Salts and crystal forms of GLP-1R agonists and their use

Abstract

Various salt forms of Compound I and Compound II, represented by the following structural formulas, and their corresponding pharmaceutical compositions are disclosed. (I), (II) Specific single crystalline forms of the 1:1 Compound I Tris salt, the 1:1 Compound II Tris salt, and the 1:1 Compound II citrate salt are characterized by different properties and physical measurements. Methods for preparing the specific crystalline forms are also disclosed. The disclosure also provides methods for treating type 2 diabetes mellitus, prediabetes, obesity, nonalcoholic fatty liver disease, nonalcoholic steatohepatitis, and cardiovascular disease in a subject. TIFF2023527997000038.tif34141

Description

Technical Field

[0001] Cross - reference to related applications This application claims the benefit of priority to International Patent Application No. PCT / CN2020 / 092530, filed on May 27, 2020. The entire content of the foregoing application is incorporated herein by reference.

Background Art

[0002] GLP - 1 is a 30 - amino - acid - long incretin hormone secreted from intestinal L cells in response to food intake. GLP - 1 has been shown to stimulate insulin secretion physiologically and in a glucose - dependent manner, decrease glucagon secretion, inhibit gastric emptying, reduce appetite, and stimulate beta - cell proliferation. In pre - clinical experiments, GLP - 1 promotes continuous beta - cell function by stimulating the transcription of genes important for glucose - dependent insulin secretion and promoting beta - cell neogenesis (Meier et al., Biodrugs. 17(2):93 - 102, 2013).

[0003] In healthy individuals, GLP - 1 plays an important role in regulating post - prandial glucose concentrations by stimulating glucose - dependent insulin secretion by the pancreas and increasing peripheral glucose uptake. GLP - 1 also suppresses glucagon secretion and reduces hepatic glucose release. In addition, GLP - 1 delays gastric emptying and slows small - intestine motility, thereby delaying food absorption. In people with T2DM, there is no normal post - prandial rise or a reduced rise of GLP - 1 (Vilsboll et al., Diabetes. 50:609 - 613, 2001).

[0004] Hoist (Physiol. Rev. 87:1409, 2007) and Meier (Nat. Rev. Endocrinol. 8:728, 2012) have described three major pharmacological activities of GLP-1 receptor agonists such as GLP-1, liraglutide, and exendin-4 in improving glycemic control in patients with T2DM by reducing fasting and postprandial glucose (FPG and PPG): (i) increased glucose-dependent insulin secretion (improvement in the first and second phases), (ii) glucagon inhibitory activity under hyperglycemic conditions, and (iii) delayed gastric emptying rate resulting in delayed absorption of dietary glucose.

[0005] International Patent Application No. PCT / CN2019 / 119373 (the entire teachings of which are incorporated herein by reference) discloses very potent GLP-1 agonists. The structures of the two agonists disclosed therein, referred to herein as "Compound I" and "Compound II", are shown below. [Chemical formula]

[0006] The chemical name of Compound I is (S)-2-((4-(3-((4-chloro-2-fluorobenzyl)-oxy)phenyl)-3,6-dihydropyridin-1(2H)-yl)methyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]-imidazole-6-carboxylic acid.

[0007] The chemical name of Compound II is (S)-2-((6-((4-cyano-2-fluorobenzyl)oxy)-3’,6’-dihydro-[2,4’-bipyridin]-1’(2’H)-yl)methyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylic acid.

[0008] To successfully develop a pharmaceutically active agent such as Compound I or Compound II, it is typically necessary to be able to facilitate isolation and purification after synthesis, be able to accommodate large-scale manufacturing, be able to be stored for extended periods with minimal water absorption, decomposition, or conversion to other solid forms, be suitable for formulation, and be readily absorbable (e.g., soluble in water and gastric juice) after administration to a subject.

Summary of the Invention

[0009] It has also been found herein that the 1:1 tris salt of Compound I, the 1:1 tris salt of Compound II, and the 1:1 citrate salt of Compound II can crystallize under defined conditions to provide the desired crystalline forms (see Examples 3-7). These three salts also have good solubility in water and simulated gastric juice, as well as solid stability (see Tables 2 and 7-1), and have a high melting point onset. The 1:1 tris salt of Compound I and the 1:1 tris salt of Compound II are also suitable for large-scale synthesis.

[0010] The designation "1:1" refers to the molar ratio between the acid (citric acid) / base (tris-(hydroxymethyl)aminomethane) and Compound I / Compound II. Due to the three carboxylic acid groups of citric acid and the two basic nitrogen atoms of Compound II, multiple possible stoichiometries are possible. The citrate salt of 1:1 Compound (I) is referred to herein as the "1:1 citrate salt of Compound II".

[0011] In one aspect, the present disclosure provides a tris salt of Compound I, wherein the molar ratio between Compound I and tris-(hydroxymethyl)aminomethane is 1:1. As described above, this salt is also referred to herein as the "1:1 tris salt of Compound I".

[0012] In another aspect, the present disclosure provides a tris salt of Compound II, wherein the molar ratio between Compound II and tris-(hydroxymethyl)aminomethane is 1:1. As described above, this salt is also referred to herein as the "1:1 Compound II tris salt".

[0013] In another aspect, the present disclosure provides a citrate salt of Compound II, wherein the molar ratio between Compound II and citric acid is 1:1. As described above, this salt is also referred to herein as the "1:1 Compound II citrate salt".

[0014] In another aspect, the present disclosure provides a pharmaceutical composition comprising a 1:1 Compound I tris salt (or a 1:1 Compound II tris salt, or a 1:1 Compound II citrate salt) and a pharmaceutically acceptable carrier.

[0015] The present disclosure provides a method for treating cardiovascular and related diseases, comprising administering to a subject in need of such treatment a therapeutically effective amount of a salt disclosed herein or the corresponding pharmaceutical composition.

[0016] The present disclosure also provides the use of a salt of the present disclosure or a pharmaceutical composition thereof in any of the methods of the present disclosure described above. In one embodiment, a salt of the present disclosure or a pharmaceutical composition thereof for use in any of the methods of the present disclosure described herein is provided. In another embodiment, a salt of the present disclosure or a pharmaceutical composition thereof for the manufacture of a medicament for any of the methods of the present disclosure described is provided.

[0017] It should be understood that any embodiment of the present disclosure that includes only what is described in the examples or claims, or only in one section of this specification, can be combined with one or more additional embodiments of the present disclosure, provided that such combination is not explicitly negated or inappropriate.

Brief Description of the Drawings

[0018]

Figure 1

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Figure 5B

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Mode for Carrying Out the Invention

[0019] The present disclosure is directed to tris salts of novel compound I (i.e., 1:1 tris salts), tris salts of novel compound II (i.e., 1:1 tris salts), and citrate salts of novel compound II (i.e., 1:1 citrate salts), and to each of the polymorphic forms thereof.

[0020] "Hydrated form" refers to a solid or crystalline form of compound I or compound II in the free base or salt state, in which water is combined as an essential part of the solid or crystal in a stoichiometric ratio (e.g., a molar ratio of 1:1 or 1:2 of compound (I): water) with the free base compound (I) or the corresponding salt. "Anhydrous form" refers to a form that does not have a stoichiometric ratio between water and the free base of compound (I) or the corresponding salt of compound (I), and in which water is substantially absent in the solid form (e.g., less than 10% by weight as determined by Karl Fischer analysis). The new solid forms disclosed in the present disclosure include hydrated forms and anhydrous forms.

[0021] As used herein, "crystalline" refers to a solid having a crystal structure in which individual molecules have a very uniform regular three-dimensional arrangement.

[0022] The disclosed crystalline compound I salts and / or compound II salts can be crystals in a single crystal form or a mixture of crystals of different single crystalline forms. A single crystal form means that compound I (or compound II) is a single crystal or a plurality of crystals in which each crystal has the same crystal form.

[0023] Regarding the crystalline forms of the compound I / compound II salts disclosed herein, at least a specific weight percentage of the 1:1 compound I / compound II salt is in the single crystal form. Examples of the specific weight percentage include 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.5%, 99.9%, or a weight percentage of 70% - 75%, 75% - 80%, 80% - 85%, 85% - 90%, 90% - 95%, 95% - 100%, 70 - 80%, 80 - 90%, 90 - 100 weight% of the compound I / compound II salt is in the single crystal form. It should be understood that all values and ranges between these values and ranges are encompassed by this disclosure.

[0024] When the crystalline compound I / compound II salt is defined as a specific percentage of one specific crystalline form of the compound I / compound II salt, the remainder is composed of an amorphous form and / or a crystalline form other than the one or more specific forms specified. Examples of the single crystal form include the 1:1 compound I tris salt (form A), the 1:1 compound II tris salt (forms B and G), and the 1:1 compound II citrate (form A), which are characterized by one or more properties as discussed herein.

[0025] Both compound I and compound II have chiral centers. The salts and polymorphic states of compound I and compound II disclosed herein are at least 80%, 90%, 99%, or 99.9 weight% pure compared to the ratio of the weight of the stereoisomer to the weight of all stereoisomers.

[0026] The crystalline compound I / compound II salts disclosed herein exhibit a strong and unique XRPD pattern with sharp peak positions at 2θ and a flat baseline, indicating a highly crystalline substance (e.g., Figure 1).

[0027] Characterization of the 1:1 Compound I Tris Salt Crystalline Form In one embodiment, the 1:1 compound I tris salt is a single crystalline form, Form A, characterized by an X-ray powder diffraction pattern that includes peaks at 2θ of 17.5°, 20.1°, 20.7°, 21.1°, and 22.6° ± 0.2. In another embodiment, Form A is characterized by an X-ray powder diffraction pattern that includes at least three (or four) peaks selected from 17.5°, 20.1°, 20.7°, 21.1°, and 22.6° ± 0.2 at 2θ. In yet another embodiment, Form A is characterized by an X-ray powder diffraction pattern that includes peaks at 2θ of 4.1°, 14.8°, 17.5°, 18.8°, 20.1°, 20.7°, 21.1°, and 22.6° ± 0.2. In yet another embodiment, Form A is characterized by an X-ray powder diffraction pattern that includes peaks at 2θ of 4.1°, 8.1°, 12.8°, 14.8°, 16.3°, 17.5°, 18.8°, 19.3°, 20.1°, 20.7°, 21.1°, 22.6°, 25.1°, and 25.8° ± 0.2. In yet another embodiment, Form A is characterized by an X-ray powder diffraction pattern that is substantially the same as FIG. 1.

[0028] In the field of crystal analysis methods, it is well known that for any given crystal form, the angular peak positions can vary slightly due to factors such as temperature fluctuations, sample movement, and the presence or absence of an internal standard. In the present disclosure, the variability of the angular peak positions is ±0.2 at 2θ. In addition, the relative peak intensities of a given crystal form can vary due to differences in crystallite size and non-random crystallite orientation in sample preparation for XRPD analysis. It is well known in the art to account for these factors without preventing the clear identification of the crystal form.

[0029] In another embodiment, the 1:1 compound I tris salt Form A is characterized by a differential scanning calorimetry (DSC) peak phase transition temperature of 173 ± 3 °C (e.g., 174.3 °C).

[0030] Characterization of the 1:1 Compound II Citrate Crystalline Form In one embodiment, the 1:1 compound II tris salt is a crystalline form, Form A, characterized by an X-ray powder diffraction pattern that includes peaks at 2θ of 5.4°, 9.4°, 12.4°, 14.3°, and 17.8° ± 0.2. In another embodiment, Form A is characterized by an X-ray powder diffraction pattern that includes at least three (or four) peaks selected from 5.4°, 9.4°, 12.4°, 14.3°, and 17.8° ± 0.2 at 2θ. In yet another embodiment, Form A is characterized by an X-ray powder diffraction pattern that includes peaks at 2θ of 5.4°, 9.4°, 10.8°, 12.4°, 14.3°, 16.2°, 17.8°, 19.6°, and 24.9° ± 0.2. In yet another embodiment, Form A is characterized by an X-ray powder diffraction pattern that includes peaks at 2θ of 5.4°, 9.4°, 10.8°, 12.4°, 14.3°, 16.2°, 17.8°, 18.8°, 19.6°, 23.6°, and 24.9° ± 0.2. In yet another embodiment, Form A is characterized by an X-ray powder diffraction pattern that is substantially the same as FIG. 4.

[0031] In another embodiment, the 1:1 compound I tris salt Form A is characterized by a differential scanning calorimetry (DSC) peak phase transition temperature of 170 ± 3 °C (e.g., 169.9 °C).

[0032] Characterization of the 1:1 Compound II Tris Salt Crystalline Form In one embodiment, the 1:1 compound II tris salt is a single crystalline form, Form B, characterized by an X-ray powder diffraction pattern that includes peaks at 2θ of 4.1°, 14.7°, 18.8°, 20.1°, and 23.1° ± 0.2. In another embodiment, Form B is characterized by an X-ray powder diffraction pattern that includes at least three (or four) peaks selected from 2θ of 4.1°, 14.7°, 18.8°, 20.1°, and 23.1° ± 0.2. In yet another embodiment, Form B is characterized by an X-ray powder diffraction pattern that includes peaks at 2θ of 4.1°, 8.2°, 14.7°, 16.4°, 18.8°, 20.1°, 20.7°, 21.3°, and 23.1° ± 0.2. In yet another embodiment, Form B is characterized by an X-ray powder diffraction pattern that includes peaks at 2θ of 4.1°, 8.2°, 14.7°, 16.4°, 18.8°, 19.1°, 20.1°, 20.7°, 21.3°, 23.1°, 24.1°, and 25.4° ± 0.2. In yet another embodiment, Form B is characterized by an X-ray powder diffraction pattern that is substantially the same as FIG. 6.

[0033] In another embodiment, the 1:1 compound II tris salt Form B is characterized by a differential scanning calorimetry (DSC) peak phase transition temperature of 168 ± 4 °C (e.g., 170.3 °C).

[0034] In one embodiment, the 1:1 compound II tris salt is a single crystalline form, Form G, characterized by an X-ray powder diffraction pattern that includes peaks at 2θ of 6.2°, 7.6°, 13.1°, 13.4°, and 18.5° ± 0.2. In another embodiment, Form G is characterized by an X-ray powder diffraction pattern that includes at least three (or four) peaks selected from 6.2°, 7.6°, 13.1°, 13.4°, and 18.5° ± 0.2 at 2θ. In yet another embodiment, Form G is characterized by an X-ray powder diffraction pattern that includes peaks at 2θ of 6.2°, 7.6°, 13.1°, 13.4°, 18.5°, 21.5°, 23.7°, and 24.1° ± 0.2. In yet another embodiment, Form G is characterized by an X-ray powder diffraction pattern that includes peaks at 2θ of 6.2°, 7.6°, 13.1°, 13.4°, 18.0°, 18.5°, 20.8°, 21.5°, 23.7°, and 24.1° ± 0.2. In yet another embodiment, Form B is characterized by an X-ray powder diffraction pattern that is substantially the same as FIG. 8.

[0035] In another embodiment, the 1:1 compound II tris salt Form G is characterized by a differential scanning calorimetry (DSC) peak phase transition temperature of 129.5 ± 4 °C.

[0036] Pharmaceutical composition In another embodiment, a pharmaceutical composition is disclosed herein. Such a pharmaceutical composition includes a salt of Compound I (or Compound II) described herein and a pharmaceutically acceptable carrier. Other pharmacologically active substances may also be present.

[0037] As used herein, "pharmaceutically acceptable carrier" includes all solvents, dispersion media, coatings, antibacterial and antifungal agents, isotonic and absorption delaying agents, and the like that are physiologically compatible. Examples of pharmaceutically acceptable carriers include one or more of water, saline, phosphate buffered saline, dextrose, glycerol, ethanol, and combinations thereof, and may include isotonic agents in the composition, such as sugars, sodium chloride, or polyhydric alcohols such as mannitol or sorbitol. Pharmaceutically acceptable substances such as wetting agents that enhance the shelf life or effectiveness of the antibody or antibody portion, or small amounts of auxiliary substances such as wetting or emulsifying agents, preservatives or buffering agents.

[0038] The compositions of the present disclosure can be in various forms. These include, for example, liquid solutions (e.g., injectable and infusible solutions), dispersions or suspensions, tablets, pills, powders, liposomes, and suppositories, and other liquid, semi-solid and solid dosage forms. The form depends on the intended mode of administration and therapeutic use.

[0039] Typical compositions are in the form of injectable or infusible solutions, such as those generally used for passive immunization of humans with antibodies. One mode of administration is parenteral (e.g., intravenous, subcutaneous, intraperitoneal, intramuscular). In another embodiment, the antibody is administered by intravenous infusion or injection. In yet another embodiment, the antibody is administered by intramuscular injection or subcutaneous injection.

[0040] Oral administration in solid dosage forms can be presented in individual units such as, for example, hard or soft capsules, pills, cachets, troches, or tablets, each containing a predetermined amount of at least one compound of the present disclosure. In another embodiment, oral administration may be in the form of a powder or granules. In another embodiment, the oral dosage form is sublingual, such as, for example, a troche. In such solid dosage forms, any one of the compounds of the above formulas is usually combined with one or more adjuvants. Such capsules or tablets may contain controlled-release formulations. In the case of capsules, tablets, and pills, the dosage form may also contain buffering agents or can be prepared using enteric coatings.

[0041] In another embodiment, oral administration may be in a liquid dosage form. Liquid dosage forms for oral administration include, for example, pharmaceutically acceptable emulsions, solutions, suspensions, syrups, and elixirs containing an inert diluent commonly used in the art (such as water). Such compositions may also contain adjuvants such as wetting agents, emulsifying agents, suspending agents, flavoring agents (such as sweetening agents), and / or fragrances.

[0042] In another embodiment, the present disclosure includes parenteral dosage forms.

[0043] "Parenteral administration" includes, for example, subcutaneous injection, intravenous injection, intraperitoneal, intramuscular injection, intracardiac injection, and infusion. Injectable preparations (i.e., sterile injectable aqueous or oily suspensions) can be formulated according to known techniques using suitable dispersing agents, wetting agents, and / or suspending agents.

[0044] In another embodiment, the present disclosure includes topical dosage forms.

[0045] Examples of "local administration" include transdermal administration, such as via a transdermal patch or iontophoresis device, intraocular administration, or intranasal or inhalation administration. Compositions for local administration also include, for example, local gels, sprays, ointments, and creams. The topical formulation may contain a compound that enhances the absorption or penetration of the active ingredient through the skin or other affected area. When the compounds of the present disclosure are administered by a transdermal device, the administration is achieved using a patch of either the reservoir and porous membrane type or the solid matrix type. Typical formulations for this purpose include gels, hydrogels, lotions, solutions, creams, ointments, powders, dressings, foams, films, skin patches, wafers, implants, sponges, fibers, bandages, and microemulsions. Liposomes can also be used. Typical carriers include alcohols, water, mineral oil, liquid petrolatum, white petrolatum, glycerin, polyethylene glycol, and propylene glycol. Penetration enhancers can be incorporated; see, for example, Finnin and Morgan, J. Pharm. Sci., 88:955-958, 1999.

[0046] Formulations suitable for local administration to the eye include, for example, eye drops, in which the compounds of the present disclosure are dissolved or suspended in a suitable carrier. A typical formulation suitable for eye or ear administration may be in the form of eye drops of a micronized suspension or solution in isotonic, pH-adjusted, sterile saline. Other formulations suitable for eye and ear administration include ointments, biodegradable (i.e., absorbable gel sponges, collagen) and non-biodegradable (i.e., silicone) implants, wafers, lenses, and particulate or vesicular systems such as niosomes or liposomes. Polymers such as crosslinked polyacrylic acid, polyvinyl alcohol, hyaluronic acid, cellulose-based polymers such as hydroxypropylmethylcellulose, hydroxyethylcellulose, or methylcellulose, or heteropolysaccharide polymers such as gellan gum may be incorporated together with a preservative such as benzalkonium chloride. Such formulations can also be delivered by iontophoresis.

[0047] For nasal administration or administration by inhalation, the compounds of the present disclosure are conveniently delivered in the form of a solution or suspension from a pump spray container that is compressed or pumped by the patient, using a suitable propellant, or as an aerosol spray from a pressurized container or nebulizer. Formulations suitable for nasal administration are typically in the form of dry powder from a dry powder inhaler (either alone, as a mixture, e.g., as a dry blend with lactose, or as mixed component particles, e.g., mixed with a phospholipid such as phosphatidylcholine), with or without the use of a suitable propellant such as 1,1,1,2-tetrafluoroethane or 1,1,1,2,3,3,3-heptafluoropropane, or as an aerosol spray from a pressurized container, pump, spray, atomizer (preferably an atomizer that generates a fine mist using electrohydrodynamics), or nebulizer. For nasal use, the powder may contain a bioadhesive, e.g., chitosan or cyclodextrin.

[0048] In another embodiment, the present disclosure includes a rectal administration form. Such rectal administration forms may be, for example, in the form of suppositories. Cocoa butter is a traditional suppository base, but various alternatives can be used as needed.

[0049] Other carrier materials and modes of administration known in the pharmaceutical art may also be used. The pharmaceutical compositions of the present disclosure can be prepared by any of the well-known techniques of pharmacy, such as effective formulations and administration procedures.

[0050] The above considerations regarding effective formulations and administration procedures are well known in the art and are described in standard texts. Pharmaceutical formulations of drugs are described, for example, in Hoover, John E., Remington’s Pharmaceutical Sciences, Mack Publishing Co., Easton, Pa., 1975, Liberman et al., Eds., Pharmaceutical Dosage Forms, Marcel Decker, New York, N.Y., 1980, and Kibbe et al., Eds., Handbook of Pharmaceutical Excipients (3 rd Ed.), American Pharmaceutical Association, Washington, 1999.

[0051] Method of treatment The "subject" is a mammal, preferably a human, but may also be an animal in need of veterinary treatment, such as a companion animal (e.g., dog, cat, etc.), a farm animal (e.g., cow, sheep, pig, horse, etc.) and a laboratory animal (e.g., rat, mouse, guinea pig, etc.).

[0052] A "treatment" regimen of a subject with an effective amount of a compound of the present disclosure may consist of a single administration or, alternatively, may include a series of applications. The length of the treatment period depends on various factors such as the severity of the disease, the age of the subject, the concentration and activity of the compound of the present disclosure, or a combination thereof. It will also be understood that the effective dosage of the compound used for treatment or prevention may increase or decrease during the course of a particular treatment or prevention regimen. Changes in dosage may be brought about and revealed by standard diagnostic assays known in the art. In some cases, chronic administration may be required.

[0053] In one aspect, the present disclosure provides salts of Compound I (or Compound II) as described herein for use in the prevention and / or treatment of the cardio-metabolic and related diseases discussed herein, including T2DM, prediabetes, NASH, and cardiovascular diseases.

[0054] In another aspect, the present disclosure provides a method for treating a disease for which an agonist of GLP-1R is an indication, in a subject in need of such prevention and / or treatment, comprising administering to the subject a therapeutically effective amount of a salt of Compound I (or Compound II) as described herein.

[0055] In another aspect, the present disclosure provides the use of a salt of Compound I (or Compound II) as described herein for the manufacture of a medicament for treating a disease or condition for which an agonist of GLP-1R is an indication.

[0056] In another aspect, the present disclosure provides a salt of Compound I (or Compound II) as described herein for use in the treatment of a disease or condition for which an agonist of GLP-1R is an indication.

[0057] In another aspect, the present disclosure provides a pharmaceutical composition for the treatment of a disease or condition for which an agonist of GLP-1R is an indication, comprising a salt of Compound I (or Compound II) as described herein.

[0058] The present disclosure also provides a pharmaceutical composition comprising a salt of Compound I (or Compound II) as described herein for use in the treatment and / or prevention of the cardiovascular and metabolic and related diseases discussed herein, including T2DM, prediabetes, NASH, and cardiovascular diseases.

[0059] In another aspect, the present disclosure relates to diabetes (including T1D and / or T2DM including prediabetes), idiopathic T1D (type 1b), latent autoimmune diabetes in adults (LADA), early-onset T2DM (EOD), young-onset atypical diabetes (YOAD), maturity-onset diabetes of the young (MODY), malnutrition-related diabetes, gestational diabetes, hyperglycemia, insulin resistance, hepatic insulin resistance, glucose tolerance disorders, diabetic neuropathy, diabetic nephropathy, kidney diseases (e.g., acute kidney injury, tubule dysfunction, inflammatory-induced changes to the proximal tubule), diabetic retinopathy, adipocyte dysfunction, visceral fat deposition, sleep apnea, obesity (including hypothalamic obesity and monogenic obesity) and related co-morbidities (e.g., osteoarthritis and urinary incontinence), eating disorders (including binge eating disorder, bulimia nervosa, and syndromic obesity such as Prader-Willi syndrome and Bardet-Biedl syndrome), weight gain due to the use of other drugs (e.g., due to the use of steroids and antipsychotics), excessive sugar craving, dyslipidemia (including hyperlipidemia, hypertriglyceridemia, increased total cholesterol, high LDL cholesterol, and low HDL cholesterol), hyperinsulinemia, NAFLD (including related diseases such as steatosis, NASH, fibrosis, cirrhosis, and hepatocellular carcinoma), cardiovascular diseases, atherosclerosis (including coronary artery disease), peripheral vascular diseases, hypertension, endothelial disorders, vascular compliance disorders, congestive heart failure, myocardial infarction (e.g., necrosis and apoptosis), stroke, hemorrhagic stroke, ischemic stroke, traumatic brain injury, pulmonary hypertension, restenosis after angioplasty, intermittent claudication, postprandial lipemia, metabolic acidosis, ketosis, arthritis, osteoporosis, Parkinson's disease, left ventricular hypertrophy, peripheral arterial disease, macular degeneration, cataracts, glomerulosclerosis, chronic kidney disease, metabolic syndrome, syndrome X, premenstrual syndrome, angina, thrombosis, atherosclerosis, transient ischemic attack, vascular restenosis, glucose metabolism disorders, states of impaired fasting plasma glucose, hyperuricemia, gout, erectile dysfunction, skin and connective tissue disorders, psoriasis, foot ulcers, ulcerative colitis, high apoB lipoproteinemia, Alzheimer's disease, schizophrenia, cognitive impairment, inflammatory bowel disease, short bowel syndrome Crohn's disease, colitis, irritable bowel syndrome, prevention or treatment of polycystic ovary syndrome, and treatment of poisoning (e.g.,Provided are salts of Compound I (or Compound II) as described herein for use in the treatment and / or prophylaxis of cardiometabolic and related diseases including alcohol and / or drug abuse).

[0060] In certain embodiments, the disease or disorder is obesity, eating disorder, weight gain due to use of other agents, excessive sugar craving, and dyslipidemia.

[0061] In certain embodiments, the disease or disorder is obesity.

[0062] In certain embodiments, the disease or disorder is prediabetes.

[0063] In certain embodiments, the disease or disorder is T2DM.

[0064] In certain embodiments, the disease or disorder is NASH.

[0065] In certain embodiments, the disease or disorder is NAFLD.

[0066] In certain embodiments, the disease or disorder is a cardiovascular disease such as hypertension.

[0067] In another aspect, the present disclosure provides a method of enhancing or stimulating GLP-1R-mediated cAMP signaling with reduced β-arrestin / arrestin-2 recruitment, comprising administering any one of the compounds of any one of the above formulas (e.g., Formulas I, II-A, III-A, and IV-A), or a pharmaceutically acceptable salt, stereoisomer, solvate, or hydrate thereof, as defined by any one of the embodiments described herein. This is based in part on the surprising discovery that, while the compounds of the present disclosure are full agonists of GLP-1R-mediated cAMP signaling, they are partial agonists of β-arrestin recruitment to the activated GLP-1R compared to the native GLP-1R ligand GLP-1 in that the maximal β-arrestin recruitment to the activated GLP-1R by the compounds of the present disclosure is lower than the maximal β-arrestin recruitment by GLP-1. Such partial and / or biased agonists of GLP-1R for cAMP signaling may provide more sustained cAMP signaling activity for better efficacy and lower side effects.

[0068] Accordingly, the methods of the present disclosure can be advantageously used for the treatment of any of the diseases or conditions described herein, such as type II diabetes (T2D) and related disorders.

[0069] In certain embodiments, the treatment induces a glycemic benefit without an attendant increase, or at least with a reduced increase, in GI side effects such as nausea, vomiting, or diarrhea. In certain embodiments, the treatment has greater tolerability compared to a control treatment having normal or enhanced β-arrestin recruitment (such as β-arrestin recruitment by GLP-1).

[0070] Administration and Dosage Typically, the compounds of the present disclosure are administered in an amount effective to treat the conditions described herein. The compounds of the present disclosure can be administered as the compound itself or, alternatively, as a pharmaceutically acceptable salt. For purposes of administration and dosage, the compound itself or its pharmaceutically acceptable salt is simply referred to as the compound of the present disclosure.

[0071] The compounds of the present disclosure are administered by any suitable route, in the form of a pharmaceutical composition adapted to such a route, at a therapeutically effective dose for the intended treatment. The compounds of the present disclosure can be administered orally, rectally, vaginally, parenterally, or topically.

[0072] The compounds of the present disclosure can be administered orally. Oral administration may involve swallowing so that the compound enters the gastrointestinal tract, or may use oral or sublingual administration where the compound enters the bloodstream directly from the mouth.

[0073] In another embodiment, the compounds of the present disclosure can also be administered directly into the bloodstream, muscle, or viscera. Suitable means for parenteral administration include intravenous, intraarterial, intraperitoneal, intrathecal, intracerebroventricular, intraurethral, intrasternal, intracranial, intramuscular, and subcutaneous. Suitable devices for parenteral administration include needles (including microneedles) syringes, needleless syringes, and infusion techniques.

[0074] In another embodiment, the compounds of the present disclosure can also be administered topically to the skin or mucosa, i.e., dermally or transdermally. In another embodiment, the compounds of the present disclosure can also be administered intranasally or by inhalation. In another embodiment, the compounds of the present disclosure can be administered rectally or vaginally. In another embodiment, the compounds of the present disclosure can also be administered directly to the eye or ear.

[0075] The dosing regimen of the compounds of the present disclosure and / or compositions containing such compounds is based on various factors including the patient type, age, weight, gender and medical condition, severity of the condition, route of administration, and the activity of the specific compound used. Accordingly, the dosing regimen may vary widely. In one embodiment, the total daily dose of the compounds of the present disclosure is typically about 0.001 to about 100 mg / kg (i.e., mg of the compounds of the present disclosure per kg of body weight) for the treatment of the indicated conditions discussed herein. In another embodiment, the total daily dose of the compounds of the present disclosure is about 0.01 to about 30 mg / kg, in another embodiment about 0.03 to about 10 mg / kg, and in yet another embodiment about 0.1 to about 3 mg / kg. It is not uncommon for the administration of the compounds of the present disclosure to be repeated multiple times (typically 4 times or less) per day. If necessary, the total daily dose can usually be increased using multiple doses per day. In certain embodiments, the patient is a human such as a human having one of the indications or disorders of treatable diseases described elsewhere herein.

[0076] For oral administration, the composition can be provided in the form of tablets containing 0.1, 0.5, 1.0, 2.5, 5.0, 10.0, 15.0, 25.0, 30.0, 50.0, 75.0, 100, 125, 150, 175, 200, 250, and 500 milligrams of the active ingredient for symptomatic adjustment of the dosage to the patient. The medicament typically contains from about 0.01 mg to 500 mg of the active ingredient, or in another embodiment from about 1 mg to about 100 mg of the active ingredient. Intravenous, the dosage may range from about 0.01 to about 10 mg / kg / min during a constant rate infusion.

[0077] Suitable subjects or patients according to the present disclosure include mammalian subjects including humans, or non-human mammals such as primates, rodents (such as mice, rats, hamsters, rabbits, etc.). In one embodiment, a human is a suitable subject. The human subject can be of either gender and at any stage of development. In certain embodiments, the human is a pediatric patient under 18 years old, 15 years old or about 14 years old, 12 years old, 10 years old, or under 5 years old.

[0078] Concurrent administration The compounds of the present disclosure can be used alone or in combination with other therapeutic agents. The present disclosure provides any of the uses, methods or compositions defined herein, and a compound of any one of the above formulas herein, or a pharmaceutically acceptable salt thereof, or a pharmaceutically acceptable solvate of the compound or salt, is used in combination with one or more other therapeutic agents contemplated herein.

[0079] Administering two or more compounds "in combination" means that all compounds are administered at times sufficiently close that each can produce a biological effect within the same time frame. The presence of one agent may alter the biological effect of other compounds. Two or more compounds can be administered simultaneously, concurrently, or sequentially. In addition, concurrent administration can be effected by mixing the compounds prior to administration or by administering the compounds as separate dosage forms at the same time but at the same or different sites of administration.

[0080] The phrases "concurrent administration", "co-administration", "simultaneous administration", and "administered simultaneously" mean that the compounds are administered in combination.

[0081] In another embodiment, the present disclosure provides a method of treatment comprising administering a compound of the present disclosure in combination with one or more other pharmaceuticals, wherein the one or more other pharmaceuticals can be selected from the agents contemplated herein.

[0082] In one embodiment, the compounds of the present disclosure are administered with antidiabetic agents including, but not limited to, biguanides (e.g., metformin), sulfonylureas (e.g., tolbutamide, glibenclamide, gliclazide, chlorpropamide, tolazamide, acetohexamide, glyclopyramide, glimepiride, or glibizide), thiazolidinediones (e.g., pioglitazone, rosiglitazone, or lobeglitazone), glitazars (e.g., saroglitazar, aleglitazar, muraglitazar, or tesaglitazar), meglitinides (e.g., nateglinide, repaglinide), dipeptidyl peptidase 4 (DPP-4) inhibitors (e.g., sitagliptin, vildagliptin, saxagliptin, linagliptin, gemigliptin, anagliptin, teneligliptin, alogliptin, treagliptin, dutogliptin, or omarigliptin), glitazones (e.g., pioglitazone, rosiglitazone, balaglitazone, riboglitazone, or lobeglitazone), sodium-glucose co-transporter 2 (SGLT2) inhibitors (e.g., empagliflozin, canagliflozin, dapagliflozin, ipragliflozin, tofogliflozin, sergliflozin etabonate, remogliflozin etabonate, or ertugliflozin), SGLTL1 inhibitors, GPR40 agonists (FFAR1 / FFA1 agonists, e.g., fasudilamide), glucose-dependent insulinotropic polypeptide (GIP) and its analogs, alpha-glucosidase inhibitors (e.g., voglibose, acarbose, or miglitol), or insulin or insulin analogs, and pharmaceutically acceptable salts of the specifically designated agents, as well as pharmaceutically acceptable solvates of the agents and salts.

[0083] In another embodiment, the compounds of the present disclosure are administered with an anti-obesity agent including, but not limited to, a pharmaceutically acceptable salt of a specifically designated agent, and a pharmaceutically acceptable solvate of the agent and the salt, a peptide YY or an analog thereof, a neuropeptide Y receptor type 2 (NPYR2) agonist, an NPYR1 or NPYR5 antagonist, a cannabinoid receptor type 1 (CB1R) antagonist, a lipase inhibitor (e.g., orlistat), a human pancreatic polypeptide (HIP), a melanocortin receptor 4 agonist (e.g., setmelanotide), a melanin-concentrating hormone receptor 1 antagonist, a farnesoid X receptor (FXR) agonist (e.g., obeticholic acid), zonisamide, phentermine (alone or in combination with topiramate), a norepinephrine / dopamine reuptake inhibitor (e.g., bupropion), an opioid receptor antagonist (e.g., naltrexone), a combination of a norepinephrine / dopamine reuptake inhibitor and an opioid receptor antagonist (e.g., a combination of bupropion and naltrexone), a GDF-15 analog, sibutramine, a cholecystokinin agonist, amylin and an analog thereof (e.g., pramlintide), leptin and an analog thereof (e.g., metreleptin), a serotonin agonist (e.g., lorcaserin), a methionine aminopeptidase 2 (MetAP2) inhibitor (e.g., beloranib or ZGN-1061), phentermine, diethylpropion, benzfetamine, an SGLT2 inhibitor (e.g., empagliflozin, canagliflozin, dapagliflozin, ipragliflozin, ipragliflozin, tofogliflozin, sergliflozin etabonate, remogliflozin etabonate, or ertugliflozin), an SGLTL1 inhibitor, a dual SGLT2 / SGLT1 inhibitor, a fibroblast growth factor receptor (FGFR) modulator, an AMP-activated protein kinase (AMPK) activator, biotin, a MAS receptor modulator, or a glucagon receptor agonist (alone or in combination with another GLP-1R agonist, e.g., liraglutide, exenatide, dulaglutide, albiglutide, lixisenatide, or semaglutide).

[0084] In another embodiment, the compounds of the present disclosure are administered with agents for treating NASH, including pharmaceutically acceptable salts of specifically designated agents, as well as pharmaceutically acceptable solvates of such agents and salts, PF-05221304, FXR agonists (e.g., obeticholic acid), PPARα / δ agonists (e.g., elafibranor), synthetic fatty acid-bile acid conjugates (e.g., alalogol), caspase inhibitors (e.g., emricasan), anti-lysyl oxidase homolog 2 (LOXL2) monoclonal antibodies (e.g., sintilimab), galectin 3 inhibitors (e.g., GR-MD-02), MAPK5 inhibitors (e.g., GS-4997), dual antagonists of chemokine receptor 2 (CCR2) and CCR5 (e.g., cenicriviroc), fibroblast growth factor 21 (FGF21) agonists (e.g., BMS-986036), leukotriene D4 (LTD4) receptor antagonists (e.g., montelukast), niacin analogs (e.g., ARI3037MO), ASBT inhibitors (e.g., volixibat), acetyl-CoA carboxylase (ACC) inhibitors (e.g., NDI010976), ketohexokinase (KHK) inhibitors, diacylglycerol acyltransferase 2 (DGAT2) inhibitors, CB1 receptor antagonists, anti-CB1R antibodies, or apoptosis signal-regulating kinase 1 (ASK1) inhibitors, but not limited thereto.

[0085] These agents and compounds of the present disclosure can be combined with pharmaceutically acceptable excipients such as physiological saline, Ringer's solution, dextrose solution, etc. Specific dosing regimens, i.e., dosage, timing, and frequency, depend on the specific individual and their medical history.

[0086] Acceptable carriers, excipients, or stabilizers are non-toxic to the recipient at the dosages and concentrations employed and include buffers such as phosphoric acid, citric acid, and other organic acids; salts such as sodium chloride; antioxidants including ascorbic acid and methionine; preservatives (octadecyl dimethyl benzyl ammonium chloride; hexamethonium chloride; benzalkonium chloride, benzetonium chloride; phenol, butyl or benzyl alcohol; alkyl parabens such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol; 3-pentanol; and m-cresol, etc.); low molecular weight (less than about 10 residues) polypeptides; proteins such as serum albumin, gelatin or Ig; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates including glucose, mannose, or dextrin; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose, or sorbitol; salt-forming counterions such as sodium; metal complexes (e.g., Zn-protein complexes); and / or nonionic surfactants such as TWEEN™, PLURONICS™ or polyethylene glycol (PEG). Liposomes containing these agents and / or compounds of the present disclosure are prepared by methods known in the art such as those described in U.S. Patent Nos. 4,485,045 and 4,544,545. Liposomes with enhanced circulation times are disclosed in U.S. Patent No. 5,013,556. Particularly useful liposomes can be produced by the reverse-phase evaporation method using a lipid composition comprising phosphatidylcholine, cholesterol, and a PEG-derivatized phosphatidylethanolamine (PEG-PE). Liposomes are extruded through a filter of defined pore size to produce liposomes of the desired diameter.

[0087] These disclosed agents and / or compounds of the present disclosure can also be encapsulated in colloidal drug delivery systems (e.g., liposomes, albumin microspheres, microemulsions, nanoparticles, and nanocapsules) or macroemulsions, such as microcapsules prepared by, for example, coacervation techniques or interfacial polymerization, e.g., hydroxy methyl cellulose or gelatin-microcapsules and poly-(methyl methacrylate) microcapsules, respectively. Such techniques are disclosed in Remington, The Science and Practice of Pharmacy, 20 th Ed., Mack Publishing (2000).

[0088] Sustained-release formulations can be used. Suitable examples of sustained-release formulations include a semipermeable matrix of a solid hydrophobic polymer containing any one of the compounds of the above formula, which matrix is in the form of a shaped article, e.g., a film or a microcapsule. Examples of sustained-release matrices include polyesters, hydrogels (e.g., poly(2-hydroxyethyl-methacrylate), or "poly(vinyl alcohol)), polylactide (U.S. Patent No. 3,773,919), copolymers of L-glutamic acid and 7-ethyl-L-glutamate, non-degradable ethylene-vinyl acetate, degradable lactic acid-glycolic acid copolymers such as those used in LUPRON DEPOT (trademark) (injectable microspheres composed of lactic acid-glycolic acid copolymer and leuprolide acetate), sucrose isobutyrate acetate, and poly-D-(-)-3-hydroxybutyric acid.

[0089] Formulations for intravenous administration must be sterile. This can be readily accomplished, for example, by filtration through a sterile filtration membrane. The compounds of the present disclosure are generally placed in a container having a sterile access port, e.g., an intravenous solution bag or vial having a stopper penetrable by a hypodermic needle.

[0090] Suitable emulsions can be prepared using commercially available fat emulsions such as Intralipid (trademark), Liposyn (trademark), Infonutrol (trademark), Lipofundin (trademark), and Lipiphysan (trademark). The active ingredient may be dissolved in a pre-mixed emulsion composition or, alternatively, in an emulsion formed by mixing an oil (e.g., soybean oil, safflower oil, cottonseed oil, sesame oil, corn oil, or almond oil) and a phospholipid (e.g., egg phospholipid, soybean phospholipid, or soy lecithin) with water. It is understood that other ingredients, such as glycerol or glucose, can be added to adjust the tonicity of the emulsion. Suitable emulsions typically contain up to 20%, for example 5 - 20%, oil. The fat emulsion can contain fat droplets of 0.1 - 1.0 μm, particularly 0.1 - 0.5 μm, and have a pH in the range of 5.5 - 8.0.

[0091] The emulsion composition may be prepared by mixing a compound of the present disclosure with Intralipid (trademark) or its components (soybean oil, egg phospholipid, glycerol, and water).

[0092] Compositions for inhalation or insufflation include solutions and suspensions, and powders, in a pharmaceutically acceptable aqueous or organic solvent or mixtures thereof. Liquid or solid compositions may contain suitable pharmaceutically acceptable excipients as described above. In some embodiments, the composition is administered via the oral or nasal respiratory route for local or systemic effects. Preferably, the composition in a sterile pharmaceutically acceptable solvent can be nebulized using a gas. The nebulized solution can be breathed directly from the nebulizer device or the nebulizer device can be attached to a face mask, tent, or intermittent positive pressure breathing apparatus. Solution, suspension, or powder compositions can be administered preferably orally or nasally from a device that delivers the formulation in an appropriate manner.

[0093] Kit Another aspect of the present disclosure provides a kit comprising any one of the compounds of the above formula, or a pharmaceutical composition comprising any one of the compounds of the above formula of the present disclosure. The kit may include, in addition to any one of the compounds of the above formula, a diagnostic or therapeutic agent of the present disclosure or its pharmaceutical composition. The kit may also include instructions for use in a diagnostic or therapeutic method. In some embodiments, the kit comprises any one of the compounds of the above formula, or its pharmaceutical composition and a diagnostic agent. In other embodiments, the kit comprises any one of the compounds of the above formula, or its pharmaceutical composition.

[0094] In yet another embodiment, the present disclosure includes a kit suitable for use in practicing the treatment methods described herein. In one embodiment, the kit includes a first dosage form comprising an amount of one or more of the compounds of the present disclosure sufficient to practice the methods of the present disclosure. In another embodiment, the kit includes an amount of one or more of the compounds of the present disclosure sufficient to practice the methods of the present disclosure, as well as a container for administration and a container for administration.

[0095] Preparation Any one of the compounds of the above formula can be prepared by the general and specific methods described below using the common general knowledge of those skilled in synthetic organic chemistry. Such common general knowledge can be found in standard references such as Comprehensive Organic Chemistry, Ed. Barton and Ollis, Elsevier, Comprehensive Organic Transformations: A Guide to Functional Group Preparations, Larock, John Wiley and Sons, and Compendium of Organic Synthetic Methods, Vol. I-XII (published by Wiley-Interscience). The starting materials used herein are either commercially available or can be prepared by conventional methods known in the art.

[0096] In the preparation of any one of the compounds of the above formula, it should be noted that some of the preparation methods described herein may require protection of remote functional groups (e.g., primary amine, secondary amine, carboxyl in any one of the precursors of the above formula). The need for such protection will vary depending on the nature of the remote functional group and the conditions of the preparation method. The need for such protection can be readily determined by those skilled in the art. The use of such protection / deprotection methods is also within the scope of the skill of those skilled in the art. For a general description of protecting groups and their use, see Greene, Protective Groups in Organic Synthesis, John Wiley & Sons, New York, 1991.

[0097] For example, certain compounds, if left unprotected, may contain primary amine or carboxylic acid functional groups that can interfere with reactions at other sites of the molecule. Thus, such functional groups can be protected by appropriate protecting groups that can be removed in the next step. Protecting groups suitable for amine and carboxylic acid protection include those commonly used in peptide synthesis (e.g., N-t-butoxycarbonyl (Boc), benzyloxycarbonyl (Cbz), and 9-fluorenylmethyleneoxycarbonyl (Fmoc) for amines, lower alkyl or benzyl esters for carboxylic acids, etc.), which generally do not react chemically under the described reaction conditions and can typically be removed without chemically altering any other functional groups of any one of the above compounds.

[0098] The schemes described below are intended to provide a general description of the methodologies used in the preparation of the compounds of the present disclosure. Some of the compounds of the present disclosure may contain one or more chiral centers with the stereochemical designations (R) or (S). It will be apparent to those skilled in the art that all synthetic transformations can be carried out in a similar manner regardless of whether the material is enantiomerically enriched or racemic. Furthermore, resolution to the desired optically active material can be carried out at any desired point in the sequence using well-known methods as described herein and in the chemical literature.

[0099] The amine compounds prepared by the methods described herein can be delivered by alkylation with a protected 2-bromoacetate in a polar aprotic solvent such as, but not limited to, DMF, DMAc, DMSO or NMP, in the presence of a suitable base such as K2CO3, Et3N, NaH or LiHMDS. Standard ester hydrolysis can be carried out to obtain the acid. Pg 2 When Pg is t-butyl, the acid can be delivered using standard acidic deprotection methods such as TFA / DCM, HCl / 1,4-dioxane, HCl / EtOAc, or other suitable conditions.

[0100] Experimental The abbreviations of the solvents are listed in the following table.

Table 1

[0101] Analysis conditions X-ray powder diffraction (XRPD) For XRPD analysis, a PANalytical Empyrean / X’ Pert3 X-ray powder diffractometer was used. The XRPD parameters used are listed in the following table.

Table 2

[0102] Single crystal X-ray diffraction (SCXRD)

Table 3

Table 4

[0103] 1 H-Nuclear magnetic resonance spectroscopy ( 1 H-NMR) 1 1H solution NMR was collected on a Bruker 400M NMR spectrometer using DMSO- d6 .

[0104] Dynamic vapor sorption (DVS) DVS was measured via an SMS (Surface Measurement System) DVS Intrinsic. The relative humidity at 25 °C was calibrated against the deliquescence points of LiCl, Mg(NO3)2, and KCl. The parameters of the DVS test are listed in the following table.

Table 5

[0105] High performance liquid chromatography (HPLC) An Agilent 1260 HPLC was used, and the detailed chromatography conditions are listed in the following table.

Table 6

[0106] Example 1. Synthesis of (S)-2-((4-(3-((4-chloro-2-fluorobenzyl)oxy)phenyl)-3,6-dihydropyridin-1(2H)-yl)methyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylic acid (Compound I)

Chemical formula

[0107] Step 2 To a solution of tert-butyl 4-(3-hydroxyphenyl)-3,6-dihydropyridine-1(2H)-carboxylate (275 mg, 1.0 mmol) in 1,4-dioxane (10 mL) was added 1-(bromomethyl)-4-chloro-2-fluorobenzene (223 mg, 1.0 mmol) and Pd2(dba)3 (91.5 mg, 0.1 mmol) and BINAP (62.2 mg, 0.1 mmol). The mixture was stirred at 100 °C for 8 h under nitrogen. The reaction was cooled to room temperature, the reaction was diluted with water (150 mL), extracted with ethyl acetate (150 mL × 3), the combined organics were washed with brine (150 mL × 3), dried, and concentrated in vacuo to give a crude product. Then, purification of the crude product by preparative TLC (PE:EA = 3:1) gave tert-butyl 4-(3-((4-chloro-2-fluorobenzyl)oxy)phenyl)-3,6-dihydropyridine-1(2H)-carboxylate (241 mg, 57.8% yield) as a yellow oil. LCMS: [M+H] +=363, Retention time (10 mM NH4HCO3) = 2.04 minutes.

[0108] Step 3 To a solution of tert-butyl 4-(3-((4-chloro-2-fluorobenzyl)oxy)phenyl)-3,6-dihydropyridine-1(2H)-carboxylate (322 mg, 0.77 mmol) in DCM (10 mL) was added HCl / 1,4-dioxane (1.2 mL). The mixture was stirred at room temperature for 2 hours. The reaction was diluted with water (50 mL) and extracted with ethyl acetate (50 mL × 3). The combined organics were washed with brine (50 mL × 3), dried, and concentrated in vacuo to give the crude product, which was purified by preparative TLC (PE:EA = 5:1) to afford 4-(3-((4-chloro-2-fluorobenzyl)oxy)phenyl)-1,2,3,6-tetrahydropyridine (198 mg) as a yellow oil. LCMS: [M+H] + =318, Retention time (10 mM NH4HCO3) = 1.61 minutes.

[0109] Step 4 To a solution of 4-(3-((4-chloro-2-fluorobenzyl)oxy)phenyl)-1,2,3,6-tetrahydropyridine (90 mg, 0.28 mmol) in 1,4-dioxane (10 mL) were added tert-butyl (S)-2-(chloromethyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate (85 mg, 0.28 mmol) and DIPEA (0.3 mL, 1.4 mol). The mixture was stirred at 90 °C for 3 hours. The reaction was diluted with water (100 mL) and extracted with ethyl acetate (100 mL × 3). The combined organic layers were washed with brine (100 mL × 3), dried, and concentrated in vacuo to give the crude product, which was purified by preparative TLC to afford tert-butyl (S)-2-((4-(3-((4-chloro-2-fluorobenzyl)oxy)phenyl)-3,6-dihydropyridine-1(2H)-yl)methyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate (80 mg, 46% yield) as a yellow solid.

[0110] LCMS: [M+H]+ = 618, retention time (10 mM NH4HCO3) = 2.36 min.

[0111] Step 5 To a solution of tert-butyl (S)-2-((4-(3-((4-chloro-2-fluorobenzyl)oxy)phenyl)-3,6-dihydropyridin-1(2H)-yl)methyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylate (80 mg, 0.13 mmol) in DCM (12 mL) was added TFA (2 mL, 26.93 mmol). The mixture was stirred at room temperature for 3 h. The reaction was concentrated in vacuo to afford the crude product, which was purified by preparative HPLC (NH4HCO3) to give (S)-2-((4-(3-((4-chloro-2-fluorobenzyl)oxy)phenyl)-3,6-dihydropyridin-1(2H)-yl)methyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylic acid (18.5 mg, 25% yield) as a white solid. LCMS: [M+H] + = 562.0, retention time (10 mM NH4HCO3) = 1.40 min.

[0112] 11H NMR (400 MHz, MeOD) δ 8.21 - 8.20 (brs, 1H), 7.97 (dd, J = 8.5, 1.4 Hz, 1H), 7.62 (d, J = 8.5 Hz, 1H), 7.54 (t, J = 8.2 Hz, 1H), 7.25 (ddd, J = 6.3, 5.6, 2.0 Hz, 3H), 7.11 - 7.02 (m, 2H), 6.89 (dd, J = 7.3, 1.9 Hz, 1H), 6.13 - 6.12 (brs, 1H), 5.28 (dd, J = 9.4, 5.0 Hz, 1H), 5.13 (s, 2H), 4.92 (d, J = 7.1 Hz, 1H), 4.77 - 4.71 (m, 1H), 4.63 (dd, J = 13.4, 8.2 Hz, 1H), 4.53 - 4.46 (m, 1H), 4.14 (d, J = 13.6 Hz, 1H), 4.03 (d, J = 13.6 Hz, 1H), 3.25 (d, J = 2.4 Hz, 2H), 2.80 (ddd, J = 22.5, 12.9, 7.2 Hz, 3H), 2.62 - 2.48 (m, 3H).

[0113] Example 2. Synthesis of (S)-2-((6-((4-cyano-2-fluorobenzyl)oxy)-3’,6’-dihydro-[2,4’-bipyridin]-1’(2’H)-yl)methyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylic acid (Compound II)

Chemical Structure

[0114] 1 H NMR (400 MHz, CDCl3) δ 7.67 - 7.63 (t, J = 7.6 Hz, 1H), 7.59 - 7.55 (t, J = 7.6 Hz, 1H), 7.49 - 7.46 (dd, J1 = 8.0 Hz, J2 = 1.2 Hz, 1H), 7.40 - 7.37 (dd, J1 = 9.2 Hz, J2 = 1.2 Hz, 1H), 6.97 - 6.95 (d, J = 7.6 Hz, 1H), 6.75 - 6.73 (d, J = 8.4 Hz, 1H), 5.48 (s, 2H).

[0115] Step 1a A mixture of 4-[(6-chloro-2-pyridyl)oxymethyl]-3-fluoro-benzonitrile (1 g, 3.81 mmol), tert-butyl 4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)-3,6-dihydro-2H-pyridine-1-carboxylate (1.29 g, 4.19 mmol), Pd(dppf)Cl2 (278.3 mg, 0.38 mmol) and NaHCO3 (479.69 mg, 5.71 mmol) in dioxane (20 mL) and H2O (4 g, 222.22 mmol) was stirred at 90 °C for 2 h under N2 until the reaction was complete as indicated by LCMS. The reaction mixture was filtered through a pad of celite using EtOAc, and the combined organics were concentrated in vacuo and purified by silica gel chromatography (hexane / EtOAc = 0 - 11%) to give the desired product tert-butyl 4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridyl]-3,6-dihydro-2H-pyridine-1-carboxylate (1.5 g, 3.63 mmol, 95.4% yield) as a pale yellow liquid. LCMS: [M+H] + = 410.1, retention time (10 mM NH4HCO3) = 2.22 min.

[0116] Step 2 To a solution of tert-butyl 4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridyl]-3,6-dihydro-2H-pyridine-1-carboxylate (1.5 g, 3.66 mmol) in DCM (20 mL) was slowly added TFA (7.40 g, 64.90 mmol, 5 mL). The reaction was stirred at 28 °C for 2 h. After completion of the reaction as determined by LCMS, the reaction mixture was concentrated in vacuo to give 3-fluoro-4-[[6-(1,2,3,6-tetrahydropyridin-4-yl)-2-pyridyl]oxymethyl]benzonitrile (1.8 g, 3.94 mmol) TFA salt as a pale yellowish liquid. The crude product was used directly in the next step without further purification. LCMS: [M+H] + = 310.1, retention time (0.01% TFA) = 1.42 min.

[0117] Step 3 A mixture of 3-fluoro-4-[[6-(1,2,3,6-tetrahydropyridin-4-yl)-2-pyridyl]oxymethyl]benzonitrile (340 mg, 0.80 mmol) and tert-butyl 2-(chloromethyl)-3-[[(2S)-oxetan-2-yl]methyl]benzimidazole-5-carboxylate (225 mg, 670 mmol, this synthesis is disclosed in International Application No. 2018 / 109607, which is incorporated herein by reference) and DIPEA (216.42 mg, 1.67 mmol) in dioxane (10 mL) was stirred at 90 °C for 1 hour until the reaction was complete as indicated by LCMS. The reaction mixture was concentrated in vacuo and purified by silica gel chromatography (hexane / EtOAc = 20:1) to give the desired product tert-butyl 2-[[4-[6-[(4-cyano-2-fluoro-phenyl)methoxy]-2-pyridyl]-3,6-dihydro-2H-pyridin-1-yl]methyl]-3-[[(2S)-oxetan-2-yl]methyl]benzimidazole-5-carboxylate (366 mg, 0.31 mmol) as a light brown solid. LCMS: [M+H] + = 610.0, retention time (10 mM NH4HCO3) = 1.87 min.

[0118] Step 4 A solution of tert-butyl 2-[[4-[6-[(4-cyano-2-fluorophenyl)methoxy]-2-pyridyl]-3,6-dihydro-2H-pyridin-1-yl]methyl]-3-[[(2S)-oxetan-2-yl]methyl]benzimidazole-5-carboxylate (180 mg, 0.30 mmol) in DCM (6 mL) was slowly added to TFA (2.96 g, 25.96 mmol, 2 mL) in DCM (2 mL) at 28 °C and stirred for 1 h. After completion of the reaction as determined by LCMS, the reaction mixture was concentrated in vacuo and the crude product was purified by preparative HPLC (10 mM NH4HCO3) to give (S)-2-((6-((4-cyanobenzyl)oxy)-3’,6’-dihydro-[2,4’-bipyridin]-1’(2’H)-yl)methyl)-1-(oxetan-2-ylmethyl)-1H-benzo[d]imidazole-6-carboxylic acid (54 mg, 33% yield) as a white solid. LCMS: + = 554.2, retention time (10 mM NH4HCO3) = 1.42 min.

[0119] 1 H NMR (400 MHz, DMSO-d6) δ 12.73-12.72 (brs, 1H), 8.27-8.26 (brs, 1H), 7.90 (d, J = 10.3 Hz, 1H), 7.81 (dd, J = 8.5, 1.5 Hz, 1H), 7.69 (dt, J = 13.2, 8.0 Hz, 4H), 7.10 (d, J = 7.5 Hz, 1H), 6.85-6.61 (m, 2H), 5.49 (s, 2H), 5.15-5.02 (m, 1H), 4.80 (dd, J = 15.2, 7.3 Hz, 1H), 4.71-4.60 (m, 1H), 4.40 (ddt, J = 11.9, 8.9, 6.0 Hz, 2H), 4.07 (d, J = 13.5 Hz, 1H), 3.91 (d, J = 13.5 Hz, 1H), 3.28-3.17 (m, 2H), 2.73 (d, J = 2.5 Hz, 2H), 2.70-2.60 (m, 1H), 2.43-2.42 (m, 2H), 2.44-2.34 (m, 1H).

[0120] Example 3.1: Preparation and Characterization of the Crystalline Form of Compound I of 3.1:1 Tris Salt (Form A) 3.1.1 Preparation Method A solution of 500.9 mg of the free form of Compound I was dissolved in 20 mL of acetone / H2O (9:1, v:v). Meanwhile, an equimolar amount of tris (107.9 mg) was dissolved in 2.5 mL of H2O. The resulting tris solution was added dropwise to the free form of the Compound I solution, and the mixture was stirred at room temperature (RT, about 1000 rpm). The resulting slurry was continuously stirred at RT for 17 hours, then cooled to 5 °C and stirred for an additional 4 hours. The solid in the mixture was isolated by centrifugation and dried in vacuo at RT for 20 hours to obtain 445.9 mg of the desired product (yield about 73.2%).

[0121] The resulting solid was further characterized by XRPD (Figure 1 and Table 1).

[0122] The results of TGA showed a 3.2% weight loss up to 150 °C (Figure 2A), and the results of DSC showed two endotherms at 112.0 °C and 174.3 °C (peak temperature) (Figure 2B).

[0123] 1 The 1H NMR spectrum showed that the molar ratio of tris / free form was 1.0:1, and no residual solvent was observed. VH-XRPD and single crystal structure determination were performed on the 1:1 Compound I tris salt (Form A), which was identified as a monohydrate that would dehydrate under conditions of ≤30% RH. Therefore, it is proposed that the 1:1 Compound I tris salt (Form A) be stored under conditions of 20 - 30 °C with a relative humidity of ≥40% RH to avoid dehydration.

Table 7

[0124] 3.1.2 Alternative Preparation Method The preparation of the 1:1 Compound I tris salt (Form A) was attempted several times via slurry / solution crystallization. The results of the preparation are summarized in Tables A and B. The results of the detailed characterization are summarized in Table C.

Table 8

Table 9

Table 10

[0125] Tris salt form A (Sample ID1) was re-prepared via a slurry of equimolar free form of Compound I and Tris in THF for 1 day, followed by vacuum drying at RT for about 2.5 days. The crystallinity of the sample decreased significantly after vacuum drying. The TGA / DSC curves showed a 7.1% weight loss up to a maximum of 150 °C, as well as two endotherms at 89.4 °C and 170.5 °C (peak temperatures). 1 The 1H NMR spectrum showed that the molar ratio of Tris / free form was 1.0:1 and the molar ratio of residual THF / free form was 0.62:1 (6.1%, wt%). The HPLC purity of Tris salt form A (Sample ID1) was measured to be 93.87 area%.

[0126] Tris salt form A (Sample ID3) was obtained via solution crystallization using 15 mg of the free form and equimolar Tris as starting materials. The TGA / DSC curves showed a 7.8% weight loss up to a maximum of 150 °C, as well as two endotherms at 98.2 °C and 174.6 °C (peak temperatures). 1 The 1H NMR spectrum showed that the molar ratio of Tris / free form was 1.0:1 and the molar ratio of residual acetone / free form was 0.04:1 (0.3%, wt%). The HPLC purity of Tris salt form A (Sample ID3) was measured to be 97.46 area%.

[0127] Tris salt form A (Sample ID4) was obtained via solution crystallization using 500 mg of the free form and equimolar Tris as starting materials. The TGA / DSC curves showed a 6.1% weight loss up to a maximum of 150 °C, as well as two endotherms at 90.9 °C and 174.2 °C (peak temperatures). 1The 1H NMR spectrum indicated that the molar ratio of tris / free form was 0.9:1 and the molar ratio of residual acetone / free form was 0.04:1 (0.3%, wt%). The HPLC purity of tris salt form A (Sample ID4) was measured to be 98.04 area%. Since the batch of this sample did not have a molar ratio of 1:1, it was not used further in the formulation study.

[0128] Tris salt form A (Sample ID5) was obtained via solution crystallization using 500 mg of the free form and equimolar tris as starting materials. The TGA / DSC curves showed a 3.2% weight loss up to a maximum of 150 °C and two endotherms at 112.0 °C and 174.3 °C (peak temperatures). 1 The 1H NMR spectrum indicated that the molar ratio of tris / free form was 1.0:1 and the molar ratio of residual acetone / free form was 0.05:1 (0.4%, wt%). The HPLC purity of tris salt form A (Sample ID5) was measured to be 98.16 area%.

[0129] 3.2 Single Crystal Growth Single crystal samples of the 1:1 compound I tris salt (form A), characterized by SCXRD, were obtained from slow evaporation experiments. The details of the experiments are described in detail below.

[0130] 15.0 mg of the 1:1 compound I tris salt (form A) starting material was added to a 3 mL glass vial, and 1.0 mL of an ACN / MeOH (1:1, v:v) solvent mixture was added. After shaking with an ultrasonic cleaner to accelerate dissolution, the suspension was filtered. The resulting clear solution was transferred to a clean 4 mL shell vial. The shell vial was sealed with a PE plug with one pinhole on top. Then, it was placed in a fume hood at room temperature for slow evaporation. After slow evaporation for 2 days, rod-shaped crystals were observed.

[0131] As demonstrated by single crystal X-ray crystallography (Figure 3), the asymmetric unit of the type A single crystal structure is composed of one Compound I anion, one tris-cation, and one water molecule, confirming that Form A is the monohydrate of the 1:1 Compound I tris-salt.

[0132] 3.3 Dynamic solubility I) Preparation of biologically relevant media and pH buffers Simulated gastric fluid (SGF) Weigh 100 mg of sodium chloride and 50 mg of Triton X-100 into a 50 mL volumetric flask. Add an appropriate amount of purified water and sonicate until all solids are completely dissolved. Add approximately 68 μL of 12 M HCl and sufficient purified water close to the target volume, and adjust the pH to 1.8. Make up the volume with purified water, mix well, and confirm the pH with a pH meter.

[0133] Fasted-state simulated intestinal fluid (FaSSIF) Weigh 170 mg of monobasic sodium phosphate, 21 mg of sodium hydroxide, and 310 mg of sodium chloride, 110 mg of SIF powder into a 50 mL volumetric flask. Add an appropriate amount of purified water and sonicate until all solids are completely dissolved. Add sufficient purified water close to the target volume and adjust the pH to 6.5. Make up the volume with purified water, mix well, and confirm the pH with a pH meter.

[0134] Fed-state simulated intestinal fluid (FeSSIF) Measure 0.41 mL of glacial acetic acid, 202 mg of sodium hydroxide, and 594 mg of sodium chloride, 560 mg of SIF powder into a 50 mL volumetric flask. Add an appropriate amount of purified water to dissolve the solids. Then add sufficient purified water close to the target volume and adjust the pH to 5.0. Make up the volume with purified water, mix well, and confirm the pH with a pH meter.

[0135] Buffer solution at pH 8.0 Weigh 302.5 mg of tris into a 50 mL volumetric flask. Add sufficient purified water close to the target volume and adjust the pH to 8.0. Make up the volume with purified water, mix well, and confirm the pH with a pH meter.

[0136] II) Dynamic solubility test The dynamic solubility was evaluated in water and three biorelevant media (SGF, FaSSIF, and FeSSIF) for 1 / 4 / 24 h. When 3 - 4 mg of the solid was suspended in 3 mL of each medium (solid load calculated using the free form: 1 mg / mL), a suspension was obtained, which was subsequently rolled (25 rpm) at 37 °C for 1 / 4 / 24 h. At each time point, approximately 1.0 mL of the suspension was sampled for centrifugation (12000 rpm, 2 min) and filtration through a 0.45 μm PTFE membrane to obtain the supernatant for HPLC and pH tests, and the residual solid was analyzed by XRPD. The detailed results are summarized in Table 2.

[0137] Based on the results of the solubility evaluation, the 1:1 compound I tris salt (form A) showed better solubility in H2O. The free form and tris salt type A showed similar solubility in FaSSIF and FeSSIF after 1 h. A morphological change was observed for the 1:1 compound I tris salt (form A) in SGF, and XRPD results after the solubility test were obtained. Since the sample was a suspension, the decrease in purity was only for reference.

Table 11

[0138] 3.4.1 Solid stability evaluation (I / II) To evaluate the physical and chemical stability of the 1:1 compound I tris salt (form A) and the amorphous free form, the samples were stored under the conditions of 60 °C for 1 day and 25 °C / 60%RH or 40 °C / 75%RH for 1 week respectively. XRPD and HPLC were used to characterize the solid, and the results are summarized in Table 3. No morphological change was observed after 1 week. The 1:1 compound I tris salt (form A) showed better chemical stability than the amorphous free form under all conditions.

Table 12

[0139] 3.4.2 Solid Stability Evaluation (II / II) The chemical stability of the 1:1 compound I tris salt (form A) was further evaluated in the solid state, and the samples were stored for 1 day and 3 days respectively under ambient conditions or at 40 °C / 75% RH. The HPLC purity and weight purity were tested three times using HPLC, and the results were summarized in Table 4. The 1:1 compound I tris salt (form A) showed better chemical stability under all conditions.

Table 13

[0140] 3.5 Solution Stability Evaluation The solution stability of the 1:1 compound I tris salt (form A) in SGF (pH 1.8) and buffer solution at pH 8.0 was evaluated at RT. A clear solution was prepared at a concentration of 0.01 mg / mL and then left at RT for 6 hours, 1 day, and 5 days. After 1 day, no significant decomposition was observed for the solution at pH 1.8, but the purity of the solution at pH 8.0 decreased slightly. The HPLC purity and pH values were summarized in Tables 5 and 6.

Table 14

Table 15

[0141] 3.6 Hygroscopicity To investigate the stability of the solid form as a function of humidity, the DVS isotherm plot of the 1:1 compound I tris salt (form A) was collected between 0% RH and 95% RH at 25 °C. A water absorption rate of 2.77% was observed at 25 °C / 80% RH. Considering that the 1:1 compound I tris salt (form A) is a hydrate and can dehydrate under low humidity, DVS was also tested at 20 and 30 °C to further confirm the suitable storage conditions and avoid dehydration. The detailed results were summarized in Table 7. No morphological changes were observed after the DVS test. Combining the results of VH-XRPD and DVS, the storage conditions of the 1:1 compound I tris salt (form A) are preferably 20 - 30 °C and relative humidity ≥ 40% RH.

Table 16

[0142] 3.7 Polymorph Screening Polymorph screening experiments on the 1:1 compound I tris salt (form A) were conducted under various conditions using different solution crystallization or solid transformation methods including anti-solvent addition, reverse anti-solvent addition, slow evaporation, slow cooling, slurry at RT, slurry at 50 °C, slurry circulation from 5 - 50 °C, vapor-solid diffusion, vapor-solution diffusion, polymer-induced crystallization, and milling.

[0143] The screening results indicate that the 1:1 compound I tris salt (form A) did not change under most of the test conditions, indicating that form A is stable.

[0144] Example 4. Preparation and Characterization of the Crystalline Forms of 1:1 Compound II Citrate (Form A) 4.1.1 Preparation Method 15 mg of the free form of compound II and 5.21 mg of citric acid in acetone were mixed. The resulting slurry was stirred at RT for 3 days and then vacuum dried at RT for about 12 hours.

[0145] The resulting solid was further characterized by XRPD (Figure 4 and Table 8).

[0146] The results of TGA (Figure 5A) showed a 2.6% weight loss up to 150 °C maximum, and the results of DSC (Figure 5B) showed one endotherm at 169.9 °C and one exotherm at 173.3 °C (peak temperature).

Table 17

[0147] 4.1.2 Alternative Preparation Method The preparation of the 1:1 compound II citrate (Form A) was attempted several times via slurry / solution crystallization. The preparation results are summarized in Tables 9 and 10. The detailed characterization results are summarized in Table 11.

Table 18

Table 19

Table 20

[0148] The 1:1 compound II citrate (Form A) (Sample ID1) was re-prepared via a slurry of equimolar starting materials and citric acid in acetone for 1 day, followed by vacuum drying at RT for approximately 1.5 days. The TGA / DSC curves showed a 1.4% weight loss up to a maximum of 150 °C, one endotherm at 171.9 °C, and one exotherm at 175.5 °C (peak temperature). 1 The H NMR spectrum showed that the molar ratio of citrate / free form was 1.0:1 (the NMR signals of citrate and the free form overlapped, and the integration of the free form was subtracted for calculation), and the molar ratio of residual acetone / free form was 0.16:1 (1.2%, wt%). The HPLC purity of the 1:1 compound II citrate (Form A) (Sample ID1) was measured to be 97.73 area%.

[0149] The 1:1 compound II citrate (Form A) (Sample ID2) was obtained via solution crystallization using 15 mg of the free form and citric acid (1:1 molar ratio, acid / free form) as starting materials. The TGA / DSC curves showed a 3.1% weight loss up to a maximum of 150 °C, as well as one endotherm at 169.9 °C and one exotherm at 173.6 °C (peak temperature). 1The 1H NMR spectrum showed that the molar ratio of citrate / free form was 1.0:1 (the integral of the free form was subtracted for calculation), and the molar ratio of residual acetone / free form was 0.09:1 (0.9%, wt%). The HPLC purity of the 1:1 compound II citrate (form A) (sample ID2) was measured to be 97.05 area%.

[0150] The 1:1 compound II citrate (form A) (sample ID3) was obtained via solution crystallization using 500 mg of the free form and citric acid (1:1 molar ratio, acid / free form) as starting materials. The TGA / DSC curves showed a 2.5% weight loss up to 150 °C, as well as one endotherm at 166.6 °C and one exotherm at 171.0 °C (peak temperature). 1 The 1H NMR spectrum showed that the molar ratio of citrate / free form was 1.0:1 (the integral of the free form was subtracted for calculation), and the molar ratio of residual acetone / free form was 0.08:1 (0.6%, wt%). The HPLC purity of the 1:1 compound II citrate (form A) (sample ID3) was measured to be 96.66 area%.

[0151] Example 5. Preparation and Characterization of the Crystalline Form of 1:1 Compound II Tris Salt (Form B) 5.1.1 Preparation Method Batch A: 1) Under a N2 atmosphere, the reactor was charged with acetone (10 L, 10.0 V). 1.0 Kg (1.0 equivalent) of the free form of compound II was added to the reactor with stirring. The temperature was adjusted to 23 °C, and the resulting mixture was stirred for 0.5 h and then filtered through a 0.2 μm microporous filter. The filtrate was collected. 2) Under a N2 atmosphere, the filtrate was transferred to the reactor. 10 L of EA (10.0 V) was added to the reactor. The temperature was adjusted to 21 °C. The tris solution (218.8 g of tris (1.0 equivalent) dissolved in 2 L of soft water (2.0 V)) was added to the reactor within 1.5 h. The mixture was stirred for 1 h. 3) The mixture was then isolated by centrifugation. The filter cake was washed with EA (2 L, 2.0 V), and the filter cake was collected. 4) When the filter cake was dried under vacuum at 70 °C for 16 hours, 1.03 Kg of the 1:1 compound II tris salt (form B) was obtained as an off-white solid.

[0152] Batch B: 1) Under a N2 atmosphere, the reactor was charged with acetone (21.5 L, 10.0 V). 2.15 Kg (1.0 equivalent) of the free form of compound II was added to the reactor with stirring. The temperature was adjusted to 25 °C, and the resulting mixture was stirred for 0.5 hour, after which the mixture was filtered through a 0.2 μm microporous filter. The filtrate was collected. 2) Under a N2 atmosphere, the filtrate was transferred to the reactor. 21.5 L of EA (10.0 V) was added to the reactor. The temperature was adjusted to 21 °C. A tris solution (470.4 g of tris (1.0 equivalent) dissolved in 4.3 L of soft water (2.0 V)) was added to the reactor within 3 hours. The mixture was stirred for 1.5 hours. 3) The mixture was then isolated by centrifugation. The cake was washed with EA (4.3 L, 2.0 V), and the filter cake was collected. 4) When the filter cake was dried under vacuum at 70 °C for 16 hours, 2.45 Kg of the 1:1 compound II tris salt (form B) was obtained as a white solid.

[0153] Batch C: 1) 3.45 Kg (1.0 eq) of the 1:1 compound II tris salt (form B) obtained from Batch A and Batch B was mixed with 3.45 L of soft water (1 V). The mixture was added to a reactor charged with 44.9 L of EA (13 V) under a N2 atmosphere with stirring. The temperature was adjusted to 25 °C, and the resulting mixture was stirred for 1 hour. 2) The mixture was then isolated by centrifugation. The cake was washed with EA (6.9 L, 2.0 V), and the filter cake was collected. 3) When the filter cake was dried under vacuum at 70 °C for 16 hours, 3.45 Kg of the 1:1 compound II tris salt (form B) was obtained as an off-white solid.

[0154] The 1:1 compound II tris salt (Form B) of Batch C was characterized by XRPD (Figure 6 and Table 5-1). The results of TGA showed a 3.7% weight loss up to a maximum of 150 °C (Figure 7A), and the results of DSC (Figure 7B) showed two endotherms at 116.1 °C and 170.3 °C (peak temperature).

[0155] VH-XRPD was performed on a sample of the 1:1 compound II tris salt (Form B) for further identification. VH-XRPD showed that the tris salt Form B converts to a new form with N2 at 10% RH and converts back after exposure to ambient conditions (40% RH). Combining the results of the characterization, the 1:1 compound II tris salt (Form B) is considered to be a hydrate and dehydrates at < 10% RH.

Table 21

[0156] 5.1.2 Alternative Preparation Methods A mixture of compound II and the free form of tris (1:0.95 molar ratio) in THF / MTBE (1:4, v:v) was stirred for 7 days and then dried under vacuum at RT for about 3 hours to obtain the 1:1 compound II tris salt (Form B).

[0157] Example 6. Preparation and Characterization of the Crystalline Form of the 1:1 Compound II Tris Salt (Form G) 6.1 Preparation Method 15 mg of the 1:1 compound II tris salt (Form B) was dissolved in 0.1 mL of NMP to form a clear solution. CH3CN was added dropwise to the solution. A precipitate was observed. The solid was isolated via centrifugation, heated to 105 °C, and then cooled to RT to obtain the 1:1 compound II tris salt (Form G).

[0158] The solid obtained was further characterized by XRPD (Figure 8 and Table 6-1).

[0159] The results of TGA (Figure 9) showed an 11.75% weight loss up to a maximum of 150 °C.

[0160] The DSC results (Figure 10) showed two endotherms at 50.9 °C and 129.5 °C (peak temperature). [Table 22]

[0161] Example 7.1: Characterization of the crystalline forms of the 1:1 tris salt of Compound II (Form B) and the 1:1 citrate salt of Compound II (Form A) The 1:1 tris salt of Compound II (Form B) and the 1:1 citrate salt of Compound II (Form A) were evaluated by dynamic solubility, solid state stability, and solution stability. The free form of Compound II was also evaluated for comparison. Additionally, the hygroscopicity of both salts was evaluated by DVS.

[0162] 7.1 Dynamic solubility The dynamic solubility was evaluated for the 1:1 citrate salt of Compound II (Form A), the 1:1 tris salt of Compound II (Form B), and the free form of Compound II in water and three biologically relevant media (SGF, FaSSIF, and FeSSIF) for 1 / 4 / 24 hours. When 3 - 4 mg of solid was suspended in 3 mL of each medium (solid loading calculated using the free form: 1 mg / mL), a suspension was obtained, which was then rolled (25 rpm) at 37 °C for 1 / 4 / 24 hours. At each time point, approximately 1.0 mL of the suspension was sampled for centrifugation (12000 rpm, 2 minutes) and filtration through a 0.45 μm PTFE membrane to obtain the supernatant for HPLC and pH testing, and the residual solid was analyzed by XRPD. The detailed results are summarized in Table 7 - 1. Based on the solubility evaluation results, the 1:1 citrate salt of Compound II (Form A) and the 1:1 tris salt of Compound II (Form B) showed similar solubility profiles in SGF, FaSSIF, and FeSSIF, which were higher than the solubility profile of the free form within 1 hour. The 1:1 tris salt of Compound II (Form B) showed better solubility in H2O. Since all samples were suspensions, the purity results were only for reference. [Table 23]

[0163] 7.2.1 Solid Stability Evaluation (I / II) To evaluate the physical and chemical stabilities of the 1:1 compound II citrate (form A), 1:1 compound II tris salt (form B), and the free form, samples were stored at 60 °C for 1 day and at 25 °C / 60% RH or 40 °C / 75% RH for 1 week each. The solids were characterized using XRPD and HPLC, and the results were summarized in Table 7-2-1. As demonstrated by the XRPD results and HPLC chromatograms, no morphological changes were observed after 1 week. The 1:1 compound II citrate (form A) and 1:1 compound II tris salt (form B) showed better chemical stability than the free form of compound II.

Table 24

[0164] 7.2.2 Solid Stability Evaluation (II / II) The chemical stabilities of the 1:1 compound II citrate (form A) and 1:1 compound II tris salt (form B) were further evaluated in the solid state, and the samples were stored at ambient conditions or 40 °C / 75% RH for 1 day and 3 days respectively. The HPLC purity and weight purity were tested three times using HPLC, and the results were summarized in Table 7-2-2. The 1:1 compound II citrate (form A) and 1:1 compound II tris salt (form B) showed similar chemical stabilities under the evaluation conditions.

Table 25

[0165] 7.3 Solution Stability Evaluation The solution stability was evaluated at RT for 1:1 compound II citrate (form A) and 1:1 compound II tris salt (form B) in SGF (pH 1.8) and buffer at pH 8.0. Clear solutions were prepared at two concentrations (0.01 mg / mL, 0.05 mg / mL) and then incubated at RT for 6 hours, 1 / 5 days, or 6 hours, 1 / 3 / 7 days. Both salts showed better stability in buffer at pH 8.0, although the purity of the solution at pH 1.8 decreased slightly after 1 day. HPLC purity and pH values were tested and the results are summarized in Tables 7-3-1 to 7-3-4.

Table 26

Table 27

Table 28

Table 29

[0166] 7.4 Hygroscopicity To investigate the stability of the solid forms as a function of humidity, DVS isotherm plots of 1:1 compound II citrate (form A) and 1:1 compound II tris salt (form B) were collected at 25 °C, 0% RH to 95% RH. The results are summarized in Table 7-4. For 1:1 compound II citrate (form A), a water uptake of 0.71% was observed at 25 °C / 80% RH. For 1:1 compound II tris salt (form B), the water uptake increased by 2.47% at 0% to 10% RH in the sorption curve of cycle 2. The water uptake increased by 0.97% at 10% to 80% RH and a plateau was observed. For both salts, no morphological changes were observed after the DVS test.

Table 30

Claims

Claim 1 A tris salt of Compound I, wherein Compound I is represented by the following structural formula, 【Chemical 1】 wherein the molar ratio between Compound I and tris(hydroxymethyl)aminomethane is 1:1, the tris salt. Claim 2 The tris salt according to Claim 1, wherein the tris salt is crystalline. Claim 3 The tris salt according to Claim 1, wherein the tris salt is in a single crystalline form. Claim 4 The tris salt according to Claim 2 or 3, wherein the tris salt is a monohydrate. Claim 5 The tris salt according to Claim 2 or 3, wherein the tris salt is non-solvated. Claim 6 The tris salt according to Claim 4, which is the tris salt form A of Compound I in a single crystalline form, characterized by an X-ray powder diffraction pattern containing at least three peaks selected from 17.5°, 20.1°, 20.7°, 21.1°, and 22.6° ± 0.2 at 2θ. Claim 7 The tris salt according to Claim 4, which is the tris salt form A of Compound I in a single crystalline form, characterized by an X-ray powder diffraction pattern containing peaks at 17.5°, 20.1°, 20.7°, 21.1°, and 22.6° ± 0.2 at 2θ. Claim 8 The tris salt according to Claim 4, which is the tris salt form A of Compound I in a single crystalline form, characterized by an X-ray powder diffraction pattern containing peaks at 4.1°, 14.8°, 17.5°, 18.8°, 20.1°, 20.7°, 21.1°, and 22.6° ± 0.2 at 2θ. Claim 9 The tris salt according to Claim 4, which is the tris salt form A of Compound I in a single crystalline form, characterized by an X-ray powder diffraction pattern containing peaks at 4.1°, 8.1°, 12.8°, 14.8°, 16.3°, 17.5°, 18.8°, 19.3°, 20.1°, 20.7°, 21.1°, 22.6°, 25.1°, and 25.8° ± 0.2 at 2θ. Claim 10 The tris salt according to any one of Claims 6 to 9, which is the tris salt form A of Compound I in a single crystalline form, characterized by a differential scanning calorimetry (DSC) peak phase transition temperature of 173 ± 3°C. Claim 11 The tris salt according to any one of Claims 6 to 10, wherein at least 90% by weight of the tris salt is the tris salt form A of Compound I in a single crystalline form. Claim 12 A tris salt of Compound II, wherein Compound II is represented by the following structural formula, [Chemical 2] A tris salt in which the molar ratio between compound II and tris(hydroxymethyl)aminomethane is 1:

1.

13. The tris salt according to claim 12, wherein the tris salt is crystalline.

14. The tris salt according to claim 13, wherein the tris salt is in a single crystalline form.

15. The tris salt according to claim 13 or 14, wherein the tris salt is a monohydrate.

16. The tris salt according to claim 13 or 14, wherein the tris salt is non-solvated.

17. The tris salt according to claim 14 or 15, wherein the tris salt is in a single crystalline form, Form B, characterized by an X-ray powder diffraction pattern comprising at least three peaks selected from 4.1°, 14.7°, 18.8°, 20.1°, and 23.1° ± 0.2 at 2θ.

18. The tris salt according to claim 14 or 15, wherein the tris salt is in a single crystalline form, Form B, characterized by an X-ray powder diffraction pattern comprising peaks at 4.1°, 14.7°, 18.8°, 20.1°, and 23.1° ± 0.2 at 2θ.

19. The tris salt according to claim 14 or 15, wherein the tris salt is in a single crystalline form, Form B, characterized by an X-ray powder diffraction pattern comprising peaks at 4.1°, 4.1°, 8.2°, 14.7°, 16.4°, 18.8°, 20.1°, 20.7°, 21.3°, and 23.1° ± 0.2 at 2θ.

20. The tris salt according to claim 14 or 15, wherein the tris salt is in a single crystalline form, Form B, characterized by an X-ray powder diffraction pattern comprising peaks at 4.1°, 8.2°, 14.7°, 16.4°, 18.8°, 19.1°, 20.1°, 20.7°, 21.3°, 23.1°, 24.1°, and 25.4° ± 0.2 at 2θ.

21. The tris salt according to any one of claims 17 - 20, wherein the tris salt is in a single crystalline form, Form B, characterized by a differential scanning calorimetry (DSC) peak phase transition temperature of 168 ± 4°C.

22. The tris salt according to any one of claims 17 - 21, wherein at least 90% by weight of the tris salt is in a single crystalline Form B.

23. The tris salt according to claim 14, wherein the tris salt is in a single crystalline form, Form G, characterized by an X-ray powder diffraction pattern comprising at least three peaks selected from 6.2°, 7.6°, 13.1°, 13.4°, and 18.5° ± 0.2 at 2θ.

24. The tris salt according to claim 14, which is in the single crystalline form, Form G, characterized by an X-ray powder diffraction pattern including peaks at 2θ of 6.2°, 7.6°, 13.1°, 13.4°, and 18.5° ± 0.

2.

25. The tris salt according to claim 14, which is in the single crystalline form, Form G, characterized by an X-ray powder diffraction pattern including peaks at 2θ of 6.2°, 7.6°, 13.1°, 13.4°, 18.5°, 21.5°, 23.7°, and 24.1° ± 0.

2.

26. The tris salt according to claim 14, which is in the single crystalline form, Form G, characterized by an X-ray powder diffraction pattern including peaks at 2θ of 6.2°, 7.6°, 13.1°, 13.4°, 18.0°, 18.5°, 20.8°, 21.5°, 23.7°, and 24.1° ± 0.

2.

27. The tris salt according to any one of claims 23 to 26, which is in the single crystalline form, Form G, characterized by a differential scanning calorimetry (DSC) peak phase transition temperature of 129.5 ± 4°C.

28. The tris salt according to any one of claims 23 to 27, wherein at least 90% by weight of the tris salt is in the single crystalline form G.

29. A citrate salt of Compound II, wherein Compound II is represented by the following structural formula, [Chemical Formula 3] wherein the molar ratio between Compound II and citric acid is 1:

1.

30. The citrate salt according to claim 29, which is crystalline.

31. The citrate salt according to claim 30, which is in the single crystalline form.

32. The citrate salt according to claim 31, which is in the single crystalline form, Form A of the citrate salt of Compound II, characterized by an X-ray powder diffraction pattern including at least three peaks selected from 5.4°, 9.4°, 12.4°, 14.3°, and 17.8° ± 0.2 at 2θ.

33. The citrate salt according to claim 31, which is in the single crystalline form, Form A of the citrate salt of Compound II, characterized by an X-ray powder diffraction pattern including peaks at 2θ of 5.4°, 9.4°, 12.4°, 14.3°, and 17.8° ± 0.

2.

34. The citrate is the citrate form A of compound II in a single crystalline form, characterized by an X-ray powder diffraction pattern having peaks at 2θ of 5.4°, 9.4°, 10.8°, 12.4°, 14.3°, 16.2°, 17.8°, 19.6°, and 24.9° ± 0.2, the citrate according to claim 31.

35. The citrate is the citrate form A of compound II in a single crystalline form, characterized by an X-ray powder diffraction pattern having peaks at 2θ of 5.4°, 9.4°, 10.8°, 12.4°, 14.3°, 16.2°, 17.8°, 18.8°, 19.6°, 23.6°, and 24.9° ± 0.2, the citrate according to claim 31.

36. The citrate is the citrate form A of compound II in a single crystalline form, characterized by a differential scanning calorimetry (DSC) peak phase transition temperature of 170 ± 3°C, the citrate according to any one of claims 32 to 35.

37. At least 90% by weight of the citrate is the citrate form A of compound II in a single crystalline form, the citrate according to any one of claims 32 to 36.

38. A pharmaceutical composition comprising the compound according to any one of claims 1 to 37 and a pharmaceutically acceptable carrier.

39. A pharmaceutical composition for treating cardiovascular metabolism and related diseases, wherein the pharmaceutical composition comprises a therapeutically effective amount of the salt according to any one of claims 1 to 37, and the diseases include type 1 diabetes, type 2 diabetes, prediabetes, idiopathic type 1 diabetes, latent autoimmune diabetes in adults, early-onset type 2 diabetes, youth-onset atypical diabetes, youth-onset adult-onset diabetes, malnutrition-related diabetes, gestational diabetes, hyperglycemia, insulin resistance, hepatic insulin resistance, glucose intolerance, diabetic neuropathy, diabetic nephropathy, kidney disease, diabetic retinopathy, adipocyte dysfunction, sleep apnea, obesity, eating disorder, dyslipidemia, hyperinsulinemia, non-alcoholic fatty liver disease, non-alcoholic steatohepatitis, fibrosis, cirrhosis, hepatocellular carcinoma, cardiovascular disease, atherosclerosis, coronary artery disease, peripheral vascular disease, hypertension, endothelial disorder, vascular compliance disorder, congestive heart failure, myocardial infarction, stroke, hemorrhagic stroke, ischemic stroke, traumatic brain injury, pulmonary hypertension, restenosis after angioplasty, intermittent claudication, postprandial lipemia, metabolic acidosis, ketosis, arthritis, osteoporosis, Parkinson's disease, left ventricular hypertrophy, peripheral arterial disease, macular degeneration, cataract, glomerulosclerosis, chronic renal failure, metabolic syndrome, syndrome X, premenstrual syndrome, angina pectoris, thrombosis, atherosclerosis, transient ischemic attack, vascular restenosis, glucose metabolism disorder, hyperuricemia, gout, erectile dysfunction, skin and connective tissue disorder, psoriasis, foot ulcer, ulcerative colitis, hyperapo B lipoproteinemia, Alzheimer's disease, schizophrenia, cognitive impairment, inflammatory bowel disease, short bowel syndrome, Crohn's disease, colitis, irritable bowel syndrome, polycystic ovary syndrome prevention or treatment, and poisoning treatment, the said pharmaceutical composition.

40. The pharmaceutical composition according to claim 39, wherein the diseases are Alzheimer's disease, type 1 diabetes, type 2 diabetes, hyperglycemia, non-alcoholic steatohepatitis, obesity, non-alcoholic fatty liver disease, or Parkinson's disease, the said pharmaceutical composition.

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