Linker-drug compounds and methods for producing the same

Abstract

This application relates to a method for producing linker-drugs. In particular, it provides a method for producing a target compound. This production method includes the steps of reacting a compound represented by formula a with a Lewis acid to obtain a crude product, and purifying the crude product to obtain a compound represented by formula A. According to this production method, quality control standards for the product can be achieved.

Description

[Technical Field]

[0001] Cross-reference of related applications This application claims priority under Chinese Patent Application No. CN202311077479.4 filed on 23 August 2023 and Chinese Patent Application No. CN202411139357.8 filed on 19 August 2024, the disclosures of which are incorporated herein by reference in their entirety. This application relates to the field of medicinal chemistry, and more particularly to a method for producing linker-drug compounds. [Background technology]

[0002] Antibody-drug conjugates (ADCs) are therapeutic molecules with targeted killing effects. ADCs are formed by conjugating a small molecule cytotoxic drug to an antibody via a chemical linker. ADCs are primarily used to treat tumors and other diseases. In ADCs, the antibody component can specifically bind to proteins expressed on the surface of tumor cells, enabling targeted delivery of cytotoxic drugs and yielding therapeutic effects and possibilities unattainable with conventional drugs. Currently, several antibody-drug conjugates have been approved for market worldwide, and numerous other candidate substances are in clinical evaluation.

[0003] Linker-drugs are compounds formed by linking a chemical linker to a low-molecular-weight cytotoxic drug. Linker-drugs function as intermediates in the production of ADCs and are important components in the construction of the entire antibody-drug conjugate.

[0004] The aforementioned description of related technologies is provided to facilitate understanding of the technical field of this application. Nothing described herein constitutes prior art to this application, nor should it be construed as a finding of the scope or content of prior art. [Overview of the project]

[0005] This application provides, in particular, linker-drug compounds and methods for producing or preparing said linker-drug compounds.

[0006] In one embodiment, the present invention provides a method for preparing a linker-drug compound represented by formula A. [ka]

[0007] In one embodiment, a method for producing a compound represented by formula A is: The process includes the steps of reacting a compound represented by formula a with a Lewis acid to obtain a crude product, and purifying the crude product to obtain a compound represented by formula A. [ka]

[0008] In some embodiments, the Lewis acid may be a bromide, TMSI (trimethyliodosilane), or TMSOTf (trimethylsilyl trifluoromethanesulfonate). In some embodiments, the Lewis acid is zinc bromide.

[0009] For example, impurities may be generated during the deprotection of compound a under acidic conditions using an acidic deprotecting agent. These impurities include the following: 1) Monodeprotection product impurities [ka] 2) Compound a impurity: [ka] 3) Drug unit impurities: [ka]

[0010] The presence of these impurities can ultimately affect the drug-to-antibody ratio (DAR) of the finally prepared ADC. Therefore, in some embodiments, the content of these impurities in the manufacturing process of the target compound can be controlled. For example, the mono-deprotected product is controlled to be less than 10%, the compound a impurity is controlled to be less than 2%, and the drug unit impurity is controlled to be less than 5%, thereby ensuring the binding efficiency between the target compound and the antibody.

[0011] Through extensive experimental research by the inventors, it has further been found that the impurity level can be reduced by using a Lewis acid (e.g., zinc bromide) as the acidic deprotecting agent compared to using a non-Lewis acid (e.g., trifluoroacetic acid) as the acidic deprotecting agent. In some embodiments, the content of the drug unit impurity can be controlled to 5% or less, or in other embodiments, to 2% or less, thereby meeting and even exceeding the applicable quality control standards. In some embodiments, when zinc bromide is used as the acidic deprotecting agent, the impurity level is significantly reduced compared to when a non-Lewis acid is used, greatly exceeding the quality control standards.

[0012] Furthermore, additional impurities can be generated even in the purification process of the crude product obtained by the above reaction, including the following.

Chemical formula

Chemical formula

Chemical formula

[0013] The presence of these impurities can significantly affect the drug-to-antibody ratio (DAR) of the resulting ADC. Specifically, the succinimide structure in the linker head of the linker-drug, represented by formula A, can undergo a Michael addition reaction with the thiol group of the antibody via a double bond to form an ADC. Ring-opening product impurities may lack this reactive function, potentially resulting in reduced or lost binding ability to the antibody, which can affect the DAR. Furthermore, the formation of drug unit impurities can cause the drug portion to separate from the linker. The separated linker may still have the ability to bind to the antibody, but can no longer deliver the drug payload, which can also affect the DAR.

[0014] Therefore, in some embodiments, impurity levels are controlled during the manufacturing process of the target compound. For example, ring-opening product impurities must be controlled to less than 10%, and drug unit impurities to less than 5%, thereby maintaining effective binding between the target compound and the antibody.

[0015] Through extensive experimental research by the inventors, it has been found that by combining preparative liquid chromatography purification under specific conditions with a nanofiltration step in the above purification process, impurity levels can be significantly reduced. In some embodiments, ring-opening product impurities are controlled to less than 10%, and drug unit impurities to less than 5%, thereby meeting applicable product quality control standards. Furthermore, in some embodiments, impurity levels can be further reduced, for example, to less than 3% for ring-opening product impurities and less than 2% for drug unit impurities, thereby significantly exceeding these standards.

[0016] In some embodiments, the purification method includes the following steps. (2-1) The crude product is purified by preparative liquid chromatography to obtain a preparative solution. (2-2) The preparative solution is concentrated and dried to obtain the compound represented by formula A. In some embodiments, an extraction step may be further included after obtaining the preparative in step (2-1) and before step (2-2).

[0017] In some embodiments, after obtaining the preparative in step (2-1) and before step (2-2), the method further includes the following extraction step. (2-i) Extract the preparative solution with an organic solvent, discard the organic phase, and obtain the aqueous phase.

[0018] In some embodiments, when purification is performed by preparative liquid chromatography in step (2-1), mobile phase A is TFA / water and mobile phase B is TFA / acetonitrile. Also, in some embodiments, the concentration step in step (2-2) includes nanofiltration.

[0019] Those skilled in the art will understand that, after obtaining the aqueous phase in step (2-i), the subsequent step (2-2) may be adjusted accordingly to include a step of concentrating and drying the aqueous phase to obtain the compound represented by formula A.

[0020] In some embodiments, the purification method includes the following steps. (2-1) The crude product is purified using preparative liquid chromatography to obtain a preparative solution. During the purification by preparative liquid chromatography, mobile phase A is TFA / water and mobile phase B is TFA / acetonitrile. (2-2) The preparative solution is concentrated and dried using nanofiltration to obtain the compound represented by formula A.

[0021] In some embodiments, the purification method includes the following steps. (2-1) The crude product is purified using preparative liquid chromatography to obtain a preparative solution. During the purification by preparative liquid chromatography, mobile phase A is TFA / water and mobile phase B is TFA / acetonitrile. (2-i) Extract the preparative solution with an organic solvent, discard the organic phase, and obtain the aqueous phase. (2-2) The aqueous phase is concentrated and dried using nanofiltration to obtain the compound represented by formula A. In some embodiments, when purification is performed by preparative liquid chromatography in step (2-1), mobile phase A is 0.2% TFA / water and mobile phase B is 0.1% TFA / acetonitrile, and in step (2-2), the concentration is performed by nanofiltration.

[0022] In some embodiments, gradient elution is used when purifying by preparative liquid chromatography in step (2-1).

[0023] In some embodiments, the gradient conditions are as follows: [Table 1]

[0024] In some embodiments, the gradient conditions are as follows: [Table 2]

[0025] In some embodiments, the gradient conditions are as follows: [Table 3]

[0026] In some embodiments, when purification is performed by preparative liquid chromatography in step (2-1), the chromatography column is UniSil 10-100 C18 450 mm × 100 mm.

[0027] In some embodiments, when purifying by preparative liquid chromatography in step (2-1), the detection wavelength is approximately 200 nm to approximately 365 nm, including, for example, 200 nm, 220 nm, 250 nm, 270 nm, 300 nm, 350 nm, or 365 nm. In some embodiments, the detection wavelength is approximately 200 nm to 230 nm.

[0028] In some embodiments, when purification is performed by preparative liquid chromatography in step (2-1), the detection wavelength is 214 nm.

[0029] In some embodiments, in step (2-2), nanofiltration is performed under conditions where the temperature is ≤ 30°C.

[0030] In some embodiments, the nanofiltration is performed using a nanofiltration apparatus.

[0031] In some embodiments, in step (2-2), drying includes freeze-drying.

[0032] In some embodiments, in step (2-i), the organic solvent comprises mixed solvent 1 and / or mixed solvent 2, where mixed solvent 1 comprises ethyl acetate and n-hexane, and mixed solvent 2 comprises dichloromethane and n-hexane.

[0033] In some embodiments, step (2-i) includes sequentially extracting the preparative with mixed solvent 1 and mixed solvent 2, discarding the organic phase, and retaining the aqueous phase.

[0034] In some embodiments, the volume ratio of the mixed solvent 1 to the preparative (mixed solvent 1 / preparative) is (1-5):1, for example, 1:1, 2:1, 3:1, 4:1, or 5:1, and in certain embodiments, it is 1:1.

[0035] In some embodiments, the volume ratio of the preparative to the mixed solvent 2 (preparative to mixed solvent 2) is (1-5):1, for example, 1:1, 2:1, 3:1, 4:1, or 5:1, and in certain embodiments, it is 5:2.

[0036] In some embodiments, the volume ratio of ethyl acetate to n-hexane (ethyl acetate / n-hexane) in the mixed solvent 1 is (1 to 5):1, for example, 1:1, 2:1, 3:1, 4:1, or 5:1, and in certain embodiments, it is 3:1.

[0037] In some embodiments, the volume ratio of ethyl acetate to n-hexane in the mixed solvent 1 is (1 to 5):1, for example, 1:1, 2:1, 3:1, 4:1, or 5:1, and in certain embodiments, it is 3:1.

[0038] In some embodiments, the volume ratio of dichloromethane to n-hexane (dichloromethane / n-hexane) in the mixed solvent 2 is (1-5):1, for example, 1:1, 2:1, 3:1, 4:1, or 5:1, and in certain embodiments, it is 3:1.

[0039] In some embodiments, the molar ratio of the Lewis acid to the compound represented by formula a (Lewis acid / compound represented by formula a) is at least 10:1, for example, 10:1, 15:1, 20:1, 25:1, 30:1, 35:1, 40:1, or 45:1, and in certain embodiments, it is 20:1.

[0040] In some embodiments, the molar ratio of the Lewis acid to the compound represented by formula a is (10 to 30):1, for example, 10:1, 15:1, 20:1, 25:1, or 30:1, and in certain embodiments, it is 20:1.

[0041] In some embodiments, the reaction is carried out in the presence of a solvent, which includes nitroalkanes (but not limited to nitromethane and nitroethane), haloalkanes (but not limited to dichloromethane, dichloroethane, and chloroform), water, ketones (but not limited to acetone and butanone), toluenes (but not limited to toluene and trifluorotoluene), esters (but not limited to ethyl acetate and dimethyl carbonate), and acetonitrile. In some embodiments, the solvent includes a mixed solvent obtained by mixing two or more of the above solvents in any proportion.

[0042] In certain embodiments, the solvent is selected from the group consisting of nitromethane, dimethyl carbonate, dichloromethane, mixed solvents of dichloromethane / ketones, mixed solvents of dichloromethane / water, and mixed solvents of dichloromethane / esters.

[0043] In some embodiments, the solvent is selected from the group consisting of nitromethane, dichloromethane / water, and dimethyl carbonate. In this specification, “dichloromethane / water” means a mixture containing dichloromethane and water.

[0044] In some embodiments, the ratio of the mass of the compound represented by formula a to the volume of the solvent is 1 g / (5 to 50 mL), for example, 1 g / 5 mL, 1 g / 10 mL, 1 g / 15 mL, 1 g / 20 mL, 1 g / 25 mL, 1 g / 30 mL, 1 g / 35 mL, 1 g / 40 mL, 1 g / 45 mL, or 1 g / 50 mL, and in certain embodiments, it is 1 g / (5 to 35 mL).

[0045] In some embodiments, the solvent is nitromethane.

[0046] In some embodiments, the ratio of the mass of the compound represented by formula a to the volume of nitromethane is 1 g / (20-50 mL), for example, 1 g / 20 mL, 1 g / 25 mL, 1 g / 30 mL, 1 g / 35 mL, 1 g / 40 mL, 1 g / 45 mL, or 1 g / 50 mL, and in certain embodiments, it is 1 g / 35 mL.

[0047] In some embodiments, the solvent is dichloromethane / water.

[0048] In some embodiments, the ratio of the mass of the compound represented by formula a to the volume of dichloromethane / water is 1 g / (10-30 mL), for example, 1 g / 10 mL, 1 g / 15 mL, 1 g / 16 mL, 1 g / 20 mL, 1 g / 25 mL, or 1 g / 30 mL, and in certain embodiments, 1 g / (15-16 mL).

[0049] In some embodiments, the volume ratio of dichloromethane to water in the dichloromethane / water mixture is (500-1000):1, for example, 500:1, 550:1, 600:1, 650:1, 700:1, 750:1, 800:1, 850:1, 900:1, 950:1, or 1000:1, and in certain embodiments, it is 750:1.

[0050] In some embodiments, the solvent is dimethyl carbonate.

[0051] In some embodiments, the ratio of the mass of the compound represented by formula a to the volume of dimethyl carbonate is 1 g / (5 to 30 mL), for example, 1 g / 5 mL, 1 g / 10 mL, 1 g / 15 mL, 1 g / 20 mL, 1 g / 25 mL, or 1 g / 30 mL, and in certain embodiments, it is 1 g / (5 to 15 mL).

[0052] In some embodiments, the reaction is carried out at a temperature of 20 to 110°C (e.g., 20°C, 30°C, 40°C, 50°C, 60°C, 70°C, 80°C, 90°C, 100°C, or 110°C). In some embodiments, the reaction is carried out at a temperature of 20 to 50°C (e.g., 20°C, 30°C, 35°C, 40°C, 45°C, or 50°C, and 40°C in certain embodiments). In some embodiments, the reaction is carried out at a temperature of 30 to 50°C (e.g., 30°C, 35°C, 40°C, 45°C, or 50°C, and 40°C in certain embodiments).

[0053] In some embodiments, the reaction takes place over a period of time from 20 minutes to 24 hours (for example, 20 minutes, 30 minutes, 40 minutes, 1 hour, 2 hours, 3 hours, 4 hours, 5 hours, 6 hours, 7 hours, 8 hours, 9 hours, 10 hours, 11 hours, 12 hours, 13 hours, 14 hours, 15 hours, 16 hours, 17 hours, 18 hours, 19 hours, 20 hours, 21 hours, 22 hours, 23 hours, or 24 hours). In some embodiments, the reaction takes place over a period of time from 20 to 120 minutes (for example, 20 minutes, 25 minutes, 30 minutes, 35 minutes, 40 minutes, 45 minutes, 50 minutes, 55 minutes, 60 minutes, 70 minutes, 80 minutes, 90 minutes, 100 minutes, 110 minutes, or 120 minutes). In some embodiments, the reaction takes place over a period of time from 20 to 60 minutes. In some embodiments, the reaction takes place over a period of time from 30 to 60 minutes. In some embodiments, the reaction is carried out for 30 minutes.

[0054] In some embodiments, after reacting the compound represented by formula a with a Lewis acid and before purification, the method further includes a post-treatment step.

[0055] In some embodiments, the post-treatment step includes concentrating the resulting reaction product under reduced pressure to obtain a crude product.

[0056] In some embodiments, the post-processing step includes crystallizing the resulting reaction product, filtering it, and drying it to obtain a crude product.

[0057] In some embodiments, the reduced-pressure concentration is carried out at a temperature of 30 to 55°C (for example, 30°C, 35°C, 40°C, 45°C, 50°C, or 55°C, and 45°C in certain embodiments).

[0058] In some embodiments, the compound represented by formula a is produced by a method comprising the following steps (3-1-1) to (3-1-3). (3-1-1) Formula b: [ka] When a compound represented by reacts with a base (e.g., triethylamine), a compound is formed: [ka] To obtain. (3-1-2) Compound: [ka] Compounds: [ka] Reacting with a compound [ka] To obtain. (3-1-3) Compound: [ka] The compound represented by formula a is obtained by reacting it with exatecan mesylate.

[0059] Alternatively, the compound represented by formula a may be produced by a method comprising the following steps (3-2-1) to (3-2-4). (3-2-1) Formula b: [ka] When a compound represented by reacts with a base (e.g., triethylamine), a compound is formed: [ka] To obtain. (3-2-2) Compound: [ka] Reacting with pentafluorophenol and DCC to form a compound: [ka] To obtain. (3-2-3) Compound: [ka] Compounds: [ka] Reacting with a compound [ka] To obtain. (3-2-4) Compound: [ka] The compound represented by formula a is obtained by reacting it with exatecan mesylate.

[0060] In some embodiments, the compound represented by formula b is produced by a method comprising the following steps (4-1) to (4-5). (4-1) Compound: [ka] Reacting with thionyl chloride and benzyl alcohol to form a compound: [ka] To obtain. (4-2) Compound: [ka] Compounds: [ka] Reacting with to form a compound: [ka] To obtain. (4-3) Compound: [ka] The compound is formed by reacting it with (Boc)2O. [ka] To obtain. (4-4) Compound: [ka] Compound obtained by reacting with hydrogen gas in the presence of Pd / C: [ka] To obtain. (4-5) Compound [ka] This is reacted with maleic anhydride to obtain the compound represented by formula b.

[0061] In some embodiments, step (4-1) is carried out by the following steps. (4-1-1) Thionyl chloride is added dropwise to benzyl alcohol, and then the compound: [ka] Add and allow to react. (4-1-2) Isopropyl ether is added to the reactant obtained in step (4-1-1) to induce crystallization, filter, and dry the filtered cake to obtain the compound: [ka] To obtain.

[0062] In some embodiments, step (4-4) is carried out by the following steps. (4-4-1) Compound [ka] It is reacted with hydrogen gas in the presence of Pd / C. (4-4-2) The reaction product obtained in step (4-4-1) is filtered, and the filtrate is concentrated under reduced pressure to obtain the crude product. (4-4-3) The crude product is recrystallized with isopropanol / water to obtain crystals. The crystals are compound: [ka] That is the case. Preferably, during recrystallization, the volume ratio of isopropanol to water (isopropanol / water) is (1-5):1, and more preferably 3:1.

[0063] In some embodiments, steps (4-5) are carried out by the following steps. (4-5-1) Compound: [ka] It is reacted with maleic anhydride. (4-5-2)n-hexane is added to the reactant obtained in step (4-5-1) and crystallized, filtered, and the filtered cake is dried to obtain the compound represented by formula b.

[0064] In another embodiment of the present invention, a method for producing a compound represented by formula A is further provided.

[0065] [ka] In one embodiment, the method includes the following steps. (1-1) The compound represented by formula a is reacted with nitromethane and zinc bromide to obtain a crude product. [ka] (1-2) The crude product is purified by preparative liquid chromatography to obtain the preparative solution. (1-3) Extract the preparative solution with an organic solvent, discard the organic phase, and obtain the aqueous phase. (1-4) The aqueous phase is concentrated and dried to obtain the compound represented by formula A. In step (1-2), when purifying by preparative liquid chromatography, mobile phase A is 0.2% TFA / water and mobile phase B is 0.1% TFA / acetonitrile, and in step (1-4), concentration is performed by nanofiltration.

[0066] In some embodiments, gradient elution is employed when purifying by preparative liquid chromatography in step (1-2).

[0067] In some embodiments, the gradient conditions are as follows: [Table 4]

[0068] In some embodiments, when purification is performed by preparative liquid chromatography in step (1-2), the chromatography column is UniSil 10-100 C18 450 mm × 100 mm.

[0069] In some embodiments, when purification is performed by preparative liquid chromatography in step (1-2), the detection wavelength is 214 nm.

[0070] In some embodiments, in step (1-4), nanofiltration is performed under conditions of a temperature ≤ 30°C.

[0071] In some embodiments, the nanofiltration is performed using a nanofiltration apparatus.

[0072] In some embodiments, in steps (1-4), drying is freeze-drying.

[0073] In some embodiments, in steps (1-3), the organic solvent is mixed solvent 1 and / or mixed solvent 2, where mixed solvent 1 is a mixed solvent consisting of ethyl acetate and n-hexane, and mixed solvent 2 is a mixed solvent consisting of dichloromethane and n-hexane.

[0074] In some embodiments, steps (1-3) are carried out by sequentially extracting the preparative solution with mixed solvent 1 and mixed solvent 2, discarding the organic phase, and obtaining the aqueous phase.

[0075] In some embodiments, the volume ratio of the mixed solvent 1 to the preparative is (1 to 5):1, for example, 1:1, 2:1, 3:1, 4:1, or 5:1, and in certain embodiments, it is 1:1.

[0076] In some embodiments, the volume ratio of the preparative to the mixed solvent 2 is (1 to 5):1, for example, 1:1, 2:1, 3:1, 4:1, or 5:1, and in certain embodiments, it is 5:2.

[0077] In some embodiments, the volume ratio of ethyl acetate to n-hexane in the mixed solvent 1 is (1 to 5):1, for example, 1:1, 2:1, 3:1, 4:1, or 5:1, and in certain embodiments, it is 3:1.

[0078] In some embodiments, the volume ratio of dichloromethane to n-hexane in the mixed solvent 2 is (1 to 5):1, for example, 1:1, 2:1, 3:1, 4:1, or 5:1, and in certain embodiments, it is 3:1.

[0079] In some embodiments, in step (1-1), the molar ratio of zinc bromide to the compound represented by formula a is (10 to 30):1, for example, 10:1, 15:1, 20:1, 25:1, or 30:1, and in certain embodiments, it is 20:1.

[0080] In some embodiments, in step (1-1), the ratio of the mass of the compound represented by formula a to the volume of nitromethane is 1 g / (20-50 mL), for example, 1 g / 20 mL, 1 g / 25 mL, 1 g / 30 mL, 1 g / 35 mL, 1 g / 40 mL, 1 g / 45 mL, or 1 g / 50 mL, and in certain embodiments, it is 1 g / 35 mL.

[0081] In some embodiments, in step (1-1), the reaction is carried out at a temperature of 30 to 50°C (for example, 30°C, 35°C, 40°C, 45°C, or 50°C, and in certain embodiments, 40°C).

[0082] In some embodiments, in step (1-1), the reaction takes place over a period of time of 20 to 60 minutes (for example, 20, 25, 30, 35, 40, 45, 50, 55, or 60 minutes, and 30 minutes in certain embodiments).

[0083] In some embodiments, in step (1-1), the reaction between the compound represented by formula a, nitromethane, and zinc bromide proceeds as follows: (1-1-1) A step of mixing the compound represented by formula a with nitromethane, dissolving it, and then adding zinc bromide and reacting it; (1-1-2) A step in which the reaction product obtained in step (1-1-1) is concentrated under reduced pressure to obtain the crude product.

[0084] In some embodiments, the reduced-pressure concentration is carried out at a temperature of 30 to 55°C (for example, 30°C, 35°C, 40°C, 45°C, 50°C, or 55°C, and 45°C in certain embodiments).

[0085] In some embodiments, the compound represented by formula a is prepared by a method comprising the following (2-1) to (2-4). (2-1) Formula b: [ka] The compound represented by triethylamine is reacted to form the compound: [ka] To obtain. (2-2) Compound: [ka] Reacting with pentafluorophenol and DCC to form a compound: [ka] To obtain. (2-3) Compound: [ka] The compound [ka] Reacting them to form a compound: [ka] To obtain. (2-4) Compound [ka] The compound represented by formula a is obtained by reacting it with exatecan mesylate.

[0086] In some embodiments, the compound represented by formula b is produced by a method comprising the following steps (3-1) to (3-5). (3-1) Compound: [ka] Reacting with thionyl chloride and benzyl alcohol to form a compound: [ka] To obtain. (3-2) Compound: [ka] Compounds: [ka] Reacting with to form a compound: [ka] To obtain. (3-3) Compound: [ka] Reacting with (Boc)2O to form a compound: [ka] To obtain. (3-4) Compound: [ka] Compound obtained by reacting with hydrogen gas in the presence of Pd / C: [ka] To obtain. (3-5) Compound: [ka] This is reacted with maleic anhydride to obtain the compound represented by formula b.

[0087] In some embodiments, step (3-1) is carried out as follows. (3-1-1) Thionyl chloride is added dropwise to benzyl alcohol, and then the compound [ka] Add and allow to react. (3-1-2) Isopropyl ether is added to the reaction product obtained in step (3-1-1) and crystallized, filtered, and the filtered cake is dried to form the compound: [ka] To obtain.

[0088] In some embodiments, steps (3-4) are carried out as follows. (3-4-1) Compound: [ka] It is reacted with hydrogen gas in the presence of Pd / C. (3-4-2) The reaction product obtained in step (3-4-1) is filtered, and the filtrate is concentrated under reduced pressure to obtain the crude product. (3-4-3) The crude product is recrystallized with isopropanol / water to obtain crystals. The crystals are compound: [ka] That is the case.

[0089] In some embodiments, during recrystallization, the volume ratio of isopropanol to water (isopropanol / water) is (1-5):1, for example, 1:1, 2:1, 3:1, 4:1, or 5:1, and in certain embodiments, it is 3:1.

[0090] In some embodiments, steps (3-5) are carried out as follows. (3-5-1) Compound: [ka] It is reacted with maleic anhydride. (3-5-2)n-hexane is added to the reactant obtained in step (3-5-1) and crystallized, filtered, and the filtered cake is dried to obtain the compound represented by formula b.

[0091] The above abstract is intended to provide a concise overview of some embodiments of this application and does not limit the scope of this disclosure. Additional embodiments and features are described in the detailed description and claims. Unless expressly stated otherwise, no features described herein are intended to be essential. The embodiments described herein may be combined in whole or in part, unless they are obviously incompatible. All scope disclosed herein includes the endpoints described herein and encompasses any sub-scopes within that scope. No description of background art constitutes prior art. The scope of this application is defined by the claims. [Brief explanation of the drawing]

[0092] The accompanying drawings are included to provide a further understanding of some embodiments of this application and constitute part of this specification. These drawings illustrate examples of embodiments and, together with the description, illustrate each aspect of this disclosure. These drawings do not limit the scope of the claims. In the drawings, unless otherwise noted, the same reference numerals indicate the same elements. The relative dimensions and proportions of the elements shown in the drawings are not necessarily drawn to actual scale and are provided for illustrative purposes only.

[0093] [Figure 1] The results of impurity detection in Step 1 of Example 1 are shown. [Figure 2] The results of impurity detection in step 1 of Example 2 are shown. [Figure 3] The results of impurity detection in Step 1 of Example 3 are shown. [Figure 4] The results of impurity detection in Step 1 of Comparative Example 1 are shown. [Figure 5] The results of impurity detection in step 2 of Example 1 are shown. [Figure 6] The results of impurity detection in step 2 of Comparative Example 2 are shown. [Figure 7]The results of impurity detection in Step 2 of Comparative Example 3 are shown. [Figure 8] The results of impurity detection in Step 2 of Comparative Example 4 are shown. [Figure 9] The results of impurity detection in Step 2 of Comparative Example 5 are shown. [Figure 10] The results of impurity detection in Step 2 of Comparative Example 6 are shown. [Figure 11] The results of impurity detection in step 2 of Comparative Example 7 are shown. [Figure 12] The results of impurity detection in step 2 of Comparative Example 8 are shown. [Figure 13] The results of impurity detection in Step 2 of Comparative Example 9 are shown. [Modes for carrying out the invention]

[0094] Embodiments of this application will be described below with reference to the accompanying drawings and examples. These drawings and examples are provided for illustrative purposes only and are not intended to limit the scope of the claims. Those skilled in the art will understand that various modifications, variations, and combinations are possible without departing from the scope of this application.

[0095] Further aspects and features of this application will become apparent from the following detailed description.

[0096] In the following experimental steps, all raw materials are commercially available unless otherwise stated.

[0097] Preliminary example 1: Compound: [ka] Manufacturing Synthesis scheme: [ka]

[0098] Step 1: Ac002 2.8 L of benzyl alcohol was added to a 5 L three-necked flask, cooled to below 10°C, and then thionyl chloride (36 mL, 0.50 mol, 2.0 eq) was added dropwise. After the addition was complete, the temperature was raised to room temperature and the reaction was allowed to proceed for 2 hours. Next, 3-amino-N-Cbz-L-alanine (60 g, 0.25 mol, 1.0 eq) was added and the reaction was allowed to proceed at room temperature for 16 hours. After confirming that the starting material (Ac001) had almost completely disappeared by TLC, the reaction was terminated. 12 L of isopropyl ether was added to the system and crystallization was carried out for 2 hours. The system was then filtered, and the resulting filtered cake was air-dried at 45°C to obtain 55.2 g of Ac002. The yield was 60.2%.

[0099] Step 2: Ac003 55 g, 0.15 mol, 1.0 eq of Ac002, 300 mL of acetonitrile, and 33 mL of purified water were added to a 1 L reaction flask and stirred to dissolve. Then, potassium carbonate (31.2 g, 0.225 mol, 1.5 eq) was added, followed by the dropwise addition of tert-butyl bromoacetate (26.42 g, 0.135 mol, 0.9 eq). After the addition was complete, the mixture was reacted at room temperature for 20 hours. After confirming that the starting material (Ac002) had almost completely disappeared by TLC, the reaction was terminated. 1.5 L of purified water was added to the system, and extraction was performed with ethyl acetate (1 L). The layers were separated, the organic layer was washed with 10% aqueous sodium chloride solution, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain the crude product. The crude product was purified by column chromatography (petroleum ether / ethyl acetate) to obtain 44.4 g of Ac003. The yield was 66%.

[0100] Step 3: Ac004 44.2 g, 0.1 mol, 1.0 eq of Ac003 and 100 mL of dichloromethane were added to a 500 mL reaction flask and stirred to dissolve. Then, triethylamine (15 mL, 0.11 mol, 1.1 eq) was added, followed by the dropwise addition of anhydrous Boc (22.9 g, 0.105 mol, 1.05 eq). The reaction was allowed to proceed at room temperature for 20 hours. After confirming that the starting material (Ac003) had almost completely disappeared by TLC, the reaction was terminated. 100 mL of purified water was added to the system and the layers were separated. The aqueous layer was extracted with dichloromethane (40 mL) and the organic layer was combined. The organic layer was washed with purified water, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain 55.3 g of a yellow oily substance, which was then used directly in the next step.

[0101] Step 4: Ac005 The yellow oily substance (55.3 g) obtained in Step 3 and 200 mL of methanol were added to a 1 L necked flask and stirred to dissolve. Next, 5% Pd / C (8.3 g, 15% m / m) was added and the hydrogenation reaction was carried out for 4 hours. After confirming that the starting material (Ac004) had almost disappeared by TLC, the reaction was terminated. The reaction mixture was filtered, and the filtrate was concentrated under reduced pressure to obtain the crude product. The crude product was recrystallized with isopropanol / water (volume ratio 3:1) to obtain 18.4 g of Ac005. The yield was 58% (2 steps).

[0102] Step 5: SM-2 In a 500 mL neck flask, 18 g of Ac005 (0.056 mol, 1.0 eq), 100 mL of dichloromethane, and 17 mL of glacial acetic acid were added and stirred to dissolve. Next, maleic anhydride (6 g, 0.062 mol, 1.1 eq) was added and the mixture was reacted at room temperature for 4 hours. After confirming that the starting material (Ac005) had almost completely disappeared by TLC, the reaction was terminated. 80 mL of n-hexane was added to the system, and the mixture was cooled to 10°C and crystallized for 2 hours. After filtration, the filtered cake was air-dried at 50°C for 4 hours to obtain 20.6 g of SM-2. The yield was 87%.

[0103] Preliminary Example 2 Compound: [ka] Preparation 1. Preparation of compound M3 Synthesis scheme: [ka] Step 1: Compound M2 SM-2 (40 g, 96 mmol, 1.0 eq), triethylamine (26.7 mL, 2.0 eq), and toluene (400 mL) were added to a 1000 mL neck flask. The mixture was reacted under reflux at 120 °C for 2 hours. The reaction was monitored by TLC to confirm that it was nearly complete. The system was cooled to 50 °C, and the solvent was removed under reduced pressure. The residue was dissolved in ethyl acetate (150 mL) and water (40 mL), and the pH was adjusted to 2-3 with 1 M hydrochloric acid while stirring in an ice bath. The layers were separated, and the aqueous layer was further extracted with ethyl acetate. The organic layers were combined, dried over anhydrous sodium sulfate, filtered, and concentrated to obtain a pale yellow oily crude product. The crude product was purified by column chromatography (DCM:MeOH=40:1) to obtain compound M2 (26.6 g); LC-MS:[M+H] + = 399.3.

[0104] Step 2: Compound M3 Compound M2 (26.5 g, 60.5 mmol, 1.0 eq), pentafluorophenol (12.2 g, 66.5 mmol, 1.1 eq), DCC (13.7 g, 66.5 mmol, 1.1 eq), and THF (300 mL) were added to a 1000 mL neck flask. The mixture was reacted at room temperature for 30 minutes (monitored by TLC). After filtering off insoluble matter, the reaction mixture was purified by preparative liquid chromatography. The resulting preparative solution was concentrated under reduced pressure using a water pump in a water bath at 35°C to remove acetonitrile, and then freeze-dried to obtain compound M3 (31.5 g). The yield was 64%. LC-MS:[M+H] + = 565.1.

[0105] 2. Synthesis of compound 5e: Synthesis scheme: Scheme 1: [ka] Scheme 2: [ka] Step 1: Compound 5a In a 25 mL neck flask, M1 (500 mg, 1.4 mmol, 1.0 eq), p-toluenesulfonic acid monohydrate (26 mg, 0.1 mmol, 0.1 eq), and THF (10 mL) were added. After stirring, the mixture was cooled to 0°C, and then L-benzyl lactate (1.2 g, 7.0 mmol, 5 eq) was gradually added. After the addition was complete, the mixture was heated to room temperature and allowed to react. The reaction was monitored by TLC. After the reaction was complete, saturated NaHCO3 aqueous solution was added, the mixture was extracted with ethyl acetate, dried over anhydrous sodium sulfate, filtered, and concentrated. The residue was purified by reverse-phase column chromatography to obtain 5a (400 mg).

[0106] LC-MS:[M+NH4] + =506.2.

[0107] 1 H NMR(400MHz,CDCl3 / CD3OD):1.39(3H,d,J=6.8Hz),3.78(2H,t,J=4.0Hz),4.17-4.27(2H,m),4.42(2H,d,J=4.0Hz),4.72-4.85(2H,m),5.1 1-5.58(2H,m),5.43(1H,s),7.06(1H,t,J=8.0Hz),7.25-7.33(6H,m),7.38(2H,t,J=8.0Hz),7.57(2H,d,J=8.0Hz),7.75(2H,d,J=8.0Hz).

[0108] Step 2: Compound 5b Compound 5a (400 mg, 0.8 mmol, 1.0 eq) and DMF (4 mL) were added to a 25 mL neck flask. After thorough stirring, the mixture was cooled to 0°C, and then DBU (137 mg, 0.9 mmol, 1.1 eq) was gradually added. After the addition was complete, the mixture was heated to room temperature and allowed to react. The reaction was monitored by TLC. After the reaction was complete, the product was designated as reaction solution (1).

[0109] In a separate 25 mL neck flask, M4 (372 mg, 0.9 mmol, 1.1 eq), PyBOP (852 mg, 1.6 mmol, 2.0 eq), and DMF (3 mL) were added and stirred at room temperature for 5 minutes. Then, the reaction mixture (1) was added and the reaction was allowed to proceed at room temperature. The reaction was monitored by HPLC. After the reaction was complete, the reaction mixture was purified by HPLC to obtain compound 5b (326 mg); LC-MS: [M + NH4] + = 679.2.

[0110] Step 3: Compound 5c 5b (4.0 g, 6.05 mmol, 1.0 eq) was added to a 100 mL neck flask and dissolved in DMF (60 mL). Next, 5% Pd / C (4 g) was added and the hydrogenation reaction was carried out at room temperature for 4 hours (the progress of the reaction was monitored by HPLC). After that, the Pd / C was filtered off, and the filtrate was left in an ice bath (approximately 0°C) without concentration to prepare for use in the next step.

[0111] Step 4: Compound 5d Scheme 1: Crude product 5c was placed in an ice bath, DIPEA (1.1 mL, 1.1 eq) was added, followed by compound M3 (3.4 g, 6.05 mmol). After the addition was complete, the temperature was raised to room temperature and the reaction was allowed to proceed for 2 hours. After confirming the completion of the reaction by HPLC, the reaction mixture was purified by HPLC to obtain a preparative. The obtained preparative was freeze-dried to obtain 5d (3.15 g); LC-MS: [MH] - = 816.3.

[0112] Scheme 2: Compound M2 (2.4 g, 6.05 mmol, 1.0 eq) was added to a necked flask and dissolved in acetonitrile (24 mL). The mixture was cooled to -10 ± 5 °C, and EEDQ (1.49 g, 6.05 mmol, 1.0 eq) was added. The mixture was reacted at the same temperature for 2 hours. Then, crude product 5c was added to the system, and the mixture was reacted at the same temperature for a further 1 hour. The reaction was monitored by HPLC to confirm completion. The reaction mixture was purified by high-performance liquid chromatography to obtain a preparative. The preparative was extracted with dichloromethane and concentrated to obtain 5d (3.0 g).

[0113] Step 5: Compound 5e In a 100 mL neck flask, 5d (2.07 g, 2.53 mmol, 1.0 eq), exatecan mesylate M5 (1.35 g, 2.53 mmol, 1.0 eq), PyBOP (1.98 g, 3.79 mmol, 1.5 eq), HOBt (0.51 g, 3.79 mmol, 1.5 eq), and DMF (40 mL) were added. Then, DIPEA (1.05 mL, 1.5 eq) was added under ice water bath conditions, and the mixture was heated to room temperature and reacted for 2 hours (monitored by HPLC). The reaction mixture was purified by preparative liquid chromatography. The resulting preparative solution was concentrated under reduced pressure using a water pump at 35°C in a water bath to remove acetonitrile, and then freeze-dried to obtain compound 5e (1.92 g). The yield was 61%. LC-MS: [M+H] + = 1235.4.

[0114] Example 1 Target compound: [ka] Manufacturing Synthesis scheme: [ka] Step 1: Compound 5e (1.0 g, 0.8 mmol, 1.0 eq) and 35 mL of nitromethane were added to a 100 mL neck flask and dissolved. Then, zinc bromide (3.64 g, 16 mmol, 20.0 eq) was added. The mixture was reacted in an oil bath at 40°C (temperature stabilized by preheating) for 30 minutes (monitored by HPLC). Subsequently, the mixture was concentrated under reduced pressure using a water pump at 45°C in a water bath to remove nitromethane and obtain a yellow solid residue.

[0115] Method for detecting impurities: The target compound was analyzed for impurities by HPLC. The results are shown in Table 1 and Figure 1.

[0116] Equipment: High-performance liquid chromatography (HPLC) Column: YMC-Triart C18-5μm-12nm 4.6mm×250mm Mobile phase A: 0.05% TFA / water (0.5 mL of TFA added to 1 L of purified water) Mobile phase B: 0.05% TFA / acetonitrile (0.5 mL of TFA added to 1 L of acetonitrile) Detection wavelength: 214nm Flow rate: 1mL / min Column temperature: 30℃ Gradient conditions: [Table 5]

[0117] Step 2: The target compound was purified by preparative liquid chromatography under acidic conditions (mobile phase: 0.2% TFA / water - 0.1% TFA / acetonitrile) to obtain a preparative solution (volume V) of the target compound. The organic solvent in the preparative solution was then sequentially extracted using ethyl acetate (0.75V) / n-hexane (0.25V) and dichloromethane (0.3V) / n-hexane (0.1V). The organic phase was discarded to obtain the aqueous phase. Subsequently, the aqueous phase was concentrated at a low temperature (≤30°C) using a nanofiltration apparatus, and then freeze-dried to obtain the target compound.

[0118] LC-MS:[M+H] +=1079.4; 1 H NMR(400MHz,DMSO-d6)δ9.39-9.02(m,1H),8.70(t,J=6.5Hz,1H),8.64(t,J=5.7Hz,1H),8.56(d,J=8.8Hz,1H),8 .34(t,J=5.7Hz,1H),8.16(d,J=8.2Hz,1H),8.01(t,J=5.5Hz,1H),7.71(d,J=10.9Hz,1H),7.30(s,1H),7.28-7. 15(m,4H),7.14(s,2H),5.53(dd,J=14.5,6.4Hz,1H),5.49-5.34(m,2H),5.22(d,J=18.8Hz,1H),5.09(d,J=18.7 Hz,1H),5.03(dd,J=9.6,3.9Hz,1H),4.73(dd,J=9.9,6.9Hz,1H),4.59(dd,J=10.1,6.5Hz,1H),4.49(ddd,J=13.2 ,8.6,4.4Hz,1H),4.14(dd,J=13.3,6.6Hz,2H),3.93(s,2H),3.84(dd,J=16.5,6.3Hz,1H),3.76(dd,J=16.9,5.7 Hz,2H),3.70(d,J=5.2Hz,2H),3.60(dd,J=16.7,5.4Hz,1H),3.52(dd,J=16.4,5.1Hz,1H),3.45(dd,J=12.8,10.1 Hz,1H),3.25-3.15(m,1H),3.14-3.05(m,1H),3.01(dd,J=13.7,4.1Hz,1H),2.73(dd,J=13.5,9.8Hz,1H),2.54- 2.47(m,1H),2.33(s,2H),2.17(d,J=5.5Hz,2H),1.91-1.79(m,2H),1.33(d,J=6.6Hz,2H),0.87(t,J=7.3Hz,2H).

[0119] The specific conditions for dispensing liquid クロマトグラフィーの are as follows: Device: Dispensing liquid クロマトグラフィー カラム:UniSil 10-100 C18 450mm×100mm Mobile phase A: 0.2% TFA / water (TFA 2mL and purified water 1L added) Mobile phase B: 0.1% TFA / acetonitrile (1 mL of TFA added to 1 L of acetonitrile) Detection wavelength: 214nm Gradient conditions: [Table 6]

[0120] Method for detecting impurities: The target compound was subjected to impurity detection by HPLC. The results are shown in Table 2 and Figure 5.

[0121] The measurement conditions for HPLC in impurity detection are as follows: Equipment: High-performance liquid chromatography (HPLC) Column: YMC-Triart C18-5μm-12nm 4.6mm×250mm Mobile phase A: 0.05% TFA / water (0.5 mL of TFA added to 1 L of purified water) Mobile phase B: 0.05% TFA / acetonitrile (0.5 mL of TFA added to 1 L of acetonitrile) Detection wavelength: 214nm Flow rate: 1mL / min Column temperature: 30℃ Gradient conditions: [Table 7]

[0122] Example 2 Synthesis scheme: Compared with Example 1, the difference is only in Step 1, and Step 2 is the same as that in Example 1. Specifically, in Step 1, compound 5e (1.0 g, 0.8 mmol, 1.0 eq) and 15 mL of dichloromethane were added to a 100 mL single-neck flask. After adding 20 μL of water and dissolving, zinc bromide (3.64 g, 16 mmol, 20.0 eq) was added. The mixture was heated and reacted in an oil bath at 40 °C (temperature stabilized by preheating) for 1 hour (monitored by HPLC). Then, it was concentrated under reduced pressure using a water pump in a water bath at 45 °C to remove dichloromethane and obtain a yellow solid residue.

[0123] Impurity detection method for Step 1: The impurity detection method in Step 1 is the same as that in Example 1. The results are shown in Table 1 and Figure 2.

[0124] Example 3 Synthesis scheme: Compared with Example 1, the difference is only in Step 1, and Step 2 is the same as that in Example 1. Specifically, in Step 1, compound 5e (1.0 g, 0.8 mmol, 1.0 eq) and 15 mL of dimethyl carbonate were added to a 100 mL single-neck flask. After dissolving, zinc bromide (3.64 g, 16 mmol, 20.0 eq) was added. The mixture was heated and reacted in an oil bath at 40 °C (temperature stabilized by preheating) for 30 minutes (monitored by HPLC). After the reaction was completed, 20 mL of dichloromethane was added for crystallization. After crystallization, it was filtered and dried to obtain a crude product solid.

[0125] Impurity detection method for Step 1: The impurity detection method in Step 1 is the same as that in Example 1. The results are shown in Table 1 and Figure 3.

[0126] Comparative Example 1 Synthesis scheme: Compared to Example 1, the only difference was in Step 1; Step 2 was identical to that of Example 1. Specifically, in Step 1, compound 5e (1.0 g, 0.8 mmol, 1.0 eq) and 10 mL of dichloromethane were added to a 100 mL necked flask and dissolved. Then, 1 mL of trifluoroacetic acid was added, and the reaction was carried out at room temperature for 30 minutes (monitored by HPLC). After the reaction was complete, the mixture was concentrated under reduced pressure to obtain the crude product.

[0127] Step 1 Impurity Detection Method: The method for detecting impurities in Step 1 is the same as in Example 1. The results are shown in Table 1 and Figure 4.

[0128] Table 1: Results of impurity detection [Table 8]

[0129] A comparison of the results of Examples 1-3 and Comparative Example 1 clearly shows that, in the manufacturing process of the target compound, only the manufacturing method for which the present application seeks protection can meet the product quality standards (mono-deprotection products must be controlled to less than 10%, compound a must be controlled to less than 2%, and drug units must be controlled to less than 5%), and furthermore, significantly exceed these quality standards (drug units can be reduced to less than 2%).

[0130] Comparative Example 2 Synthesis scheme: Compared to Example 1, Step 1 is identical to that of Example 1, with the only difference being Step 2. Specifically, in Step 2, the target compound was purified by preparative liquid chromatography (mobile phase: pure water-acetonitrile) to obtain a preparative solution. The specific conditions for preparative liquid chromatography were the same as in Example 1, except that mobile phase A was replaced with pure water and mobile phase B was replaced with acetonitrile.

[0131] Step 2: Method for detecting impurities: The method for detecting impurities in Step 2 is the same as in Example 1. The results are shown in Table 2 and Figure 6.

[0132] Comparative Example 3 Synthesis scheme:

[0133] Compared to Example 1, Step 1 was identical to that of Example 1, with the only difference being Step 2. Specifically, in Step 2, the target compound was purified by preparative liquid chromatography under acidic conditions (mobile phase: 0.1% TFA / water-acetonitrile) to obtain a preparative solution (volume V) of the target compound. Next, the organic solvent in the preparative solution was sequentially extracted using ethyl acetate (0.75V) / n-hexane (0.25V) and dichloromethane (0.3V) / n-hexane (0.1V). The organic phase was discarded to obtain the aqueous phase. Subsequently, the aqueous phase was concentrated at a low temperature (≤30°C) using a nanofiltration apparatus, and then freeze-dried to obtain the target compound. The specific conditions for preparative liquid chromatography were the same as in Example 1, except that mobile phase A was replaced with 0.1% TFA / water and mobile phase B was replaced with acetonitrile.

[0134] Step 2: Method for detecting impurities: The method for detecting impurities in Step 2 is the same as in Example 1. The results are shown in Table 2 and Figure 7.

[0135] Comparative Example 4 Synthesis scheme: Compared to Example 1, Step 1 was identical to that of Example 1, with the only difference being Step 2. Specifically, in Step 2, the target compound was purified by preparative liquid chromatography under acidic conditions (mobile phase: 0.1% TFA / water-acetonitrile) to obtain a preparative solution (volume V) of the target compound. Next, the organic solvent in the preparative solution was sequentially extracted using ethyl acetate (0.75V) / n-hexane (0.25V) and dichloromethane (0.3V) / n-hexane (0.1V). The organic phase was discarded to obtain the aqueous phase. Subsequently, the aqueous phase was concentrated under reduced pressure in a water bath at 35°C using an oil pump, and then freeze-dried to obtain the target compound. The specific conditions for preparative liquid chromatography were the same as in Example 1, except that mobile phase A was replaced with 0.1% TFA / water and mobile phase B was replaced with acetonitrile.

[0136] Step 2: Method for detecting impurities: The method for detecting impurities in Step 2 is the same as in Example 1. The results are shown in Table 2 and Figure 8.

[0137] Comparative Example 5 Synthesis scheme: Compared to Example 1, Step 1 was identical to that of Example 1, with the only difference being Step 2. Specifically, in Step 2, the target compound was purified by preparative liquid chromatography under acidic conditions (mobile phase: 0.1% TFA / water - 0.1% TFA / acetonitrile) to obtain a preparative solution (volume V) of the target compound. Next, the organic solvent in the preparative solution was sequentially extracted using ethyl acetate (0.75V) / n-hexane (0.25V) and dichloromethane (0.3V) / n-hexane (0.1V). The organic phase was discarded to obtain the aqueous phase. Subsequently, the aqueous phase was concentrated at a low temperature (≤30°C) using a nanofiltration apparatus, and then freeze-dried to obtain the target compound. The specific conditions for preparative liquid chromatography were the same as in Example 1, except that mobile phase A was replaced with 0.1% TFA / water.

[0138] Step 2: Method for detecting impurities: The method for detecting impurities in Step 2 is the same as in Example 1. The results are shown in Table 2 and Figure 9.

[0139] Comparative Example 6 Synthesis scheme: Compared with Example 1, Step 1 is the same as that in Example 1, and the difference is only in Step 2. Specifically, in Step 2, after purification by preparative liquid chromatography under acidic conditions (mobile phase: 0.1% TFA / water - 0.1% TFA / acetonitrile), a fractionation solution (volume V) of the target compound was obtained. Next, the organic solvent in the fractionation solution was sequentially extracted using ethyl acetate (0.75V) / n - hexane (0.25V) and dichloromethane (0.3V) / n - hexane (0.1V). The organic phase was discarded to obtain an aqueous phase. Subsequently, the aqueous phase was concentrated under reduced pressure at 35 °C in a water bath using an oil pump, and then freeze - dried to obtain the target compound. Regarding the specific conditions of preparative liquid chromatography, except that mobile phase A was replaced with 0.1% TFA / water, other conditions were the same as those in Example 1.

[0140] Impurity detection method for Step 2: The impurity detection method in Step 2 is the same as that in Example 1. The results are shown in Table 2 and Figure 10.

[0141] Comparative Example 7 Synthesis scheme: Compared with Example 1, Step 1 is the same as that in Example 1, and the difference is only in Step 2. Specifically, in Step 2, after purification by preparative liquid chromatography under acidic conditions (mobile phase: 0.2% TFA / water - 0.1% TFA / acetonitrile), a fractionation solution (volume V) of the target compound was obtained. Next, the organic solvent in the fractionation solution was sequentially extracted using ethyl acetate (0.75V) / n - hexane (0.25V) and dichloromethane (0.3V) / n - hexane (0.1V). The organic phase was discarded to obtain an aqueous phase. Subsequently, the aqueous phase was concentrated under reduced pressure at 35 °C in a water bath using an oil pump, and then freeze - dried to obtain the target compound. The specific conditions of preparative liquid chromatography are the same as those in Example 1.

[0142] Impurity detection method for Step 2: The method for detecting impurities in Step 2 is the same as in Example 1. The results are shown in Table 2 and Figure 11.

[0143] Comparative Example 8 Synthesis scheme: Compared to Example 1, Step 1 was identical to that of Example 1, with the only difference being Step 2. Specifically, in Step 2, the target compound was purified by preparative liquid chromatography under acidic conditions (mobile phase: 0.2% TFA / water - 0.2% TFA / acetonitrile) to obtain a preparative solution (volume V) of the target compound. Next, the organic solvent in the preparative solution was sequentially extracted using ethyl acetate (0.75V) / n-hexane (0.25V) and dichloromethane (0.3V) / n-hexane (0.1V). The organic phase was discarded to obtain the aqueous phase. Subsequently, the aqueous phase was concentrated at a low temperature (≤30°C) using a nanofiltration apparatus, and then freeze-dried to obtain the target compound. The specific conditions for preparative liquid chromatography were the same as in Example 1, except that mobile phase B was replaced with 0.2% TFA / acetonitrile.

[0144] Step 2: Method for detecting impurities: The method for detecting impurities in Step 2 is the same as in Example 1. The results are shown in Table 2 and Figure 12.

[0145] Comparative Example 9 Synthesis scheme: Compared to Example 1, Step 1 was identical to that of Example 1, with the only difference being Step 2. Specifically, in Step 2, the target compound was purified by preparative liquid chromatography under acidic conditions (mobile phase: 0.2% TFA / water - 0.2% TFA / acetonitrile) to obtain a preparative solution (volume V) of the target compound. Next, the organic solvent in the preparative solution was sequentially extracted using ethyl acetate (0.75V) / n-hexane (0.25V) and dichloromethane (0.3V) / n-hexane (0.1V). The organic phase was discarded to obtain the aqueous phase. Subsequently, the aqueous phase was concentrated under reduced pressure in a water bath at 35°C using an oil pump, and then freeze-dried to obtain the target compound. The specific conditions for preparative liquid chromatography were the same as in Example 1, except that mobile phase B was replaced with 0.2% TFA / acetonitrile.

[0146] Step 2: Method for detecting impurities: The method for detecting impurities in Step 2 is the same as in Example 1. The results are shown in Table 2 and Figure 13.

[0147] Table 2: Results of impurity detection [Table 9]

[0148] A comparison of the results of Example 1 and Comparative Examples 2-9 clearly shows that, in the manufacturing process of the target compound, only the manufacturing method for which this application seeks protection can meet the product quality standards (ring-opening products must be controlled to less than 10%, and drug units must be controlled to less than 5%), and furthermore, significantly exceed these quality standards (ring-opening products can be reduced to 3% or less, and drug units to 2% or less).

[0149] This application is not limited to the specific methods, experimental procedures, or reagents described herein, which may be modified as appropriate. The descriptions and examples herein are provided for illustrative purposes only and do not limit the scope of this application. The scope of this application is defined by the claims.

Claims

1. A method for producing a compound represented by formula A, 【Chemistry 1】 The steps include: reacting a compound represented by formula a with a Lewis acid to obtain a crude product; The steps include: purifying the crude product to obtain a compound represented by formula A; Includes, 【Chemistry 2】 Preferably, the Lewis acid is a bromide, TMSI, or TMSOTf. A method for producing a Lewis acid, wherein the Lewis acid is zinc bromide.

2. The aforementioned purification process The crude product is purified by preparative liquid chromatography to obtain a preparative solution (2-1), (2-2) The steps are to concentrate and dry the aforementioned preparative solution to obtain a compound represented by formula A, Includes, Optionally, the manufacturing method further includes an extraction step after obtaining the preparative in step (2-1) and before step (2-2), Optionally, after obtaining the preparative in step (2-1) and before step (2-2), The process further includes the step (2-i) of extracting the aforementioned preparative solution with an organic solvent and discarding the organic phase to obtain an aqueous phase, In step (2-1), when purifying by preparative liquid chromatography, mobile phase A contains TFA / water and mobile phase B contains TFA / acetonitrile. The manufacturing method according to claim 1, wherein in step (2-2), the concentration includes nanofiltration concentration.

3. The manufacturing method further includes one or more technical features selected from the group consisting of (i) to (iii) below, (i) In step (2-1), when purifying by preparative liquid chromatography, gradient elution is used, and the gradient conditions are: And, (ii) In step (2-1), when purifying by preparative liquid chromatography, the detection wavelength is 200-365 nm. (iii) In step (2-2), nanofiltration is carried out under conditions of temperature ≤ 30°C. Preferably, the nanofiltration is performed using a nanofiltration apparatus, as described in claim 2.

4. The manufacturing method further includes one or more technical features selected from the group consisting of (i) to (ii) below, (i) In step (2-2), drying is freeze-drying, (ii) In step (2-i), the organic solvent is mixed solvent 1 and / or mixed solvent 2, where mixed solvent 1 comprises ethyl acetate and n-hexane, and mixed solvent 2 comprises dichloromethane and n-hexane. Preferably, in step (2-i), the preparative is sequentially extracted with the mixed solvent 1 and the mixed solvent 2, the organic phase is discarded, and the aqueous phase is obtained. Preferably, the volume ratio of the mixed solvent 1 to the preparative is (1-5):1, and more preferably 1:

1. Preferably, the volume ratio of the preparative solution to the mixed solvent 2 is (1-5):1, and more preferably 5:

2. Preferably, in the mixed solvent 1, the volume ratio of ethyl acetate to n-hexane is (1-5):1, more preferably 3:

1. Preferably, in the mixed solvent 2, the volume ratio of dichloromethane to n-hexane is (1-5):1, and more preferably 3:1, the manufacturing method according to claim 2 or 3.

5. The manufacturing method further includes one or more technical features selected from the group consisting of (i) to (iv) below, (i) The molar ratio of the Lewis acid to the compound represented by formula a is 10:1 or greater. (ii) The reaction is carried out in the presence of a solvent, the solvent being selected from the group consisting of nitroalkanes (e.g., nitromethane, nitroethane), haloalkanes (e.g., dichloromethane, dichloroethane, chloroform), water, ketones (e.g., acetone, butanone, etc.), toluenes (e.g., toluene, trifluorotoluene), esters (e.g., ethyl acetate, dimethyl carbonate), acetonitrile, and mixed solvents obtained by mixing two or more of these solvents in any proportion, preferably selected from the group consisting of nitromethane, dimethyl carbonate, dichloromethane, a mixture of dichloromethane / ketones, a mixture of dichloromethane / water, and a mixture of dichloromethane / esters. (iii) The above reaction is carried out at a temperature of 20 to 110°C. (iv) The manufacturing method according to any one of claims 1 to 4, wherein the reaction is carried out over a period of 20 minutes to 24 hours.

6. After reacting the compound represented by formula a with a Lewis acid and before purification, the production method includes a post-treatment step, Preferably, the post-processing is (i) A step of concentrating the obtained reactants under reduced pressure to obtain the crude product, or (ii) The step of crystallizing the reaction product obtained, filtering it, and drying it to obtain the crude product. Includes, Preferably, the reduced-pressure concentration is carried out at a temperature of 30-55°C, according to the manufacturing method according to any one of claims 1 to 5.

7. The compound represented by formula a is Formula b: 【Transformation 3】 The compound represented by reacts with a base to form a compound: 【Chemistry 4】 Steps to obtain (3-1-1), Compound: 【Transformation 5】 Compound: 【Transformation 6】 Reacting with it to form a compound: 【Transformation 7】 Steps to obtain (3-1-2), Compound: 【Transformation 8】 The steps are: reacting with exatecan mesylate to obtain a compound represented by formula a (3-1-3), Manufactured by a method including, Alternatively, the compound represented by formula a is Formula b: 【Chemistry 9】 The compound represented by reacts with a base to form a compound: 【Chemistry 10】 Steps to obtain (3-2-1), Compound: 【Chemistry 11】 The compound obtained by reacting it with pentafluorophenol and DCC is: 【Chemistry 12】 Steps to obtain (3-2-2), Compound: 【Chemistry 13】 Compound: 【Chemistry 14】 Reacting with it to form a compound: 【Chemistry 15】 The steps to obtain (3-2-3), Compound: 【Chemistry 16】 The steps are: reacting with exatecan mesylate to obtain a compound represented by formula a (3-2-4), A manufacturing method according to any one of claims 1 to 6, manufactured by a method including the following:

8. The compound represented by formula b is Compound: 【Chemistry 17】 The compound is formed by reacting it with thionyl chloride and benzyl alcohol. [Chemistry 18] Steps to obtain (4-1), Compound: 【Chemistry 19】 Compound: 【Chemistry 20】 Reacting with it to form a compound: 【Chemistry 21】 Steps to obtain (4-2), Compound: 【Chemistry 22】 (Boc) ２ Compound formed by reacting with O: 【Chemistry 23】 Steps to obtain (4-3), compound 【Chemistry 24】 The compound obtained by reacting Pd / C with hydrogen gas: 【Chemistry 25】 Steps to obtain (4-4), Compound: 【Chemistry 26】 The steps are: reacting with maleic anhydride to obtain a compound represented by formula b (4-5), The manufacturing method according to claim 7, manufactured by a method including the following.

9. Step (4-1) is, Thionyl chloride is added dropwise to benzyl alcohol, and then the compound: 【Chemistry 27】 The step of adding and reacting (4-1-1), The reaction product obtained in step (4-1-1) was crystallized by adding isopropyl ether, filtered, and the filtered cake was dried to obtain the compound: 【Chemistry 28】 Steps to obtain (4-1-2), The manufacturing method according to claim 8, which is carried out by a step including the following:

10. Step (4-4) is, Compound: 【Chemistry 29】 The step of reacting with hydrogen gas in the presence of Pd / C (4-4-1), Step (4-4-2) involves filtering the reaction product obtained in step (4-4-1) and concentrating the filtrate under reduced pressure to obtain the crude product, The crude product is recrystallized with isopropanol / water to obtain crystals (4-4-3), This is carried out by steps including, The aforementioned crystal is a compound: 【Transformation 30】 The manufacturing method according to claim 8, wherein, preferably, when recrystallizing, the volume ratio of isopropanol to water is (1-5):1, and more preferably 3:

1.

11. Steps (4-5) are: Compound: 【Chemistry 31】 The step of reacting with maleic anhydride (4-5-1), Step (4-5-2) involves adding n-hexane to the reactant obtained in step (4-5-1) to crystallize it, filtering it, and drying the filtered cake to obtain the compound represented by formula b, The manufacturing method according to claim 8, which is carried out by a step including the following:

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