WO2025113498A1 - Cd73 inhibitor alkaline salt, preparation method therefor, and use thereof - Google Patents

Abstract

Disclosed are a CD73 inhibitor alkaline salt, a preparation method therefor, and a use thereof. The CD73 inhibitor is a compound having the structure shown in formula (I). An alkaline salt of the compound shown in the formula (I) can greatly improve physical and chemical properties such as the chemical stability, solubility, and hygroscopicity of an amorphous free acid of the compound represented by formula (I), meet industrial production requirements, and satisfy development requirements for clinical medicine preparations. The alkaline salt of the compound represented by formula (I) can be widely applied in the preparation of drugs for treating tumors, immune-related diseases, and metabolic diseases mediated at least in part by CD73.

Description

A basic salt of a CD73 inhibitor and its preparation method and application Technical Field

The present invention belongs to the field of drug development, and specifically relates to a basic salt of a CD73 inhibitor and a preparation method and application thereof.

Background Art

CD73, also known as Ecto-5'-nucleotidase (eNT), is a 70kDa protein molecule. Under normal circumstances, it is expressed on vascular endothelial cells and some blood cells. It is anchored to the cell membrane surface through glycosylphosphatidylinositol (GPI) and regulates the metabolism of adenosine triphosphate (ATP) together with CD39. Among them, CD39 (also known as extracellular nucleoside triphosphate hydrolase-NTPDase1) can catalyze ATP to generate adenosine monophosphate (AMP), and only produce a small amount of adenosine diphosphate (ADP). The main function of CD73 is to catalyze the conversion of extracellular nucleotides (such as 5'AMP) into their corresponding nucleosides (such as adenosine).

The nucleosides produced by CD73, especially adenosine, are considered to be internal regulatory molecules for many different physiological functions. Adenosine can regulate the cardiovascular system, central nervous system, respiratory system, kidneys, fat cells, platelets and immune system. In the immune system, extracellular adenosine can act on many different immune cells and mediate anti-inflammatory responses. In many tissues, adenosine can also promote the process of fibrosis.

CD73 expression has been found in many tumor cells, including leukemia, bladder cancer, glioma, glioblastoma, ovarian cancer, melanoma, prostate cancer, thyroid cancer, esophageal cancer and breast cancer. At the same time, CD73 expression has also been found on the surface of immunosuppressive cells (including regulatory T cells Treg and myeloid suppressor cells MDSC). High expression of CD73 has also been found to be associated with angiogenesis, invasion, resistance to chemotherapy, tumor metastasis and shorter survival of cancer patients in various tumors including breast cancer and melanoma.

Mechanism-based studies have shown that malignant tumor cells release a large amount of ATP under chemotherapy and other stresses, which is quickly converted into adenosine and accumulated in the tumor microenvironment. The release of extracellular ATP due to cell death or intracellular stress activates the immune response, but the ATP metabolite adenosine has immunosuppressive activity. One important point is that adenosine in the tumor inhibits infiltrating effector T lymphocytes by activating adenosine receptors (such as A2A), thereby promoting tumor development. Therefore, the accumulation of extracellular adenosine in tumor tissue is an important mechanism for tumor immune escape.

Reducing CD73 expression with interfering RNA or overexpressing CD73 in tumor cells can regulate tumor growth and migration; CD73 knockout mice are less likely to produce organ transplant rejection and spontaneous tumors; genetic deletion of the A2A receptor gene can induce T cell-dependent tumor rejection. In mouse models, treatment with antibodies that bind to mouse CD73 can inhibit breast tumor growth and migration.

Therefore, targeting CD73 represents a potential therapeutic strategy that can enhance the efficacy of anti-tumor therapy and provide a new therapeutic strategy for limiting the further development of tumors. At the same time, targeting CD73 can also be used to treat other diseases mediated by adenosine, such as enhancing immune response, enhancing immune effect, enhancing inflammatory response and treating neurological disorders, neurodegenerative diseases and central nervous system diseases, such as depression, Parkinson's disease, sleep disorders, fibrosis and other immune-inflammatory diseases.

Shanghai Abbisko Therapeutics Co., Ltd. has invented a small molecule compound with CD73 inhibitory effect (WO2018113584A1) during its long-term research. Its representative compounds are as follows:

The Chinese name is: (((((2R,3S,4R,5R)-5-(6-chloro-4-(((R)-6-fluoro-2,3-dihydro-1H-inden-1-yl)amino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-3,4-dihydroxytetrahydrofuran-2-yl)methoxy)(hydroxy)phosphino)methyl)phosphonic acid (compound of formula (I)). The compound has a strong inhibitory effect on the enzymatic and cytological activity of CD73 and can be used to treat diseases that are at least partially mediated by CD73. Tumors, immune-related diseases, metabolic diseases, such as: prostate cancer, colon cancer, rectal cancer, pancreatic cancer, gastric cancer, endometrial cancer, cervical cancer, brain cancer, liver cancer, bladder cancer, ovarian cancer, testicular cancer, head cancer, neck cancer, skin cancer, mesothelial lining cancer, white blood cell cancer, esophageal cancer, breast cancer, muscle cancer, connective tissue cancer, lung cancer, adrenal cancer, thyroid cancer, kidney cancer, bone cancer, brain tumor, glioblastoma, mesothelioma, renal cell carcinoma, sarcoma, choriocarcinoma, epidermal basal cell carcinoma, testicular seminoma, etc.

The applicant obtained the free acid lyophilized powder of the compound of formula (I) according to the preparation method disclosed in the specification, which was identified as an amorphous form. During the later pharmaceutical research process, it was found that its physicochemical properties were poor and unstable, and could not meet the needs of subsequent clinical preparation development and industrial production.

Therefore, in order to meet the needs of clinical research and marketed drug preparations, it is urgently necessary to develop an aggregated form of the compound of formula (I) that is suitable for drug development to overcome the defects of the existing technology.

Summary of the invention

In order to solve the problems existing in the prior art, the inventors have conducted in-depth research on the different aggregation forms of the compound of formula (I), and developed several basic salts of the compound of formula (I), which greatly improved the physicochemical properties of the free state of the compound of formula (I), such as chemical stability, solubility and hygroscopicity. The basic salt raw materials of the compound of formula (I) meet the requirements of industrial production and can meet the needs of clinical drug preparation development. In addition, the inventors accidentally discovered several crystalline free acids of the compound of formula (I) during the salt screening process, which have good stability and hygroscopicity and can be used for medium- and long-term storage in the industrial production process, or as raw materials for the production of basic salts. The basic salts and crystalline free acids of the compound of formula (I) have very important clinical application value and are expected to accelerate the development of a new generation of CD73 small molecule inhibitors.

The first aspect of the present invention provides a basic salt of a compound of formula (I),

Wherein, the basic salt is sodium salt, potassium salt, calcium salt, arginine salt, meglumine salt or ammonium salt.

As a preferred embodiment, the basic salt is a sodium salt or a calcium salt.

As a further preferred embodiment, the molar ratio of the compound of formula (I) to the base in each molecule of the basic salt of the compound of formula (I) is 1:1 to 1:5.

As a further preferred embodiment, the molar ratio of the compound of formula (I) to the base in each molecule of the basic salt of the compound of formula (I) is 1:1, 1:2, 1:3, 1:4 or 1:5.

As a further preferred embodiment, the basic salt of the compound of formula (I) is the sodium salt of the compound of formula (I), and the molar ratio of the compound of formula (I) to sodium atoms in each molecule of the sodium salt of the compound of formula (I) is 1:1, 1:3 or 1:5.

As a further preferred embodiment, the molar ratio of the compound of formula (I) to sodium atoms in each molecule of the sodium salt of the compound of formula (I) is 1:3.

As a preferred embodiment, the sodium salt of the compound of formula (I) is an amorphous or crystalline compound.

As a preferred embodiment, the basic salt of the compound of formula (I) is the trisodium salt of the amorphous form of the compound (I).

As a preferred embodiment, the basic salt of the compound of formula (I) is a crystalline trisodium salt of the compound of formula (I).

As a further preferred embodiment, the trisodium salt of the crystalline form of the compound of formula (I) is form A', and its X-ray powder diffraction pattern (XRPD) includes five or more peaks at diffraction angles (2θ) of 3.22±0.2°, 6.38±0.2°, 9.55±0.2°, 12.72±0.2°, 19.10±0.2°, 20.01±0.2°, 20.53±0.2°, 23.13±0.2°, 25.50±0.2°, 26.27±0.2°, 26.66±0.2°, 28.80±0.2°, 29.43±0.2°, 32.06±0.2° and 35.37±0.2°.

As the most preferred embodiment, the X-ray powder diffraction pattern of the crystalline form A' includes substantially the same peaks at the diffraction angle (2θ) as shown in FIG. 27 , and its X-ray powder diffraction data are shown in the following table:

This crystalline form of the trisodium salt of the compound of formula (I) is designated Form A'.

As a further preferred embodiment, the trisodium salt of the crystalline form of the compound of formula (I) is form B', and its X-ray powder diffraction pattern (XRPD) includes five or more peaks at diffraction angles (2θ) of 3.15±0.2°, 6.27±0.2°, 9.34±0.2°, 12.34±0.2°, 12.59±0.2°, 15.64±0.2°, 17.06±0.2°, 18.90±0.2°, 23.60±0.2° and 25.93±0.2°.

As the most preferred embodiment, the X-ray powder diffraction pattern of the crystalline form B' includes substantially the same peaks at the diffraction angle (2θ) as shown in FIG40 , and its X-ray powder diffraction data are shown in the following table:

This crystalline form of the trisodium salt of the compound of formula (I) is designated Form B'.

As a further preferred embodiment, the trisodium salt of the crystalline form of the compound of formula (I) is form C', and its X-ray powder diffraction pattern (XRPD) includes five or more peaks at diffraction angles (2θ) of 11.4±0.2°, 17.09±0.2°, 17.68±0.2°, 19.01±0.2°, 19.59±0.2°, 19.97±0.2°, 20.81±0.2°, 21.65±0.2°, 22.39±0.2°, 22.81±0.2°, 24.80±0.2°, 27.06±0.2°, 27.73±0.2°, 30.53±0.2° and 32.62±0.2°.

As the most preferred embodiment, the X-ray powder diffraction pattern of the crystalline form C' includes substantially the same peaks at the diffraction angle (2θ) as shown in FIG44 , and its X-ray powder diffraction data are shown in the following table:

This crystalline form of the trisodium salt of the compound of formula (I) is designated Form C'.

As a further preferred embodiment, the trisodium salt of the crystalline form of the compound of formula (I) is form D', and its X-ray powder diffraction pattern (XRPD) includes five or more peaks at diffraction angles (2θ) of 11.97±0.2°, 17.97±0.2°, 18.92±0.2°, 19.41±0.2°, 20.36±0.2°, 20.97±0.2°, 21.66±0.2°, 22.88±0.2°, 23.24±0.2°, 24.15±0.2°, 24.77±0.2°, 25.11±0.2°, 25.51±0.2°, 26.21±0.2° and 30.10±0.2°.

As the most preferred embodiment, the X-ray powder diffraction pattern of the crystalline form D' comprises substantially the same peaks at the diffraction angle (2θ) as shown in FIG31 , and its X-ray powder diffraction data are shown in the following table:

This crystalline form of the trisodium salt of the compound of formula (I) is designated Form D'.

As a further preferred embodiment, the trisodium salt of the crystalline form of the compound of formula (I) is form E', and its X-ray powder diffraction pattern (XRPD) includes five or more peaks at diffraction angles (2θ) of 3.15±0.2°, 3.24±0.2°, 9.79±0.2°, 12.35±0.2°, 12.41±0.2°, 14.30±0.2°, 18.61±0.2°, 19.91±0.2°, 20.80±0.2°, 23.94±0.2°, 24.89±0.2°, 31.39±0.2° and 34.67±0.2°.

As the most preferred embodiment, the X-ray powder diffraction pattern of the crystalline form E' includes substantially the same peaks at the diffraction angle (2θ) as shown in FIG32 , and its X-ray powder diffraction data are shown in the following table:

This crystalline form of the trisodium salt of the compound of formula (I) is designated Form E'.

As a further preferred embodiment, the trisodium salt of the crystalline form of the compound of formula (I) is form F', and its X-ray powder diffraction pattern (XRPD) includes peaks at diffraction angles (2θ) of 3.12±0.2°, 5.81±0.2°, 6.27±0.2°, 9.43±0.2°, 18.94±0.2° and 25.58±0.2°.

As the most preferred embodiment, the X-ray powder diffraction pattern of the crystalline form F' includes substantially the same peaks at the diffraction angle (2θ) as shown in FIG. 45 , and its X-ray powder diffraction data are shown in the following table:

This crystalline form of the trisodium salt of the compound of formula (I) is designated Form F'.

As a further preferred embodiment, the trisodium salt of the crystalline form of the compound of formula (I) is form A1', and its X-ray powder diffraction pattern (XRPD) includes five or more peaks at diffraction angles (2θ) of 3.19±0.2°, 9.55±0.2°, 12.71±0.2°, 13.52±0.2°, 14.43±0.2°, 16.37±0.2°, 19.07±0.2°, 19.99±0.2°, 20.48±0.2°, 24.02±0.2°, 25.06±0.2°, 25.43±0.2°, 26.21±0.2°, 26.60±0.2° and 29.40±0.2°.

As the most preferred embodiment, the X-ray powder diffraction pattern of the crystalline form A1' includes substantially the same peaks at the diffraction angle (2θ) as shown in FIG36 , and its X-ray powder diffraction data are shown in the following table:

The crystalline form of the trisodium salt of the compound of formula (I) is designated as Form A1'.

As a further preferred embodiment, the basic salt of the compound of formula (I) is a calcium salt of the compound of formula (I), and the molar ratio of the compound of formula (I) to calcium atoms per molecule of the calcium salt of the compound of formula (I) is 1:1.

The second aspect of the present invention provides a method for preparing a basic salt of a compound of formula (I), comprising the following steps:

1) dissolving or dispersing the free acid of the compound of formula (I) in water or an organic solvent, and adding an alkaline solution to the above system to carry out a salt-forming reaction; or, adding the free acid of the compound of formula (I) to an alkaline solution to carry out a salt-forming reaction;

2) collecting the solid product basic salt of the compound of formula (I) precipitated in the above-mentioned salt-forming reaction process, or obtaining the solid product basic salt of the compound of formula (I) by creating supersaturation in the salt-forming system;

The free acid of the compound of formula (I) is an anhydrate, a hydrate or a solvate;

The basic salt of the compound of formula (I) is a sodium salt, potassium salt, calcium salt, arginine salt, meglumine salt or ammonium salt of the compound of formula (I);

The alkaline solution is selected from an aqueous solution or an organic solvent solution of sodium hydroxide, potassium hydroxide, calcium hydroxide, arginine, meglumine or ammonia.

The third aspect of the present invention provides a method for preparing a basic salt of a compound of formula (I), comprising the following steps:

1) dissolving or dispersing pentasodium salt of the compound of formula (I) in water or an organic solvent;

2) stirring or suspending, and collecting the trisodium salt solid of the compound of formula (I) precipitated in the above process.

As a further preferred embodiment, the method for creating supersaturation in the salt-forming system in step 2) of the above preparation method comprises one or more of the following: volatilizing the solvent, adding an anti-solvent or lowering the temperature.

As a further preferred embodiment, the organic solvent in the above preparation method is selected from alcohols, chloroalkanes, ketones, ethers, cyclic ethers, esters, alkanes, cycloalkanes, benzenes, amides or sulfoxides, or mixtures thereof, or aqueous solutions thereof.

As a further preferred embodiment, the organic solvent is selected from methanol, ethanol, n-propanol, isopropanol, dichloromethane, heptane, acetonitrile, acetone, methyl ethyl ketone, toluene, 1,4-dioxane, tetrahydrofuran, 2-methyltetrahydrofuran, N,N-dimethylformamide, dimethyl sulfoxide, ethyl acetate, isopropyl acetate, methyl tert-butyl ether or 2-methoxyethyl ether, or a mixture thereof, or an aqueous solution thereof.

The fourth aspect of the present invention provides a crystalline form of a free acid of a compound of formula (I):

As a preferred embodiment, the free acid of the crystalline form of the compound of formula (I) is form A, and its X-ray powder diffraction pattern (XRPD) includes five or more peaks at diffraction angles (2θ) of 5.56±0.2°, 6.55±0.2°, 7.24±0.2°, 9.83±0.2°, 11.13±0.2°, 13.14±0.2°, 15.81±0.2°, 18.01±0.2°, 19.75±0.2°, 20.73±0.2°, 22.38±0.2°, 23.10±0.2°, 23.37±0.2° and 26.28±0.2°.

As the most preferred embodiment, the X-ray powder diffraction (XRPD) of the crystalline form A includes substantially the same peaks as those at the diffraction angle (2θ) shown in FIG. 9 , and its X-ray powder diffraction data are shown in the following table:

The crystalline form of the free acid of the compound of formula (I) is designated as Form A.

As a preferred embodiment, the free acid of the crystalline form of the compound of formula (I) is form B, and its X-ray powder diffraction pattern (XRPD) includes five or more peaks at diffraction angles (2θ) of 4.76±0.2°, 7.21±0.2°, 15.17±0.2°, 16.58±0.2°, 17.03±0.2°, 17.90±0.2°, 19.12±0.2°, 21.32±0.2°, 22.05±0.2°, 23.54±0.2°, 24.32±0.2°, 24.58±0.2°, 25.73±0.2°, 26.08±0.2° and 26.70±0.2°.

As the most preferred embodiment, the X-ray powder diffraction (XRPD) of the crystalline form B includes substantially the same peaks as those at the diffraction angle (2θ) shown in FIG8 , and its X-ray powder diffraction data are shown in the following table:

The crystalline form of the compound of formula (I) free acid is designated as Form B.

The fourth aspect of the present invention provides a pharmaceutical composition, which comprises a clinically effective amount of a basic salt of a compound of formula (I), or a crystalline free acid of a compound of formula (I); and a pharmaceutically acceptable carrier.

In a fifth aspect, the present invention provides a basic salt of a compound of formula (I), or a crystalline free acid of a compound of formula (I), or a pharmaceutical composition comprising a clinically effective amount of a basic salt of a compound of formula (I) or a crystalline free acid of a compound of formula (I) in the preparation of a medicament for treating tumors, immune-related diseases and disorders or metabolic diseases mediated at least in part by CD73.

As a further preferred embodiment, the tumor is selected from prostate cancer, colon cancer, rectal cancer, pancreatic cancer, gastric cancer, endometrial cancer, cervical cancer, brain cancer, liver cancer, bladder cancer, ovarian cancer, testicular cancer, head cancer, neck cancer, skin cancer, mesothelial lining cancer, white blood cell cancer, esophageal cancer, breast cancer, muscle cancer, connective tissue cancer, lung cancer, adrenal cancer, thyroid cancer, kidney cancer, bone cancer, brain tumor, glioblastoma, mesothelioma, renal cell carcinoma, sarcoma, choriocarcinoma, epidermal basal cell carcinoma and testicular seminoma.

As a further preferred embodiment, the tumor is selected from skin cancer, colon cancer, pancreatic cancer, breast cancer, prostate cancer, lung cancer, leukemia cancer, brain tumor, ovarian cancer and sarcoma.

As a further preferred embodiment, the skin cancer is melanoma and basal cell carcinoma; the white blood cell cancer is lymphoma and leukemia; the lung cancer is small cell lung cancer and non-small cell carcinoma; and the sarcoma is Kaposi's sarcoma.

As a further preferred embodiment, the immune-related diseases and disorders are selected from rheumatoid arthritis, renal failure, lupus erythematosus, asthma, psoriasis, ulcerative colitis, pancreatitis, allergies, fibrosis, anemia fibromyalgia, Alzheimer's disease, congestive heart failure, stroke, aortic valve stenosis, arteriosclerosis, osteoporosis, Parkinson's disease, infection, Crohn's disease, ulcerative colitis, allergic contact dermatitis and eczema, systemic sclerosis and multiple sclerosis.

In a sixth aspect, the present invention provides a basic salt of a compound of formula (I), or a crystalline free acid of a compound of formula (I), or a pharmaceutical composition comprising a clinically effective amount of a basic salt of a compound of formula (I) or a crystalline free acid of a compound of formula (I) for use as a drug for treating tumors, immune diseases, disorders or metabolic diseases mediated at least in part by CD73.

As a further preferred embodiment, the tumor is selected from prostate cancer, colon cancer, rectal cancer, pancreatic cancer, gastric cancer, endometrial cancer, cervical cancer, brain cancer, liver cancer, bladder cancer, ovarian cancer, testicular cancer, head cancer, neck cancer, skin cancer, mesothelial lining cancer, white blood cell cancer, esophageal cancer, breast cancer, muscle cancer, connective tissue cancer, small cell lung cancer, adrenal cancer, thyroid cancer, kidney cancer, bone cancer, brain tumor, glioblastoma, mesothelioma, renal cell carcinoma, sarcoma, choriocarcinoma, epidermal basal cell carcinoma, fibroblast ... Basal cell carcinoma and testicular seminoma; the immune-related diseases and disorders are selected from rheumatoid arthritis, renal failure, lupus erythematosus, asthma, psoriasis, ulcerative colitis, pancreatitis, allergies, fibrosis, anemia, fibromyalgia, Alzheimer's disease, congestive heart failure, stroke, aortic valve stenosis, arteriosclerosis, osteoporosis, Parkinson's disease, infection, Crohn's disease, ulcerative colitis, allergic contact dermatitis and eczema, systemic sclerosis and multiple sclerosis.

The present invention also relates to a method for treating tumors, immune diseases and disorders and metabolic diseases that are at least partially mediated by CD73, comprising administering to a patient in need thereof a therapeutically effective amount of a basic salt of a compound of formula (I), or a crystalline form of a free acid of a compound of formula (I), or a pharmaceutical composition comprising a clinically effective amount of a basic salt of a compound of formula (I) or a crystalline form of a free acid of a compound of formula (I).

As a further preferred embodiment, the tumor is selected from prostate cancer, colon cancer, rectal cancer, pancreatic cancer, gastric cancer, endometrial cancer, cervical cancer, brain cancer, liver cancer, bladder cancer, ovarian cancer, testicular cancer, head cancer, neck cancer, skin cancer, mesothelial lining cancer, white blood cell cancer, esophageal cancer, breast cancer, muscle cancer, connective tissue cancer, small cell lung cancer, adrenal cancer, thyroid cancer, kidney cancer, bone cancer, brain tumor, glioblastoma, mesothelioma, renal cell carcinoma, sarcoma, choriocarcinoma, epidermal Basal cell carcinoma and testicular seminoma; the immune diseases and disorders are selected from rheumatoid arthritis, renal failure, lupus erythematosus, asthma, psoriasis, ulcerative colitis, pancreatitis, allergies, fibrosis, anemia fibromyalgia, Alzheimer's disease, congestive heart failure, stroke, aortic valve stenosis, arteriosclerosis, osteoporosis, Parkinson's disease, infection, Crohn's disease, ulcerative colitis, allergic contact dermatitis and eczema, systemic sclerosis and multiple sclerosis.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1 is an X-ray powder diffraction pattern of the free acid of the compound of formula (I). The abscissa represents the 2θ value (degrees), and the ordinate represents the peak intensity.

Figure 2 is the DSC spectrum of the free acid of the compound of formula (I). The abscissa represents temperature (°C), and the ordinate represents heat flow (w/g).

Figure 3 is the mDSC spectrum of the free acid of the compound of formula (I). The abscissa represents temperature (°C), the first ordinate on the left represents reversible heat flow (w/g), the second ordinate on the left represents total heat flow (w/g), and the right ordinate represents irreversible heat flow (w/g).

Figure 4 is a TGA spectrum of the free acid of the compound of formula (I). The abscissa represents temperature (°C), and the ordinate represents weight (%).

Fig. 5 is the ¹ H-NMR spectrum of the free acid of the compound of formula (I). The abscissa represents the displacement of the compound (ppm), and the ordinate represents the signal intensity.

Figure 6 is an X-ray powder diffraction pattern of the pentasodium salt of the compound of formula (I). The abscissa represents the 2θ value (degrees), and the ordinate represents the peak intensity.

Fig. 7 is the ¹ H-NMR spectrum of the pentasodium salt of the compound of formula (I). The abscissa represents the displacement of the compound (ppm), and the ordinate represents the signal intensity.

Figure 8 is an X-ray powder diffraction pattern of the free acid crystalline form B of the compound of formula (I). The abscissa represents the 2θ value (degrees), and the ordinate represents the peak intensity.

Figure 9 is an X-ray powder diffraction pattern of the free acid crystalline form A of the compound of formula (I). The abscissa represents the 2θ value (degrees), and the ordinate represents the peak intensity.

Figure 10 is an X-ray powder diffraction pattern of the monosodium salt of the amorphous form (I) compound. The abscissa represents the 2θ value (degrees) and the ordinate represents the peak intensity.

Figure 11 is an X-ray powder diffraction pattern of the trisodium salt of the amorphous form (I) compound. The abscissa represents the 2θ value (degrees), and the ordinate represents the peak intensity.

Figure 12 is an X-ray powder diffraction pattern of the monopotassium salt of the amorphous form (I) compound. The abscissa represents the 2θ value (degrees) and the ordinate represents the peak intensity.

Figure 13 is an X-ray powder diffraction pattern of the monocalcium salt of the amorphous form (I) compound. The abscissa represents the 2θ value (degrees) and the ordinate represents the peak intensity.

Figure 14 is an X-ray powder diffraction pattern of the arginine salt of the amorphous form (I) compound. The abscissa represents the 2θ value (degrees), and the ordinate represents the peak intensity.

Figure 15 is an X-ray powder diffraction pattern of the amorphous form (I) compound meglumine salt. The abscissa represents the 2θ value (degrees) and the ordinate represents the peak intensity.

Figure 16 is an X-ray powder diffraction pattern of the diammonium salt of the amorphous form (I) compound. The abscissa represents the 2θ value (degrees) and the ordinate represents the peak intensity.

Figure 17 is the ¹ H-NMR spectrum of the meglumine salt of the amorphous form (I) compound. The abscissa represents the compound displacement (ppm), and the ordinate represents the signal intensity.

Figure 18 is a DSC spectrum of the free acid crystalline form B of the compound of formula (I). The abscissa represents temperature (°C), and the ordinate represents heat flow (w/g).

Figure 19 is a TGA spectrum of the free acid crystalline form B of the compound of formula (I). The abscissa represents temperature (°C), and the ordinate represents weight (%).

Figure 20 is a ¹ H-NMR spectrum of the free acid form B of the compound of formula (I). The abscissa represents the compound displacement (ppm), and the ordinate represents the signal intensity.

Figure 21 is the mDSC spectrum of the trisodium salt of the compound of formula (I). The horizontal axis represents temperature (°C), the left vertical axis represents reversible heat flow (w/g), the right vertical axis represents irreversible heat flow (w/g), and the curves on the figure represent the total heat flow (W/g), irreversible heat flow (W/g) and reversible heat flow (W/g) from bottom to top.

Figure 22 is a TGA spectrum of the trisodium salt of the compound of formula (I). The abscissa represents temperature (°C), and the ordinate represents weight (%).

Figure 23 is the ¹ H-NMR spectrum of the trisodium salt of the compound of formula (I). The abscissa represents the compound displacement (ppm), and the ordinate represents the signal intensity.

Figure 24 is the mDSC spectrum of the monocalcium salt of the compound of formula (I). The abscissa represents temperature (°C), and the ordinate represents heat flow (w/g). The left first ordinate represents reversible heat flow (w/g), the left second ordinate represents total heat flow (w/g), and the right ordinate represents irreversible heat flow (w/g).

Figure 25 is a TGA spectrum of the monocalcium salt of the compound of formula (I). The abscissa represents temperature (°C), and the ordinate represents weight (%).

Figure 26 is the ¹ H-NMR spectrum of the monocalcium salt of the compound of formula (I). The abscissa represents the compound displacement (ppm), and the ordinate represents the signal intensity.

Figure 27 is the XRPD spectrum of the trisodium salt of the compound of formula (I) in Form A', where the abscissa represents the 2θ value (degrees) and the ordinate represents the peak intensity.

Figure 28 is the DSC spectrum of the trisodium salt form A' of the compound of formula (I), where the abscissa represents temperature (°C) and the ordinate represents heat flow (w/g).

Figure 29 is the TGA spectrum of the trisodium salt of the compound of formula (I) in form A', where the abscissa represents temperature (°C) and the ordinate represents weight (%).

FIG30 is ^{a 1} H-NMR spectrum of the trisodium salt of the compound of formula (I) in form A′, wherein the abscissa represents the compound displacement (ppm) and the ordinate represents the signal intensity.

Figure 31 is the XRPD spectrum of the trisodium salt form D' of the compound of formula (I), where the abscissa represents the 2θ value (degrees) and the ordinate represents the peak intensity.

Figure 32 is the XRPD spectrum of the trisodium salt of the compound of formula (I) in form E', where the abscissa represents the 2θ value (degrees) and the ordinate represents the peak intensity.

Figure 33 is the DSC spectrum of the trisodium salt of the compound of formula (I) in form E', where the abscissa represents temperature (°C) and the ordinate represents heat flow (w/g).

Figure 34 is the TGA spectrum of the trisodium salt of the compound of formula (I) in form E', where the abscissa represents temperature (°C) and the ordinate represents weight (%).

FIG35 is a ¹ H-NMR spectrum of the trisodium salt of the compound of formula (I) in form E', wherein the abscissa represents the compound displacement (ppm) and the ordinate represents the signal intensity.

Figure 36 is the XRPD spectrum of the trisodium salt form A1' of the compound of formula (I), where the abscissa represents the 2θ value (degrees) and the ordinate represents the peak intensity.

Figure 37 is the DSC spectrum of the trisodium salt of the compound of formula (I) in form A1', where the abscissa represents temperature (°C) and the ordinate represents heat flow (w/g).

Figure 38 is the TGA spectrum of the trisodium salt of the compound of formula (I) in form A1', where the abscissa represents temperature (°C) and the ordinate represents weight (%).

FIG39 is a ¹ H-NMR spectrum of the trisodium salt of the compound of formula (I) in form A1′, wherein the abscissa represents the compound displacement (ppm) and the ordinate represents the signal intensity.

Figure 40 is the XRPD spectrum of the trisodium salt form B' of the compound of formula (I), where the abscissa represents the 2θ value (degrees) and the ordinate represents the peak intensity.

Figure 41 is the DSC spectrum of the trisodium salt form B' of the compound of formula (I), where the abscissa represents temperature (°C) and the ordinate represents heat flow (w/g).

Figure 42 is the TGA spectrum of the trisodium salt of the compound of formula (I) in form B', where the abscissa represents temperature (°C) and the ordinate represents weight (%).

FIG43 is a ¹ H-NMR spectrum of the trisodium salt of the compound of formula (I) in form B′, wherein the abscissa represents the compound displacement (ppm) and the ordinate represents the signal intensity.

Figure 44 is the XRPD spectrum of the trisodium salt of the compound of formula (I) in form C', where the abscissa represents the 2θ value (degrees) and the ordinate represents the peak intensity.

Figure 45 is the XRPD spectrum of the trisodium salt form F' of the compound of formula (I), where the abscissa represents the 2θ value (degrees) and the ordinate represents the peak intensity.

Figure 46 is the DSC spectrum of the trisodium salt form F' of the compound of formula (I), where the abscissa represents temperature (°C) and the ordinate represents heat flow (w/g).

Figure 47 is the TGA spectrum of the trisodium salt form F' of the compound of formula (I), where the abscissa represents temperature (°C) and the ordinate represents weight (%).

Figure 48 is the 1H-NMR spectrum of the trisodium salt form F' of the compound of formula (I), where the abscissa represents the compound displacement (ppm) and the ordinate represents the signal intensity.

Figure 49 is an X-ray powder diffraction pattern of a sample of the free acid of the amorphous form (I) compound after DVS test. The abscissa represents the 2θ value (degrees), and the ordinate represents the peak intensity.

Figure 50 is an X-ray powder diffraction pattern of a sample of the free acid crystalline form B of the compound of formula (I) after DVS test. The abscissa represents the 2θ value (degrees), and the ordinate represents the peak intensity.

Figure 51 is an X-ray powder diffraction pattern of amorphous form (I) trisodium salt sample after DVS test. The abscissa represents 2θ value (degrees), and the ordinate represents peak intensity.

Figure 52 is an X-ray powder diffraction pattern of a sample of the monocalcium salt of the amorphous form (I) compound after DVS test. The abscissa represents the 2θ value (degrees), and the ordinate represents the peak intensity.

Figure 53 is an XRPD overlay of the trisodium salt form A1' of the compound of formula (I) before and after the DVS test. The abscissa represents the 2θ value (degrees), and the ordinate represents the peak intensity.

Figure 54 is an XRPD overlay of the trisodium salt of the compound of formula (I) in Form E' before and after DVS testing. The abscissa represents the 2θ value (degrees), and the ordinate represents the peak intensity.

DETAILED DESCRIPTION

The present invention studies the different aggregation forms of the compound of formula (I), provides a basic salt and crystalline free acid of the compound of formula (I), greatly improves the physicochemical properties of the free state of the compound of formula (I), such as chemical stability, solubility and hygroscopicity, so that the basic salt and crystalline free acid of the compound of formula (I) can meet the needs of clinical drug preparation development, have very important clinical application value, can be widely used in the preparation of treatments for tumors, immune-related diseases and metabolic diseases mediated at least in part by CD73, and are expected to accelerate the development of a new generation of CD73 inhibitor drugs. On this basis, the present invention is completed.

Detailed Description: Unless stated otherwise, the following terms used in the specification and claims have the following meanings.

In the present invention, the compound of formula (I) and the free acid of the compound of formula (I) have the same meaning and have the structure shown in the following formula (I):

"Pharmaceutical composition" means a mixture containing one or more compounds described herein or their physiologically/pharmaceutically acceptable salts, free acids or prodrugs and other chemical components, as well as other components such as physiologically/pharmaceutically acceptable carriers and excipients. The purpose of a pharmaceutical composition is to facilitate administration to an organism, facilitate the absorption of the active ingredient, and thus exert biological activity.

"Clinically effective dose" refers to the dose or concentration that can cause the minimum pharmacological effect clinically, also known as the minimum effective dose or threshold dose.

The basic salts and free acids of the compound of formula (I) show polymorphism or monocrystalline phenomena. For example, the trisodium salt shows polymorphism. These "polymorphs" differ in their X-ray powder diffraction patterns, physicochemical and pharmacokinetic properties and thermodynamic stability.

As used herein, "salt" refers to a compound prepared by reacting an organic acid or base drug with a pharmaceutically acceptable inorganic or organic acid or base.

(1) Methods and Materials

The reagents in the examples of the present invention are known and can be purchased on the market, or can be synthesized by or according to methods known in the art.

Unless otherwise specified, all reactions of the present invention are carried out under continuous magnetic stirring, the solvent is a dry solvent, and the temperature unit is Celsius (° C.) The environmental conditions refer to 20-25° C. and 60-80% RH.

Unless otherwise specified, the various crystal forms referred to in the present invention may be crystalline-free or crystalline-containing. If it is a crystalline-containing form, preferably each sub-crystal contains 1, 2, 3, 4 or 5 molecules of crystalline water, and more preferably each sub-crystal contains 1 or 2 molecules of crystalline water.

(2) HPLC test method (HPLC)

The stability and solubility of the compound were tested using HPLC. The instrument model used was: Agilent 1260 infinityII. Wavelength: 220 nm

Chromatographic column: XBridge BEH Shield RP18 (150mm×4.6mm, 2.5μm); detector: DAD; column temperature: 40℃; flow rate: 1.0mL/min; diluent: acetonitrile:water (v:v＝1:1); mobile phase A: 10mM ammonium acetate aqueous solution; mobile phase B: acetonitrile.

(3) X-ray powder diffraction test method (XRPD)

The crystal form of the sample was characterized by XRPD. The instrument model used was Bruker D8 Advance. The specific instrument parameters were as follows: X-ray optical path: reflection mode; detector: LYNXEYE_XE_T (1D mode); opening angle: Max; radiation source: Cu/K-Alpha1 Main beam path slit: dual main motorized slit 10.0mm SollerMount soller axis 2.5°; Secondary beam path slit: detector OpticsMount soller slit 2.5° dual secondary motorized slit 5.2mm; Scanning mode: continuous scanning; Scan type: dual optical path mode; Step length: 0.02°; Time per step: 0.12s/step; Scanning range: 3° to 40°; Sample rotation speed: 15rpm; Sample plate: single crystal silicon wafer, flat plate.

(4) Melting point and melting enthalpy test method (DSC)

The experimental method for characterizing the sample by differential scanning calorimetry (DSC) is to take a small amount of sample powder, place it in an aluminum pan that is compatible with the instrument and can be pressed, and then press the aluminum pan with the sample, and then send it to the instrument for detection. The instrument model used for differential scanning calorimetry in this patent is TA Discovery 2500 or Q2000, and the scanning parameters are set as follows: using nitrogen atmosphere, heating rate of 10℃/min, temperature range of 30 to 250℃.

(5) Modulated differential scanning calorimetry (mDSC)

Instrument: TA Discovery 2500; Sample: Tzero pan and Tzero sealed cover with 0.7mm diameter hole; Pan temperature range: 0℃/30℃ to 250℃; Heating rate: 2℃/min; Nitrogen flow rate: 50mL/min; Temperature adjustment range: +/-1℃/min; Sample amount: about 0.5-2mg.

(6) Thermogravimetric test method (TGA)

The experimental method of characterizing the sample by thermogravimetric analysis (TGA) is to take a small amount of sample powder, place it in an aluminum pan that matches the instrument, load the sample, and send it to the instrument for detection. The instrument model used in the thermogravimetric analysis method in this patent is TA Discovery 5500 or Q5000, and the scanning parameters are set to use a nitrogen atmosphere and a heating rate of 10℃/min. The termination temperature is 300℃ or the remaining sample weight is less than 80% of the initial weight.

(7) Residual solvent test method ( ¹ H-NMR)

The residual solvent was tested by nuclear magnetic resonance (1H-NMR). The instrument model used was Bruker Avance-AV 400M, the probe was 5mm PABBO BB/19F-1H/D Z-GRD Z108618/0406, the number of scans was 8 times, the temperature was 297.6K, and the relaxation delay was 1 second.

(8) Ion content test method (IC)

The sodium ion content was tested using ion chromatography (IC) method, and the instrument model used was Metrohm 940professional IC.

Detector: conductivity detector; eluent (anion): _3.2mmol /L _Na2CO3 +1.0mmol/L _NaHCO3 ; eluent (cation): 2.5mmol/L; MSA suppressor supplement: 0.5% _H2SO4 ; chromatographic column: Anion A SUPP 5-150 or Cation Column _C4-150 ; column temperature: 30°C; flow rate: 0.7mL/min (anion) or 0.9mL/min (cation); diluent: acetonitrile: water (v:v=1:1).

(9) Water content test method (KF)

The moisture content of samples was tested using the Karl Fischer (KF) coulometric method, and the instrument model used was Mettler Toledo Coulometric KF Titrator C30.

(10) Polarized light microscopy (PLM)

The samples were characterized by polarizing light microscopy (PLM), the instrument model used was BX53LED OLYMPUS, and the light source type was cross-polarized light. The samples were dispersed with silicone oil.

(11) Dynamic moisture sorption (DVS)

The experimental method for characterizing samples using the dynamic moisture sorption method (DVS) is to take a small amount of sample powder, place it in a precision sample tray that matches the instrument, load the sample, and send it to the instrument for detection. The instrument model used in the dynamic moisture sorption method in this patent is DVS Intrinsic. The experimental parameters are set to use nitrogen as the carrier gas, set the constant temperature to 25°C, and use the gradient mode for the test. The humidity change is 40%-95%-0%-95%-40% RH. The humidity change of each gradient in the range of 0% to 90% is 10%, and the humidity change of the gradient in the range of 90% to 95% is 5%. The gradient end point is judged by the dm/dt method, and the gradient end point is when dm/dt is less than 0.002% and maintained for 10 minutes, or the longest maintenance time of each gradient is 240 minutes. After the test is completed, the sample is analyzed by XRPD to confirm whether the solid form has changed.

The present invention is further described in detail and completely by the accompanying drawings and the following examples, which are only used to illustrate specific embodiments of the present invention and should not be interpreted as limiting the scope of the present invention in any way.

Example 1

(Preparation of the free acid of the compound of formula (I) and its pentasodium salt)

(1) Preparation method of free acid of compound of formula (I)

Step 1: Synthesis of (2R,3R,4R,5R)-2-(acetoxymethyl)-5-(4,6-dichloro-1H-pyrazolo[3,4-b]pyridin-1-yl)tetrahydrofuran-3,4-diyl diacetate

4,6-dichloro-1H-pyrazolo[3,4-b]pyridine (3.1 g, 16.49 mmol) was placed in hexamethyldisilazane (30 mL), and ammonium sulfate (22 mg, 0.16 mmol) was added. The reaction solution was heated to 140°C and stirred for 3 hours. The hexamethyldisilazane was removed by rotary evaporation under reduced pressure, and the residue was dissolved in acetonitrile (60 mL). β-D-furanose 1,2,3,5-tetraacetate (5.8 g, 18.14 mmol) was added. The reaction mixture was cooled to 0°C in an ice bath, and trimethylsilyl trifluoromethanesulfonate (5.5 g, 24.7 mmol) was added dropwise under ice bath cooling and stirring. The reaction solution was slowly heated to room temperature and stirred overnight. The reaction solution was evaporated under reduced pressure below 30°C to remove acetonitrile. The residue was diluted with ethyl acetate, washed with saturated sodium bicarbonate, separated, and the aqueous phase was extracted with ethyl acetate. The organic phases were combined and evaporated under reduced pressure. The residue was separated by column chromatography (ethyl acetate/petroleum ether=15/85) to obtain (2R,3R,4R,5R)-2-(acetoxymethyl)-5-(4,6-dichloro-1H-pyrazolo[3,4-b]pyridin-1-yl)tetrahydrofuran-3,4-diyl diacetate (5.6 g, yield: 76%). MS m/z (ESI): 446.2, 448.2 [M+H] ⁺ .

Step 2: Synthesis of (2R,3R,4R,5R)-2-(acetoxymethyl)-5-(6-chloro-4-(((R)-6-fluoro-2,3-dihydro-1H-inden-1-yl)amino)-1H-pyrazolo[3,4-b]pyridin-1-yl)tetrahydrofuran-3,4-diyl diacetate

(2R,3R,4R,5R)-2-(Acetoxymethyl)-5-(4,6-dichloro-1H-pyrazolo[3,4-b]pyridin-1-yl)tetrahydrofuran-3,4-diyl diacetate (0.40 g, 0.90 mmol) was dissolved in N-methylpyrrolidone (2.0 mL), (R)-6-fluoro-2,3-dihydro-1H-inden-1-amine hydrochloride (0.25 g, 1.35 mmol) and N,N-diisopropylethylamine (0.46 g, 3.60 mmol) were added, and the reaction was stirred in a sealed tube at 100°C for 44.3 hours. After the reaction is completed, water is added to dilute, and the mixture is extracted with ethyl acetate. The organic phase is washed with water and concentrated. The crude product is separated by column chromatography (EA/PE=4/6) to obtain (2R,3R,4R,5R)-2-(acetoxymethyl)-5-(6-chloro-4-(((R)-6-fluoro-2,3-dihydro-1H-inden-1-yl)amino)-1H-pyrazolo[3,4-b]pyridin-1-yl)tetrahydrofuran-3,4-diyl diacetate (0.44 g, yield 85%). MS m/z(ESI):561.3[M+H] ⁺ .

Step 3: Synthesis of (2R,3R,4S,5R)-2-(6-chloro-4-(((R)-6-fluoro-2,3-dihydro-1H-inden-1-yl)amino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol

(2R,3R,4R,5R)-2-(Acetoxymethyl)-5-(6-chloro-4-(((R)-6-fluoro-2,3-dihydro-1H-inden-1-yl)amino)-1H-pyrazolo[3,4-b]pyridin-1-yl)tetrahydrofuran-3,4-diyl diacetate (0.44 g, 0.78 mmol) was dissolved in methanol (3.0 mL), potassium carbonate (0.54 g, 3.90 mmol) was added, and the reaction was stirred at room temperature for 1 hour. After the reaction was completed, methanol was concentrated under reduced pressure, and the crude product was separated by column chromatography (DCM/MEOH=14/1), and concentrated to give (2R,3R,4S,5R)-2-(6-chloro-4-(((R)-6-fluoro-2,3-dihydro-1H-inden-1-yl)amino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-5-(hydroxymethyl)tetrahydrofuran-3,4-diol (0.27 g, yield 76%). MS m/z (ESI): 435.2 [M+H] ⁺ .

Step 4: Preparation of (((((2R,3S,4R,5R)-5-(6-chloro-4-(((R)-6-fluoro-2,3-dihydro-1H-inden-1-yl)amino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-3,4-dihydroxytetrahydrofuran-2-yl)methoxy)hydroxyphosphino)methyl)phosphocarboxylic acid

((3aR,4R,6R,6aR)-6-(6-chloro-4-(((R)-6-fluoro-2,3-dihydro-1H-inden-1-yl)amino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-2,2-dimethyltetrahydrofuro[3,4-d][1,3]dioxazol-4-yl)methanol (0.27 g, 0.62 mmol) was dissolved in anhydrous trimethyl phosphate (3.0 mL), cooled to 0°C, and a solution of methylenebisphosphonic dichloride (0.62 g, 2.50 mmol) in trimethyl phosphate (2.0 mL) was slowly added dropwise. After the addition was completed, the reaction was continued at 0°C for 30 minutes. After the reaction was completed, water was added to quench the reaction. After stirring for 10 minutes, the mixture was directly purified by reverse phase (H ₂ O/ACN=4/1) to obtain the target compound (((((2R,3S,4R,5R)-5-(6-chloro-4-(((R)-6-fluoro-2,3-dihydro-1H-inden-1-yl)amino)-1H-pyrazolo[3,4-b]pyridin-1-yl)-3,4-dihydroxytetrahydrofuran-2-yl)methoxy)hydroxyphosphino)methyl)phosphinocarboxylic acid (89.4 mg, yield 24%). MS m/z(ESI):593.0[M+H] ⁺ .

¹ H NMR (400MHz, MeOH-d ₄ )δ8.19(s,1H),7.28(dd,J＝8.3,5.1Hz,1H),7.00(ddd,J＝17.9,8.9,2.4Hz,2H),6.50(s,1H),6.33( d,J＝3.7Hz,1H),5.32(t,J＝7.3Hz,1H),4.71(dd,J＝5.2,3.8Hz,1H),4.57(t,J＝5.2Hz,1H),4.30(ddd ,J＝11.0,7.3,3.8Hz,1H),4.26–4.08(m,2H),3.05(ddd,J＝16.1,8.7,3.9Hz,1H),2.93(dt,J＝15.9,8 .0Hz,1H),2.70(dtd,J＝11.9,7.6,4.0Hz,2H),2.42(t,J＝21.0Hz,2H),2.07(dq,J＝12.9,8.0Hz,1H).

(2) Preparation method of pentasodium salt of compound of formula (I) (API:Na=1:5)

The free acid of the compound of formula (I) was dispersed in water, an excess of sodium hydroxide aqueous solution was added, and the mixture was stirred at room temperature until it became clear, and methanol was slowly added until solids precipitated, and the mixture was stirred at room temperature overnight, and the solids were collected. The molar ratio of API:Na was confirmed to be 1:5 by ion chromatography.

(3) Characterization of the free acid and pentasodium salt (API:Na=1:5) of the compound of formula (I)

The relevant properties of the compound of formula (I) and its sodium salt prepared above were measured by HPLC, XRPD, DSC, TGA, ¹ H-NMR, IC, and KF as follows. The specific test methods are detailed in the definition:

Example 2

(Screening of free acid crystal forms of compound of formula (I))

(1) 25℃ 3-week suspension experiment

About 40 mg of the free acid of the compound of formula (I) was weighed, 0.1-0.5 mL of solvent was added, and the mixture was suspended at 25° C. and 300 rpm with magnetic stirring for 3 weeks. The obtained suspension was centrifuged at 14,000 rpm using a 0.45 μm nylon filter centrifuge tube, and the obtained solid part was characterized by XRPD. The experimental results are shown in the table below.

(2) 50℃ 3-week suspension experiment

About 50 mg of the free acid of the compound of formula (I) was weighed, 0.1-0.5 mL of solvent was added, and the mixture was suspended at 50° C. with magnetic stirring at 300 rpm for 3 weeks. The obtained suspension was centrifuged at 14,000 rpm using a 0.45 μm nylon filter centrifuge tube, and the obtained solid part was characterized by XRPD. The experimental results are shown in the table below.

(3) Antisolvent drop crystallization experiment

Weigh about 40 mg of the free acid of the compound of formula (I), add 0.1-5 mL of a good solvent to fully dissolve at ambient temperature (about 22-25° C.), and pass the resulting solution through a 0.45 μm nylon filter syringe filter to obtain a clear solution. Slowly add 1-4 times the volume of the anti-solvent to the clear solution until a large amount of solid precipitates. The resulting sample is suspended at 25° C. for one week, and the resulting solid is characterized by polarizing microscopy (PLM). The experimental results are shown in the table below.

Example 3

(Salt type screening test)

The screening experiment was carried out on the selected counterions (basic cations) and screening solvents by the suspension method. About 50 mg of the free acid of the compound of formula (I) was weighed and placed in a 2 mL glass bottle with 1-3 equivalents of counterions, and an appropriate volume of screening solvent was added to carry out the screening experiment by the suspension method. The obtained sample was suspended at 50°C for 2 hours, then naturally cooled to 25°C, and suspended at 25°C for 1 week. The obtained suspension was centrifuged at 14,000 rpm through a 0.45 μm nylon filter membrane. The experimental results are shown in the table below. The obtained solid was characterized by XRPD, and the obtained good solid was characterized by nuclear magnetic resonance ( ¹ H-NMR) or ion chromatography (IC) for salt formation ratio.

(1) Experiment number RC0:

About 30 mg of pentasodium salt (API:Na=1:5) was weighed and suspended in 0.2 mL of ethanol at 25°C for 3 days. The obtained solid was an amorphous trisodium salt (API:Na=1:3).

(2) Screening solvent: ethanol

(3) Screening solvent - acetone

(4) Screening solvent - dichloromethane

(5) Salt type characterization

The following 7 amorphous salt forms were obtained from the above screening experiment, which were characterized by ¹ H-NMR, IC and HPLC (notes), respectively. The results are shown in the table below.

Example 4

(Preparation of Free Acid Crystal Form B of Compound of Formula (I))

The free acid form B of the compound of formula (I) was prepared and obtained according to the following steps.

1) Weigh about 300 mg of the free acid of the compound of formula (I) and suspend it in 1 mL of isopropanol.

2) The above suspension was suspended at 25° C. at 300 rpm for 4 days.

3) Add 5 mg of Form B seed crystals (obtained in Example 2, Experiment EQ10) to the above suspension.

4) Add 1 mL of isopropanol to the above suspension, and continue to suspend the resulting suspension for 2 weeks.

5) The solid part was collected by filtration and dried at 22-25°C/10-20%RH for about 20 hours. A total of 215 mg of free acid form B of the compound of formula (I) was obtained with a purity of 95.2% and a yield of 70%.

Through characterization of the above product, XRPD diagram shows that its crystallinity is moderate, see Figure 8, DSC shows that it has no melting point after dehydration/desolvation at 31°C and 142°C, see Figure 18. TGA shows that it loses about 4.5% of weight at 100°C and loses 2.2% of weight between 100°C and 160°C, see Figure 19. ¹ H-NMR shows that it contains 0.3 equivalents of isopropanol, equivalent to about 2.8%, see Figure 20.

Example 5

(Preparation of trisodium salt of amorphous form (I) compound)

(1) Preparation method 1:

The trisodium salt of the amorphous form (I) compound was prepared and obtained according to the following steps:

1) Weigh about 90 mg of pentasodium salt of the compound of formula (I) (API:Na=1:5) and suspend it in 0.6 mL of ethanol at 25°C for 2 days.

2) The solid portion was collected by filtration and dried under 22-25°C/40-60%RH conditions for about 2 hours.

3) A total of 60 mg of trisodium salt of amorphous form (I) of the compound was obtained as off-white powder with a purity of 99.8% and a yield of 72%.

(2) Preparation method 2:

The trisodium salt of the amorphous form (I) compound was prepared and obtained according to the following steps:

1) Weigh 3.2 equivalents of sodium hydroxide (about 109 mg), add 3 mL of ethanol, and stir at 50°C for 30 min to obtain a clear solution.

2) At 50° C., the free acid solution of the compound of formula (I) was slowly added into the sodium hydroxide solution, and the obtained sample was converted into a suspension.

3) Add 5 mg of amorphous trisodium salt seeds (the product of the above method 1) to the above suspension.

4) The suspension was stirred at 50°C for 2 hours, cooled to 25°C, and stirred at 25°C for 2 days.

5) The solvent of the suspension was removed by a rotary evaporator, and the obtained solid was suspended in 3 mL of ethanol for another 3 days.

6) The solid part was collected by filtration and dried under vacuum at 35° C. for about 2 hours. A total of 502 mg of trisodium salt of amorphous form (I) compound was obtained as off-white powder with a purity of 97.1% and a yield of 88%.

Through the analysis of the above product, XRPD shows that it is close to amorphous, see Figure 11, and DSC shows that it has no obvious Tg, see Figure 21. ¹ H-NMR or IC shows that its stoichiometric ratio is 1:3, and TGA shows that the weight loss at 245°C is about 14.1%, see Figure 22. ¹ H-NMR shows that it contains 0.7 equivalents of ethanol, equivalent to about 4.2%, see Figure 23.

Note: The free acid of the compound of formula (I) in the above method 2 can be replaced by the free acid crystalline form B isopropanol-water solvate.

Example 6

(Scale-up preparation of amorphous monocalcium salt)

The amorphous form of compound (I) monocalcium salt was prepared and obtained according to the following steps.

1) Weigh 500 mg of the free acid of the compound of formula (I), add 5 mL of ethanol, and stir at 50° C. for 0.5 hour to obtain a nearly clear solution.

2) Weigh 1.05 equivalents of calcium hydroxide (about 62 mg) and add it to the above clear solution.

3) About 5 mg of amorphous monocalcium salt seeds (RC5A product in Example 3.1) were added to the above suspension.

4) The suspension was stirred at 50°C for 2 hours, cooled to 25°C, and stirred at 25°C for 2 days.

5) The solid portion was collected by filtration and dried under vacuum at 30°C for about 1 hour.

6) A total of 511 mg of amorphous form (I) compound monocalcium salt was obtained with a purity of 97% and a yield of 95%.

Through characterization of the above product, the XRPD pattern shows that it is amorphous, see Figure 13. DSC shows that it has no obvious Tg, see Figure 24. ¹ H-NMR or IC shows that its stoichiometric ratio is 1:1, and TGA shows that the weight loss is about 8.6% at 150°C, see Figure 25. ¹ H-NMR shows that it contains 0.7 equivalents of ethanol, equivalent to about 4.6%, see Figure 26.

Example 7

(Preparation of the trisodium salt crystal form of the compound of formula (I))

For the trisodium salt of the compound of formula (I), more than one hundred screening experiments were conducted under different screening conditions, and the following seven crystalline forms of the trisodium salt of the compound of formula (I) were obtained.

7.1 Preparation of Trisodium Salt Form A' of Compound of Formula (I)

(1) Preparation method 1:

1) Weigh 50 mg of the trisodium salt of the compound of formula (I) and add 0.2 mL of tetrahydrofuran/water (v:v=95:5).

2) Suspend at 50°C with magnetic stirring at 400 rpm for 1 week.

3) The obtained suspension was centrifuged and filtered at 14,000 rpm using a 0.45 μm nylon filter membrane centrifuge tube to obtain the trisodium salt form A' of the compound of formula (I).

(2) Preparation method 2:

1) Weigh about 500 mg of the trisodium salt of amorphous form (I) compound and add it into an 8 mL glass bottle.

2) Add 5 mL of a mixed solvent of tetrahydrofuran:water (v:v=95:5), and stir at 50° C. to obtain a suspension.

3) Add 5 mg of Form A' seed crystals (product of method 1) to the resulting suspension.

4) The obtained suspension was suspended at 50°C with a magnetic stirrer at 400 rpm for 1 week.

5) The obtained solid was collected by suction filtration and vacuum dried at 50°C for about 2 hours to obtain 440 mg of the trisodium salt of the compound of formula (I) in the form A' as an off-white powder with a yield of 80%.

The product was characterized by XRPD as shown in Figure 27, DSC at 26.8°C for dehydration/desolvation; no melting point after dehydration/desolvation, as shown in Figure 28. TGA showed a weight loss of about 14.5% at 200°C, as shown in Figure 29. ¹ H-NMR showed that it contained 0.3 equivalents of tetrahydrofuran, equivalent to about 3.2%, as shown in Figure 30.

7.2 Preparation of the trisodium salt of the compound of formula (I) in form D'

30 mg of trisodium salt of the compound of formula (I) Form A’ was weighed and placed at about 22-25°C/60-80% RH for 12 hours. Form A’ was completely transformed into Form D’. Its XRPD pattern is shown in Figure 31.

7.3 Preparation of the trisodium salt of the compound of formula (I) in form E'

30 mg of the trisodium salt of the compound of formula (I) crystalline form A' was weighed and placed at about 22-25°C/60-75% RH for 20 hours, and then placed at 22-25°C, 20-30% RH for 30 minutes to obtain crystalline form E' with a purity of 97.9%. Its XRPD characterization is shown in Figure 32. DSC dehydrated at 22.6°C, and there was no melting point after dehydration, as shown in Figure 33. TGA showed a weight loss of about 10.6% at 200°C, as shown in Figure 34. ¹ H-NMR showed that it contained 0.1 equivalents of tetrahydrofuran equivalent to about 1.1%, as shown in Figure 35, and KF measured its water content to be 9.3%.

7.4 Preparation of the trisodium salt of the compound of formula (I) Form A1'

Weigh 200 mg of the trisodium salt of the compound of formula (I) in Form A', place it in an open container at 22-25°C/75%RH for 2 days, and then place it at 22-25°C/20-30%RH for about 30 minutes, and the resulting form A1' has a low crystallinity and a purity of 97.8%. Its XRPD diagram is shown in Figure 36. DSC dehydrates at 23.9°C, and there is no melting point after dehydration, see Figure 37. TGA shows a weight loss of about 16.1% at 245°C, see Figure 38. ¹ H-NMR shows that it contains 0.1 equivalents of tetrahydrofuran, equivalent to about 1.1%, see Figure 39, and KF measures its water content to be 11.5%.

7.5 Preparation of Trisodium Salt Form B' of Compound of Formula (I)

(1) Preparation method 1:

1) Weigh 50 mg of the trisodium salt of the compound of formula (I) and add 0.2 mL of methyl pyrrolidone.

2) Suspend at 25°C with magnetic stirring at 400 rpm for 2 weeks.

3) The obtained suspension was centrifuged and filtered at 14,000 rpm using a 0.45 μm nylon filter membrane centrifuge tube to obtain the trisodium salt form B' of the compound of formula (I).

(2) Preparation method 2:

1) Weigh 50 mg of the trisodium salt of the compound of formula (I) and add 0.2 mL of methyl pyrrolidone.

2) Suspend at 50°C with magnetic stirring at 400 rpm for 1 week.

3) The obtained suspension was centrifuged and filtered at 14,000 rpm using a 0.45 μm nylon filter membrane centrifuge tube to obtain the trisodium salt form B' of the compound of formula (I).

The product was characterized by XRPD as shown in Figure 40, DSC at 19.1°C for dehydration/desolvation; no melting point after dehydration/desolvation, as shown in Figure 41. TGA showed a weight loss of about 20.7% at 245°C, as shown in Figure 42. ¹ H-NMR showed that it contained 0.55 equivalents of methylpyrrolidone, equivalent to about 8.5%, as shown in Figure 43.

7.6 Preparation of the trisodium salt of the compound of formula (I) Form C'

1) Weigh 50 mg of the trisodium salt of the compound of formula (I) and add 0.2 mL of 1,4-dioxane/water (V:V=95:5).

2) Suspend at 25°C with magnetic stirring at 400 rpm for 2 weeks.

3) The obtained suspension was centrifuged and filtered at 14,000 rpm using a 0.45 μm nylon filter membrane centrifuge tube to obtain the trisodium salt form C' of the compound of formula (I), characterized by XRPD, as shown in Figure 44.

7.7 Preparation of the trisodium salt of the compound of formula (I) Form F'

Weigh 200 mg of the trisodium salt of the compound of formula (I) in form C', and dry it in vacuum at 50°C for 1 hour to transform form C' into form F', and its XRPD characterization is shown in Figure 45. DSC desolventizes at 30.0°C, and there is no melting point after desolventizes, see Figure 46. TGA shows a weight loss of about 15.3% at 230°C, see Figure 47. ¹ H-NMR shows that it contains 1.5 equivalents of 1,4-dioxane, equivalent to about 16.8%, see Figure 48.

Example 8

(Solid physical and chemical stability)

This experiment examines the solid stability of the free acid of the amorphous form (I), the free acid crystalline form B (isopropanol-water solvate) of the compound of formula (I), the trisodium salt of the amorphous form (I), the monocalcium salt of the amorphous form (I), the trisodium salt crystalline form A1' of the compound of formula (I), and the trisodium salt crystalline form E' of the compound of formula (I) under the following experimental conditions.

1) BS1: solid, 25°C/92%RH, open conditions, 1 week.

2) BS2: solid, 40°C/75%RH, open conditions, 1 week.

3) BS3: solid, 60°C, sealed container, 1 week.

4) BS4: solid, light (visible light, 1.2 million lumens per hour).

5) BS5: solid, 25°C/92%RH, open conditions, 8 weeks.

6) BS6: solid, 40°C/75%RH, open conditions, 8 weeks.

7) BS7: solid, 60°C, sealed container, 8 weeks.

The test results are as follows:

(1) Changes in chemical purity

(2) Changes in physical state

(3) Appearance color change

From the above experimental results, it can be seen that the purity of amorphous free acid and free acid crystal form B decreases significantly under 40°C/75% RH, 60°C and light conditions, and they are chemically unstable.

The crystalline free acid form B is physically unstable and transforms into an amorphous form under the conditions of 25°C/92%RH, 40°C/75%RH, 60°C and light, and is physically unstable.

Compared with the free acid crystalline form B, the amorphous monocalcium salt and the amorphous trisodium salt exhibit excellent chemical purity stability. The chemical properties are stable under 25°C/92%RH, 40°C/75%RH, 60°C and light conditions.

The trisodium salt forms A1’ and E’ are chemically stable at 25°C/92%RH, 40°C/75%RH, and 60°C.

Compared with the amorphous trisodium salt, the chemical purity of the amorphous monocalcium salt is slightly reduced (about 1%) after exposure to light, but the light degradation problem of the monocalcium salt can be effectively avoided under light-shielding conditions.

Compared with the amorphous trisodium salt, the crystalline trisodium salts A1' and E' transformed into amorphous form under 25°C/92%RH and 40°C/75%RH conditions, deliquesced, and were accompanied by a slight color change, and were physically unstable.

Therefore, compared with the amorphous free acid, the free acid crystalline form B and the amorphous monocalcium salt, the amorphous trisodium salt has a significant advantage in chemical stability. Compared with the crystalline trisodium salts A1' and E', the amorphous trisodium salt has a significant advantage in physical stability.

Example 9

(Solubility Test)

A certain amount of the free acid of the amorphous form (I) compound; the free acid crystalline form B (isopropanol-water solvate) of the compound of formula (I); and the trisodium salt of the amorphous form (I) compound were accurately weighed and placed in 2 mL glass bottles (the masses of the weighed free acid and candidate salt were equivalent to 10 mg of anhydrous free acid, respectively), 1 mL of solvents of different pH values were added (the preparation method is shown in the table below) to obtain clear solutions, and the pH values of the clear solutions were measured by a pH meter.

2.3 mg of amorphous form (I) compound monocalcium salt was accurately weighed and placed in a 2 mL glass bottle. The mass of the amorphous form (I) compound monocalcium salt was equivalent to 2 mg of free acid anhydrous crystalline form of the compound of formula (I). The resulting suspension was stirred at 400 rpm for 24 hours at 37°C. The sample was centrifuged at 14,000 rpm for 5 min at 37°C. The concentration of the supernatant was determined by HPLC and the pH value of the supernatant was determined by a pH meter. The residual solid portion after the solubility experiment was characterized by XRPD. The test results are shown in the table below.

At ambient temperature (about 22-25° C.), pH 7.4 phosphate buffer solution (50 mM) was added to 200 mg of trisodium salt form A1′ of the compound of formula (I) (equivalent to nearly 150 mg of anhydrous free acid), the pH value of the resulting solution was adjusted to 7.8, and the solution was then stirred at 25° C. for 3 days without obvious precipitation.

The free acid of the amorphous form (I), the free acid form B of the compound of formula (I), and the trisodium salt of the amorphous form (I) all have high solubility (>10 mg/mL) in the five aqueous buffers of different pH values. The pH drift increases with increasing concentration in the pH 4.5 acetate buffer, pH 6.8 phosphate buffer, and pH 7.4 phosphate buffer.

The trisodium salt form A1' of the compound of formula (I) has a very high solubility of about 150 mg/mL in pH 7.4 phosphate buffer, which is much higher than the solubility of the monocalcium salt of the amorphous form of the compound (I).

The monocalcium salt of amorphous form (I) of the compound has a low solubility (~0.5 mg/mL) in these five aqueous buffers and the solid obtained after the solubility experiment is still amorphous.

Example 10

(Hygroscopicity test)

The hygroscopicity of the amorphous free acid, free acid form B, amorphous trisodium salt and amorphous monocalcium salt was evaluated by dynamic moisture sorption (DVS) test at 25 °C.

From the data analysis in the above table, we can see that:

1) The free acid of the amorphous form (I) compound has strong hygroscopicity, and its weight gain from moisture absorption between 0% RH and 80% RH at 25° C. is about 14.7%. After DVS test, the obtained solid is still amorphous, as shown in FIG49 .

2) The free acid crystal form B of the compound of formula (I) has strong hygroscopicity, and its weight gain from moisture absorption between 0% RH and 80% RH at 25° C. is about 13.1%. After DVS test, the free acid crystal form B transforms into an amorphous form, as shown in FIG50 .

3) The trisodium salt of the amorphous form (I) compound has strong hygroscopicity, and its weight gain from moisture absorption between 0% RH and 80% RH at 25° C. is about 29.1%. After DVS test, the obtained solid is amorphous, as shown in FIG51 .

4) The monocalcium salt of the amorphous form (I) compound has strong hygroscopicity, and its weight gain from moisture absorption between 0% RH and 80% RH at 25° C. is about 14.1%. After the DVS test, the obtained solid is still amorphous, as shown in FIG52 .

5) The trisodium salt of the compound of formula (I) in form A1' has strong hygroscopicity, and its weight gain from moisture absorption between 0% RH and 80% RH at 25°C is about 35.9. After DVS test, the obtained solid is amorphous, as shown in Figure 53.

6) The trisodium salt of the compound of formula (I) in the form E' has strong hygroscopicity, and its weight gain from moisture absorption at 0% RH to 80% RH at 25°C is about 33.0%. After DVS test, the obtained solid is amorphous, as shown in Figure 54.

All documents mentioned in the present invention are cited as references in the present invention, just as each document is cited as references separately. In addition, it should be understood that after reading the above teachings of the present invention, those skilled in the art can make various changes or modifications to the present invention, and these equivalent forms also fall within the scope of the present invention.

Claims (29)

A basic salt of a compound of formula (I),
Wherein, the basic salt is sodium salt, potassium salt, calcium salt, arginine salt, meglumine salt or ammonium salt. The basic salt of the compound of formula (I) according to claim 1, characterized in that the basic salt is a sodium salt or a calcium salt. The basic salt of the compound of formula (I) according to claim 1, characterized in that the molar ratio of the compound of formula (I) to the base in each molecule of the basic salt of the compound of formula (I) is 1:1 to 1:5; preferably, the molar ratio of the compound of formula (I) to the base is 1:1, 1:2, 1:3, 1:4 or 1:5. The basic salt of the compound of formula (I) according to claim 1, characterized in that the basic salt of the compound of formula (I) is a sodium salt of the compound of formula (I), and the molar ratio of the compound of formula (I) to sodium atoms in each molecule of the sodium salt of the compound of formula (I) is 1:1, 1:3 or 1:5. The basic salt of the compound of formula (I) according to claim 4, characterized in that the molar ratio of the compound of formula (I) to sodium atoms in each molecule of the sodium salt of the compound of formula (I) is 1:3. The basic salt of the compound of formula (I) according to claim 4, characterized in that the sodium salt of the compound of formula (I) is an amorphous or crystalline compound. The basic salt of the compound of formula (I) according to claim 6, characterized in that the basic salt of the compound of formula (I) is the trisodium salt of the amorphous form of the compound (I). The basic salt of the compound of formula (I) according to claim 6, characterized in that the basic salt of the compound of formula (I) is a crystalline trisodium salt of the compound of formula (I). The basic salt of the compound of formula (I) according to claim 8, characterized in that the crystalline form of the trisodium salt of the compound of formula (I) is form A', and its X-ray powder diffraction pattern (XRPD) includes five or more peaks at diffraction angles (2θ) of 3.22±0.2°, 6.38±0.2°, 9.55±0.2°, 12.72±0.2°, 19.10±0.2°, 20.01±0.2°, 20.53±0.2°, 23.13±0.2°, 25.50±0.2°, 26.27±0.2°, 26.66±0.2°, 28.80±0.2°, 29.43±0.2°, 32.06±0.2° and 35.37±0.2°; Preferably, the X-ray powder diffraction pattern of the crystalline form A' comprises peaks at diffraction angles (2θ) substantially the same as those shown in Figure 27. The basic salt of the compound of formula (I) according to claim 8, characterized in that the crystalline form of the trisodium salt of the compound of formula (I) is Form B', and its X-ray powder diffraction pattern (XRPD) includes five or more peaks at diffraction angles (2θ) of 3.15±0.2°, 6.27±0.2°, 9.34±0.2°, 12.34±0.2°, 12.59±0.2°, 15.64±0.2°, 17.06±0.2°, 18.90±0.2°, 23.60±0.2° and 25.93±0.2°; Preferably, the X-ray powder diffraction pattern of the crystalline form B' comprises peaks at diffraction angles (2θ) substantially the same as those shown in Figure 40. The basic salt of the compound of formula (I) according to claim 8, characterized in that the crystalline form of the trisodium salt of the compound of formula (I) is form C', and its X-ray powder diffraction pattern (XRPD) includes five or more peaks at diffraction angles (2θ) of 11.4±0.2°, 17.09±0.2°, 17.68±0.2°, 19.01±0.2°, 19.59±0.2°, 19.97±0.2°, 20.81±0.2°, 21.65±0.2°, 22.39±0.2°, 22.81±0.2°, 24.80±0.2°, 27.06±0.2°, 27.73±0.2°, 30.53±0.2° and 32.62±0.2°; Preferably, the X-ray powder diffraction pattern of the crystalline form C' comprises peaks at diffraction angles (2θ) substantially the same as those shown in Figure 44. The basic salt of the compound of formula (I) according to claim 8, characterized in that the crystalline form of the trisodium salt of the compound of formula (I) is form D', and its X-ray powder diffraction pattern (XRPD) includes five or more peaks at diffraction angles (2θ) of 11.97±0.2°, 17.97±0.2°, 18.92±0.2°, 19.41±0.2°, 20.36±0.2°, 20.97±0.2°, 21.66±0.2°, 22.88±0.2°, 23.24±0.2°, 24.15±0.2°, 24.77±0.2°, 25.11±0.2°, 25.51±0.2°, 26.21±0.2° and 30.10±0.2°; Preferably, the X-ray powder diffraction pattern of the crystalline form D' comprises peaks at diffraction angles (2θ) substantially the same as those shown in Figure 31. The basic salt of the compound of formula (I) according to claim 8, characterized in that the crystalline form of the trisodium salt of the compound of formula (I) is form E', and its X-ray powder diffraction pattern (XRPD) includes five or more peaks at diffraction angles (2θ) of 3.15±0.2°, 3.24±0.2°, 9.79±0.2°, 12.35±0.2°, 12.41±0.2°, 14.30±0.2°, 18.61±0.2°, 19.91±0.2°, 20.80±0.2°, 23.94±0.2°, 24.89±0.2°, 31.39±0.2° and 34.67±0.2°; Preferably, the X-ray powder diffraction pattern of the crystalline form E' comprises peaks at diffraction angles (2θ) substantially the same as those shown in Figure 32. The basic salt of the compound of formula (I) according to claim 8, characterized in that the crystalline form of the trisodium salt of the compound of formula (I) is a crystalline form F', and its X-ray powder diffraction pattern (XRPD) includes peaks at diffraction angles (2θ) of 3.12±0.2°, 5.81±0.2°, 6.27±0.2°, 9.43±0.2°, 18.94±0.2° and 25.58±0.2°; Preferably, the X-ray powder diffraction pattern of the crystalline form F' comprises peaks at diffraction angles (2θ) substantially the same as those shown in Figure 45. The basic salt of the compound of formula (I) according to claim 8, characterized in that the crystalline form of the trisodium salt of the compound of formula (I) is form A1', and its X-ray powder diffraction pattern (XRPD) includes five or more peaks at diffraction angles (2θ) of 3.19±0.2°, 9.55±0.2°, 12.71±0.2°, 13.52±0.2°, 14.43±0.2°, 16.37±0.2°, 19.07±0.2°, 19.99±0.2°, 20.48±0.2°, 24.02±0.2°, 25.06±0.2°, 25.43±0.2°, 26.21±0.2°, 26.60±0.2° and 29.40±0.2°; Preferably, the X-ray powder diffraction pattern of the crystalline form A1' comprises peaks substantially the same as those at diffraction angles (2θ) shown in Figure 36. The basic salt of the compound of formula (I) according to claim 1, characterized in that the basic salt of the compound of formula (I) is a calcium salt of the compound of formula (I), and the molar ratio of the compound of formula (I) per molecule to the calcium atom in the calcium salt of the compound of formula (I) is 1:1. A method for preparing a basic salt of the compound of formula (I) according to claim 1, characterized in that it comprises the following steps: 1) dissolving or dispersing the free acid of the compound of formula (I) in water or an organic solvent, and adding an alkaline solution to the above system to carry out a salt-forming reaction; or, adding the free acid of the compound of formula (I) to an alkaline solution to carry out a salt-forming reaction; 2) collecting the solid product basic salt of the compound of formula (I) precipitated in the above-mentioned salt-forming reaction process, or obtaining the solid product basic salt of the compound of formula (I) by creating supersaturation in the salt-forming system; The free acid of the compound of formula (I) is an anhydrate, a hydrate or a solvate; The basic salt of the compound of formula (I) is a sodium salt, potassium salt, calcium salt, arginine salt, meglumine salt or ammonium salt of the compound of formula (I); The alkaline solution is selected from an aqueous solution or an organic solvent solution of sodium hydroxide, potassium hydroxide, calcium hydroxide, arginine, meglumine or ammonia. A method for preparing a basic salt of a compound of formula (I) according to claim 6, characterized in that it comprises the following steps: 1) dissolving or dispersing pentasodium salt of the compound of formula (I) in water or an organic solvent; 2) stirring or suspending, and collecting the trisodium salt solid of the compound of formula (I) precipitated in the above process. The preparation method according to claim 17 is characterized in that the method of creating supersaturation in the salt-forming system in step 2) comprises one or more of the following: volatilizing the solvent, adding an anti-solvent or cooling. The preparation method according to claim 17 or 18, characterized in that the organic solvent is selected from alcohols, chloroalkanes, ketones, ethers, cyclic ethers, esters, alkanes, cycloalkanes, benzenes, amides or sulfoxide organic solvents, or mixtures thereof, or aqueous solutions thereof; Preferably, the organic solvent is selected from methanol, ethanol, n-propanol, isopropanol, dichloromethane, heptane, acetonitrile, acetone, methyl ethyl ketone, toluene, 1,4-dioxane, tetrahydrofuran, 2-methyltetrahydrofuran, N,N-dimethylformamide, dimethyl sulfoxide, ethyl acetate, isopropyl acetate, methyl tert-butyl ether or 2-methoxyethyl ether, or a mixture thereof, or an aqueous solution thereof. A crystalline form of the free acid of the compound of formula (I):
The crystalline form of the free acid of the compound of formula (I) according to claim 21, characterized in that the free acid of the crystalline form of the compound of formula (I) is Form A, and its X-ray powder diffraction pattern (XRPD) includes five or more peaks at diffraction angles (2θ) of 5.56±0.2°, 6.55±0.2°, 7.24±0.2°, 9.83±0.2°, 11.13±0.2°, 13.14±0.2°, 15.81±0.2°, 18.01±0.2°, 19.75±0.2°, 20.73±0.2°, 22.38±0.2°, 23.10±0.2°, 23.37±0.2° and 26.28±0.2°; Preferably, the X-ray powder diffraction pattern of the crystalline form A is substantially the same as the peak at the diffraction angle (2θ) shown in FIG. 9 . The crystalline form of the free acid of the compound of formula (I) according to claim 21, characterized in that the free acid of the crystalline form of the compound of formula (I) is Form B, and its X-ray powder diffraction pattern (XRPD) includes five or more peaks at diffraction angles (2θ) of 4.76±0.2°, 7.21±0.2°, 15.17±0.2°, 16.58±0.2°, 17.03±0.2°, 17.90±0.2°, 19.12±0.2°, 21.32±0.2°, 22.05±0.2°, 23.54±0.2°, 24.32±0.2°, 24.58±0.2°, 25.73±0.2°, 26.08±0.2° and 26.70±0.2°; Preferably, the X-ray powder diffraction pattern of the crystalline form B is substantially the same as the peak at the diffraction angle (2θ) shown in FIG8 . A pharmaceutical composition comprising: A clinically effective amount of a basic salt of a compound of formula (I) according to any one of claims 1 to 16, or a crystalline form of a free acid of a compound of formula (I) according to any one of claims 21 to 23; and a pharmaceutically acceptable carrier. Use of a basic salt of a compound of formula (I) as described in any one of claims 1 to 16, or a crystalline free acid of a compound of formula (I) as described in any one of claims 21 to 23, or a pharmaceutical composition as described in claim 24 in the preparation of a medicament for treating tumors, immune-related diseases and disorders or metabolic diseases mediated at least in part by CD73. The use according to claim 25, characterized in that the tumor is selected from prostate cancer, colon cancer, rectal cancer, pancreatic cancer, gastric cancer, endometrial cancer, cervical cancer, brain cancer, liver cancer, bladder cancer, ovarian cancer, testicular cancer, head cancer, neck cancer, skin cancer, mesothelial lining cancer, white blood cell cancer, esophageal cancer, breast cancer, muscle cancer, connective tissue cancer, lung cancer, adrenal cancer, thyroid cancer, kidney cancer, bone cancer, brain tumor, glioblastoma, mesothelioma, renal cell carcinoma, sarcoma, choriocarcinoma, epidermal basal cell carcinoma and testicular seminoma; Preferably, the tumor is selected from the group consisting of skin cancer, colon cancer, pancreatic cancer, breast cancer, prostate cancer, lung cancer, leukemia cancer, brain tumor, ovarian cancer and sarcoma. The use according to claim 25 is characterized in that the immune-related diseases and disorders are selected from rheumatoid arthritis, renal failure, lupus erythematosus, asthma, psoriasis, ulcerative colitis, pancreatitis, allergies, fibrosis, anemia fibromyalgia, Alzheimer's disease, congestive heart failure, stroke, aortic valve stenosis, arteriosclerosis, osteoporosis, Parkinson's disease, infection, Crohn's disease, ulcerative colitis, allergic contact dermatitis and eczema, systemic sclerosis and multiple sclerosis. A basic salt of a compound of formula (I) as described in any one of claims 1 to 16, or a crystalline free acid of a compound of formula (I) as described in any one of claims 21 to 23, or a pharmaceutical composition as described in claim 24, which is used as a drug for treating tumors, immune diseases and disorders or metabolic diseases mediated at least in part by CD73. According to the pharmaceutical composition of claim 28, the tumor is selected from prostate cancer, colon cancer, rectal cancer, pancreatic cancer, gastric cancer, endometrial cancer, cervical cancer, brain cancer, liver cancer, bladder cancer, ovarian cancer, testicular cancer, head cancer, neck cancer, skin cancer, mesothelial lining cancer, white blood cell cancer, esophageal cancer, breast cancer, muscle cancer, connective tissue cancer, small cell lung cancer, adrenal cancer, thyroid cancer, kidney cancer, bone cancer, brain tumor, glioblastoma, mesothelioma, renal cell carcinoma, sarcoma, choriocarcinoma, Epidermal basal cell carcinoma and testicular seminoma; the immune-related diseases and disorders are selected from rheumatoid arthritis, renal failure, lupus erythematosus, asthma, psoriasis, ulcerative colitis, pancreatitis, allergies, fibrosis, anemia, fibromyalgia, Alzheimer's disease, congestive heart failure, stroke, aortic valve stenosis, arteriosclerosis, osteoporosis, Parkinson's disease, infection, Crohn's disease, ulcerative colitis, allergic contact dermatitis and eczema, systemic sclerosis and multiple sclerosis.