing agent, which is HBTU, TBTU, EEDQ, EDCl, T4P, DMTMM, TSTU or HATU, preferably HBTU, TBTU, HATU, more preferably HATU; Preferably, the compound a-1 is: Preferably, the compound of formula I-iii is:

20. The production method according to claim 19, wherein (2) The dehydrative condensation of the compound of formula I-iii with compound a- 1 further comprises a base, which is DIPEA or TEA, preferably DIPEA; 21. The production method according to claim 20, wherein (3) The dehydrative condensation of the compound of formula I-iii with compound a- l is carried out in an organic solvent, which is DMAc, DMSO, MeCN, DMF, NMP or HMPA, preferably DMAc, DMSO, MeCN, DMF or NMP, more preferably DMSO; (4) The molar volume ratio of compound a-1 to the organic solvent is 0.07-0.12 mol / L, preferably 0.1 mol / L; (5) The molar volume ratio of the compound of formula I-iii to the organic solvent is 0.07-0.12 mol / L, preferably 0.l mol / L; (6) The molar ratio of compound a-1 to the compound of formula I-iii is 1:0.8-1:1.5, preferably 1:1; ​ ​ ​ (7) the molar ratio of the compound a-1 and the condensing agent is 1:1.1-1:1.8, preferably 1:1.2; (8) the molar ratio of the base and the compound a-1 is 1:1-3:1, preferably 2:1; (9) the reaction temperature of the dehydration condensation of the compound of formula I-iii and compound a-1 is -30--10°C, preferably -25--15°C, more preferably -20°C; (10) the reaction time of the dehydration condensation of the compound of formula I-iii and compound a-1 is 0.2-1 hour, or 0.5 hour; (11) the dehydration condensation of the compound of formula I-iii and compound a-1 further comprises a post-treatment step, which is extraction, HPLC separation; wherein: the post-treatment step is preferably adding methyl tert-butyl ether in the reaction solution of the reaction, and then performing HPLC preparation after extraction; the condition of the HPLC is more preferably mobile phase A (0.1% phosphoric acid aqueous solution): mobile phase B (methanol) = 50%:50%, flow rate 1120 mL / min, elution time 5 min.

22. The preparation method of any one of claims 19-21, wherein, the preparation method further comprises performing a deprotection reaction of compound a-i to prepare the compound a-1, or the preparation method of the compound of formula I, wherein the compound a-1 is prepared by a method comprising a deprotection reaction of compound a-i, wherein the definitions of groups L3, Tr, Rx, p, R1, R2 are as described in any one of claims 1-4; which comprises any one of the following characteristics: wherein the protecting group is a -Boc group, (1) the deprotection reaction of compound a-i further comprises an organic acid, and the organic acid is hydrochloric acid, zinc bromide, TMSOTf or trifluoroacetic acid, preferably hydrochloric acid or trifluoroacetic acid, more preferably trifluoroacetic acid; Preferably, the compounds a-i are preferably 23. The production method according to claim 22, wherein (2) the deprotection reaction of compound a-i is carried out in an organic solvent, and the organic solvent is methanol, dichloromethane, 1,4-dioxane, ethyl acetate, preferably methanol, dichloromethane, ethyl acetate, more preferably dichloromethane; (3) the molar volume ratio of compound a-i to the organic solvent is 0.1-0.4 mol / L, preferably 0.23 mol / L; (4) the volume ratio of the organic acid to the organic solvent is 1:2-1:5, preferably 1:2; (5) the volume molar ratio of the organic acid to compound a-i is 3:1-0.8:1, preferably 2.5:1-1:1, more preferably 2:1-2.5:1, still preferably 2.5:1, 2.1:1 or 2:1; (6) the reaction temperature of the deprotection reaction of compound a-i is -20-20°C, preferably -20--10°C, more preferably 0-10°C, still more preferably 0°C; (7) the reaction time of the deprotection reaction of compound a-i is 0.5-4 hours, or 1 hour; (8) the deprotection reaction of compound a-i further comprises a post-treatment step, which is distillation, extraction, HPLC separation. ​ ​ 24. The production method according to claim 22 or 23, characterized by, The preparation method further comprises an addition reaction of compound a and compound f to prepare the compound a-i, or The compounds a-i are prepared by a process comprising the addition reaction of compound a with compound f, wherein the definitions of groups L3, Tr, Rx, p, R1, R2 are as described in any one of claims 1-4.

25. The production method according to claim 24, wherein It comprises any one of the following characteristics: (1) The addition reaction further comprises an activating agent, which is DMAP, HOAt, HOSu or HOBt, preferably DMAP, HOSu or HOBt, more preferably HOSu; (2) The addition reaction further comprises a base, which is DIPEA or NMM, preferably DIPEA; (3) The addition reaction is carried out in an organic solvent, which is DMAc, DMSO, MeCN, DMF, NMP or DIPEA, preferably DMF; (4) The molar volume ratio of the compound a to the organic solvent is 0.07-0.2 mol / L, preferably 0.18 mol / L; (5) The molar volume ratio of the compound f to the organic solvent is 0.07-0.2 mol / L, preferably 0 18 mol / L; (6) The molar ratio of the compound f to the compound a is 1:0.8-1:1.5, 1:0.9-1:1, more preferably 1:1; (7) The molar ratio of the activating agent to the compound a is 0.2:1-1.5:1, preferably 0.5:1-1.5:1, more preferably 0.5:1, 1:1 or 1.5:1; (8) The molar ratio of the base to the compound a-1 is 1:1-3:1, preferably 2:1; (9) The reaction temperature of the addition reaction is 10-15℃; (10) The reaction time of the addition reaction is 2-5 hours, or 3 hours; (11) The addition reaction further comprises a post-treatment step, which is extraction, filtration.

26. A method for preparing compound a-1, characterized in that, The preparation method comprises a deprotection reaction of compound a-i, wherein the protecting group is a -Boc group, wherein: the definitions of groups L3, Tr, Rx, p, R1, R2 are as claimed in any one of claims 1-4, Preferably, the preparation conditions of the compound a-1 are as claimed in claim 23; Preferably, the preparation method further comprises an addition reaction of compound a and compound f to prepare the compound a-i. In the production method of the compound a-1, the compound a-i is produced by a method including addition reaction of the compound a with the compound f, wherein: the definitions of groups L3, Tr, Rx, p, R1 and R2 are as claimed in any one of claims 1-4, Preferably, a preparation method of the compound a-i is as claimed in claim 25.

27. A method of making a compound a-i comprising adding compound a to compound f to obtain said compound a-i, wherein: the definitions of groups L3, Tr, Rx, p, R1and R2are as claimed in any one of claims 1-4, Preferably, a preparation method for the compound a-i is as claimed in claim 25.

28. A method for preparing a compound as shown in Formula I, characterized in that, The preparation method comprises substituting the compounds shown in formulae I-VI, wherein the definitions of groups L2, L3, Tr, Rx, p, R1, R2 are as claimed in any of claims 1-4.

29. The production method according to claim 28, wherein The preparation method further comprises a condensation reaction of a compound represented by formula I-VII and compound e to prepare the compound represented by formula I-VI, or The compounds of formulae I-VI are prepared by a process comprising condensation of a compound of formula I-VII with a compound e, 30. The preparation method according to claim 29, characterized in that, The preparation method further comprises a deprotection reaction of a compound represented by formula I-X and piperidine to prepare the compound represented by formula I-VII, or The preparation method further comprises a condensation reaction of a compound represented by formula I-VII and a compound e to prepare the compound represented by formula I-VI, or The preparation method further comprising a deprotection reaction of a compound represented by formula I-X and piperidine to prepare a compound represented by formula I-VII, or The compounds of formulae I-VII are prepared by a process comprising deprotection of a compound of formula I-X with piperidine, 31. The production method according to claim 30, wherein the preparation method further comprises condensation reaction of compound a-1 with compound a-2 to prepare the compound of formula I-X, or The compounds of formula I-X are prepared by a process comprising condensation of compound a-1 with compound a-2, 32. The production method according to claim 31, wherein the preparation method further comprises substitution reaction of compound I-VIII with piperidine to prepare the compound a-1, or The compound a-1 is prepared by a method comprising a substitution reaction of compounds I-VIII with piperidine, 33. The production method according to claim 32, wherein the preparation method further comprises substitution reaction of compound a with compound a-3 to prepare the compound of formula I-VII, or the preparation method further comprises substitution reaction of compound a with compound a-3 to prepare the compound of formula I-VII, or The compounds of formulae I-VII are prepared by a process comprising a substitution reaction of compound a with compound a-3, 34. A compound having the structure: ###00019### 34

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