Crystalline form i of melanocortin receptor agonist compound and method for preparing same

Abstract

To provide a stable crystalline form of a novel compound having excellent selective agonistic activity for melanocortin receptors, particularly melanocortin-4 receptor (MC4R), and a method for producing the same.SOLUTION: The present invention relates to crystalline form I of a compound represented by chemical formula 1, a pharmaceutically acceptable salt thereof, or a solvate thereof, the crystalline form I having three or more characteristic peaks selected from the following diffraction angles (2θ values) in an X-ray powder diffraction (XRPD) pattern: 8.240±0.2°, 9.363±0.2°, 10.2693±0.2°, 10.5969±0.2°, 12.050±0.2°, 12.841±0.2°, 13.503±0.2°, 15.5738±0.2°, 16.6030±0.2°, 17.009±0.2°, 17.305±0.2°, 18.364±0.2°, 18.7390±0.2°, 19.188±0.2°, 19.476±0.2°, 20.001±0.2°, 20.477±0.2°, 20.665±0.2°, 21.348±0.2°, 21.976±0.2°, 22.580±0.2°, 23.896±0.2°, 4.334±0.2°, 24.812±0.2°, 25.243±0.2°, 25.833±0.2°, 26.646±0.2°, 27.82±0.2°, 28.316±0.2°, 28.609±0.2°, 29.692±0.2°, 30.185±0.2°, and 30.875±0.2°.SELECTED DRAWING: None

Description

[Technical Field]

[0001] This application claims the benefit of priority based on Korean Patent Application No. 10-2020-0180807, filed December 22, 2020, and all contents disclosed in the documents of this Korean patent application are incorporated herein by reference.

[0002] The present invention relates to crystalline form I of a novel compound that exhibits excellent enhancing activity against melanocortin receptors, a method for preparing the same, and a pharmaceutical composition containing the same. [Background technology]

[0003] Leptin is a hormone secreted by adipocytes. Its secretion level increases with increasing body fat content. It regulates various physiological functions, including appetite, body fat content, and energy metabolism, by regulating the function of various neuropeptides produced in the hypothalamus (Schwartz et al., Nature 404, 661-671 (2000)). Leptin signaling, which regulates appetite and body weight, is mediated by various downstream factors, most notably melanocortin, agouti-related peptide (AgRP), and neuropeptide Y (NPY).

[0004] When blood leptin levels increase as a result of a calorie surplus in the body, the pituitary gland increases secretion of the protein hormone proopiomelanocortin (POMC) and decreases production of AgRP and NPY. POMC neurons produce the small peptide hormone alpha-MSH (melanocyte stimulating hormone), which acts as an agonist to the melanocortin-4 receptor (MC4R) in second-order neurons, thereby inducing appetite reduction. On the other hand, when leptin levels decrease as a result of a calorie deficit, expression of AgRP, an MC4R antagonist, and NPY increase, resulting in increased appetite. Thus, in response to changes in leptin, alpha-MSH and AgRP hormones mediate appetite regulation by acting as agonists and antagonists to MC4R.

[0005] Alpha-MSH hormones bind to three MCR subtypes in addition to MC4R, inducing various physiological responses. Five MCR subtypes have been identified to date. MC1R is primarily expressed in skin cells and is involved in skin pigmentation. MC2R is primarily expressed in the adrenal gland and is known to be involved in the production of glucocorticoid hormones, with ACTH (adrenocorticotropic hormone) derived from POMC as its only ligand. MC3R and MC4R, primarily expressed in the central nervous system, are involved in regulating appetite, energy metabolism, and fat storage efficiency. MC5R, expressed in various tissues, is known to regulate exocrine function (Wikberg, et al., Pharm Res 42(5)393-420(2000)). In particular, activation of the MC4R receptor has been proven to be a major target for the development of anti-obesity drugs, as it effectively reduces body weight by reducing appetite and increasing energy metabolism (Review: Wikberg, Eur. J. Pharmacol 375, 295-310 (1999)); Wikberg, et al., Pharm Res 42(5) 393-420 (2000); Douglas et al., Eur J Pharm 450, 93-109 (2002); O'Rahilly et al., Nature Med 10, 351-352 (2004)).

[0006] The role of MC4R in appetite and body weight regulation was first demonstrated by experiments using agouti mice, an animal model in which agouti protein is overexpressed. In agouti mice, a genetic mutation causes high levels of agouti protein to be expressed in the central nervous system, where it acts as an MC4R antagonist in the hypothalamus, inducing obesity (Yen, TT et al., FASEB J. 8, 479-488 (1994); Lu D., et al., Nature 371, 799-802 (1994)). Subsequent research has shown that the hypothalamic neurons do indeed express AgRP (agouti-related peptide), which is similar to agouti protein, and this is also known to be involved in appetite regulation as an antagonist of MC4R (Shutter, et al., Genes Dev., 11, 593-602 (1997); Ollman, et al., Science 278, 135-138 (1997)).

[0007] When alpha-MSH, an in vivo MC4R agonist, is administered intracerebrally to animals, it has been shown to have an appetite-reducing effect, whereas when treated with the MC4R antagonists SHU9119 (peptide) or HS014 (peptide), it has been observed to increase appetite again (Kask et al., Biochem. Biophys. Res. Comm. 245, 90-93 (1998)). Furthermore, animal studies using Melanotan II (MTII, Ac-Nle-c[Asp-His-DPhe-Arg-Trp-Lys]-NH2) and its analogous agonist HP228 have shown that cerebral, intraperitoneal, or subcutaneous administration suppresses appetite, reduces body weight, and increases energy metabolism (Thiele TE, et al. Am J Physiol 274(1 Pt 2), R248-54(1998); Lee MD, et al. FASEB J 12, A552(1998); Murphy B., et al. J Appl Physiol 89, 273-82(2000)). Conversely, administration of the representative compound SHU9119 to animals resulted in significant and sustained increases in food intake and body weight, providing pharmacological evidence that MCR agonists can be used as therapeutic agents for obesity. The significant appetite-reducing effect observed upon MTII administration was not observed in MC4R KO (knock-out) mice, and these experimental results also demonstrate that the appetite-reducing effect is primarily mediated by activation of MC4R (Marsh, et al., Nat Genet 21, 119-122 (1999)).

[0008] The main type of obesity treatments developed to date are appetite suppressants that act on the central nervous system, most of which are drugs that modulate the activity of neurotransmitters. Examples include the noradrenaline agents phentermine and mazindol, and the serotonergic agents fluoxetine and sibutramine. However, these neurotransmitter modulators exert a wide range of effects on various physiological functions in addition to appetite suppression due to their numerous receptor subtypes. Therefore, these modulators have the major drawback of lacking selectivity for each subtype and causing various side effects when administered for a long period of time.

[0009] On the other hand, melanocortin agonists are neuropeptides, not neurotransmitters, and have the advantage of being able to induce weight loss by suppressing appetite without affecting other physiological functions, given that all functions except energy metabolism are normal in MC4R gene knockout mice. In particular, the receptor used is a G-protein coupled receptor (GPCR), which is one of the most successful receptors for new drugs developed to date, making it relatively easy to ensure selectivity for subtype receptors, which sets them apart from existing receptors.

[0010] As examples of utilizing such melanocortin receptors as a site of action, International Publication Nos. WO2008 / 007930 and WO2010 / 056022 disclose compounds that act as melanocortin receptor agonists.

[0011] Furthermore, the inventors of the present invention have conducted extensive research and invented a novel compound represented by the following Chemical Formula 1, which has excellent selective enhancing activity against melanocortin receptors, particularly against melanocortin-4 receptors (MC4R), and a method for producing the same (Korean Patent Application No. 10-2019-0141649 (filed November 7, 2019)).

[0012] [ka] (R1 is C2-C5 alkyl.)

[0013] On the other hand, the crystalline structure of a pharmaceutically active ingredient can sometimes affect the chemical stability of the drug. Different crystallization and storage conditions can change the crystalline structure of the compound, sometimes resulting in the production of different crystalline forms. Generally, amorphous drug products do not have a regular crystalline structure and sometimes have other defects such as poor product stability, small particle size, difficult filtration, tendency to agglomeration, and poor flowability. Therefore, it is necessary to improve various physical properties of the product. Thus, it is necessary to research a crystalline structure with high purity and good chemical stability for a compound. [Prior art documents] [Patent documents]

[0014] [Patent Document 1] International Patent Application Publication No. WO2008 / 007930 [Patent Document 2] International Patent Application Publication No. WO2010 / 056022 Summary of the Invention [Problem to be solved by the invention]

[0015] An object of the present invention is to provide a stable crystalline form of a novel compound that has excellent selective stimulating activity for melanocortin receptors, particularly melanocortin-4 receptor (MC4R), and a method for producing the same.

[0016] Another object of the present invention is to provide a pharmaceutical composition containing a stable crystalline form of said novel compound. [Means for solving the problem]

[0017] To achieve the above objectives, In one aspect, the present invention provides Crystalline Form I of a compound of Formula 1, a pharmaceutically acceptable salt thereof, or a solvate thereof: The X-ray powder diffraction (XRPD) pattern showed the following diffraction angles (2θ values): 8.240±0.2°, 9.363±0.2°, 10.2693±0.2°, 10.5969±0.2°, 12.050±0.2°, 12.841±0.2°, 13.503±0.2°, 15.5738±0.2°, 16.6030±0.2°, 17.009±0.2°, 17.305±0.2°, 18.364±0.2°, 18.7390±0.2°, 19.188±0.2°, 19.476±0.2°, 20.001±0.2°, 20.477 and 30.875±0.2°.

[0018] [ka]

[0019] In the above Chemical Formula 1, R1 is C2-C5 alkyl.

[0020] The compounds of Formula 1 may have asymmetric carbon centers and asymmetric axes or planes and may exist as cis or trans isomers, R or S isomers, racemates, partial stereoisomeric mixtures, and individual partial stereoisomers, all of which isomers and mixtures are included within the scope of the compounds of Formula 1.

[0021] For convenience, in this specification, unless otherwise specified, the compound of Chemical Formula 1 is used to mean the compound of Chemical Formula 1, its pharmaceutically acceptable salts, its isomers and solvates.

[0022] In one embodiment according to the present invention, R1 in Formula 1 is C2-C5 alkyl. In another embodiment according to the present invention, R1 in Formula 1 is linear or branched C2-C5 alkyl, for example, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, or tert-butyl.

[0023] In another embodiment according to the present invention, R1 in Chemical Formula 1 is C2-C4 alkyl. In another embodiment according to the present invention, R1 in Chemical Formula 1 is linear or branched C2-C4 alkyl, for example, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, or tert-butyl. Specifically, R1 may be isopropyl.

[0024] In one embodiment according to the present invention, the pharmaceutically acceptable salts include, but are not limited to, acid addition salts formed with inorganic acids such as hydrochloric acid, sulfuric acid, nitric acid, phosphoric acid, hydrobromic acid, hydroiodic acid, etc.; organic carboxylic acids such as tartaric acid, formic acid, citric acid, acetic acid, trichloroacetic acid, trifluoroacetic acid, gluconic acid, benzoic acid, lactic acid, fumaric acid, maleic acid, etc.; sulfonic acids such as methanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid, or naphthalenesulfonic acid, etc.

[0025] In one embodiment according to the present invention, the solvate may include hydrates; solvates with organic solvents such as methanol, ethanol, 2-propanol, 1,2-propanediol, 1,3-propanediol, n-butanol, 1,4-butanediol, tert-butanol, acetic acid, acetone, butyl acetate, methyl acetate, ethyl acetate, propyl acetate, t-butyl acetate, isobutyl acetate, methyl ethyl ketone, 2-pentanone, tetrahydrofuran, acetonitrile, chloroform, toluene, and mixtures thereof.

[0026] In one embodiment according to the present invention, crystalline Form I has an X-ray powder diffraction pattern of 8.240±0.2°, 9.363±0.2°, 10.2693±0.2°, 10.5969±0.2°, 12.050±0.2°, 12.841±0.2°, 13.503±0.2°, 15.5738±0.2°, 16. 6030±0.2°, 17.009±0.2°, 17.305±0.2°, 18.364±0.2°, 18.7390±0.2°, 19.188±0.2°, 19.476±0.2°, 20.001±0.2°, 20.477±0.2°, 20.665±0.2°, 21.348±0.2°, 21 The chromatic aberration may have 3 or more, 5 or more, 7 or more, 9 or more, 10 or more, 13 or more, 15 or more, 17 or more, 20 or more, 23 or more, 25 or more, 27 or more, or 30 or more characteristic peaks selected from 0.976±0.2°, 22.580±0.2°, 23.896±0.2°, 4.334±0.2°, 24.812±0.2°, 25.243±0.2°, 25.833±0.2°, 26.646±0.2°, 27.82±0.2°, 28.316±0.2°, 28.609±0.2°, 29.692±0.2°, 30.185±0.2°, and 30.875±0.2°.

[0027] In another embodiment according to the present invention, crystalline Form I has an X-ray powder diffraction pattern of 8.240±0.2°, 9.363±0.2°, 10.2693±0.2°, 10.5969±0.2°, 12.050±0.2°, 12.841±0.2°, 13.503±0.2°, 15.5738±0.2°, 16.6030±0.2°, 17.009±0.2°, 17.305±0.2°, 18.364±0.2°, 18.7390±0.2°, 19.188±0.2°, 19.476±0.2°, 20.001±0.2°. 0.2°, 20.477±0.2°, 20.665±0.2°, 21.348±0.2°, 21.976±0.2°, 22.580±0.2°, 23.896±0.2°, 4.334±0.2°, 24.812±0.2°, 25.243±0.2°, 25.833±0.2°, 26.646±0.2°, 27.82±0.2°, 28.316±0.2°, 28.609±0.2°, 29.692±0.2°, 30.185±0.2° and 30.875±0.2°.

[0028] In one embodiment according to the present invention, the crystalline form I may have the X-ray powder diffraction (XRPD) pattern shown in FIG.

[0029] The crystalline form I according to the present invention may exhibit an endothermic peak at 155-165° C. in a differential scanning calorimetry (DSC) profile. The melting endotherm may begin at an onset temperature of 159.14° C.

[0030] In one embodiment according to the present invention, the crystalline form I may have a DSC profile as shown in FIG.

[0031] Crystalline Form I according to the present invention may exhibit no observable weight loss in a thermogravimetric analysis (TGA) profile when heated at a temperature of 270° C. or less. The term "no observable weight loss" may include, for example, a weight loss of 5% or less, 4% or less, 3% or less, 2% or less, 1% or less, or 0% (i.e., no weight loss at all).

[0032] In one embodiment according to the present invention, the crystalline form I may have a TGA profile as shown in FIG.

[0033] Crystalline Form I according to the present invention may exhibit a total weight gain observed at 0-90% RH of 5% w / w or less, e.g., 4% w / w or less, 3% w / w or less, 2% w / w or less, 1% w / w or less, 0.5% w / w or less, or 0.2% w / w, as a result of moisture sorption analysis (DVS).

[0034] In one embodiment according to the present invention, the crystalline form I may have a DVS profile as shown in FIG.

[0035] The stability test results (HPLC) of the crystalline form I of the present invention showed that it was chemically stable for 4 weeks under accelerated conditions (40°C, 75% RH) and harsh conditions (80°C), indicating that it is a crystalline form that is stable to heat and humidity.

[0036] In this specification, X-ray powder diffraction (XRPD) analysis was performed using a PANalytical X' Pert Pro MPD system, Malvern Panalytical Ltd.

[0037] Differential scanning calorimetry (DSC) analysis was performed using a DSC1, Mettler-Toledo AG.

[0038] Thermogravimetric analysis (TGA) was performed using a TGA / DSC 1, Mettler-Toledo AG.

[0039] The results of moisture sorption analysis (DVS) were obtained using an automated gravimetric vapor sorption analyzer equipped with a Cahn D200 ultra-microbalance (DVS Intrinsic apparatus, Surface Measurement Systems Ltd., UK).

[0040] The stability analysis was performed using HPLC (Agilent Technologies, Inc.) and the results shown are shown below.

[0041] Crystalline Form I may have higher purity and be more physically and chemically stable than the crude compound of Formula 1, the amorphous compound of Formula 1, or other crystalline forms of the compound of Formula 1.

[0042] Furthermore, Crystal Form I of the compound of Chemical Formula 1 may have superior melanocortin-4 receptor activating ability and preventive or therapeutic effects against diseases such as obesity, diabetes, inflammation, and erectile dysfunction compared to known melanocortin-4 receptor agonists, but the effects of the present invention are not limited thereto.

[0043] In another aspect, the present invention provides a method for preparing crystalline Form I, comprising the steps of dissolving the compound of Formula 1 in a crystallization solvent to prepare a mixed solution, and obtaining crystals from the mixed solution.

[0044] First, the compound represented by the formula 1 is dissolved in a crystallization solvent.

[0045] The compound of Chemical Formula 1 for preparing Crystalline Form I can be the compound of Chemical Formula 1, a salt thereof, an isomer thereof, or a solvate thereof.

[0046] The compound of Chemical Formula 1 can be obtained by the manufacturing method described in the specification of Korean Patent Application No. 10-2019-0141649 (filed on November 7, 2019).

[0047] The crystallization solvent is not particularly limited as long as it is a solvent suitable for crystallization of a compound. In one embodiment, the crystallization solvent includes an organic solvent.

[0048] The organic solvent may include dialkyl ether solvents such as diethyl ether, dipropyl ether, dibutyl ether, diisoamyl ether, ethyl methyl ether, methyl propyl ether, methyl butyl ether, and ethyl propyl ether; cyclic ether solvents such as tetrahydrofuran and tetrahydropyran; aromatic ring-containing ether solvents such as diphenyl ether and anisole; amide solvents such as dimethylacetamide; ketone solvents such as methyl isobutyl ketone; ester solvents such as isopropyl acetate; alcohol solvents such as 1-pentyl alcohol; toluene solvents such as toluene; or mixtures thereof.

[0049] In one embodiment according to the present invention, the organic solvent may be an ether solvent.

[0050] In another embodiment according to the present invention, the organic solvent can include methyl tert-butyl ether (MTBE).

[0051] In one embodiment, the crystallization solvent may be used in an amount that completely dissolves the compound of Formula 1.

[0052] In one specific example, the crystallization solvent can be used in an amount of 0.5 to 10 mL, 0.5 to 5 mL, 0.8 to 3 mL, 1.0 to 2.5 mL, 1.5 to 2 mL, 1.6 to 1.8 mL, or 1.625 mL per 1 g of the compound of Chemical Formula 1.

[0053] The dissolution of the crude compound of Formula 1 in the solvent can be carried out at a temperature of 15 to 30°C, specifically 23 to 28°C, without stirring or with stirring.

[0054] In one embodiment according to the present invention, 0.6 g of the compound of Chemical Formula 1 is dissolved in 0.975 mL of MTBE at 25° C. to obtain a mixed solution in which the compound of Chemical Formula 1 is dissolved.

[0055] Next, crystals are obtained from the mixed solution in which the compound of Formula 1 is dissolved.

[0056] The crystals can be obtained, for example, by cooling the solution, evaporating the solvent, adding an antisolvent to supersaturate, or using methods such as slurry conversion.

[0057] The method may further include at least one of stirring and filtering the mixed solution in any order.

[0058] In one embodiment according to the present invention, the mixed solution is cooled, stirred, and filtered to obtain crystals.

[0059] The cooling may be performed so that the temperature of the mixed solution to which the acid has been added dropwise is 0° C. to 10° C. Specifically, the cooling may be performed so that the temperature of the mixed solution is 0° C. to 5° C., or 3° C.

[0060] The stirring can be carried out for, but is not limited to, for example, 1 hour to 72 hours, 10 hours to 48 hours, 15 hours to 36 hours, 20 hours to 24 hours, or 21 hours.

[0061] The preparation method of the present invention may further include a step of adding a non-polar organic solvent at any step of the preparation method of Crystal Form I. In one embodiment, the method may further include a step of adding a non-polar organic solvent to the solution before preparing the mixed solution, during preparing the mixed solution, after preparing the mixed solution, before cooling, after cooling, or simultaneously with cooling, or before stirring, after stirring, or simultaneously with stirring the cooled mixed solution. Adding the non-polar organic solvent can increase the rate of crystallization particle formation, thereby improving the yield or production stability of the obtained Crystal Form I, but the present invention is not limited thereto.

[0062] The non-polar organic solvent can be used without any particular limitation as long as it is an organic solvent having non-polar properties, and examples thereof include hexane, heptane, cyclohexane, carbon tetrachloride, benzene, and chloroform.

[0063] The thus obtained crystalline form I may have higher purity and be more physically and chemically stable than the crude compound of Chemical Formula 1, the amorphous compound of Chemical Formula 1, or any crystalline form of Chemical Formula 1, but the effects of the present invention are not limited thereto.

[0064] In yet another aspect, the present invention provides a pharmaceutical composition comprising: (i) said crystalline Form I; and (ii) a pharmaceutically acceptable carrier.

[0065] Since crystalline form I of the present invention exhibits excellent enhancing activity on melanocortin receptors, particularly the melanocortin-4 receptor (MC4R), the present invention can also provide a pharmaceutical composition for enhancing melanocortin receptor function, which contains as an active ingredient crystalline form I. Specifically, the pharmaceutical composition can be a composition for enhancing melanocortin-4 receptor function.

[0066] Furthermore, the pharmaceutical composition may exhibit excellent effects in the prevention or treatment of obesity, diabetes, inflammation, and erectile dysfunction, and may be a composition for the prevention or treatment of obesity, diabetes, inflammation, or erectile dysfunction, but the use of the present invention is not limited to these diseases.

[0067] As used herein, the term "carrier" refers to a compound that facilitates the introduction of a compound into cells or tissues.

[0068] When crystalline Form I of the present invention is administered for clinical purposes, the total daily dose administered to a host in a single dose or in separate doses is preferably in the range of 0.01 to 10 mg / kg body weight, but the specific dose level for an individual patient may vary depending on the particular compound used, the patient's body weight, sex, health condition, diet, time of drug administration, administration method, excretion rate, drug mixture, and severity of disease, etc.

[0069] The crystalline Form I of the present invention can be administered by any route depending on the purpose. For example, the amorphous compound of the present invention can be administered by injection or orally.

[0070] The pharmaceutical compositions of the present invention can be in various oral administration forms such as tablets, pills, powders, capsules, granules, syrups or emulsions, or in parenteral administration forms such as injectable preparations for intramuscular, intravenous or subcutaneous administration.

[0071] Injectable preparations can be prepared using suitable dispersing agents, wetting agents, suspending agents, or excipients according to known techniques.

[0072] Excipients that can be used in the pharmaceutical formulation of the present invention may include, but are not limited to, sweeteners, binders, solubilizers, solubilizers, wetting agents, emulsifiers, isotonicity agents, adsorbents, disintegrants, antioxidants, preservatives, lubricants, fillers, flavorings, etc. For example, excipients that can be used include lactose, dextrose, sucrose, mannitol, sorbitol, cellulose, glycine, silica, magnesium aluminum silicate, starch, gelatin, tragacanth gum, alginic acid, sodium alginate, methylcellulose, sodium carboxymethylcellulose, water, ethanol, polyethylene glycol, polyvinylpyrrolidone, sodium chloride, calcium chloride, orange essence, strawberry essence, vanilla flavor, etc.

[0073] When the pharmaceutical composition of the present invention is in the form of oral administration, examples of the carrier to be used include, but are not limited to, cellulose, calcium silicate, corn starch, lactose, sucrose, dextrose, calcium phosphate, stearic acid, magnesium stearate, calcium stearate, gelatin, talc, etc.

[0074] When the pharmaceutical composition of the present invention is in the form of an injectable preparation, the carrier may include, but is not limited to, water, saline, aqueous glucose solution, aqueous saccharide solution, alcohol, glycol, ether, oil, fatty acid, fatty acid ester, glyceride, etc.

[0075] In yet another aspect, there is provided crystalline Form I as described above for use in enhancing the activity of melanocortin receptors, particularly melanocortin-4 receptors (MC4R).

[0076] In one embodiment, there is provided crystalline Form I as described above for use in the treatment or prevention of obesity, diabetes, inflammation or erectile dysfunction.

[0077] In yet another aspect, there is provided a method for enhancing the activity of melanocortin receptors, particularly melanocortin-4 receptors (MC4R), comprising the step of administering crystalline Form I described above to a subject.

[0078] In yet another aspect, there is provided a method of treating obesity, diabetes, inflammation or erectile dysfunction comprising administering to a subject crystalline Form I described above. [Effects of the Invention]

[0079] Crystalline Form I of the present invention exhibits excellent enhancing activity on melanocortin receptors, particularly melanocortin-4 receptor (MC4R), and can therefore be usefully used in the prevention or treatment of obesity, diabetes, inflammation, and erectile dysfunction.

[0080] Crystalline Form I of the present invention exhibits an on-target effect on the melanocortin-4 receptor, and therefore exhibits weight loss and food intake reduction effects, does not affect anxiety and depression, and can be administered without safety issues such as side effects and mutation induction due to hERG (human ether-a-go-go related gene) inhibition.

[0081] Furthermore, crystalline Form I according to the present invention has superior purity, yield, physical and chemical stability compared to the crude compound of Formula 1, the amorphous compound of Formula 1, or any other crystalline form of Formula 1.

[0082] Specifically, the crystalline form I may have superior solubility, storage stability, and production stability compared to the compound of Chemical Formula 1, the amorphous compound of Chemical Formula 1, or any other crystalline form of Chemical Formula 1. [Brief explanation of the drawings]

[0083] [Figure 1] 1 is a graph showing the XRPD results of Example 1. [Figure 2] 1 is a graph showing the results of DSC in Example 1. [Figure 3] 1 is a graph showing the TGA results of Example 1. [Figure 4] 1 is a graph showing the DVS results of Example 1. DETAILED DESCRIPTION OF THE INVENTION

[0084] The present invention will be described in more detail below with reference to Production Examples and Examples, although these Examples are merely illustrative of the present invention and are not intended to limit the scope of the present invention.

[0085] Preparation Example 1: Preparation of methyl (2S,4S)-4-(N-((1s,4R)-4-methylcyclohexyl)isobutyramido)pyrrolidine-2-carboxylate hydrochloride [ka]

[0086] The title compound was obtained through the following steps A, B, C, D and E.

[0087] Step A: Preparation of 1-(tert-butyl) 2-methyl (2S,4S)-4-azidopyrrolidine-1,2-dicarboxylate Under nitrogen, 1-(tert-butyl) 2-methyl(2S,4R)-4-((methylsulfonyl)oxy)pyrrolidine-1,2-dicarboxylate (48.5 g, 150 mmol) was dissolved in N,N'-dimethylformamide (250 mL) and sodium azide (19.5 g, 300 mL) was added. The mixture was stirred at 80°C for 16 hours, and the reaction solvent was concentrated under reduced pressure. Water was added, and the mixture was extracted twice with ethyl acetate. The organic layer was washed with aqueous sodium chloride and water, dried over anhydrous magnesium sulfate, and filtered. The filtrate was concentrated under reduced pressure to give crude 1-(tert-butyl) 2-methyl(2S,4S)-4-azidopyrrolidine-1,2-dicarboxylate (39.59 g, 98%), which was used in the next step without purification.

[0088] MS [M+H] = 271 (M+1) 1 H NMR (400 MHz, CD3OD) δ 4.43-4.37 (m, 1H), 4.35-4.27 (br, 1H), 3.77 (s, 1.8H), 3.76 (s, 1.2H), 3.73-3.66 (m, 1H), 3.44-3.38 (m, 1H), 2.63-2.49 (m, 1H), 2.19-2.11 (m, 1H), 1.50 (s, 4.5H), 1.44 (s, 4.5H)

[0089] Step B: Preparation of 1-(tert-butyl) 2-methyl (2S,4S)-4-aminopyrrolidine-1,2-dicarboxylate 1-(tert-butyl) 2-methyl(2S,4S)-4-azidopyrrolidine-1,2-dicarboxylate (24.59 g, 91.0 mmol) obtained in Step A was dissolved in tetrahydrofuran (180 mL), and 1 M trimethylphosphine tetrahydrofuran solution (109.2 mL, 109.2 mmol) was slowly added at 0°C. The mixture was stirred at the same temperature for 1 hour and then at room temperature for 3 hours. The reaction solvent was concentrated under reduced pressure, and dichloromethane (100 mL) and water (150 mL) were added and stirred for approximately 30 minutes. The layers were separated and extracted again with dichloromethane. The organic layer was dried over anhydrous magnesium sulfate and filtered. The filtrate was concentrated under reduced pressure to obtain crude 1-(tert-butyl) 2-methyl(2S,4S)-4-aminopyrrolidine-1,2-dicarboxylate (20.62 g, 93%), which was used in the next step without purification.

[0090] MS [M+H] = 245 (M+1) 1H NMR (400 MHz, CD3OD) δ 4.27 (m, 1H), 3.77 (s, 1.8H), 3.76 (s,1.2H), 3.75-3.67 (m, 1H), 3.50-3.42 (m, 1H), 3.22-3.17 (m, 1H), 2.58-2.47 (m,1H), 1.82-1.71 (m, 1H), 1.48 (s, 4.5H), 1.42 (s, 4.5H)

[0091] Step C: Preparation of 1-(tert-butyl) 2-methyl(2S,4S)-4-(((1s,4R)-4-methylcyclohexyl)amino)pyrrolidine-1,2-dicarboxylate 1-(tert-butyl) 2-methyl(2S,4S)-4-aminopyrrolidine-1,2-dicarboxylate (20.62 g, 84.4 mmol) obtained in Step B was dissolved in dichloroethane (150 mL) and 4-methylcyclohexanone (9.5 mL, 101.3 mmol) was added. Sodium triacetoxyborohydride (26.8 g, 126.6 mmol) was added at 0°C, and the mixture was stirred at room temperature for 16 hours. The reaction solvent was concentrated under reduced pressure, water was added, and the mixture was extracted twice with ethyl acetate. The organic layer was washed with aqueous sodium chloride solution, dried over anhydrous magnesium sulfate, and filtered. The filtrate was concentrated under reduced pressure and purified by column chromatography to obtain 1-(tert-butyl) 2-methyl(2S,4S)-4-(((1S,4R)-4-methylcyclohexyl)amino)pyrrolidine-1,2-dicarboxylate (22.9 g, 80%).

[0092] MS [M+H] = 341 (M+1) 1H NMR (400 MHz, CD3OD) δ 4.26 (m, 1H), 3.76 (s, 1.8H), 3.75 (s, 1.2H), 3.78-3.71 (m, 1H), 3.49-3.40 (m, 1H), 3.22-3.16 (m, 1H), 2.69-2.60(br, 1H), 2.58-2.46 (m, 1H), 1.87-1.77 (m, 1H), 1.73-1.63 (m, 1H), 1.62-1.35(m, 8H), 1.48 (s, 4.5H), 1.42 (s, 4.5H), 0.96 (d, 3H)

[0093] Step D: Preparation of 1-(tert-butyl) 2-methyl(2S,4S)-4-(N-((1s,4R)-4-methylcyclohexyl)isobutyramido)pyrrolidine-1,2-dicarboxylate 1-(tert-butyl) 2-methyl(2S,4S)-4-(((1s,4R)-4-methylcyclohexyl)amino)pyrrolidine-1,2-dicarboxylate (37.29 g, 109.5 mmol) obtained in Step C was dissolved in dichloromethane (500 ml), triethylamine (61.1 ml, 438.1 mmol) was added, and isobutyryl chloride (11.7 ml, 219 mmol) was slowly added at 0°C. After stirring at room temperature for 16 hours, the reaction solvent was concentrated under reduced pressure, followed by the addition of aqueous sodium bicarbonate solution and extraction twice with ethyl acetate. The organic layer was washed with aqueous sodium chloride solution and water, dried over anhydrous magnesium sulfate, and filtered. The filtrate was concentrated under reduced pressure and purified by column chromatography to give 1-(tert-butyl) 2-methyl(2S,4S)-4-(N-((1s,4R)-4-methylcyclohexyl)isobutyramido)pyrrolidine-1,2-dicarboxylate (38.79 g, 86%).

[0094] MS [M+H] = 411 (M+1) 1H NMR (400 MHz, CD3OD) δ 4.27 (m, 1H), 3.76 (s, 1.8H), 3.75 (s, 1.2H), 3.78-3.72 (m, 1H), 3.50-3.41 (m, 1H), 3.33-3.14 (m, 1H), 2.69-2.60 (m, 2H), 2.57-2.43 (m, 1H), 1.87-1.79 (m, 1H), 1.70-1.61 (m, 1H), 1.60-1.32 (m, 8H), 1.47 (s, 4.5H), 1.41 (s, 4.5H), 1.10 (dd, 6H), 0.99 (d, 3H)

[0095] Step E: Preparation of methyl (2S,4S)-4-(N-((1s,4R)-4-methylcyclohexyl)isobutyramido)pyrrolidine-2-carboxylate hydrochloride 1-(tert-butyl) 2-methyl(2S,4S)-4-(N-((1s,4R)-4-methylcyclohexyl)isobutyramido)pyrrolidine-1,2-dicarboxylate (34.0 g, 82.8 mmol) obtained in Step D was dissolved in dichloromethane (200 mL), and then 4N hydrochloric acid in 1,4-dioxane (82.8 mL, 331.3 mmol) was added at 0°C. After stirring at room temperature for 6 hours, the reaction solvent was concentrated under reduced pressure to obtain a crude product (28.7 g, 99%), which was used in the next step without purification.

[0096] MS[M+H] = 311 (M+1)

[0097] Production Example 2: Production of (3S,4R)-1-(tert-butyl)-4-(4-chlorophenyl)pyrrolidine-3-carboxylic acid [ka]

[0098] The title compound was obtained by the method described in International Publication No. WO2004 / 092126.

[0099] MS[ M+H] = 282 (M+1) 1H NMR (400 MHz, CD3OD) δ 7.43-7.33 (m, 4H), 3.90-3.69 (m, 3H), 3.59 (dd, J = 11.2, 10.0 Hz, 1H), 3.29 (dd, J = 11.2, 11.2 Hz, 1H), 3.18-3.09 (m, 1H), 1.44 (s, 9H)

[0100] Production Example 3: Production of N-((3S,5S)-1-((3S,4R)-1-(tert-butyl)-4-(4-chlorophenyl)pyrrolidine-3-carbonyl)-5-(morpholine-4-carbonyl)pyrrolidin-3-yl)-N-((1s,4R)-4-methylcyclohexyl)isobutyramide (MC70) [ka]

[0101] The title compound was obtained through the following steps A, B and C.

[0102] Step A: Preparation of methyl (2S,4S)-1-((3S,4R)-1-(tert-butyl)-4-(4-chlorophenyl)pyrrolidine-3-carbonyl)-4-(N-((1s,4R)-4-methylcyclohexyl)isobutyramido)pyrrolidine-2-carboxylate Methyl (2S,4S)-4-(N-((1s,4R)-4-methylcyclohexyl)isobutyramido)pyrrolidine-2-carboxylate hydrochloride (28.7 g, 82.73 mmol) obtained in Production Example 1, (3S,4R)-1-(tert-butyl)-4-(4-chlorophenyl)pyrrolidine-3-carboxylic acid (24.5 g, 86.87 mmol) obtained in Production Example 2, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (22.2 g, 115.83 mmol), and 1-hydroxybenzotriazole hydrate (15.7 g, 115.83 mmol) were dissolved in N,N'-dimethylformamide (400 ml), and N,N'-diisopropylethylamine (72.0 ml, 413.66 mmol) was slowly added. The mixture was stirred at room temperature for 16 hours, and the reaction solvent was concentrated under reduced pressure. Then, 0.5N aqueous sodium hydroxide was added and the mixture was extracted twice with ethyl acetate. The organic layer was washed twice with aqueous sodium chloride and water, dried over anhydrous magnesium sulfate, and filtered. The filtrate was concentrated under reduced pressure and purified by column chromatography to give methyl (2S,4S)-1-((3S,4R)-1-(tert-butyl)-4-(4-chlorophenyl)pyrrolidine-3-carbonyl)-4-(N-((1s,4R)-4-methylcyclohexyl)isobutyramido)pyrrolidine-2-carboxylate (41.19 g, 87%).

[0103] MS [M+H] = 575 (M+1)

[0104] Step B: Preparation of (2S,4S)-1-((3S,4R)-1-(tert-butyl)-4-(4-chlorophenyl)pyrrolidine-3-carbonyl)-4-(N-((1s,4R)-4-methylcyclohexyl)isobutyramido)pyrrolidine-2-carboxylic acid Methyl (2S,4S)-1-((3S,4R)-1-(tert-butyl)-4-(4-chlorophenyl)pyrrolidine-3-carbonyl)-4-(N-((1S,4R)-4-methylcyclohexyl)isobutyramido)pyrrolidine-2-carboxylate (39.4 g, 68.62 mmol) obtained in Step A was dissolved in methanol (450 mL), and 6N aqueous sodium hydroxide solution (57.2 mL, 343.09 mmol) was added. The mixture was stirred at room temperature for 16 hours, and the pH was adjusted to approximately 5 with 6N aqueous hydrochloric acid. The reaction solution was then concentrated under reduced pressure. The concentrated solution was dissolved in dichloromethane, and the undissolved solid was filtered through a paper filter. The filtrate was concentrated under reduced pressure to obtain the crude title compound (38.4 g, 99%), which was used in the next step without purification.

[0105] MS [M+H] = 561 (M+1)

[0106] Step C: Preparation of N-((3S,5S)-1-((3S,4R)-1-(tert-butyl)-4-(4-chlorophenyl)pyrrolidine-3-carbonyl)-5-(morpholine-4-carbonyl)pyrrolidin-3-yl)-N-((1s,4R)-4-methylcyclohexyl)isobutyramide (2S,4S)-1-((3S,4R)-1-(tert-butyl)-4-(4-chlorophenyl)pyrrolidine-3-carbonyl)-4-(N-((1s,4R)-4-methylcyclohexyl)isobutyramido)pyrrolidine-2-carboxylic acid (38.4 g, 68.60 mmol) obtained in Step B, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (18.4 g, 96.04 mmol), and 1-hydroxybenzotriazole hydrate (13.0 g, 96.04 mmol) were dissolved in N,N'-dimethylformamide (200 mL), and then morpholine (5.9 mL, 68.80 mmol) and N,N'-diisopropylethylamine (59.7 mL, 343.02 mmol) were slowly added thereto. The mixture was stirred at room temperature for 16 hours, concentrated under reduced pressure, and then 0.5N aqueous sodium hydroxide was added and extracted twice with ethyl acetate. The organic layer was washed twice with aqueous sodium chloride and water, dried over anhydrous magnesium sulfate, and filtered. The filtrate was concentrated under reduced pressure and purified by column chromatography to give N-((3S,5S)-1-((3S,4R)-1-(tert-butyl)-4-(4-chlorophenyl)pyrrolidine-3-carbonyl)-5-(morpholine-4-carbonyl)pyrrolidin-3-yl)-N-((1s,4R)-4-methylcyclohexyl)isobutyramide (37.05 g, 86%, MC70).

[0107] MS [M+H] = 630 (M+1)

[0108] Example 1: Preparation of Crystalline Form I of N-((3S,5S)-1-((3S,4R)-1-(tert-butyl)-4-(4-chlorophenyl)pyrrolidine-3-carbonyl)-5-(morpholine-4-carbonyl)pyrrolidin-3-yl)-N-((1s,4R)-4-methylcyclohexyl)isobutyramide [ka]

[0109] 0.6 g of the compound (MC70) prepared in Preparation Example 3 was dissolved in 0.975 mL of MTBE at room temperature of 25° C. for 30 minutes. After dissolution was complete, the mixture was cooled to 3° C., stirred for about 21 hours, and filtered to obtain the title compound (crystalline form I of MC70).

[0110] The compound of Example 1 was analyzed by XRPD (FIG. 1), DSC (FIG. 2), TGA (FIG. 3), and DVS (FIG. 4) using the following methods, and the resulting graphs are shown in FIGS. 1 to 4, respectively.

[0111] Experimental Example 1. XRPD evaluation Powder XRPD diffraction patterns were obtained using a PANalytical X'Pert Pro MPD system equipped with a monochromatized radiation source and a Ni filter as a solid-state detector in the following manner.

[0112] Approximately 20-30 mg of sample was placed on a glass sample holder with a flat surface. The instrument generator was set to 45 kV (acceleration voltage) and 40 mA (filament emission), and measurements were performed in reflection mode (not-spin). Bragg angles (2θ) were measured in the range of 4-40° with a step size of 0.026° and a time per step of 51 seconds.

[0113] The obtained crystalline form I was measured by XRPD, and the results are shown in FIG.

[0114] As can be seen from the spectrum shown in FIG. 1, the crystalline form I of the present invention is a crystalline substance, and the specific XRPD values ​​are shown in Table 1 below.

[0115] [Table 1]

[0116] Experimental Example 2. Differential Scanning Calorimetry (DSC) DSC measurements were performed using a Mettler Toledo DSC1 system. Approximately 2–5 mg of sample was weighed and placed in a 40 μL aluminum crucible (flat-bottomed aluminum pan with a pinhole lid) to form a pinhole. The sample was then heated from 25°C to 350°C at a rate of 10°C / min for DSC measurements. During the measurements, nitrogen gas was supplied to the instrument at a rate of 70 mL / min to prevent the inflow of oxygen and other gases. Data collection and evaluation were performed using the software STARe.

[0117] The results of DSC measurement of the obtained crystalline form I are shown in FIG.

[0118] As can be seen from Figure 2, crystalline form I exhibits one endothermic peak (peak 162.13°C) at approximately 159.14°C (Onset). The temperature value may have an error of ±5°C.

[0119] Experimental Example 3: Thermogravimetric Analysis (TGA) TGA measurements were performed using a Mettler Toledo TGA / DSC 1 module. Approximately 4–8 mg of sample was weighed and placed in a 100 μL Al crucible (flat-bottomed aluminum crucible). The sample was then heated from 30°C to 350°C at a rate of 10°C / min and subjected to TGA measurements. Nitrogen gas was supplied to the instrument at a rate of 80 mL / min during the measurements to prevent the inflow of oxygen and other gases. Data collection and evaluation were performed using the software STARe.

[0120] The results of TGA measurement of the obtained crystalline form I are shown in FIG.

[0121] As can be seen from Figure 3, no weight loss was observed in crystalline form I prior to degradation, confirming that it is an anhydrous substance. A melting endotherm was observed with an onset temperature of 159.14°C, confirming that it is a thermally stable compound up to 270°C. The temperature values ​​have an error of ±5°C.

[0122] Experimental Example 4. Moisture Sorption Analysis (DVS) DVS was measured using an automated gravimetric vapor sorption analyzer equipped with a Cahn D200 ultra-microbalance (DVS Intrinsic apparatus, Surface Measurement Systems Ltd., UK).

[0123] First, a 10-20 mg sample was placed in a sample pan and suspended by a hangdown wire. Measurements were started when the weight and humidity reached equilibrium. The relative humidity was increased from 0% to 90% and then decreased back to 0% for two cycles, with the relative humidity increasing by 10% increments using the dm / dt method. The experiment was conducted at room temperature (25°C).

[0124] The results of DVS analysis of the obtained crystalline form I are shown in FIG.

[0125] As shown in Figure 4, the DVS analysis confirmed that the total weight increase observed between 0 and 90% RH was 0.2% w / w. After the DVS analysis, XRPD analysis was performed, and the resulting XRPD pattern matched that of the initial compound, confirming that no physical changes had occurred.

[0126] Experimental Example 5. Stability evaluation Approximately 10 to 30 mg of sample was stored for 4 weeks under accelerated conditions (40°C, 75% RH) in an open state, or under harsh conditions in a sealed state in an oven at 80°C. To compare the sample with a sample stored at room temperature, HPLC analysis was performed using the method in Table 2 below.

[0127] [Table 2]

[0128] The stability of the obtained crystalline form I was evaluated, and the results are shown in Table 3 below.

[0129] [Table 3]

[0130] As can be seen from Table 3 above, the content of crystalline form I according to the present invention was observed to decrease by about 1.3% for up to 4 weeks under accelerated conditions (40°C, 75% RH) and in a sealed state (harsh conditions, 80°C), and the content was observed to decrease by less than 1% for 4 weeks under accelerated conditions. This confirms that crystalline form I according to the present invention exhibits excellent stability against heat and humidity, as it exhibits chemical stability for 4 weeks.

Claims

1. Crystalline Form I of a compound of the following formula 1, a pharmaceutically acceptable salt thereof, or a solvate thereof: The X-ray powder diffraction pattern (XRPD) showed the following diffraction angles (2θ values): 8.240±0.2°, 9.363±0.2°, 10.2693±0.2°, 10.5969±0.2°, 12.050±0.2°, 12.841±0.2°, 13.503±0.2°, 15.5738±0.2°, 16.6030±0.2°, 17.009±0.2°, 17.305±0.2°, 18.364±0.2°, 18.7390±0.2°, 19.188±0.2°, 19.476±0.2°, 20.001±0.2°, 20.4 77±0.2°, 20.665±0.2°, 21.348±0.2°, 21.976±0.2°, 22.580±0.2°, 23.896±0.2°, 4.334±0.2°, 24.812±0.2°, 25.243±0.2°, 25.833±0.2°, 26.646±0.2°, 27.82±0.2°, 28.316±0.2°, 28.609±0.2°, 29.692±0.2°, 30.185±0.2°, and 30.875±0.2°. 【Chemical 1】 (In the above Chemical Formula 1, R １ is C ２ -C ５ alkyl)

2. The R １ is C ２ ~C ４ 2. The crystalline form I of claim 1, wherein the alkyl is alkyl.

3. 3. Crystalline Form I of claim 2, wherein the compound of Formula 1 is N-((3S,5S)-1-((3S,4R)-1-(tert-butyl)-4-(4-chlorophenyl)pyrrolidine-3-carbonyl)-5-(morpholine-4-carbonyl)pyrrolidin-3-yl)-N-((1s,4R)-4-methylcyclohexyl)isobutyramide.

4. 2. The crystalline form I of claim 1, wherein the pharmaceutically acceptable salt is selected from the group consisting of hydrochloride, sulfate, nitrate, phosphate, hydrobromide and hydroiodide.

5. A method for preparing crystalline form I according to any one of claims 1 to 4, comprising the steps of: dissolving the compound of Formula 1 in a crystallization solvent to prepare a mixed solution; and obtaining crystals from the mixed solution.

6. 6. The method for preparing crystalline Form I according to claim 5, wherein the crystallization solvent comprises an organic solvent.

7. 7. The method for preparing Crystalline Form I according to claim 6, wherein the organic solvent comprises diethyl ether, dipropyl ether, dibutyl ether, diisoamyl ether, ethyl methyl ether, methyl propyl ether, methyl butyl ether, ethyl propyl ether, tetrahydrofuran, tetrahydropyran, diphenyl ether, anisole, dimethylacetamide, methyl isobutyl ketone, isopropyl acetate, 1-pentyl alcohol, toluene, or a mixture thereof.

8. 8. The method for preparing crystalline Form I according to claim 7, wherein the organic solvent comprises methyl tertiary butyl ether.

9. 6. The method for preparing crystalline Form I according to claim 5, wherein the step of obtaining the crystals comprises at least one of cooling, stirring, and filtering the mixed solution.

10. 10. The method for preparing crystalline Form I according to claim 9, wherein the step of obtaining the crystals comprises cooling the mixed solution to a temperature of 0-10°C.

11. A pharmaceutical composition comprising crystalline form I of any one of claims 1 to 4 and a pharmaceutically acceptable carrier.

12. A pharmaceutical composition for enhancing the activity of melanocortin-4 receptor, comprising crystalline form I of any one of claims 1 to 4 and a pharmaceutically acceptable carrier.

13. The pharmaceutical composition of claim 12, wherein the composition is for the prevention or treatment of obesity, diabetes, inflammation, or erectile dysfunction.

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