Crystalline forms of nitroxoline prodrugs, pharmaceutical compositions containing the same, and methods for their preparation and applications

Abstract

The present invention relates to crystalline forms of nitroxoline prodrugs, pharmaceutical compositions containing the same, and their preparation and application. The X-ray powder diffraction pattern of crystalline form A contains characteristic peaks at 5.74±0.2°, 6.78±0.2°, 10.86±0.2°, 13.54±0.2°, 16.70±0.2°, and 22.65±0.2°; the X-ray powder diffraction pattern of crystalline form B contains characteristic peaks at 5.44±0.2°, 10.90±0.2°, 14.09±0.2°, 16.17±0.2°, 17.92±0.2°, 20.66±0.2°, and 23.13±0.2°. Compared with nitroxoline, the crystalline forms A and B in the present application are less fouling, have lower requirements on equipment, and are more suitable for industrial production; the stability characteristics lead to more stability in quality control and drug efficacy for industrial production.

Description

[Technical Field]

[0001] The present invention relates to crystalline forms of nitroxoline prodrugs, pharmaceutical compositions containing same, and methods for their preparation and applications. [Background technology]

[0002] Nitroxoline is a commercially available antibacterial drug that has long been used to treat urinary tract infections. Recent discoveries have shown that nitroxoline is also highly effective in inhibiting angiogenesis and the growth and invasion of cancer cells, and it is now being developed for antitumor applications.

[0003] Human pharmacokinetic studies have shown that nitroxoline can be rapidly absorbed into the blood circulation. However, due to a significant first-pass effect on the drug in the liver, its biological half-life is very short (a single-arm, open-label, multicenter Phase II clinical trial conducted by Jiangsu Yahong Meditech Co., Ltd., China, showed that its half-life was 1.22–1.44 hours), thus requiring frequent dosing. To maintain continuous drug exposure, nitroxoline drugs are commonly prescribed for administration three times a day (TID) or four times a day (QID), which not only leads to economic losses and is unhelpful for patient compliance, but also, more seriously, increases the drug's sustained damage to the body. Meanwhile, nitroxoline's very poor water solubility often necessitates its preparation as an immediate-release formulation to improve solubility, substantially increasing production costs.

[0004] A prodrug is a compound obtained by chemical modification of an active drug, which is converted into the original drug by the action of enzymes in vivo to exert its effectiveness. Prodrugs have been widely used in drug research and development, and have been successfully developed for many different drugs with excellent efficacy in applications. The prodrug strategy can solve some of the deficiencies of active agents due to their own physical and chemical properties, such as: 1) eliminating the unpleasant odor of the drug; 2) increasing the blood concentration of the drug; 3) improving the lipid solubility or water solubility of the drug; 4) extending the duration of action of the drug; 5) changing the administration route of the drug, etc.

[0005] Drug polymorphism is not negligible and has become an important part in drug research and development and drug quality control. Research on drug polymorphism can promote the selection of biological activity of drug compounds, help improve the stability, solubility and other properties of drugs, and be beneficial to the development of drug preparations, drug storage, improvement of drug product quality, etc. It can also improve the bioavailability of compounds and enhance clinical efficacy. [Prior art documents] [Patent documents]

[0006] [Patent Document 1] WO2020 / 063824 [Non-patent literature]

[0007] [Non-Patent Document 1] Journal of Heterocyclic Chemistry, 1971, Vol. 8, p. 821 Summary of the Invention [Problem to be solved by the invention]

[0008] However, there are no reports relating to nitroxoline prodrugs and their crystalline forms in the prior art. [Means for solving the problem]

[0009] The technical problem solved by the present invention is to provide crystalline forms of nitroxoline prodrugs, pharmaceutical compositions containing same, and methods for their preparation and their applications.

[0010] The present inventors have studied a number of nitroxoline prodrugs (particularly the nitroxoline prodrug compounds described in the examples of WO2020 / 063824) and found that the nitroxoline prodrug ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate has superior water solubility, gastrointestinal stability, and pharmacokinetics to other compounds. Furthermore, the present inventors have found that the amorphous form of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate has low stability, which is not conducive to the preparation of pharmaceutical formulations. Based on this, through further research, the present inventors have obtained the crystalline forms of the present invention and their preparation methods.

[0011] The present invention provides crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate, the X-ray powder diffraction pattern of which, obtained using Cu-Kα radiation and expressed in 2θ angles, contains characteristic peaks at 5.74±0.2°, 6.78±0.2°, 10.86±0.2°, 13.54±0.2°, 16.70±0.2° and 22.65±0.2°.

[0012] In some preferred embodiments, the X-ray powder diffraction pattern of crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate was obtained by using Cu-Kα radiation and is expressed in 2θ angles and includes characteristic peaks at 5.74±0.2°, 6.78±0.2°, 8.25±0.2°, 10.86±0.2°, 13.54±0.2°, 14.92±0.2°, 16.70±0.2°, 17.23±0.2°, 18.10±0.2°, 19.56±0.2°, 22.65±0.2°, and 27.22±0.2°.

[0013] In some preferred embodiments, the X-ray powder diffraction pattern of crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate was obtained using Cu-Kα radiation, is shown in 2θ angles, and is as shown in FIG.

[0014] In some preferred embodiments, differential scanning calorimetry of crystalline form A shows an endothermic peak at 101.4°C.

[0015] In some preferred embodiments, differential scanning calorimetry of crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate shows an endothermic peak at 101.4°C.

[0016] The present invention also provides a method for preparing the aforementioned crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate, comprising the steps of: The method includes the steps of mixing a solution I containing ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and a normal solvent with an anti-solvent to precipitate a solid, and performing solid-liquid separation to obtain crystalline form A.

[0017] In the above preparation method, the positive solvent may be a mild solvent capable of dissolving ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate. The positive solvent is preferably one or more of an ester solvent, a C1-C6 alcohol solvent, a ketone solvent, a nitrile solvent, an ether solvent, and a lower halogenated alkane solvent.

[0018] Here, the ester solvent is preferably ethyl acetate.

[0019] Here, the C1 to C6 alcohol solvent is preferably one or more of methanol, ethanol, isopropanol and isobutanol, more preferably methanol and / or ethanol, and even more preferably methanol or ethanol.

[0020] Here, the ketone solvent is preferably one or more of acetone, methyl ethyl ketone and methyl isobutyl ketone, more preferably acetone or methyl isobutyl ketone, even more preferably acetone.

[0021] Here, the nitrile solvent is preferably acetonitrile.

[0022] Here, the ether solvent is preferably tetrahydrofuran and / or 1,4-dioxane, more preferably tetrahydrofuran.

[0023] Here, the lower halogenated alkane solvent is preferably dichloromethane.

[0024] Here, the positive solvent is more preferably an ester solvent, and here, the ester solvent is preferably a C1 to C5 ester solvent, more preferably ethyl acetate.

[0025] In the above preparation method, preferably after filtration, Solution I is obtained.

[0026] In the above preparation method, the anti-solvent may be an anti-solvent that can promote the crystallization or precipitation of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate in Solution I. The anti-solvent is preferably one or more of an ether solvent, an alcohol, a lower alkane solvent, and water, more preferably one or more of an ether solvent, a lower alkane solvent, and water.

[0027] Here, the ether solvent is preferably one or more of methyl tert-butyl ether, diethyl ether and petroleum ether, more preferably petroleum ether and / or methyl tert-butyl ether, even more preferably petroleum ether or methyl tert-butyl ether.

[0028] Here, the alcohol is preferably a C1 to C6 alcohol, more preferably isopropanol.

[0029] Here, the lower alkane solvent is preferably one or more of n-heptane, n-hexane and n-octane, more preferably n-heptane.

[0030] The anti-solvent here is more preferably an ether solvent, and the ether solvent here is preferably petroleum ether.

[0031] In particular, in the above preparation method, the positive solvent is more preferably an ester solvent, where the ester solvent is preferably a C1-C5 ester solvent, more preferably ethyl acetate; the anti-solvent is more preferably an ether solvent, where the ether solvent is preferably petroleum ether.

[0032] In the above preparation method, the volume ratio of the positive solvent to the anti-solvent is preferably 1:20 to 2:1, more preferably 1:10 to 1:2, for example, 1:5 or 1:9, even more preferably 0.3 to 0.5.

[0033] In the above preparation method, the mixing step may be accomplished by stirring.

[0034] In the above preparation method, the mixing temperature may be room temperature.

[0035] In the above preparation method, the temperature for precipitation of the solid may be room temperature.

[0036] The present invention also provides a second method for preparing the aforementioned crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate, comprising: Mixing Solution II containing ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and a solvent at room temperature to 50°C, and centrifuging to obtain Crystalline Form A; The solvent is one or more of a C1-C6 alcohol solvent, an ester solvent, an ether solvent, a lower alkane solvent, a lower halogenated alkane solvent, a ketone solvent, an aromatic hydrocarbon solvent, a nitrile solvent, dimethyl sulfoxide, and water, preferably one or more of a C1-C6 alcohol solvent, an ester solvent, an ether solvent, a lower alkane solvent, a ketone solvent, an aromatic hydrocarbon solvent, a nitrile solvent, dimethyl sulfoxide, and water.

[0037] Here, the C1 to C6 alcohol solvent is preferably one or more of methanol, ethanol and isopropanol, more preferably isopropanol and / or methanol.

[0038] Here, the ester solvent is preferably one or more of methyl acetate, ethyl acetate and isopropyl acetate, more preferably isopropyl acetate and / or ethyl acetate.

[0039] Here, the ether solvent is preferably one or more of methyl ethyl ether, diethyl ether, methyl isopropyl ether, methyl tert-butyl ether, cyclopentyl methyl ether, anisole, tetrahydrofuran, 2-methyltetrahydrofuran, and 1,4-dioxane, more preferably one or more of methyl tert-butyl ether, cyclopentyl methyl ether, anisole, tetrahydrofuran, 2-methyltetrahydrofuran, and 1,4-dioxane.

[0040] Here, the lower alkane solvent is preferably n-heptane.

[0041] Here, the lower halogenated alkane solvent is preferably dichloromethane.

[0042] Here, the ketone solvent is preferably one or more of methyl ethyl ketone, methyl isobutyl ketone and acetone.

[0043] Here, the aromatic hydrocarbon solvent is preferably toluene.

[0044] Here, the nitrile solvent is preferably acetonitrile.

[0045] In some preferred embodiments, in the second preparation method as described above, the solvent is a mixed solvent of C1-C6 alcohol and water, a mixed solvent of ether and lower alkane, a mixed solvent of ketone and lower alkane, a mixed solvent of ketone and ether, a mixed solvent of ester and C1-C6 alcohol, a mixed solvent of aromatic hydrocarbon and lower alkane, a mixed solvent of ketone and C1-C6 alcohol, or a mixed solvent of ether and ester, preferably a mixed solvent of C1-C6 alcohol and water, more preferably a mixed solvent of isopropanol and water or methanol and water; The volume ratio of the former to the latter in the mixed solvent is preferably 1:8 to 1:1, more preferably 1:4 to 1:2.

[0046] In some preferred embodiments, in the second preparation method as described above: Any crystalline or amorphous form of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate is suspended in a solvent at a temperature of 50°C, stirred, and centrifuged to obtain crystalline form A; The solvent is one or more of a C1-C6 alcohol solvent, an ester solvent, a ketone solvent, an ether solvent, a lower alkane solvent, an aromatic hydrocarbon solvent, a nitrile solvent, and water, preferably a mixed solvent of two solvents, more preferably a mixed solvent of a C1-C6 alcohol and water, a mixed solvent of a ketone and an ether, a mixed solvent of an ester and a C1-C6 alcohol, a mixed solvent of a ketone and a C1-C6 alcohol, a mixed solvent of an ether and an ester, a mixed solvent of an aromatic hydrocarbon and a lower alkane, a mixed solvent of an ether and a lower alkane, a mixed solvent of two ethers, or a mixed solvent of a nitrile and an ether, more preferably a mixed solvent of isopropanol / water, methyl isobutyl ketone / methyl tert-butyl ether, ethyl acetate / isopropanol, toluene / n-heptane, 2-methyltetrahydrofuran / n-heptane, butanone / isopropanol, acetonitrile / cyclopentyl methyl ether, anisole / isopropyl acetate, or 1,4-dioxane / cyclopentyl methyl ether; The volume ratio of the former to the latter in the mixed solvent is preferably 1:4 to 1:2.

[0047] The present invention also provides a third method for preparing the aforementioned crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate, comprising: A method comprising the steps of: subjecting a solution III comprising ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and a solvent to one to five cycles of 50°C → 5°C → 50°C until a solid precipitates, preferably three cycles, and performing solid-liquid separation to obtain crystalline form A; or heating solution III to 50°C to dissolve, hot filtering, cooling the filtrate to 5°C to -20°C, and performing solid-liquid separation to obtain crystalline form A; The solvent is preferably one or more, preferably one or two of a C1-C6 alcohol solvent, an ester solvent, a ketone solvent, an ether solvent, a lower alkane solvent, an aromatic hydrocarbon solvent, and water.

[0048] Here, the C1 to C6 alcohol solvent is preferably one or more of methanol, ethanol, and isopropanol.

[0049] Here, the ester solvent is preferably one or more of methyl acetate, ethyl acetate and isopropyl acetate.

[0050] Here, the ketone solvent is preferably one or more of methyl ethyl ketone, methyl propyl ketone and acetone.

[0051] Here, the ether solvent is preferably one or more of methyl ethyl ether, diethyl ether, methyl isopropyl ether, methyl tert-butyl ether, cyclopentyl methyl ether and anisole.

[0052] Here, the lower alkane solvent is preferably n-heptane.

[0053] Here, the aromatic hydrocarbon solvent is preferably toluene.

[0054] In some preferred embodiments, in the third preparation method as described above, the solvent is a mixed solvent of C1-C6 alcohol and water, a mixed solvent of C1-C6 alcohol and ether, a mixed solvent of ketone and ester, a mixed solvent of aromatic hydrocarbon and ester, or a mixed solvent of ketone and lower alkane; the C1-C6 alcohol solvent is preferably ethanol and / or isopropanol, and the ketone solvent is preferably butanone and / or methyl isobutyl ketone; the ester solvent is preferably isopropyl acetate; the ether solvent is preferably cyclopentyl methyl ether; the aromatic hydrocarbon solvent is preferably toluene; and the lower alkane solvent is preferably n-heptane; The volume ratio of the former to the latter in the mixed solvent is preferably 1:20 to 2:1, more preferably 1:10 to 1:2.

[0055] The present invention also provides a fourth method for preparing the aforementioned crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate, comprising: The method includes placing an opened first container containing any crystalline or amorphous form of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate solid into a second container containing a solvent, sealing the second container and allowing it to stand at room temperature, and collecting the product to obtain crystalline Form A when the solid becomes wet or a solid precipitate is observed; wherein the solvent is one or more of a C1-C6 alcohol solvent, an ether solvent, a ketone solvent, an ester solvent, an aromatic hydrocarbon solvent, dimethyl sulfoxide, and water.

[0056] Here, the C1 to C6 alcohol solvent is preferably one or more of methanol, ethanol, isopropanol and isobutanol, more preferably ethanol and / or isopropanol.

[0057] Here, the ether solvent is preferably one or more of methyl tert-butyl ether, tetrahydrofuran, 2-methyltetrahydrofuran, 1,4-dioxane and anisole, more preferably tetrahydrofuran.

[0058] Here, the ketone solvent is preferably one or more of acetone, methyl ethyl ketone and methyl isobutyl ketone, more preferably acetone.

[0059] Here, the ester solvent is preferably ethyl acetate.

[0060] Here, the aromatic hydrocarbon solvent is preferably toluene.

[0061] The present invention also provides a fifth method for preparing the aforementioned crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate, comprising: placing an opened first container containing Solution IV, which contains ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and an anti-solvent, into a second container containing an anti-solvent, sealing the second container, allowing it to stand at room temperature, and collecting the product to obtain crystalline Form A when a solid becomes wet or a solid precipitate is observed; The positive solvent is preferably one or more of a C1-C6 alcohol solvent, an ether solvent, and a ketone solvent; and the anti-solvent is one or more of a lower alkane solvent, an ether solvent, an alcohol solvent, and water.

[0062] In the case of a positive solvent, the C1-C6 alcohol solvent is preferably one or more of methanol, ethanol, isopropanol and isobutanol, more preferably ethanol.

[0063] In the case of positive solvent, the ether solvent is preferably one or more of tetrahydrofuran, 2-methyltetrahydrofuran and 1,4-dioxane, more preferably 1,4-dioxane.

[0064] In the case of a positive solvent, the ketone solvent is preferably one or more of acetone, methyl ethyl ketone and methyl isobutyl ketone, more preferably methyl isobutyl ketone.

[0065] In the case of the anti-solvent, the lower alkane solvent is preferably n-heptane.

[0066] In the case of the antisolvent, the ether solvent is preferably methyl tert-butyl ether.

[0067] In the case of the anti-solvent, the alcohol solvent is preferably isopropanol.

[0068] In some preferred embodiments, in the fifth preparation method as described above, the volume ratio of positive solvent to anti-solvent is preferably 1:20 to 2:1, more preferably 1:10 to 1:2, such as 1:8 or 1:4.

[0069] The present invention also provides a sixth method for preparing the aforementioned crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate, comprising: volatilizing a solution V comprising ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and a solvent at room temperature, and collecting the precipitated solid to obtain crystalline form A; The solvent is one or more of a C1-C6 alcohol solvent, a ketone solvent, an ester solvent, an ether solvent, a lower alkane solvent, an aromatic hydrocarbon solvent, a nitrile solvent, a lower halogenated alkane solvent, and water, preferably one or more of a C1-C6 alcohol solvent, a ketone solvent, an ether solvent, and a lower halogenated alkane solvent.

[0070] Here, the C1 to C6 alcohol solvent is preferably one or more of methanol, ethanol and isopropanol, more preferably isopropanol.

[0071] Here, the ketone solvent is preferably one or more of methyl ethyl ketone, methyl isobutyl ketone and acetone, more preferably methyl ethyl ketone and / or methyl isobutyl ketone.

[0072] Here, the ester solvent is preferably one or more of methyl acetate, ethyl acetate and isopropyl acetate.

[0073] Here, the ether solvent is preferably one or more of methyl ethyl ether, diethyl ether, methyl isopropyl ether, methyl tert-butyl ether, cyclopentyl methyl ether, anisole and 1,4-dioxane, more preferably 1,4-dioxane.

[0074] Here, the lower alkane solvent is preferably n-heptane.

[0075] Here, the aromatic hydrocarbon solvent is preferably toluene.

[0076] Here, the nitrile solvent is preferably acetonitrile.

[0077] Here, the lower halogenated alkane solvent is preferably dichloromethane.

[0078] The present invention also provides crystalline form B of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate, the X-ray powder diffraction pattern of which was obtained using Cu-Kα radiation and is expressed in 2θ angles, and contains characteristic peaks at 5.44±0.2°, 10.90±0.2°, 14.09±0.2°, 16.17±0.2°, 17.92±0.2°, 20.66±0.2°, and 23.13±0.2°.

[0079] In some preferred embodiments, the X-ray powder diffraction pattern of crystalline form B of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate is obtained by using Cu-Kα radiation and is expressed in 2θ angles and includes characteristic peaks at 5.44±0.2°, 6.19±0.2°, 10.90±0.2°, 14.09±0.2°, 14.88±0.2°, 16.17±0.2°, 17.92±0.2°, 20.66±0.2°, 21.69±0.2°, and 23.13±0.2°.

[0080] In some preferred embodiments, the X-ray powder diffraction pattern of crystalline form B of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate is obtained by using Cu-Kα radiation and is expressed in 2θ angles and includes characteristic peaks at 5.44±0.2°, 6.19±0.2°, 8.06±0.2°, 10.90±0.2°, 12.18±0.2°, 14.09±0.2°, 14.88±0.2°, 16.17±0.2°, 17.92±0.2°, 20.66±0.2°, 21.69±0.2°, 23.13±0.2°, 24.42±0.2°, and 26.03±0.2°.

[0081] In some preferred embodiments, the X-ray powder diffraction pattern of crystalline form B of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate was obtained using Cu-Kα radiation and is shown in 2θ angles as shown in FIG.

[0082] In some preferred embodiments, differential scanning calorimetry of crystalline form B of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate shows an endothermic peak at 101.5°C.

[0083] The present invention also provides a method for preparing the aforementioned crystalline form B of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate, comprising: The method comprises the steps of: mixing a solution A containing ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and a solvent with an anti-solvent to precipitate a solid; and performing solid-liquid separation to obtain a crystalline form B; the positive solvent is an ester solvent, the ester solvent is preferably one or more of methyl acetate, ethyl acetate and isopropyl acetate, more preferably ethyl acetate; The method is provided wherein the anti-solvent is an alkane solvent, preferably the alkane solvent is one or more of n-hexane, n-heptane and n-octane, more preferably n-heptane.

[0084] In some preferred embodiments, the volume ratio of positive solvent to anti-solvent is from 1:20 to 2:1, preferably from 1:10 to 1:2.

[0085] In some preferred embodiments, crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate is dissolved in a positive solvent to obtain solution A.

[0086] The present invention also provides a pharmaceutical composition comprising the aforementioned crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate or crystalline form B of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and auxiliary materials.

[0087] In the aforementioned pharmaceutical compositions, auxiliary materials generally refer to pharmaceutically acceptable carriers, diluents or excipients.

[0088] The present invention also provides use of the aforementioned crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate or crystalline form B of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate, or a pharmaceutical composition containing the same, in the preparation of a medicament for treating an infectious disease or cancer.

[0089] Here, the infectious disease is preferably a systemic infection, a reproductive system infection, or a urinary system infection.

[0090] Here, the cancer is preferably bladder cancer or prostate cancer.

[0091] In this specification, room temperature refers to 10 to 35°C, preferably 15 to 30°C.

[0092] As used herein, alcohol solvent refers to a type of organic compound formed by replacing one or more hydrogens in a hydrocarbon molecule with a hydroxyl group, typically a straight or branched chain alcohol compound having 1 to 6 carbons, such as one or more of methanol, ethanol, n-propanol, isopropanol, n-butanol, sec-butanol, primary butanol, and tert-butanol.

[0093] As used herein, ketone solvent refers to a compound in which a carbonyl group is bonded to two alkyl groups, typically a straight- or branched-chain ketone compound having 1 to 6 carbons, such as one or more of acetone, butanone (also known as methyl ethyl ketone), methyl isopropyl ketone, and methyl isobutyl ketone.

[0094] As used herein, ester solvent refers to a compound formed by esterifying an inorganic or organic acid with an alcohol and removing water, typically a straight or branched chain ester compound having 1 to 6 carbons, such as one or more of ethyl formate, methyl acetate, ethyl acetate, isopropyl acetate, and isobutyl acetate.

[0095] As used herein, ether solvent refers to a product in which hydrogen on the hydroxyl group of an alcohol or phenol is replaced by an alkyl group, typically a straight-chain, branched-chain, or cyclic ether compound having 1 to 6 carbons, such as one or more of diethyl ether, methyl tert-butyl ether, tetrahydrofuran, 2-methyltetrahydrofuran, 1,4-dioxane, and cyclopentyl methyl ether.

[0096] As used herein, a lower alkane solvent refers to a hydrocarbon that is liquid at room temperature, typically a straight or branched chain alkane or cycloalkane compound having 4 to 10 carbon atoms, such as one or more of n-pentane, n-heptane, n-octane, and cyclohexane.

[0097] As used herein, a lower halogenated alkane solvent refers to a hydrocarbon compound that typically has 1 to 10 carbon atoms and contains one or more of fluorine, chlorine, bromine, and iodine, which is liquid at room temperature, preferably a halogen-substituted straight-chain or branched-chain alkane compound having 1 to 6 carbon atoms, such as one or more of dichloromethane, dichloroethane, chloroform, bromoethane, and bromobutane.

[0098] As used herein, aromatic hydrocarbon solvent refers to a hydrocarbon containing a benzene ring structure in its molecule that is liquid at room temperature, such as toluene and / or xylene.

[0099] In this specification, a nitrile solvent refers to a compound containing a cyano group in the molecule, and typically refers to a straight-chain or branched-chain nitrile compound having 1 to 6 carbon atoms, preferably acetonitrile.

[0100] In this specification, the amorphous form of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate generally refers to an amorphous crude product of ((5-nitroquinolin)olin-8-yl)oxy)methyl-isobutyryl-L-prolinate, and may of course be pure ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate.

[0101] In this specification, solution I may be a solution prepared from any crystalline or amorphous form of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and a positive solvent.

[0102] In this specification, solution II may be a solution prepared from any crystalline or amorphous form of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and a solvent.

[0103] In this specification, solution III may be a solution prepared from any crystalline or amorphous form of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and a solvent.

[0104] In this specification, solution IV may be a solution prepared from any crystalline or amorphous form of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and a positive solvent.

[0105] In this specification, solution V may be a solution prepared from any crystalline or amorphous form of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and a solvent.

[0106] In this specification, solution A may be a solution prepared from any crystalline or amorphous form of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and a positive solvent.

[0107] As used herein, the positive solvent refers to a mild solvent capable of dissolving ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate. The positive solvent is preferably one or more of an ester solvent, a C1-C6 alcohol solvent, a ketone solvent, a nitrile solvent, an ether solvent, and a lower halogenated alkane solvent.

[0108] As used herein, the term "anti-solvent" refers to an anti-solvent that can promote the crystallization or precipitation of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate in solution. The anti-solvent is preferably one or more of an ether solvent, a lower alkane solvent, and water.

[0109] "Pharmaceutically acceptable," as used herein, means generally safe, free from biological or other undesirable toxicity, and useful for preparing pharmaceutical compositions acceptable for veterinary and human pharmaceutical use.

[0110] "Carrier," as used herein, refers to a diluent, adjuvant, or excipient with which a compound is administered. Pharmaceutically acceptable carriers can be liquids, such as water and oils, including those of petroleum, animal, vegetable, or synthetic origin, such as peanut oil, soybean oil, mineral oil, rapeseed oil, and the like. Pharmaceutically acceptable carriers can also be physiological saline, gum arabic, gelatin, starch paste, talc, keratin, silica gel, urea, and the like. In addition, auxiliary substances, stabilizers, thickeners, lubricants, coloring agents, and the like can also be used.

[0111] Those skilled in the art will understand that the pharmaceutical compositions of the present invention may be formulated into various formulations well known in the art according to the particular route of administration, such as oral preparations (powders, tablets, capsules, soft capsules, liquid medicines, syrups, elixirs, powders, sachets, granules), topical preparations (creams, ointments, lotions, gels, balms, plasters, pastes, sprays, aerosols, etc.), or injectable preparations (solutions, suspensions, emulsions). Among the pharmaceutical compositions according to the present invention, particular mention may be made of those suitable for oral, parenteral (intravenous or subcutaneous) or nasal administration, such as tablets or dragees, sublingual tablets, gelatin capsules, troches, suppositories, creams, ointments, skin gels, injections, drinkable suspensions, etc.

[0112] The pharmaceutical composition according to the present invention may contain pharmaceutically acceptable carriers, adjuvants or diluents, such as fillers, disintegrants, lubricants, suspending agents, binders, sweeteners, flavoring agents, preservatives, matrices, etc. Fillers include, for example, starch, pregelatinized starch, lactose, mannitol, chitin, microcrystalline cellulose, sucrose, etc. Disintegrants include, for example, starch, pregelatinized starch, microcrystalline cellulose, sodium carboxymethyl starch, cross-linked polyvinylpyrrole, low-substituted hydroxypropyl cellulose, cross-linked sodium carboxymethyl cellulose, etc. Lubricants include, for example, magnesium stearate, sodium lauryl sulfate, talc, silicon dioxide, etc. Suspending agents include, for example, polyvinylpyrrolidone, microcrystalline cellulose, sucrose, agar, hydroxypropyl methylcellulose, etc. Binders include, for example, starch slurry, polyvinylpyrrolidone, hydroxypropyl methylcellulose, etc. The compositions of the invention may be prepared by any method known in the art so as to provide quick, sustained or delayed release of the active ingredient after administration to the patient.

[0113] The pharmaceutical compositions of the present invention may be administered to an individual animal, e.g., a mammal (rat, mouse, domestic animal, or human) by a variety of routes, all routes of administration being contemplated, for example, the route of administration may be oral, topical, rectal, or by intravenous, intramuscular, transdermal, intrathecal, epidural, or intracerebroventricular injection.

[0114] The dosage of the active ingredient of the present invention may vary according to the condition and weight of the individual, the nature and severity of the disease, the drug form, the administration route, and the administration period, and may be selected by those skilled in the art. The dosage may vary from 1 to 1500 mg / day, and the drug may be administered daily in a single dose or in divided doses.

[0115] The positive improvement effect of the present invention is that in the prior art, nitroxoline API is dark yellow in color, easily stained, and the manufacturing process requires high industrial equipment, making it difficult to clean. Compared with nitroxoline, crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate or crystalline form B of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate prepared in the present invention is less susceptible to staining, has lower equipment requirements, and is more suitable for industrial production. Furthermore, crystalline form A and crystalline form B prepared in the present invention have stable properties, which further contribute to quality control in industrial production and stability in drug efficacy. Furthermore, compared with crystalline form B, crystalline form A has better stability, which further contributes to quality control in industrial production and stability in drug efficacy. [Brief explanation of the drawings]

[0116] [Figure 1] 1 is an XRPD pattern of crystalline form A prepared in Example 2. [Figure 2] 1 is a TGA / DSC spectrum of crystalline form A prepared in Example 2. [Figure 3] 1 is a DVS spectrum of crystalline form A prepared in Example 2. [Figure 4] 1 is a comparative XRPD pattern of crystalline form A prepared in Example 2 before and after DVS test. [Figure 5] 1 is a PLM photograph of crystalline form A prepared in Example 2. [Figure 6] 1 is an XRPD pattern of crystalline form B prepared in Example 11. [Figure 7] 1 is a TGA / DSC spectrum of crystalline form B prepared in Example 11. [Figure 8] 1 is a PLM photograph of crystalline form B prepared in Example 11. DETAILED DESCRIPTION OF THE INVENTION

[0117] The present invention will be described in more detail below with reference to examples, which are only used to illustrate the technical solutions of the present invention, but do not limit the essence and scope of the present invention.

[0118] In the following examples, the structures of the compounds were identified by nuclear magnetic resonance (NMR) and / or mass spectrometry (MS). The NMR shifts (δ) are 10 -6 The values ​​are given in units of (ppm). NMR determinations were performed using a Bruker 400M nuclear magnetic resonance spectrometer, and the solvent for the determinations was deuterated dimethyl sulfoxide (dimethyl sulfoxide-d6).

[0119] In the following examples, MS determination was performed using a liquid chromatography-mass spectrometer (Thermo, Ultimate3000 / MSQ); LC / MS determination was performed using an Agilent 6250 liquid chromatography-mass spectrometer; 200-300 mesh Yantai Huanghai silica gel was generally used as the support for silica gel column chromatography.

[0120] In the following examples, nitroxoline and L-Boc proline were purchased from Accela Chemical Reagent Co., Ltd.

[0121] Laboratory Equipment 1. X-ray powder diffraction (XRPD) Equipment model:Empyrean; X-rays: Cu, kα, Kα1(Å): 1.540598; Kα2(Å): 1.544426; Kα2 / Kα1 intensity ratio: 0.50; X-ray tube settings: Voltage: 45 kV, Current: 40 mA Scanning style: Continuous Scanning range (2 theta degrees): 3~40 Scan time per step (seconds): 17.8 Scan step size (2°): 0.0167 Test time: 5 minutes 30 seconds

[0122] 2.Thermogravimetric analysis (TGA) Device model: TA Q5000 / Discovery 5500 Method: Linear heating Temperature range: Room temperature to 350°C Scan rate: 10°C / min Shielding gas: Nitrogen Sample amount: 1 to 3 mg

[0123] 3. Differential Scanning Calorimetry (DSC) Equipment model: TA Q2000 / Discovery 2500 Method: Linear heating Temperature range: 25℃~260℃ Scan rate: 10°C / min Shielding gas: Nitrogen Sample amount: 1 to 3 mg

[0124] 4. Modulated Differential Scanning Calorimetry (mDSC) Testing method: conventional mDSC Amplitude (℃): 1.0 Modulate Time (sec): 60 Scan rate (℃ / min): 3.0 Shielding gas: Nitrogen Sample amount: 1 to 3 mg

[0125] 5. Dynamic Vapor Sorption (DVS) DVS curves were obtained on a DVS Intrinsic from SMS (Surface Measurement Systems). Relative humidity at 25°C was corrected for the deliquescence points of LiCl, Mg(NO3)2, and KCl. Test temperature: 25℃ Sample amount: 10 to 20 mg Shielding gas and flow rate: Nitrogen, 200 mL / min dm / dt:0.002% / min; Minimum dm / dt equilibration time: 10 minutes Maximum equilibration time: 180 minutes RH range: 0%RH~95%RH~0%RH RH gradient: 10%

[0126] 6. Polarized Light Microscopy (PLM) Acquisition was performed with an Axio Scope A1 microscope at room temperature. [Example]

[0127] Preparation of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate

[0128] [ka]

[0129] Step 1: Preparation of 5-nitro-8-chloromethoxyquinoline Aqueous sodium bicarbonate (60 mL, 3.5 mol / L) and tetrabutylammonium hydrogen sulfate (1.78 g, 5.24 mmol) were added to a solution of nitroxoline (10.00 g, 52.59 mmol) in dichloromethane (100 mL) at room temperature. The reaction mixture was stirred at room temperature for 20 minutes. Chloromethyl chlorosulfonate (10.42 g, 63.15 mmol) was added dropwise to the reaction mixture, which was then stirred at room temperature for 16 hours. The reaction mixture was filtered, and the organic phase was separated and washed successively with saturated potassium carbonate solution and saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: dichloromethane) to give 5-nitro-8-chloromethoxyquinoline (2.5 g, 20% yield).

[0130] Step 2: Preparation of 1-(tert-butyl) 2-(((5-nitroquinolin-8-yl)oxy)methyl) (S)-pyrrolidine-1,2-dicarboxylate 8-Chloromethoxy-5-nitroquinoline (1.5 g, 6.3 mmol) and L-Boc proline (2.02 g, 9.4 mmol) were dissolved in 15 mL of DMF at room temperature, and potassium carbonate (1.73 g, 12.6 mmol) was added. The reaction solution was stirred at room temperature for 3 hours, 70 mL of water was added, and the reaction solution was extracted with ethyl acetate (50 mL × 2). The organic phase was washed with saturated brine, dried over anhydrous sodium sulfate, filtered, and concentrated under reduced pressure to obtain the crude product. The crude product was purified by silica gel column chromatography (PE:EA = 1:1) to obtain the product 1-(tert-butyl) 2-(((5-nitroquinolin-8-yl)oxy)methyl)(S)-pyrrolidine-1,2-dicarboxylate (2.6 g, 98% yield).

[0131] Step 3: Preparation of ((5-nitroquinolin-8-yl)oxy)methyl L-prolinate hydrochloride 1-(Tert-butyl) 2-(((5-nitroquinolin-8-yl)oxy)methyl)(S)-pyrrolidine-1,2-dicarboxylate (2.6 g, 6.4 mmol) was placed in HCl / dioxane (30 mL) at 0° C. and stirred at room temperature for 20 minutes. The reaction solution was concentrated under reduced pressure to give the product ((5-nitroquinolin-8-yl)oxy)methyl L-prolinate hydrochloride (2.3 g, 97% yield).

[0132] Step 4: Preparation of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate ((5-Nitroquinolin-8-yl)oxy)methyl L-prolinate hydrochloride (150 mg, 0.43 mmol) was added to anhydrous dichloromethane (5 mL) at room temperature. After cooling in an ice bath, isobutyryl chloride (103.7 mg, 0.90 mmol) was added, and triethylamine (180 mg, 1.72 mmol) was added dropwise slowly between 0 and 10 °C, followed by stirring for 20 min. The reaction solution was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (PE:EA = 1:1 to 0:1) to obtain ((5-nitroquinolin-8-yl)oxy)methyl isobutyryl L-prolinate (85 mg, 49.3% yield). 1 H-NMR (400 MHz, dimethyl sulfoxide-d6): δ: 9.05 (d, J = 4.0 Hz,1H), 9.00 (d, J = 8.8 Hz, 1H), 8.56 (d, J=8.8 Hz, 1H), 7.89-7.86 (dd, J = 4.0 Hz,8.8Hz,1H),7.55 (d, J=8.8 Hz, 1H), 6.24-6.11 (m, 2H), 4.36-4.33 (m, 1H), 3.59-3.68 (m, 2H), 2.51-2.66(m, 1H), 2.14~2.19(m, 1H),1.92-1.85(m, 2H), 1.83-1.78(m, 1H) ,0.95 (d, J=6.8 Hz .3H), 0.89 (d, J=6.8 Hz .3H). MS calculated: 387.3; MS found: 388.2 [M+H] + . [Example]

[0133] Preparation Example 1 of Crystalline Form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate 260 g of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate (purity 98.5%) prepared according to Example 1 was dissolved in 800 mL of ethyl acetate and filtered through diatomaceous earth to obtain a clear filtrate. The filtrate was concentrated to 400 mL under reduced pressure, and petroleum ether (100 mL) was added with stirring. Stirring was continued for 20 minutes at 20-30°C until small particles precipitated from the solution. Petroleum ether (900 mL) was slowly added dropwise, and a large amount of pale yellow solid precipitated. Stirring was continued for 2 hours at 20-30°C, and the reaction solution was filtered to obtain the wet product (300 g) as a pale yellow solid. The wet product was dried under reduced pressure at 30-40°C to obtain a solid (240 g, yield 92.3%, LCMS purity: 99.6%).

[0134] The X-ray powder diffraction (XRPD) pattern of the solid is shown in Figure 1, and its XRPD diffraction peak data is shown in Table 1 below. The TGA / DSC spectrum is shown in Figure 2. The TGA result shows that the solid has a mass loss of 0.8% when heated to 150°C; the DSC result shows that the solid has an endothermic peak at 101.4°C (onset temperature). The DVS spectrum is shown in Figure 3, which shows that the vapor sorption of the solid at 25°C / 80% RH is 0.17%, indicating that the sample has almost no hygroscopicity. The comparative XRPD patterns before and after the DVS test are shown in Figure 4, which shows that the crystalline form of the solid does not change before and after the DVS test. The PLM result is shown in Figure 5, which shows that the solid is composed of irregular crystalline particles. This crystalline form is defined as Form A.

[0135] [Table 1A]

[0136] [Table 1B] [Example]

[0137] Preparation Example 2 of Crystalline Form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate Approximately 30 mg aliquots of the solid prepared according to Example 2 were weighed and placed in glass vials, and 0.5 mL of the solvent listed in Table 2 below was added to each. The resulting suspension was left under magnetic stirring (approximately 1000 rpm) at room temperature for 6 days, and then centrifuged at 10,000 rpm to obtain a solid. XRPD testing identified it as the same as the crystals prepared in Example 2, and both were crystalline form A.

[0138] [Table 2] [Example]

[0139] Preparation Example 3 of Crystalline Form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate Approximately 30 mg aliquots of the solid prepared according to Example 2 were weighed and placed in glass vials, and 0.5 mL of the solvent listed in Table 3 below was added to each. The resulting suspension was placed under magnetic stirring (approximately 1000 rpm) at 50°C for 3 days, and then centrifuged at 10,000 rpm to obtain a solid. XRPD testing identified it as the same as the crystals obtained in Example 2, and both were crystalline form A.

[0140] [Table 3] [Example]

[0141] Preparation Example 4 of Crystalline Form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate Approximately 30 mg aliquots of the solid prepared according to Example 2 were weighed and placed in glass vials, and 0.5 mL of each of the solvents listed in Table 4 below was added. The resulting suspensions were subjected to temperature cycling (50 → 5°C, 0.1°C / min, 5 → 50°C, 0.375°C / min, 3 cycles) under stirring, and then centrifuged at 10,000 rpm to obtain solids. XRPD testing identified them as the same as the crystals obtained in Example 2, and both were crystalline form A.

[0142] [Table 4] [Example]

[0143] Preparation Example 5 of Crystalline Form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate Approximately 30 mg of the solid prepared according to Example 2 was weighed into a glass vial, and 0.5 mL of the solvent listed in Table 5 below was added. The solution was stirred at 50°C for 2 hours and filtered (PTFE filter membrane with a pore size of 0.45 μm; manufacturer: Titan Chemical Co.; model: Syringe filter hydrophobic polytetrafluoroethylene (PTFE) 0.45 μm 13 mm). The filtrate was then placed in a biochemical incubator (manufacturer: Shanghai Yiheng Scientific Instrument Co., Ltd.; model: BPC-70F), and the temperature was reduced from 50°C to 5°C at a cooling rate of 0.1°C / min. When the solution was still clear, the clear sample was transferred to -20°C and allowed to stand overnight to obtain a solid, which was removed using a spatula. XRPD testing identified it as the same crystals obtained in Example 2, and both were crystalline form A.

[0144] [Table 5] [Example]

[0145] Preparation Example 6 of Crystalline Form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate Approximately 30 mg aliquots of the solid prepared according to Example 2 were weighed and placed in glass vials, and 1.0 mL of each of the normal solvents listed in Table 6 below was added, and the solutions were filtered. The antisolvents listed in Table 6 below were added dropwise to the clear solution at room temperature under stirring until solids precipitated. After approximately 9 mL of the antisolvent was added, if no solids precipitated, the addition was stopped, and the solution was centrifuged at 10,000 rpm to obtain the solids. XRPD testing identified them as the same crystals obtained in Example 2, and both were crystalline form A.

[0146] [Table 6] [Example]

[0147] Preparation Example 7 of Crystalline Form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate An aliquot of approximately 30 mg of the solid prepared according to Example 2 was weighed and placed in a 3 mL glass vial, and approximately 4 mL of the solvent listed in Table 7 below was added to another 20 mL glass vial. The opened 3 mL glass vial was placed in the 20 mL glass vial, and the 20 mL glass vial was then sealed. The 20 mL glass vial was left at room temperature until the solid surface became wet, or left for 7 days, after which an XRPD test was performed. The XRPD test identified it as the same crystals as those obtained in Example 2, and both were crystalline form A.

[0148] [Table 7] [Example]

[0149] Preparation Example 8 of Crystalline Form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate An approximately 30 mg aliquot of the solid prepared according to Example 2 was weighed and dissolved in 0.5-1.0 mL of the antisolvent listed in Table 8 below, and the solution was filtered. The resulting filtrate was transferred to a 3 mL glass vial, and approximately 4 mL of the antisolvent listed in Table 8 below was added to another 20 mL glass vial. The opened 3 mL glass vial containing the filtrate was placed in a 20 mL glass vial, which was then sealed and allowed to stand at room temperature. When solid precipitation was observed, the solid was collected and subjected to XRPD analysis. XRPD analysis identified it as the same crystals obtained in Example 2, and both were crystalline form A.

[0150] [Table 8] [Example]

[0151] Preparation Example 9 of Crystalline Form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate Approximately 30 mg of the solid prepared according to Example 2 was weighed into a 3 mL glass vial, and 1.0 to 2.0 mL of the solvent listed in Table 9 below was added. The solution was shaken and filtered (PTFE filter membrane with a pore size of 0.45 μm; manufacturer: Titan Chemical Co.; model: Syringe filter hydrophobic polytetrafluoroethylene (PTFE) 0.45 μm 13 mm), and the filtrate was collected. The glass vial containing the clear solution was sealed with parafilm, several small holes were drilled on it, and the glass vial was left at room temperature to allow slow evaporation. When solid precipitation was observed, the resulting solid was collected and subjected to XRPD analysis. XRPD analysis identified it as the same crystals obtained in Example 2, and both were crystalline form A.

[0152] [Table 9] [Example]

[0153] Preparation of crystalline form B of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate 30.5 mg of the solid prepared in Example 2 was dissolved in 1 mL of ethyl acetate. After filtration, n-heptane was slowly added, and a solid precipitated when 7 mL was added. The solid was obtained by filtration and drying.

[0154] The X-ray powder diffraction (XRPD) pattern of the solid is shown in Figure 6, and the XRPD diffraction peak data is shown in Table 10 below. The TGA / DSC spectrum is shown in Figure 7 and shows that the solid has a mass loss of 1.2% when heated to 150°C, with an endothermic peak at 101.5°C (onset temperature). PLM results show that the sample is needle-like, approximately 100 μm in length (see Figure 8). This crystalline form is defined as crystalline form B.

[0155] [Table 10]

[0156] Test Example 1: Determination of water solubility of the compound ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate obtained in Example 1 The compound obtained in Example 1 can slowly release the active ingredient nitroxoline after entering the human body, which can simultaneously inhibit methionine aminopeptidase MetAP2 and silent mating signaling regulation 2 homolog in vascular endothelial cells, exerting a synergistic inhibitory effect on tumor angiogenesis. Meanwhile, nitroxoline also has an inhibitory effect on tumor cell proliferation. Furthermore, the released active ingredient nitroxoline exerts a bacteriostatic effect by inhibiting methionine aminopeptidase MetAP in bacteria.

[0157] The present inventors first conducted a study on the water solubility of nitroxoline and the compound obtained in Example 1.

[0158] Experimental equipment: 96-well filter plate (MSHVN4510 or MSHVN4550, Millipore); electronic digital vortex (MS3 Digital, IKA); water-circulating multipurpose vacuum pump (SHB-III, Zhengzhou Greatwall Science, Industry and Trade Co., Ltd.); balance (XSLT05, METTLER TOLEDO); ThermoMixer Comfort (Eppendorf AG 22331 Hamburg); liquid chromatography (LC-30AD, Shimadzu); mass spectrometer (API4000, Applied); sampler (CTC Anylytics AG System). Nitroxoline was synthesized by Wisdom Pharmaceutical Co., Ltd. according to the method disclosed in Journal of Heterocyclic Chemistry, 1971, Vol. 8, p. 821.

[0159] Experimental procedure: 500 μL of phosphate buffer (pH = 1.2, 4.5, 6.8, or 7.4) was added to a glass vial, and 2 mg of compound powder was added. The vial was sealed with a cap and placed on a vortex mixer (VORTEX-GENIE2) to mix thoroughly at room temperature for 24 hours. The solution was then vacuum filtered, and the filtrate was treated to determine the compound concentration by LC / MS / MS.

[0160] The solubility results of the compounds obtained in Example 1 are shown in Table 11 below.

[0161] [Table 11]

[0162] Conclusion: Compared with nitroxoline (5-nitro-8-hydroxyquinoline), the water solubility of the compound obtained in Example 1 is several times higher in a pH 7.4 buffer solution, and its water solubility remains almost unchanged at different pH levels, which can be considered essentially unchanged. This feature is particularly important in the development of drug formulations.

[0163] Test Example 2: Determination of the stability of the compound obtained in Example 1 in liver microsomes and plasma The compound obtained in Example 1 is expected to be degraded to nitroxoline in vivo, resulting in anticancer effects. Liver microsomal enzymes and plasma metabolic enzymes are important pathways for compound metabolism in vivo. Therefore, in vitro experiments were conducted to determine the stability of the compound obtained in Example 1 in liver microsomes and plasma.

[0164] 1. Stability determination in liver microsomes Experimental equipment: thermostatic oscillator (SHA-B, Guohua Instrument); centrifuge (5810R, Eppendorf); mass spectrometer (API4000, Applied); liquid chromatograph (LC-30AD, Shimadzu); sampler (CTC Analytics AGSystem, CTC).

[0165] Experimental Procedure: A coenzyme-free reaction solution was prepared by adding 25 μg / mL alamethicin (Aldrich Reagents), 5 mM magnesium chloride, and 0.5 mg / mL microsomes (XENOTECH) to 100 mM phosphate buffer. To a portion of the reaction solution, 1 mM reduced nicotinamide adenine dinucleotide phosphate (Aldrich Reagents) and 5 mM uridine diphosphate glucuronic acid (Aldrich Reagents) were added to prepare a coenzyme-free reaction solution. The working solution of the compound obtained in Example 1 was then added to the two reaction solutions to a final compound concentration of 2 μM. Immediately after thorough mixing, 50 μL of the solution was collected as a 0-minute sample, and another 50 μL was collected 30 minutes after incubating the remaining samples at 37°C. Proteins in all collected samples were immediately precipitated, and the supernatants were collected by centrifugation. The compound concentrations in the supernatants were determined by LC / MS / MS.

[0166] The stability results of the compound obtained in Example 1 in microsomes are shown in Table 12 below.

[0167] [Table 12]

[0168] Conclusion: The above data indicate that the compound obtained in Example 1 can be rapidly converted to nitroxoline after entering the body, which reduces the possibility of unnecessary biological toxicity, and the compound has advantages and properties for drug development.

[0169] Test Example 3: Pharmacokinetic assay of the compound obtained in Example 1 in rats In this experiment, nitroxoline and the compound obtained in Example 1 were administered to rats by single intravenous or oral administration, and then the changes in the concentration of compound nitroxoline in rat plasma were studied, and the in vivo pharmacokinetic behavior of nitroxoline and the compound obtained in Example 1 in rats was evaluated.

[0170] 1. Laboratory equipment Tandem quadrupole mass spectrometer (API4000, Applied Biosystems, USA), liquid chromatography (1200, Agilent), autosampler (CTC Analytics HTC PAL), Analyst v1.6.2, Applied Biosystems, USA, refrigerated centrifuge (1-15PK, Sigma), vortex (VX-III, Beijing Targin Technology Co., Ltd.).

[0171] 2. Pharmacokinetics Experiments Male SD rats (Beijing Vital River Laboratory Animal Technology Co., Ltd., Experimental Animal Manufacturing Permit Number: SCXK(Beijing)2016-0006, Experimental Animal Certification Number: 11400700325643), weighing 180-250 g and 6-7 weeks old, were used in groups of three. They were fasted overnight before drug administration, allowed free access to water, and given food 4 hours after drug administration. The compound to be tested was placed in an EP tube, and 1.017 mL of dimethyl sulfoxide, 2.035 mL of Solutol®, and sterile water for injection (volume ratio of the three was 1:2:17, v:v:v) were added. The EP tube was sonicated for 20 minutes to completely dissolve the compound (compound formulation concentration: 0.005 mmol / mL). The intravenous dose was 0.01 mmol / kg, and the oral dose was 0.1 mmol / kg. 0.3 ml of whole blood was collected from the retro-orbital venous plexus before drug administration (hour 0) and at 0.0833, 0.25, 0.5, 1, 2, 4, 6, 8, 10, 24, 28, 32, and 48 hours after drug administration (sampling times were adjusted accordingly) and placed in a centrifuge tube containing EDTA-K2 (Aldrich Reagents) for anticoagulation. After sample collection, the tube was placed in crushed ice. The tube was centrifuged at 5000 rpm for 5 minutes within 0.5 hours of sample collection. Total washed plasma was isolated and placed in a separate washed centrifuge tube. Stabilizing solution was added at a ratio of 100:3 (plasma / stabilizing solution, v / v). The tube was then placed in a -20°C refrigerator until testing.

[0172] Preparation of stabilization solution: 200 mg of vitamin C (Aldrich Reagents) was dissolved in 8 mL of physiological saline, and then 2 mL of formic acid was added and mixed thoroughly.

[0173] 3. Determining Sample Concentration Standard curve: A series of working solutions for the standard curve was prepared, and 5 μL of each solution was added to 50 μL of blank rat plasma. 150 μL of internal standard working solution (2 ng / mL diphenhydramine (Aldrich Reagents) solution in acetonitrile) was added, and the resulting solution was vortexed for 1 minute. After centrifugation at 12,000 rpm for 10 minutes at 4°C, 100 μL of the supernatant was collected in a sample tube, and 10 μL was injected into the LC / MS system for analysis.

[0174] Sample to be tested: 50 μL of the plasma sample to be tested was added with 5 μL of diluted working solution, followed by 150 μL of internal standard working solution (2 ng / mL diphenhydramine solution in acetonitrile), and the resulting solution was vortexed for 1 minute. After centrifugation at 12,000 rpm for 10 minutes at 4°C, 100 μL of the supernatant was collected in a sample tube, and 10 μL was injected into the LC / MS system for determination. Pharmacokinetic parameters were calculated using WinNonlin V6.2 non-compartmental model.

[0175] The test results are shown in Tables 13 to 14 below.

[0176] [Table 13]

[0177] [Table 14]

[0178] Conclusion: Compared with nitroxoline, the compound obtained in Example 1 has significantly improved absorption or half-life in rats. As a result, the drug molecule has good compliance improvement in terms of reducing the dosage or administration frequency.

[0179] Test Example 4: Pharmacokinetic assay of the compound obtained in Example 1 in dogs Nitroxoline is mainly metabolized in the liver by phase II metabolism, which has a high metabolic rate, and therefore has a short in vivo half-life. In this experiment, nitroxoline and the compound obtained in Example 1 were administered intravenously or orally to dogs in a single dose, and the changes in the concentration of the nitroxoline compound in dog plasma were investigated, and the in vivo pharmacokinetic behavior of nitroxoline and the compound obtained in Example 1 was evaluated.

[0180] 1. Laboratory equipment Tandem quadrupole mass spectrometer (API5500, Applied Biosystems, USA), liquid chromatography (1200, Agilent), autosampler (CTC Analytics HTC PAL), Analyst v1.6.2, Applied Biosystems, USA.

[0181] 2. Pharmacokinetics Experiments Male beagles (Beijing Marshall Bioresources Co., Ltd., Laboratory Animal Manufacturing Permit Number: SCXK(Beijing)2016-0001, Laboratory Animal Certification Number: 11400600001728), weighing 10-13 kg and aged 20-22 months, were used in groups of three. They were fasted overnight before drug administration, allowed free access to water, and were given food 4 hours after drug administration. The compound to be tested was placed in an EP tube, and dimethyl sulfoxide, Solutol®, and sterile water for injection (volume ratio of the three was 1:2:17, v:v:v) were added. The EP tube was sonicated for 20 minutes to completely dissolve the compound (compound formulation concentration: 0.005 mmol / mL). The intravenous dose was 0.01 mmol / kg, and the oral dose was 0.1 mmol / kg. 0.3 ml of whole blood was collected from the jugular vein before drug administration (time 0) and at 0.0833, 0.25, 0.5, 0.75, 1, 2, 4, 6, 8, 10, and 12 hours after drug administration (sampling times were adjusted accordingly) into centrifuge tubes containing EDTA-K2 (Aldrich Reagents) for anticoagulation. After sample collection, the tubes were placed in crushed ice. The tubes were centrifuged at 1530 g for 10 minutes within 0.5 hours of sample collection. Total washed plasma was isolated, placed in separate washed centrifuge tubes, and placed in a -20°C refrigerator until testing.

[0182] 3. Determining Sample Concentration A series of solutions for the standard curve were prepared. To 10 μL of the standard curve solution and sample, 1000 μL of internal standard working solution (5 ng / mL verapamil (Aldrich Reagents) solution, 50 ng / mL glibenclamide (Aldrich Reagents) and 50 ng / mL diclofenac (Aldrich Reagents) in acetonitrile) was added, and the resulting solution was vortexed for 5 minutes. After centrifugation at 4°C and 3700 rpm for 10 minutes, 60 μL of the supernatant was collected in a sample tube and thoroughly mixed with 120 μL of water. 10 μL of the mixed solution was injected into the LC / MS system for determination. Pharmacokinetic parameters were calculated using WinNonlin V6.2 non-compartmental model.

[0183] The test results are shown in Tables 15 to 18 below.

[0184] [Table 15]

[0185] [Table 16]

[0186] [Table 17]

[0187] [Table 18]

[0188] Conclusion: Compared with nitroxoline, it can be seen from the data that the compound obtained in Example 1 has good absorption in beagle dogs, indicating that the administered dosage of drug molecules can be effectively reduced by using prodrug molecules.

[0189] Test Example 5 Stability test of crystalline form A of the present invention The crystalline forms obtained in Examples 2 to 10 were packaged in double-layer PE bags, aluminum foil bags, and cardboard tubes, sealed, and stored at 40°C and 75% relative humidity; 25°C and 60% relative humidity; and 5±3°C for 6 months. Test items (appearance, moisture, related substance 1, related substance 2, content) were tested, and the specific test methods were as follows. The results are shown in Table 19. It can be seen from Table 19 that the impurity content of crystalline form A was relatively low and did not increase substantially under each condition, indicating that crystalline form A has good stability under long-term conditions.

[0190] 1. Water 1.1. Equipment and Facilities Moisture meter electronic balance 1.2 Reagents Karl Fischer Titration: Analytical Grade Absolute methanol: analytical grade 1.3 Detection methods Pretitration was performed by pumping approximately 50 mL of methanol into the titration beaker; Approximately 1.0 g of the test sample was accurately weighed and placed in a titration beaker, stirred to dissolve, and the water content of the test sample was titrated; The mass of test sample added, the concentration of the Karl Fischer titer and the volume of Karl Fischer titer consumed during the determination were recorded; the moisture content of a second test sample was determined using the same method. 1.4 Calculation formula

[0191]

number

[0192] V - volume of Karl Fischer titration consumed for the test sample, mL; F - equivalent mass of water per mL of Karl Fischer titration, mg / mL; W - weighed sample amount of test sample, g.

[0193] 2 Related Substances 1 2.1 Chromatography conditions: High-performance liquid chromatography (HPLC) Column: Waters XBridge C18 4.6 x 150 mm, 5 μm Detector: UV or equivalent detector Wavelength: 210nm Column temperature: 35℃ Flow rate: 1.0mL / min Injection volume: 10μL Needle cleaning: acetonitrile Mobile phase gradient:

[0194] [Table 19]

[0195] 2.2 Reagents and reference materials Acetonitrile (chromatography grade) Phosphoric acid (chromatography grade) Dichloromethane (chromatography grade) 5-nitro-8-hydroxyquinoline reference substance: Same structural formula as 5-nitro-8-hydroxyquinoline Reference substance of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate: Same structural formula as ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate Impurity 1 Reference Substance:

[0196] [ka]

[0197] Impurity 2 Reference Substance:

[0198] [ka]

[0199] 2.3 Preparation of solutions 1) Mobile phase A: 0.1% phosphoric acid aqueous solution 1.0 mL of phosphoric acid was accurately measured and added to 1000 mL of ultrapure water and mixed thoroughly. 2) Mobile phase B: Acetonitrile 3) Diluent (blank solution): Acetonitrile 4) System compatibility: Approximately 25 mg of 5-nitro-8-hydroxyquinoline reference material and approximately 25 mg of Impurity 2 reference material were accurately weighed into a 50 mL volumetric flask. Diluent was added to dissolve the materials and the solution was diluted to the mark. The solution was shaken well and labeled as Solution 1. Approximately 5 mg of Impurity 1 reference material was weighed and placed in a 50 mL volumetric flask. The material was completely dissolved using dichloromethane that had been preheated to 40°C in a water bath, and the solution was diluted to the mark. The solution was shaken well and labeled as Solution 2. Approximately 50 mg of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate reference material was accurately weighed and placed in a 100 mL volumetric flask. An appropriate amount of diluent was added to dissolve the material, and then 1.0 mL of Solution 1 and 5.0 mL of Solution 2 were accurately added, and the solution was diluted to the mark by adding diluent. The solution was shaken well and labeled as the system suitability solution. 5) Sensitivity solution 1.0 mL of the system suitability solution was accurately measured and placed in a 100 mL volumetric flask. Diluent was added to dilute the solution to the mark and the solution was shaken well. 1.0 mL of the above solution was accurately measured and placed in a 20 mL volumetric flask. Diluent was added to dilute the solution to the mark and the solution was shaken well. 6) Test sample solution Approximately 25 mg of test sample was accurately weighed and placed in a 50 mL volumetric flask. Diluent was added to dissolve the substance, the solution was diluted to the mark, and the solution was shaken well. 2.4 Calculation: Blanks subtracted

[0200]

number

[0201] A U - Peak area of ​​impurities in the test sample solution; A T - total peak area of ​​the test sample solution. 2) Total impurities (%) = Σ single impurity

[0202] [Table 20]

[0203] 3 Related Substance 2 (D-Isobutyrylproline) 3.1 Chromatography conditions: High-performance liquid chromatography (HPLC) Column: Synergi Hydro RP 4.6 x 250 mm, 4 μm Detector: UV or equivalent detector Wavelength: 210nm Column temperature: 35℃ Flow rate: 1.0mL / min Injection volume: 5μL Needle cleaning: acetonitrile Mobile phase gradient:

[0204] [Table 21]

[0205] 3.2 Reagents and reference materials Acetonitrile (chromatography grade) Phosphoric acid (chromatography grade) ultra pure water Reference substance for Related Substance 2: D-Isobutyrylproline 3.3 Preparation of solutions 1) Mobile phase A: 0.1% phosphoric acid aqueous solution 2) Mobile phase B: Acetonitrile 3) Diluent (blank solution): 50% acetonitrile aqueous solution 4) Reference substance solution Approximately 40 mg of Related Substance 2 reference substance was accurately weighed and placed in a 100 mL volumetric flask. Diluent was added to dissolve the substance, the solution was diluted to the mark, and the solution was shaken well. 1.0 mL of the above solution was accurately measured and placed in a 100 mL volumetric flask. Diluent was added to dilute the solution to the mark, and the solution was shaken well. Two solutions were prepared in parallel and designated as RS1 / RS2, respectively. 5) Test sample solution Approximately 20 mg of test sample was accurately weighed and placed in a 10 mL volumetric flask. Diluent was added to dissolve the test sample, and the solution was diluted to the mark and shaken well. Two solutions were prepared in parallel and designated as S1 / S2, respectively. 3.4 Calculation: Only the peak of related substance 2 was integrated

[0206]

number

[0207] A RS2 - average peak area of ​​reference solution 2;

[0208]

number

[0209] - average peak area of ​​five injections of reference solution 1; As - peak area of ​​related substance 2 in the test sample solution; M RS1 - weighed sample amount of related substance 2 in reference solution 1, mg; Ms - weighed sample amount of the test sample in the test sample solution, mg; M RS2 - weighed sample amount of related substance 2 in reference solution 2, mg;

[0210] [Table 22]

[0211] 4 content 4.1 Chromatography conditions: High-performance liquid chromatography (HPLC) Column: Waters XBridge C18 4.6 x 150 mm, 5 μm Detector: UV or equivalent detector Wavelength: 210nm Column temperature: 35℃ Flow rate: 1.0mL / min Injection volume: 5μL Needle cleaning: acetonitrile Mobile phase: 10mM KH2PO4 aqueous solution (pH2.8): ACN=57:43 (V / V) Run Time: 10 minutes 4.2 Reagents and reference materials Potassium dihydrogen phosphate (chromatography grade) Acetonitrile (chromatography grade) Phosphoric acid (chromatography grade) Reference substance of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate: Same structural formula as ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate 4.3 Preparation of solutions 1) Mobile phase: 10mM KH2PO4 aqueous solution (pH2.8): ACN=57:43 (V / V) 1.36 g of KH2PO4 was weighed and placed in 1000 mL of ultrapure water and completely dissolved. The pH was adjusted to 2.8 using phosphoric acid, and the solution was filtered using a 0.45 μm filter membrane. 2) Diluent (blank solution): acetonitrile:ultrapure water = 43:57 (v / v) 430 mL of acetonitrile and 570 mL of ultrapure water were measured, mixed thoroughly, and sonicated. 3) Reference substance solution Approximately 20 mg of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate reference material was accurately weighed and placed in a 100 mL volumetric flask. Diluent was added to dissolve the material, the solution was diluted to the mark, and the solution was shaken well. Two solutions were prepared in parallel. 4) Test sample solution Approximately 20 mg of test sample was accurately weighed and placed in a 100 mL volumetric flask. Diluent was added to dissolve the test sample, and the solution was diluted to the mark and shaken well. Two solutions were prepared in parallel. 4.4 Calculation

[0212]

number

[0213] A RS2 - average peak area of ​​reference solution 2;

[0214]

number

[0215] - average peak area of ​​five injections of reference solution 1; As - the peak area of ​​((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate in the test sample solution; M RS1 - weighed sample amount of reference substance ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate in reference solution 1, mg; Ms - weighed sample amount of the test sample in the test sample solution, mg; M RS2 - weighed sample amount of reference substance ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate in reference solution 2, mg; P - content of the reference substance ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate, %.

[0216] [Table 23]

[0217] [Table 24]

Claims

1. A crystal having crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate, characterized in that its X-ray powder diffraction pattern, obtained using Cu-Kα radiation and expressed in 2θ angles, contains characteristic peaks at 5.74±0.2°, 6.78±0.2°, 10.86±0.2°, 13.54±0.2°, 16.70±0.2°, and 22.65±0.2°.

2. 2. The crystal having crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate according to claim 1, characterized in that its X-ray powder diffraction pattern, obtained using Cu-Kα radiation and expressed in 2θ angles, contains characteristic peaks at 5.74±0.2°, 6.78±0.2°, 8.25±0.2°, 10.86±0.2°, 13.54±0.2°, 14.92±0.2°, 16.70±0.2°, 17.23±0.2°, 18.10±0.2°, 19.56±0.2°, 22.65±0.2° and 27.22±0.2°.

3. 3. The crystals of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate having crystalline form A according to claim 1 or 2, characterized in that differential scanning calorimetry of the crystals having crystalline form A shows an endothermic peak at 101.4°C.

4. mixing a solution I containing ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and a positive solvent with an anti-solvent to precipitate a solid, and performing solid-liquid separation to obtain crystals having crystalline form A; The positive solvent is C 1 ~C 5 Ester solvent, C 1 ~C 6 one or more of an alcohol solvent, a ketone solvent, a nitrile solvent, an ether solvent, and a lower halogenated alkane solvent; 1 ~C 5 The ester solvent is ethyl acetate; 1 ~C 6 The alcohol solvent is one or more of methanol, ethanol, isopropanol, and isobutanol; the ketone solvent is one or more of acetone, methyl ethyl ketone, and methyl isobutyl ketone; the nitrile solvent is acetonitrile; the ether solvent is tetrahydrofuran and / or 1,4-dioxane; the lower halogenated alkane solvent is dichloromethane; The anti-solvent is one or more of an ether solvent, an alcohol, a lower alkane solvent, and water; wherein the ether solvent is one or more of methyl tert-butyl ether, diethyl ether, and petroleum ether; and the alcohol is C 1 ~C 6 the lower alkane solvent is one or more of n-heptane, n-hexane, and n-octane; A method for preparing crystals having crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate according to any one of claims 1 to 3, characterized in that the volume ratio of the positive solvent to the antisolvent is from 1:10 to 1:

2.

5. 5. The method of claim 4, wherein the positive solvent is ethyl acetate and the anti-solvent is petroleum ether.

6. 6. The method according to claim 4, wherein the volume ratio of positive solvent to anti-solvent is between 0.3 and 0.

5.

7. mixing a solution II containing ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and a solvent at a temperature ranging from room temperature to 50°C, and centrifuging to obtain crystals having crystalline form A; The solvent is C 1 ~C 6 one or more of alcohol solvents, ester solvents, ether solvents, lower alkane solvents, lower halogenated alkane solvents, ketone solvents, aromatic hydrocarbon solvents, nitrile solvents, dimethyl sulfoxide, and water; where C 1 ~C 6 The method for preparing crystals having crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate according to any one of claims 1 to 3, characterized in that the alcohol solvent is one or more of methanol, ethanol, and isopropanol; the ester solvent is one or more of methyl acetate, ethyl acetate, and isopropyl acetate; the ether solvent is one or more of methyl ethyl ether, diethyl ether, methyl isopropyl ether, methyl tert-butyl ether, cyclopentyl methyl ether, anisole, tetrahydrofuran, 2-methyltetrahydrofuran, and 1,4-dioxane; the lower alkane solvent is n-heptane; the lower halogenated alkane solvent is dichloromethane; the ketone solvent is one or more of methyl ethyl ketone, methyl isobutyl ketone, and acetone; the aromatic hydrocarbon solvent is toluene; and the nitrile solvent is acetonitrile.

8. The solvent is C 1 ~C 6 Mixed solvents of alcohol and water, mixed solvents of ether and lower alkane, mixed solvents of ketone and lower alkane, mixed solvents of ketone and ether, mixed solvents of ester and C 1 ~C 6 Mixed solvents with alcohols, mixed solvents with aromatic hydrocarbons and lower alkanes, mixed solvents with ketones and C 1 ~C 6 a mixed solvent with an alcohol or a mixed solvent of an ether and an ester; 8. The method according to claim 7, wherein the volume ratio of the former to the latter in the mixed solvent is 1:4 to 1:

2.

9. suspending any crystalline or amorphous form of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate in a solvent at a temperature of 50°C, and stirring and centrifuging to obtain crystals having crystalline form A; The solvent is C 1 ~C 6 Mixed solvents of alcohol and water, mixed solvents of ketone and ether, mixed solvents of ester and C 1 ~C 6 Mixed solvents with alcohol, ketone and C 1 ~C 6 a mixed solvent with an alcohol, a mixed solvent of an ether and an ester, a mixed solvent of an aromatic hydrocarbon and a lower alkane, a mixed solvent of an ether and a lower alkane, a mixed solvent of two ethers, or a mixed solvent of a nitrile and an ether; 8. The method according to claim 7, wherein the volume ratio of the former to the latter in the mixed solvent is from 1:4 to 1:

2.

10. the solvent is a mixed solvent of isopropanol / water, methyl isobutyl ketone / methyl tert-butyl ether, ethyl acetate / isopropanol, toluene / n-heptane, 2-methyltetrahydrofuran / n-heptane, butanone / isopropanol, acetonitrile / cyclopentyl methyl ether, anisole / isopropyl acetate, or 1,4-dioxane / cyclopentyl methyl ether; 10. The method according to claim 9, wherein the volume ratio of the former to the latter in the mixed solvent is from 1:4 to 1:

2.

11. A method for producing crystals having crystalline form A, comprising: subjecting a solution III comprising ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and a solvent to one to five cycles of 50°C → 5°C → 50°C until a solid precipitates; and performing solid-liquid separation to obtain crystals having crystalline form A; or heating solution III to 50°C to dissolve the solution, performing hot filtration, cooling the filtrate to 5°C to -20°C, and performing solid-liquid separation to obtain crystals having crystalline form A; The solvent is C 1 ~C 6 one or two of an alcohol solvent, an ester solvent, a ketone solvent, an ether solvent, a lower alkane solvent, an aromatic hydrocarbon solvent, and water; where C 1 ~C 6 4. A method for preparing crystals having crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate according to claim 1, wherein the alcohol solvent is one or more of methanol, ethanol, and isopropanol; the ester solvent is one or more of methyl acetate, ethyl acetate, and isopropyl acetate; the ketone solvent is one or more of methyl ethyl ketone, methyl propyl ketone, and acetone; the ether solvent is one or more of methyl ethyl ether, diethyl ether, methyl isopropyl ether, methyl tert-butyl ether, cyclopentyl methyl ether, and anisole; the lower alkane solvent is n-heptane; and the aromatic hydrocarbon solvent is toluene.

12. The solvent is C 1 ~C 6 Alcohol and water mixed solvent, C 1 ~C 6 a mixed solvent of an alcohol and an ether, a mixed solvent of a ketone and an ester, a mixed solvent of an aromatic hydrocarbon and an ester, or a mixed solvent of a ketone and a lower alkane; wherein C 1 ~C 6 The alcohol solvent is ethanol and / or isopropanol; the ketone solvent is butanone and / or methyl isobutyl ketone; the ester solvent is isopropyl acetate; the ether solvent is cyclopentyl methyl ether; the aromatic hydrocarbon solvent is toluene; and the lower alkane solvent is n-heptane.

12. The method according to claim 11, wherein the volume ratio of the former to the latter in the mixed solvent is from 1:10 to 1:

2.

13. placing an opened first container containing any crystalline or amorphous form of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate solid into a second container containing a solvent, sealing the second container and allowing it to stand at room temperature, and when the solid becomes wet or a solid precipitate is observed to be present, collecting the product to obtain crystals having crystalline form A; The solvent is C 1 ~C 6 one or more of an alcohol solvent, an ether solvent, a ketone solvent, an ester solvent, an aromatic hydrocarbon solvent, dimethyl sulfoxide, and water; 1 ~C 6 4. A method for preparing crystals having crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate according to claim 1, wherein the alcohol solvent is one or more of methanol, ethanol, isopropanol, and isobutanol; the ether solvent is one or more of methyl tert-butyl ether, tetrahydrofuran, 2-methyltetrahydrofuran, 1,4-dioxane, and anisole; the ketone solvent is one or more of acetone, methyl ethyl ketone, and methyl isobutyl ketone; the ester solvent is ethyl acetate; and the aromatic hydrocarbon solvent is toluene.

14. placing an opened first container containing Solution IV comprising ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and an anti-solvent into a second container containing an anti-solvent, sealing the second container and allowing it to stand at room temperature, and when the solid becomes wet or a solid precipitate is observed, collecting the product to obtain crystals having crystalline form A; The positive solvent is C 1 ~C 6 The solvent is one or more of an alcohol solvent, an ether solvent, and a ketone solvent; the anti-solvent is one or more of a lower alkane solvent, an ether solvent, an alcohol solvent, and water; and in the case of a positive solvent, C 1 ~C 6 The alcohol solvent is one or more of methanol, ethanol, isopropanol, and isobutanol; for the positive solvent, the ether solvent is one or more of tetrahydrofuran, 2-methyltetrahydrofuran, and 1,4-dioxane; for the positive solvent, the ketone solvent is one or more of acetone, methyl ethyl ketone, and methyl isobutyl ketone; for the anti-solvent, the lower alkane solvent is n-heptane; for the anti-solvent, the ether solvent is methyl tert-butyl ether; for the anti-solvent, the alcohol solvent is isopropanol; A method for preparing crystals having crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate according to any one of claims 1 to 3, characterized in that the volume ratio of the positive solvent to the antisolvent is from 1:10 to 1:

2.

15. volatilizing a solution V comprising ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and a solvent at room temperature; and collecting the precipitated solid to obtain crystals having crystalline form A; The solvent is C 1 ~C 6 one or more of an alcohol solvent, a ketone solvent, an ester solvent, an ether solvent, a lower alkane solvent, an aromatic hydrocarbon solvent, a nitrile solvent, a lower halogenated alkane solvent, and water; where C 1 ~C 6 4. A method for preparing crystals having crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate according to any one of claims 1 to 3, characterized in that the alcohol solvent is one or more of methanol, ethanol, and isopropanol; the ketone solvent is one or more of methyl ethyl ketone, methyl isobutyl ketone, and acetone; the ester solvent is one or more of methyl acetate, ethyl acetate, and isopropyl acetate; the ether solvent is one or more of methyl ethyl ether, diethyl ether, methyl isopropyl ether, methyl tert-butyl ether, cyclopentyl methyl ether, anisole, and 1,4-dioxane; the lower alkane solvent is n-heptane; the aromatic hydrocarbon solvent is toluene; the nitrile solvent is acetonitrile; and the lower halogenated alkane solvent is dichloromethane.

16. A crystal having crystalline form B of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate, characterized in that its X-ray powder diffraction pattern, obtained using Cu-Kα radiation and expressed in 2θ angles, contains characteristic peaks at 5.44±0.2°, 10.90±0.2°, 14.09±0.2°, 16.17±0.2°, 17.92±0.2°, 20.66±0.2°, and 23.13±0.2°.

17. 17. The crystal having crystalline form B of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate according to claim 16, characterized in that its X-ray powder diffraction pattern, obtained using Cu-Kα radiation and expressed in 2θ angles, contains characteristic peaks at 5.44±0.2°, 6.19±0.2°, 10.90±0.2°, 14.09±0.2°, 14.88±0.2°, 16.17±0.2°, 17.92±0.2°, 20.66±0.2°, 21.69±0.2° and 23.13±0.2°.

18. 18. The crystal having crystalline form B of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate according to claim 17, characterized in that its X-ray powder diffraction pattern, obtained using Cu-Kα radiation and expressed in 2θ angles, contains characteristic peaks at 5.44±0.2°, 6.19±0.2°, 8.06±0.2°, 10.90±0.2°, 12.18±0.2°, 14.09±0.2°, 14.88±0.2°, 16.17±0.2°, 17.92±0.2°, 20.66±0.2°, 21.69±0.2°, 23.13±0.2°, 24.42±0.2° and 26.03±0.2°.

19. The method comprises the steps of: mixing a solution A containing ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate and a solvent with an anti-solvent to precipitate a solid; and performing solid-liquid separation to obtain crystals having a crystalline form B; the positive solvent is one or more of methyl acetate, ethyl acetate, and isopropyl acetate; the anti-solvent is an alkane solvent, the alkane solvent being one or more of n-hexane, n-heptane, and n-octane; 19. A method for preparing crystals having crystalline form B of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate according to any one of claims 16 to 18, characterized in that the volume ratio of the positive solvent to the antisolvent is from 1:10 to 1:

2.

20. 20. The method according to claim 19, characterized in that crystals of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate according to any one of claims 1 to 3 having crystalline form A are dissolved in a positive solvent to obtain solution A.

21. A pharmaceutical composition comprising a crystal having crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate according to any one of claims 1 to 3, or a crystal having crystalline form B of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate according to any one of claims 16 to 18, and a pharmaceutically acceptable carrier, diluent or excipient.

22. 22. Use of a crystal having crystalline form A of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate according to any one of claims 1 to 3, or a crystal having crystalline form B of ((5-nitroquinolin-8-yl)oxy)methyl-isobutyryl-L-prolinate according to any one of claims 16 to 18, or a pharmaceutical composition according to claim 21, in the preparation of a medicament for treating an infectious disease or cancer; wherein the infectious disease is a systemic infection, a reproductive system infection, or a urinary system infection; and the cancer is bladder cancer or prostate cancer.

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