RU2781383C2 - Sulfonamide compounds and their use - Google Patents

Abstract

FIELD: biotechnology.

SUBSTANCE: invention relates to a cocrystal of benzoic acid and 5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole-2-yl)propyl)sulfamoyl)benzamide, where the powder X-ray diffractogram of the cocrystal contains five or more peaks at (2θ ± 0.2°) 6.8°, 7.8°, 11.2°, 13.4°, 13.7°, 16.0°, 17.1°, 17.8°, and 23.2°. The invention also relates to a pharmaceutical composition having inhibitory activity against ribonucleotide reductase, containing (i) a cocrystal of benzoic acid and 5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole-2-yl)propyl)sulfamoyl)benzamide, and (ii) a pharmaceutically acceptable carrier. The invention relates to a method for inhibition of ribonucleotide reductase in vivo, including the administration of therapeutically effective amount of a pharmaceutical composition according to the invention to a human subject who needs it. A cocrystal of benzoic acid and 5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole-2-yl)propyl)sulfamoyl)benzamide is used for the production of a ribonucleotide reductase inhibitor as a drug for inhibition of activity against ribonucleotide reductase.

EFFECT: cocrystal of benzoic acid and 5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole-2-yl)propyl)sulfamoyl)benzamide, which has properties of stability, lower electrostatic character, which is a powerful and selective inhibitor of ribonucleotide reductase.

18 cl, 20 tbl, 12 dwg, 10 ex

Description

FIELD OF TECHNOLOGY TO WHICH THE INVENTION RELATES

[0001] The present invention relates to new crystalline forms of sulfonamide compounds having ribonucleotide reductase inhibitory activity, to pharmaceutical compositions containing the crystalline form of the compounds, and to methods for their preparation and use.

BACKGROUND OF THE INVENTION

[0002] Ribonucleotide reductase (hereinafter also referred to as "RNR") consists of a heterooligomer of M1 large subunit and M2 small subunit, with the expression of both subunits required for enzymatic activity. RNR recognizes ribonucleoside-5'-diphosphate (hereinafter also referred to as "NDP") as a substrate and catalyzes its reduction to 2'-deoxyribonucleoside-5'-diphosphate (hereinafter also referred to as "dNDP"). RNR is a rate-limiting enzyme in the de novo dNTP synthesis pathway and plays an essential role in DNA synthesis and repair.

[0003] The enzymatic activity of RNR is closely associated with cell proliferation, and its enzymatic activity has been reported to be particularly high in cancer. In various types of solid tumors and blood cancers, numerous correlations have been reported with overexpression of the M2 RNR subunit and its effect on cancer prognosis. In addition, cell growth inhibition is achieved through RNR inhibition, and an in vivo antitumor effect has been described in cell lines derived from several types of cancer and in non-clinical models. Thus, there is a reasonable suggestion that RNR is an important molecular target for cancer treatment.

[0004] It is generally accepted that hydroxyurea (hereinafter also referred to as "HU") and 3-aminopyridine-2-carboxaldehydethiosemicarbazone (hereinafter also referred to as "3-AP") exhibit RNR inhibitory activity. These compounds, however, differ in structure from the sulfonamide compounds of the present invention. Although HU has been clinically used for more than 30 years, its RNR inhibitory activity is weak and its effect is limited. Resistance to HU application is also considered a problem. 3-AP is able to chelate metal ions, especially iron (Fe) ions, thereby inhibiting RNR, but it has been suggested that 3-AP off-targets various other Fe ion-containing proteins, leading to side effects such as hypoxia , shortness of breath, methemoglobinemia and the like in clinical cases.

[0005] Accordingly, there is a need to develop a potent RNR inhibitor that does not chelate metal ions and can be used to treat RNR-related diseases such as cancer.

[0006] In addition, there is a need to develop an RNR inhibitor that is easy to handle. It is known that the ability to accumulate an electrostatic charge and the hygroscopicity of a biologically active compound affect the handling of the compound when it is included in a potential pharmaceutical composition. For example, active compounds that are exposed to static electricity can cause problems in the manufacturing process such as reduced yield and uneven packaging. Thus, an active chemical compound having a low electrostatic charge is preferred. Hygroscopic compounds present problems with their absorption of moisture, which leads to changes in the mass of the compound depending on the amount of water present in the environment, making it difficult to accurately assess the biological effectiveness of the compound and ensure the uniformity of pharmaceutical compositions containing the compound. Accordingly, it is desirable to use an active chemical compound with low hygroscopicity.

SUMMARY OF THE INVENTION

[0007] One of the objectives of the present invention is to provide a new, stable and less electrostatic crystalline form, co-crystal and/or salt of a compound that is a potent and selective RNR inhibitor and can be used as an antitumor agent or therapeutic agent in the treatment of other diseases associated with with RNR.

[0008] As a result of conducting extensive research to identify a chemical compound that satisfies the above requirements, the present inventors have found stable, low electrostatic and non-hygroscopic forms of sulfonamide compounds having excellent RNR inhibitory activity and useful as a therapeutic agent for the treatment tumors and other diseases associated with RNR. These sulfonamide compounds exist in crystalline form such as co-crystals and/or as a salt.

[0009] Various aspects of the present invention are contained in the following paragraphs [1] - [28].

[1] A crystalline form of a compound selected from the group consisting of the following:

5-bromo-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole -2-yl)propyl)sulfamoyl)benzamide;

5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole -2-yl)propyl)sulfamoyl)benzamide;

5-chloro-2-(N-((1S,2R)-2-(3-ethyl-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4 -oxadiazol-2-yl)propyl)sulfamoyl)benzamide;

5-chloro-2-(N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4 -oxadiazol-2-yl)propyl)sulfamoyl)benzamide;

5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole-2- yl) propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide;

5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole- 2-yl)propyl)-4-hydroxy-4-methylchroman-8-sulfonamide;

N-((1S,2R)-2-(3-bromo-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl) propyl)-5-chloro-4-hydroxy-4-methylchroman-8-sulfonamide;

5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole- 2-yl)propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide; and

5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole-2- yl)propyl)-4-hydroxychroman-8-sulfonamide.

[2] The crystalline form according to [1], wherein the chemical purity of the crystalline form is 90% or more.

[3] Crystalline form according to [1] or item [2], where the optical purity of the crystalline form is 100% e.i.

[4] The crystalline form according to any one of [1] to [3], wherein the crystalline form is stable under conditions of about 40° C. and about 75% relative humidity for about 4 weeks.

[5] The crystalline form according to any one of [1] to [4], wherein the crystalline form further comprises benzoic acid.

[6] The crystal form according to any one of [1] to [5], wherein the crystal form is a co-crystal of benzoic acid and the compound.

[7] A pharmaceutical composition comprising (i) a crystalline form according to any one of paragraphs [1]-[6] and (ii) a pharmaceutically acceptable carrier.

[8] A method for inhibiting ribonucleotide reductase in vivo , comprising administering to a person in need thereof a therapeutically effective amount of the pharmaceutical composition according to [7]. In some embodiments, the inhibition of ribonucleotide reductase occurs in a human tumor cell.

[9] An antitumor agent containing a crystalline form according to any one of [1] to [6] as an active ingredient. In some embodiments, the implementation of the anticancer agent is an oral anticancer agent.

[10] The use of a crystalline form according to any one of [1] to [6] to obtain a ribonucleotide reductase inhibitor.

[11] Use of the crystalline form according to any one of [1] to [6] as a medicine.

[12] Use of the crystalline form according to any one of [1] to [6] as an antitumor agent. In some embodiments, the implementation of the anticancer agent is an oral anticancer agent.

[13] The crystalline form according to any one of [1]-[6] for use as a ribonucleotide reductase inhibitor.

[14] The crystalline form according to any one of paragraphs [1]-[6] for use in the prevention or treatment of tumors.

[15] The crystalline form according to any one of [1]-[6] for use in the prevention or treatment of tumors by oral administration of the compound.

[16] The crystalline form of any one of [1]-[6], wherein the compound or benzoic acid may be substituted with one or more radioactive isotopes or a non-radioactive isotope.

[17] A benzoic acid salt of a compound selected from the group consisting of the following:

5-bromo-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole -2-yl)propyl)sulfamoyl)benzamide;

5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole -2-yl)propyl)sulfamoyl)benzamide;

5-chloro-2-(N-((1S,2R)-2-(3-ethyl-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4 -oxadiazol-2-yl)propyl)sulfamoyl)benzamide;

5-chloro-2-(N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4 -oxadiazol-2-yl)propyl)sulfamoyl)benzamide;

5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole-2- yl) propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide;

5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole- 2-yl)propyl)-4-hydroxy-4-methylchroman-8-sulfonamide;

N-((1S,2R)-2-(3-bromo-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl) propyl)-5-chloro-4-hydroxy-4-methylchroman-8-sulfonamide;

5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole- 2-yl)propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide; and

5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole-2- yl)propyl)-4-hydroxychroman-8-sulfonamide.

[18] Pharmaceutical composition containing (i) benzoic acid salt p.[17] and (ii) a pharmaceutically acceptable carrier.

[19] A method for inhibiting ribonucleotide reductase in vivo , comprising administering to a human subject in need thereof a therapeutically effective amount of the pharmaceutical composition of [18]. In some embodiments, inhibition of ribonucleotide reductase occurs in a tumor cell of a human subject.

[20] An antitumor agent containing a benzoic acid salt according to [17] as an active ingredient. In some embodiments, the implementation of the anticancer agent is an oral anticancer agent.

[21] Application of salt of benzoic acid p.[17] to obtain a ribonucleotide reductase inhibitor.

[22] Application of salt of benzoic acid p.[17] as a medicine.

[23] Application of the salt of benzoic acid p.[17] as an antitumor agent. In some embodiments, the implementation of the anticancer agent is an oral anticancer agent.

[24] Salt of benzoic acid p. [17] for use as a ribonucleotide reductase inhibitor.

[25] Salt of benzoic acid p. [17] for use in the prevention or treatment of tumors.

[26] Salt of benzoic acid p. [17] for use in the prevention or treatment of tumors by oral administration of the compound.

[27] The benzoic acid salt of [17], wherein the benzoic acid may be substituted with one or more radioactive isotopes or non-radioactive isotopes.

[28] The present invention relates to crystalline forms according to any of [1] to [6], co-crystals of [6] and/or benzoic acid salts of [17], which act as RNR inhibitors.

[00010] The benzoic acid crystalline forms, co-crystals and/or salts of the present invention exhibit excellent RNR inhibitory activity and stability, and are easy to handle because they are non-hygroscopic and/or non-electrostatic. Accordingly, the crystalline forms, co-crystals and/or salts of benzoic acid described herein are suitable for use in the treatment of diseases associated with RNR.

BRIEF DESCRIPTION OF THE DRAWINGS

[00011] Figure 1 is a graph illustrating diurnal variation in relative tumor volume (hereinafter also referred to as "RTV") following treatment with an exemplary compound described herein.

[00012] Figure 2 is a graph illustrating daily RTV variations following treatment with an exemplary compound described herein.

[00013] Figure 3 is a graph illustrating diurnal variations in RTV following treatment with an exemplary compound described herein.

[00014] Figure 4 is a graph illustrating diurnal variations in RTV following treatment with an exemplary compound described herein.

[00015] Figure 5 shows an exemplary crystal form described in this application. An exemplary crystalline form had a crystal size of 0.15 x 0.20 x 0.25 mm, was colorless, and plate-shaped.

[00016] FIG. 6 is an X-ray powder diffraction (XRD) pattern of an exemplary co-crystal.

[00017] Figure 7 shows a differential scanning calorimetry (DSC) curve of an exemplary co-crystal.

[00018] FIG. 8 shows isothermal moisture adsorption/desorption curves of a co-crystal of an exemplary compound (A) and a free form of the same compound (B).

[00019] FIG. 9 shows a simulation result of an X-ray diffraction pattern based on a single crystal analysis result of an exemplary benzoic acid co-crystal.

[00020] FIGS. 10-12 show optical purity measurements of an exemplary crystalline form.

DETAILED DESCRIPTION OF THE INVENTION

[00021] The present invention relates to crystalline forms, co-crystals or benzoic acid salts of a compound selected from the group consisting of the following:

5-bromo-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole -2-yl)propyl)sulfamoyl)benzamide;

5-chloro-2-(N-((1S,2R)-2-(2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl )propyl)sulfamoyl)benzamide;

5-bromo-2-(N-((1S,2R)-2-(2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl )propyl)sulfamoyl)benzamide;

5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole -2-yl)propyl)sulfamoyl)benzamide;

5-chloro-2-(N-(1S,2R)-2-(2-fluoronaphthalen-1-yl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2- yl) propyl) sulfamoyl) benzamide;

5-chloro-2-(N-((1S,2R)-2-(3-ethyl-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4 -oxadiazol-2-yl)propyl)sulfamoyl)benzamide;

5-chloro-2-(N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4 -oxadiazol-2-yl)propyl)sulfamoyl)benzamide;

5-bromo-2-(N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4 -oxadiazol-2-yl)propyl)sulfamoyl)benzamide;

2-(N-((1S,2R)-2-(3-bromo-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole-2 -yl)propyl)sulfamoyl)-5-chlorobenzamide;

5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole-2- yl)propyl)-6-(pyrrolidine-1-carbonyl)pyridine-2-sulfonamide;

5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole-2- yl) propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide;

5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole- 2-yl)propyl)-4-hydroxy-4-methylchroman-8-sulfonamide;

N-((1S,2R)-2-(3-bromo-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl) propyl)-5-chloro-4-hydroxy-4-methylchroman-8-sulfonamide;

5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole- 2-yl)propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide;

5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole-2- yl) propyl)-4-hydroxychroman-8-sulfonamide;

3-chloro-6-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole -2-yl)propyl)sulfamoyl)-N,N-dimethylpicolinamide;

4-amino-2-methoxy-N-((1S,2R)-2-(8-methylnaphthalene-1-yl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazol- 2-yl)propyl)benzenesulfonamide;

4-amino-N-((1S,2R)-2-(2,3-dihydro-1H-inden-4-yl)-1-(5-oxo-4,5-dihydro-1,3,4- oxadiazol-2-yl)propyl)-2-methoxybenzenesulfonamide; and

5-chloro-2-(((1S,2R)-methyl-d3-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4 -oxadiazol-2-yl)propyl)sulfamoyl)benzamide.

[00022] In some embodiments, the compound is selected from the group consisting of the following:

5-bromo-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole -2-yl)propyl)sulfamoyl)benzamide;

5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole -2-yl)propyl)sulfamoyl)benzamide;

5-chloro-2-(N-((1S,2R)-2-(3-ethyl-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4 -oxadiazol-2-yl)propyl)sulfamoyl)benzamide;

5-chloro-2-(N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4 -oxadiazol-2-yl)propyl)sulfamoyl)benzamide;

5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole-2- yl) propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide;

5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole- 2-yl)propyl)-4-hydroxy-4-methylchroman-8-sulfonamide;

N-((1S,2R)-2-(3-bromo-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl) propyl)-5-chloro-4-hydroxy-4-methylchroman-8-sulfonamide;

5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole- 2-yl)propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide; and

5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole-2- yl)propyl)-4-hydroxychroman-8-sulfonamide.

[00023] In additional embodiments, the compound is selected from the group consisting of the following:

5-bromo-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole -2-yl)propyl)sulfamoyl)benzamide;

5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole -2-yl)propyl)sulfamoyl)benzamide;

5-chloro-2-(N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4 -oxadiazol-2-yl)propyl)sulfamoyl)benzamide;

5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole- 2-yl)propyl)-4-hydroxy-4-methylchroman-8-sulfonamide; and

5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole-2- yl)propyl)-4-hydroxychroman-8-sulfonamide.

[00024] In some embodiments, the compound is 5-bromo-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4.5 -dihydro-1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide. In some embodiments, the present invention provides a crystalline form of 5-bromo-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4.5 -dihydro-1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide or benzoic acid co-crystal and compound.

[00025] In some embodiments, the compound is 5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4.5 -dihydro-1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide. In some embodiments, the present invention provides crystalline forms of 5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4.5 -dihydro-1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide or benzoic acid co-crystal and compound.

[00026] In some embodiments, the compound is 5-chloro-2-(N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4 ,5-dihydro-1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide. In some embodiments, the present invention relates to the crystalline form of 5-chloro-2-(N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4 ,5-dihydro-1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide or benzoic acid co-crystal and compound.

[00027] In some embodiments, the compound is 5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5- dihydro-1,3,4-oxadiazol-2-yl)propyl)-4-hydroxy-4-methylchroman-8-sulfonamide. In some embodiments, the present invention relates to the crystalline form of 5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5- dihydro-1,3,4-oxadiazol-2-yl)propyl)-4-hydroxy-4-methylchroman-8-sulfonamide or benzoic acid co-crystal and compound.

[00028] In some embodiments, the compound is 5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro- 1,3,4-oxadiazol-2-yl)propyl)-4-hydroxychroman-8-sulfonamide. In some embodiments, the present invention provides a crystalline form of 5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro- 1,3,4-oxadiazol-2-yl)propyl)-4-hydroxychroman-8-sulfonamide or benzoic acid co-crystal and compound.

[00029] The crystalline forms of the compounds, co-crystals and/or salts of benzoic acid of the present invention and their intermediates can be isolated and purified using well-known separation and purification methods such as recrystallization, crystallization, distillation and column chromatography.

[00030] When optical isomers, stereoisomers, tautomers or rotational isomers are possible in the form of a crystalline form of the compounds, co-crystals or salts of benzoic acid of the present invention, but are not expressly represented, it is understood that the crystalline forms of the compounds, co-crystals and/or salts of benzoic acid encompass these isomers singly or as mixtures thereof. For example, unless otherwise stated, when the crystalline form of the compound, co-crystal and/or benzoic acid salt of the present invention is present as a racemate, the possible enantiomers and/or diastereomers that can be separated from the racemate are also considered to be covered by the present invention. Enantiomers and/or diastereomers can usually be obtained by well known synthetic methods.

[00031] As used herein and unless otherwise indicated, the term "crystal" and related terms used herein, when used to describe a compound, substance, modification, material, component, or product, means that the compound, substance , modification, material, component, or product is essentially crystalline, as determined by X-ray diffraction analysis. See, for example, Remington: The Science and Practice of Pharmacy, 21st edition, ^Lippincott , Williams and Wilkins, Baltimore, MD (2005); The United States Pharmacopeia, ^23rd ed. 1843-1844 (1995).

[00032] As used herein and unless otherwise indicated, the term "crystalline form" and related terms herein refer to solid forms that are crystalline. Crystalline forms include single-component crystalline forms and multicomponent crystalline forms, and may optionally include, without limitation, co-crystals, salts (including pharmaceutically acceptable salts), polymorphs, solvates, hydrates, and/or other molecular complexes. In some embodiments, the crystalline form of the substance may be substantially free of amorphous forms and/or other crystalline forms. In some embodiments, the crystalline form of the substance may contain less than about 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10 %, 11%, 12%, 13%, 14%, 15%, 20%, 25%, 30%, 35%, 40%, 45% or 50% of one or more amorphous forms and/or other crystalline forms by weight .

[00033] The term "co-crystal" refers to a molecular complex derived from a number of co-crystal formers known in the art. Unlike a salt, a co-crystal generally does not involve hydrogen transfer between the co-crystal former and the drug, and instead involves intermolecular interactions, such as hydrogen bonding, aromatic ring grouping, or dispersion forces, between the co-crystal former and the sulfonamide compound in the crystal structure. In some embodiments, the present invention relates to a co-crystal containing or consisting of benzoic acid and a sulfonamide compound as described herein.

[00034] As used herein and unless otherwise indicated, the terms "polymorphs", "polymorphic forms" and related terms in this document refer to two or more crystalline forms that consist essentially of the same molecule, molecules and /or ions. Like different crystalline forms, different polymorphs may have different physical properties, such as, for example, melting point, specific heat of fusion, solubility, solubility properties and/or vibrational spectra, as a result of the arrangement or conformation of molecules and/or ions in the crystal lattice. Differences in physical properties can affect pharmaceutical parameters such as storage stability, compressibility and density (important in formulating and manufacturing a product) and dissolution rate (an important bioavailability factor). Differences in stability may be due to changes in chemical reactivity (for example, different oxidation that causes the dosage form to discolor faster when containing one polymorph compared to a form containing the other polymorph) or mechanical changes (for example, tablets crumble during storage , since the kinetically preferred polymorph is converted to the thermodynamically more stable polymorph) or both (eg tablets of one polymorph are more susceptible to degradation at high humidity). As a result of solubility/dissolution differences, in the extreme case, some polymorphic transformations can lead to loss of activity or, at the other extreme, to toxicity. In addition, physical properties may be important in processing (for example, one polymorph may be more likely to form solvates or may be difficult to filter and wash out of impurities, and the particle shape and size distribution may be different between polymorphs).

[00035] As used herein and unless otherwise indicated, the term "amorphous", "amorphous form" and related terms used herein means that the substance, component or product in question is not essentially crystalline, as defined by X-ray diffraction data. In particular, the term "amorphous form" describes a disordered solid form, i.e., a solid form devoid of long-range crystalline order.

[00036] The term "pharmaceutically acceptable salt" refers to salts derived from various organic and inorganic counterions known in the art. The pharmaceutically acceptable salt may be safe for animal or human consumption.

[00037] In one aspect, the invention provides a crystalline form of a sulfonamide compound as described above. In some embodiments, the crystal structure of the sulfonamide compound described herein includes a leaf-like, lamellar, and/or planar crystal structure.

[00038] Methods for determining crystalline forms and amorphous forms include, without limitation, thermogravimetric analysis (TGA), differential scanning calorimetry (DSC), X-ray powder diffractometry (XRPD), single crystal X-ray diffractometry, vibrational spectroscopy, for example, infrared (IR) and Raman spectroscopy, nuclear magnetic resonance (NMR) spectroscopy in solid phase and solution, optical microscopy, optical heating microscopy, scanning electron microscopy (SEM), electron crystallography and quantification, particle size analysis (PSA), surface area analysis, solubility determinations , dissolution determinations, elemental analysis and Karl Fischer analysis. Characteristic unit cell parameters can be determined using one or more methods such as, but not limited to, x-ray diffraction and neutron diffraction, including single crystal diffraction and powder diffraction. Suitable methods for analyzing powder diffraction data include profile refinement, such as Rietveld refinement, which can be used, for example, to analyze diffraction peaks associated with a single phase in a sample containing more than one solid phase. Other methods suitable for analyzing powder diffraction data include unit cell indexing, which allows one skilled in the art to determine unit cell parameters from a sample containing crystalline powder.

[00039] In an X-ray powder diffraction spectrum, the diffraction angle and overall pattern can be important in identifying the identity of the crystal due to the nature of the data. The relative intensity of the powder X-ray diffraction spectrum may vary depending on the direction of crystal growth, particle size or measurement conditions and, therefore, should not be interpreted strictly. In addition, some values obtained from different spectra may cause some errors depending on the crystal growth direction, particle size, measurement conditions, and the like. For example, the diffraction angle may be the diffraction angle (2θ±0.2°) in an X-ray powder diffraction spectrum, which means that the diffraction angle may individually be in the range of ±0.2° of any particular value, unless otherwise indicated.

[00040] In some embodiments, the invention provides a crystalline form of 5-bromo-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4 ,5-dihydro-1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide. In additional embodiments, the X-ray diffraction pattern of this crystal form has two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, or nine peaks at (2θ±0.2, 0 .3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 or 1.0°) 6.8°, 7.8°, 11.2°, 13.4 °, 13.7°, 16.0°, 17.1°, 17.8° and 23.2°. In other embodiments, the crystalline form of 5-bromo-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1 ,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, or 161° C. to about 160, 161, 162, 163, 164, 165, 166, 167, 168, 169 or 170°C according to DSC. In some embodiments, the crystalline form of 5-bromo-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1 ,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide has an endothermic peak from 155°C to 168°C, from 158°C to 162°C, from 159°C to 161°C, from 159°C up to 165°C or from 160°C to 163°C according to DSC. In additional embodiments, the crystalline form of 5-bromo-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1 ,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169 or 170°C according to DSC. In further embodiments, the optical purity of a given crystalline form is 100% ee.

[00041] In some embodiments, the invention relates to the crystalline form of 5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4 ,5-dihydro-1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide. In additional embodiments, the X-ray diffraction pattern of this crystal form has two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, or nine peaks at (2θ±0.2, 0 .3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 or 1.0°) 6.8°, 7.8°, 11.2°, 13.4 °, 13.7°, 16.0°, 17.1°, 17.8° and 23.2°. In other embodiments, the crystalline form of 5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1 ,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, or 161° C. to about 160, 161, 162, 163, 164, 165, 166, 167, 168, 169 or 170°C according to DSC. In some embodiments, the crystalline form of 5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1 ,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide has an endothermic peak from 155°C to 168°C, from 158°C to 162°C, from 159°C to 161°C, from 159°C up to 165°C or from 160°C to 163°C according to DSC. In additional embodiments, the crystalline form of 5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1 ,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169 or 170°C according to DSC. In further embodiments, the optical purity of a given crystalline form is 100% ee.

[00042] In some embodiments, the invention relates to the crystalline form of 5-chloro-2-(N-((1S,2R)-2-(3-ethyl-6-fluoro-2-methylphenyl)-1-(5-oxo -4,5-dihydro-1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide. In additional embodiments, the X-ray diffraction pattern of this crystal form has two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, or nine peaks at (2θ±0.2, 0 .3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 or 1.0°) 6.8°, 7.8°, 11.2°, 13.4 °, 13.7°, 16.0°, 17.1°, 17.8° and 23.2°. In other embodiments, the crystalline form of 5-chloro-2-(N-((1S,2R)-2-(3-ethyl-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro -1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, or 161°C to about 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, or 170°C according to DSC. In some embodiments, the crystalline form of 5-chloro-2-(N-((1S,2R)-2-(3-ethyl-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro -1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide has an endothermic peak from 155°C to 168°C, from 158°C to 162°C, from 159°C to 161°C, from 159 °C to 165°C or 160°C to 163°C according to DSC. In additional embodiments, the crystalline form of 5-chloro-2-(N-((1S,2R)-2-(3-ethyl-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro -1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169 or 170°C according to DSC. In further embodiments, the optical purity of a given crystalline form is 100% ee.

[00043] In some embodiments, the invention relates to a co-crystal of benzoic acid and 5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo -4,5-dihydro-1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide. In additional embodiments, the X-ray diffraction pattern of this co-crystal has two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, or nine peaks at (2θ±0.2, 0, 3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 or 1.0°) 6.8°, 7.8°, 11.2°, 13.4° , 13.7°, 16.0°, 17.1°, 17.8° and 23.2°. In other embodiments, the co-crystal has an endothermic peak from about 154, 155, 156, 157, 158, 159, 160, or 161°C to about 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, or 170°C according to DSC. In some embodiments, a co-crystal of benzoic acid and 5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro -1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide has an endothermic peak from 155°C to 168°C, from 158°C to 162°C, from 159°C to 161°C, from 159 °C to 165°C or 160°C to 163°C according to DSC. In additional embodiments, this co-crystal has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, or 170°C as measured by DSC. In additional embodiments, the optical purity of a given benzoic acid co-crystal is 100% ee.

[00044] In some embodiments, the invention relates to the crystalline form of 5-chloro-2-(N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo -4,5-dihydro-1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide. In additional embodiments, the X-ray diffraction pattern of this crystal form has two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, or nine peaks at (2θ±0.2, 0 .3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 or 1.0°) 6.8°, 7.8°, 11.2°, 13.4 °, 13.7°, 16.0°, 17.1°, 17.8° and 23.2°. In other embodiments, the crystalline form of 5-chloro-2-(N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro -1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, or 161°C to about 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, or 170°C according to DSC. In some embodiments, the crystalline form of 5-chloro-2-(N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro -1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide has an endothermic peak from 155°C to 168°C, from 158°C to 162°C, from 159°C to 161°C, from 159 °C to 165°C or 160°C to 163°C according to DSC. In additional embodiments, the crystalline form of 5-chloro-2-(N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro -1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169 or 170°C according to DSC. In further embodiments, the optical purity of a given crystalline form is 100% ee.

[00045] In some embodiments, the invention relates to the crystalline form of 5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5- dihydro-1,3,4-oxadiazol-2-yl)propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide. In additional embodiments, the X-ray diffraction pattern of this crystal form has two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, or nine peaks at (2θ±0.2, 0 .3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 or 1.0°) 6.8°, 7.8°, 11.2°, 13.4 °, 13.7°, 16.0°, 17.1°, 17.8° and 23.2°. In other embodiments, the crystalline form of 5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3, 4-oxadiazol-2-yl)propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, or 161° C. to about 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, or 170°C according to DSC. In some embodiments, the crystalline form of 5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3, 4-oxadiazol-2-yl)propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide has an endothermic peak from 155°C to 168°C, from 158°C to 162°C, from 159° C to 161°C, 159°C to 165°C, or 160°C to 163°C according to DSC. In additional embodiments, the crystalline form of 5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3, 4-oxadiazol-2-yl)propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, or 170°C according to DSC. In further embodiments, the optical purity of a given crystalline form is 100% ee.

[00046] In some embodiments, the invention relates to the crystalline form of 5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4, 5-dihydro-1,3,4-oxadiazol-2-yl)propyl)-4-hydroxy-4-methylchroman-8-sulfonamide. In additional embodiments, the X-ray diffraction pattern of this crystal form has two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, or nine peaks at (2θ±0.2, 0 .3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 or 1.0°) 6.8°, 7.8°, 11.2°, 13.4 °, 13.7°, 16.0°, 17.1°, 17.8° and 23.2°. In other embodiments, the crystalline form of 5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1, 3,4-oxadiazol-2-yl)propyl)-4-hydroxy-4-methylchroman-8-sulfonamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, or 161° C. to about 160, 161, 162, 163, 164, 165, 166, 167, 168, 169 or 170°C according to DSC. In some embodiments, the crystalline form of 5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1, 3,4-oxadiazol-2-yl)propyl)-4-hydroxy-4-methylchroman-8-sulfonamide has an endothermic peak from 155°C to 168°C, from 158°C to 162°C, from 159°C to 161°C, 159°C to 165°C or 160°C to 163°C according to DSC. In additional embodiments, the crystalline form of 5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1, 3,4-oxadiazol-2-yl)propyl)-4-hydroxy-4-methylchroman-8-sulfonamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169 or 170°C according to DSC. In further embodiments, the optical purity of a given crystalline form is 100% ee.

[00047] In some embodiments, the invention relates to the crystalline form of N-((1S,2R)-2-(3-bromo-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro- 1,3,4-oxadiazol-2-yl)propyl)-5-chloro-4-hydroxy-4-methylchroman-8-sulfonamide. In additional embodiments, the X-ray diffraction pattern of this crystal form has two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, or nine peaks at (2θ±0.2, 0 .3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 or 1.0°) 6.8°, 7.8°, 11.2°, 13.4 °, 13.7°, 16.0°, 17.1°, 17.8° and 23.2°. In other embodiments, the crystalline form N-((1S,2R)-2-(3-bromo-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4- oxadiazol-2-yl)propyl)-5-chloro-4-hydroxy-4-methylchroman-8-sulfonamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, or 161°C to about 160, 161, 162, 163, 164, 165, 166, 167, 168, 169 or 170°C according to DSC. In some embodiments, the crystalline form N-((1S,2R)-2-(3-bromo-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4- oxadiazol-2-yl)propyl)-5-chloro-4-hydroxy-4-methylchroman-8-sulfonamide has an endothermic peak from 155°C to 168°C, from 158°C to 162°C, from 159°C to 161°C, 159°C to 165°C or 160°C to 163°C according to DSC. In further embodiments, the crystalline form N-((1S,2R)-2-(3-bromo-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4- oxadiazol-2-yl)propyl)-5-chloro-4-hydroxy-4-methylchroman-8-sulfonamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169 or 170°C according to DSC. In further embodiments, the optical purity of a given crystalline form is 100% ee.

[00048] In some embodiments, the invention relates to the crystalline form of 5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4, 5-dihydro-1,3,4-oxadiazol-2-yl)propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide. In additional embodiments, the X-ray diffraction pattern of this crystal form has two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, or nine peaks at (2θ±0.2, 0 .3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 or 1.0°) 6.8°, 7.8°, 11.2°, 13.4 °, 13.7°, 16.0°, 17.1°, 17.8° and 23.2°. In other embodiments, the crystalline form of 5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1, 3,4-oxadiazol-2-yl)propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, or 161°C up to about 160, 161, 162, 163, 164, 165, 166, 167, 168, 169 or 170°C according to DSC. In some embodiments, the crystalline form of 5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1, 3,4-oxadiazol-2-yl)propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide has an endothermic peak from 155°C to 168°C, from 158°C to 162°C, from 159°C to 161°C, 159°C to 165°C, or 160°C to 163°C according to DSC. In additional embodiments, the crystalline form of 5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1, 3,4-oxadiazol-2-yl)propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, or 170°C according to DSC. In further embodiments, the optical purity of a given crystalline form is 100% ee.

[00049] In some embodiments, the invention relates to the crystalline form of 5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5- dihydro-1,3,4-oxadiazol-2-yl)propyl)-4-hydroxychroman-8-sulfonamide. In additional embodiments, the X-ray diffraction pattern of this crystal form has two or more, three or more, four or more, five or more, six or more, seven or more, eight or more, or nine peaks at (2θ±0.2, 0 .3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9 or 1.0°) 6.8°, 7.8°, 11.2°, 13.4 °, 13.7°, 16.0°, 17.1°, 17.8° and 23.2°. In other embodiments, the crystalline form of 5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3, 4-oxadiazol-2-yl)propyl)-4-hydroxychroman-8-sulfonamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, or 161°C to about 160, 161, 162, 163, 164, 165, 166, 167, 168, 169, or 170°C according to DSC. In some embodiments, the crystalline form of 5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3, 4-oxadiazol-2-yl)propyl)-4-hydroxychroman-8-sulfonamide has an endothermic peak from 155°C to 168°C, from 158°C to 162°C, from 159°C to 161°C, from 159 °C to 165°C or 160°C to 163°C according to DSC. In additional embodiments, the crystalline form of 5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3, 4-oxadiazol-2-yl)propyl)-4-hydroxychroman-8-sulfonamide has an endothermic peak of about 154, 155, 156, 157, 158, 159, 160, 161, 162, 163, 164, 165, 166, 167, 168, 169 or 170°C according to DSC. In further embodiments, the optical purity of a given crystalline form is 100% ee.

[00050] In some embodiments, the X-ray diffraction pattern of the crystalline form of the present invention has two or more peaks at (2Ɵ±0.2°) 6.8°, 7.8°, 11.2°, 13.4°, 13.7 °, 16.0°, 17.1°, 17.8° and 23.2°. In some embodiments, the implementation of the crystalline form has an endothermic peak from 155°C to 168°C according to DSC. In additional embodiments, the implementation of the crystalline form of the present invention may be characterized by a combination of the above described embodiments, which do not contradict each other. For example, the crystalline form may be characterized by a combination of the X-ray diffraction peaks described above and the DSC endothermic peak described above.

[00051] The term "about" used for the maximum temperature of the endothermic peak on the DSC curve means that the temperature value is in the range of ±2, 3, 4 or 5 °C from this value.

[00052] Unless otherwise indicated herein, the term "about" when used in connection with a numerical value or range of values that is given to characterize a particular solid form, such as a particular temperature or temperature range; weight change, such as, for example, weight change with temperature or humidity; the content of the solvent or water in terms of, for example, weight or percentage; or a peak position such as, for example, in an IR or Raman spectroscopy or XRPD analysis; indicate that the value or range of values may vary to an extent considered acceptable to a person skilled in the art, while still describing this particular solid form. For example, in specific embodiments, the term "about" when used in this context and, unless otherwise indicated, indicates that a numerical value or range of values may vary between 25, 20, 15, 10, 9, 8, 7, 6, 5, 4, 3, 2, 1, 0.5 or 0.25% of the stated value or value range.

[00053] In some embodiments, the crystalline form, co-crystal, or salt of benzoic acid described herein may comprise a plurality of crystals (crystalline polymorphs) having spatially regular atomic arrangements and different physicochemical properties. In additional embodiments, the crystalline form, co-crystal, or salt of benzoic acid described herein may be a mixture with crystalline polymorphs.

[00054] In some embodiments, in the crystalline form, co-crystal, or benzoic acid salt of the present invention, the molar ratio of sulfonamide compound to benzoic acid is from about 0.4 to about 1.6, from about 0.6 to about 1.4, from about 0.8 to about 1.2, about 0.9 to about 1.1, or about 1 to about 1. In additional embodiments, in the crystalline form, co-crystal, or benzoic acid salt of the present invention, the molar ratio of sulfonamide compound to benzoic acid is from about 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1, 1.1, 1.2, 1.3 to about 1.3, 1.2, 1.1, 1.0, 0.9, 0.8, 0.7, 0.6, 0.5 or 0.4. In other embodiments, the molar ratio of sulfonamide compound to benzoic acid is about 1:1.

[00055] In accordance with some embodiments of the crystalline forms, co-crystals or salts of benzoic acid of the present invention, the sulfonamide compound or benzoic acid may be substituted with one or more radioactive isotopes or non-radioactive isotopes.

[00056] In some embodiments, the optical purity of the crystalline form, co-crystal, or salt of benzoic acid described herein is at least about 70, 80, 85, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 99.5 or 99.8%. The term "optical purity" or "chiral purity" as used herein refers to the ratio of the observed optical rotation of a sample having a mixture of enantiomers to the optical rotation of a single pure enantiomer. In further embodiments, the optical purity is about 95% or greater. In other embodiments, the optical purity is about 98% or greater. In still other embodiments, the optical purity is about 100%. In some embodiments, optical purity may be measured by HPLC. In some embodiments, the present invention relates to methods for improving the optical purity of a sulfonamide compound described herein by mixing the sulfonamide compound with benzoic acid or by forming a crystalline form, co-crystal or salt of benzoic acid using the methods described herein.

[00057] In some embodiments, the crystalline form, co-crystal, or benzoic acid salt of a compound described herein may be physically and/or chemically pure. In some embodiments, the crystalline form, co-crystal, or benzoic acid salt of a compound described herein can be at least about 100, 99, 98, 97, 96, 95, 94, 93, 92, 91, 90, 89, 88, 87, 86, 85, 84, 83, 82, 81 or 80% physically and/or chemically pure. In some embodiments, the crystalline form, co-crystal, or benzoic acid salt of a compound described herein is substantially pure. As used herein and unless otherwise indicated, a sample containing a particular crystalline form or an amorphous form that is "substantially pure", such as essentially free of other solid forms and/or other chemical compounds, contains in particular less than about 25%, 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.75%, 0, 5%, 0.25% or 0.1% wt. one or more other solid forms and/or other chemical compounds.

[00058] In some embodiments, the crystalline form, co-crystal, or salt of benzoic acid described herein is stable at about 30, 40, 50, 60, 70, or 80°C and about 65, 70, 75, 80, or 85 %RH for about 1, 2, 3, 4, 5, 6, 7, 8, 12, 16, 24, 36, 48, 60, 72, 84, 96, 108, 120, 132, 144, or 156 weeks or more and for about 160, 156, 150, 138, 126, 114, 102, 90, 78, 66, 54, 42, 30, 24, 18, 12, or 6 weeks or less. The state can be a closed state or an open state. As used herein, the term "closed" may mean that the lid of the sample bottle is closed or sealed during the stability experiment, and "open" may mean that the lid is open. In additional embodiments, the crystalline form, co-crystal, or salt of benzoic acid described herein is stable at about 40°C and about 75% relative humidity for about 1, 2, 3, 4, 5, 6, 7, 8 , 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 or 22 weeks. In some embodiments, the state is a closed state. In other embodiments, the crystalline form, co-crystal, or salt of benzoic acid described herein is stable at about 40° C. and about 75% relative humidity for about 4 weeks. In some embodiments, the state is a closed state. Thus, the crystalline forms, co-crystals or salts of benzoic acid described herein exhibit excellent storage stability over a long period of time. Wherein, the term "stable" means that (i) the change in optical purity is about 1.0%, 0.5%, 0.3%, 0.1%, 0.05%, or 0.01% or less compared with the original optical purity, (ii) the increase in the amount of impurities is about 1.0%, 0.5%, 0.3%, 0.1%, 0.05% or 0.01% or less compared to the original the amount of impurities, and/or (iii) the X-ray diffraction pattern retains 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, or 90% or more of the initial peaks at (2θ±0.2°) .

[00059] Another aspect of the present invention relates to a process for preparing the crystalline form, co-crystal and/or salt of benzoic acid described herein, the method comprising: dissolving the benzoic acid and the sulfonamide compound described herein in a solvent system containing at least at least two solvents; and supersaturating the solvent system until a crystalline form, co-crystal and/or benzoic acid salt is formed and recovered from the solvent system. In another aspect, the present invention relates to a method for obtaining a crystalline form of a sulfonamide compound described herein from a non-crystalline form, the method comprising: dissolving benzoic acid and a sulfonamide compound in a solvent system containing at least two solvents; and supersaturating the solvent system until the product crystallizes out of the solvent system.

[00060] In some embodiments, the method may also include adding or mixing benzoic acid with a sulfonamide compound described herein. In additional embodiments, from about 0.8, 0.9, 1, 2, 3, 4, or 5 to about 1, 2, 3, 4, 5, 6, 7, 8, or 9 molar equivalents of benzoic acid relative to the sulfonamide compound may be mixed with a sulfonamide compound. In some embodiments, the method may also include adding or mixing about 0.8 to about 5, about 2 to about 5, about 3 to about 5, about 1 to about 4, or about 1 to 6 molar equivalents of benzoic acid with a sulfonamide compound as described herein. In further embodiments, about 1, 2, 3, 4, 5, 6, or 7 molar equivalents of benzoic acid relative to the sulfonamide compound may be mixed with the sulfonamide compound.

[00061] In additional embodiments, the dissolution step may include adding or mixing the solvent system with benzoic acid and the sulfonamide compound, wherein the solvent system is present in an amount of from about 10 to about 40 times, from about 20 to about 30 times, or from about 10 to about 30 times larger in volume, including about 10, 11, 12, 13, 14, 15, 18, 20, 22, 25, 27, 30, 35, 38, or 40 times. In other embodiments, the dissolution step may include adding a solvent system with benzoic acid and a sulfonamide compound, wherein the solvent system is present at about 10, 15, 20, 25, 30, 35, 40 times to about 60, 55, 50, 45 times , 40, 35 or 30 times larger in volume. In further embodiments, at least one of the at least two solvents in the solvent system is heptane or toluene. In other embodiments, the solvent system comprises heptane and toluene.

[00062] In other embodiments, in a solvent system or in a mixed solution of heptane and toluene, the proportion of toluene is from about 5, 6, 7, 8, 9, 10, or 11 to about 10, 15, 17, or 20%, including about 5, 8, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19 and 20%. In other embodiments, in the mixed solution of heptane and toluene, the proportion of toluene is 5 to 20%, 10 to 20%, 5 to 10%, or 8 to 15%.

[00063] In some embodiments, the supersaturation step includes heating the solvent system. In other embodiments, the supersaturation step comprises stirring the slurry from the dissolution step above room temperature, or from about 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 53 , 54, 55, or 56°C to about 50, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, or 66°C for about 1, 2, 3, 4, or 5 hours to about 4, 5, 6, 7, 8, 9, 10, 15, 20, 30, or 40 hours. In yet other embodiments, the supersaturation step comprises stirring the slurry at a temperature of from about 40°C to about 60°C, from about 50°C to about 60°C, from about 40°C to about 50°C, or from about 38°C. C to about 65°C for about 1 hour to about 40 hours, from about 3 hours to about 4 hours, from about 4 hours to about 5 hours, or from about 3 hours to about 10 hours. In additional embodiments, the step supersaturation involves stirring the slurry at a temperature of from about 40°C to about 60°C for about 3 hours to about 10 hours.

[00064] In some embodiments, the method for preparing the crystalline form, co-crystal and/or salt of benzoic acid described herein further comprises washing the product from the supersaturation step to remove remaining excess benzoic acid.

[00065] Although prodrugs of crystalline forms, cocrystals or salts of benzoic acid as described herein are also included in the present invention, the term "prodrug" refers to compounds that are converted to the compound described herein or its salt by reaction with an enzyme or gastric acid under physiological conditions in vivo , i.e. compounds that are converted to the compound of the present invention or a salt thereof by enzymatic oxidation, reduction or hydrolysis and the like, or compounds that are converted to the compound described herein or a salt thereof by gastric acid. In addition, a prodrug of a compound described here or a salt thereof may be a compound that is converted to a compound described here or a salt thereof under physiological conditions such as those described in Hirokawa Shoten 1990 annual "Development of Pharmaceuticals" Vol. 7, Molecular Design, pages 163-198.

[00066] In addition, the crystalline forms, co-crystals or salts of benzoic acid of the present invention may be solvates (eg, hydrates, etc.) or non-solvates, and both forms are covered by the compound described here or its salt. Compounds labeled with isotopes (eg, deuterium, ³ H, ¹⁴ C, ³⁵ S, ¹²⁵ I, etc.) and the like are also covered by the compounds described in this application, or their salt.

[00067] The crystalline form, co-crystal or salt of benzoic acid described herein exhibit RNR inhibitory activity. The crystalline form, co-crystal or salt of benzoic acid described herein is suitable as a drug for the prevention or treatment of RNR-related diseases without causing side effects based on off-target effects of protein-requiring iron ions due to its excellent RNR inhibitory activity and its structure and the absence of chelation of metal ions. "RNR-associated disease" includes diseases whose incidence can be reduced or whose symptoms are in remission or amelioration and/or cure by removing or suppressing and/or inhibiting RNR functions. Such diseases include, for example, malignant tumors. The malignancies considered are not particularly limited and may include head and neck cancer, gastrointestinal cancer (e.g., esophageal cancer, gastric cancer, duodenal cancer, liver cancer, bile duct cancer (e.g., gallbladder or biliary tract cancer, etc.). .d.), pancreatic cancer, colorectal cancer (e.g., colon cancer, rectal cancer, etc.), etc.), lung cancer (e.g., non-small cell lung cancer, small cell lung cancer, mesothelioma and etc.), breast cancer, genital cancer (e.g. ovarian cancer and uterine cancer (e.g. cervical cancer, endometrial cancer, etc.), etc.), genitourinary cancer (e.g. cancer kidney, bladder cancer, prostate cancer, testicular tumor, etc.), hematopoietic tissue tumors (for example, leukemia, malignant lymphoma, multiple myeloma, etc.), bone and soft tissue tumors, skin cancer, brain tumors brain and the like.

[00068] "RNR" as used herein includes human RNR or non-human RNR.

[00069] Accordingly, the present invention provides an RNR inhibitor that includes the crystalline form, co-crystal or salt of benzoic acid described herein as an active ingredient. In addition, the present invention relates to the use of the crystalline form, co-crystal or salt described herein for the preparation of RNR inhibitors. The present invention also provides the use of the crystalline form, co-crystal or salt of benzoic acid described herein as RNR inhibitors. In addition, the present invention provides the crystalline form, co-crystal or salt of benzoic acid described herein for use in the prevention or treatment of disorders associated with RNR inhibitors.

[00070] In another embodiment, the present invention provides a medicinal product containing the crystalline form, co-crystal or salt of benzoic acid described herein, as an active ingredient. In addition, the present invention relates to the use of the crystalline form, co-crystal or salt of benzoic acid described herein for the preparation of a drug. In addition, the present invention provides the use as drugs of the crystalline form, co-crystal or salt of benzoic acid described in this document. In addition, the present invention provides a crystalline form, co-crystal or salt of benzoic acid described herein for use as a medicine.

[00071] In yet another embodiment, the present invention provides a pharmaceutical composition comprising the crystalline form, co-crystal or salt of benzoic acid described herein and a pharmaceutically acceptable carrier. The crystalline form, co-crystal or salt of benzoic acid described herein has low or no hygroscopicity and low electrostatic charge storage capacity, resulting in excellent processing properties. As the hygroscopicity decreases, the problem of maintaining and controlling the moisture present in the storage state is solved, and during the manufacture of a solid preparation such as a tablet or capsule, the quality of the preparation (uniformity) can be easily controlled by weight control. In addition, since the ability to store electrostatic charge is low, adhesion to production equipment and packaging is negligible.

[00072] In some embodiments, the drug or pharmaceutical composition is a therapeutic agent for diseases associated with RNR, and in a more preferred embodiment, the drug or pharmaceutical composition is an antineoplastic agent.

[00073] In further embodiments, the present invention relates to administering an effective amount of the crystalline form, co-crystal, or salt of benzoic acid described herein to a subject in need thereof to provide a method of inhibiting RNR activity. In addition, the present invention includes administering an effective amount of the crystalline form, co-crystal, or salt of benzoic acid described herein to a subject to provide a method for preventing, ameliorating, or treating diseases associated with RNR. In some embodiments, the method for preventing, alleviating, or treating diseases associated with RNR is a method for preventing, alleviating, or treating tumors. In this method, the subjects include a human or non-human animals (eg, cows, pigs, horses, dogs, cats, and the like) in need of such a method. In additional embodiments, the implementation of the subject may be a subject suffering from a disease associated with the RNR described in this document.

[00074] When using a crystalline form, co-crystal or salt of benzoic acid described herein as a pharmaceutical agent, it is optionally formulated together with a pharmaceutically acceptable carrier, and various dosage forms can be selected in accordance with prophylactic or therapeutic purposes, and as dosage forms, for example, oral agents, injections, suppositories, ointments and any similar patches are used. Because the crystalline form, co-crystal or salt of benzoic acid described herein has excellent oral absorption, oral agents are preferred. These dosage forms can be obtained by conventional methods known in the art.

[00075] With regard to pharmaceutically acceptable carriers, various conventional organic or inorganic carrier substances are used as pharmaceutical materials, formulated as: excipients, binders, disintegrants, lubricants, colorants for solid preparations; and solvents, solubilizers, suspending agents, isotonizing agents, buffers, emollients for liquid formulations, and the like. In addition, pharmaceutical additives, which include preservatives, antioxidants, colors, sweeteners, flavors and stabilizers, can also be used if necessary.

[00076] With regard to pharmaceutically acceptable carriers and pharmaceutical additives, they usually include, for example, as an excipient: lactose, sucrose, sodium chloride, glucose, starch, calcium carbonate, kaolin, microcrystalline cellulose, silicic acid and the like; as binders: water, ethanol, propanol, simple syrup, glucose solution, starch solution, gelatin solution, carboxymethylcellulose, hydroxypropylcellulose, hydroxypropylstarch, methylcellulose, ethylcellulose, shellac, calcium phosphate, polyvinylpyrrolidone, and the like; as disintegrants: dry starch, sodium alginate, powdered agar, sodium hydrogen carbonate, calcium carbonate, sodium lauryl sulfate, stearic acid monoglyceride, lactose and the like; as lubricants: purified talc stearate, borax, polyethylene glycol and the like; as dyes: titanium oxide, iron oxide and the like; as flavorings: sucrose, orange peel, citric acid, tartaric acid and the like.

[00077] A solid preparation for oral use can be obtained by adding an excipient to the crystalline form, co-crystal or salt of benzoic acid described herein, and if necessary, binders, disintegrants, lubricants, coloring or flavoring agents and the like can also be added, followed by incorporation into tablets, coated tablets, granules, powders, capsules and the like.

[00078] Injectable forms can be prepared by adding pH adjusters, buffers, stabilizers, isotonics, local anesthetics, and the like to the crystalline form, co-crystal, or salt of benzoic acid described herein, followed by administration to subcutaneous, intramuscular, and intravenous formulations. injection forms in the usual way.

[00079] Upon receipt of a rectal suppository, the suppository can be made in the usual way after adding an excipient and, if necessary, a surfactant and the like to the active ingredient described herein. When prepared as an ointment such as a paste, cream and gel, the base, stabilizer, wetting agent, preservative and the like are mixed as needed and prepared in a conventional manner. For example, white petrolatum, paraffin, glycerin, cellulose derivatives, polyethylene glycol, silicone, bentonite, and the like can be used as the base. As preservatives, methyl parahydroxybenzoate, ethyl parahydroxybenzoate, propyl parahydroxybenzoate and the like can be used. In the case of making a patch, an ointment, cream, gel, paste or the like can be applied to the support in a known manner. As the substrate, a woven fabric made of cotton, staple fiber, synthetic fiber, non-woven fabric, soft vinyl chloride film, polyethylene, polyurethane or the like, or foam sheet can be used.

[00080] The amount of crystalline form, co-crystal or salt of benzoic acid described herein to be included in each unit dosage form described above may be, in general, per unit dosage form, from about 0.05 mg, 0 .1 mg, 1 mg, 5 mg, 10 mg or 20 mg up to about 100 mg, 500 mg or 1000 mg for an oral formulation, from about 0.01 mg or 0.1 mg to about 200 mg, 300 mg or 500 mg for an injectable form, and from about 1 mg, 5 mg, or 10 mg to about 100 mg, 500 mg, or 1000 mg for suppositories, provided that these amounts may be varied depending on the symptoms of the patients in whom they are to be used, or from their dosage forms.

[00081] In addition, the daily dose of the drug with dosage form is in relation to the crystalline form, co-crystal or salt described here from 0.05 mg per day to 5000 mg per day for an adult (body weight 50 kg), preferably from 0. 1 mg to 2000 mg, and preferably the above amounts are administered one or two to three times a day, provided that they can be changed depending on the patient's symptoms, weight, age or sex, and the like.

[00082] Similar substances described in Fig.10 - Fig.12 are also included in this application. Similar substances of the present invention can be synthesized by known methods or can be obtained from commercially available products. Similar substances can be identified by comparing high performance liquid chromatography retention times, mass spectra and analysis using a photodiode array detector (PDA) between similar substances thus obtained and similar substances detected in accordance with the present invention.

[00083] In addition, these analogous substances can be quantified by an external standard method or an internal standard method.

[00084] When these analogous substances are possibly contained as impurities in a medicinal product or pharmaceutical preparation, these analogous substances are regulated in accordance with ICH-Q3A guidance of the International Council for the Harmonization of Specifications for Pharmaceutical Preparations for Human Use. The method of the present invention is very useful because it is possible to confirm compliance with the guideline standard.

[00085] Similar substances are listed in the following table.

[00086] [Table 1]

Name Chemical structure OPC:
(2S,3R)-2-((2-carbamoyl-4-chlorophenyl)sulfonamido)-3-(6-fluoro-2,3-dimethylphenyl)butanoic acid

SAC:
5-((1S,2R)-1-(5-chloro-1,1-dioxido-3-oxobenzo[d]isothiazol-2(3H)-yl)-2-(6-fluoro-2,3-dimethylphenyl )propyl)-1,3,4-oxadiazol-2(3H)-one

O.P.H.:
5-chloro-2-(N-((2S,3R)-3-(6-fluoro-2,3-dimethylphenyl)-1-hydrazinyl-1-oxobutan-2-yl)sulfamoyl)benzamide

deme:
5-chloro-2-(N-((1S,2R)-2-(2-fluoro-5-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole-2 -yl)propyl)sulfamoyl)benzamide

OPD1

OPHD

Examples

[00087] The present invention, in accordance with some embodiments, is described below in more detail using experimental examples, but the present invention is not intended to be limited to these examples.

[00088] The various reagents used in the examples were commercially available products unless otherwise stated. Column Biotage Ltd. SNAP-ULTRA (registered trademark) packed with silica was used for silica gel column chromatography, or a column manufactured by Biotage SNAP KP-C18-HS (registered trademark) packed with silica was used for reverse phase silica gel column chromatography. HPLC purification by preparative reverse phase column chromatography was performed under the following conditions. The injection volume and gradient were adjusted accordingly.

Column: YMC-Actus Triart C18, 30 × 50 mm, 5 µm

UV detection: 254 nm

Column flow rate: 40 ml/min

Mobile phase: water/acetonitrile (0.1% formic acid)

Injection volume: 1.0 ml

Gradient: Water/Acetonitrile (10% to 90%)

[00089] AL400 (400MHz; JEOL (JEOL)) and Mercury400 (400MHz; Agilent Technologies) were used for NMR spectra, and tetramethylsilane was used as an internal standard when tetramethylsilane was included in the heavy solvent, otherwise the determination was carried out using NMR solvent as an internal standard, giving all δ values in ppm. In addition, LCMS spectra were determined under the following conditions using a Waters ACQUITY SQD spectrometer (quadrupole).

Column: Waters ACQUITY UPLC (registered trademark) BEH C18, 2.1 × 50 mm, 1.7 µm

MS detection: with negative ESI

UV detection: 254 nm and 280 nm

Column flow rate: 0.5 ml/min

Mobile phase: water/acetonitrile (0.1% formic acid)

Injection volume: 1 µl

[Table 2] Gradient

Time (min) Water Acetonitrile 0 95 5 0.1 95 5 2.1 5 95 3.0 STOP

Abbreviations have the meanings given below.

c: singlet

e: doublet

t: triplet

square: quartet

dd: doublet of doublets

dt: doublet of triplets

td: triplet of doublets

tt: triplet of triplets

ddd: doublet of doublets of doublets

ddt: doublet of doublets of triplets

dtd: doublet of triplets of doublets

tdd: triplet of doublets of doublets

m: multiplet

wide: wide

brs: widened singlet

DMSO-d ₆ : deuterated dimethyl sulfoxide

CDCl ₃ : heavy chloroform

CD ₃ OD: heavy methanol

CDI: 1,1'-carboxymethylsulfonyldiimidazole

DAST: N,N-diethylaminosulfur trifluoride

DIBAL-H: diisobutylaluminum hydride

DMF: dimethylformamide

DMSO: dimethyl sulfoxide

THF: tetrahydrofuran

WSC=EDCI=1-ethyl-3-(3-dimethylaminopropyl)carbodiimide

HOBt=1-hydroxybenzotriazole

[00090] Reference Example A1: 2-(1-bromoethyl)-1-fluoro-3,4-dimethylbenzene

[00091] [Formula I]

[00092] (Step 1) 1-(6-fluoro-2,3-dimethylphenyl)ethanol

[00093] After adding dropwise a solution of methylmagnesium bromide in diethyl ether (3.0 M, 70 ml) to a THF solution of 6-fluoro-2,3-dimethylbenzaldehyde (22.0 g) (300 ml) at 0°C, the reaction the mixture was stirred at room temperature for 1 hour. Saturated aqueous ammonium chloride solution (150 ml) was added dropwise under ice bath conditions, and ethyl acetate (200 ml) was added, and the resulting product was separated into different layers. The organic layer was washed successively with HCl (1M, 200 ml), water (200 ml), and brine (200 ml), and then dried over anhydrous magnesium sulfate and concentrated under reduced pressure to give 1-(6-fluoro-2,3 -dimethylphenyl)ethanol (23.7 g).

[00094] (Step 2) Phosphorus tribromide (26.5 ml) was added dropwise at 0°C to a chloroform (120 ml) solution of 1-(6-fluoro-2,3-dimethylphenyl)ethanol (23.7 g) obtained in the above step 1, and the reaction solution was stirred for 30 min at 0°C. The reaction mixture was added to an ice-bath-cooled saturated aqueous sodium hydrogen carbonate solution (1 L). After adding chloroform (500 ml) to the mixture, the resulting product was separated into different layers, and the organic layer was washed successively with water (200 ml) and brine (200 ml). The organic layer was dried over anhydrous magnesium sulfate to obtain the title compound (29.5 g) by concentration under reduced pressure.

[00095] Reference Example A2 and A3: Aldehyde and methylmagnesium bromide were reacted with each other as a starting material in the same manner as in Reference Example A1, Step 1 and Step 2, and then the reaction product was reacted with phosphorus tribromide to obtain a compound Reference Example A2 and A3 shown below.

[00096] [Table 3]

Reference Example Raw material synthesized compound A2

A3

[00097] Reference Example A4: 2-(1-bromoethyl)-4-ethyl-1-fluoro-3-methylbenzene

[00098] [Formula II]

[00099] (Step 1) 2-bromo-3-ethyl-6-fluorobenzaldehyde

[000100] To a THF (150 ml) solution of 2-bromo-1-ethyl-4-fluorobenzene (14.4 g) was added dropwise a THF solution of lithium diisopropylamide (1.5 M, 54 ml) at -78°C. After the reaction solution was stirred for 30 minutes, DMF (6.5 ml) was added, and the mixture was further stirred for 20 minutes. Water (50 ml) and hydrochloric acid (6 M, 50 ml) were added dropwise successively to the reaction solution, and the mixture was extracted twice with hexane (100 ml). The combined organic layer was washed twice with brine (50 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure to give 2-bromo-3-ethyl-6-fluorobenzaldehyde (14.5 g).

[000101] (Step 2) 3-ethyl-6-fluoro-2-methylbenzaldehyde

[000102] Water (90 ml), tripotassium phosphate (32.0 d), methylboronic acid (6.4 g) and the addition of [bis(diphenylphosphino)ferrocene]palladium(II) dichlorodichloromethane (1.75 g), and the reaction solution was refluxed at 110° C. for 2 hours. allowed to cool to room temperature and the mixture was further stirred for 2 h after adding hexane (90 ml). The reaction solution was filtered through CELITE, and the filtrate was separated after the residue was washed with hexane. The organic layer was washed twice with saturated brine (100 ml) and, after drying over anhydrous sodium sulfate, concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: hexane/ethyl acetate) to give 3-ethyl-6-fluoro-2-methylbenzaldehyde (8.4 g).

[000103] (Step 3) In accordance with the methods described in Reference Example A1, Steps 1 and 2, using 3-ethyl-6-fluoro-2-methylbenzaldehyde (8.4 g) obtained in the above Step 2, carried out the same operation to obtain the compound of Reference Example A4.

[000104] Reference B1: (2S,3R)-2-amino-3-(6-fluoro-2,3-dimethylphenyl)butanoic acid

[000105] [Formula III]

[000106] The DMF solution (50 ml) of 2-(1-bromoethyl)-1-fluoro-3,4-dimethylbenzene (14.0 g) obtained in Reference Example A1 was added dropwise to the DMF solution (50 ml) ( S)-2-[o-[(N-benzylprolyl)amino]phenyl]benzylideneamino-acetate-(2-)-N,N,N-nickel(II) (14.5 g) and potassium hydroxide (16.3 d), and the mixture was stirred at the same temperature for 1 hour. A saturated ammonium chloride solution (50 ml) and ethyl acetate (50 ml) were added to the reaction solution, the layers were separated, and the aqueous layer was extracted twice with ethyl acetate (50 ml). The combined organic layers were washed successively with water (50 ml) and brine (50 ml), dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: ethyl acetate/hexane). The resulting compound was dissolved in methanol (120 ml), hydrochloric acid (3 M, 90 ml) was added and the mixture was stirred at 80°C for 45 min. Methanol was distilled off under reduced pressure, and chloroform (50 ml) and water (50 ml) were added to the residue. The aqueous layer was washed with chloroform (50 ml) and concentrated under reduced pressure. The residue was purified by reverse phase silica gel column chromatography (methanol/water) to give the title compound (2.0 g). ¹ H NMR (CD ₃ OD) δ: 7.03 (dd, J=8.2, 5.7 Hz, 1H), 6.79 (dd, J=11.7, 8.4 Hz, 1H), 3.74-3.87 (m, 2H), 2.29 (s, 3H), 2.25 (s, 3H), 1.40 (dd, J=6.8, 2.4Hz, 3H)

[000107] Reference Examples B2-B4:

[000108] After the reaction of the alkylating agent obtained in Reference Examples A2-A4 and (S)-2-[o-[(N-benzylprolyl)amino]phenyl]benzylideneaminoacetate-(2-)-N,N,N-nickel (II) the compounds of Reference Examples B2-B4 shown below were obtained by acid hydrolysis.

[000109] [Table 4]

Reference Example Raw material
(Reference example number or structural formula) synthesized compound B2 A2

B3 A3

B4 A4

[000110] Reference C1: 5-chloro-8-(chlorosulfonyl)-4-methyl-d3-chroman-4-yl acetate

[000111] [Formula IV]

[000112] (Step 1) 8-bromo-5-chloro-4-methylchroman-4-ol

[000113] THF (50 ml) was added to a diethyl ether (1.0 M, 63 ml) solution of methyl d3-magnesium iodide and a THF solution (50 ml) of 8-bromo-5-chlorochroman-4-one ( 7.5 g) was added dropwise at room temperature. The reaction solution was stirred for 10 minutes at the same temperature, and the layer was separated. In an ice bath, hydrochloric acid (1 M, 50 ml) was slowly added dropwise, and ethyl acetate (50 ml) was added in separate layers. The aqueous layer was extracted with ethyl acetate (50 ml) and the combined organic layer was washed with brine (50 ml), dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: hexane/ethyl acetate) to give 8-bromo-5-chloro-4-methylchroman-4-ol (7.7 g).

[000114] (Step 2) 8-bromo-5-chloro-4-methyl-d3-chroman-4-yl acetate

[000115] An acetonitrile solution (12 ml ) scandium(III) trifluoromethanesulfonate (340 mg) at -40°C, and the reaction solution was stirred for 30 minutes at the same temperature. Saturated aqueous sodium hydrogen carbonate solution (100 ml) and ethyl acetate (100 ml) were added successively to the reaction solution, and the layers were separated. The aqueous layer was extracted with ethyl acetate (100 ml) and the combined organic layers were washed twice with saturated aqueous sodium hydrogen carbonate (100 ml) and once with brine (100 ml). The organic layer was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: hexane/ethyl acetate) to give 8-bromo-5-chloro-4-methyl-d3-chroman-4-ylacetate (8.9 g).

[000116] (Step 3) 8-(benzylthio)-5-chloro-4-methyl-d3-chroman-4-yl acetate

[000117] 4 ,5-bis(diphenylphosphino)-9,9-dimethylxanthene (600 mg), tris(dibenzylideneacetone)dipalladium (0) (480 mg), N,N-diisopropylethylamine (7.2 ml) and benzyl mercaptan (2.8 ml) , and the reaction solution was stirred for 2 hours at 90°C. The reaction solution was allowed to cool to room temperature and filtered through CELITE. After washing the residue with hexane (50 ml), water (50 ml) was added to the filtrate and the layer was separated. The organic layer was washed with brine (50 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The reaction solution was concentrated under reduced pressure, and the residue was purified by silica gel column chromatography (eluent: hexane/ethyl acetate) to obtain 8-(benzylthio)-5-chloro-4-methyl-d3-chroman-4-ylacetate (6.3 g) .

[000118] (Step 4) To an acetonitrile solution (100 ml) of 8-(benzylthio)-5-chloro-4-methyl-d3-chroman-4-yl acetate (6.3 g) obtained in the above Step 3, water (3 ml), acetic acid (4.3 ml) and 1,3-dichloro-5,5-dimethylhydantoin (7.2 g) were added at 0° C., and the reaction solution was stirred for 30 minutes at the same temperature . A saturated aqueous solution of sodium hydrogen carbonate (70 ml) and ethyl acetate (70 ml) were added to the reaction solution, and the layers were separated. The aqueous layer was extracted with ethyl acetate (70 ml). The combined organic layers were washed with brine (70 ml), dried over anhydrous sodium sulfate, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: hexane/ethyl acetate) to give the title compound (5.3 g).

[000119] Reference Examples C2-C4: According to the method of Reference Example C1, steps 1-4, the compounds of Reference Example C2 were synthesized with the starting materials listed in the following table. According to the method of Reference Examples C1, steps 3 and 4, the compounds of Reference Examples C3 and C4 were synthesized.

[000120] [Table 5]

Reference Example Raw material synthesized compound C2

C3

C4

[000121] Reference D1: 5-((1S,2R)-1-amino-2-(6-fluoro-2,3-dimethylphenyl)propyl)-1,3,4-oxadiazole-2-(3H)- he monohydrochloride

[000122] [Formula V]

[000123] (Step 1) (2S,3R)-2-((tert-butoxycarbonyl)amino)-3-(6-fluoro-2,3-dimethylphenyl)butanoic acid

[000124] Water (9 ml), 1,4-dioxane (9 ml) and triethylamine (955 μl) were added sequentially to (2S,3R)-2-amino-3-(6-fluoro-2,3-dimethylphenyl) butanoic acid (515 mg) obtained in Reference Example B1, and the mixture was cooled to 0°C. Di-tert-butyl dicarbonate (650 mg) was added to the reaction solution at the same temperature, and the mixture was stirred for 45 minutes. The reaction solution was added to hydrochloric acid (1 M, 20 ml) and extracted with ethyl acetate (20 ml). The organic layer was washed with brine (20 ml), dried over anhydrous magnesium sulfate, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: hexane/ethyl acetate/2% acetic acid) to give (2S,3R)-2-((tert-butoxycarbonyl)amino)-3-(6-fluoro-2,3-dimethylphenyl) butanoic acid (745 mg).

[000125] (Step 2) tert-butyl-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4 -oxadiazol-2-yl)propyl)carbamate

[000126] To a THF solution (14.0 ml) of (2S,3R)-2-(tert-butoxycarbonylamino)-3-(6-fluoro-2,3-dimethylphenyl)butanoic acid (440 mg) above step 1, CDI (302 mg) was added and the reaction solution was stirred at room temperature for 20 minutes. The reaction solution was cooled to 0°C, hydrazine monohydrate (200 μl) was added, and the mixture was stirred at the same temperature for 30 minutes. The reaction solution was added to water (20 ml) and extracted with ethyl acetate (20 ml). The organic layer was washed with brine (20 ml), dried over anhydrous sodium sulfate and concentrated under reduced pressure. CDI (560 mg) was added to a 1,4-dioxane solution (14 ml) of the resulting residue, and the reaction solution was stirred at room temperature for 30 minutes. The reaction solution was added to water (20 ml) and extracted with ethyl acetate (20 ml). The organic layer was washed with brine (20 ml), dried over anhydrous sodium sulfate and concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography (eluent: hexane/ethyl acetate) to give t-butyl-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4, 5-dihydro-1,3,4-oxadiazol-2-yl)propyl)carbamate (356 mg).

[000127] (Step 3) tert-butyl-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4 -oxadiazol-2-yl)propyl)carbamate (550 mg) obtained in the above Step 2 was dissolved in hydrochloric acid-1,4-dioxane (4 M, 5.0 ml), and the reaction solution was stirred at room temperature in over 1.5 hours. The reaction solution was concentrated under reduced pressure to give the title compound.

[000128] Reference Example D2 and D3: According to the method described in Reference Example D1, steps 1-3, the following compounds of Reference Examples D2 and D3 were synthesized.

[000129] [Table 6]

Reference Example Raw material
(Reference example number or structural formula) synthesized compound D2 Reference Example B2

D3 Reference Example B3

[000130] Example 1: 5-bromo-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1 ,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide

[000131] (Step 1) To a solution in 1,4-dioxane (5.0 ml) and water (5.0 ml) (2S,3R)-2-amino-3-(6-fluoro-2,3- dimethylphenyl)butanoic acid (300 mg) obtained in Reference Example B1 was added with triethylamine (570 µl) and then cooled to 0°C. 4-Bromo-2-cyanobenzene-1-sulfonyl chloride (362 mg) was added to the reaction solution, and the mixture was stirred at the same temperature for 45 minutes. The reaction solution was added to hydrochloric acid (1 M, 15 ml) and extracted with ethyl acetate (15 ml). The organic layer was washed with brine (20 ml), dried over anhydrous magnesium sulfate, and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: hexane/ethyl acetate/2% acetic acid) to give (2S,3R)-2-(4-bromo-2-cyanophenylsulfonamido)-3-(6-fluoro-2,3-dimethylphenyl ) butanoic acid (465 mg).

[000132] (Step 2) To THF solution (5.0 ml) of (2S,3R)-2-(4-bromo-2-cyanophenylsulfonamido)-3-(6-fluoro-2,3-dimethylphenyl)butanoic acid (465 mg) obtained in the above Step 1 was added CDI (210 mg) and the reaction solution was stirred at room temperature for 20 minutes. The reaction solution was cooled to 0°C, hydrazine monohydrate (200 μl) was added, and the mixture was stirred at the same temperature for 20 minutes. The reaction solution was added to water (20 ml) and extracted with ethyl acetate (20 ml). The organic layer was washed with brine (20 ml), dried over anhydrous sodium sulfate and concentrated under reduced pressure. CDI (211 mg) was added to a 1,4-dioxane solution (4.0 ml) of the obtained residue, and the reaction solution was stirred at 45°C for 1 hour. The reaction solution was added to water (20 ml) and extracted with ethyl acetate (20 ml ). The organic layer was washed with brine (20 ml), dried over anhydrous sodium sulfate and concentrated under reduced pressure. The resulting residue was purified by silica gel column chromatography (eluent: hexane/ethyl acetate) to give 4-bromo-2-cyano-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-( 5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)propyl)benzenesulfonamide (386 mg).

[000133] (Step 3) In DMSO solution (5.0 ml) 4-bromo-2-cyano-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1- Aqueous hydrogen peroxide (1.0 ml) and potassium carbonate (420 mg), and the reaction solution was stirred at 60°C for 2.5 hours. The reaction solution was slowly added to hydrochloric acid (1 M, 15 ml) in an ice bath, and then extracted with ethyl acetate (15 ml) . The organic layer was washed with brine (20 ml), dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by silica gel column chromatography (eluent: hexane/ethyl acetate) to give the title compound.

[000134] Examples 2-4

[000135] The compounds of examples 2-4 were synthesized in accordance with the method of example 1, steps 1-3. Required starting materials are listed in the following table.

[000136] [Table 7]

Example Raw material _ArSO2Cl Name of the synthesized compound 2 Reference Example B1

5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole -2-yl)propyl)sulfamoyl)benzamide 3 Reference Example B4

5-chloro-2-(N-((1S,2R)-2-(3-ethyl-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4 -oxadiazol-2-yl)propyl)sulfamoyl)benzamide four Reference Example B2

5-chloro-2-(N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4 -oxadiazol-2-yl)propyl)sulfamoyl)benzamide

[000137] Example 5: 5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3, 4-oxadiazol-2-yl)propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide, isomer A and isomer B

[000138] (Step 1) 5-chloro-8-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro- 1,3,4-oxadiazol-2-yl) propyl) sulfamoyl)-4-methyl-d3-chroman-4-yl acetate

[000139] To a pyridine solution (1.5 ml) of 5-((1S,2R)-1-amino-2-(6-fluoro-2,3-dimethylphenyl)propyl)-1,3,4-oxadiazole-2 (3H)-one monohydrochloride (45 mg) obtained in Reference Example D1 was added 5-chloro-8-(chlorosulfonyl)-4-methyl-d3-chroman-4-ylacetate (80 mg) obtained in Reference Example C1, and the reaction solution was stirred at room temperature for 12 hours. The reaction solution was concentrated under reduced pressure, and the resulting residue was purified by silica gel column chromatography (eluent: hexane/ethyl acetate) to obtain the title compound (59 mg) as a diastereomeric mixture 1: one.

[000140] (Step 2) 1:1 diastereomeric mixture of 5-chloro-8-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4 ,5-dihydro-1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)-4-methyl-d3-chroman-4-ylacetate (59 mg) obtained from step 1 above was dissolved in methanol (2, 0 ml) and water (1.0 ml), lithium hydroxide (5 mg) was added, and the reaction solution was stirred at 55°C for 1 hour. After the reaction solution was concentrated, hydrochloric acid (1 M, 10 ml) was added to the residue and ethyl acetate (10 ml) and the layers were separated. The aqueous layer was extracted with ethyl acetate (10 ml) and the combined organic layers washed with brine (10 ml). The organic layer was dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by reverse phase HPLC (water/acetonitrile) and the fractions were concentrated to give each of two diastereomeric products. The substance eluted first was designated Compound A, and the substance eluted later was designated Compound B.

[000141] Examples 6-8: Following the method of Example 5, Step 2, the following compounds of Examples 6-8 were synthesized. In the case of separation of diastereomers, the compound eluted earlier was designated as A and the compound eluted later as B. The ratio of diastereomers of the mixture was 1:1, unless otherwise indicated. Required starting materials are listed in the following table.

[000142] [Table 8]

Example Raw material _ArSO2Cl Name of the synthesized compound 6A and 6B Reference Example D2 Reference Example C2 5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole- 2-yl) propyl)-4-hydroxy-4-methylchroman-8-sulfonamide 7A and 7B Reference Example D3 Reference Example C2 N-((1S,2R)-2-(3-bromo-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl) propyl)-5-chloro-4-hydroxy-4-methylchroman-8-sulfonamide 8A and 8B Reference Example D2 Reference Example C1 5-chloro-N-((1S,2R)-2-(3-chloro-6-fluoro-2-methylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole- 2-yl) propyl)-4-hydroxy-4-methyl-d3-chroman-8-sulfonamide

[000143] Example 9: 5-fluoro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3, 4-oxadiazol-2-yl)propyl)-4-hydroxychroman-8-sulfonamide, isomer A and isomer B

[000144] (Step 1) When using 5-((1S,2R)-1-amino-2-(6-fluoro-2,3-dimethylphenyl)propyl)-1,3,4-oxadiazole-2(3H )-one monohydrochloride (40 mg) obtained from Reference Example D1 and 5-fluoro-4-oxochroman-8-sulfonyl chloride (60 mg) obtained from Reference Example C3 gave 5-fluoro-N-((1S, 2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)propyl)-4-oxochroman-8 -sulfonamide (44 mg) according to the method of example 5, step 1.

[000145] (Step 2) Sodium borohydride (13.5 mg) was added to an ethanol solution (2.0 ml) of 5-fluoro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl )-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)propyl)-4-oxochroman-8-sulfonamide (44 mg) obtained from step 1 above, and reaction the solution was stirred at room temperature for 30 minutes. After the reaction solution was concentrated under reduced pressure, water (10 ml) and ethyl acetate (10 ml) were added to the residue, separated, and the aqueous layer was extracted with ethyl acetate (10 ml). The combined organic layer was washed with brine (10 ml), dried over anhydrous sodium sulfate and concentrated under reduced pressure. The residue was purified by reverse phase HPLC (water/acetonitrile) and the fractions were concentrated to give each of two diastereomeric products. The substance eluted first was designated Compound A, and the substance eluted later was designated Compound B.

[000146] Example 10: Following the method of Example 5 Step 1 and Example 9 Step 2, the compounds of Example 10 shown below were synthesized. In the case of separation of diastereomers, the compound eluted first was designated A and the compound eluted later was designated B. The ratio of diastereomers of the mixture was 1:1 unless otherwise indicated. Required starting materials are listed in the following table.

[000147] [Table 9]

Example Raw material _ArSO2Cl Name of the synthesized compound 10A and 10B Reference Example D1 Reference Example C4 5-chloro-N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazole-2- yl) propyl)-4-hydroxychroman-8-sulfonamide

[000148] As a comparative example, a compound of the following formula was prepared.

[000149] [Formula VI]

[000150] ¹ H NMR (CD ₃ OD) δ: 7.54 (d, J=8.4 Hz, 2H), 7.17-7.29 (m, 5H), 7.08-7.14 ( m, 2H), 4.55-4.61 (m, 1H), 3.00-3.13 (m, 2H), 2.39 (s, 3H)

[000151] The following are the structural formulas and physical properties of the compounds of Examples 1-10.

[000152] [Table 10]

Example Structural formula Values of physical properties one

¹ H NMR (CD ₃ OD) δ: 7.74-7.78 (m, 3H), 6.97 (dd, J=8.2, 5.7 Hz, 1H), 6.71 (dd, J =11.7, 8.4 Hz, 1H), 4.78-4.81 (m, 1H), 3.51-3.61 (m, 1H), 2.20 (s, 3H), 2, 17 (s, 3H), 1.44 (d, J=7.0 Hz, 3H); LC/MS RT 1.67 min, m/z [MH] ^- 525, 527 2

¹ H NMR (CD ₃ OD) δ: 7.84 (d, J=8.4 Hz, 1H), 7.62 (d, J=2.2 Hz, 1H), 7.58 (dd, J= 8.4, 2.2 Hz, 1H), 6.98 (dd, J=8.2, 5.7 Hz, 1H), 6.72 (dd, J=11.7, 8.4 Hz, 1H ), 4.82 (d, J=11.4 Hz, 1H), 3.50-3.60 (m, 1H), 2.20 (s, 3H), 2.17 (s, 3H), 1 .45 (d, J=7.0 Hz, 3H); LC/MS RT 1.65 min, m/z [MH] ^- 481, 483 3

¹ H NMR (CD ₃ OD) δ: 7.84 (d, J=8.4 Hz, 1H), 7.62 (d, J=2.2 Hz, 1H), 7.58 (dd, J= 8.4, 2.2 Hz, 1H), 6.98 (dd, J=8.4, 5.9 Hz, 1H), 6.75 (dd, J=11.7, 8.8 Hz, 1H ), 4.78 (d, J=11.0 Hz, 1H), 3.50-3.60 (m, 1H), 2.52-2.59 (m, 2H), 2.24 (s, 3H), 1.46 (d, J=7.0 Hz, 3H), 1.06 (t, J=7.5 Hz, 3H); LC/MS RT 1.73 min, m/z [MH] ^- 495, 497 four

¹ H NMR (CD ₃ OD) δ: 7.86 (d, J=8.4 Hz, 1H), 7.63 (d, J=1.8 Hz, 1H), 7.59 (dd, J= 8.4, 2.2 Hz, 1H), 7.25 (dd, J=8.8, 5.1 Hz, 1H), 6.88 (t, J=10.0 Hz, 1H), 4, 80 (d, J=11.4 Hz, 1H), 3.55-3.65 (m, 1H), 2.37 (s, 3H), 1.47 (d, J=7.0 Hz, 3H ); LC/MS RT 1.68 min, m/z [MH] ^- 501, 503 5A

¹ H NMR (CD ₃ OD) δ: 7.61 (d, J=8.4 Hz, 1H), 7.02 (d, J=8.5 Hz, 1H), 6.96 (dd, J= 8.2, 5.7 Hz, 1H), 6.70 (dd, J=11.7, 8.4 Hz, 1H), 4.71 (d, J=11.4 Hz, 1H), 4, 40-4.46 (m, 1H), 4.26 (td, J=10.8, 2.6 Hz, 1H), 3.62-3.71 (m, 1H), 2.23 (s, 3H), 2.17 (s, 3H), 2.05-2.13 (m, 2H), 1.50 (d, J=7.0 Hz, 3H); LC/MS RT 1.64 min, m/z [MH] ^- 527, 529 5B

¹ H NMR (CD ₃ OD) δ: 7.59 (d, J=8.8 Hz, 1H), 7.02 (d, J=8.4 Hz, 1H), 6.97 (dd, J= 8.4, 5.9 Hz, 1H), 6.71 (dd, J=11.7, 8.4 Hz, 1H), 4.75 (d, J=11.4 Hz, 1H), 4, 41-4.48 (m, 1H), 4.32 (td, J=10.7, 2.7 Hz, 1H), 3.63-3.73 (m, 1H), 2.22 (s, 3H), 2.18 (s, 3H), 2.05-2.13 (m, 2H), 1.48 (d, J=7.0 Hz, 3H); LC/MS RT 1.71 min, m/z [MH] ^- 527, 529 6A

¹ H-NMR (CDCl ₃ ) δ: 8.15 (1H, s), 7.66 (1H, d, J=8.4 Hz), 7.19 (1H, dd, J=8.6.5 .1 Hz), 6.99 (1H, d, J=8.6 Hz), 6.78 (1H, dd, J=10.8, 9.0 Hz), 5.52 (1H, d, J =11.0 Hz), 4.87(1H, t, J=10.4 Hz), 4.47-4.44(1H, m), 4.28-4.25(1H, m), 3 .48 (1H, s), 3.29 (1H, s), 2.37 (3H, s), 2.32-2.28 (1H, m), 2.09-2.06 (1H, m ), 1.78 (3H, s), 1.54 (3H, d, J=7.0 Hz); LC/MS RT 1.68 min, m/z [MH] ^- 544, 546 6B

¹ H-NMR (CDCl ₃ ) δ: 8.68 (1H, br s), 7.64 (1H, d, J=8.8 Hz), 7.19 (1H, dd, J=8.8 , 4.9 Hz), 7.01 (1H, d, J=8.8 Hz), 6.78 (1H, dd, J=10.8, 8.8 Hz), 5.47-5.42 (1H, m), 4.81 (1H, t, J=10.9 Hz), 4.45-4.42 (1H, m), 4.32 (1H, t, J=10.9 Hz) , 3.53(1H, br s), 3.40(1H, br s), 2.35(3H, s), 2.33-2.27(1H, m), 2.15-2 .10(1H, m), 1.78(3H, s), 1.59-1.58(3H, m); LC/MS RT 1.74 min, m/z [MH] ^- 544, 546 7A

¹ H NMR (CD ₃ OD) δ: 7.61 (d, J=8.4 Hz, 1H), 7.42 (dd, J=9.0, 5.3 Hz, 1H), 7.03 ( d, J=8.4 Hz, 1H), 6.81 (dd, J=11.2, 9.0 Hz, 1H), 4.69 (d, J=11.4 Hz, 1H), 4, 39-4.47(m, 1H), 4.22-4.33(m, 1H), 3.63-3.78(m, 1H), 2.44(s, 3H), 2.17- 2.24 (m, 1H), 2.05-2.15 (m, 1H), 1.75 (s, 3H), 1.52 (d, J=6.6 Hz, 3H); LC/MS RT 1.70 min, m/z [MH] ^- 588, 590 7B

¹ H NMR (CD ₃ OD) δ: 7.60 (d, J=8.6 Hz, 1H), 7.43 (dd, J=8.9, 5.1 Hz, 1H), 7.02 ( d, J=8.6 Hz, 1H), 6.82 (dd, J=11.2, 8.9 Hz, 1H), 4.74 (d, J=11.4 Hz, 1H), 4, 39-4.47 (m, 1H), 4.33 (td, J=10.8, 2.6 Hz, 1H), 3.65-3.77 (m, 1H), 2.44 (s, 3H), 2.22-2.31 (m, 1H), 2.05-2.12 (m, 1H), 1.75 (s, 3H), 1.50 (d, J=7.0 Hz , 3H); LC/MS RT 1.76 min, m/z [MH] ^- 588, 590 8A

¹ H NMR (CD ₃ OD) δ: 7.61 (d, J=8.4 Hz, 1H), 7.24 (dd, J=8.8, 5.1 Hz, 1H), 7.03 ( d, J=8.8 Hz, 1H), 6.87 (dd, J=11.0, 8.8 Hz, 1H), 4.69 (d, J=11.4 Hz, 1H), 4, 41-4.47 (m, 1H), 4.26 (td, J=10.9, 2.4 Hz, 1H), 3.65-3.72 (m, 1H), 2.39 (s, 3H), 2.15-2.25 (m, 1H), 2.05-2.13 (m, 1H), 1.52 (d, J=6.6 Hz, 3H); LC/MS RT 1.68 min, m/z [MH] ^- 547, 549 8B

¹ H NMR (CD ₃ OD) δ: 7.60 (d, J=8.4 Hz, 1H), 7.24 (dd, J=9.0, 4.9 Hz, 1H), 7.02 ( d, J=8.4 Hz, 1H), 6.87 (dd, J=11.2, 9.0 Hz, 1H), 4.74 (d, J=11.4 Hz, 1H), 4, 41-4.46 (m, 1H), 4.29-4.36 (m, 1H), 3.64-3.74 (m, 1H), 2.38 (s, 3H), 2.22- 2.29 (m, 1H), 2.05-2.13 (m, 1H), 1.50 (d, J=7.0 Hz, 3H); LC/MS RT 1.74 min, m/z [MH] ^- 547, 549 9A

¹ H NMR (CD ₃ OD) δ: 7.73 (dd, J=8.8, 6.2 Hz, 1H), 6.96 (dd, J=8.4, 5.9 Hz, 1H), 6.66-6.77 (m, 2H), 4.69 (d, J=11.4 Hz, 1H), 4.84-4.90 (m, 1H), 4.53-4.60 ( m, 1H), 4.35 (ddd, J=13.1, 10.9, 2.4 Hz, 1H), 3.62-3.71 (m, 1H), 2.21 (s, 3H) , 2.17 (s, 3H), 1.95-2.12 (m, 2H), 1.49 (d, J=7.0 Hz, 3H); LC/MS RT 1.57 min, m/z [MH] ^- 494 9B

¹ H NMR (CD ₃ OD) δ: 7.77 (dd, J=8.8, 6.2 Hz, 1H), 6.97 (dd, J=8.2, 5.7 Hz, 1H), 6.68-6.78 (m, 2H), 4.85-4.93 (m, 1H), 4.74 (d, J=11.4 Hz, 1H), 4.51-4.60 ( m, 1H), 4.33 (td, J=11.5, 3.3 Hz, 1H), 3.62-3.71 (m, 1H), 2.22 (s, 3H), 2.18 (s, 3H), 1.96-2.09 (m, 2H), 1.47 (d, J=6.6 Hz, 3H); LC/MS RT 1.61 min, m/z [MH] ^- 494 10A

¹ H NMR (CD ₃ OD) δ: 7.66 (d, J=8.4 Hz, 1H), 7.05 (d, J=8.4 Hz, 1H), 6.96 (dd, J= 8.1, 5.5 Hz, 1H), 6.69 (dd, J=11.7, 8.4 Hz, 1H), 4.86-4.93 (m, 1H), 4.71 (d , J=11.4 Hz, 1H), 4.53-4.61 (m, 1H), 4.29-4.39 (m, 1H), 3.63-3.71 (m, 1H), 2.21 (s, 3H), 2.17 (s, 3H), 2.01-2.06 (m, 2H), 1.49 (d, J=7.0 Hz, 3H); LC/MS RT 1.61 min, m/z [MH] ^- 510, 512 10B

¹ H NMR (CD ₃ OD) δ: 7.70 (d, J=8.4 Hz, 1H), 7.07 (d, J=8.4 Hz, 1H), 6.97 (dd, J= 8.3, 5.7 Hz, 1H), 6.71 (dd, J=11.7, 8.3 Hz, 1H), 4.92-4.95 (m, 1H), 4.75 (d , J=11.4 Hz, 1H), 4.53-4.60 (m, 1H), 4.26-4.39 (m, 1H), 3.58-3.75 (m, 1H), 2.22 (s, 3H), 2.18 (s, 3H), 1.95-2.14 (m, 2H), 1.47 (d, J=7.0 Hz, 3H); LC/MS RT 1.67 min, m/z [MH] ^- 510, 512

[000153] Experimental Example: The compound of the present invention was evaluated using the following test method.

[000154] Experimental Example 1: Human RNR Inhibition Effect

[000155] The inhibitory activity against the ribonucleotide reduction reaction (hereinafter referred to as the RNR reaction) of the test compound was determined by measuring the formation of deoxycytidine diphosphate (hereinafter referred to as dCDP) from cytidine diphosphate (hereinafter referred to as CDP) by the following method.

[000156] Human M1 subunit and human M2 subunit (mutant lacking amino-terminal 59 amino acids), which are fused with a histidine tag at the amino-terminus, are overexpressed in Escherichia coli and solubilized after harvest, and histidine-labeled human M1 and M2 proteins are purified on a nickel chelate column .

[000157] The method described in Cancer Research 64, 1-6 (2004) was used to measure the RNR inhibitory activity of a test compound.

[000158] First, test compounds were serially diluted with DMSO. Then, human M1 protein and human M2 protein were added to an aqueous solution of albumin prepared from 0.02% fetal bovine serum, DMSO solution of the compound of the present invention or DMSO control solution (final DMSO concentration was 1%) was added, and the mixture was allowed to stand for 20 min. The reaction buffer was then added [50 mM HEPES buffer (pH 7.2) at final concentration, 4 mM magnesium acetate at final concentration, 100 mM potassium chloride at final concentration, 6 mM dithiothreitol at final concentration, 2 mM adenosine triphosphate at final concentration, 0.24 mM nicotinamide adenine dinucleotide phosphate at final concentration] and 10 μM CDP at final concentration, and incubated at 37°C for 30 min to carry out the RNR reaction. Immediately after the reaction, the reaction was stopped by heating at 100° C. for 15 minutes, followed by centrifugation at 10,000 rpm for 10 minutes. After centrifugation, a portion (5 µl) of the obtained supernatant was analyzed by high performance liquid chromatography (Shimadzu Corporation, Prominence) using Shim-pack XR-ODS (manufactured by Shimadzu GLC Co., 3.0 x 100 mm). Elution was performed at a measurement wavelength of 265 nm at a flow rate of 0.5 ml/min with a 9 min concentration gradient from a 12:13 mixture of mobile phase A (10 mM potassium dihydrogen phosphate (pH 6.7), 10 mM tetrabutylammonium, 0.25% methanol) and mobile phase B (50 mM potassium dihydrogen phosphate (pH 6.7), 5.6 mM tetrabutylammonium, 30% methanol) in the same 2:3 mixture to measure CDP of substrate (RT 5.9 min) and reaction product dCDP (CT 6.2 min).

[000159] The inhibitory activity of a test compound was determined using the following equation, and the concentrations of test compounds inhibiting the RNR response by 50% are shown as IC ₅₀ (μM) in Tables 19-1 to 19-3.

[000160] [Mathematical formula 1]

Degree of Inhibition (%) = (1 - (Amount of dCDP generated by addition of test compound (pmol) / Amount of control dCDP generated (pmol))) × 100

[000161] As a result, it can be seen from the table below that the compound of the present invention has an excellent RNR inhibitory action. In contrast, Comparative Example 1 had an IC _{50 of} 43 μM and did not exhibit RNR inhibitory activity as found in the Compound Examples of the present invention.

[000162] [Table 11]

Example RNR enzyme inhibitory activity, IC ₅₀ (μM) Example RNR enzyme inhibitory activity, IC ₅₀ (μM) Example RNR enzyme inhibitory activity, IC ₅₀ (μM) one 0.06 6A 0.10 9A 0.12 2 0.14 6B 0.18 9B 0.05 3 0.18 7A 0.08 10A 0.08 four 0.25 7B 0.06 10B 0.07 5A 0.03 8A 0.08 5B 0.08 8B 0.14

[000163] Experimental Example 2: Inhibitory effect on cell proliferation of a human breast cancer cell line

[000164] HCC 1806 cells derived from a human breast cancer cell line were passaged daily at a cell density not exceeding 80% in Roswell Park Memorial Institute ATCC recommended medium (RPMI-1640) containing 10% fetal bovine serum (FBS) . To start testing the cell proliferation inhibition activity, HCC 1806 cells were suspended in the above medium after seeding at 180 μl in each well of a 96-well flat-bottomed plate, so that the number of cells per well was 2000, the cells were cultured at 37°C for 1 day in an incubator containing 5% carbon dioxide gas. The next day, the test compound was dissolved in DMSO and 20 µl of drug additive solution serially diluted with distilled water to 10-fold final concentration was added to each well of the cell culture plate, and the cells were cultured at 37°C for 72 h in an incubator containing 5 % gaseous carbon dioxide. After culturing for 72 hours, 20 μl of glutaraldehyde was added to each well and left to stand for 30 minutes, then the plate was washed 10 times with water and dried. 100 µl of a dye solution (0.05% crystal violet in 20% methanol) was added to each well and kept for 30 min, then the plate was washed 10 times with water and dried. 100 μl of the extract solution (0.1 N NaH ₂ PO ₄ : 100% ethanol=1:1) was added to each well and mixed, and the mixture was measured at a wavelength of 540 nm using a plate reader (MTP-450, manufactured by Corona Electric Co., Ltd.). The degree of growth inhibition was calculated by the following formula, and the concentration (IC ₅₀ (μM)) of the test compound for 50% inhibition was determined. The results are shown in Table 11.

[000165] Degree of growth inhibition (%) = {(C-B) - (T-B)} / (C-B) × 100

[000166] T: Optical density of the well to which the test compound was added

[000167] C: Optical density of wells to which test compound was not added

[000168] B: Optical density of wells to which cell suspension was not added

[000169] As a result, as can be seen from the following table, it was found that the compounds of the present invention have a growth inhibitory activity against cancer cells.

[000170] [Table 12]

Example number Cell Growth Inhibition IC ₅₀ (µM) Example number Cell Growth Inhibition IC ₅₀ (µM) Example number Cell Growth Inhibition IC ₅₀ (µM) one 0.16 6A 0.06 9A 0.29 2 0.20 6B 0.82 9B 0.78 3 0.64 7A 0.06 10A 0.13 four 0.56 7B 0.67 10B 0.40 5A 0.05 8A 0.14 5B 0.46

[000171] Experimental Example 3 Inhibitory effect on cell proliferation of human cancer cell lines

[000172] According to the method of Experimental Example 2, the inhibitory effect on cell proliferation was evaluated on various cancer cell lines as described in Tables 12 and 13.

[000173] As a result, as can be seen from the following table, it was found that the compounds of the present invention have growth inhibitory activity against various types of human-derived cancer cells.

[000174] [Table 13]

cell line NUGC-3 NCI-H460 CFPAC-1 A673 GB-1 HLE MSTO-211H DU145 type of carcinoma Stomach cancer Lung cancer Pancreas cancer Ewing's sarcoma glioblastoma Liver cancer Mesothelioma prostate cancer Cultural environment RPMI-1640+10% FBS Recommended ATCC RPMI-1640+10% FBS IMDM+10% FBS DMEM+10% FBS DMEM+10% FBS DMEM+10% FBS Recommended ATCC RPMI-1640+10% FBS EMEM + 0.1 mM non-essential amino acid + 1 mM sodium pyruvate + 10% FBS Number of cells (cells/well) 2000 1000 2000 2000 3000 3000 6000 5000 Example 2 1.22 0.73 0.94 1.09 1.57 0.79 0.70 1.04 Example 10A 0.71 0.35 0.35 0.61 1.12 0.42 0.39 0.53 Example 3 3.11 1.50 1.71 2.56 5.22 1.74 1.54 1.84 Example 1 1.12 0.57 0.54 0.92 1.56 0.56 0.65 0.73 Example 6A 0.40 0.25 0.33 0.32 0.64 0.26 0.32 0.31 Example 7A 0.36 0.18 0.23 0.25 0.46 0.20 0.27 0.26 Example 5A 0.27 0.13 0.17 0.18 0.37 0.14 0.17 0.17 Example 8A 0.51 0.31 0.36 0.40 0.85 0.29 0.37 0.34

[000175] [Table 14]

cell line A2780 ACHN HCT116 RPMI7932 NCI-H2228 NCI-H2170 type of carcinoma ovarian cancer kidney cancer colorectal cancer Melanoma Lung cancer Lung cancer Cultural environment RPMI-1640+10% FBS EMEM+10% FBS McCoy 5A+10% FBS RPMI-1640+10% FBS Recommended ATCC RPMI-1640+10% FBS Recommended ATCC RPMI-1640+10% FBS Number of cells (cells/well) 2000 2000 1000 4000 5000 5000 Example 2 0.83 0.75 0.91 2.67 1.27 1.89 Example 10A 0.40 0.38 0.48 1.23 0.88 1.10 Example 3 2.08 1.50 2.30 4.74 3.21 3.90 Example 1 0.63 0.68 0.75 1.74 1.35 1.41 Example 6A 0.30 0.22 0.28 0.72 0.73 0.57 Example 7A 0.19 0.17 0.27 0.51 0.48 0.52 Example 5A 0.13 0.13 0.22 0.43 0.50 0.49 Example 8A 0.38 0.32 0.38 0.65 0.74 0.88

[000176] Experimental Example 4: Evaluation of antitumor activity using human blood cancer cell line (MV-4-11), subcutaneous transplantation model (in vivo)

[000177] The human blood cancer cell line MV-4-11 was transplanted subcutaneously into a nude mouse, and at the time when the tumor volume of the tumor transplanted nude mice reached about 100-300 mm ³ , four mice were assigned to each group by random stratification in this way so that the average value of the tumor volume for each group was the same (day 0), and the compound of the present invention was administered orally daily at a dose of 100 mg/kg/day once a day for 14 days.

[000178] In order to compare the chronological change in tumor proliferation upon administration of each test compound, the relative tumor volume (RTV), taking the tumor volume at the time of assignment to the group as 1, as the tumor proliferation rate was calculated according to the following formula, and the change in the mean RTV values of each individual are presented in Fig.1 - Fig.4.

[000179] RTV = (tumor volume on day of tumor volume measurement) / (tumor volume at grouping)

[000180] The mean RTV in the compound of the present invention group on the last day of evaluation was less than the mean RTV in the control group, and when a statistically significant difference (Student's t-test) was shown, the compound of the present invention was found to be significantly effective, and a statically significant difference is marked with * in the figure (*: p<0.05).

[000181] Thus, it was found that the compound of the present invention has a significant antitumor effect.

[000182] Experimental Example 5: Formation of an illustrative sample of a crystalline form, co-crystal or salt

[000183] A compound selected from Examples 1-10 above was attempted to crystallize with a total of 47 secondary compounds (sodium hydroxide, potassium hydroxide, L-arginine, calcium hydroxide, choline hydroxide, diethylamine, L-lysine, diethanolamine, tromethamine, ethylenediamine , oxalic acid, dibenzoyl-L-tartaric acid, maleic acid, glutamic acid, malonic acid, fumaric acid, L-tartaric acid, 4-hydroxybenzamide, nicotinamide, isonicotinamide, saccharin, galactaric acid, citric acid, D-glucuronic acid, L -malic acid, ethylenediamine, glutaric acid, succinic acid, urea, arginine, benzoic acid, ascorbic acid, methylparaben, vanillin, glycolic acid, hydrochloric acid, hippuric acid, L-lactic acid, methanesulfonic acid, phosphoric acid, p-toluenesulfonic acid monohydrate acids, sulfuric acid, L-alanine, glycine, meglumine, L-proline, L-serine, L-valine), but only the combination with benzoic acid formed an illustrative a sample of a crystalline form, co-crystal, or salt.

[000184] Initially, 93 mg of a compound selected from examples 1-10 was dissolved in 500 μl of isopropyl acetate and the solution was stirred at 60°C; 233 μl of a solution of benzoic acid in methanol (concentration: 0.8672 mmol/ml) was added to the sample. The mixture was stirred at 60°C for 10 min, filtered through a preheated syringe filter (0.2 μm, nylon) and then cooled to ambient temperature, followed by storage at reduced temperature, evaporation and stirring of the gel obtained by evaporation in 1 ml heptane at ambient conditions for 30 days, producing an illustrative sample containing 1 equivalent of benzoic acid. However, with this production method, more than 30 days of production time was required, and only about 10 mg of an illustrative sample was obtained.

[000185] In another example, 3 g of an illustrative sample was obtained by suspending a similar amount of the same compound in a heptane-toluene solvent mixture and stirred at a temperature of 40°C or higher. For example, 1518 mg of benzoic acid, as well as 57 ml of heptane and 3 ml of toluene were added to 3000 mg of a compound selected from examples 1-10. The suspension was stirred for about 4.5 hours while heating at 60°C. The suspension was collected by filtration, washed with a mixture of heptane and toluene in a ratio of 19:1 and heated at 60°C. The solid was recovered and dried to give 3420.6 mg (91% recovery) of a product with a chemical purity of 98.67% and an optical purity of 100% ee. ¹ H-NMR(CDCl ₃ ) δ: 8.77 (s, 1H), 8.09 (d, J=8.0 Hz, 2H), 7.89 (d, J=8.0 Hz, 1H) , 7.60 (t, J=7.2 Hz, 1H), 7.54 (d, J=2 Hz, 1H), 7.40-7.51 (m, 3H), 6.91 (dd, J=6.0Hz, 8.4Hz, 1H), 6.87(d, 9.6Hz, 1H), 6.67(dd, J=8.0Hz, 8.4Hz, 1H), 6.55 (broad s, 1H), 6.34 (broad s, 1H), 4.88 (dd, J=9.6 Hz, 10.4 Hz, 1H), 3.40-3.55 (m, 1H), 2.15 (s, 6H), 1.42 (d, J=6.8 Hz, 3H).

[000186] The suspension was filtered and the solid was washed with a mixture of heptane and toluene and heated at 60°C. As a result of evaluating the physicochemical properties of the obtained crystalline form, co-crystal or salt, it was unexpectedly found that this product has excellent storage stability, moreover, it has low hygroscopicity and electrostatic charge storage capacity, and is easy to handle.

[000187] In another example, 506 mg of benzoic acid, as well as 19 ml of heptane and 1 ml of toluene were added to 1000 mg of a compound selected from examples 1-10. The suspension was stirred for about 3.5 hours while heating at 60°C. The suspension was collected by filtration, washed with a mixture of heptane and toluene in a ratio of 19:1 and heated at 60°C. The solid was recovered and dried to obtain 947 mg (extraction rate: 75.6%) of an exemplary sample.

[000188] In addition, among the 47 kinds of secondary compounds used for the crystallization study, there were secondary compounds containing a basic functional group (or cation), but the present compound, which is acidic, did not form a crystal with these basic secondary compounds. Thus, it was unexpected that the acidic compound formed a crystalline form, a co-crystal or a salt with only benzoic acid, which is an acidic compound. In addition, it is impossible to predict what physico-chemical properties the resulting crystalline form, co-crystal or salt has.

[000189] Experimental Example 6: Structural Characteristics

[000190] Single crystal analysis: 5 ml of heptane, 3 ml of n-propyl acetate and 203.3 mg of benzoic acid were added to 100 mg of the sample from Experimental Example 5 using the compound of Example 2 (hereinafter also referred to as "illustrative sample of the present invention") and dissolved at 80°C. 810 mg of benzoic acid was added to the dissolved product and dissolved. A small amount of product was added and the mixture was left to stand overnight at 60°C. Thereafter, the temperature was lowered to 50°C and left to stand for 6 days to obtain crystals for single crystal analysis.

[000191] The measurement was carried out under the following measurement conditions and data processing was performed using the CrysAlisPro measurement and data processing software, the CrystalStructure structure analysis software package, and the PDXL integrated powder radiography software.

Device: XtaLAB PRO MM 007

X-ray source: Cu Kα (λ=1.54184 Å)

Tube voltage - tube current: 40 kV - 30 mA

Measurement temperature: -173°C (when using a low temperature spray device)

Collimator diameter: 0.3mm

Camera Length: 39.71mm

Vibration angle: 0.25°

Exposure time: 0.25s/sample

Total number of measurements: 14476 plates

Total measurement time: 1 h 35 min

[000192] The results are shown below and in Figure 5.

Crystal system: monoclinic system

Space group: P21/n (No. 4)

Lattice constant: a=13.89023 (9) Å

b=7.77623 (4) Å

c=14.00408 (9) Å

α=90°

β=110.5202 (7)°

γ=90°

Unit cell volume: 1416.653 (16) Å 3

[000193] The X-ray powder diffraction data of an exemplary sample of the present invention was measured in accordance with the following test conditions.

Device: EMPYREAN by PANalytical

Reflection method (intensive method)

Target: Cu

X-ray tube current: 40mA

X-ray tube voltage: 45KV

Scan range: 2θ= 5.0-40.0°

Pitch: 2θ= 0.0131°

Average time/step: 8.670 s

Scanning speed: 0.0015°/s

Slit divergence: 1°

Diffusion Slit: 2.0mm

Receiving slot: 8.0 mm

[000194] The X-ray diffraction spectrum is shown in Fig.6. The X-ray powder diffraction profile of the sample has diffraction angles (2θ±0.2°) of 6.8°, 7.8°, 11.2°, 13.4°, 13.7°, 16.0°, 17.1°, 17.8° and 23.2°.

[000195] Experimental Example 7: Differential Scanning Calorimetry (DSC Analysis)

[000196] DSC analysis of an illustrative sample of the present invention was carried out in accordance with the following test conditions.

Device: DSC 1 STAR SYSTEM from METTLER TOLEDO

Sample: approximately 1 mg

Sample container: made of aluminum

Temperature rise range: 25-290 °C

Temperature rise rate: 10 °C/min.

Atmospheric gas: nitrogen

Nitrogen gas flow rate: 50 ml/min.

[000197] The operation of the devices, including data processing, was carried out in accordance with the manual supplied with the DSC device. The resulting DSC curve is shown in Fig.7. As seen in the DSC results, an endothermic peak (top of the peak) was observed at 162°C.

[000198] Figure 9 shows the result of XRD profile simulation based on the single crystal analysis result and the characteristic peaks of the diffraction angle (2θ±0.2°) (6.8°, 7.8°, 11.2°, 13.4° , 13.7°, 16.0°, 17.1°, 17.8° and 23.2°) correspond to the above example.

[000199] Experimental Example 8: Hygroscopicity

[000200] Moisture adsorption/desorption testing was carried out in accordance with the following conditions.

[000201] Approximately 10-15 mg of the sample was placed in a special quartz holder, and the weight of each sample at each humidity was continuously measured and recorded under the following conditions. The operation of the device, including data processing, was carried out in accordance with the method and procedure indicated in the instructions for each device.

Device: VTI SA+ (manufactured by TA Instruments Inc.)

Drying temperature: 60°C

Heating rate: 1°C/min

Equilibrium drying: in the range not exceeding 300 min, confirm that there is no decrease of 0.01% of the mass. within 5 minutes

Measurement temperature: 25°C

Moisture equilibrium: in the range not exceeding 120 min, it is confirmed that there is no increase of 0.01% of the mass. within 5 minutes

Relative Humidity Program: Increase by 5% RH from 5% RH to 95% RH, and decrease by 5% RH from 95% RH to 5% RH

[000202] When the free form of the compound was originally obtained as an amorphous substance, the free form had hygroscopicity (FIG. 8) and also had a high ability to accumulate an electrostatic charge. In this case, the expression "free form" refers to an amorphous substance, represented by a drug substance alone. However, as described above, the crystalline form, co-crystal or free form salt of the compound of Example 2 was obtained, and this sample did not have hygroscopicity (FIG. 8) and had a low ability to accumulate an electrostatic charge.

[000203] As shown in Figure 8, the hygroscopicity of the resulting product (milled into product) exhibited a weight change of about 0.1% at 95% relative humidity in the moisture adsorption/desorption test, and accordingly the product exhibited little or no hygroscopicity. These physicochemical properties are superior to the hygroscopicity and ability to accumulate an electrostatic charge of the free form of the same compound.

[000204] The resulting solid product of Example 2 had a low ability to accumulate an electrostatic charge, and slight sticking to a spatula or glass bottle was observed.

[000205] As a comparative example, the above compound was spray dried to remove residual solvent. 2 g of the compound was dissolved in 18 g of ethanol, and spray drying was carried out under the following conditions.

After heating the input heat: progression

Waste heat temperature: 78°C

Delivery flow rate: 5 ml/min

Spray nitrogen flow rate 350 l/h

[000206] The obtained spray-dried product was dried under reduced pressure in an atmosphere at 90° C. for 1 hour to obtain a free form of the compound, which was amorphous, had a high ability to store electrostatic charge, and exhibited strong adhesion when transferred with a spatula or in a glass bottle. . In addition, as shown in FIG. 8, a weight gain of about 3.8% was observed at 95% relative humidity in the moisture adsorption/desorption test. This comparative example was also used for the stability test below.

[000207] Experimental Example 9: Stability Test

[000208] Initially, chemical purity was measured by the following methods.

[000209] The amount of similar substances in the sample solution was measured using HPLC analysis. The operation of the device, including data processing, was carried out in accordance with the manual supplied with the device (i.e. Prominence-i). Approximately 2 mg of an illustrative sample of the present invention was dissolved in 4 ml of a mixture (17:3) mobile phase A/mobile phase B, and the resulting mixture was used as a sample solution.

Column: CAPCELL PAK ADME by Shiseido (4.6 × 150 mm, 3 µm)

UV detection: 220 nm

Column temperature: 40°C

Column flow rate: 1.0 ml/min

Mobile phase: A; 5 mM phosphate buffer, pH 6.2/Acetonitrile (4:1)

B: acetonitrile

Injection volume: 5 µl

Sample cooler temperature: 5°C

Collection time: 40 min

[000210] [Table 15] HPLC Gradient

Time (min) Mobile phase A
about.% Mobile phase B
about.% 0~10 85→68 15→32 10~22 68 32 22~32 68→35 32→65 32~40 35 65 40~41 35→85 65→15 41~50 85 fifteen

[000211] The stability of the solid illustrative sample of the present invention and the free form obtained above was tested after the sample was stored at 40°C/75% RH (closed and open) for 2 weeks or 4 weeks. The open state means that the glass bottle cap has been removed and the bottle covered with a Kimwipe.

Measurement points: 2 weeks and 4 weeks

Stored amount: approximately 50-60 mg

Storage container: brown glass bottle

[000212] [Table 16]

Initial value 40°C/75% RH (open) 40°C/75% RH (closed) 2 weeks 4 weeks 2 weeks 4 weeks Comparative Example
Chemical purity (%) 98.49 98.35 98.14 98.42 98.16 Comparative Example
Total Similar Substances (%) 1.51 1.65 1.86 1.58 1.84 Illustrative Sample
Chemical purity (%) 98.67 98.67 98.60 98.67 98.62 Illustrative Sample
Total Similar Substances (%) 1.33 1.33 1.40 1.33 1.38

[000213] After storage, the sample gave only a small amount of similar substances, showing excellent stability compared to the comparative example having a free form. Thus, it was confirmed that the sample of the present invention has excellent stability.

[000214] Optical purity was also measured under the following HPLC conditions.

Detector: UV at 240 nm

Column: CHIRALPAK IE (4.6 x 250 mm, 5 µm)/Daicel Corp.

Column temperature: 40°C

Mobile phase: n-hexane/ethanol/ethanolamine/acetic acid (65:35:0.5:0.2)

Flow rate: 1.0 ml/min

Injection volume: 5 µl

Sample temperature: 5°C

Measurement time interval: 40 min

[000215] Limit of quantification (LOQ):

[000216] Compound - RR, RS, SS: 0.05%

[000217] Optical purity was also measured with the addition of 0.2% optical isomer.

[000218] The stability of the solid illustrative sample of the present invention and the free form obtained above was tested before and after the sample was stored at 40°C/75% RH (closed and open) for 24 hours. After 24 hours, no significant no change in optical purity was observed.

[000219] The results of the measurement of optical purity are shown in Fig.10 - Fig.12.

[000220] Experimental Example 10: Dissolution Test

[000221] Approximately 15 mg of an illustrative sample of the present invention was added to 1 ml of each solvent and left to swell at 37° C. for 2 hours, protected from light. After centrifugation, each supernatant was filtered and 0.7 ml of a mixture of water and acetonitrile (1:1) was added to 0.1 ml of the filtrate. The dissolution test was carried out with 5 µl of solvent according to the following conditions.

Detector: UV spectrophotometer (wavelength: 230 nm)

Column: CAPCELPAK ADME (4.6 × 150 mm, 3 mm)

Column temperature: constant temperature approx. 40°C

Mobile phase A: A: mixture of 5 mM phosphate buffer (pH 6.52) and acetonitrile (4:1)

B: Acetonitrile

After mobile phase: Gradient control was performed by mixing mobile phases A and B as indicated in the following table below.

Flow rate: 1.0 ml/min

Measurement time interval: within 12 minutes after injection

[000222] [Table 17] HPLC Gradient

Time after administration
sample (min) Mobile phase A (vol.%) Mobile phase B (vol.%) 0-4 70→30 30→70 4-12 thirty 70 12-13 30→70 70→30 13-18 70 thirty

[000223] The results of the dissolution test are shown in Table 18 below.

[000224] [Table 18] Solubility Data

Solvent Solubility (mg/ml) free acid Illustrative Sample water 0.23 0.23 The first dissolution test fluid,
pH 1.2 0.23 0.23 Acetate buffer (50 mM, pH 4.0) 0.25 0.26 Second dissolution test fluid, pH 6.8 0.80 0.61 0.5% HPMC 0.19 0.13

[000225] No significant change was observed in the XRD pattern after the 2 hour dissolution study, and no significant difference was observed between the free acid form and the illustrative sample of the present invention.

[000226] Experimental Example 11: Solid State Stability Test

[000227] The stability of the solid illustrative sample of the present invention and the free form obtained above was tested after the sample was stored at 40°C/75% RH (open), 70°C (closed) and 80°C (closed) after 2 weeks or 4 weeks. The open state means that the glass bottle cap has been removed and the bottle sealed with a Kimwipe, 70°C and 80°C (closed).

[000228] Measurement points: 2 weeks and 4 weeks

[000229] Stored amount: about 30 mg

[000230] Storage container: brown glass bottle

[000231] The amount of similar substances in the sample solution was measured using HPLC analysis. The operation of the device, including data processing, was carried out in accordance with the manual supplied with the device (i.e. Prominence-i). Approximately 2 mg of an illustrative sample of the present invention was dissolved in 4 ml of a mixture (17:3) mobile phase A/mobile phase B, and the resulting mixture was used as a sample solution.

Column: CAPCELL PAK ADME by Shiseido (4.6 × 150 mm, 3 µm)

UV detection: 220 nm

Column temperature: 40°C

Column flow rate: 1.0 ml/min

Mobile phase: A; 5 mM phosphate buffer, pH 6.5/Acetonitrile (4:1)

B: acetonitrile

Injection volume: 5 µl

Sample cooler temperature: 5°C

Collection time: 40 min

[000232] [Table 19] HPLC Gradient

Time (min) Mobile phase A
about.% Mobile phase B
about.% 0~10 85→68 15→32 10~22 68 32 22~32 68→35 32→65 32~40 35 65 40~41 35→85 65→15 41~50 85 fifteen

[000233] [Table 20]

Initial value 40°C, 75% RH
(open) 70°C (closed) 80°C (closed) 4 weeks 2 weeks 4 weeks 2 weeks 4 weeks Comparative Example
Chemical purity (%) 98.33 97.97 97.31 96.39 95.90 92.92 Comparative Example
Total Similar Substances (%) 1.67 2.03 2.69 3.61 4.10 7.08 Illustrative Sample
Chemical purity (%) 99.69 99.69 99.67 99.68 99.66 99.66 Illustrative Sample
Total Similar Substances (%) 0.31 0.31 0.33 0.32 0.34 0.34

Claims (18)

1. Co-crystal of benzoic acid and 5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1 ,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide, where the powder X-ray diffraction pattern of the co-crystal contains five or more peaks at (2θ ± 0.2°) 6.8°, 7.8°, 11.2° , 13.4°, 13.7°, 16.0°, 17.1°, 17.8° and 23.2°. 2. A co-crystal according to claim 1, wherein the co-crystal has an endothermic peak of 155 to 168° C. as measured by DSC. 3. A co-crystal according to claim 2, wherein the co-crystal has an endothermic peak at 162° C. measured by DSC. 4. Co-crystal according to claim 1, wherein the X-ray powder diffraction pattern of the co-crystal contains peaks at (2θ ± 0.2°) 6.8°, 7.8°, 11.2°, 13.4°, 13.7°, 16 .0°, 17.1°, 17.8° and 23.2°. 5. A co-crystal according to claim 1, wherein the X-ray powder diffraction pattern of the co-crystal contains peaks at (2θ ± 0.2°) 6.8°, 7.8°, 11.2°, 13.4°, 13.7°, 16 0°, 17.1°, 17.8° and 23.2°, and wherein the co-crystal has an endothermic peak at 162° C. measured by DSC. 6. Co-crystal of benzoic acid and 5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3-dimethylphenyl)-1-(5-oxo-4,5-dihydro-1 ,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide, where the molar ratio of benzoic acid to 5-chloro-2-(N-((1S,2R)-2-(6-fluoro-2,3- dimethylphenyl)-1-(5-oxo-4,5-dihydro-1,3,4-oxadiazol-2-yl)propyl)sulfamoyl)benzamide is 1:1, and where the X-ray powder diffraction pattern of the co-crystal contains five or more peaks at (2θ ± 0.2°) 6.8°, 7.8°, 11.2°, 13.4°, 13.7°, 16.0°, 17.1°, 17.8° and 23, 2°. 7. A co-crystal according to claim 6, wherein the co-crystal has an endothermic peak of 155 to 168° C. as measured by DSC. 8. A co-crystal according to claim 7, wherein the co-crystal has an endothermic peak at 162° C. measured by DSC. 9. Co-crystal according to claim 6, wherein the X-ray powder diffraction pattern of the co-crystal contains peaks at (2θ ± 0.2°) 6.8°, 7.8°, 11.2°, 13.4°, 13.7°, 16 .0°, 17.1°, 17.8° and 23.2°. 10. Co-crystal according to claim 6, wherein the X-ray powder diffraction pattern of the co-crystal contains peaks at (2θ ± 0.2°) 6.8°, 7.8°, 11.2°, 13.4°, 13.7°, 16 0°, 17.1°, 17.8° and 23.2°, and wherein the co-crystal has an endothermic peak at 162° C. measured by DSC. 11. A pharmaceutical composition having ribonucleotide reductase inhibitory activity, comprising (i) a co-crystal according to any one of claims 1 to 10, and (ii) a pharmaceutically acceptable carrier. 12. A method for inhibiting ribonucleotide reductase in vivo , comprising administering to a human subject in need thereof a therapeutically effective amount of a pharmaceutical composition according to claim 11. 13. The method of claim 12, wherein the inhibition of ribonucleotide reductase occurs in the tumor cell of the human subject. 14. An antitumor agent having ribonucleotide reductase inhibitory activity, containing a co-crystal according to any one of claims 1 to 10 as an active ingredient. 15. The antitumor agent of claim 14, wherein the antitumor agent is an oral antitumor agent. 16. The use of a co-crystal according to any one of claims 1 to 10 for the production of a ribonucleotide reductase inhibitor. 17. The use of a co-crystal according to any one of claims 1 to 10 as a drug for inhibiting ribonucleotide reductase activity. 18. The use of a co-crystal according to any one of claims 1 to 10 for the preparation of an oral antitumor agent having ribonucleotide reductase inhibitory activity.

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