Application and preparation method of L-serine or pharmaceutically acceptable salt thereof

Abstract

The invention relates to application and a preparation method of L-serine or pharmaceutically acceptable salt thereof. Specifically, one aspect of the present invention relates to the use of a medicament for the preparation of L-serine or a pharmaceutically acceptable salt thereof, said medicament comprising a pharmaceutically acceptable carrier, in a unit form, for the treatment of communication disorders and / or social interaction disorders in a subject with autism spectrum disorders.

Description

Technical Field

[0001] One aspect of the present invention relates to the use of L-serine or a pharmaceutically acceptable salt thereof and a method for preparing the same. Furthermore, another aspect of the present invention relates to a preparation and a kit comprising L-serine or a pharmaceutically acceptable salt thereof, wherein the preparation is in unit dose form. Background Art

[0002] During the development of children, balanced development of excitatory synapses / inhibitory synapses is achieved during the critical stage of reorganizing the signal transmission system of the cerebral cortex. On the other hand, if synaptic pruning is not performed correctly during the critical period, neurodevelopmental disorders characterized by reciprocal social interaction disorders / lack, communication disorders and / or repetitive behaviors will occur. Therefore, in order to treat or improve the neurodevelopmental disorders, early intervention is required before the end of the critical period.

[0003] However, the causes and symptoms of the neurodevelopmental disorders are very diverse, making it difficult to find a therapeutic target substance or therapeutic mechanism. In addition, in patients with neurodevelopmental disorders, risperidone and aripiprazole are limitedly used in patients with severe aggression and anxiety, but these drugs cannot improve social interaction disorders and / or communication disorders. Although drugs are being developed to improve social interaction disorders and / or communication disorders, there are currently no approved drugs.

[0004] On the other hand, L-serine participates in a variety of metabolic systems and plays an important role in the normal growth and development of nerve cells. Phospholipids such as phosphatidylserine and sphingosine derived from L-serine are the main components of the myelin sheath surrounding nerve cells, enabling rapid transmission of neural information. In addition, mutations in genes (PHGDH, PSAT, PSPH) related to L-serine synthesis lead to severe neurodevelopmental disorders and malformations, and this phenomenon suggests that most of the L-serine used to maintain normal brain function is synthesized in the brain. However, it is not clear whether L-serine can improve autism spectrum disorders or social interaction disorders and / or communication disorders.

[0005] In addition, L-serine is widely contained in food, but since it is an amino acid that is difficult to cross the blood-brain barrier, it has limitations in reaching the brain only through diet. Therefore, in order to use L-serine as an active ingredient in medicine, high doses of L-serine are required. L-serine can be prepared by a variety of methods, such as (i) fermentation of Escherichia coli using formaldehyde or fermentation using Gram-negative bacteria, or (ii) preparing DL-serine mixtures using mercury and performing optical resolution. However, if L-serine is prepared using a fermentation method, there is a problem of building a new clean area in the preparation factory and conducting harmful bacteria analysis tests. In addition, formaldehyde is a carcinogen and mercury is very toxic to the human body, so it is difficult to use these substances to prepare L-serine for human administration. In addition to the method, other types of L-serine preparation methods are known to have low production yields.

[0006] [Prior art literature]

[0007] (Patent Document 1) U.S. Patent No. 5382517

[0008] (Patent Document 2) China Published Patent No. 105037060

[0009] (Patent Document 3) Japanese Patent Publication No. 1993-140056 Summary of the invention

[0010] Technical issues

[0011] There is a need to develop a therapeutic approach that can treat symptoms of communication impairment and / or social interaction impairment in subjects with autism spectrum disorders.

[0012] Furthermore, in order to use L-serine or its pharmaceutically acceptable salt which is difficult to be directly delivered to the central nervous system as medicine, it is required to develop a preparation containing high-dose or high-concentration L-serine. In order to develop such preparation, a preparation method is required which can produce high-concentration L-serine or its pharmaceutically acceptable salt in large quantities with high yield without using harmful substances such as formaldehyde.

[0013] Technical Solution

[0014] [1] One aspect of the present invention relates to a method for producing L-serine, comprising: a) a step of chelating glycine and a copper compound to form a glycine-copper complex; b) a step of reacting the complex of step a) with a carbon source to form a complex into which a hydroxymethyl group is introduced; c) a step of treating the complex of step b) with a reducing agent to form DL-serine; and d) a step of optically resolving the DL-serine of step c) to separate L-serine.

[0015] [2] In [1], the copper compound in step a) may be selected from CuSO 4 , CuOAc 2 , CuCl 2 、CuI、CuBr、CuBr 2 and any one or more of the group consisting of hydrates thereof.

[0016] [3] In [1] or [2], the step a) may be performed by adding glycine, copper, a base and purified water.

[0017] [4] In [3], the amount of purified water added may be 6 to 30 times the weight of glycine.

[0018] [5] In any one of [1] to [4], the glycine-copper complex of step a) may have the structure of the following Chemical Formula 2:

[0019] [Chemical formula 2]

[0020]

[0021] [6] In any one of [1] to [5], the step b) may be performed by adding a carbon source, an alkali, and purified water.

[0022] [7] In any one of [1] to [6], the carbon source may be at least one selected from the group consisting of formaldehyde, polyoxymethylene, 1,3-dioxolane, 1,3-dioxane, and hexamidine.

[0023] [8] In any one of [1] to [7], the complex of step b) may have the structure of the following Chemical Formula 3:

[0024] [Chemical formula 3]

[0025]

[0026] [9] In any one of [1] to [8], the reducing agent in step c) may be sodium sulfide (Na 2 S).

[0027]

[10] In any one of [1] to [9], after the step c) is performed, the method may further include a step of performing reflux cooling using purified water.

[0028]

[11] In any one of [1] to

[10] , in the step d), at least one acid selected from the group consisting of tartaric acid, malic acid, mandelic acid and camphorsulfonic acid may be used.

[0029]

[12] Another aspect of the present invention provides a use of L-serine or a pharmaceutically acceptable salt thereof for preparing a medicament for improving or treating communication disorders and / or reciprocal social interaction disorders in subjects with autism spectrum disorders, wherein the medicament comprises a pharmaceutically acceptable carrier and is in unit dose form.

[0030]

[13] In

[12] , the subject may have childhood autism.

[0031]

[14] In

[12] or

[13] , the clinical global impression-severity scale (CGI-S) of the subject can be above 5, and the anti-social behavior scale (ASB) of the Vineland Adaptive Behavior Scales-II (VABS-II) can be above 48.

[0032]

[15] In any of

[12] to

[14] , the subject may be under 18 years old, under 13 years old, under 11 years old, under 7 years old, less than 7 years old, 2 to 7 years old, or 2 to 6 years old.

[0033]

[16] In any of

[12] to

[15] , the subject may have a body weight of 10 kg to 100 kg or 10 kg to 60 kg.

[0034]

[17] In any of

[12] to

[16] , the daily dosage of L-serine or a pharmaceutically acceptable salt thereof administered to the subject may be 1 g to 60 g, 2 g to 30 g, 4 g to 30 g, 2 g to 28 g or 2 g to 14 g.

[0035]

[18] In any of

[12] to

[16] , the daily dose of L-serine or a pharmaceutically acceptable salt thereof administered to the subject may be 100 mg / kg to 600 mg / kg, 140 mg / kg to 580 mg / kg, 140 mg / kg to 572 mg / kg, 200 mg / kg to 400 mg / kg, 280 mg / kg to 580 mg / kg, 280 mg / kg to 572 mg / kg, 70 mg / kg to 280 mg / kg, 140 mg / kg to 290 mg / kg or 140 mg / kg to 286 mg / kg.

[0036]

[19] In any of

[12] to

[18] above, the drug may be administered to the subject twice a day.

[0037]

[20] In any one of

[12] to

[19] , the pharmaceutical agent may be in an oral dosage form.

[0038]

[21] In any of

[12] to

[20] , the pharmaceutical agent may be in the form of a syrup.

[0039]

[22] In any of

[12] to

[21] , the agent may include one or more of a sweetener, a thickener, a pH adjuster, a preservative, and a solvent.

[0040]

[23] In any one of

[12] to

[22] , the concentration of L-serine or a pharmaceutically acceptable salt thereof contained in the agent may be 10 mg / ml to 500 mg / ml.

[0041]

[24] In any one of

[12] to

[23] , it may be characterized in that each volume of the unit dosage form is 10 ml to 100 ml.

[0042]

[25] In any of

[12] to

[24] , the content of L-serine or a pharmaceutically acceptable salt thereof contained in the unit dosage form may be 1 g to 100 g, 1 g to 60 g, 2 g to 30 g, 4 g to 30 g, 2 g to 28 g or 1 g to 14 g.

[0043]

[26] In any one of

[12] to

[25] , the feature may be that the L-serine is prepared by the following method:

[0044] a) a step of chelating glycine and a copper compound to form a glycine-copper complex;

[0045] b) reacting the complex obtained in step a) with a carbon source to form a complex into which a hydroxymethyl group is introduced;

[0046] c) treating the complex obtained in step b) with a reducing agent to form DL-serine; and

[0047] d) a step of optically resolving the DL-serine obtained in step c) to separate L-serine.

[0048]

[27] Another aspect of the present invention provides a preparation comprising 1 g to 100 g of L-serine or a pharmaceutically acceptable salt thereof, a thickener, a pH adjuster, a preservative and a solvent, wherein the preparation is in a unit dosage form.

[0049]

[28] In the above

[27] , it may be characterized in that each volume of the preparation is 10ml to 100ml.

[0050]

[29] In

[27] or

[28] , the preparation can be administered to improve or treat communication disorders and / or social interaction disorders in a subject with autism spectrum disorder.

[0051]

[30] In any of

[27] to

[29] , the daily dose of L-serine or a pharmaceutically acceptable salt thereof administered to the subject may be 100 mg / kg to 600 mg / kg, 140 mg / kg to 580 mg / kg, 140 mg / kg to 572 mg / kg, 200 mg / kg to 400 mg / kg, 280 mg / kg to 580 mg / kg, 280 mg / kg to 572 mg / kg, 70 mg / kg to 280 mg / kg, 140 mg / kg to 290 mg / kg or 140 mg / kg to 286 mg / kg.

[0052]

[31] One aspect of the present invention provides a kit comprising a preparation containing 1 g to 100 g of L-serine or a pharmaceutically acceptable salt thereof, wherein the preparation is used to improve or treat communication disorders and / or social interaction disorders in a subject with autism spectrum disorder, and the preparation is in unit dosage form.

[0053] Effects of the Invention

[0054] In addition, L-serine of the present invention or its pharmaceutically acceptable salt or the preparation (for example, unit dosage form) comprising said L-serine or its pharmaceutically acceptable salt, is excellent in improving the effect of social interaction disorder and / or communication disorder. And then, L-serine or its pharmaceutically acceptable salt and said preparation are excellent in improving the effect of autism spectrum disorder symptom.

[0055] The method for preparing L-serine or a pharmaceutically acceptable salt thereof of the present invention is a method for synthesizing L-serine or a pharmaceutically acceptable salt thereof from glycine without using toxic substances or bacteria such as Escherichia coli, and has the advantage of being industrially applicable. BRIEF DESCRIPTION OF THE DRAWINGS

[0056] Figure 1 One embodiment of a method for producing L-serine or a pharmaceutically acceptable salt thereof is shown.

[0057] Figure 2 The results of analyzing L-serine obtained by the preparation method of the present invention using Differential Scanning Calorimeter (DSC) are shown.

[0058] Figure 3 Shown are the results of a 36-month stability test of a formulation containing L-serine.

[0059] Figure 4 The diagram shows an overview of social experiments and social cognition experiments in autism spectrum disorder models and the results of the social experiments and social cognition experiments. Figure 4 A is a diagram showing an overview of the social experiment. Figure 4 B is a diagram showing an overview of the social cognition experiment. Figure 4 C and Figure 4 D is a graph showing the results of a social experiment. Figure 4 E and Figure 4 F is a graph showing the results of a social cognition experiment. Figure 4 * indicates p value < 0.05, ** indicates p value < 0.01, *** indicates p value < 0.001, **** indicates p value < 0.0001, and ns indicates not statistically significant.

[0060] Figure 5 A diagram showing a cross-shaped maze used in an anxiety experiment in an autism spectrum disorder model and a diagram showing the results of the anxiety experiment. Figure 5 A is a schematic diagram of a cross-shaped maze used in anxiety experiments in autism spectrum disorder models. Figure 5 B is a heat map showing the activity of mice during the behavioral test. Figure 5 C and Figure 5 D are graphs showing the results of the anxiety experiment. Figure 5 * indicates p value < 0.05, ** indicates p value < 0.01, *** indicates p value < 0.001, and ns indicates not statistically significant.

[0061] Figure 6The graph shows the results of an experiment to confirm the normalizing effect of dopamine secretion in an autism spectrum disorder model using patch clamp. Figure 6 A is a graph showing the results of analyzing the spontaneous action potential discharge rate in the midbrain VTA region in the control group and each experimental group. Figure 6 B is a graph showing the results of potential frequency analysis in the control group and each experimental group. Figure 6 The * indicates p-value < 0.05, and the ** indicates p-value < 0.01.

[0062] Figure 7 Experiments to confirm the normalizing effect of synaptic pruning in models of autism spectrum disorders. Figure 7 A is a picture of the overall cortical area (prelimbic cortex), Figure 7 B is a diagram of the density distribution of dendritic spines of pyramidal neurons in layer 5 of the cortex observed using a confocal microscope system. Figure 7 C is a graph showing the dendritic spine density of cortical layer 5 pyramidal neurons among the groups. Figure 7 ** indicates p-value < 0.01, **** indicates p-value < 0.0001.

[0063] Figure 8 The figure shows the experimental results of confirming the intrinsic excitability of mPFC pyramidal neurons in an autism spectrum disorder model using patch clamp. Figure 8 A is a graph showing the discharge rate of action potentials corresponding to current injection, Figure 8 B is shown Figure 8 A graph of the discharge rate variation of A. Figure 8 ** indicates p-value < 0.01, *** indicates p-value < 0.001, and **** indicates p-value < 0.0001.

[0064] Fig. 9 This is a graph showing the difference in spatial working memory in an autism spectrum disorder model at different doses of L-serine.

[0065] Fig.10 This is a graph showing individual recognition differences at different doses of L-serine in an autism spectrum disorder model.

[0066] Fig.11 This is a graph showing the plasma concentration-time curve of beagle dogs to which L-serine was orally or intravenously administered.

[0067] Fig.12 A diagram showing an overview of the Phase II clinical trial.

[0068] Fig.13To show the trends (Line plot) of three groups at different time points regarding the changes in scores of the Korean-Vineland Adaptive Behavior Scale-II (K-VABS-II) Adaptive Behavior Composite (ABC) at 12 weeks, 24 weeks, and 36 weeks after the start of the Phase II clinical trial compared with the start of the Phase II clinical trial (FAS group).

[0069] Fig.14 The K-VABS-II ABC and the Korean-Parenting Stress Index-4 at 12 weeks after AST-001 syrup administration are shown in comparison with the start of the Phase II clinical trial. th Figure 3 shows the effect difference (FAS group) between the control group (placebo) and the treatment group (high-dose group and low-dose group) of Parental Distress values ​​of the K-PSI-4-SF (hereinafter referred to as K-PSI-4-SF), as well as the 90% confidence interval and statistical significance.

[0070] Fig.15 This graph analyzes the change in K-VABS-II 2-Domain Composite (the average of the standard scores in the two domains of communication and sociality, hereinafter referred to as 2DC) scores (FAS group) at 12 weeks, 24 weeks, and 36 weeks after the start of the Phase II clinical trial compared to the start of the clinical trial.

[0071] Fig.16 A graph showing the trend (Line plot) of three groups of different time points regarding the change in Clinical Global Impression-Severity (CGI-S) scores (FAS group) at 2 weeks, 4 weeks, 8 weeks, 12 weeks, 14 weeks, 16 weeks, 24 weeks and 36 weeks compared to the start of the Phase II clinical trial.

[0072] Fig.17 This is a graph analyzing the changes in K-VABS-II ABC (PPS population) at 12, 24, and 36 weeks compared to the start of the Phase II clinical trial.

[0073] Fig.18 This is a graph showing an overview of the 52-week continuous dosing clinical trial.

[0074] Fig.19The figure is a graph showing the different time point trends (Line plot) of the CGI-S scores of two groups (FAS group) of patients participating in the 52-week continuous dosing clinical trial from the start point of the Phase II clinical trial to the end point of the 52-week continuous dosing clinical trial.

[0075] Fig. 20 This is a graph showing the different time point trends (Lineplot) of the changes in CGI-S scores of two groups (FAS group) from the start of the Phase II clinical trial to the end of the 52-week continuous dosing clinical trial.

[0076] Fig.21 A graph showing the predicted results of K-VABS-II ABC score morphology at 12 weeks of AST-001 dosing in a simulation analysis for modeling the dosing to be used in a Phase III clinical trial. DETAILED DESCRIPTION

[0077] definition

[0078] In the present specification, the term "subject" includes humans, mice, rats, dogs, cats, horses, cows, primates without limitation. The subject may be a human.

[0079] In this specification, the term "unit dose" means a daily or single dose of an active ingredient (e.g., L-serine or a pharmaceutically acceptable salt thereof) administered to a subject in order to provide a desired result. A unit dose may be administered to a subject as a single dose or a series of subdoses.

[0080] In this specification, the term "unit dosage form" means a pharmaceutical composition or preparation comprising a daily dosage or a single dosage of an active ingredient to be administered to a subject in order to obtain a desired result and a pharmaceutically acceptable carrier. The unit dosage form can be prepared into a dosage form known to be suitable for oral or parenteral administration. For example, oral dosage forms include oral tablets, capsules, lozenges, syrups, powders, and suspensions, but are not limited thereto.

[0081] In the present specification, the term "purified" means that when the isolate is isolated, the purity of the isolate is 90% or more, preferably 95% or more, more preferably 99% or more, and most preferably 99.9% or more.

[0082] In this specification, the term "communication disorder" refers to a disease in which there is a problem with communication ability, which may occur in a variety of diseases such as neurological diseases (e.g., brain damage caused by trauma), mental disorders (e.g., panic disorder, post-traumatic stress disorder), autism spectrum disorders, language disorders, etc.

[0083] In the present specification, the term "reciprocal social interaction disorder" includes social (verbal communication or non-verbal communication) disorder, social interaction disorder, social cognition disorder or social relationship disorder.

[0084] Verbal communication disorders include difficulty understanding and using spoken language, difficulty initiating or maintaining a conversation, or repetitive or inflexible use of language (echoespeech). Nonverbal communication disorders refer to: (i) difficulty interpreting and using facial expressions, gestures, eye contact, and body language, (ii) typical use of eye contact and facial expressions, or (iii) difficulty understanding the nonverbal cues of others.

[0085] Impairment in social interactivity refers to: (i) difficulty engaging in social interactions, (ii) difficulty responding appropriately to social overtures from others, or (iii) reduced sharing of interests, emotions, or activities with others.

[0086] Social cognitive impairment refers to: (i) difficulty understanding the perspectives or feelings of others (theory of mind), (ii) difficulty recognizing social cues or nuances of social interactions, or (iii) limited ability to predict and respond to the behavior of others.

[0087] Social relationship impairments include: (i) difficulties developing, maintaining, and understanding relationships, (ii) difficulties making friends or participating in group activities, or (iii) a preference for solitary activities or interactions with a narrow range of interests.

[0088] L-serine or a pharmaceutically acceptable salt thereof

[0089] L-serine used in the present invention is a compound represented by the following Chemical Formula 1.

[0090] [Chemical formula 1]

[0091]

[0092] The pharmaceutically acceptable salt of L-serine is a salt commonly used in the pharmaceutical industry, and may be an inorganic salt, an inorganic acid salt, an organic acid salt, a sulfonate salt, etc. of L-serine.

[0093] Preparation method of L-serine or its pharmaceutically acceptable salt

[0094] One aspect of the present invention relates to a method for producing L-serine, comprising: a) chelating glycine and a copper compound to form a glycine-copper complex; b) reacting the complex of step a) with a carbon source to form a complex into which a hydroxymethyl group is introduced; c) treating the complex of step b) with a reducing agent to form DL-serine; and d) performing optical resolution on the DL-serine of step c) to separate L-serine. One embodiment of the production method of the present invention is Figure 1 Shown in.

[0095] In the present invention, the step a) is a step of chelating glycine and a copper compound to form a glycine-copper complex.

[0096] The step a) can be performed by adding glycine, copper, a base and purified water.

[0097] The copper that forms a complex with glycine in step a) can be a copper complex that is generally used in various industrial preparation methods and is easily available, and can be selected from the group consisting of CuSO 4 , CuOAc 2 , CuCl 2 、CuI、CuBr、CuBr 2 and any one or a mixture of more than one of the group consisting of its hydrates.

[0098] The glycine used in the step a) can be used in an amount of 5kg to 700kg, 10kg to 600kg, 20kg to 500kg, 30kg to 400kg, 40kg to 300kg, 50kg to 200kg, 60kg to 150kg or 70kg to 100kg, depending on the size of the preparation device. The amount of the copper compound can be 0.3 to 1 equivalent, preferably 0.4 to 0.7 equivalent, and most preferably 0.45 to 0.55 equivalent relative to the amount of glycine used.

[0099] The base used in the step a) can be selected from the group consisting of ammonia water, ammonia gas, sodium hydroxide, potassium hydroxide, calcium hydroxide, triethylamine, diethylamine, sodium carbonate, potassium carbonate, sodium bicarbonate, potassium bicarbonate and mixtures thereof. The amount of the base can be 0.6 to 4 equivalents, preferably 0.8 to 3 equivalents, and most preferably 1.0 to 2.5 equivalents relative to the amount of glycine.

[0100] At this time, the amount of purified water can be 6 to 30 times, preferably 4 to 20 times, and most preferably 3 to 10 times the weight of glycine, but is not limited thereto. In the case of excessive use of purified water, the process is difficult to handle, and the amount of wastewater discarded after the process increases, so using the minimum amount is important to ensure economic efficiency.

[0101] In addition, when the temperature is increased, it becomes difficult to form a glycine-copper complex, and the yield decreases. Therefore, the temperature is preferably room temperature to 40°C.

[0102] The glycine-copper complex of step a) may have a structure of the following Chemical Formula 2.

[0103] [Chemical formula 2]

[0104]

[0105] In the present invention, step b) is a step of reacting the complex obtained in step a) with a carbon source to form a complex having a hydroxymethyl group introduced therein.

[0106] The step b) can be performed by adding a carbon source, an alkali and purified water.

[0107] Specifically, the step b) is a process of introducing a carbon into the complex of the step a) and then introducing a hydroxyl group to convert it into a copper complex of racemic serine having a structure of the following chemical formula 3.

[0108] [Chemical formula 3]

[0109]

[0110] The base used in the step b) may be any one or more selected from the group consisting of aqueous ammonia, ammonia gas, sodium hydroxide, potassium hydroxide, calcium hydroxide, triethylamine, diethylamine, sodium carbonate, potassium carbonate, sodium bicarbonate, potassium bicarbonate and mixtures thereof.

[0111] The amount of purified water used in the step b) can be 6 to 25 times, preferably 5 to 20 times, and most preferably 4 to 15 times, compared to the weight of the copper glycine complex having the structure of Chemical Formula 2, but is not limited thereto.

[0112] The carbon source used in the b) step can be selected from any one or more of the group consisting of formaldehyde dilute solution, paraformaldehyde, 1,3-dioxolane, 1.3-dioxane and hexamidine, and can be preferably paraformaldehyde or hexamidine. The formaldehyde can be used as a dilute solution, but not limited to a 15% to 35% aqueous solution. The carbon source can be 1 to 5.0 equivalents, preferably 1.5 to 4.0 equivalents, and most preferably 2.0 to 3.5 equivalents compared to the content of the glycine copper complex having the structure of Chemical Formula 2, but is not limited thereto.

[0113] Although the reaction can be carried out at room temperature, in order to facilitate the reaction, the reaction temperature can be raised to room temperature to below 40°C and stirred. The reaction time can be 1 to 18 hours, preferably 2 to 14 hours, and most preferably 3 to 12 hours.

[0114] In the present invention, step c) is a step of treating the complex obtained in step b) with a reducing agent to form DL-serine.

[0115] The reducing agent used in step c) can be sodium borohydride (NaBH 4 , Sodium borohydride), lithium aluminum hydride (LiAlH 4 , Lithium aluminum hydride) or sodium sulfide (Na 2 S, sodium sulfide), most preferably sodium sulfide. The content of the copper complex of racemic serine having the structure of Chemical Formula 3 can be 1 to 5 equivalents, preferably 1.2 to 4.0 equivalents, and most preferably 1.5 to 3.5 equivalents, but is not limited thereto.

[0116] The solvent used in the step c) may be any one or more selected from the group consisting of purified water, methanol, ethanol, isopropanol, butanol and hexanol, and may preferably be purified water, methanol, ethanol or isopropanol, and most preferably may be purified water or methanol. In addition, when the organic solvent is used, in order to smoothly stir, 10% to 40% of the total amount of the solvent may be mixed for use. The total amount of the solvent used may be 6 to 30 times, preferably 5 to 20 times, and most preferably 4 to 12 times the weight of the copper complex of racemic serine having the structure of Chemical Formula 3, but is not limited thereto.

[0117] Although the reaction can be carried out at room temperature, the reaction temperature may be raised to room temperature to below 50° C. and stirred for a smooth reaction. The reaction time may be 3 to 24 hours, preferably 4 to 18 hours, and most preferably 6 to 12 hours.

[0118] After the step c) is performed, in order to completely remove the copper removed from the complex, a step of reflux cooling using purified water may be further included. The amount of purified water used at this time may be 2 to 10 times, preferably 4 to 7 times, the amount of DL-serine obtained in the step c).

[0119] In the present invention, step d) is a step of optically resolving the DL-serine obtained in step c) to separate L-serine.

[0120] In the step d), in order to selectively obtain only L-serine, various acids as optical resolution-promoting substances may be used to obtain L-serine monoacid salt of the following Chemical Formula 4.

[0121] [Chemical formula 4]

[0122]

[0123] The acid used in the step d) may be any one or more selected from the group consisting of tartaric acid, malic acid, mandelic acid and camphor sulphonic acid, or any one or more selected from the group consisting of (+)-tartaric acid, (-)-malic acid, (-)-mandelic acid and (+)-camphor-10-sulfonic acid. The acid may be 0.85 to 1.5 equivalents, preferably 0.90 to 1.25 equivalents, and most preferably 0.95 to 1.15 equivalents of the racemic DL-serine obtained in the step c). The reaction temperature may be 40 to 100°C, preferably 50 to 90°C, and most preferably 55 to 85°C.

[0124] The reaction solvent in the step d) can be any one or more selected from the group consisting of ethyl acetate, toluene, methyl tert-butyl ether, acetic acid, acetone, tetrahydrofuran, isopropanol, methanol and ethanol, and can be preferably ethyl acetate, acetic acid or isopropanol. The amount of solvent used is 6 to 20 times, preferably 5 to 15 times, and most preferably 4 to 12 times, compared to the weight of DL-serine, but is not limited thereto. The reaction time can be 1 to 12 hours, preferably 2 to 10 hours, and most preferably 3 to 6 hours.

[0125] As a process of filtering the generated L-serine salt and washing the excess acid, any filtration method such as reduced pressure filtration, centrifugal separation, and filter dryer that is feasible in industrial production methods can be used. An organic solvent of 1 to 8 times, preferably 2 to 7 times, and most preferably 3 to 6 times the amount of DL-serine used can be used. At this time, in order to prevent the yield from decreasing, the moisture content of the organic solvent is preferably 1% or less. The organic solvent can be selected from any one or more of the group consisting of acetone, methanol, ethanol, isopropanol, butanol, hexanol, ethyl acetate, toluene, methyl tert-butyl ether, dichloromethane, cyclohexane, and pentane, and can be preferably acetone, methanol, ethanol, and isopropanol.

[0126] In order to obtain only pure L-serine, the process may further include neutralizing the L-serine salt of Chemical Formula 4 with a base, wherein the base may be selected from the group consisting of ammonia water, ammonia gas, sodium hydroxide, potassium hydroxide, calcium hydroxide, triethylamine, diethylamine, sodium carbonate, potassium carbonate, sodium bicarbonate, potassium bicarbonate and mixtures thereof. The pH at this time may be 6 to 7.

[0127] The drying temperature may be between 20°C and 80°C, preferably between 30°C and 70°C, and most preferably between 40°C and 60°C. Vacuum or hot air drying may be used but are not limited thereto.

[0128] Use of L-serine or its pharmaceutically acceptable salt

[0129] In another aspect of the present invention, the present invention relates to the use of serine or a pharmaceutically acceptable salt thereof for preparing a medicament for improving or treating communication disorders and / or reciprocal social interaction disorders in subjects with autism spectrum disorders, wherein the medicament comprises a pharmaceutically acceptable carrier and the medicament is in unit dose form.

[0130] The autism spectrum disorder is known as a neurodevelopmental disorder syndrome with core symptoms such as anxiety and social interaction. Non-limiting examples of autism spectrum disorders include autism, childhood autism, pervasive developmental disorder and Asperger's syndrome, Angelman syndrome, fragile X syndrome, fragile X-associated tremor / ataxia syndrome (FXTAS), Rett syndrome, Landau-Klevner syndrome, Prader-Willi syndrome, tardive dyskinesia and Williams syndrome, but are not limited thereto.

[0131] In addition, the subject with autism spectrum disorder can meet the autism spectrum disorder requirements of DSM-5 (Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition, Text Revision) and / or ADI-R (Autism Diagnostic Observation Inventory-Revised).

[0132] In one embodiment of the present invention, the autism spectrum disorder may be childhood autism. The childhood autism is a disease name classified as Korean disease classification code F84.0 or International Classification of Disease (ICD) F84.0.

[0133] In another embodiment of the present invention, the subject's clinical global impression-severity scale (CGI-S) is above 5, and the Vineland Adaptive Behavior Scales-II (VABSII) antisocial behavior scale (ASB) is above 48.

[0134] In one embodiment of the present invention, the subject may be an adult, or the subject may be 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17 or 18 years old, preferably under 18, under 13, under 11, under 7, less than 7, 2 to 7 or 2 to 6 years old. In addition, the individual may be a child, and the age may be 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17 or 18 years old, or may be under 18, under 13, under 11, under 7, less than 7, 2 to 7 or 2 to 6 years old.

[0135] In another embodiment of the present invention, the subject's body weight may be 10 kg to 100 kg, 10 kg to 90 kg, 10 kg to 80 kg, 10 kg to 70 kg or 10 kg to 60 kg.

[0136] In one embodiment of the present invention, the unit dose of L-serine or a pharmaceutically acceptable salt thereof may be a daily dosage or a single dosage of L-serine or a pharmaceutically acceptable salt thereof.

[0137] In another embodiment of the present invention, the daily dosage of L-serine or a pharmaceutically acceptable salt thereof can be selected from 1g, 2g, 3g, 4g, 5g, 6g, 7g, 8g, 9g, 10g, 11g, 12g, 13g, 14g, 15g, 16g, 17g, 18g, 19g, 20g, 21g, 22g, 23g, 24g, 25g, 26g, 27g, 28g, 29g, 30g, 31g, 32g, 33g, 34g, 35g, 36g, 37g, 38g, 39g, 40g, 41g, 42g, 43g, 44g, 45g, 46g, 47g, 48g, 49g, 50g, 51g, 52g 98g, 99g, 100g, 101g, 102g, 103g, 104g, 105g, 106g, 107g, 108g, 109g, 110g, 111g, 112g, 113g, 114g, 115g, 116g, 117g, 118g, 119g, 120g, 121g, 122g, 123g, 124g, 125g, 126g, 127g, 128g, 129g, 130g, 131g, 132g, 133g, 134g, 135g, 136g, 137g, 138g, 139g, 140g, 141g, 142g, 143g In addition, according to the body weight of the individual, the daily dosage of the L-serine or its pharmaceutically acceptable salt can be 2g, 4g, 7g, 8g, 10g, 12g 14g, 20g or 28g, and the total daily dosage of the L-serine or its pharmaceutically acceptable salt can be the daily dosage for each body weight described in Table 4 and Table 15.

[0138] In one embodiment of the present invention, the daily dosage of L-serine or a pharmaceutically acceptable salt thereof may be 2 g to 15 g, or 2 g to 14 g, or 4 g to 30 g, or 4 g to 28 g.

[0139] In another embodiment of the present invention, the daily dosage of the L-serine or its pharmaceutically acceptable salt can be 100mg / kg to 600mg / kg, or can be 140mg / kg to 580mg / kg, 140mg / kg to 572mg / kg, 200mg / kg to 400mg / kg, 280mg / kg to 580mg / kg or 280mg / kg to 572mg / kg. In addition, the daily dosage of the L-serine or its pharmaceutically acceptable salt can be 180mg / kg, 300mg / kg, 480mg / kg or 600mg / kg. Further, according to the body weight of the individual, the total daily dosage of the L-serine or its pharmaceutically acceptable salt can be about 285.7 mg / kg to about 400 mg / kg, about 333.3 mg / kg to about 533.3 mg / kg, about 378.4 mg / kg to about 560 mg / kg, about 392.2 mg / kg to about 526.3 mg / kg, about 466.7 mg / kg to about 538.5 mg / kg, about 142.9 mg / kg to about 200 mg / kg. kg, about 166.7 mg / kg to about 266.7 mg / kg, about 189.2 mg / kg to about 280 mg / kg, about 196.1 mg / kg to about 263.2 mg / kg, about 233.3 mg / kg to about 269.2 mg / kg, or about 200 mg / kg to about 400 mg / kg, or about 285 mg / kg to about 400 mg / kg, about 333 mg / kg to about 534 mg / kg, about 378 mg / kg to about 5 The dosage may be about 300 mg / kg, about 200 mg / kg to about 400 mg / kg, about 150 mg / kg to about 200 mg / kg, about 166 mg / kg to about 267 mg / kg, about 189 mg / kg to about 280 mg / kg, about 196 mg / kg to about 264 mg / kg, about 233 mg / kg to about 270 mg / kg, about 200 mg / kg to about 400 mg / kg, or about 300 mg / kg to about 400 mg / kg. 07.7 mg / kg to 400 mg / kg, about 400 mg / kg to about 571.4 mg / kg, about 352.9 mg / kg to about 571.4 mg / kg, about 408.2 mg / kg to about 571.4 mg / kg, about 280 mg / kg, about 311.1 mg / kg, about 350 mg / kg, about 400 mg / kg, about 466.7 mg / kg or about 560 mg / kg can be the daily dosage for each body weight recorded in Tables 4 and 15.

[0140] In another embodiment of the present invention, L-serine or a pharmaceutically acceptable salt thereof can be administered to the subject more than once a day, once a day, twice a day, three times a day, four times a day, or five times a day, and can be preferably administered twice a day.

[0141] In another embodiment of the present invention, the single administration amount of L-serine or a pharmaceutically acceptable salt thereof may be 70 mg / kg to 280 mg / kg, 140 mg / kg to 290 mg / kg or 140 mg / kg to 286 mg / kg. In addition, the L-serine or its pharmaceutically acceptable salt can be administered in an amount of about 142.9 mg / kg to about 200 mg / kg, about 166.7 mg / kg to about 266.7 mg / kg, about 189.2 mg / kg to about 280 mg / kg, about 196.1 mg / kg to about 263.2 mg / kg, about 233.3 mg / kg to about 269.2 mg / kg, about 71.4 mg / kg to about 100 mg / kg, about 83.3 mg / kg to about 133.3 mg / kg, about 94.6 mg / kg to about 140 mg / kg, about 98 mg / kg to about 131.6 mg / kg, or about 116.7 mg / kg to about 134.6 mg / kg, or can be about 142 mg / kg to about 200 mg / kg, about 166 mg / kg to about 267 mg / kg, about 189 mg / kg to about 280 mg / kg. kg, about 196 mg / kg to about 264 mg / kg, about 233 mg / kg to about 270 mg / kg, about 71 mg / kg to about 100 mg / kg, about 83 mg / kg to about 134 mg / kg, about 94 mg / kg to about 140 mg / kg, about 98 mg / kg to about 132 mg / kg, about 116 mg / kg to about 135 mg / kg, about 153.8 mg / kg to about 200 mg / kg, about 200 mg / kg to about 285.7 mg / kg, about 176.5 mg / kg to about 285.7 mg / kg, about 204.1 mg / kg to about 285.7 mg / kg, about 140 mg / kg, about 155.6 mg / kg, about 175 mg / kg, about 200 mg / kg, about 233.3 mg / kg or about 280 mg / kg can be the single dose for each body weight described in Tables 4 and 15.

[0142] In one embodiment of the present invention, the medicament comprising L-serine or a pharmaceutically acceptable salt thereof can be a non-oral dosage form or an oral dosage form, and can be preferably an oral dosage form. For the oral dosage form, as long as it is a known oral dosage form in the technical field to which this case belongs, there is no restriction (for example, oral tablets, capsules, lozenges, syrups, powders or suspensions, etc.). The medicament can be a syrup dosage form.

[0143] In another embodiment of the present invention, the medicament may include one or more of a sweetener, a thickener, a pH adjuster, a solvent, a preservative and a flavoring agent.

[0144] As for the sweetener, any substance can be used as long as it is a substance used in the technical field to improve the taste, stability and miscibility of pharmaceutical compositions. It can include but is not limited to sorbitol, maltodextrin, fructose, glycerol, propylene glycol, peanut oil, dehydrated sorbitan, stevia, white sugar, etc. Preferably, one or more sweeteners selected from white sugar, sorbitol, acesulfame potassium and sucralose can be used.

[0145] As for the thickener, any substance can be used as long as it is a substance used in the technical field to improve the stability and consistency of the pharmaceutical composition to which the present invention belongs. It can be used but not limited to carboxymethyl cellulose (CMC), hydroxypropyl methyl cellulose (HPMC), polyvinyl pyrrolidone, polyethylene oxide, gum arabic, sorbitol, polysorbate, etc., preferably carboxymethyl cellulose can be used.

[0146] As for the pH adjuster, any substance can be used as long as it is a substance used in the technical field to adjust the pH of the pharmaceutical composition to which the present invention belongs. It can be used but not limited to alkaline pH adjusters, acidic pH adjusters, phosphate buffer, sodium bicarbonate, ammonia solution, sodium chloride, etc., preferably, an acidic pH adjuster can be used, and more preferably, citric acid, acetic acid, carbonic acid, lactic acid, etc. can be used.

[0147] In the present invention, the solvent included in the preparation containing L-serine or a pharmaceutically acceptable salt thereof is a commonly used solvent in the technical field to which the present invention belongs, and purified water, ethanol solvent, acetone, etc. can be used, preferably purified water.

[0148] As for the preservative, any substance can be used as long as it is a substance used to maintain the stability and effectiveness of the pharmaceutical composition in the technical field to which the present invention belongs. Sodium benzoate, benzyl alcohol, parahydroxybenzoate, formaldehyde, etc. can be used, and preferably methyl parahydroxybenzoate and / or propyl parahydroxybenzoate can be used.

[0149] As for the flavoring agent, any substance can be used as long as it is a substance used in the technical field to which the invention belongs to give fragrance to pharmaceutical compositions or improve user experience, and peppermint oil, lemon essential oil, vanilla essence, spices, etc. can be used.

[0150] In another embodiment of the present invention, the concentration of L-serine or a pharmaceutically acceptable salt thereof contained in the preparation may be 10 mg / ml to 500 mg / ml.

[0151] In one embodiment of the present invention, the preparation may be in unit dosage form with each volume ranging from 10 ml to 100 ml.

[0152] In another embodiment of the present invention, the content of L-serine or a pharmaceutically acceptable salt thereof contained in the unit dosage form may be 1 g to 100 g, 1 g to 60 g, 2 g to 30 g, 4 g to 30 g, 2 g to 28 g or 1 g to 14 g.

[0153] In another embodiment of the present invention, the L-serine contained in the preparation can be prepared by the following method:

[0154] a) a step of chelating glycine and a copper compound to form a glycine-copper complex;

[0155] b) reacting the complex obtained in step a) with a carbon source to form a complex into which a hydroxymethyl group is introduced;

[0156] c) treating the complex obtained in step b) with a reducing agent to form DL-serine; and

[0157] d) a step of optically resolving the DL-serine obtained in step c) to separate L-serine.

[0158] The materials used in the steps a) to d) and the details of each step are as described above.

[0159] Preparation and kit containing L-serine or a pharmaceutically acceptable salt thereof

[0160] One aspect of the present invention relates to a preparation comprising L-serine or a pharmaceutically acceptable salt thereof, which can be in a unit dosage form.

[0161] In one embodiment of the present invention, the preparation is a preparation comprising 1 g to 100 g of L-serine or a pharmaceutically acceptable salt thereof, a thickener, a pH adjuster, a preservative and a solvent, and the preparation may be in a unit dosage form.

[0162] In another embodiment of the present invention, it may be characterized in that each volume of the preparation is 10 ml to 100 ml.

[0163] In another embodiment of the present invention, the preparation can be administered to improve or treat the communication disorder and / or social interaction disorder of the experimenter with autism spectrum disorder.At this moment, 1 daily dosage or 1 dosage of the L-serine or its pharmaceutically acceptable salt administered to the experimenter is as mentioned above.

[0164] Another aspect of the present invention relates to a kit comprising a preparation comprising 1 g to 100 g of L-serine or a pharmaceutically acceptable salt thereof, the preparation being used to improve or treat a communication disorder and / or social interaction disorder in a subject with an autism spectrum disorder, the preparation being in unit dosage form. The kit may include instructions describing a method for administering the preparation comprising the L-serine or a pharmaceutically acceptable salt thereof to a subject.

[0165] The present invention will be described in more detail below according to the following examples. However, the following examples are only used to illustrate the present invention, and the scope of the present invention is not limited thereto.

[0166] Preparation Example 1. Preparation of L-glycine copper complex

[0167] Preparation Example 1.1. Using anhydrous CuSO 4 Preparation of L-glycine copper complex

[0168] 75.0 kg of glycine (source: EOS, China) and 1000 L of purified water were added to the reactor at room temperature and stirred for 10 minutes. Then, anhydrous CuSO 4 (Source: China Yongchun Applied Materials Co., Ltd.) 156kg and NaOH (Source: Xingchang Chemical) 80kg were stirred at room temperature for 6 hours. After the reaction was completed, the temperature was lowered to about 10°C, the solid formed was filtered under reduced pressure, and then washed 3 times with 100kg of purified water. The blue solid as the target compound was obtained by vacuum drying at 50°C for 10 hours (yield: 83.5%).

[0169] Preparation Example 1.2. Using 5-hydrate CuSO 4 (CuSO 4 ·H 2 O) Preparation of L-glycine copper complex

[0170] 75.0 kg of glycine (source: EOS, China) and 1200 L of purified water were added to the reactor at room temperature and stirred for 10 minutes. Then, CuSO 4 ·H 2 O (source: China Yongchun Applied Materials Co., Ltd.) 249kg and NaOH (source: Xingchang Chemical) 80kg were stirred at room temperature for 6 hours. After the reaction was completed, the temperature was lowered to about 10°C, the solid formed was filtered under reduced pressure, and then washed 3 times with 100kg of purified water. It was vacuum dried at 50°C for 10 hours to obtain a blue solid as the target compound (yield: 82.0%).

[0171] Preparation Example 2. Preparation of DL-serine copper complex

[0172] Preparation Example 2-1. Preparation of DL-serine copper complex

[0173] 60 kg of the copper glycine complex obtained in Preparation Example 1, 15 kg of paraformaldehyde (source: EOS, China) and 500 kg of purified water were added to the reactor at room temperature and stirred for 10 minutes. Then, 45 kg of sodium hydroxide (NaOH) was slowly added while maintaining the room temperature. After stirring at room temperature for 1 hour, the temperature was raised to about 40°C and stirred for 12 hours, filtered and cooled to about room temperature. After the formed solid was filtered under reduced pressure, it was washed 3 times with 100 kg of purified water. After vacuum drying at 50°C for 10 hours, a blue solid as the target compound was obtained (yield: 77.0%).

[0174] Preparation Example 2-2. Preparation of DL-serine copper complex

[0175] The target compound was obtained as a blue solid (yield: 74.0%) in the same manner as in Example 2-1, except that 12.5 kg of Hexamidine (imported by Rayleigh Chemical, China) was used instead of paraformaldehyde in Preparation Example 2-1.

[0176] Preparation Example 3. Preparation of DL-serine

[0177] 30 kg of the substance obtained in Preparation Example 2 and 240 kg of purified water were added to the reactor at room temperature and stirred for 10 minutes. Then, sodium sulfide (Na 2 S) (Source: China EOS Company) 8.8kg. After stirring at room temperature for 2 hours, stir at about 45°C for 8 hours, and then cool to about 5°C. After adding the formed solid to 150kg of purified water, increase the temperature and reflux for 2 hours, then slowly lower the temperature and cool to about 5°C. After vacuum filtration, wash once with 10kg of purified water and twice with 20kg of ethanol. Vacuum dry at 50°C for 10 hours to obtain a white solid as the target compound (yield: 79%).

[0178] Preparation Example 4. Preparation of L-serine

[0179] Preparation Example 4.1. Preparation of L-serine using camphorsulfonic acid

[0180] 10 kg of DL-serine and 23.2 kg of L-(-)-camphorsulfonic acid (source: China EOS) obtained in Preparation Example 3 were added to the reactor, and 100 kg of acetic acid (source: Mita Junyao) was added for dissolution. Heat to about 80°C to 90°C and stir for 3 hours. The mixture was then cooled to about 5°C to 10°C, stirred for 1 hour, and filtered to obtain a white solid L-serine-camphorsulfonate. The obtained solid was dissolved in 120 L of anhydrous ethanol, 28 to 30% ammonia water was added, and the pH was adjusted to about 6.0 to 7.0. The reactants were stirred at room temperature for 1.5 hours, the temperature was lowered to about 5°C to 10°C, and further stirred for 3 hours and filtered to obtain a crude L-serine product as a white solid. Then, 15 kg of the crude product, 1 kg of anhydrous ethanol and purified water were added, stirred at room temperature for 2 hours, filtered, and washed with 2 kg of anhydrous ethanol. After vacuum drying at 50°C for 6 hours, 4.97 kg of L-serine was obtained. The chemical purity and optical purity of the product were measured to be 99.5% and 99.7%, respectively.

[0181] Preparation Example 4.2. Preparation of L-serine using tartaric acid

[0182] Using (+)-tartaric acid instead of camphorsulfonic acid, 4.87 kg of L-serine was obtained in the same manner as in Preparation Example 4.1. The chemical purity and optical purity of the product were measured to be 99.4% and 99.7%, respectively.

[0183] Experimental Example 1. Confirmation test of L-serine

[0184] The L-serine obtained in Preparation Example 4 was analyzed based on the L-serine analysis method specified in the United States Pharmacopoeia, and it was confirmed that all of them met the requirements.

[0185] The L-serine obtained in Preparation Example 4 was tested by infrared spectroscopy (ATR method) in the Korean Pharmacopoeia General Test Method. It was confirmed that the L-serine at 2930, 1676, 1439, 1389, 1257, 1093, 658 cm -1 Main absorption was observed near each frequency, and it was confirmed that when L-serine was dissolved in water, 1 mL of ninhydrin solution (1->1000) was added to 5 mL and heated for 3 minutes, the solution exhibited purple color.

[0186] In addition, the obtained L-serine was analyzed by differential scanning calorimetry (DSC), and the result confirmed that an endothermic peak appeared at 226°C, which is consistent with the literature value (refer to Figure 2 ).

[0187] Preparation Example 5. Preparation of a preparation containing L-serine

[0188] Preparation Example 5.1. Preparation of oral syrup

[0189] A preparation containing the L-serine obtained in Preparation Example 4 (containing 100 mg / ml L-serine, sodium carboxymethylcellulose, methyl parahydroxybenzoate, propyl parahydroxybenzoate, citric acid hydrate, potassium citrate hydrate, white sugar, D-sorbitol solution, apple mint flavor SJ-G (22005221), and purified water) was prepared. The preparation contains 10 g of the main ingredient L-serine per 100 mL, and is a sweet colorless syrup. The preparation can be packaged in aluminum foil at a dosage of 2 g / 20 mL.

[0190] The results of the 36-month stability test (room temperature (25°C)) of the preparation are shown in Figure 3 The preparation had no changes in properties during the 36 months, and the content of the main component L-serine, the pH of the preparation and the specific gravity did not change much (refer to Figure 3 ).

[0191] Preparation Example 5.2. Stability of a composition containing high concentration of L-serine

[0192] After the purified water is poured into the beaker, the temperature is raised to above 50°C (about 60°C). 0.5mg / ml to 100mg / ml of thickener (agar, bentonite, carbomer, sodium carboxymethylcellulose, carrageenan, sodium microcrystalline carboxymethylcellulose, guar gum, hydroxyethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methylcellulose, pectin, polyethylene oxide, povidone, corn starch, potato starch, wheat starch, xanthan gum or gelatin) is added to the purified water, and the temperature is raised or stirred (maintained at above 50°C). After the thickener is dissolved, a preservative, a pH adjuster (citric acid, acetic acid, carbonic acid or lactic acid) and an acidulant are added to adjust the pH to 5.0 to 7.0. Then, after cooling to a temperature below 30° C., L-serine (50 mg / ml, 75 mg / ml, 100 mg / ml, 250 mg / ml, 300 mg / ml or 500 mg / ml) obtained in Preparation Example 4 and a sorbitol solution as a diluent were added. After the L-serine and the diluent were dissolved, purified water was added.

[0193] The stability changes of the preparations were observed under 4-week accelerated stability test conditions (40±2°C, RH 75±5%). As a result, no changes in properties were observed in all preparations, and the change rate of L-serine content was only 0.31% to 2.8%, indicating excellent preparation stability.

[0194] Example 1. In vivo autism spectrum disorder animal model experiment

[0195] Example 1.1. Effects of improving sociality, social cognition and anxiety

[0196] Pregnant C57BL / 6 mice were subcutaneously injected with VPA (valproic acid) on gestational day 12. Among the mice born from pregnant mice to which VPA was subcutaneously administered, male mice were selected for use as autism spectrum disorder mouse model animals.

[0197] The critical period for mice is 3 weeks of age, so social experiments, social cognition experiments, and anxiety experiments were conducted on 3-week-old normal mice and 3-week-old autism spectrum disorder mice to confirm baseline characteristics. 500 mg / kg of L-serine (hereinafter referred to as AST-001) was orally administered to autism spectrum disorder mice with confirmed baseline characteristics during 2 weeks. Social experiments, social cognition experiments, and anxiety experiments were performed again on autism spectrum disorder mice that were orally administered AST-001 for 2 weeks.

[0198] Social Experiment

[0199] A social object (strange mouse) and a non-social object (empty cage) were placed in the two rooms of a 3-chamber, and a behavioral experiment was conducted to confirm whether the experimental subject mouse moved toward the social object (refer to Figure 4 A). Starting from 3 weeks of age, water was orally administered once a day to the normal group and the autism spectrum disorder group for 2 weeks, and AST-001 500 mg / kg (dissolved in water) was orally administered once a day to the autism spectrum disorder mice in the drug group. 5-week-old normal mice (n=6) and 5-week-old autism spectrum disorder mice (n=6) were used in the experiment. 5-week-old normal mice showed a social preference index for moving in the direction of social objects, while 5-week-old autism spectrum disorder mice did not show a social preference index. However, in the behavioral experiment of the group where AST-001 was orally administered for 2 weeks, it was confirmed that autism spectrum disorder mice had a social preference index that preferred social objects more (refer to Figure 4 C and Figure 4 Specifically, the autism spectrum disorder mice that were not administered AST-001 had little difference in the time they spent interacting with social objects and the time they spent interacting with empty cages, and this difference was not statistically significant, and they had a low preference for social objects (refer to Figure 4 C and Figure 4 In contrast, the control group and ASD mice treated with AST-001 500 mg / kg spent significantly more time interacting with social objects than with the empty cage, and this difference was statistically significant, indicating a high preference for social objects (refer to Figure 4 C and Figure 4 D).

[0200] Social cognition experiment

[0201] A social cognition experiment was conducted to measure whether the subject mouse moved toward the unfamiliar mouse when placed between an unfamiliar mouse and a familiar mouse (refer to Figure 4 B). Starting from 3 weeks of age, during a period of 2 weeks, the normal group and the autism spectrum disorder group were orally given water once a day, and the autism spectrum disorder mice in the drug group were orally administered AST-001 500 mg / kg (dissolved in water) once a day. 5-week-old normal mice (n=6) and 5-week-old autism spectrum disorder mice (n=6) were used in the experiment. 5-week-old normal mice showed a greater preference for moving in the direction of unfamiliar mice, while 5-week-old autism spectrum disorder mice did not show a preference for unfamiliar mice. However, in the social cognition experiment of the group where AST-001 was orally administered for 2 weeks, the preference of autism spectrum disorder mice for unfamiliar mice increased (refer to Figure 4 E and Figure 4 Specifically, the autism spectrum disorder mice that were not administered AST-001 had little difference in the interaction time with familiar mice and the interaction time with unfamiliar mice, and this difference was not statistically significant, and the preference for unfamiliar mice was low (refer to Figure 4 E and Figure 4 In contrast, the control group and the autism spectrum disorder mice treated with AST-001 500 mg / kg spent significantly more time interacting with the unfamiliar mice than with the familiar mice, and this difference was statistically significant, indicating a high preference for the unfamiliar mice (refer to Figure 4 E and Figure 4 F). That is, it was confirmed that oral administration of AST-001 improved the social cognition of mice with autism spectrum disorders.

[0202] Anxiety experiment

[0203] An anxiety test was conducted to determine whether the experimental mice showed anxiety behavior of avoiding the open arm space when placed in an elevated plus maze (refer to Figure 5 A). Starting from 3 weeks of age, during 2 weeks, the normal group and the autism spectrum disorder group were orally given water once a day, and the autism spectrum disorder mice in the drug group were orally administered AST-001 500mg / kg (dissolved in water) once a day. 5-week-old normal mice (n=6) and 5-week-old autism spectrum disorder mice (n=6) were used in the experiment. 5-week-old autism spectrum disorder mice showed a greater tendency to avoid open spaces than 5-week-old normal mice. However, in the anxiety experiment of the group where AST-001 was orally administered for 2 weeks, the anxiety level of the autism spectrum disorder mice decreased, and the time they wanted to go to the open space increased (refer to Figure 5 B to Figure 5 Specifically, the autism spectrum disorder mice that were not administered AST-001 spent very little time in the open space (open arm) of the cross maze and spent more time in the closed space (closed arm) of the cross maze (refer to Figure 5 C and Figure 5 However, the control group and the autism spectrum disorder mice treated with AST-001 500mg / kg spent significantly longer time in the open space of the cross maze than the autism spectrum disorder mice not treated with AST-001, and spent less time in the closed space of the cross maze (refer to Figure 5 C and Figure 5 D). Therefore, it was confirmed that oral administration of AST-001 improved the anxiety level of mice with autism spectrum disorders.

[0204] Example 1.2. Effect of normalizing dopamine secretion

[0205] The autism spectrum disorder mouse model was established by the method described in Example 1.1. 3-week-old male autism spectrum disorder mice were divided into a group (n=6) that was orally administered with AST-001 250 mg / kg for 2 weeks and a group (n=6) that was orally administered with AST-001 500 mg / kg for 2 weeks. AST-001 was orally administered. The 3-week-old male normal mouse group and the autism spectrum disorder mouse group (n=6) were used as the control group (n=6), and water was orally administered once a day for 2 weeks for behavioral experiments.

[0206] Spontaneous action potentials of dopaminergic neurons in the midbrain VTA region of 5-week-old normal mice and 5-week-old male autism spectrum disorder mice were confirmed by patch clamp. Compared with normal mice, autism spectrum disorder mice showed a decrease in spontaneous action potential discharge rate (refer to Figure 6 A). Therefore, it can be seen that the dopamine secretion of mice with autism spectrum disorders is reduced, which indicates that behavioral abnormalities related to the dopamine pathway, such as attention, reward, and cognition, occur.

[0207] However, after 2 weeks of oral administration of AST-001, the spontaneous action potential discharge rate of mice with autism spectrum disorder was similar to that of normal mice (ref. Figure 6 A and Figure 6 B). Therefore, it was confirmed that administration of 250 mg / kg to 500 mg / kg of AST-001 increased dopamine secretion in autism spectrum disorder mice.

[0208] Example 1.3. Normalization effect of synaptic pruning

[0209] A mouse model of autism spectrum disorder was established by the method described in Example 1.1. Autism spectrum disorder is known to be caused by errors in synaptic pruning during critical periods, resulting in excessive growth of dendrites beyond the required period. Therefore, an experiment was conducted to confirm whether the administration of AST-001 can normalize the number of dendrites. For reference, the confocal microscopy observation results of the cortical region can be found in Figure 7 A.

[0210] We examined whether the dendritic spine density of 3-week-old male autism spectrum disorder mice (n=8) reached a level similar to that of normal male control mice (n=3) after oral administration of AST-001 400 mg / kg for 2 weeks. The number of dendrites in autism spectrum disorder mice was significantly reduced, reaching a level similar to that of the control group (refer to Figure 7 B and Figure 7 C). Thus, it was confirmed that oral administration of AST-001 normalized synaptic pruning during the critical period of autism spectrum disorder.

[0211] Example 1.4. Effect of normalizing nerve cell activity

[0212] Autism spectrum disorder model animals BTBR T+tf / J (BTBR) and C57BL / 6 mice (used as a control group) were purchased from Jackson Laboratory. When the intrinsic excitability of mPFC pyramidal neurons was measured by the action potential discharge rate based on current injection using patch clamp, the action potential discharge rate of BTBR mice was reduced compared with the control group (refer to Figure 8 However, after oral administration of 500 mg / kg of AST-001 to BTBR mice for 2 weeks, the action potential discharge rate was measured, showing that the action potential discharge rate of BTBR mice and control mice was at a similar level (refer to Figure 8 B).

[0213] Example 1.5. Effect of improving individual recognition and spatial memory

[0214] Spatial memory experiment

[0215] The autism spectrum disorder mouse model was established in the same manner as in Example 1.1. After 3-week-old male autism spectrum disorder mice were orally administered 180 mg / kg, 300 mg / kg, 480 mg / kg or 600 mg / kg of L-serine over a 28-day period, a spatial memory experiment was performed. Normal mice were used as a control group. Eight mice were used in each experimental group.

[0216] The Y-maze test was used to evaluate the spatial working memory of mice. The spontaneous change value, which is a measure of spatial memory, increased with increasing doses of L-serine (refer to Fig. 9 ).

[0217] Individual identification experiment

[0218] The autism spectrum disorder mouse model was established in the same manner as in Example 1.1. Three-week-old male autism spectrum disorder mice were orally administered 180 mg / kg, 300 mg / kg, 480 mg / kg, or 600 mg / kg / day of L-serine over a period of 21 days, and then an individual recognition experiment was performed. Eight mice were used in each experimental group.

[0219] Since mice tend to be more interested in novel objects than familiar objects, a NOR (Novel object recognition) experiment was conducted to confirm whether mice can recognize novel individuals.

[0220] The higher the dosage of AST-001, the higher the DI (discrimination index) value of autism spectrum disorder mice in recognizing / memorizing new objects (refer to Fig.10 ).

[0221] Example 2. Pharmacokinetic Experiment

[0222] Example 2.1. Pharmacokinetics experiment in beagle dogs

[0223] 4g or 8g of L-serine were orally or intravenously administered to beagles (1.8-2.6 years old, 8.6-11.6kg, n=20). Blood was collected before (baseline blood collection) and after (main blood collection) administration of L-serine. During the experiment, beagles were kept in a fasting state, and the plasma L-serine concentration was analyzed by LC-MS / MS.

[0224] The plasma concentration-time curves of L-serine after oral or intravenous administration to beagle dogs are shown in Fig.11 shown.

[0225] The area under the plasma concentration-time curve (AUC) was calculated from the plasma drug concentration curves at different times after oral administration of 4 g or 8 g of L-serine. inf ) were 627.7±108.4μg·hr / mL and 1329.0±317.8μg·hr / mL, respectively. The drug elimination half-life (t 1 / 2) were 3.8±1.4 hours and 4.7±1.8 hours respectively. The bioavailability of oral administration was 106.5% and 84.2% after 4g and 8g administration per individual respectively.

[0226] Example 2.2. Pharmacokinetics experiment in adult males

[0227] Healthy adult males were used as subjects and pharmacokinetic experiments were performed according to the experimental design shown in Table 1. Blood was collected 24 hours before administration of AST-001 (L-serine main component) and the concentration of L-serine was measured. AST-001 powder 60g is a white homogeneous powder without foreign matter, packaged in HDPE bottles or bottles of the same material, and the composition of the drug is that each 1000mg dose contains 1000mg of the main component L-serine.

[0228] [Table 1]

[0229]

[0230] The baseline L-serine in the 24 hours before AST-001 administration generally did not show a time-dependent pattern, and no tendency to change over time during the day was observed. After a single oral administration of AST-001 10g, 15g, 20g, or 30g, after systemic exposure to L-serine, Cmax, AUClast, and AUCinf showed a linear increase in the pattern as the AST-001 dose increased. On the other hand, when AST-001 was repeatedly administered 15g twice a day for 7 days, a steady state was reached, with an average half-life of about 11 hours. Compared to a single administration of AST-001 15g, the average accumulation index was about 1.7 times higher in the steady state.

[0231] After comprehensive safety evaluations including adverse reactions, clinical laboratory tests, vital signs, electrocardiograms, and physical examinations, the results showed that AST-001 has excellent safety and drug tolerance in healthy adults with a single oral administration of up to 30 g per day and repeated oral administration of 15 g twice a day for 7 days. Therefore, it was confirmed that AST-001 can be safely administered within the dose range of 10g-30g per day for indications requiring repeated administration.

[0232] The results of the pharmacokinetic analysis of AST-001 in the repeated-dose study in healthy adult males are shown in Table 2 below.

[0233] [Table 2]

[0234]

[0235]

[0236] Tmax: Median,

[0237] C max , AUC tau ,t 1 / 2 , CL ss / F and Vz / F: Mean ± SD

[0238] Example 2.3. Population pharmacokinetic simulation

[0239] The population pharmacokinetic model of AST-001 was established by applying zero-order absorption and first-order elimination to the two-compartmental model, and the residual model used a proportional error model. For baseline L-serine, the model was reflected in the form of additional injection of L-serine in a steady state. The following pharmacokinetic parameters were reflected in the model by applying allometric scaling for body weight.

[0240] V1=V1×(weight / 70)1

[0241] V2=V2×(body weight / 70)1

[0242] Q=Q×(weight / 70)0.75

[0243] CL=CL×(weight / 70)0.75

[0244] After 15 g BID administration to healthy adults, the AUC tau (about 882.8h*μg / mL) was set as the target exposure. Population pharmacokinetic simulation analysis was performed using the NONMEM version 7.4 program, and the FOCEINTERACTION option was used to analyze the blood sampling time and L-serine blood concentration data.

[0245] In addition, the analysis results showed that the baseline L-serine showed little change over time and was detected to be constant within the concentration range of 5-30μg / mL. The average AUC of L-serine without baseline correction after a single dose of AST-001 10g, 20g, and 30g last The average AUC of L-serine without baseline correction was 613.3 h*μg / mL, 1134.6 h*μg / mL, and 1525.5 h*μg / mL, respectively. When 15 g of AST-001 was administered twice a day for 1 week, tau The average half-life of L-serine exogenously administered using AST-001 is 6.5 to 14.0 hours.

[0246] When simulating the dose and usage for each weight range, the exposure of the recommended usage of 400 mg / kg and 100 mg / kg is within the target AUC tau Taking into account the drug exposure observed in clinical trials conducted on healthy adults, the analysis showed that the 4g / day to 28g / day dose of AST-001 in children showed similar effects to the 400mg / kg / day dose of AST-001 (see Table 3).

[0247] [Table 3]

[0248]

[0249] Example 3. Phase II clinical trial

[0250] A multicenter, randomized, double-blind, placebo-controlled, treatment exploratory Phase II clinical trial was conducted on 151 children with autism spectrum disorders to explore the efficacy and safety of AST-001 syrup. Fig.12 shown.

[0251] The AST-001 syrup used in the clinical trial is a colorless or light yellow clear syrup with a fragrance and sweet taste, which is the preparation prepared in the preparation example 5.1. As a placebo, a syrup of the same properties was used. AST-001 syrup-placebo 100mL does not contain the main ingredient. Although it is prepared with the same components as the test drug, in order to show a taste similar to the test drug, more sweetener (white sugar) is added than the test drug.

[0252] Subjects who were judged to be suitable for participation in this clinical trial were randomly assigned to the placebo group, high-dose group, and low-dose group at a ratio of 1:1:1 during the main study administration period (week 0 to week 12). Patients assigned to the high-dose group and the low-dose group were orally administered AST-001 syrup for 12 weeks using the administration method and dosage according to different body weights as shown in Table 4. Subjects who received the high dose were administered with a dose equivalent to the low dose during the first 2 weeks, and then the dose was increased to the high dose to confirm safety.

[0253] [Table 4]

[0254]

[0255]

[0256] The control group receiving the 12-week placebo administration received AST-001 syrup administration at a high dose in Table 4 during the additional 12-week extension study (low dose administration for the first 2 weeks, high dose administration after safety confirmation), and the corresponding group was classified as a high-dose short-term administration group. In the main study, patients who were administered a high dose of AST-001 syrup continued to receive a high-dose AST-001 administration and were classified as a high-dose long-term administration group. In the main study, patients who were administered a low dose of AST-001 syrup continued to receive a low-dose AST-001 syrup administration and were classified as a low-dose long-term administration group. After the 12-week extension study period, the effectiveness and abnormal reactions of the patients during the 12-week (i.e., follow-up period) were tracked and observed.

[0257] The baseline characteristics of the patients participating in the clinical trial (FAS population) are shown in Table 5.

[0258] [Table 5]

[0259]

[0260]

[0261] a: Analysis was performed using Fisher's exact test.

[0262] Effectiveness

[0263] The main efficacy evaluation variable in the Phase II clinical trial of AST-001 syrup is the change in the K-VABS-II ABC score at 12 weeks after administration compared to the baseline. The K-VABS-II is an appropriate and reproducible assessment tool for evaluating the functional adaptation level of autism spectrum disorders, consisting of 4 main areas of communication, life skills, sociality, and motor skills and an optional maladaptive behavior indication area. Each question consists of 3 Likert scales (0 points: completely unable. 1 point: sometimes or partially. 2 points: generally able). The main evaluation variable K-VABS-II Adaptive Behavior Composite (K-VABS-II ABC) score is a calculated score (standard score) for the main area, and the higher the score, the higher the level of adaptive behavior. CGI-S is an OSU (Ohio State University, Ohio State University) autism CGI-S scale, which evaluates the overall severity through 7 Likert scales (1 point (normal, completely unsick) to 7 points (the most seriously ill patients)), and the lower the score, the lower the severity.

[0264] exist Fig.13 The figure shows the changes in K-VABS-IIABC scores of the three groups at 12 weeks, 24 weeks and 36 weeks compared with the start of the clinical trial. Fig.14The difference in the effect of the treatment group compared to the control group (placebo) on the K-VABS-II ABC and the main area and the K-PSI-4-SF parental distress values at the time point of 12 weeks of AST-001 syrup administration is shown in Fig.15 The change amounts of the three groups of K-VABS-II 2DC scores at the 12-week, 24-week, and 36-week time points compared to the clinical start time point are shown.

[0265] Furthermore, in Fig.16 The change amounts of the three groups of CGI-S scores at the 2-week, 4-week, 8-week, 12-week, 14-week, 16-week, 24-week, and 36-week time points compared to the start time point of the Phase II clinical trial are shown.

[0266] All analyses were performed on the FAS (Full Analysis Set) group. The FAS group included all patients who signed consent to receive the clinical trial drug more than once and had their efficacy evaluation variable data collected more than once after the baseline.

[0267] <Efficacy Analysis at 12 Weeks after AST-001 Syrup Administration>

[0268] The change amount of the K-VABS-II ABC score (LS mean ± SE) at 12 weeks after AST-001 syrup administration compared to the baseline was 1.66 ± 0.48 points for the control group (placebo) and 3.08 ± 0.49 points for the high-dose group. Compared with the change amount of the control group, the change amount of the high-dose group increased more significantly (see Fig.14 ). The difference in the change amount was 1.43 ± 0.69 points [90% confidence interval (CI): 0.28, 2.58], showing statistical significance (p = 0.042) (see Fig.14 ). Therefore, it was confirmed that with the administration of AST-001 syrup, the improvement in the functional adaptation level was statistically significant.

[0269] With the administration of AST-001 syrup, the K-VABS-II ABC score, the K-VABS-II communication score, the K-VABS-II Daily Living Skills score, the K-VABS-II social score, the K-VABS-II Motor Skills score, and the K-PSI-4-SF parental distress value were all improved (see Fig.14 ). Especially in the high-dose AST-001 syrup administration group, compared with the control group, the scores in the K-VABS-II communication and motor skills areas were significantly improved (p value 0.007; p value 0.049, see Fig.14)。In addition, in the high-dose AST-001 syrup administration group, compared with the control group, the K-PSI-4-SF parental distress values were significantly improved (p-value 0.027, refer to Fig.14 ).

[0270] Furthermore, when analyzing the change in K-VABS-II 2DC scores at the 12-week point of the start of the clinical trial, the change in the low-dose AST-001 syrup administration group and the high-dose AST-001 syrup administration group was higher compared with the control group (refer to Fig.15 ), and the high-dose administration group was statistically significant compared with the control group (p-value 0.026).

[0271] In addition, the baseline average CGI-S score and the average CGI-S score at 12 weeks of AST-001 syrup administration reflecting age were significantly improved in both the high-dose and low-dose administration groups compared with the control group (refer to Fig.16 ). The difference in the change in scores between the two groups was statistically significant (p-value for the high-dose group and the control group: 0.046; p-value for the low-dose group and the control group: 0.017).

[0272] Furthermore, the age of the patients participating in the clinical trial was used as a benchmark of 7 years old or 6 years old, and a subgroup efficacy analysis was performed. The results of the K-VABS-II ABC analysis are shown in Table 6 as follows.

[0273] [Table 6]

[0274]

[0275] As confirmed in Table 6 above, in patients under 7 years old and patients less than 7 years old receiving high-dose and low-dose AST-001 syrup administration, it was analyzed that the change in the 12-week K-VABS-II ABC score was significantly improved compared with the baseline value. Therefore, it was shown that the effect of improving autism spectrum disorder was significant in the patient group under 7 years old or less than 7 years old, suggesting that early treatment is effective in improving the disease during the critical period of synaptic pruning.

[0276] In addition, in the FAS population, a group of patients (PPS) who completed the clinical trial without major violations of the clinical trial protocol was selected. Taking the PPS population as the object, the change in the K-VABS-II ABC score at the 12-week point, which is the main efficacy evaluation variable, was analyzed. In the PPS population, the degree of improvement in the high-dose group compared with the control group and the low-dose group compared with the control group was more statistically significant than the analysis in the FAS population (p-value = 0.024; p-value = 0.075) (refer to Fig.17 ).

[0277] <Efficacy Analysis at 24 Weeks after AST-001 Syrup Administration>

[0278] During the extension study from 12 weeks to 24 weeks, in the intervals when AST-001 syrup was administered to the high-dose long-term administration group, low-dose long-term administration group, and high-dose short-term administration group, it was confirmed in all the administration groups that for the changes in K-VABS-II ABC and the main area, 2DC, and CGI-S scores, the improvements at the 24-week time point were greater than those at the clinical start time point and the 12-week time point (see Fig.13 , Fig.15 and Fig.16 ). In particular, in the high-dose short-term administration group where AST-001 high-dose was administered after 12 weeks of placebo administration, it was confirmed that the changes in K-VABS-II ABC and the main area, 2DC, and CGI-S scores were more improved at the 24-week time point than at the 12-week time point. Compared with the clinical trial start time point, there were no statistically significant differences in the K-VABS-II ABC and CGI-S scores among the three groups at the 24-week time point (p-value = 0.555; p-value = 0.441).

[0279] <Analysis of Efficacy at 36 Weeks after AST-001 Syrup Administration>

[0280] On the other hand, during the follow-up study from 24 weeks to 36 weeks after the start of the clinical trial, it was a period of follow-up observation without administering drugs to all patients in the high-dose long-term administration group, low-dose long-term administration group, and high-dose short-term administration group. There were no statistically significant differences in the K-VABS-II ABC and CGI-S scores among the high-dose long-term administration group, low-dose long-term administration group, and high-dose short-term administration group (p-value = 0.731; p-value = 0.611).

[0281] However, when observing the drug discontinuation effect during the 12-week period after the end of AST-001 syrup administration (i.e., from 24 weeks to 36 weeks), the examination results of each main area of K-VABS-II at the 36-week time point showed that except for the main area of motor skills, all other main areas showed a slightly decreasing tendency, and it was confirmed that the effective results improved until the 24-week time point after administration were due to the effect of AST-001 syrup.

[0282] In addition, when analyzing the changes in K-VABS-II ABC scores and CSI-S scores at the 36-week time point after the start of the clinical trial respectively, it was confirmed that in the order of the high-dose long-term administration group of AST-001 syrup, the low-dose long-term administration group of AST-001 syrup, and the high-dose short-term administration group of AST-001 syrup, the values of the changes in K-VABS-II ABC and the changes in CSI-S scores increased, and the improvement degrees also increased in this order (see Fig.13 and Fig.16).

[0283] As a result, it was confirmed that the core symptoms of autism spectrum disorder were improved when AST-001 syrup was orally administered at low or high doses to children with autism spectrum disorder.

[0284] Security

[0285] Table 7 shows the adverse reactions after administration (TEAE, Treatment Emergent Adverse Event) that occurred in more than 5% of the patients participating in the clinical trial.

[0286] [Table 7]

[0287]

[0288] The most common adverse reactions during the 36-week clinical trial were cough, nasopharyngitis, fever and diarrhea. Most of the adverse reactions were mild. The analysis showed that there was no statistically significant difference in the adverse reactions between the high-dose group, low-dose group and control group (p = 0.115). Five people (3.3%) had major adverse reactions that were not causally related to AST-001 syrup. In the clinical trial, 12 people (24.0%) reported adverse drug reactions (ADRs) related to AST-001 syrup in the control group, 6 people (11.8%) reported in the low-dose group, and 7 people (14.0%) reported in the high-dose group. The analysis showed that there was no statistically significant difference between these groups (p = 0.209).

[0289] In summary, it is shown that compared with the control group, the administration of AST-001 syrup is excellent in improving the symptoms of autism spectrum disorder such as communication and motor skills, and it is also safe and tolerable. In addition, the high-dose short-term administration group, which was administered with a high dose of AST-001 syrup after 12 weeks of administration to the control group, showed a similar level of development to the low-dose long-term administration group and the high-dose long-term administration group. It was further shown that compared with the control group, the AST-001 syrup administration group improved parenting stress by improving the parental distress sub-area of ​​K-PSI-4-SF. Therefore, it was confirmed that the administration of AST-001 syrup can improve the core symptoms of autism spectrum disorder, and the stress of parents who monitor patients with autism spectrum disorder can also be improved.

[0290] Example 4.52-week continuous administration clinical trial

[0291] The Phase II clinical trial of Example 3 was completed. Among the 75 subjects (responders) who had a treatment response to AST-001 syrup, 61 patients who were interested in participating in the continuous dosing trial were selected. Each subject received a maximum of 52 weeks of dosing. The clinical trial overview is shown in Fig.18 middle.

[0292] The subjects are patients whose K-VABS-II ABC scores at the 24-week point (End of Treatment, EOT) after administration in the Phase II clinical trial increased by more than 4 points compared with the baseline, or patients whose comprehensive improvement (CGI-Improvement, hereinafter referred to as CGI-I) evaluation results at the 24-week point (EOT) were 1 point (great improvement) or 2 points (great improvement). CGI-I is the OSU autism CGI-I scale, which means the use of 7 Likert scales (1 point: great improvement to 7 points: very poor) to evaluate the overall improvement of the Phase II clinical trial compared with the baseline visit. The lower the score, the higher the improvement. The above-selected subjects are assigned to one of the dosing groups shown in Table 8. At this time, the high-dose dosage of AST-001 syrup and the low-dose dosage of AST-001 syrup are the dosages recorded in Table 4.

[0293] [Table 8]

[0294] Dosing group assigned in Phase II clinical trials usage High-dose long-term administration group AST-001 Syrup High Dose Low-dose long-term administration group AST-001 Syrup Low Dose High-dose short-term administration group AST-001 Syrup High Dose

[0295] The baseline status of the patients at the start of the 52-week continuous dosing trial is shown in Table 9 below. There was no statistically significant difference between the low-dose group and the high-dose group in terms of age, gender, prevalence of comorbid mental illness, and baseline CGI-S.

[0296] [Table 9]

[0297]

[0298]

[0299] Effectiveness

[0300] The mean CGI-S scores measured at the start point (i.e., baseline), 8 weeks, 16 weeks, 24 weeks, 32 weeks, 40 weeks, and 52 weeks of the 52-week continuous dosing clinical trial and the changes in CGI-S scores compared to baseline are recorded in Table 10.

[0301] [Table 10]

[0302]

[0303] Furthermore, the CGI-S scores and CGI-S score change trends of each group from the start of the Phase II clinical trial to the end of the 52-week continuous administration clinical trial were Fig.19 and Fig. 20 Displayed in.

[0304] During the 52-week continuous dosing clinical trial, both the low-dose group and the high-dose group showed a tendency to improve CGI-S, and the high-dose group showed a higher level of improvement than the low-dose group. At the baseline of the continuous dosing trial, the CGI-S score (severity) of the low-dose group was higher, and accordingly, by the midpoint, the low-dose group showed a slightly larger change than the high-dose group, but at the end of the continuous dosing (52 weeks), the high-dose group showed a higher improvement than the low-dose group. In addition, after the high-dose group showed a higher improvement by the end of the 24-week dosing (EOT) of the Phase II trial, it deteriorated by the baseline of the continuous dosing trial, which was the dosing interruption period, and with the start of re-administration of AST-001 syrup, the severity decreased again and was maintained, showing further improvement at the 52-week point.

[0305] The mean CGI-I scores and CGI-I response rates (proportion of responders with a CGI-I score of 1 or 2) measured at the start point (i.e., baseline), 8 weeks, 16 weeks, 24 weeks, 32 weeks, 40 weeks and 52 weeks of the 52-week continuous dosing clinical trial are recorded in Table 11 below.

[0306] [Table 11]

[0307]

[0308] At the baseline of the 52-week continuous dosing trial, the CGI-I score of the low-dose group was lower than that of the high-dose group. In contrast, at the 52-week point, the CGI-I score of the high-dose group showed a lower result than that of the low-dose group, indicating that the degree of improvement during the continuous dosing trial was greater in the high-dose group than in the low-dose group. In addition, both the low-dose group and the high-dose group achieved a score of about 2 points, which means improvement, in the CGI-I evaluation at all time points of the continuous dosing trial, showing the improvement effect of long-term dosing.

[0309] In addition, among the 48 subjects with CGI-I response at the 24-week point in the Phase II clinical trial, 43 continued to have CGI-I response at the baseline point in the continuous dosing trial. Approximately 90% of the subjects maintained the improvement effect in CGI-I of the Phase II clinical trial. As of the end point of the 52-week continuous dosing clinical trial, approximately 94% of the subjects continued to have CGI-I response.

[0310] Furthermore, when the age of the patients participating in the clinical trial was divided into those aged 8 years and above and those aged 7 years and below, and the effectiveness of AST-001 syrup administration was analyzed, the improvement in CGI-S compared to the baseline of the Phase II clinical trial was higher in the group of subjects aged 7 years and below than in the group of subjects aged 8 years and above (see Table 12).

[0311] [Table 12]

[0312]

[0313] In addition, in the group of subjects under 7 years old, the improvement in CGI-I at the time of 52 weeks of continuous administration was higher than that in the group of subjects over 8 years old (see Table 13).

[0314] [Table 13]

[0315]

[0316]

[0317] That is, in terms of the change in CGI-S scores and the average CGI-I scores compared to the baseline of the Phase II clinical trial, the effect of improving autism spectrum disorders in the patient group under 7 years old was greater than that in the patient group over 8 years old, suggesting that early treatment during the critical period of synaptic pruning is effective in improving the disease.

[0318] Security

[0319] Among the abnormal reactions reported during the 52-week continuous dosing clinical trial, no severe adverse reactions or deaths were evaluated to be related to AST-001. Since the 52-week continuous dosing clinical trial was conducted under the COVID-19 epidemic situation, COVID-19 was the most common abnormal reaction among the 41 abnormal reactions reported (5 cases in the low-dose group and 6 cases in the high-dose group) (see Table 14 below).

[0320] [Table 14]

[0321]

[0322] In addition, a total of 2 cases of adverse drug reactions (ADR) were both grade 1 (mild) adverse reactions, and "recovery" was confirmed after the adverse reactions. Therefore, it is shown that children with autism spectrum disorders can safely take the drug without interruption during the longest period of continuous administration of 52 weeks.

[0323] In summary, AST-001 syrup can be safely administered to children with autism spectrum disorders over a long period of time, and shows a tendency that the longer the administration period, the better the improvement in effectiveness. Not only does the therapeutic response shown in the Phase II clinical trial continue, but the supplementary improvement effect brought about by long-term administration has also been confirmed.

[0324] Example 5. Phase III clinical trial usage / dose modeling analysis

[0325] Considering that the AST-001 syrup used in the clinical trial has a dosage form of 2g / 20mL per packet, a modeling analysis was conducted to increase the convenience of taking for children. The aim was to evaluate whether the effect confirmed in the Phase II clinical trial could be achieved even if the single-dose dosage for each weight range was set to an even number.

[0326] For modeling analysis, a population pharmacokinetic / pharmacodynamic model was established using the experimental data obtained from the clinical trial of healthy adult males in Example 2.2 and the Phase II clinical trial in Example 3. The population pharmacokinetic / pharmacodynamic analysis of AST-001 used the Monolix (version 2021R1) program, and the parameter estimation was performed by the stochastic approximate expectation maximization algorithm, using the inspection time points and KVABS-II-ABC score data based on the clinical trial scheme of Example 2.2 and Example 3. The model obtained by removing the production of endogenous L-serine from the model established in Example 2.3 was used as a population pharmacokinetic model. The pharmacodynamic model explored the turnover model, the effect compartment model, and the direct model, and the effect of the drug explored the linear model and the Emax model. The morphology of the K-VABS-II-ABC score changes over time, including the placebo group, explored constant, linear, and exponential models.

[0327] Finally, a simulation analysis predicting the morphology of K-VABS-II-ABC scores after 12 weeks of AST-001 administration by weight range was performed based on various scenarios using a pharmacokinetic / pharmacodynamic model.

[0328] The simulation analysis used the Simulx (2021R1 version) program and the "httk" package of R software to generate and analyze data of 1,000 to 1,200 virtual children aged 2 to 12 years. The weight-based doses 1, 3, and 4 of the simulation scenario were analyzed using the data of 1,000 virtual children, and the weight-based dose 2 was analyzed using the data of 1,200 virtual children. At the 12-week point, the proportion of subjects whose KVABS-II-ABC score change increased by 2 points or more (= target achievement rate) was explored, and the dose in the scenario that showed a target achievement rate similar to the dosing method for each weight range in the Phase II clinical trial was predicted to be a dose that can both improve the convenience of administration and show the effects confirmed in the Phase II clinical trial.

[0329] [Population pharmacokinetic / pharmacodynamic analysis results]

[0330] The pharmacodynamic profile of AST-001 syrup was appropriately described by a model that linearly presented drug effects according to effect site concentration, independent of drug administration, and changes in K-VABS-II-ABC scores were included in the linear progressive model. The final model was analyzed to appropriately predict the pharmacodynamic profile of AST-001 syrup after administration through GOF and VPC.

[0331] [Simulation analysis results]

[0332] The results of 12 weeks of BID dosing of AST-001 syrup were compared with the results of the simulation of the dosing regimen in each weight range in the Phase II clinical trial. The results of the weight-based dosing scenario 4 achieved the most similar efficacy target. The results of the analysis that predicted the 12-week K-VABS-II-ABC score morphology at this time are shown in Fig.21 middle.

[0333] Specifically, the median change in K-VABS-II-ABC scores from week 0 to week 12 was predicted to be 1.67 to 3.74 in the AST-001 syrup treatment group and 0.92 to 1.15 in the placebo group (refer to Fig.21 In addition, when 2 g BID was administered in the 10 kg to 13 kg body weight range, the proportion of children whose K-VABS-II-ABC score changed by 2 points or more (target achievement rate) was 40.5%, which was 2.1 times that of the placebo group. Similarly, in other weight ranges, the target achievement rate of the AST-001 syrup treatment group was confirmed to be 1.8 to 2.7 times that of the placebo group (refer to Fig.21 ).

[0334] Therefore, the analysis shows that, considering the convenience of children's administration and the effect of AST-001 syrup, if AST-001 syrup is administered in the dosage for each weight range as shown in Table 15, it will show significant efficacy clinically compared to placebo.

[0335] [Table 15]

[0336]

Claims

1. A use of L-serine or a pharmaceutically acceptable salt thereof for preparing a medicament for improving or treating communication disorders and / or social interaction disorders in subjects with autism spectrum disorders, wherein: The medicament comprises a pharmaceutically acceptable carrier, The medicament is in unit dosage form.

2. The use according to claim 1, wherein The subject has childhood autism.

3. The use according to any one of claim 1 or claim 2, wherein The subject's Clinical Global Impression-Severity Scale is 5 or above, and the Vineland Adaptive Behavior Scale-Second Edition Antisocial Behavior Scale is 48 or above.

4. The use according to any one of claims 1 to 3, wherein The subject is under 18 years old, under 13 years old, under 11 years old, under 7 years old, less than 7 years old, 2 to 7 years old, or 2 to 6 years old.

5. The use according to any one of claims 1 to 4, wherein The subject has a body weight of 10 kg to 100 kg or 10 kg to 60 kg.

6. The use according to any one of claims 1 to 5, wherein The daily dosage of L-serine or a pharmaceutically acceptable salt thereof administered to the subject is 1 g to 60 g, 2 g to 30 g, 4 g to 30 g, 2 g to 28 g or 2 g to 14 g.

7. The use according to any one of claims 1 to 5, wherein The daily dosage of L-serine or a pharmaceutically acceptable salt thereof administered to the subject is 100 mg / kg to 600 mg / kg, 140 mg / kg to 580 mg / kg, 140 mg / kg to 572 mg / kg, 200 mg / kg to 400 mg / kg, 280 mg / kg to 580 mg / kg, 280 mg / kg to 572 mg / kg, 70 mg / kg to 280 mg / kg, 140 mg / kg to 290 mg / kg or 140 mg / kg to 286 mg / kg.

8. The use according to any one of claims 1 to 7, wherein The drug was administered to the subject twice a day.

9. The use according to any one of claims 1 to 8, wherein The medicament is in oral dosage form.

10. The use according to any one of claims 1 to 9, wherein The medicine is in the form of a syrup.

11. The use according to any one of claims 1 to 10, wherein The medicament includes one or more of a sweetener, a thickener, a pH adjuster, a preservative and a solvent.

12. The use according to any one of claims 1 to 11, wherein The concentration of L-serine or a pharmaceutically acceptable salt thereof contained in the pharmaceutical agent is 10 mg / ml to 500 mg / ml.

13. The use according to any one of claims 1 to 12, characterized in that: Each volume of the unit dosage form is 10 ml to 100 ml.

14. The use according to any one of claims 1 to 13, wherein The unit dosage form contains L-serine or a pharmaceutically acceptable salt thereof in an amount of 1 g to 100 g, 1 g to 60 g, 2 g to 30 g, 4 g to 30 g, 2 g to 28 g or 1 g to 14 g.

15. The use according to any one of claims 1 to 14, characterized in that The L-serine is prepared by the following method: a) chelating glycine and a copper compound to form a glycine-copper complex; b) reacting the complex obtained in step a) with a carbon source to form a complex into which a hydroxymethyl group is introduced; c) treating the complex obtained in step b) with a reducing agent to form DL-serine; and d) a step of optically resolving the DL-serine obtained in step c) to separate L-serine.

16. A preparation comprising 1 g to 100 g of L-serine or a pharmaceutically acceptable salt thereof, a thickener, a pH adjuster, a preservative and a solvent, wherein: The preparation is in unit dosage form.

17. The preparation according to claim 16, characterized in that Each volume of the preparation is 10 ml to 100 ml.

18. The preparation according to claim 16 or claim 17, wherein The formulation is administered to improve or treat communication disorders and / or social interaction disorders in a subject with autism spectrum disorder.

19. The preparation according to claim 18, wherein The daily dosage of L-serine or a pharmaceutically acceptable salt thereof administered to the subject is 100 mg / kg to 600 mg / kg, 140 mg / kg to 580 mg / kg, 140 mg / kg to 572 mg / kg, 200 mg / kg to 400 mg / kg, 280 mg / kg to 580 mg / kg, 280 mg / kg to 572 mg / kg, 70 mg / kg to 280 mg / kg, 140 mg / kg to 290 mg / kg or 140 mg / kg to 286 mg / kg.

20. A kit comprising a preparation containing 1 g to 100 g of L-serine or a pharmaceutically acceptable salt thereof, wherein: The preparation is used to improve or treat communication disorders and / or social interaction disorders in subjects with autism spectrum disorders, and the preparation is in unit dosage form.

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