Amorphous melanocortin-4 receptor agonist

Abstract

To provide a novel amorphous compound having an excellent agonistic activity, which is selective for a melanocortin receptor, in particular, melanocortin-4 receptor (MC4R), or a salt thereof.SOLUTION: The present invention relates to an amorphous compound represented by a chemical formula 1, a method for preparing the same, and a pharmaceutical composition comprising the same.SELECTED DRAWING: None

Description

Technical Field

[0001] This application claims the benefit of priority based on Korean Patent Application No. 10-2020-0142399, filed on October 29, 2020, and all the contents disclosed in the document of the Korean Patent Application are incorporated herein by reference in their entirety.

[0002] The present invention relates to an amorphous novel compound having excellent enhancing activity against melanocortin receptors, a method for producing the same, and a pharmaceutical composition containing the same.

Background Art

[0003] Leptin protein is a hormone secreted by adipocytes, and its secretion amount increases as the content of body fat increases. By regulating the functions of various neuropeptides produced in the hypothalamus, it regulates various biological functions including appetite, body fat content, and energy metabolism (Schwartz, et al., Nature 404, 661 - 671 (2000)). The signal transduction of appetite and body weight regulation by leptin protein is carried out by regulating various factors downstream, and the most representative ones are melanocortin, AgRP (agouti-related peptide), and neuropeptide Y (NPY) hormones.

[0004] As a result of excessive calories in the body, when the concentration of leptin in the blood increases, the secretion of the proopiomelanocortin (POMC) protein hormone in the pituitary gland increases, and the production of AgRP and NPY decreases. The small peptide hormone alpha-MSH (melanocyte stimulating hormone) is produced from POMC neurons, and this hormone ultimately induces a decrease in appetite as an agonist of the melanocortin-4 receptor (MC4R) in secondary neurons. On the other hand, as a result of calorie deficiency, when the concentration of leptin decreases, the expression of AgRP, an MC4R antagonist, increases, and the expression of NPY also increases, ultimately promoting appetite. That is, in response to changes in leptin, the alpha-MSH hormone and the AgRP hormone play roles of enhancement and antagonism to MC4R, respectively, and are involved in appetite regulation.

[0005] The alpha-MSH hormone binds to three MCR subtypes in addition to MC4R and induces various physiological responses. To date, five MCR subtypes have been identified. Among them, MC1R is mainly expressed in skin cells and is involved in skin pigmentation regulation. MC2R is mainly expressed in the adrenal gland and is known to be involved in the production of glucocorticoid hormones. Only ACTH (adrenocorticotropic hormone) derived from POMC is its ligand. MC3R and MC4R, which are mainly expressed in the central nervous system, are involved in regulating appetite, energy metabolism, and the efficiency of body fat storage. MC5R, which is 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 demonstrated as the main target for the development of anti-obesity drugs because it shows the effect of efficiently reducing body weight by inducing a decrease in appetite and an increase in 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] In appetite and weight regulation, the role of MC4R was primarily demonstrated by experiments on animal models with abnormal expression of agouti protein (agouti mouse). In the case of Agouti mice, genetic mutations cause the agouti protein to be highly expressed in the central nervous system, where it acts as an antagonist of MC4R 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 results have observed that AgRP (agouti-related peptide), which is similar to the agouti protein, is actually expressed in hypothalamic neurons, and these are also known to be involved in appetite regulation as antagonists 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 shows an effect of reducing appetite. In contrast, when animals are treated with SHU9119 (peptide) or HS014 (peptide), which are MC4R antagonists, a phenomenon of increased appetite is observed (Kask et al., Biochem. Biophys. Res. Comm. 245, 90-93 (1998)). Moreover, in animal studies using Melanotan II (MTII, Ac-Nle-c[Asp-His-DPhe-Arg-Trp-Lys]-NH2) and its similar agonist HP228, after intracerebral, intraperitoneal or subcutaneous administration, effects such as appetite suppression, weight loss, and increased energy metabolism have been confirmed (Thiele T.E., et al. Am J Physiol 274(1 Pt 2), R248-54(1998); Lee M.D., et al. FASEB J 12, A552(1998); Murphy B., et al. J Appl Physiol 89, 273-82(2000)). Conversely, administration of the representative SHU9119 to animals shows significant and continuous feed intake and weight gain, providing pharmacological evidence that MCR agonists can be used as anti-obesity agents. The significant appetite-reducing effect that appears during MTII administration does not appear in MC4R KO (knock-out) mice, and this experimental result also proves that the appetite-reducing effect is mainly achieved by the activation of MC4R (Marsh, et al., Nat Genet 21, 119-122(1999)).

[0008] As obesity therapeutics developed to date, appetite suppressants that act on the central nervous system are the main type. Among them, most are drugs that regulate the action of neurotransmitters. Examples include phentermine and mazindol, which are noradrenalin agents, and fluoxetine and sibutramine, which are serotonergic agents. However, in the case of the neurotransmitter regulators, due to numerous subtypes of receptors, in addition to inhibiting appetite, they also have a wide range of effects on various physiological functions. Therefore, in the case of the regulators, there is a lack of selectivity for each subtype, and when administered for a long time, there are significant drawbacks accompanied by various side effects.

[0009] On the other hand, melanocortin agonists are neuropeptides rather than neurotransmitters. Considering that all other functions except energy metabolism are normal in MC4R gene KO mice, they have the advantage as an action point in that they can induce only weight loss due to appetite inhibition without affecting other physiological functions. In particular, since its receptor belongs to the G-protein coupled receptor (GPCR), which is the most successful category among the new drug action points developed to date, the aspect that it is relatively easy to ensure selectivity for subtype receptors is greatly different from the existing action points.

[0010] As an example of utilizing such a melanocortin receptor as an action point, International Publication Nos. WO2008 / 007930 and WO2010 / 056022 disclose compounds as agonists of the melanocortin receptor.

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

[0012] [Chemical Formula]

[0013] On the other hand, the salt form and crystal structure of a pharmaceutically active ingredient sometimes affect the chemical stability of the drug. Different salt forms of a compound may change the filterability, product stability, storage stability, etc. of the compound, and different crystallization conditions and storage conditions of the compound may change the crystal structure of the compound, and sometimes may cause production accompanied by different crystal forms. Therefore, it is necessary to study the salt form and / or crystal structure having high purity and good chemical stability for a single compound. [Summary of the Invention] [Problems to be Solved by the Invention]

[0014] An object of the present invention is to provide an amorphous novel compound or a salt thereof having excellent selective enhancing activity against melanocortin receptors, particularly melanocortin-4 receptor (MC4R).

[0015] Another object of the present invention is to provide a method for producing the amorphous compound or a salt thereof.

[0016] Another object of the present invention is to provide a pharmaceutical composition containing the amorphous compound or a salt thereof. [Means for Solving the Problems]

[0017] To achieve the above object, On one side, the present invention provides an amorphous compound of the following chemical formula 1, or a salt thereof.

[0018]

Chemical formula

[0019] In the above chemical formula 1, R1 is C2-C5 alkyl.

[0020] The compounds according to the present invention can have an asymmetric carbon center and an asymmetric axis or plane, and can exist as cis or trans isomers, R or S isomers, racemates, mixtures of diastereoisomers and individual diastereoisomers, and all these isomers and mixtures are included within the scope of the present invention.

[0021] In this specification, for convenience, unless otherwise specified, the compound of chemical formula 1 is used in the sense of including both the compound of chemical formula 1, its pharmaceutically acceptable salts and its isomers.

[0022] In one specific example according to the present invention, R1 of the above chemical formula 1 is C2-C5 alkyl. In another specific example according to the present invention, R1 of the above chemical formula 1 is linear or branched C2-C5 alkyl, for example, ethyl, n-propyl, iso-propyl, n-butyl, iso-butyl, sec-butyl or tert-butyl.

[0023] In another specific example according to the present invention, R1 of the above chemical formula 1 is C2 or C3 alkyl. In another specific example according to the present invention, R1 of the above chemical formula 1 is linear or branched C2 or C3 alkyl, for example, ethyl, n-propyl or iso-propyl.

[0024] In one specific example according to the present invention, the pharmaceutically acceptable salts include, but are not limited to, acid addition salts formed by inorganic acids such as hydrochloric acid, sulfuric acid, nitric acid, phosphoric acid, hydrobromic acid, hydroiodic acid, 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, and sulfonic acids such as methanesulfonic acid, benzenesulfonic acid, p-toluenesulfonic acid or naphthalenesulfonic acid.

[0025] In another specific example according to the present invention, the pharmaceutically acceptable salt of the amorphous compound of Chemical Formula 1 can be the hydrochloride compound of the following Chemical Formula 2.

[0026]

Chemical formula

[0027] In Chemical Formula 2, R2 is C2-C5 alkyl.

[0028] In still another specific example according to the present invention, the compound of Chemical Formula 1 or its pharmaceutically acceptable salt can be N-((3S,5S)-1-((3S,4R)-1-(tert-butyl)-4-(4-chlorophenyl)pyrrolidine-3-carbonyl)-5-(morpholine-4-carbonyl)pyrrolidine-3-yl)-N-((1s,4R)-4-methylcyclohexyl)isobutylamide hydrochloride of the following Chemical Formula 3.

[0029]

Chemical formula

[0030] The amorphous compound according to the present invention or its pharmaceutically acceptable salt can be recognized as having no characteristic peaks at 2θ angles of 4° to 40° in the X-ray diffraction (XRD) pattern.

[0031] In one specific example according to the present invention, the amorphous compound or a pharmaceutically acceptable salt thereof can have an XRD pattern shown in FIG. 1.

[0032] The amorphous compound or a pharmaceutically acceptable salt thereof according to the present invention can have no exothermic peak when heated at a temperature of 350° C. or lower in a differential scanning calorimetry (DSC) profile. Further, the amorphous compound or a pharmaceutically acceptable salt thereof can have an endothermic peak in at least one of the temperature ranges of 20 to 130° C. and 210 to 350° C. in the DSC profile. Further, the amorphous compound or a pharmaceutically acceptable salt thereof can have a glass transition temperature (Tg) in a temperature range of 145° C. to 165° C., for example, 150° C. to 160° C., or at a temperature of 156° C. in the DSC profile.

[0033] In one specific example according to the present invention, the amorphous compound or a pharmaceutically acceptable salt thereof can have a DSC profile shown in FIG. 2.

[0034] The amorphous compound or a pharmaceutically acceptable salt thereof according to the present invention can have a weight loss of 10% or less, for example, 1% to 10%, 5% to 9%, or 7% when heated at a temperature of 180° C. or lower in a thermogravimetric analysis (TGA) profile.

[0035] In one specific example according to the present invention, the amorphous compound or a pharmaceutically acceptable salt thereof can have a TGA profile shown in FIG. 3. In the present specification,

[0036] The results of X-ray diffraction (XRD) analysis were shown using a PANalytical X’ Pert Pro MPD system, Malvern Panalytical Ltd.

[0037] Differential scanning calorimetry (DSC) analysis shows the results obtained using DSC1, Mettler-Toledo AG.

[0038] Thermogravimetric analysis (TGA) shows the results obtained using TGA / DSC 1, Mettler-Toledo AG.

[0039] Stability analysis shows the results obtained using HPLC, Agilent Technologies, Inc.

[0040] The amorphous compound of Formula 1 or a pharmaceutically acceptable salt thereof can have a higher purity compared to the crude compound or a pharmaceutically acceptable salt thereof, and can be physically and chemically more stable.

[0041] In addition, the compound of Formula 2, in the form of the hydrochloride salt of the compound of Formula 1, the amorphous hydrochloride salt compound of Formula 2 can have a higher purity compared to the compound of Formula 1, and can be physically and chemically more stable. Moreover, compared to known melanocortin-4 receptor agonists, the enhancing ability for the melanocortin-4 receptor and the preventive or therapeutic effects on diseases such as obesity, diabetes, inflammation, erectile dysfunction, etc. can be further excellent, but the effects of the present invention are not limited thereto.

[0042] In another aspect, the present invention provides a method for producing the amorphous compound of Formula 1 or a pharmaceutically acceptable salt thereof, comprising the steps of dissolving the crude compound represented by Formula 1 in an organic solvent to produce a mixed solution, cooling the mixed solution, and dropwise adding an acid to the cooled mixed solution.

[0043] First, the crude compound represented by Formula 1 is dissolved in an organic solvent.

[0044] For the production of the amorphous compound of Chemical Formula 1 or a pharmaceutically acceptable salt thereof, the crude compound of Chemical Formula 1 can be a compound of Chemical Formula 1, a salt thereof, or an isomer.

[0045] The compound of Chemical Formula 1 can be obtained by the production method described in the specification of Korean Application No. 10-2019-0141649 (filed on November 7, 2019).

[0046] The solvent for dissolving the crude compound of Chemical Formula 1 can be used without particular limitation as long as it can dissolve the crude compound of Chemical Formula 1. In one specific example, the solvent can be an organic solvent, and specifically, it can include an ether-based organic solvent.

[0047] The ether-based organic solvents include, but are not limited to, for example, dialkyl ether-based 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-based solvents such as tetrahydrofuran and tetrahydropyran; and aromatic ring-containing ether-based solvents such as diphenyl ether and anisole. The organic solvent can be used alone as one solvent, or two or more of the solvents can be mixed and used.

[0048] In one specific example according to the present invention, the organic solvent can include methyl tert-butyl ether (MTBE).

[0049] For 1 g of the crude compound of Chemical Formula 1, 10 to 30 mL or 15 to 25 mL of the organic solvent can be used.

[0050] The dissolution of the crude compound of Chemical Formula 1 in the solvent can be carried out without stirring or with stirring at a temperature of 15 to 30 °C, specifically 23 to 28 °C.

[0051] In one specific example according to the present invention, for 1 g of the crude compound of Chemical Formula 1, 19 mL of MTBE can be used to obtain a mixed solution in which the crude compound of Chemical Formula 1 is dissolved at 25°C.

[0052] Next, the mixed solution in which the crude compound of Chemical Formula 1 is dissolved is cooled.

[0053] The cooling can be performed such that the temperature of the mixed solution becomes 0°C or lower. Specifically, the cooling can be performed such that the temperature of the mixed solution becomes -10 to 0°C, specifically, -5 to 0°C.

[0054] The production method of the present invention can further include the step of adding a nonpolar organic solvent to the mixed solution before, after, or simultaneously with the cooling of the mixed solution. By adding the nonpolar organic solvent to increase the formation rate of crystallization particles, the yield or production stability of the amorphous compound obtained can be made excellent, but the present invention is not limited thereto.

[0055] The nonpolar organic solvent can be used without particular limitation as long as it is an organic solvent having nonpolar characteristics. For example, hexane, heptane, cyclohexane, carbon tetrachloride, benzene, chloroform, etc. can be used.

[0056] In one specific example according to the present invention, after adding heptane to the mixed solution, it can be cooled to -5 to 0°C.

[0057] The step of adding the nonpolar organic solvent can be any step in the production method of the amorphous compound of the present invention.

[0058] Next, an acid is added dropwise to the cooled mixed solution.

[0059] The acid includes hydrochloric acid, and the addition of the acid can be performed in the presence of ethyl acetate.

[0060] In one specific example according to the present invention, the addition of the acid may be achieved by adding a hydrochloric acid-ethyl acetate solution to the mixed solution.

[0061] The concentration of hydrochloric acid in the hydrochloric acid-ethyl acetate solution can be 0.5 M to 6 M, specifically, 1 M to 5 M, 3 M to 5 M, for example, 4 M.

[0062] The precipitate formed by the addition of the acid can be filtered and washed to obtain an amorphous compound of Chemical Formula 1, specifically, an amorphous compound of Chemical Formula 2 or an amorphous pharmaceutically acceptable salt thereof.

[0063] The amorphous compound of Chemical Formula 1 obtained as described above can have a higher purity and can be physically and chemically more stable compared to the crude compound 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) the amorphous compound of Chemical Formula 1, or a salt of the amorphous compound of Chemical Formula 1; and (ii) a pharmaceutically acceptable carrier.

[0065] The amorphous compound of Chemical Formula 1 according to the present invention, or a salt of the amorphous compound of Chemical Formula 1, exhibits excellent enhancing effects on melanocortin receptors, particularly, melanocortin-4 receptor (MC4R). Therefore, the present invention can provide a pharmaceutical composition for enhancing the function of melanocortin receptors containing the above-mentioned amorphous compound or a salt thereof as an active ingredient. Specifically, the pharmaceutical composition can be a composition for enhancing the function of melanocortin-4 receptor.

[0066] In addition, the pharmaceutical composition can exhibit excellent effects on the prevention or treatment of obesity, diabetes, inflammation, and erectile dysfunction, and can be a composition for preventing or treating obesity, preventing or treating diabetes, preventing or treating inflammation, or preventing or treating erectile dysfunction, but the uses of the present invention are not limited only to these diseases.

[0067] As used herein, "carrier" means a compound that facilitates the introduction of a compound into a cell or tissue.

[0068] When administering the amorphous compound of the present invention for clinical purposes, the total daily dose administered to the host in a single volume or divided volumes is preferably in the range of 0.01 to 10 mg per kg of body weight. However, the specific dose level for an individual patient can vary depending on the specific compound used, the patient's body weight, gender, health status, diet, time of drug administration, method of administration, excretion rate, drug combination, and severity of the disease, etc.

[0069] The amorphous compound 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 composition of the present invention can be in various oral dosage forms such as tablets, pills, powders, capsules, granules, syrups or emulsions, or in parenteral dosage forms such as injectable preparations for intramuscular, intravenous or subcutaneous administration.

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

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

[0073] When the pharmaceutical composition of the present invention is in the form of an oral administration, examples of carriers 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, examples of the carrier include, but are not limited to, water, saline, glucose aqueous solution, pseudo-sugar aqueous solution, alcohol, glycol, ether, oil, fatty acid, fatty acid ester, glyceride, etc.

[0075] In yet another aspect, there is provided the above amorphous compound of Chemical Formula 1 or its amorphous salt compound for use in enhancing the function of melanocortin receptors, particularly melanocortin-4 receptor (MC4R).

[0076] In one embodiment, there is provided the above amorphous compound of Chemical Formula 1 or its amorphous salt compound 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 function of a melanocortin receptor, particularly the melanocortin-4 receptor (MC4R), which includes the step of administering to a subject the above-described amorphous compound of Chemical Formula 1 or an amorphous salt compound thereof.

[0078] In yet another aspect, there is provided a method for treating obesity, diabetes, inflammation or erectile dysfunction, which includes the step of administering to a subject the above-described amorphous compound of Chemical Formula 1 or an amorphous salt compound thereof.

Advantages of the Invention

[0079] The amorphous compound of Chemical Formula 1 or a salt thereof according to the present invention exhibits excellent enhancing effects on melanocortin receptors, particularly the melanocortin-4 receptor (MC4R), and thus can be usefully used for the prevention or treatment of obesity, diabetes, inflammation and erectile dysfunction.

[0080] The amorphous compound of Chemical Formula 1 or a salt thereof according to the present invention shows an on-target effect on the melanocortin-4 receptor, exhibits weight loss and diet reduction effects, does not affect anxiety and depression, and can be administered without safety problems such as side effects on hERG (human ether-a-go-go related gene) inhibition and induction of mutations.

[0081] In addition, the amorphous compound of Chemical Formula 1 or a salt thereof according to the present invention is superior in purity, yield, physical and chemical stability compared to the crude compound of Chemical Formula 1.

[0082] Specifically, the amorphous compound of Chemical Formula 1 or a salt thereof can be superior in solubility, storage stability and production stability compared to the crude compound of Chemical Formula 1.

Brief Description of the Drawings

[0083]

Figure 1

Figure 2

Figure 3

Mode for Carrying Out the Invention

[0084] Hereinafter, the present invention will be described more specifically with reference to Production Examples and Examples. However, these Examples are illustrative of the present invention, and the scope of the present invention is not limited thereto.

[0085] Production Example 1: Production of Methyl (2S,4S)-4-((1S,4R)-4-Methylcyclohexyl)isobutylamide)pyrrolidine-2-carboxylate Hydrochloride

Chemical formula

[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. Stirring was carried out at 80 °C for 16 hours. After concentrating the reaction solvent under reduced pressure, water was added, and extraction was performed twice with ethyl acetate. The organic layer was washed with an aqueous sodium chloride solution and water, and then 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-azidopyrrolidine-1,2-dicarboxylate (39.59 g, 98%), which was used in the next step without purification.

[0088] MS [M+H] = 271 (M+1) 11H 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 the above step A was dissolved in tetrahydrofuran (180 ml), and then 1M tetrahydro solution of trimethylphosphine (109.2 ml, 109.2 mmol) was gradually added at 0 °C. After stirring at the same temperature for 1 hour, the mixture was stirred at room temperature for 3 hours. After concentrating the reaction solvent under reduced pressure, dichloromethane (100 ml) and water (150 ml) were added and stirred for about 30 minutes. After liquid separation and extraction with dichloromethane once again, 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%) and used in the next step without purification.

[0090] MS [M+H] = 245 (M+1) 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 The 1-(tert-butyl) 2-methyl (2S,4S)-4-aminopyrrolidine-1,2-dicarboxylate (20.62 g, 84.4 mmol) obtained in the previous 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 an aqueous sodium chloride solution and then 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) 1 H 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)isobutylamide)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 the previous step C was dissolved in dichloromethane (500 ml), triethylamine (61.1 ml, 438.1 mmol) was added, and then isobutyryl chloride (11.7 ml, 219 mmol) was gradually added at 0 °C. After stirring at room temperature for 16 hours, the reaction solvent was concentrated under reduced pressure, an aqueous sodium hydrogen carbonate solution was added, and extraction was performed twice with ethyl acetate. The organic layer was washed with an 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 obtain 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) 1 H 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)isobutylamide)pyrrolidine-2-carboxylate hydrochloride 1-(tert-Butyl) 2-methyl (2S,4S)-4-(N-((1s,4R)-4-methylcyclohexyl)isobutylamide) pyrrolidine-1,2-dicarboxylate (34.0 g, 82.8 mmol) obtained in the above step D was dissolved in dichloromethane (200 ml), and then 4N hydrochloric acid 1,4-dioxane solution (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 crude (28.7 g, 99%), which was used in the next step without purification.

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

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

Chemical formula

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

[0099] MS[ M+H] = 282 (M+1) 1 H 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: 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)isobutylamide

Chemical formula

[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)isobutylamide)pyrrolidine-2-carboxylate Methyl (2S,4S)-4-(N-((1s,4R)-4-methylcyclohexyl)isobutylamide)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 gradually added. Stirring was carried out at room temperature for 16 hours. After concentrating the reaction solvent under reduced pressure, 0.5 N aqueous sodium hydroxide solution was added and extraction was performed twice with ethyl acetate. The organic layer was washed twice each with aqueous sodium chloride solution and water, and then dried over anhydrous magnesium sulfate and filtered. The filtrate was concentrated under reduced pressure and purified by column chromatography to obtain methyl (2S,4S)-1-((3S,4R)-1-(tert-butyl)-4-(4-chlorophenyl)pyrrolidine-3-carbonyl)-4-(N-((1s,4R)-4-methylcyclohexyl)isobutylamide)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)isobutylamide)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)isobutylamide)pyrrolidine-2-carboxylate (39.4 g, 68.62 mmol) obtained in the previous step A was dissolved in methanol (450 ml), and then 6N aqueous sodium hydroxide solution (57.2 ml, 343.09 mmol) was added. The mixture was stirred at room temperature for 16 hours, adjusted to about pH 5 with 6N aqueous hydrochloric acid solution, and then the reaction solution was 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)isobutylamide (2S,4S)-1-((3S,4R)-1-(tert-Butyl)-4-(4-chlorophenyl)pyrrolidine-3-carbonyl)-4-(N-((1s,4R)-4-methylcyclohexyl)isobutylamido)pyrrolidine-2-carboxylic acid (38.4 g, 68.60 mmol), 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 gradually added in sequence. Stirring was carried out at room temperature for 16 hours, and after the reaction solution was concentrated under reduced pressure, 0.5 N aqueous sodium hydroxide solution was added and extracted twice with ethyl acetate. The organic layer was washed twice each with aqueous sodium chloride solution and water, and then dried over anhydrous magnesium sulfate and filtered. The filtrate was concentrated under reduced pressure and purified by column chromatography to obtain 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)isobutylamide (37.05 g, 86%, Production Example 3).

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

[0108] Example 1 Preparation of amorphous compound 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)isobutylamide hydrochloride Based on 1 g of the compound (MC70) finally produced in Production Example 3 above, 19 mL of MTBE was used to dissolve the compound (MC70) at 25°C. After the dissolution was complete, 1 mL of heptane was added, and then the mixture was cooled to -5 to 0°C. After reaching the set temperature, 1 equivalent of 4M HCl / EtOAc was added dropwise, followed by stirring for about 90 minutes. The mixture was then filtered to obtain amorphous N-((3S,5S)-1-((3S,4R)-1-(tert-butyl)-4-(4-chlorophenyl)pyrrolidin-3-carbonyl)-5-(morpholine-4-carbonyl)pyrrolidin-3-yl)-N-((1s,4R)-4-methylcyclohexyl)isobutylamide hydrochloride (Example 1). (Yield: about 90%)

[0109] Amorphous characteristics 1) X-ray diffraction (XRD) The powder XRD diffraction pattern was obtained by the following method using a PANalytical X’Pert Pro MPD system equipped with a monochromatized radiation source and a Ni filter as a solid-state detector.

[0110] After uniformly placing about 20 - 30 mg of the sample on a glass sample holder so as to have a flat surface, the generator of the instrument was set to 45 kV (acceleration voltage) and 40 mA (filament emission), and then measurement was carried out in reflection mode (not-spin). The Bragg angle (2θ) in the range of 4 - 40° was measured under the conditions of a step size of 0.026° and a time per step of 51 seconds.

[0111] The XRD results are illustrated in Figure 1. As shown in Figure 1, amorphous Example 1 had no diffraction peaks and showed a broad range of noise typical of an amorphous sample.

[0112] 2) Differential scanning calorimetry (DSC) DSC was measured using a Mettler Toledo DSC1 system. A sample of approximately 2 - 5 mg was weighed and placed in a 40 μL Al crucible (flat-bottomed aluminum pan with one pin-hole lid) to form one pin hole. Next, the sample was heated from 25 °C to 350 °C at a rate of 10 °C / min to measure DSC. During the measurement, nitrogen gas was supplied into the apparatus 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.

[0113] The DSC results are shown in Figure 2. An endothermic peak was observed at approximately 33.2 °C (Onset), and the glass transition temperature was observed at approximately 156.5 °C (Midpoint ISO). After approximately 220 °C, an endothermic peak due to decomposition was shown. The temperature values have an error of ±5 °C.

[0114] 3) Thermogravimetric analysis (TGA) TGA was measured using a Mettler Toledo TGA / DSC 1 module. A sample of approximately 4 - 8 mg was weighed and placed in a 100 μL Al crucible (flat-bottomed aluminum crucibles). Next, the sample was heated from 30 °C to 350 °C at a rate of 10 °C / min to measure TGA. During the measurement, nitrogen gas was supplied into the apparatus at a rate of 80 mL / min to prevent the inflow of oxygen and other gases. Data collection and evaluation were performed using the software STARe.

[0115] The TGA results are shown in Figure 3. As shown in Figure 3, a weight loss of approximately 5.6% was observed at a temperature below 100 °C. Next, a weight loss of approximately 1.1% was observed at approximately 150 °C - 180 °C. After approximately 220 °C, a weight loss due to decomposition occurred. The temperature values have an error of ±5 °C.

[0116] Experimental Example 1. Stability Evaluation A sample of about 10 to 30 mg was stored in an unsealed state under accelerated conditions (40 °C, 75% RH), and under severe conditions, it was stored in a sealed state in an oven at 80 °C for 4 weeks. To compare the sample with the sample stored at room temperature, HPLC analysis was performed by the method shown in Table 1.

[0117]

Table 1

[0118] The stability results of Example 1 evaluated by the above method are shown in Table 2 below.

[0119]

Table 2

Claims

1. An amorphous compound of the following chemical formula 1 or a pharmaceutically acceptable salt thereof. 【Chemical 1】 In the above chemical formula 1, R １ is C ２ -C ５ alkyl.

2. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, having the X-ray diffraction pattern shown in FIG.

1.

3. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, having the DSC profile shown in FIG.

2.

4. The compound according to claim 1 or a pharmaceutically acceptable salt thereof, having the TGA profile shown in FIG.

3.

5. The pharmaceutically acceptable salt of the compound of the above chemical formula 1 is selected from the group consisting of hydrochloride, sulfate, nitrate, phosphate, hydrobromide and hydroiodide salts of the said compound, the compound according to claim 1 or a pharmaceutically acceptable salt thereof.

6. 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)isobutylamide hydrochloride, the compound according to claim 1 or a pharmaceutically acceptable salt thereof.

7. A method for producing an amorphous compound according to any one of claims 1 to 6 or a pharmaceutically acceptable salt thereof, comprising: dissolving a compound represented by the following chemical formula 1 in an organic solvent to produce a mixed solution; cooling the said mixed solution; and adding an acid dropwise to the cooled mixed solution. 【Chemical Formula 2】 In the above chemical formula 1, R １ is C ２ -C ５ alkyl.

8. The production method according to claim 7, wherein the organic solvent includes an ether-based organic solvent.

9. The production method according to claim 7, further comprising adding a non-polar organic solvent to the mixed solution before, after or simultaneously with the cooling of the mixed solution.

10. A pharmaceutical composition comprising the compound according to any one of claims 1 to 6 and a pharmaceutically acceptable carrier.

11. A pharmaceutical composition for enhancing melanocortin-4 receptor function, comprising the compound according to any one of claims 1 to 6 and a pharmaceutically acceptable carrier.

12. The pharmaceutical composition according to claim 11, wherein the composition is for the prevention or treatment of obesity, diabetes, inflammation or erectile dysfunction.

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