Crystal form of GLP-1r agonist, preparation method therefor and use thereof

Abstract

Disclosed in the present application are preparation methods for and uses of a crystal form 1, crystal form 2, amorphous form 1, and amorphous form 2 of a compound represented by formula (I). Specifically, the compound is used for preventing or treating GLP-1-mediated related diseases.

Description

Crystal forms, preparation methods, and applications of GLP-1R agonists

[0001] Citation of relevant applications

[0002] This disclosure claims the full benefits of Chinese Patent Application No. 202510088613.3, filed on January 20, 2025 with the State Intellectual Property Office of the People's Republic of China, entitled "Crystal Form of GLP-1R Agonist and Its Preparation Method and Use", the entire contents of which are incorporated herein by reference.

[0003] field

[0004] This disclosure generally relates to the field of drug crystal technology, and specifically to new crystal forms, their preparation methods, and uses.

[0005] background

[0006] Glucagon-like peptide-1 (GLP-1) is a peptide hormone secreted by enteroendocrine cells in the gut in response to diet. GLP-1 is thought to play a role in postprandial blood glucose regulation by directly increasing diet-induced insulin secretion from pancreatic β-cells and by promoting satiety through delaying food transport through the intestines. GLP-1 mediates intracellular signaling via the GLP-1 receptor (GLP-1R), a member of the G protein-coupled receptor family located on the cell membrane, which, upon activation, leads to the accumulation of the second messenger cyclic adenosine monophosphate (cAMP). Nonalcoholic steatohepatitis (NASH) is associated with features of metabolic syndrome, including obesity, type 2 diabetes, insulin resistance, and cardiovascular disease.

[0007] Compound (I) is a GLP-1R agonist compound that has been extensively studied for its association with diseases related to diabetes, obesity, and NASH. Its chemical name is 3-((1S,2S)-1-(2-((S)-3-(3-(1-cyclopropyl-4-fluoro-1H-indazol-5-yl)-2-oxo-2,3-dihydro-1H-imidazol-1-yl)-2-(4-fluoro-3,5-dimethylphenyl)-4-methyl-4,5,6,7-tetrahydro-2H-pyrazolo[4,3-c]pyridin-5-carbonyl)-5-(tetrahydro-2H-pyran-4-yl)-1H-indole-1-yl)-2-methylcyclopropyl)-1,2,4-oxadiazol-5(4H)-one, and its structural formula is shown in Formula (I).

[0008] Currently, there is still a gap in the field of preparing crystal forms for compound (I). Therefore, there is a need to develop crystal forms with higher performance, especially higher drug-like properties.

[0009] Overview

[0010] Firstly, a crystal form 1 having the structure of formula (I) is provided.

[0011] The X-ray powder diffraction pattern, expressed in terms of diffraction angle 2θ, has at least three characteristic diffraction peaks selected from 4.7°±0.2°, 10.8°±0.2°, 13.4°±0.2°, and 14.8°±0.2°.

[0012] In some embodiments, crystal form 1 further has at least one characteristic diffraction peak selected from 15.9°±0.2°, 17.4°±0.2°, 19.3°±0.2° and 20.2°±0.2° in the X-ray powder diffraction pattern expressed in diffraction angle 2θ.

[0013] In some embodiments, the X-ray powder diffraction pattern, expressed in terms of diffraction angle 2θ, further includes at least one characteristic diffraction peak selected from 13.6°±0.2°, 21.3°±0.2°, 21.7°±0.2°, and 23.6°±0.2°.

[0014] In some embodiments, the X-ray powder diffraction pattern of the crystal form 1, expressed in terms of diffraction angle 2θ, is shown in Figure 1.

[0015] In some embodiments, crystal form 1 is a monoclinic crystal system and has space group P21.

[0016] In some embodiments, crystal form 1 has the following unit cell parameters: α=90°, β=98°, γ=90°.

[0017] In some embodiments, the crystal form 1 has a DSC pattern as shown in FIG2.

[0018] In some embodiments, the crystal form 1 has single crystal parameters as shown in Table 2.

[0019] In some embodiments, the crystal form 1 has the single crystal parameters shown in FIG5.

[0020] Secondly, a method for preparing crystal form 1 is provided, comprising:

[0021] The compound of formula (I) was dissolved in an organic solvent and stirred for 6 to 12 hours. A solid was precipitated and separated to obtain crystal form 1.

[0022] In some embodiments, the organic solvent is selected from any one or a mixture of ethyl acetate, acetonitrile, tetrahydrofuran, acetone, or DMF.

[0023] In some embodiments, the stirring is carried out at a temperature of 10–25°C.

[0024] Thirdly, crystal form 2 with structure (I) is provided.

[0025] The X-ray powder diffraction pattern, expressed in terms of diffraction angle 2θ, has at least three characteristic diffraction peaks selected from 7.0°±0.2°, 14.0°±0.2°, 16.3°±0.2°, and 17.9°±0.2°.

[0026] In some embodiments, crystal form 2 further has at least one characteristic diffraction peak selected from 11.0°±0.2°, 14.7°±0.2°, 15.7°±0.2° and 17.0°±0.2° in the X-ray powder diffraction pattern expressed in diffraction angle 2θ.

[0027] In some embodiments, crystal form 2 further has at least one characteristic diffraction peak selected from 9.9°±0.2°, 13.3°±0.2°, 18.6°±0.2° and 19.1°±0.2° in the X-ray powder diffraction pattern expressed in diffraction angle 2θ.

[0028] In some embodiments, the X-ray powder diffraction pattern of the crystal form 2, expressed in terms of diffraction angle 2θ, is shown in Figure 3.

[0029] In some embodiments, the crystal form 2 has a DSC pattern as shown in FIG4.

[0030] Water vapor in the environment has a significant impact on the permeability of drugs and the stability of formulations. When the relative humidity in the air increases to a sufficiently high level, the hygroscopicity of the crystal form, or even the drug based on that crystal form, does not increase dramatically; in some embodiments, it increases virtually nothing.

[0031] Fourthly, a method for preparing crystal form 2 is provided, which includes:

[0032] The compound of formula (I) was dissolved in an organic solvent and stirred for 6 to 12 hours. A solid was precipitated and separated to obtain crystal form 2.

[0033] In some embodiments, the organic solvent is ethanol.

[0034] In some embodiments, the stirring is carried out at a temperature of 10–25°C;

[0035] Fifthly, an amorphous 1 as shown in formula (I) is provided.

[0036] In the X-ray powder diffraction pattern expressed as diffraction angle 2θ, there are diffuse patches at 8° to 30°; more preferably, there are diffuse patches at 8° to 25°.

[0037] In some embodiments, the X-ray powder diffraction pattern of the amorphous 1 is shown in Figure 6.

[0038] In some embodiments, the DSC spectrum of the amorphous 1 is shown in Figure 7.

[0039] In some implementations, the TGA pattern of the amorphous 1 is shown in Figure 8.

[0040] In some embodiments, the specific crystal form and amorphous form of the compound (I) having the structure of formula (I) disclosed herein have at least one of the following excellent properties: for example, high stability, good crystallinity, low hygroscopicity, uniform particle size distribution, good flowability, good formulation processability, good dissolution, high solubility and bioavailability, meeting pharmaceutical requirements, stable storage, and simple preparation method.

[0041] Sixthly, an amorphous 2 having a sodium salt of a compound of formula (I) (as shown in formula (II)) is provided.

[0042] In some embodiments, the X-ray powder diffraction pattern of the amorphous 2 is shown in Figure 10.

[0043] In some embodiments, the DSC spectrum of the amorphous 2 is shown in Figure 11.

[0044] In some implementations, the TGA pattern of the amorphous 2 is shown in Figure 12.

[0045] In a seventh aspect, a pharmaceutical composition is provided comprising the crystal form 1 or crystal form 2 or amorphous form 1 or amorphous form 2 as described in this disclosure and optionally a pharmaceutically acceptable carrier.

[0046] Pharmaceutically acceptable carriers in this disclosure include diluents or excipients or other additives, examples of which include, but are not limited to, wetting agents, disintegrants, lubricants, binders, surfactants, etc. Examples of other additives include, but are not limited to, shellac, gum arabic, talc, titanium dioxide, sugars (e.g., sucrose), gelatin, water, polysaccharides such as lactose or glucose, paraffin (e.g., petroleum fractions), vegetable oils (e.g., peanut oil or sesame oil), and pharmaceutically acceptable organic solvents such as alcohols (e.g., ethanol or glycerol), natural mineral powders (e.g., kaolin, clay, talc, and chalk), synthetic mineral powders (e.g., highly dispersed silica and silicates), emulsifiers (e.g., lignin, sulfite solutions, methylcellulose, starch, and polyvinylpyrrolidone), magnesium stearate, stearic acid, sodium lauryl sulfate, etc.

[0047] The pharmaceutical compositions disclosed herein can be prepared in various dosage forms, including but not limited to, pharmaceutical formulations suitable for oral administration, such as solid oral formulations including tablets, coated tablets, capsules, granules, powders, pills, etc., or liquid oral formulations including solutions, syrups, suspensions, emulsions, etc.; pharmaceutical formulations suitable for parenteral administration, such as intravenous infusion formulations, intramuscular or subcutaneous injection formulations, rectal suppositories, nasal inhalation formulations, or transdermal patches for topical administration. Additional excipients may also be included in the formulation manufacturing process, preferably those that do not cause a change in crystal form.

[0048] Eighthly, the use of the crystalline or amorphous forms described in this disclosure in the preparation of medicaments for the prevention or treatment of diseases in subjects in need.

[0049] In some embodiments, the use of crystal form 1 or crystal form 2 or amorphous form 1 or amorphous form 2 of this disclosure in the preparation of medicaments for the prevention or treatment of GLP-1-mediated diseases.

[0050] Ninth aspect, providing a method for preventing or treating a disease, comprising administering to an individual who requires the method a therapeutically effective amount or a preventively effective amount of a pharmaceutical composition of crystal form 1 or crystal form 2 or amorphous form 1 or amorphous form 2 or containing a therapeutically effective amount or a preventively effective amount of crystal form 1 or crystal form 2 or amorphous form 1 or amorphous form 2.

[0051] In some embodiments, the pharmaceutical composition comprises a preventive and / or therapeutically effective amount of the crystal form 1.

[0052] In some implementations, the GLP-1-mediated diseases or symptoms are selected from: type 1 diabetes, type 2 diabetes, prediabetes, weight management, overweight, hyperglycemia, insulin resistance, impaired glucose tolerance, diabetic neuropathy, nephropathy, diabetic retinopathy, adipocyte dysfunction, visceral fat deposition, sleep apnea, obesity, eating disorders, excessive sugar consumption, dyslipidemia, hyperinsulinemia, non-alcoholic fatty liver disease, fibrosis, cirrhosis, hepatocellular carcinoma, cardiovascular disease, hypertension, endothelial dysfunction, arthritis, osteoporosis, Parkinson's disease, metabolic syndrome, hyperuricemia, gout, erectile dysfunction, skin and connective tissue diseases, psoriasis, foot ulcers, ulcerative colitis, Alzheimer's disease, and schizophrenia.

[0053] In some implementations, the GLP-1-mediated diseases or symptoms are selected from: long-term weight management, chronic kidney disease, non-alcoholic steatohepatitis, malnutrition-related diabetes, gestational diabetes, hepatic insulin resistance, diabetic nephropathy, atherosclerosis, coronary artery disease, peripheral vascular disease, hypertension, endothelial dysfunction, impaired vascular compliance, congestive heart failure, myocardial infarction, stroke, hemorrhagic stroke, ischemic stroke, traumatic brain injury, pulmonary hypertension, and restenosis after angioplasty. Intermittent claudication, postprandial lipemia, metabolic acidosis, ketosis, left ventricular hypertrophy, peripheral artery disease, macular degeneration, cataracts, glomerulosclerosis, chronic renal failure, syndrome X, premenstrual syndrome, angina pectoris, thrombosis, atherosclerosis, transient ischemic attack, restenosis, impaired glucose metabolism, impaired fasting glucose, hyperapolipoprotein B lipoproteinemia, cognitive impairment, inflammatory bowel disease, short bowel syndrome, Crohn's disease, colitis, irritable bowel syndrome, polycystic ovary syndrome, and substance addiction.

[0054] In a tenth aspect, this disclosure relates to the combined use of the crystal form 1, crystal form 2, amorphous form 1, amorphous form 2 and their pharmaceutical compositions described herein with other pharmaceuticals.

[0055] In some embodiments, the crystal form of this disclosure has superior properties that make it more suitable for the preparation of pharmaceuticals. For example, high solubility, high purity and stability, and high bioavailability.

[0056] In some embodiments, the crystal form of this disclosure is a solid block or powder crystal with a small specific surface area, which is beneficial for improving dissolution, facilitating rapid drug absorption, and having lower filtration resistance and better solid flowability in the formulation.

[0057] In some embodiments, the crystal form disclosed herein undergoes simple post-processing and has a fixed and relatively high melting point. The crystal form exhibits only 0.1% or less mass change within a relative humidity range of 0-80%, demonstrating extremely low hygroscopicity and better preservation of the active pharmaceutical ingredient. Formulations containing this crystal form exhibit superior quality, safety, and stability during manufacturing and storage processes; problems such as uneven active ingredient content and increased impurities are avoided, thereby eliminating the need for specialized and expensive packaging. Attached Figure Description

[0058] Figure 1 is an XRPD image of the free crystal form 1 prepared according to Example 1.

[0059] Figure 2 shows the DSC spectrum of the free crystal form 1 prepared according to Example 1.

[0060] Figure 3 is an XRPD image of the free crystal form 2 prepared according to Example 2.

[0061] Figure 4 shows the DSC spectrum of the free crystal form 2 prepared according to Example 2.

[0062] Figure 5 is a single-crystal unit cell structure diagram of the free state crystal form 1 prepared according to Example 1.

[0063] Figure 6 is an XRPD image of the free amorphous 1 prepared according to Example 4.

[0064] Figure 7 shows the DSC spectrum of the free amorphous 1 prepared according to Example 4.

[0065] Figure 8 shows the TGA spectrum of the free amorphous 1 prepared according to Example 4.

[0066] Figure 9 shows the stability test results of the amorphous 1 prepared according to Example 4.

[0067] Figure 10 shows the XRPD diagram of amorphous compound 2 of formula (II).

[0068] Figure 11 shows the DSC spectrum of amorphous compound 2 of formula (II).

[0069] Figure 12 shows the TGA spectrum of amorphous compound 2 of formula (II). Detailed Implementation

[0070] Various exemplary embodiments of this disclosure are now described in detail. This detailed description should not be considered as a limitation of this disclosure, but rather as a more detailed description of certain aspects, features, and implementations of this disclosure.

[0071] It should be understood that the terminology used in this disclosure is for describing particular embodiments only and is not intended to limit the disclosure. Furthermore, for numerical ranges in this disclosure, it should be understood that an upper and lower limit of the range and each intermediate value between them are specifically disclosed. Every smaller range between any stated value or intermediate value within a stated range, and any other stated value or intermediate value within said range, is also included in this disclosure. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0072] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art described herein. While this disclosure describes only preferred methods and materials, any methods and materials similar or equivalent to those described herein may be used in implementation or testing of this disclosure. All references to literature in this specification are incorporated by reference in their entirety to disclose and describe the methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail. Unless otherwise stated, “%” means percentage based on weight.

[0073] In this paper, the starting material compound (I) was prepared by the method described in US18 / 884965 or PCT / IB2024 / 058942.

[0074] The term "individual" as used herein includes mammals. A mammal can be any mammal, such as a human, primate, bird, mouse, rat, poultry, dog, cat, cow, horse, goat, camel, sheep, or pig. Humans are preferred as mammals.

[0075] The term "room temperature" as used in this article generally refers to 4-30℃, and preferably 20±5℃.

[0076] In this document, the term "crystal form" refers to a specific lattice configuration of a crystalline substance. It is known in the art that crystal form is related to stability, dissolution, and mechanical properties in pharmaceuticals. Different crystal forms of the same substance typically possess different lattices (e.g., unit cells) with distinct physical properties. Different crystal forms can be characterized by methods known in the art. For example, they can be identified using solid-state characterization methods such as X-ray powder diffraction (XRPD). Other characterization methods include differential scanning calorimetry (DSC), pyrolysis gravimetric analysis (TGA), dynamic vapor adsorption (DVS), solid-state NMR, etc. Crystal forms can be characterized using any of the above methods, or a combination of two or more methods.

[0077] Unless otherwise specified herein, all other experimental methods are well-known in the field and can be found in reference to, for example, the Pharmacopoeia of the People's Republic of China. The parameter settings for the detection are as follows:

[0078] The specific conditions for XRPD (X-ray powder diffraction) measurement are: Bruker D8, Cu-Kα radiation, detection range 3°-40°2θ, step size 0.02°2θ, scan rate 0.2s.step-1, current and voltage 40mA, 40KV.

[0079] The TGA (Thermogravimetric Analysis) test conditions are as follows: 1-15 mg of sample is placed in a platinum crucible, and segmented high-resolution detection is performed at 10.00℃ / min from room temperature to 350℃; for the TGA graph, the temperature points and weight loss values ​​are retained.

[0080] The test conditions for differential calorimetry analysis are as follows: 1-10 mg of sample is placed in a perforated platinum crucible, equilibrated at 0 °C, and then heated to 300 °C at a rate of 10 °C / min; for the DSC chart, the temperature points and enthalpy values ​​are retained.

[0081] The dynamic moisture adsorption analysis method is as follows: 1-10 mg of sample is placed in a platinum crucible, and the weight change is detected during the process of relative humidity changing from 0% to 80% to 0%.

[0082] Polarized light microscopy (PLM) images were acquired using an XP-500E polarized light microscope. A small amount of powder sample was placed on a glass slide, a small amount of mineral oil was added to disperse the sample, a coverslip was placed on the stage, and the sample was observed and photographed.

[0083] The test conditions for Fourier transform infrared spectroscopy were: Bruker tensor 27, ATR method, and acquisition range of 600 cm⁻¹. -1 -4000cm -1 4cm resolution -1 .

[0084] In this article, the terms “performance” or “crystal properties” or “property” or “pharmaceutical properties” include its physical and chemical properties, and evaluation indicators include, but are not limited to, melting point, solubility, dissolution rate, mechanical properties, stability, pharmacokinetic or pharmacodynamic properties, hygroscopicity, flowability, compressibility, and competitiveness.

[0085] In this article, the term "effective amount" refers to an amount of compound or combination of compounds that can alleviate, reduce or eliminate one or more symptoms of a particular disease or condition, or prevent, alter or delay the occurrence of one or more symptoms of a particular disease or condition.

[0086] In this document, the term "effective amount" refers to the amount of an active agent that elicits the biological or medical response sought by the investigator, veterinarian, physician, dietitian, or other clinician, caregiver, or subject (including one or more of the following):

[0087] (1) Disease prevention: For example, preventing subjects who may be susceptible to a disease but have not yet shown the pathology or symptoms of the disease from developing the disease;

[0088] (2) Suppressing disease: For example, suppressing the development of a disease in an individual who is experiencing or exhibiting the pathology or symptoms of a disease (i.e., preventing the further development of the pathology and / or symptoms).

[0089] (3) Improve symptoms: For example, improve the condition of an individual who is experiencing or exhibiting the pathology or symptoms of a certain condition (i.e., reverse the pathology and / or symptoms).

[0090] In this article, the terms "prevention" or "avoidance" refer to preventing the occurrence or appearance of one or more symptoms associated with a specific disease, and do not necessarily mean completely preventing the disease from developing. For example, the term "prevention" refers to treating an individual for preventative purposes at least one symptom that may eventually manifest as a disease or condition but has not yet manifested. Such individuals can be identified by known risk factors associated with the subsequent development of the disease. Alternatively, preventative treatment can be carried out as a preventative measure without prior identification of risk factors. Delaying the onset of at least one symptom can also be considered prevention.

[0091] In this article, the term "treatment" refers to the treatment of an individual who has already exhibited at least one symptom of a condition, or who has previously exhibited at least one symptom of a condition, or who is identified as being at risk of developing the condition. For example, "treatment" can include relieving, reducing, or improving symptoms of a condition; preventing additional symptoms; improving or preventing underlying metabolic causes of symptoms; suppressing the condition (e.g., preventing its progression); alleviating symptoms; promoting the remission of symptoms; alleviating a secondary symptom caused by a primary condition; or preventing and / or therapeutically stopping symptoms of a condition. For example, "treatment" when referring to a condition includes reducing the severity of one or more symptoms associated with a specific condition. Therefore, treating a condition does not necessarily mean reducing the severity of all symptoms associated with the condition, nor does it necessarily mean completely reducing the severity of one or more symptoms associated with the condition. Studies have shown that even a moderate reduction in weight or related parameters (such as BMI, waist circumference, and body fat percentage) can lead to health improvements, such as lowering blood pressure, improving lipid profiles, or reducing sleep apnea.

[0092] Metabolic diseases refer to a class of diseases caused by abnormal metabolism of substances or energy, resulting in metabolic disorders. They are usually diseases that affect multiple systems throughout the body. In this article, metabolic diseases specifically refer to diseases related to abnormal lipid metabolism, preferably including obesity, overweight, and weight-related conditions.

[0093] BMI (Body Mass Index) is an internationally recognized standard for measuring a person's fatness and health status. It is calculated by dividing weight (in kilograms) by the square of height (in meters), using the formula: BMI = weight (kg) / height. 2 (m 2 ).

[0094] "Obesity" refers to an individual's BMI ≥ 30 kg / m². 2 .

[0095] "Overweight" refers to an individual's weight exceeding the healthy weight range, with a BMI of 25 kg / m². 2 ≤BMI<30kg / m 2 The preferred value is 27 kg / m 2 ≤BMI<30kg / m2 .

[0096] "Weight management" refers to the behaviors, techniques, and physiological processes that help an individual achieve and maintain a healthy weight. A healthy weight for a particular patient can be determined in consultation with a healthcare professional; in one implementation, "weight management" refers to weight loss and / or changes in body fat composition.

[0097] Long-term weight management therapy helps patients achieve their healthy weight goals. In one embodiment, "long-term weight management" means that the subject achieves their healthy weight goal and maintains their weight within that target range for a certain period of time. In another embodiment, "long-term weight management" means that the subject achieves their healthy weight goal and maintains their weight within that target range for at least 13 weeks. In yet another embodiment, the subject achieves their healthy weight goal and maintains their weight within that target range for at least 26 weeks. In yet another embodiment, "long-term weight management" means that the subject achieves their healthy weight goal and maintains their weight within that target range for at least 52 weeks. In yet another embodiment, "long-term weight management" means that the subject achieves their healthy weight goal and maintains their weight within that target range for at least 104 weeks.

[0098] Example 1

[0099] Take about 30 mg of compound (I) sample, dissolve and clarify it with 1.5 mL of LDMF at 0-15℃, keep it at 15-25℃ and stir for 2-6 h; filter, rinse the filter cake with 0.3 mL of LDMF, collect the filter cake to obtain purified wet product 1, transfer the purified wet product to a forced-air drying oven to dry, control the temperature at 40-50℃, dry to constant weight, and obtain crystal form 1 sample.

[0100] The sample from Example 1 was characterized as follows.

[0101] 1. XRPD spectral analysis

[0102] For specific results, please refer to Figure 1 and Table 1, where Figure 1 is the XRPD pattern of crystal form 1.

[0103] Table 1:

[0104] 2. DSC spectrum analysis

[0105] The differential scanning calorimetry (DSC) analysis results are shown in Figure 2. As can be seen from Figure 2, the extrapolated onset temperature of crystal form 1 is 266℃, and the peak temperature is 268℃.

[0106] 3. Single-crystal unit cell structure analysis

[0107] The results of the single-crystal cell structure analysis are shown in Figure 5. The results show that the crystal form 1 disclosed herein is a monoclinic crystal system with space group P21 and has the cell parameters shown in Table 2 below.

[0108] Table 2 - Cell Parameters

[0109] Example 2

[0110] Take about 30 mg of compound (I) sample, dissolve and clarify it in 1.5 mL of ethanol at 0-15℃, keep it at 15-25℃ and stir for 2-6 h; filter, rinse the filter cake with 0.3 mL of acetonitrile, collect the filter cake to obtain purified wet product 1, transfer the purified wet product to a forced-air drying oven to dry, control the temperature at 40-50℃, dry to constant weight, and obtain crystal form 2 sample.

[0111] The sample from Example 2 was characterized as follows.

[0112] 1. XRPD spectral analysis

[0113] For specific results, please refer to Figure 3 and Table 3, where Figure 3 is the XRPD pattern of crystal form 2.

[0114] Table 3

[0115] 2. DSC spectrum analysis

[0116] The differential scanning calorimetry (DSC) analysis results are shown in Figure 4. As can be seen from Figure 4, the extrapolated onset temperature for crystal form 2 is 277℃, and the peak temperature is 280℃.

[0117] Example 3

[0118] Solvent screening experiment: Referring to Experiment 1 and Experiment 2, the solvent was replaced, and the following screening results were obtained:

[0119] Example 4

[0120] 30 mg of the compound was dissolved and clarified in 3 mL of dichloromethane, and concentrated at 40-50°C until no obvious droplets were observed in the condenser. It was then dissolved and clarified in 7.5 mL of acetonitrile:water (4:1), and freeze-dried to obtain amorphous 1.

[0121] The sample from Example 4 was characterized as follows.

[0122] 1. XRPD spectral analysis

[0123] See Figure 6 for the specific results, where Figure 6 is the XRPD pattern of amorphous 1.

[0124] 2. DSC spectrum analysis

[0125] See Figure 7 for the specific results, where Figure 7 is the DSC spectrum of amorphous 1.

[0126] 2. TGA Spectrum Analysis

[0127] See Figure 8 for specific results, where Figure 8 is the TGA spectrum of amorphous 1.

[0128] Example 5

[0129] solubility of amorphous 1

[0130] The amorphous 1 prepared in Example 4 was added to the above solution or water and vortexed for 15 min at room temperature to mix. The equilibrium solubility of amorphous 1 in hydrochloric acid solutions at pH 1.0, pH 1.2 and pH 2.0, acetate buffer solutions at pH 3.0, pH 4.0 and pH 4.5, phosphate buffer solutions at pH 5.0, phosphate buffer solutions at pH 6.8 and pH 7.4, water, and anhydrous sodium carbonate solution was determined. The results are shown in Table 4.

[0131] Table 4. Equilibrium solubility of amorphous 1 in media with different pH values.

[0132] Example 6

[0133] This example is a stability experiment of the amorphous material 1 prepared in Example 4.

[0134] The sample was placed in an open environment at 60℃ and XRPD was tested on day 0, day 10 and day 30. The results showed that the characteristic peaks of amorphous 1 did not change.

[0135] Under the conditions specified in the packaging of this product (inner packaging: double-layer polyethylene bag + desiccant, outer packaging: aluminum foil bag), XRPD was tested at 25℃-60% in an environment, on day 0 and at month 6. The results showed that the characteristic peak of amorphous 1 did not change.

[0136] The test results are shown in Figure 9.

[0137] Example 7: In vitro activity evaluation

[0138] (1).hGLP-1RcAMPAssay

[0139] 1. Cell line and reagent preparation

[0140] 1) Cell line: Flp-In-293-hGLP1R

[0141] 2) Culture medium: DMEM + 10% FBS + 1X Penicillin-Streptomycin + 200μg / mL HB

[0142] 3) Test buffer: 1X HBSS + 20mM HEPES + 0.1% BSA + 500μM IBMX

[0143] 2. Agonist Testing

[0144] a) Flpin-293-GLP1R cells were seeded in a 384-well assay plate (6007680-50, PE) using complete culture medium, with 2,000 cells per well.

[0145] b) Prepare the 4X complex working solution using the detection buffer.

[0146] c) Add 5 μL of 4X compound working solution to the cell plate and incubate at 37°C for 30 minutes.

[0147] d) Dilute the Eu-cAMP tracer (1 / 50) with lysis buffer, and then add 10 μl / well to the detection plate.

[0148] e) Dilute Ulight-anti-cAMP (1 / 150) with lysis buffer, then add 10 μl / well to the detection plate.

[0149] f) Incubate at a constant temperature for 1 hour.

[0150] g) Read the tablet at wavelengths of 665 nm and 615 nm on the Envision2105 tablet reader.

[0151] 3. Data Analysis

[0152] The formula for calculating 3.1% Activity is as follows:

[0153] %Activity=100-(Signal) cmpd -Signal Ave_PC ) / (Signal Ave_VC -Signal Ave_PC )×100.

[0154] 3.2 Calculate EC 50 And plot the effect-dose curve of cmpds:

[0155] Y=Bottom+(Top-Bottom) / (1+10^((LogEC 50 -X)*HillSlope)).

[0156] X: Logarithm of agonist concentration;

[0157] Y: Percentage of activity.

[0158] Table 5 shows the bioactivity of amorphous 1 in the hGLP-1R agonist cAMP stimulation assay as follows.

[0159] Table 5: Bioactivity of amorphous 1 in hGLP-1R agonist cAMP stimulation assay (EC50)

[0160] Example 8: Pharmacokinetic Study

[0161] 1. PK study of single subcutaneous administration in SD rats

[0162] PK studies of amorphous 1 or Orforglipron administered via single subcutaneous injection: Animals: Male Sprague-Dawley (SD) rats; Solvent formulation: 80% MCT + 20% benzyl alcohol; Dosage: 30 mg / kg; Concentration: 30 mg / mL; Volume: 1 mL / kg; Administration site: Subcutaneous injection on the back of the skin. Blood collection time points: Before administration, 1 h, 2 h, 8 h, 24 h, 48 h, 72 h, 96 h, 120 h, 144 h, 216 h, 312 h, 480 h, and 696 h. Sample collection: Blood samples were collected, and after protein precipitation, they were analyzed by liquid chromatography-mass spectrometry (LC / MS / MS). Pharmacokinetic parameters were calculated using PhoenixWinNonlin 8.0 software, and the data are shown in Table 6.

[0163] Table 6. Drug exposure of amorphous 1 after a single subcutaneous injection Note: AUC 0-696h It is an important parameter in pharmacokinetics, referring to the area under the curve from 0 to the end of the study (696 hours), representing the amount of drug exposed in the body during that time period.

[0164] The structure of Orforglipron is shown above.

[0165] 2. Single-dose oral administration pharmacokinetic study in Sprague-Dawley (SD) rats

[0166] Amorphous 1 and orforglipron single-dose oral pharmacokinetic study protocol: Animals: male Sprague-Dawley (SD) rats; Solutol:PEG400:Tween80:Saline = 10:40:2:48 (v / v / v / v); Dosage: 5 mg / kg; Dosage concentration: 0.5 mg / mL; Dosage volume: 10 mL / kg; Blood sample collection time points: before administration, 0.25 h, 0.5 h, 1 h, 2 h, 3 h, 4 h, 8 h, 12 h, 18 h, 24 h, 32 h, 40 h, 48 h; Sample collection: Approximately 0.15 mL of blood was collected from the jugular sinus at each time point, and the collected whole blood was placed in EDTA-K2 anticoagulant tubes. Blood samples were collected, and after protein precipitation, they were analyzed by liquid chromatography-mass spectrometry (LC / MS / MS). Pharmacokinetic parameters were calculated using Phoenix WinNonlin 8.0 software, and the data are shown in Table 7.

[0167] Table 7: Pharmacokinetic parameters of different compounds after a single oral administration Note: C max This refers to the highest blood drug concentration in plasma after administration.

[0168] Example 9: In vivo pharmacodynamic study

[0169] Animal selection: Within one week prior to administration of the test substance, 28 male cynomolgus monkeys were anesthetized with salbutamol (5.0 mg / kg, intramuscular injection, IM) and then intravenously injected with 0.5 g / kg (1 mL / kg) of 50% glucose within 30 seconds. Blood samples were collected at 10 time points: 45 min before glucose injection (without anesthesia), 6 min before glucose injection (under anesthesia), and 1 min, 5 min, 10 min, 15 min, 20 min, 30 min, 40 min, and 60 min after glucose injection. At each time point, 10 μL of whole blood was collected from the saphenous vein for blood glucose testing. Approximately 1.5 mL of whole blood was collected from each animal at each time point and placed in a vacuum blood collection tube containing a coagulant. The tube was immediately and gently inverted several times to mix thoroughly, placed at room temperature, and then centrifuged at 4°C (2000 g, 10 min) to obtain serum. The collected serum samples were stored in an ultra-low temperature freezer until all samples were tested for blood glucose, insulin, and C-peptide. For screening cynomolgus monkeys, 15 monkeys were selected for formal experiments based on the trends in blood glucose, insulin, and C-peptide changes.

[0170] Formal Experiment:

[0171] Grouping: The 15 selected cynomolgus monkeys were randomly divided into 3 groups of 5 each.

[0172] Sample preparation: Weigh an appropriate amount of the compound disclosed herein and dissolve it in the solvent: 5% DMSO / 10% Cremophor EL / 20% PEG400 / 65% 100mM Glycine-NaOH pH10 to prepare a suitable concentration for intravenous bolus injection.

[0173] Administration method: 5 minutes before glucose injection, administer the test sample via intravenous bolus injection: Draw the corresponding volume of test sample for each animal into the syringe and administer the drug via bolus injection, which should be completed within 30 seconds.

[0174] Intravenous glucose tolerance test: The intravenous glucose tolerance test begins 5 minutes after the test sample is administered via intravenous bolus. The animals are kept under anesthesia during the test (using acetaminophen (5.0 mg / kg, IM) for anesthesia). 5 minutes after the bolus administration of the test sample, 0.5 g / kg (1.0 mL / kg) of 50% glucose is injected intravenously and the test is completed within 30 seconds.

[0175] Sample collection: Blood was collected 45 minutes before glucose injection (without anesthesia) and 6 minutes before glucose injection (under anesthesia). Blood collection points after glucose injection (under anesthesia): 1 minute, 5 minutes, 10 minutes, 15 minutes, 20 minutes, 30 minutes, 40 minutes, and 60 minutes. At each time point, 10 μL of whole blood was collected from the saphenous vein for blood glucose testing. Approximately 1.5 mL of whole blood was collected from the saphenous vein of all animals at each time point and placed in a vacuum blood collection tube containing a coagulant. The tube was immediately and gently inverted several times to mix thoroughly, then placed at room temperature and centrifuged at 4°C (2000g, 10 minutes) to obtain serum. The collected serum samples were stored in an ultra-low temperature freezer until all samples were analyzed.

[0176] Sample analysis: Detection of insulin, C-peptide, and blood glucose in serum samples.

[0177] Experimental results: The amorphous 1 disclosed herein can significantly increase the secretion of insulin and C-peptide in cynomolgus monkeys, while reducing blood glucose concentration.

[0178] Example 10: Dissolution Test

[0179] The prescribed amounts of the amorphous 2 (i.e., the amorphous 2 of the sodium salt of compound (I)), copovidone VA64, and poloxamer 407 of this disclosure are dissolved in ethanol, dried by rotary evaporation at 40-60°C, and the resulting dried product is then mechanically pulverized, sieved through a 140-mesh sieve, and dried at 60°C to obtain solid dispersed particles. Microcrystalline cellulose, mannitol, croscarmellose sodium, and magnesium stearate excipients are further added to prepare tablets.

[0180] Dissolution test: The above tablets were dissolved in a phosphate buffer solution containing 0.7% Tween 80 at pH 6.8 using a NanoChromCore 120C 184.6×50mm 3μm column or an equivalent column. The mobile phase was 10mM dipotassium hydrogen phosphate solution-acetonitrile = 40:60. The flow rate was 1.0ml / min; the column temperature was 35℃; and the detection wavelength was 254nm. An appropriate amount of amorphous 2 reference standard was accurately weighed, dissolved in acetonitrile by sonication, and diluted with acetonitrile to prepare a solution with a concentration of approximately 10μg / ml, which served as the reference solution. 5μl of each of the reference solution and the test solution were accurately injected into the liquid chromatograph, and the chromatograms were recorded. The dissolution rate was calculated by peak area using the external standard method. The results showed that the dissolution rate of amorphous 2 tablets in each batch was between 86% and 106% at 60 min, meeting the dissolution requirements for clinical drugs.

[0181] Although this disclosure has been described with reference to exemplary embodiments, it should be understood that this disclosure is not limited to the exemplary embodiments disclosed. Various adjustments or changes may be made to the exemplary embodiments of this disclosure without departing from the scope or spirit of this disclosure. The scope of the claims should be interpreted in the broadest possible sense to cover all modifications and equivalent structures and functions.

Claims

1. Crystal form 1 of the compound shown in formula (I), in, The X-ray powder diffraction pattern, expressed in terms of diffraction angle 2θ, has at least three characteristic diffraction peaks selected from 4.7°±0.2°, 10.8°±0.2°, 13.4°±0.2°, and 14.8°±0.2°.

2. The crystal form 1 as described in claim 1, wherein, In the X-ray powder diffraction pattern expressed in diffraction angle 2θ, it further has at least one characteristic diffraction peak selected from 15.9°±0.2°, 17.4°±0.2°, 19.3°±0.2° and 20.2°±0.2°.

3. Crystal form 1 as described in claim 1 or 2, wherein, In the X-ray powder diffraction pattern expressed in diffraction angle 2θ, it further has at least one characteristic diffraction peak selected from 13.6°±0.2°, 21.3°±0.2°, 21.7°±0.2° and 23.6°±0.2°.

4. Crystal form 1 as described in any one of claims 1-3, wherein, The X-ray powder diffraction pattern of crystal form 1, expressed in terms of diffraction angle 2θ, is shown in Figure 1.

5. The crystal form 1 as described in claim 1, wherein, The crystal form 1 is a monoclinic crystal system and has the space group P21.

6. The crystal form 1 as described in claim 5, wherein, The crystal form 1 has the following unit cell parameters: α=90°, β=98°, γ=90°.

7. The method for preparing crystal form 1 according to any one of claims 1-6, comprising: The compound of formula (I) was dissolved in an organic solvent and stirred for 6 to 12 hours. A solid was precipitated and separated to obtain crystal form 1.

8. The preparation method according to claim 7, wherein, The organic solvent is selected from any one of ethyl acetate, acetonitrile, tetrahydrofuran, acetone, or DMF, or a mixture thereof.

9. The preparation method according to claim 7 or 8, wherein, The stirring is carried out at a temperature of 10–25°C.

10. Crystal form 2 of the compound shown in formula (I), in, The X-ray powder diffraction pattern, expressed in terms of diffraction angle 2θ, has at least three characteristic diffraction peaks selected from 7.0°±0.2°, 14.0°±0.2°, 16.3°±0.2°, and 17.9°±0.2°.

11. The crystal form 2 as described in claim 10, wherein, In the X-ray powder diffraction pattern expressed in diffraction angle 2θ, it further has at least one characteristic diffraction peak selected from 11.0°±0.2°, 14.7°±0.2°, 15.7°±0.2° and 17.0°±0.2°.

12. The crystal form 2 as described in claim 10 or 11, wherein, In the X-ray powder diffraction pattern expressed in diffraction angle 2θ, it further has at least one characteristic diffraction peak selected from 9.9°±0.2°, 13.3°±0.2°, 18.6°±0.2° and 19.1°±0.2°.

13. Crystal form 2 as described in any one of claims 10-12, wherein, The X-ray powder diffraction pattern of crystal form 2, expressed in terms of diffraction angle 2θ, is shown in Figure 3.

14. The method for preparing crystal form 2 according to any one of claims 10-13, in, The preparation method includes: The compound of formula (I) was dissolved in an organic solvent and stirred for 6 to 12 hours. A solid was precipitated and separated to obtain crystal form 2.

15. The preparation method according to claim 14, wherein, The organic solvent is ethanol.

16. The preparation method according to claim 14 or 15, wherein, The stirring is carried out at a temperature of 10–25°C.

17. The amorphous form of the compound shown in formula (I), in, In the X-ray powder diffraction pattern expressed as diffraction angle 2θ, there are diffuse patches at 8° to 30°; more preferably, there are diffuse patches at 8° to 25°.

18. The amorphous 1 as claimed in claim 17, wherein, The X-ray powder diffraction pattern of the amorphous 1, expressed in terms of diffraction angle 2θ, is shown in Figure 6.

19. The amorphous 1 as described in claim 17 or 18, wherein, The amorphous 1 has a DSC pattern as shown in Figure 7.

20. The amorphous 1 as described in any one of claims 17-19, wherein, The amorphous 1 has a TGA pattern as shown in Figure 8.

21. The amorphous form of the compound shown in formula (II), in, The X-ray powder diffraction pattern of the amorphous 2, expressed in terms of diffraction angle 2θ, is shown in Figure 10.

22. A pharmaceutical composition comprising crystal form 1 as described in any one of claims 1-6, crystal form 2 as described in any one of claims 10-13, amorphous form 1 as described in claims 17-20, or amorphous form 2 as described in claim 21, and at least one pharmaceutically acceptable carrier.

23. Use of the crystal form 1 of any one of claims 1-6, or the crystal form 2 of any one of claims 10-13, or the amorphous form 1 of claims 17-20, or the amorphous form 2 of claim 21, or the pharmaceutical composition of claim 22 in the preparation of a medicament for the prevention or treatment of GLP-1-mediated diseases.

24. A method for preventing or treating GLP-1-mediated diseases, comprising administering to an individual in need of the method any therapeutically effective amount or a preventatively effective amount of crystal form 1 as described in any one of claims 1-6, or crystal form 2 as described in any one of claims 10-13, or amorphous form 1 as described in claims 17-20, or amorphous form 2 as described in claim 21, or any combination thereof of the pharmaceutical composition described in claim 22.

25. The use as described in claim 23 or the method as described in claim 24, wherein the GLP-1-mediated disease or symptom is selected from: type 1 diabetes, type 2 diabetes, prediabetes, weight management, overweight, hyperglycemia, insulin resistance, impaired glucose tolerance, diabetic neuropathy, nephropathy, diabetic retinopathy, adipocyte dysfunction, visceral fat deposition, sleep apnea, obesity, eating disorders, excessive sugar consumption, dyslipidemia, hyperinsulinemia, non-alcoholic fatty liver disease, fibrosis, cirrhosis, hepatocellular carcinoma, cardiovascular disease, hypertension, endothelial dysfunction, arthritis, osteoporosis, Parkinson's disease, metabolic syndrome, hyperuricemia, gout, erectile dysfunction, skin and connective tissue diseases, psoriasis, foot ulcers, ulcerative colitis, Alzheimer's disease, and schizophrenia.

26. The use as described in claim 23 or the method as described in claim 24, wherein the GLP-1-mediated disease or symptom is selected from: long-term weight management, chronic kidney disease, non-alcoholic steatohepatitis, malnutrition-related diabetes, gestational diabetes, hepatic insulin resistance, diabetic nephropathy, atherosclerosis, coronary artery disease, peripheral vascular disease, hypertension, endothelial dysfunction, impaired vascular compliance, congestive heart failure, myocardial infarction, stroke, hemorrhagic stroke, ischemic stroke, traumatic brain injury, and pulmonary hypertension. Restenosis after angioplasty, intermittent claudication, postprandial lipemia, metabolic acidosis, ketosis, left ventricular hypertrophy, peripheral artery disease, macular degeneration, cataracts, glomerulosclerosis, chronic renal failure, syndrome X, premenstrual syndrome, angina pectoris, thrombosis, atherosclerosis, transient ischemic attack, restenosis, impaired glucose metabolism, impaired fasting glucose, hyperapolipoprotein B lipoproteinemia, cognitive impairment, inflammatory bowel disease, short bowel syndrome, Crohn's disease, colitis, irritable bowel syndrome, polycystic ovary syndrome, and substance addiction.

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