An intermediate of PNU-159682 and a process for its preparation

Abstract

The application discloses a preparation method of PNU-159682 and an intermediate of PNU-159682 and a preparation method of the intermediate. Specifically, the application provides a preparation method of compound 7, a preparation method of compound 5, compound 3 and a preparation method of the compound 3 and compound 4. The preparation method has the advantages of low cost, high yield, mild reaction condition and suitability for industrial large-scale production.

Description

An intermediate of PNU-159682 and its preparation method

[0001] This application claims priority to Chinese Patent Application No. 2023114246133, filed on October 30, 2023, and Chinese Patent Application No. 2023114206297, filed on October 30, 2023. This application incorporates the entirety of the aforementioned Chinese patent applications. Technical Field

[0002] The invention belongs to the field of organic synthesis, and particularly relates to an intermediate of PNU-159682 and a preparation method thereof. Background Art

[0003] Antibody drug conjugates (ADCs) are a new type of highly effective biopharmaceutical that connects an antibody to a biologically active small molecule cytotoxic payload via a linker. Primarily used for the treatment of cancer and tumors, they have become a hot topic of development, with multiple ADCs in various stages of clinical development. The principle is to link a cytotoxin to an antibody. The antibody then recognizes specific antigens on the cancer cell surface, allowing the cytotoxin to enter the cancer cell via endocytosis, thereby delivering the cytotoxin to the target site and achieving targeted treatment of malignant tumors. Based on their mechanism of action, toxins suitable for ADCs are categorized as microtubule inhibitors, DNA-damaging agents, and RNA polymerase inhibitors.

[0004] Among them, PNU-159682 (CAS: 202350-68-3, structural formula below) is a metabolite of the anthracycline neomycin and a DNA topoisomerase II inhibitor with excellent cytotoxicity. In ADC synthesis, PNU-159682 is a more potent and more tolerable ADC cytotoxin than doxorubicin and can be used in EDV nanocell technology to overcome drug resistance.

[0005] The synthesis of PNU-159682 requires the use of the key intermediate compound 5 (structural formula shown below). However, the prior art only provides the preparation method of the hydrochloride salt of compound 5, which mostly uses doxorubicin hydrochloride as the starting material.

[0006] Patent BE904431A discloses the following method, the synthesis route of which is shown below. This patented method requires doxorubicin hydrochloride as a raw material, which is very expensive. Furthermore, this method has complex post-processing operations and requires column purification, which not only results in large losses, low yield, and poor atom economy, but also has a two-step column purification yield of 55%. Furthermore, the patent states that the reaction is quenched after 15 minutes, which is relatively short. Furthermore, this reaction generates localized heat, making it very uncontrollable for scaled-up production. Furthermore, as the feed amount increases, a large amount of insoluble matter is produced, which is not conducive to industrial scale-up production.

[0007] Patent EP0434960A1 discloses the following method, the synthesis route of which is shown below. This patented method also uses expensive doxorubicin hydrochloride as a raw material, which is very costly. The patent discloses that a large amount of iodide is required, 15 equivalents, and the reaction time is long, requiring 36 hours. The post-processing operation is complex, requiring column purification, and the overall yield is low at only 45%. The atom economy is poor, the cycle is long, and it is not suitable for industrial scale-up.

[0008] Patent CN115043895A discloses the following method for preparing Compound Ic, which has a similar structure to Compound 5. This method uses doxorubicin hydrochloride as a starting material, undergoing carbonyl protection and reductive amination. This method is actually carried out in two steps, resulting in a low yield of only 45%. Furthermore, this method, which uses doxorubicin hydrochloride as a starting material, is costly and therefore unsuitable for scale-up production.

[0009] In summary, the related preparation methods in the prior art are complex and expensive, which is very unfavorable for the large-scale production of the key intermediate compound 5. Therefore, designing and implementing a synthetic method suitable for industrial production with mild conditions, simple operation and high yield has become the focus of research and development for those skilled in the art.

[0010] Summary of the Invention

[0011] The present invention addresses the low yield, high cost, and unsuitability for industrial production of key intermediates of PNU-159682 in the prior art. To address this issue, the present invention provides an intermediate of PNU-159682 and a method for its preparation. The preparation method of the present invention offers the advantages of low cost, high yield, mild reaction conditions, and suitability for large-scale industrial production.

[0012] The present invention provides a method for preparing compound 5, which comprises the following steps S2, S3 and S4:

[0013] Step S2: In a solvent, in the presence of a base and a reducing agent, compound A-2 and / or its salt is subjected to a reductive amination reaction with compound 2s as shown in the following formula to obtain compound 3,

[0014] Step S3: Compound 3 is subjected to the following reaction in a solvent under the action of hydrogen bromide to obtain compound 4:

[0015] Among them, each R 1 is methyl, ethyl, n-propyl, isopropyl, n-butyl or n-pentyl;

[0016] In step S3, the reaction temperature is 30-50°C;

[0017] In step S3, the molar ratio of hydrogen bromide to compound 3 is (2-4):1;

[0018] Step S4: In a solvent, compound 4 is subjected to a hydrolysis reaction as shown in the following formula under the action of a base to obtain compound 5.

[0019] In a preferred embodiment, certain conditions and operations are defined as follows, and the definitions of conditions and operations not mentioned are as described in any embodiment of the present invention (this paragraph is hereinafter referred to as "in a preferred embodiment").

[0020] In a preferred embodiment, each R 1 It is a methyl group.

[0021] In a preferred embodiment, the salt of compound A-2 is selected from One or more of

[0022] In a preferred embodiment, step S2 comprises the following steps: in a solvent, in the presence of a base and a reducing agent, subjecting compound 2 and compound 2s to a reductive amination reaction as shown in the following formula to obtain compound 3:

[0023] In a preferred embodiment, in step S2, the solvent is an ether solvent, which can be selected from one or more of ethylene glycol dimethyl ether, 1,4-dioxane and tetrahydrofuran, such as tetrahydrofuran.

[0024] In a preferred embodiment, in step S2, the molar volume ratio of the "compound A-2 and / or its salt" to the solvent is (50-150) mmol:1L, for example (80-100) mmol:1L, and another example is 91 mmol:1L.

[0025] In a preferred embodiment, in step S2, the base is an inorganic base; for example, the inorganic base is selected from one or more of sodium bicarbonate, potassium bicarbonate, sodium carbonate, potassium carbonate and calcium carbonate, tripotassium phosphate, trisodium phosphate, dipotassium hydrogen phosphate and disodium hydrogen phosphate; another example is sodium bicarbonate.

[0026] In a preferred embodiment, in step S2, the molar ratio of the base to the "compound A-2 and / or its salt" is (1-3):1, for example (2.5-3.5):1, and another example is 3:1.

[0027] In a preferred embodiment, in step S2, the reducing agent is selected from one or more of sodium borohydride, potassium borohydride, lithium borohydride, sodium cyanoborohydride and lithium aluminum hydride, such as sodium cyanoborohydride.

[0028] In a preferred embodiment, in step S2, the molar ratio of the reducing agent to the "compound A-2 and / or its salt" is (1-5):1, for example (1-3):1, and another example is 1.5:1.

[0029] In a preferred embodiment, in step S2, the molar ratio of the compound 2s to the "compound A-2 and / or its salt" is (1-5):1, for example (2-4):1, and another example is 3:1.

[0030] In a preferred embodiment, in step S2, the temperature of the reductive amination reaction is 10-25°C, for example 25°C.

[0031] In a preferred embodiment, in step S2, the reductive amination reaction is carried out under light-shielding conditions.

[0032] In a preferred embodiment, the preparation method of compound 3 comprises the following steps: adding compound A-2 and / or its salt to a solvent, cooling (for example, to 0°C), adding a base, stirring for 3 to 8 minutes, adding 2-[(1S)-1-methoxy-2-oxoethoxy]acetaldehyde and a reducing agent, stirring at 10 to 25°C, and after the reaction is completed, post-processing to obtain the product.

[0033] In step S2, the progress of the reaction can be monitored using conventional monitoring methods for such reactions in the art (e.g., HPLC, TLC, or LC-MS), with the disappearance or non-reaction of "Compound A-2 and / or its salt" being the endpoint of the reaction. The reductive amination reaction can be performed for 10 to 20 hours, e.g., 15 to 20 hours, or even 16 hours.

[0034] In a preferred embodiment, in step S2, after the reductive amination reaction is completed, the following post-processing steps are further included. The post-processing steps are conventional post-processing steps for reactions of this type in the art, for example, including one or more of filtration, extraction, washing, drying, concentration, and purification. The extraction solvent may be methyltetrahydrofuran. The washing solvent may be water. The purification may be performed by column chromatography, and the eluents for the column chromatography may be dichloromethane and methanol, with the volume ratio of dichloromethane to methanol being 80:1.

[0035] In a preferred embodiment, in step S3, the solvent is a ketone organic solvent, such as acetone.

[0036] In a preferred embodiment, in step S3, the molar volume ratio of the compound 3 to the solvent is (50-150) mmol:1L, for example (50-133) mmol:1L, and another example is 66 mmol:1L.

[0037] In a preferred embodiment, in step S3, the hydrogen bromide is added in the form of an aqueous hydrogen bromide solution; for example, an aqueous hydrogen bromide solution, wherein the molar concentration of hydrogen bromide in the aqueous hydrogen bromide solution may be 0.15 to 0.3 mol / L, for example, 0.15 mol / L.

[0038] In a preferred embodiment, in step S3, the molar ratio of hydrogen bromide to compound 3 is 2.3:1 or 3.4:1.

[0039] In a preferred embodiment, in step S3, the reaction temperature is 35-45°C, for example 40°C.

[0040] In a preferred embodiment, in step S3, the reaction is carried out under light-proof conditions.

[0041] In a preferred embodiment, in step S3, the reaction is carried out under an inert atmosphere, such as a nitrogen atmosphere.

[0042] In a preferred embodiment, the preparation method of compound 4 comprises the following steps: adding a solvent to compound 3, stirring to dissolve, adding hydrogen bromide, reacting at 30-40°C for 4-6 hours, and directly proceeding to the next step of reaction after TLC detection shows complete conversion without post-treatment.

[0043] In step S3, the progress of the reaction can be monitored using conventional monitoring methods for such reactions in the art (e.g., HPLC, TLC, or LC-MS), with the disappearance or no further reaction of compound 3 being considered the endpoint of the reaction. The reaction time can be 4 to 6 hours, for example, 4 hours.

[0044] In a preferred embodiment, in step S3, after the reaction is completed, the next reaction is directly carried out without any post-treatment.

[0045] In a preferred embodiment, when the reaction in step S3 is directly carried out to the next step without any post-treatment, the amounts of the base and the solvent used in step S4 are calculated based on the yield of step S3 being 100%.

[0046] In a preferred embodiment, the preparation method of compound 5 comprises the following steps: adding a solvent to compound 3, stirring to dissolve, adding an acid, reacting at 35-45°C for 4-6 hours, and after TLC detection of complete conversion, adding a base to the reaction system, stirring at 35-45°C for 18-22 hours. After the reaction is completed, diluting, extracting, concentrating, beating, filtering, and drying to obtain the product.

[0047] In a preferred embodiment, in step S4, the solvent is a ketone organic solvent, such as acetone.

[0048] In a preferred embodiment, in step S4, the molar volume ratio of the compound 4 to the solvent is (50-150) mmol:1L, for example (50-133) mmol:1L, and another example is 66 mmol:1L.

[0049] In a preferred embodiment, in step S4, the base is selected from one or both of sodium formate and sodium acetate; for example, sodium formate.

[0050] In a preferred embodiment, in step S4, the molar ratio of the base to the compound 4 is (3.5-13.5):1, for example (4-8.5):1, for example (5-8.5):1, and more for example 6:1.

[0051] In a preferred embodiment, in step S4, the temperature of the hydrolysis reaction is 30-50°C, such as 35-45°C, such as 40°C.

[0052] In a preferred embodiment, in step S4, the hydrolysis reaction is carried out under light-proof conditions.

[0053] In a preferred embodiment, in step S4, the hydrolysis reaction is carried out under an inert atmosphere, such as a nitrogen atmosphere.

[0054] In a preferred embodiment, the preparation method of compound 5 comprises the following steps: adding a base to compound 4, stirring at 30-50°C for 18-24 hours, and after the reaction is completed, diluting, extracting, concentrating, beating, filtering, and drying to obtain the compound 5.

[0055] In step S4, the progress of the reaction can be monitored using conventional monitoring methods for such reactions in the art (e.g., HPLC, TLC, or LC-MS), with the disappearance or no further reaction of compound 4 being considered the endpoint of the reaction. The hydrolysis reaction time can be 15 to 25 hours, e.g., 18 to 22 hours, and further e.g., 20 hours.

[0056] In a preferred embodiment, in step S4, after the hydrolysis reaction is completed, the following post-processing steps are further included. The post-processing steps are conventional post-processing steps for reactions of this type in the art, for example, including one or more of dilution, extraction, drying, concentration, pulping, and filtration. The dilution solvent may be water. The extraction solvent may be dichloromethane. The pulping solvent may be methyl tert-butyl ether.

[0057] In a preferred embodiment, the preparation method of compound 5 further includes a preparation method of compound 2, which comprises the following steps S1: in a solvent, in the presence of a catalyst and an additive, compound 1, Br2 and a reaction reagent are reacted as shown in the following formula to obtain compound 2,

[0058] The reaction reagent is R 1 OH; each R 1 is methyl, ethyl, n-propyl, isopropyl, n-butyl or n-pentyl.

[0059] In a preferred embodiment, in step S1, the solvent is an ether solvent; preferably, the ether solvent is ethylene glycol dimethyl ether, 1,4-dioxane or tetrahydrofuran, such as 1,4-dioxane.

[0060] In a preferred embodiment, in step S1, the reaction reagent is methanol, ethanol, 1-propanol, 1-butanol or 1-pentanol; for example, methanol.

[0061] In a preferred embodiment, in step S1, the volume ratio of the reaction reagent to the solvent is 1:(0.5-1.5), for example 1:(0.5-1), and another example is 1:1.

[0062] In a preferred embodiment, in step S1, the molar volume ratio of the compound 1 to the solvent is (100-300) mmol:1L, for example (200-260) mmol:1L, and another example is 236 mmol:1L.

[0063] In a preferred embodiment, in step S1, the additive is trimethyl orthoformate, triethyl orthoformate or triisopropyl orthoformate, for example, trimethyl orthoformate.

[0064] In a preferred embodiment, in step S1, the molar ratio of the additive to the compound 1 is (17-20):1, for example (18-19):1, and another example is 18.5:1.

[0065] In a preferred embodiment, in step S1, the catalyst is a sulfonic acid, such as p-toluenesulfonic acid, benzenesulfonic acid, 4-ethylbenzenesulfonic acid, 4-chlorobenzenesulfonic acid, xylenesulfonic acid, 2,3-dimethylbenzenesulfonic acid, 2,4-dimethylbenzenesulfonic acid, 2,5-dimethylbenzenesulfonic acid, 2,6-dimethylbenzenesulfonic acid or methanesulfonic acid, and p-toluenesulfonic acid is another example.

[0066] In a preferred embodiment, in step S1, the molar ratio of the catalyst to the compound 1 is (0.1-0.3):1, for example, 0.1:1.

[0067] In a preferred embodiment, in step S1, the Br2 is added in the form of a dichloromethane solution of Br2, and the volume ratio of dichloromethane to Br2 is (1-20):1, for example (6-18):1, and another example is 16.9:1.

[0068] In a preferred embodiment, in step S1, the molar ratio of Br2 to compound 1 is (1-3):1, for example (1-2):1, and another example is 1.3:1.

[0069] In a preferred embodiment, in step S1, the Br2 is added dropwise.

[0070] In a preferred embodiment, step S1 includes the following steps:

[0071] (I) mixing compound 1, a catalyst, a reaction reagent and a solvent to obtain a mixed solution;

[0072] (II) mixing the mixed solution obtained in step (I) with a reaction reagent (for example, at 0° C.) to obtain a mixed solution;

[0073] (III) adding Br2 to the mixed solution obtained in step (II) to react and obtain compound 2.

[0074] In a preferred embodiment, in step S1, the reaction temperature is -10 to 10°C, such as -5 to 5°C, and another example is 0°C.

[0075] In a preferred embodiment, in step S1, the reaction is carried out under light-proof conditions.

[0076] In a preferred embodiment, in step S1, the reaction is carried out under an inert atmosphere, such as a nitrogen atmosphere.

[0077] In a preferred embodiment, in step S1, the preparation method of compound 2 includes the following steps: adding compound 1 and a catalyst to a mixture consisting of a solvent and a reaction reagent, adding the reaction reagent after cooling (for example, 0°C) under the protection of an inert gas, adding Br2 dropwise, stirring at -5 to 5°C for 3 to 5 hours, and after the reaction is completed, post-treating to obtain the product.

[0078] In step S1, the progress of the reaction can be monitored using conventional monitoring methods for such reactions in the art (e.g., TLC or LC-MS), with the disappearance or no further reaction of compound 1 being considered the endpoint of the reaction. The reaction time can be 3 to 6 hours, e.g., 3 to 5 hours, or even 4 hours.

[0079] In a preferred embodiment, in step S1, after completion of the reaction, the following post-processing steps are further performed. The post-processing steps are conventional post-processing steps for such reactions in the art, for example, comprising one or more of quenching, crystallization, filtration, washing, and drying. The quenching agent may be propylene oxide. The crystallization solvent may be methyl tert-butyl ether. The washing solvent may be methyl tert-butyl ether.

[0080] The present invention provides a compound 3 or compound 4:

[0081] Each R 1 The definition of is as described in any embodiment of the present invention.

[0082] In a preferred embodiment, the compound 3 is

[0083] The present invention provides a method for preparing compound 5, which comprises the following step S4: in a solvent, subjecting compound 4 to a reaction as shown in the following formula under the action of a base to obtain compound 5.

[0084] The base is selected from one or both of sodium formate and sodium acetate. The conditions and operations of the preparation method of compound 5 can be as described in any one of the present invention.

[0085] In a preferred embodiment, the preparation method of compound 5 further includes the preparation method of compound 4, which comprises the following step S3: in a solvent, compound 3 is subjected to the reaction shown in the following formula under the action of hydrogen bromide to obtain compound 4,

[0086] Each R 1 The definition of is as described in any embodiment of the present invention;

[0087] In step S3, the reaction temperature is 30-50°C;

[0088] In step S3, the molar ratio of the hydrogen bromide to the compound 3 is (2-4):1.

[0089] The conditions and operations of the preparation method of compound 4 can be as described in any one of the present invention.

[0090] In a preferred embodiment, the preparation method of compound 4 further includes a preparation method of compound 3, which comprises the following step S2: in a solvent, in the presence of a base and a reducing agent, subjecting compound A-2 and / or its salt to a reductive amination reaction with compound 2s as shown in the following formula to obtain compound 3,

[0091] The salt of compound A-2, each R 1 The definition of is as described in any embodiment of the present invention;

[0092] The conditions and operations for the preparation of compound 3 may be as described in any of the schemes of the present invention.

[0093] In a preferred embodiment, the preparation method of compound 3 further includes a preparation method of compound 2, which comprises the following steps S1: in a solvent, in the presence of a catalyst and an additive, compound 1, Br2 and a reaction reagent are reacted as shown in the following formula to obtain compound 2,

[0094] The reaction reagents and each R 1 The definition of is as described in any embodiment of the present invention;

[0095] The conditions and operations for the preparation of compound 2 may be as described in any of the schemes of the present invention.

[0096] The present invention provides a method for preparing compound 3, comprising the following step S2: in a solvent, in the presence of a base and a reducing agent, subjecting compound A-2 and / or its salt to a reductive amination reaction with compound 2s as shown in the following formula to obtain compound 3,

[0097] The salt of compound A-2, each R 1 The definition of is as described in any embodiment of the present invention;

[0098] The conditions and operations for the preparation of compound 3 may be as described in any of the schemes of the present invention.

[0099] The present invention provides a crystalline form of the above-mentioned compound 3-1, whose X-ray powder diffraction pattern expressed in 2θ angles has diffraction peaks at 5.6°±0.2°, 7.3°±0.2°, 7.9°±0.2°, 8.8°±0.2° and 12.7°±0.2°.

[0100] In a preferred embodiment, the X-ray powder diffraction pattern of the crystalline form of compound 3-1 expressed in 2θ angles has diffraction peaks at 5.6°±0.2, 7.3°±0.2°, 7.9°±0.2°, 8.8°±0.2°, 10.0°±0.2°, 12.3°±0.2° and 12.7°±0.2°; preferably, there are diffraction peaks at 5.6°±0.2°, 6.5°±0.2°, 7.3°±0.2°, 7.9°±0.2°, 8.8°±0.2°, 10.0°±0.2°, 12.3°±0.2°, 12.7°±0.2°, 15.9°±0.2°, 16.9°±0.2° and 18.7°±0.2°.

[0101] The present invention provides a crystalline form of the above-mentioned compound 3-1, which is a single crystal, belongs to the monoclinic system, space group P21, and has unit cell parameters: α=90°,β=97.774(4)°,γ=90°,unit cell volume The number of asymmetric units in the unit cell is Z = 16, and the charge density is 1.412Mg / m 3 .

[0102] The present invention provides a method for preparing compound 7, which comprises the following steps: S6: in a solvent, in the presence of a catalyst and an organic base, subjecting compound 6 to a ring-closure reaction as shown in the following formula to obtain compound 7:

[0103] The organic base is 2,6-diisopropylaniline and / or N,N-diisopropylethylamine (DIEA / DIPEA);

[0104] The solvent is a mixed solvent consisting of a nitrile solvent and an ether solvent.

[0105] The present invention provides a method for preparing compound 7, which comprises the following steps: preparing compound 4 from compound 3, hydrolyzing compound 4 to obtain compound 5, and oxidizing compound 5 to obtain compound 6; and further comprising step S6: subjecting compound 6 to a ring-closure reaction as shown in the following formula in a solvent in the presence of a catalyst and an organic base to obtain compound 7.

[0106] Among them, each R 1is methyl, ethyl, n-propyl, isopropyl, n-butyl or n-pentyl;

[0107] The organic base is 2,6-diisopropylaniline and / or N,N-diisopropylethylamine (DIEA / DIPEA);

[0108] The solvent is a mixed solvent consisting of a nitrile solvent and an ether solvent.

[0109] In a preferred embodiment, in the method for preparing compound 7, certain conditions and operations have the following definitions, and the definitions of the conditions and operations not mentioned are as described in any embodiment of the present invention (this paragraph is hereinafter referred to as "in a preferred embodiment").

[0110] In a preferred embodiment, in step S6, the nitrile solvent is acetonitrile.

[0111] In a preferred embodiment, in step S6, the ether solvent is one or more of tetrahydrofuran, dioxane and ethylene glycol dimethyl ether, such as tetrahydrofuran.

[0112] In a preferred embodiment, in step S6, the volume ratio of the ether solvent to the nitrile solvent is (0.2-1):1; for example, (0.5-0.7):1, and another example is 0.6:1.

[0113] In a preferred embodiment, in step S6, the molar volume ratio of the compound 6 to the solvent is 1 mmol:(0.1-0.3)L, for example, 1 mmol:(0.1-0.2)L, and another example is 1 mmol:0.16L.

[0114] In a preferred embodiment, in step S6, the organic base is N,N-diisopropylethylamine.

[0115] In a preferred embodiment, in step S6, the molar ratio of the organic base to the compound 6 is (1-5):1, for example (1.5-3.5):1, and for example 3:1.

[0116] In a preferred embodiment, in step S6, the catalyst is a triazine compound, such as cyanuric chloride.

[0117] In a preferred embodiment, in step S6, the molar ratio of the catalyst to the compound 6 is (1-3):1, for example (1-2):1, and for example 1.5:1.

[0118] In a preferred embodiment, in step S6, the temperature of the ring-closure reaction is 5-25°C, such as 10-18°C, and another example is 15°C.

[0119] In a preferred embodiment, in step S6, the reaction is carried out under an inert atmosphere, such as a nitrogen atmosphere.

[0120] In a preferred embodiment, in step S6, the ring-closure reaction is carried out under light-shielding conditions.

[0121] In step S6, the progress of the ring-closure reaction can be monitored using conventional monitoring methods in the art (e.g., HPLC, TLC, or LC-MS). The reaction endpoint is generally determined by the disappearance or non-reaction of compound 6. The reaction time can be 2 to 7 hours, e.g., 2 to 5 hours, and further e.g., 2 hours.

[0122] In a preferred embodiment, in step S6, after the ring-closure reaction is completed, the following post-processing steps are further included. The post-processing steps are conventional post-processing operations for such reactions in the art, for example, including one or more of quenching, extraction, filtration, washing, drying, concentration and purification.

[0123] The quenching reagent may be an aqueous solution of 3-amino-1,2-propanediol (eg, 0.45 mol / L).

[0124] The quenching may include the following steps: reacting the mixture obtained after the ring-closure reaction with an aqueous solution of 3-amino-1,2-propanediol (eg, 0.45 mol / L).

[0125] The extraction solvent may be dichloromethane. The washing solution may be saturated saline. The drying agent may be anhydrous sodium sulfate. The purification may be performed by column chromatography, and the eluents for the column chromatography may be dichloromethane and methanol, wherein the volume ratio of dichloromethane to methanol may be (120-500):1.

[0126] In a preferred embodiment, the preparation method of compound 7 comprises the following steps: under the protection of inert gas, compound 6 and a solvent are mixed, the temperature is controlled to 10-18°C, protected from light, an organic base is added and stirred to dissolve, cyanuric chloride is added, the temperature is controlled to 10-18°C and the reaction is carried out for 2-7 hours, after the reaction is complete, 3-amino-1,2-propanediol aqueous solution is added to quench, water and dichloromethane are added to extract, and after the insoluble matter is precipitated, the compound is filtered with diatomaceous earth, the organic phase is washed, dried, concentrated, and column chromatographed to obtain the compound.

[0127] In a preferred embodiment, the preparation method of compound 7 also includes the preparation method of compound 6, and the preparation method of compound 6 includes the following step S5: in a solvent, in the presence of an oxidant, subjecting compound 5 to an oxidation reaction as shown in the following formula to obtain compound 6,

[0128] In a preferred embodiment, in step S5, the solvent is a chlorinated alkane organic solvent, such as one or more of dichloromethane, 1,2-dichloroethane and chloroform, and another example is dichloromethane.

[0129] In a preferred embodiment, in step S5, the molar volume of the compound 5 and the solvent is (1-30) mmol:1L, for example (10-20) mmol:1L, and another example is 15.6 mmol:1L.

[0130] In a preferred embodiment, in step S5, the oxidant is selected from one or more of Jones reagent, Collins reagent, pyridinium chlorochromate, pyridinium dichromate, manganese dioxide, Dess-Martin periodinane, potassium permanganate, periodic acid, osmium tetroxide, 30% hydrogen peroxide, meta-chloroperbenzoic acid and tert-butyl hydroperoxide; for example, meta-chloroperbenzoic acid (m-CPBA).

[0131] In a preferred embodiment, in step S5, the molar ratio of the oxidant to the compound 5 is (1-3):1, for example (1-2):1, and another example is 1.2:1.

[0132] In a preferred embodiment, in step S5, the oxidant is added at a temperature of -10 to 0°C, for example, at -5°C.

[0133] In a preferred embodiment, in step S5, the oxidation reaction is carried out under light-proof conditions.

[0134] In a preferred embodiment, in step S5, the temperature of the oxidation reaction is -20 to 25°C, such as 0 to 5°C, and another example is 0°C.

[0135] In step S5, the progress of the oxidation reaction can be monitored by conventional monitoring methods in the art (e.g., HPLC, TLC, or LC-MS), and the disappearance or no further reaction of compound 5 is used as the reaction endpoint. The reaction time can be 1 to 20 hours, for example, 1 to 3 hours, and for example, 2 hours.

[0136] In a preferred embodiment, in step S5, after the oxidation reaction is completed, the following post-processing steps are further included, and the post-processing steps are conventional post-processing operations for such reactions in the art, for example, including one or more of quenching, concentration, and purification. The quenching reagent can be thioanisole. The purification can be column chromatography, and the eluents for the column chromatography can be dichloromethane and methanol, and the volume ratio of dichloromethane to methanol can be (10-100):1.

[0137] In a preferred embodiment, the preparation method of compound 6 comprises the following steps: adding a solvent to compound 5, cooling it (for example, to -5°C), adding an oxidant, stirring at 0-5°C for 1-3 hours, and after the reaction is completed, performing post-treatment to obtain the compound.

[0138] The present invention provides a crystalline form of compound 7, which is a single crystal, belongs to the monoclinic system, space group P21, and unit cell parameters: α=90°,β=92.23(3)°,γ=90°,unit cell volume The number of asymmetric units in the unit cell is Z = 4, and the charge density is 1.516Mg / m 3 ;

[0139] The present invention provides a method for preparing ADC, which comprises the following steps:

[0140] (1) Preparation method of compound 7: comprising the following steps S6: in a solvent, in the presence of a catalyst and an organic base, compound 6 undergoes a ring-closure reaction as shown in the following formula to obtain compound 7

[0141] The conditions and operations of the preparation method of compound 7 can be as described in any scheme of the present invention;

[0142] (2) preparing the ADC compound using compound 7 as a raw material;

[0143] The ADC compounds include but are not limited to the following drugs:

[0144] Anti-CD22-NMS249, huXBR1-402-PNU, NAV-001, NBE-002, SOT102, EGFR VEDVsPNU+EDVs40mer, ADC-1, NBE-001, PNU-159682-P1C1TM or SOT107.

[0145] The structural formula of Anti-CD22-NMS249 is as follows. The synthesis method of its payload+linker is referred to patent WO2009099741A1.

[0146] The structural formula of NBE-002 is as follows. The synthesis method of its payload+linker is based on patent WO2016102679 A1.

[0147] Without violating the common sense in the art, the above-mentioned preferred conditions can be arbitrarily combined to obtain preferred embodiments of the present invention.

[0148] The reagents and raw materials used in the present invention are commercially available.

[0149] The positive progress effect of the present invention is:

[0150] 1) The present invention provides a method for preparing a PNU-159682 intermediate, using daunorubicin hydrochloride as a raw material to synthesize a novel compound 3. Compound 3 is further deprotected and hydrolyzed to obtain the target intermediate. This method not only overcomes the high cost disadvantage of the existing technology, but also uses these intermediates to design mild reaction conditions, high safety, high yield, and low post-processing difficulty, providing a guarantee for the subsequent preparation of high-quality drugs. The method is suitable for laboratory preparation research and is particularly suitable for industrial large-scale production.

[0151] 2) Although the prior art has reported the preparation of compound 2, the prior art has not reported the direct reductive amination reaction using compound 2 as a raw material. This may be because glycoside compounds are generally unstable under acidic and alkaline conditions, which is not conducive to the deprotection of ketal. as well as The methoxy group in the fragment is unstable and easily hydrolyzed. Based on the structural characteristics of this type of compound, the inventors unexpectedly discovered that by controlling the ratio between the substrate and the reaction reagent in the reaction system, as well as the reaction temperature, the stability of both the glycoside and methoxy sites can be simultaneously solved, further improving the reaction yield.

[0152] 3) The present invention provides a method for preparing PNU-159682. The inventors have found in experiments that by selecting a base and regulating the combination of solvents, the formation of by-products is effectively reduced, and the technical problem of low yield in the prior art is effectively solved. It also solves the technical problem of the inability to scale up production in the prior art.

[0153] 4) Furthermore, using daunorubicin hydrochloride as a raw material, a novel intermediate compound 3 was synthesized. Compound 3 was further deprotected and hydrolyzed to obtain the target intermediate. This not only overcomes the high cost defect of the existing technology, but also the reaction conditions designed using these intermediates are mild, safe, high yield, and low post-processing difficulty, providing a guarantee for the subsequent preparation of high-quality drugs. It is suitable for laboratory preparation research and is especially suitable for industrial large-scale production.

[0154] 5) Although the prior art has reported the preparation of compound 2, the prior art has not reported the direct reductive amination reaction using compound 2 as a raw material. This may be because glycoside compounds are generally unstable under acidic and alkaline conditions, which is not conducive to the deprotection of ketal. as well as The methoxy group in the fragment is unstable and easily hydrolyzed. Based on the structural characteristics of this type of compound, the inventors unexpectedly discovered that by controlling the ratio between the substrate and the reaction reagent in the reaction system, as well as the reaction temperature, the stability of both the glycoside and methoxy sites can be simultaneously solved, further improving the reaction yield. BRIEF DESCRIPTION OF THE DRAWINGS

[0155] Figure 1 is an XPRD diagram of the crystalline form of compound 3-1;

[0156] Figure 2 is a crystal structure diagram of the crystalline form of compound 3-1;

[0157] Figure 3 is a unit cell stacking diagram of the crystalline form of compound 3-1;

[0158] Figure 4: Crystal structure of the crystalline form of compound 7;

[0159] Figure 5: Unit cell stacking diagram of the crystalline form of compound 7. DETAILED DESCRIPTION

[0160] The present invention is further illustrated by way of examples below, but the present invention is not limited to the scope of the examples. Experimental methods in the following examples where specific conditions are not specified were performed according to conventional methods and conditions, or selected according to the product specifications.

[0161] Unless otherwise stated, the units used in the following experimental descriptions represent the following definitions: N: mol / L; V: ml / g.

[0162] Example 1: Preparation of Compound 2-1

[0163] Compound 1 (100 g, 177.31 mmol, 1.0 eq) and p-toluenesulfonic acid (3.05 g, 17.73 mmol, 0.1 eq) were dissolved in methanol (750 mL, 7.5 V) and 1,4-dioxane (750 mL, 7.5 V). The mixture was protected from light and cooled to 0°C under nitrogen. Trimethyl orthoformate (347.7 g, 3280 mmol, 18.5 eq) was added. Br2 (liquid bromine 11.8 mL, 1.3 eq) was diluted with dichloromethane (200 mL). Then add it dropwise to the reaction mixture, stir it at 0°C for 4h. After the reaction is completed, add propylene oxide (32mL) to quench the mixture, stir it at 0°C for 1h, add methyl tert-butyl ether (7.2L) to crystallize, stir it at 0°C for 0.5h, filter, wash the filter cake with methyl tert-butyl ether, and dry it to obtain compound 2-1 (127.44g, 98%, purity 95.6%).

[0164] 1H NMR (400MHz, DMSO) δ7.88(q,J=7.7Hz,2H),7.62(d,J=7.7Hz,1H),5.45(s,1H),5.27(s,1H),4.8 7(d,J=4.1Hz,1H),4.57(s,1H),4.18(q,J=6.3Hz,1H),4.00(d,J=18.1Hz,5H),3.60(s,1H),3.4 9(d,J=6.2Hz,6H),3.07(d,J=18.6Hz,1H),2.79(d,J=18.7Hz,1H),2.38(d,J=14.9Hz,1H),2.00 (dd,J=14.9,4.9Hz,1H),1.88(t,J=14.0Hz,1H),1.68(d,J=11.8Hz,1H),1.18(d,J=6.4Hz,3H).

[0165] Example 2: Study on the influence of reaction conditions on the reaction in the preparation of compound 2-1

[0166] 2.1: Effect of trimethyl orthoformate CH(OCH3)3 on the reaction in the preparation of compound 2-1

[0167] Referring to the preparation method of Example 1, the amount of trimethyl orthoformate was varied to investigate its effect on the reaction. The IPC purity of the raw material and product in the reaction solution after 4 and 6 hours of reaction was tested by HPLC, and the results are shown in Table 1.

[0168] Table 1

[0169] From the results in Table 1, it can be seen that as the amount of CH(OCH3)3 decreases, the corresponding reaction is slower, the reaction is more complicated, and the raw materials cannot be completely converted; as the amount increases, the purity decreases; 18.5eq is the optimal, the raw materials are completely converted, but the purity decreases as the reaction time continues to increase.

[0170] 2.2: Study on the effect of p-toluenesulfonic acid TsOH on the reaction in the preparation of compound 2-1

[0171] Referring to the preparation method of Example 1, the amount of p-toluenesulfonic acid TsOH was varied to investigate its effect on the reaction. The IPC purity of the raw material and product in the reaction solution after 3 hours, 4 hours, and 6 hours of reaction was tested by HPLC, and the results are shown in Table 2.

[0172] Table 2

[0173] From the results in Table 2, it can be seen that the reaction speed becomes faster when TsOH is added, but the product will decompose when the reaction time is prolonged. When 0.1eq of p-toluenesulfonic acid is added and the reaction time is 4h, the reaction effect is the best.

[0174] 2.3: Study on the effect of reaction temperature on the reaction in the preparation of compound 2-1

[0175] The reaction temperature was changed to investigate its effect on the reaction effect, referring to the preparation method of Example 1. The IPC purity of the raw material and product in the reaction solution after 5 hours of reaction was tested by HPLC. The results are shown in Table 3.

[0176] Table 3

[0177] From the results in Table 3, we can see that at -15-5°C, the temperature is too low and the product purity is very low. At 10-20°C, the reaction proceeds very quickly and the stability deteriorates. The best results are obtained at a temperature of -5-5°C.

[0178] Example 3: Preparation of Compound 3-1

[0179] Compound 2-1 (100 g, 136.35 mmol, 1 eq) was added to tetrahydrofuran (1.5 L), protected from light, cooled to 0 ° C, added with sodium bicarbonate (34.36 g, 409.05 mmol, 3.0 eq), stirred for 5 min, and 2-[(1S)-1-methoxy-2-oxoethoxy] acetaldehyde (54.04 g, 409.05 mmol, 3.0 eq) and sodium cyanoborohydride (12.85 g, 204.53 mmol, 1.5 eq) were added dropwise. After the addition, the mixture was stirred at 0 ° C for 1 h and at 25 ° C for 16 h. After the reaction was completed, it was filtered and extracted with methyltetrahydrofuran (1 L). The aqueous phase was extracted with methyltetrahydrofuran (500 mL × 2), the organic phases were combined, washed with water (1 L), dried, concentrated and spin-dried, and purified by column (DCM: MeOH =80:1), and the filtrate was concentrated to dryness to obtain compound 3-1 (98.51 g, yield 96%, purity 95.9%).

[0180] 1H NMR (400MHz, DMSO) δ13.91(d,J=5.7Hz,1H),13.14(d,J=6.5Hz,1H),7.87–7.79(m,1H),7.76(t,J=7.4Hz,1H),7.55(t,J=6.6Hz,1H),5.28(d ,J=2.4Hz,1H),4.89(s,1H),,4.36(d,J=3.0Hz,1H),4.09(t,J=8.7Hz,1H),4.04–3.98(m,3H),3.93(s,3H),3.87–3.79(m,1H),3.78–3.69(m, 2H),3.62–3.52(m,2H),3.50(d,J=2.1Hz,3H),3.49(s,3H),3.45–3.37(m,1H),3.24(d,J=7.2Hz,3H),3.03(d,J=18.8Hz,1H),2.77(d,J=18. 7Hz,1H),2.61(d,J=9.6Hz,1H),2.45(s,1H),2.39(d,J=12.6Hz,2H),2.25(dd,J=11.0,5.4Hz,1H),1.86–1.77(m,2H),1.13(t,J=4.6Hz,3H).

[0181] Example 4: Study on the Effect of Different Solvents on the Reaction in the Preparation of Compound 3-1

[0182] Referring to the preparation method of Example 3, the solvent was changed to investigate its effect on the reaction effect. The IPC purity of the raw material and product in the reaction solution after 4 hours of reaction was tested by HPLC. The results are shown in Table 4.

[0183] Table 4

[0184] As shown in Table 4, THF is the best solvent for the reaction.

[0185] Example 5: Preparation of Compounds 4 and 5

[0186] Compound 3-1 (100 g, 132.87 mmol, 1.0 eq) was dissolved in acetone (2 L), added to an aqueous hydrogen bromide solution (0.15 mol / L, 2 L), protected from light, and heated to 40 ° C under nitrogen protection. The reaction was completed after stirring for 4 hours; sodium formate (54.22 g, 797.23 mmol, 6 eq) was added and stirred at 40 ° C for 20 hours. After the reaction was completed, the reaction solution was diluted with water (3 L), extracted with DCM (5 L), and the aqueous phase was extracted with DCM (2 L × 3). The organic phases were combined, dried, concentrated, and slurried with methyl tert-butyl ether (1 L), filtered, and dried to obtain compound 5 (81.25 g, yield 95%, purity 92%).

[0187] 1 H NMR (400MHz, CDCl3) δ13.93 (s, 1H), 13.19 (s, 1H), 8.00 (d, J = 7.6Hz, 1H), 7.77 (t, J = 8.0Hz, 1H),7.38(d,J=8.4Hz,1H),6.08(s,7H),5.55(s,1H),5.27(s,1H),4.74(d,J=13.8Hz,3H),4.50(s,1H ),4.08(s,3H),3.97–3.91(m,2H),3.76(s,21H),3.60–3.47(m,2H),3.39(s,3H),3.23(d,J=18.7Hz,1H ),3.05(d,J=4.9Hz,2H),2.97(d,J=18.7Hz,1H),2.61(dd,J=11.1,3.5Hz,1H),2.50(s,3H),2.37(d,J =14.2Hz, 2H), 2.16 (dd, J = 14.6, 3.5Hz, 1H), 1.79 (d, J = 7.0Hz, 2H), 1.38 (d, J = 6.5Hz, 3H), 0.07 (s, 3H).

[0188] Example 6: Study on the effects of different conditions on the reaction during the preparation of compounds 4 and 5

[0189] 6.1: Study on the Effect of Reaction Temperature on the Deketalization Reaction in the Preparation of Compound 4

[0190] Referring to the preparation method of Example 5 (i.e., Compound 3-1 to Compound 4), the reaction temperature was varied to investigate its effect on the reaction performance. The IPC purity of the raw materials and products in the reaction solutions after 2, 4, and 6 hours of reaction were measured by HPLC, and the results are shown in Table 5.

[0191] Table 5

[0192] From the results in Table 5, it can be seen that the lower the temperature, the slower the reaction. The reaction is fastest at 40°C, and the raw materials are completely converted in 4 hours. Prolonging the reaction time reduces the purity and significantly increases the impurities.

[0193] 6.2: Study on the Effect of Deprotection Reagents on the Deketalization Reaction in the Preparation of Compound 4

[0194] Referring to the preparation method of Example 5 (i.e., compound 3-1 to compound 4), the amount of hydrobromic acid used as the deprotection reagent was varied to examine its effect on the reaction. The IPC purity of the starting material and product in the reaction solution after 4 and 6 hours of reaction was determined by HPLC, as shown in Table 6.

[0195] Table 6

[0196] From the results in Table 6, it can be seen that acid is a deketalization catalyst, but it will affect the stability of the substrate. It can be seen that with less acid, the reaction conversion is slower; with the same amount of acid, the reaction time is extended and the reaction is more complicated (side reactions such as deglycosidation); with too much acid, the side reactions increase and the purity decreases.

[0197] 6.3: Study on the influence of reaction conditions on the preparation of compound 5

[0198] Referring to the preparation method of Example 5 (i.e., compound 3-1 to compound 5), the reaction volume was changed to investigate its effect on the reaction effect. The IPC purity of the product in the reaction solution after the reaction was completed was tested by HPLC, and the results are shown in Table 7.

[0199] Table 7

[0200] From the results in Table 7, it can be seen that the reaction volume has an impact on the control of reaction impurities.

[0201] 6.4: Study on the Effect of Sodium Formate Equivalent on the Reaction in the Preparation of Compound 5

[0202] Referring to the preparation method of Example 5 (i.e., compound 4 to compound 5), the amount of sodium formate was changed to investigate its effect on the reaction effect. The IPC purity of the product in the reaction solution after the reaction was completed was tested by HPLC, and the results are shown in Table 8.

[0203] Table 8

[0204] Example 7: Preparation method of the crystalline form of compound 3-1.

[0205] Single crystal cultivation: 5 mg of compound 3-1 prepared in Example 3 was weighed and dissolved in 0.5 mL of acetonitrile, and liquid phase diffusion was performed with 1.5 mL of water to obtain a crystalline sample.

[0206] The XPRD pattern of the crystalline form of compound 3-1 is shown in Figure 1; the X-ray powder diffraction pattern expressed in 2θ angles has diffraction peaks at 5.6°±0.2°, 6.5°±0.2°, 7.3°±0.2°, 7.9°±0.2°, 8.8°±0.2°, 10.0°±0.2°, 12.3°±0.2°, 12.7°±0.2°, 15.9°±0.2°, 16.9°±0.2° and 18.7°±0.2°.

[0207] Single crystal detection method of compound 3-1: Bruker D8 VENTURE double microfocus single crystal X-ray diffractometer, PHOTON III charge integrating pixel array detector, CuKα radiation, wavelength The distance between the crystal and the detector is d = 40 mm, the tube voltage is 50.0 kV, the tube current is 1.100 mA, and the number of independent diffraction points is 39319.

[0208] The crystal structure diagram of the compound 3-1 crystal form is shown in Figure 2. The unit cell stacking diagram of the compound 3-1 crystal form is shown in Figure 3.

[0209] The crystallographic parameters of compound 3-1 are shown in Table 9.

[0210] Table 9

[0211] Example 8: Preparation of Compound 6

[0212] Compound 5 (100 g, 155.37 mmol, 1.0 eq) was added to DCM (10 L, 100 V), protected from light, cooled to -5°C, and m-chloroperbenzoic acid (m-CPBA) (32.17 g, 186.44 mmol, 1.2 eq) was added and the temperature was raised to 0°C. The mixture was stirred for 2 h and monitored by TLC / LCMS / HPLC. After the reaction was complete, thioanisole (4 mL, 0.2 eq) was added and stirred for 1 h to quench the reaction. The mixture was concentrated and purified by column chromatography (DCM:MEOH=100:1~10:1) to obtain compound 6 (91.21 g, yield 89%, purity 95%).

[0213] 1H NMR (400MHz, CDCl3) δ13.87(d,J=5.9Hz,1H),13.09(s,1H),7.89(d,J=7.4Hz,1H),7.70(t, J=8.1Hz,1H),7.30(t,J=8.8Hz,1H),5.61(d,J=3.2Hz,1H),5.21(d,J=1.8Hz,1H),4.77(d,J=3.6Hz,1H),4.67(s,2H ),4.62(d,J=11.3Hz,1H),4.18(s,2H),4.00(s,3H),3.87(dd,J=13.0,6.4Hz,1H),3.75(d,J=12.6Hz,1H),3.51(t,J= 14.5Hz,2H),3.42–3.35(m,3H),3.24(ddd,J=37.4,19.0,12.4Hz,5H),3.09–2.98(m,1H),2.92(t,J=17.7Hz,1H),2. 77(td,J=12.8,4.0Hz,1H),2.33(d,J=14.8Hz,1H),2.15–2.04(m,1H),1.93(d,J=9.6Hz,1H),1.29(t,J=12.2Hz,3H).

[0214] Example 9: Preparation of Compound 7

[0215] Under the protection of inert gas (nitrogen), compound 6 (10 g, 15.2 mmol, 1.0 eq) and a mixture of acetonitrile / tetrahydrofuran (1.5 L / 0.9 L) were added to the reaction flask, the temperature was controlled to 15 ° C, and the mixture was protected from light. N, N-diisopropylethylamine (5.9 g, 45.6 mmol, 3.00 eq) was added and stirred to dissolve. Then, cyanuric chloride (4.2 g, 22.8 mmol, 1.5 eq) was added and the temperature was controlled to 15 ° C for 2 h. After the reaction was complete, , add 3-amino-1,2-propanediol aqueous solution (0.2 L, 0.45 mol / L) and stir for 10 min, then add water (1.8 L) and dichloromethane (1.8 L) and stir for 10 min. After the insoluble matter is precipitated, filter with celite, wash the organic phase twice with saturated brine, dry over anhydrous sodium sulfate, filter, and concentrate, and perform column chromatography (DCM / MeOH = 500: 1 ~ 120: 1) to give compound 7 (6 g, yield 62%, purity 95%).

[0216] 1H NMR (400MHz, CDCl3) δ13.86(s,1H),13.20(s,1H),8.02(d,J=7.6Hz,1H),7.80(t,J=8.1Hz,1H),7.42(d,J=8.4Hz,1H),5.50(t,J=5.4Hz,1H),5 .31(s,1H),4.86(d,J=16.3Hz,1H),4.79(s,2H),4.74(s,1H),4.50(s, 1H),4.11(s,4H),4.07–4.00(m,1H),3.95(t,J=9.3Hz,1H),3.61(d,J=1 1.7Hz,1H),3.49(s,3H),3.42(dd,J=12.3,6.1Hz,1H),3.24(d,J=18.8Hz,1H),3.09(s,1H),3.01(t,J=14.4Hz,1H),2.80(dt,J=17.9,10.3Hz, 2H), 2.53(d,J=14.7Hz,1H), 2.14(dd,J=14.6,3.5Hz,1H), 2.04(dd,J=10.1,4.7Hz,1H), 1.77(dt,J=15.0,6.0Hz,1H), 1.41(t,J=12.5Hz,3H).

[0217] Example 10: Study on the influence of reaction conditions on the reaction in the preparation of compound 7

[0218] 9.1: Study on the Effect of Base on the Reaction in the Preparation of Compound 7

[0219] Referring to the preparation method of Example 9, the type of base was changed to investigate its effect on the reaction effect. The IPC purity of the product in the reaction solution after 1 hour and 3 hours of reaction was tested by HPLC. The results are shown in Table 10.

[0220] Table 10

[0221] 9.2: Study on the Effect of Solvent on the Reaction in the Preparation of Compound 7

[0222] Referring to the preparation method of Example 9, the type of solvent was changed to investigate its effect on the reaction effect. The IPC purity of the product in the reaction solution after the reaction was tested by HPLC, and the results are shown in Table 11.

[0223] Table 11

[0224] Example 11: Preparation of single crystal of compound 7 (3 molecules of chloroform solvate)

[0225] 5 mg of compound 7 prepared in Example 9 was completely dissolved in 30 μL of chloroform and allowed to stand to cool to obtain the product.

[0226] Single crystal detection method of compound 7: using a Bruker D8 VENTURE (liquid metal target) single crystal X-ray diffractometer, PHOTON III charge integrating pixel array detector, 250W anode liquid gallium target X-ray generator, GaKα radiation, wavelength The distance between the crystal and the detector is d = 37 mm, the tube voltage is 69.7 kV, the tube current is 2.855 mA, and the number of independent diffraction points is 12210.

[0227] The crystal structure diagram of the crystalline form of Compound 7 is shown in FIG4 ; the unit cell stacking diagram of the crystalline form of Compound 7 is shown in FIG5 .

[0228] The crystallographic parameters of the crystalline form of compound 7 are shown in Table 12.

[0229] Table 12

Claims

1. A method for preparing compound 5, characterized in that: The preparation method of compound 5 comprises the following steps S2, S3 and S4: Step S2: In a solvent, in the presence of a base and a reducing agent, compound A-2 and / or its salt is subjected to a reductive amination reaction with compound 2s as shown in the following formula to obtain compound 3, Step S3: Compound 3 is subjected to the following reaction under the action of hydrogen bromide in a solvent to obtain compound 4: Among them, each R 1 is methyl, ethyl, n-propyl, isopropyl, n-butyl or n-pentyl; In step S3, the reaction temperature is 30-50°C; In step S3, the molar ratio of hydrogen bromide to compound 3 is (2-4):1; Step S4: In a solvent, compound 4 is subjected to a hydrolysis reaction as shown in the following formula under the action of a base to obtain compound 5:

2. The method for preparing compound 5 according to claim 1, characterized in that: It meets one or more of the following conditions: (1) Each R 1 is methyl; (2) The salt of compound A-2 is selected from One or more of (3) In step S2, the solvent is an ether solvent, and the ether solvent can be selected from one or more of ethylene glycol dimethyl ether, 1,4-dioxane and tetrahydrofuran, such as tetrahydrofuran; (4) In step S2, the molar volume ratio of the "compound A-2 and / or its salt" to the solvent is (50-150) mmol:1L, for example (80-100) mmol:1L, and another example is 91 mmol:1L; (5) In step S2, the base is an inorganic base; for example, the inorganic base is selected from one or more of sodium bicarbonate, potassium bicarbonate, sodium carbonate, potassium carbonate and calcium carbonate, tripotassium phosphate, trisodium phosphate, dipotassium hydrogen phosphate and disodium hydrogen phosphate; another example is sodium bicarbonate; (6) In step S2, the molar ratio of the base to the "compound A-2 and / or its salt" is (1-3):1, for example (2.5-3.5):1, and for example 3:1; (7) In step S2, the reducing agent is selected from one or more of sodium borohydride, potassium borohydride, lithium borohydride, sodium cyanoborohydride and lithium aluminum hydride, such as sodium cyanoborohydride; (8) In step S2, the molar ratio of the reducing agent to the "compound A-2 and / or its salt" is (1-5):1, for example (1-3):1, and for example 1.5:1; (9) In step S2, the molar ratio of the compound 2s to the "compound A-2 and / or its salt" is (1-5):1, for example (2-4):1, Another example is 3:1; (10) In step S2, the temperature of the reductive amination reaction is 10 to 25° C., for example, 25° C.; (11) In step S2, the reductive amination reaction is carried out under light-proof conditions; (12) In step S2, the reductive amination reaction time is 10 to 20 hours, for example 15 to 20 hours, and for example 16 hours; (13) In step S2, after the reductive amination reaction is completed, the following post-treatment steps are further included: one or more of filtration, extraction, washing, drying, concentration and purification; the extraction solvent may be methyltetrahydrofuran; the washing solvent may be water; the purification may be column chromatography, the eluent of the column chromatography may be dichloromethane and methanol, and the volume ratio of dichloromethane to methanol may be 80:1; (14) In step S3, the solvent is a ketone organic solvent, such as acetone; (15) In step S3, the molar volume ratio of the compound 3 to the solvent is (50-150) mmol:1 L, for example (50-133) mmol:1 L, and for example 66 mmol:1 L; (16) In step S3, the hydrogen bromide is added in the form of an aqueous hydrogen bromide solution; for example, an aqueous hydrogen bromide solution, wherein the molar concentration of hydrogen bromide in the aqueous hydrogen bromide solution may be 0.15 to 0.3 mol / L, for example, 0.15 mol / L; (17) In step S3, the molar ratio of hydrogen bromide to compound 3 is 2.3:1 or 3.4:1; (18) In step S3, the reaction temperature is 35 to 45° C., for example, 40° C.; (19) In step S3, the reaction is carried out under light-proof conditions; (20) In step S3, the reaction is carried out under an inert atmosphere, such as a nitrogen atmosphere; (21) In step S3, the reaction time is 4 to 6 hours, for example 4 hours; (22) In step S3, after the reaction is completed, the next step of reaction is directly carried out without any post-treatment; (23) In step S4, the solvent is a ketone organic solvent, such as acetone; (24) In step S4, the molar volume ratio of the compound 4 to the solvent is (50-150) mmol:1 L, for example (50-133) mmol:1 L, and for example 66 mmol:1 L; (25) In step S4, the base is selected from one or both of sodium formate and sodium acetate; for example, sodium formate; (26) In step S4, the molar ratio of the base to the compound 4 is (3.5-13.5):1, for example (4-8.5):1, for example (5-8.5):1, and more preferably 6:1; (27) In step S4, the temperature of the hydrolysis reaction is 30 to 50° C., for example, 35 to 45° C., for example, 40° C.; (28) In step S4, the hydrolysis reaction is carried out under light-proof conditions; (29) In step S4, the hydrolysis reaction is carried out under an inert atmosphere, such as a nitrogen atmosphere; (30) In step S4, the hydrolysis reaction time is 15 to 25 hours, for example 18 to 22 hours, and for example 20 hours; and (31) In step S4, after the hydrolysis reaction is completed, the following post-treatment steps are also included: one or more of dilution, extraction, drying, concentration, pulping and filtration; the dilution solvent can be water; the extraction solvent can be dichloromethane; the pulping solvent can be methyl tert-butyl ether.

3. The method for preparing compound 5 according to claim 1, characterized in that: The step S2 comprises the following steps: in a solvent, in the presence of a base and a reducing agent, subjecting compound 2 and compound 2s to a reductive amination reaction as shown in the following formula to obtain compound 3:

4. The method for preparing compound 5 according to claim 3, characterized in that: When compound 2 is used as a reaction substrate in step S2, the preparation method of compound 5 also includes a preparation method of compound 2, and the preparation method of compound 2 includes the following steps S1: in a solvent, in the presence of a catalyst and an additive, compound 1, Br2 and a reaction reagent are reacted as shown in the following formula to obtain compound 2, The reaction reagent is R 1 OH; each R 1 The definition as claimed in claim 1 or 2.

5. The method for preparing compound 5 according to claim 4, characterized in that: The preparation method of the compound 2 satisfies one or more of the following conditions: (1) In step S1, the solvent is an ether solvent; preferably, the ether solvent is ethylene glycol dimethyl ether, 1,4-dioxane or tetrahydrofuran, such as 1,4-dioxane; (2) In step S1, the reaction reagent is methanol, ethanol, 1-propanol, 1-butanol or 1-pentanol; for example, methanol; (3) In step S1, the volume ratio of the reaction reagent to the solvent is 1:(0.5-1.5), for example 1:(0.5-1), and for example 1:1; (4) In step S1, the molar volume ratio of the compound 1 to the solvent is (100-300) mmol:1L, for example (200-260) mmol:1L, and for example 236 mmol:1L; (5) In step S1, the additive is trimethyl orthoformate, triethyl orthoformate or triisopropyl orthoformate, for example, trimethyl orthoformate; (6) In step S1, the molar ratio of the additive to the compound 1 is (17-20):1, for example (18-19):1, and for example 18.5:1; (7) In step S1, the catalyst is a sulfonic acid, such as p-toluenesulfonic acid, benzenesulfonic acid, 4-ethylbenzenesulfonic acid, 4-chlorobenzenesulfonic acid, xylenesulfonic acid, 2,3-dimethylbenzenesulfonic acid, 2,4-dimethylbenzenesulfonic acid, 2,5-dimethylbenzenesulfonic acid, 2,6-dimethylbenzenesulfonic acid or methanesulfonic acid, Another example is p-toluenesulfonic acid; (8) In step S1, the molar ratio of the catalyst to the compound 1 is (0.1-0.3):1, for example 0.1:1; (9) In step S1, the Br2 is added in the form of a dichloromethane solution of Br2, and the volume ratio of dichloromethane to Br2 is (1-20):1, for example (6-18):1, and another example is 16.9:1; (10) In step S1, the molar ratio of Br2 to compound 1 is (1-3):1, for example (1-2):1, and for example 1.3:1; (11) In step S1, the Br2 is added dropwise; (12) Step S1 includes the following steps: (I) mixing compound 1, a catalyst, a reaction reagent and a solvent to obtain a mixed solution; (II) mixing the mixed solution obtained in step (I) with a reaction reagent (for example, mixing at 0° C.) to obtain a mixed solution; (III) adding Br2 to the mixed solution obtained in step (II) to react to obtain compound 2; (13) In step S1, the reaction temperature is -10 to 10°C, for example -5 to 5°C, and for example 0°C; (14) In step S1, the reaction is carried out under light-proof conditions; (15) In step S1, the reaction is carried out under an inert atmosphere, such as a nitrogen atmosphere; (16) The reaction time is 3 to 6 hours, for example 3 to 5 hours, for example 4 hours; and (17) In step S1, after the reaction is completed, the following post-treatment steps are further included: one or more of quenching, crystallization, filtration, washing and drying; the quenching reagent may be propylene oxide; the crystallization solvent may be methyl tert-butyl ether; the washing solvent may be methyl tert-butyl ether.

6. A method for preparing compound 5, characterized in that: The preparation method of the compound 5 comprises the following step S4: in a solvent, subjecting the compound 4 to a hydrolysis reaction as shown in the following formula under the action of a base to obtain the compound 5, The base is selected from one or both of sodium formate and sodium acetate. The conditions and operations of the preparation method of compound 5 can also be as described in claim 2.

7. The method for preparing compound 5 according to claim 6, characterized in that: The preparation method of the compound 5 further includes the preparation method of the compound 4, and the preparation method of the compound 4 includes the following step S3: in a solvent, compound 3 is subjected to a reaction shown in the following formula under the action of hydrogen bromide to obtain compound 4, Each R 1 as defined in claim 1 or 2; In step S3, the reaction temperature is 30-50°C; In step S3, the molar ratio of hydrogen bromide to compound 3 is (2-4):1; The conditions and operations of the preparation method of compound 4 may also be as described in claim 2.

8. A method for preparing compound 3, characterized in that: The preparation method of the compound 3 comprises the following step S2: in a solvent, in the presence of a base and a reducing agent, subjecting the compound A-2 and / or its salt to a reductive amination reaction with the compound 2s as shown in the following formula to obtain the compound 3, The salt of compound A-2 is defined as in claim 2 or 3, and each R 1 as defined in claim 1 or 2; The conditions and operations of the preparation method of compound 3 may be as described in claim 2 or 3.

9. A compound 3, a compound 3-1 or a crystal form thereof, a compound 4 or a crystal form of compound 7, characterized in that: Each R 1 as defined in claim 1 or 2; Preferably, the compound 3 is The X-ray powder diffraction pattern of the crystalline form of the compound 3-1 expressed in 2θ angle has diffraction peaks at 5.6°±0.2°, 7.3°±0.2°, 7.9°±0.2°, 8.8°±0.2° and 12.7°±0.2°; preferably, at 5.6°±0.2, 7.3°±0.2°, 7.9°±0.2°, 8.8°±0.2°, 10.0°±0.2°, 12.3°±0 .2° and 12.7°±0.2° have diffraction peaks; more preferably, at 5.6°±0.2°, 6.5°±0.2°, 7.3°±0.2°, 7.9°±0.2°, 8.8°±0.2°, 10.0°±0.2°, 12.3°±0.2°, 12.7°±0.2°, 15.9°±0.2°, 16.9°±0.2° and 18.7°±0.2° have diffraction peaks; The crystal form of compound 7 is a single crystal, belongs to the monoclinic system, space group P21, and the unit cell parameters are: α=90°,β=92.23(3)°,γ=90°,unit cell volume The number of asymmetric units in the unit cell is Z = 4, and the charge density is 1.516Mg / m 3 ; 10. A method for preparing compound 7, characterized in that: The preparation method of compound 7 comprises the following steps: preparing compound 4 from compound 3, obtaining compound 5 from compound 4 through hydrolysis reaction, and obtaining compound 6 from compound 5 through oxidation reaction; and further comprises S6: in a solvent, in the presence of a catalyst and an organic base, subjecting compound 6 to a ring-closing reaction as shown in the following formula to obtain compound 7, Among them, each R 1 is methyl, ethyl, n-propyl, isopropyl, n-butyl or n-pentyl; The organic base is 2,6-diisopropylaniline and / or N,N-diisopropylethylamine; The solvent is a mixed solvent consisting of a nitrile solvent and an ether solvent.

11. The method for preparing compound 7 according to claim 10, characterized in that: The preparation method of compound 7 satisfies one or more of the following conditions: (1) In step S6, the nitrile solvent is acetonitrile; (2) In step S6, the ether solvent is one or more of tetrahydrofuran, dioxane and ethylene glycol dimethyl ether, such as tetrahydrofuran; (3) In step S6, the volume ratio of the ether solvent to the nitrile solvent is (0.2-1):1; for example (0.5-0.7):1, and for example 0.6:1; (4) In step S6, the molar volume ratio of the compound 6 to the solvent is 1 mmol:(0.1-0.3) L, for example, 1 mmol:(0.1-0.2) L, and for example, 1 mmol:0.16 L; (5) In step S6, the organic base is N,N-diisopropylethylamine; (6) In step S6, the molar ratio of the organic base to the compound 6 is (1-5):1, for example (1.5-3.5):1, and for example 3:1; (7) In step S6, the catalyst is a triazine compound, such as cyanuric chloride; (8) In step S6, the molar ratio of the catalyst to the compound 6 is (1-3):1, for example (1-2):1, and for example 1.5:1; (9) In step S6, the temperature of the ring-closure reaction is 5 to 25° C., for example, 10 to 18° C., and for example, 15° C.; (10) In step S6, the reaction is carried out under an inert atmosphere, such as a nitrogen atmosphere; (11) In step S6, the ring-closure reaction is carried out in a light-proof condition; (12) In step S6, the reaction time is 2 to 7 hours, for example 2 to 5 hours, and for example 2 hours; and (13) In step S6, after the ring-closure reaction is completed, the following post-treatment steps are further included: one or more of quenching, extraction, filtration, washing, drying, concentration and purification; the quenching reagent can be an aqueous solution of 3-amino-1,2-propanediol; the extraction solvent can be dichloromethane; the washing solution can be saturated brine; the drying desiccant can be anhydrous sodium sulfate; the purification can be column chromatography, and the eluent of the column chromatography can be dichloromethane and methanol, and the volume ratio of dichloromethane to methanol can be (120-500):

1.

12. The method for preparing compound 7 according to claim 10, characterized in that: The preparation method of compound 7 includes the preparation method of compound 6, the preparation method of compound 6 The method comprises the following steps S5: in a solvent, in the presence of an oxidant, subjecting compound 5 to an oxidation reaction as shown in the following formula to obtain compound 6, Preferably, the preparation method of compound 6 satisfies one or more of the following conditions: (1) In step S5, the solvent is a chlorinated alkane organic solvent, such as one or more of dichloromethane, 1,2-dichloroethane and chloroform, and another example is dichloromethane; (2) In step S5, the molar volume of the compound 5 and the solvent is (1-30) mmol:1 L, for example (10-20) mmol:1 L, and for example 15.6 mmol:1 L; (3) In step S5, the oxidant is selected from one or more of Jones reagent, Collins reagent, pyridinium chlorochromate, pyridinium dichromate, manganese dioxide, Dess-Martin periodinane, potassium permanganate, periodic acid, osmium tetroxide, 30% hydrogen peroxide, meta-chloroperbenzoic acid and tert-butyl hydroperoxide; for example, meta-chloroperbenzoic acid; (4) In step S5, the molar ratio of the oxidant to the compound 5 is (1-3):1, for example (1-2):1, and for example 1.2:1; (5) In step S5, the oxidant is added at a temperature of -10 to 0°C, for example, at -5°C; (6) In step S5, the oxidation reaction is carried out under light-proof conditions; (7) In step S5, the temperature of the oxidation reaction is -20 to 25°C, for example, 0 to 5°C, and for example, 0°C; (8) In step S5, the reaction time is 1 to 20 hours, for example 1 to 3 hours, and for example 2 hours; and (9) In step S5, after the oxidation reaction is completed, the following post-treatment steps are further included: one or more of quenching, concentration and purification; the quenching reagent can be anisole; the purification can be column chromatography, and the eluent of the column chromatography can be dichloromethane and methanol, and the volume ratio of dichloromethane to methanol can be (10-100):

1.

13. A method for preparing ADC, characterized in that: It includes the following steps: (1) Preparation method of compound 7: The method comprises the following steps S6: in a solvent, in the presence of a catalyst and an organic base, compound 6 undergoes a ring-closing reaction as shown in the following formula to obtain compound 7; The conditions and operations of the preparation method of compound 7 may be as described in any one of claims 1 to 8; (2) using compound 7 as a raw material to prepare the ADC compound; The ADC compounds include the following drugs: Anti-CD22-NMS249, huXBR1-402-PNU, NAV-001, NBE-002, SOT102, EGFR VEDVsPNU+EDVs40 mer, ADC-1, NBE-001, PNU-159682-P1C1TM or SOT107.

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