Depot composition with controlled initial release through hydrophobic ion pairing gelation for treatment of obesity and diabetes for providing improved medical therapy of semaglutide or tirzepatide, and method for preparing same

Abstract

The present invention relates to a depot composition comprising calcium chloride inside microspheres. The depot composition according to the present invention can control excessive release of an active ingredient contained inside microspheres at the initial stage of depot injection by using calcium chloride as a cationic salt. In addition, the composition has excellent suspendability and a significantly small amount of residual solvent, and thus even when administered as an injection by a user, the effect of the active ingredient can be uniformly and continuously obtained. In addition, an improved medical therapy of semaglutide can be provided by adjusting the dose of semaglutide or tirzepatide by means of the depot formulation.

Description

Depot composition with controlled initial release through hydrophobic ion pairing gelation for the treatment of obesity and diabetes to provide improved medical therapy of semaglutide or tirzepatide and method for preparing the same

[0001] The present invention provides improved medical therapy of semaglutide or tirzepatide.

[0002] In addition, the present invention relates to a depot composition, which controls the initial release of microspheres through hydrophobic ion pairing gelation and significantly increases the amount of residual solvent removed, so that even if an excessive amount of drug is administered over a long period of time without repeated administration of the depot composition, there are no sudden side effects, and it provides high user convenience and excellent therapeutic effects through excellent anti-diabetes and anti-obesity effects.

[0003]

[0004] Depot preparations are formulations designed to provide prolonged drug effects and are primarily used for long-term administration of medications such as hormones. As sustained-release formulations, depot preparations have the advantage of minimizing the number of injections required. However, it is challenging to achieve consistent and sustained drug release throughout the entire post-injection period. In particular, if the initial release of the drug within the depot preparation is high, excessive drug release during the initial injection can lead to adverse effects due to the excessive drug dose and a reduction in the sustained drug effect over the specified period. Therefore, controlling the initial release of depot preparations is crucial.

[0005] GLP-1 receptor agonists, such as semaglutide, were initially developed to lower blood sugar levels in diabetics. They have also been shown to be effective in reducing weight in obese patients, leading to their use in the treatment of both diabetes and obesity. Semaglutide is an analog of the incretin hormone secreted by the intestines. It increases the amount of insulin secreted after a meal and slows the emptying of stomach contents, allowing food to move more slowly from the stomach to the small intestine. Furthermore, semaglutide can lower blood sugar and aid in weight loss through various appetite-suppressing effects on the central nervous system.

[0006] Meanwhile, tirzepatide is a dual agonist that acts as both a GLP-1 receptor agonist and a GIP receptor agonist, and like semaglutide, it has glycemic control and anti-obesity effects. GIP (glucose-dependent insulin-releasing polypeptide) is an incretin hormone and, like GLP-1, is secreted from the small intestine in response to food. It has been recognized as an obesity hormone because it induces glucagon secretion along with insulin secretion. However, by acting together with GLP-1, it counteracts the glucagon secretion induced by GIP, thereby reducing fat accumulation. Therefore, tirzepatide, a dual GLP-1 / GIP receptor agonist, may have the same glycemic control and anti-obesity effects as semaglutide.

[0007] However, when the body is exposed to excessive amounts of GLP-1 and GLP-1 / GIP receptor agonists, side effects such as nausea, diarrhea, increased heart rate, or headache may occur. Therefore, controlling the initial release in depot formulations containing GLP-1 and GLP / GIP receptor agonists as active ingredients is one of the more important tasks. In the case of the already approved semaglutide-containing products Ozempic (for the treatment of diabetes) and Wigovi (for the treatment of obesity) and the tirzepatide-containing products Maunzaro (for the treatment of diabetes) and Zebbound (for the treatment of obesity), the drug is gradually increased to a higher dose after the body adapts to the drug from a low dose in advance. This is because of the side effects that occur when exposed to high doses of the drug, and this is why controlling the initial exposure is important.

[0008] Meanwhile, Japanese Patent No. 5681626 discloses a controlled-release formulation comprising microspheres containing a lipid component containing a sterol and a polylactide-co-glycolide (PLGA) polymer, and a phospholipid component, capable of controlling the release rate. However, the lipid component is not sufficiently effective in suppressing initial release, and its strong hydrophobicity can deteriorate the microspheres' properties or cause them to clump together, resulting in poor suspension.

[0009] Furthermore, according to Korean patent application No. 10-2011-0145945, a novel synthetic polymer containing a poly(lactide-co-glycolide) (PLGA) polymer, which forms microspheres, is synthesized with a polyvalent ion to provide a novel synthetic polymer with a polyvalent ion. However, this novel synthetic polymer requires mandatory toxicity testing for clinical use, which complicates approval. Therefore, the practical application of this polymer in depot formulations is limited.

[0010] Furthermore, Korean Patent No. 10-409413 discloses a method for significantly suppressing initial over-release and minimizing organic solvent usage by manufacturing microspheres using an underwater drying method and then heating and drying them above the glass transition temperature (Tg) of a biodegradable polymer. However, this manufacturing method has the disadvantage that the bioactive substance can be denatured by heat.

[0011] Thus, since conventional slow-release microspheres have limitations in providing a depot composition with sufficiently controlled initial release, there is still a need for the development of a depot composition with controlled initial release and the ability to maintain the drug for a desired period of time.

[0012] Additionally, improved medical therapies are needed to address gastrointestinal side effects, particularly vomiting, abdominal pain, and nausea, that are adverse effects of semaglutide or tirzepatide in preventing or treating obesity or preventing or treating diabetes.

[0013]

[0014] The present invention is intended to solve the above-mentioned problems, and an object of the present invention is to provide a depot composition and a method for producing the same in which excessive initial release is suppressed and the amount of residual solvent is significantly reduced.

[0015] In addition, the present invention aims to provide an improved medical therapy of semaglutide and / or tirzepatide that can exhibit a long-term therapeutic effect, including semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof.

[0016] The inventors of the present invention have endeavored to develop a depot composition capable of controlling excessive initial drug release. As a result, they have discovered that incorporating calcium chloride into the aqueous phase of microspheres can suppress initial drug release. Furthermore, they have discovered that incorporating a plasticizer into the oil phase of microspheres can significantly increase the amount of residual solvent removed. These microspheres, which can be used as depots, can technically contain a large amount of drug while preventing initial drug release over a targeted period of time. Furthermore, they have confirmed that these microspheres possess excellent formulation properties with long-term therapeutic efficacy, thereby completing the present invention.

[0017]

[0018] The present invention relates to a pharmaceutical composition for preventing or treating obesity or diabetes, comprising semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof as an active ingredient,

[0019] The dosage of GLP-1 receptor agonists for the prevention or treatment of obesity is 12 to 96 mg / 4 weeks,

[0020] A pharmaceutical composition is provided in which the dosage of a GLP-1 receptor agonist for the prevention or treatment of diabetes is 4 to 80 mg / 4 weeks.

[0021]

[0022] In addition, the present invention includes O / W2 type microspheres including an oil layer (O layer) and a second aqueous layer (W2 layer), or W1 / O / W2 type microspheres including a first aqueous layer (W1 layer), an oil layer (O layer), and a second aqueous layer (W2 layer),

[0023] In the above O / W2 type microspheres, the oil layer (O layer) or the first aqueous layer (W1 layer) in the above W1 / O / W2 type microspheres comprises semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof.

[0024] A depot composition is provided, wherein the oil layer (O layer) of the O / W2 type microparticles or W1 / O / W2 type microparticles comprises a biodegradable polymer, and the second aqueous layer (W2 layer) comprises calcium chloride.

[0025]

[0026] In addition, the present invention comprises a step of preparing an oil phase (O layer) solution or mixing a first water phase 1 (W1 layer) solution and the oil phase (O layer) solution to prepare a W1 / O emulsion; and

[0027] It includes a step of preparing an O / W2 emulsion or a W1 / O / W2 emulsion by adding the above oil layer (O layer) solution or W1 / O emulsion to a second water phase 2 (W2) solution,

[0028] In the above O / W2 emulsion, the oil layer (O layer) solution or in the above W1 / O / W2 emulsion, the first water layer (W1 layer) solution comprises semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof;

[0029] A method for producing a depot composition is provided, wherein the oil layer (O layer) solution in the O / W2 emulsion or W1 / O / W2 emulsion contains a biodegradable polymer, and the second aqueous layer (W2 layer) solution contains calcium chloride.

[0030]

[0031] The microspheres of the depot composition of the present invention contain calcium chloride in their aqueous layer, thereby controlling the release of excessive amounts of the active ingredient contained within the microspheres during the initial injection of the depot. Furthermore, they exhibit excellent suspending properties and significantly lower residual solvent levels, allowing users to consistently and safely obtain the effects of the active ingredient even when administered as an injection.

[0032] Additionally, the dosage of semaglutide and / or tirzepatide can be adjusted through the depot formulation, thereby providing improved medical therapy of semaglutide and / or tirzepatide.

[0033]

[0034] Figures 1a to c are SEM photographs of microspheres manufactured according to Examples 1-3(a), 1-9(b), and 1-13(c) of the present invention.

[0035] Figure 2 is an SEM photograph of microspheres manufactured according to Comparative Example 1-1 of the present invention.

[0036] Figure 3 is an SEM photograph of microspheres manufactured according to Comparative Example 2-7 of the present invention.

[0037] Figure 4a shows the animal PK profiles of microspheres manufactured according to Comparative Example 1-1, Examples 1-3, 1-5, and Comparative Example 2-7 of the present invention. In addition, Figure 4b shows the animal PK profiles of microspheres manufactured according to Comparative Example 1-6 and Example 2-6 of the present invention.

[0038] Figures 5a to e show DSC results of microspheres manufactured according to 504H polymer (a) and Comparative Examples 1-1 (b), 2-7 (c), and Examples 1-3 (d), 1-6 (e) of the present invention.

[0039] Figures 6a to e show the FT-IR results of microspheres manufactured according to 504H polymer (a) and Comparative Examples 1-1 (b), 2-7 (c), and Examples 1-3 (d), 1-6 (e) of the present invention.

[0040] Figures 7a to 7b show the blood sugar (Glucose) reduction effect (a) and blood glucose achromatoplast reduction effect (b) in a diabetic animal model according to the dose of semaglutide depot.

[0041] Figure 8 shows the body weight reduction effect in an obesity-induced animal model according to the dose of semaglutide depot.

[0042]

[0043] Hereinafter, the configuration of the present invention will be described in detail.

[0044] In the present invention, the numerical limitation indicates more than or less than, unless otherwise stated.

[0045]

[0046] The present invention provides a pharmaceutical composition for preventing or treating obesity or diabetes, comprising a GLP-1 receptor agonist or a GLP-1 / GIP receptor dual agonist as an active ingredient.

[0047] In the present invention, the GLP-1 receptor agonist is semaglutide or a pharmaceutically acceptable salt thereof, and the GLP-1 / GIP receptor dual agonist is tirzepatide or a pharmaceutically acceptable salt thereof.

[0048] In the present invention, semaglutide or tirzepatide is a glucagon-like peptide-1 (GLP-1) receptor agonist that participates in the action of GLP-1, a hormone that plays a role in lowering blood sugar, and can be used for the prevention or treatment of obesity or diabetes. Among these, tirzepatide is a glucose-dependent insulin-releasing polypeptide, a GIP hormone, and can be used for the prevention or treatment of obesity or diabetes by acting together with GLP-1.

[0049] In one embodiment, semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof may each be selected from the group consisting of acetate, benzoate, hydroxynaphthoate, napadisylate, or pamoate.

[0050] In one specific example, the appropriate dosage of semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof varies depending on the patient's body weight, age, sex, health condition, diet, administration time, administration method, excretion rate, and severity of the disease. For example, the daily dosage of semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof of the present invention may be 0.357 to 3.428 mg, preferably 1 to 1.714 mg, for obesity treatment, and may be 0.142 to 2.857 mg, preferably 0.428 to 1.142 mg, for diabetes treatment, based on semaglutide or tirzepatide.

[0051] In one specific embodiment, a suitable administration cycle of semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof may be determined depending on the dosage. For example, the semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof of the present invention may be administered once every month to once every six months, preferably once every month, once every two months, once every three months, or once every six months, and most preferably once every month.

[0052] When the pharmaceutical composition according to the present invention is used for the prevention or treatment of obesity, the dosage of semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof is 12 to 96 mg / 4 weeks, respectively. In addition, when the pharmaceutical composition is used for the prevention or treatment of diabetes, the dosage of semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof is 4 to 80 mg / 4 weeks, respectively.

[0053] In the present invention, obesity can be prevented or treated without side effects by administering 12 to 96 mg of semaglutide or a pharmaceutically acceptable salt thereof or tirzepatide or a pharmaceutically acceptable salt thereof once every four weeks. Side effects in this case refer to safety events such as gastrointestinal abnormalities, and gastrointestinal abnormalities may refer to symptoms selected from the group consisting of nausea, vomiting, diarrhea, and constipation.

[0054] In one specific example, the dosage of semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof for the prevention or treatment of obesity may be specifically 12 to 96 mg / 4 weeks, 15 to 86 mg / 4 weeks, 19 to 77 mg / 4 weeks, 24 to 60 mg / 4 weeks, or 28 to 48 mg / 4 weeks, respectively.

[0055] In addition, in the present invention, diabetes can be prevented or treated without side effects by administering 4 to 80 mg / 4 weeks of semaglutide or a pharmaceutically acceptable salt thereof once every 4 weeks; or tirzepatide or a pharmaceutically acceptable salt thereof.

[0056] In one specific example, the dosage of semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof for the prevention or treatment of diabetes may be specifically 4 to 80 mg / 4 weeks, 6 to 64 mg / 4 weeks, 8 to 48 mg / 4 weeks, 10 to 40 mg / 4 weeks, or 12 to 32 mg / 4 weeks, respectively.

[0057] In one specific embodiment, the pharmaceutical composition according to the present invention can be administered to a subject as a formulation for parenteral administration.

[0058] Specifically, the pharmaceutical composition of the present invention can be formulated into various forms for parenteral administration, such as microneedle formulations and depot formulations. Such pharmaceutical compositions can be administered intramuscularly, transdermally, intravenously, and / or subcutaneously.

[0059] In one specific embodiment, semaglutide or a pharmaceutically acceptable salt thereof according to the present invention; or tirzepatide or a pharmaceutically acceptable salt thereof may be administered to a subject in the form of a depot composition.

[0060] The above depot composition is a sustained-release formulation capable of releasing the drug over an extended period of time. Sustained-release formulations can gradually release the drug within the formulation to the intended level over a certain period of time, while depot compositions can control the initial burst release, in which an excessive amount of the drug is released at the beginning of the injection.

[0061] If the body is exposed to excessive amounts of semaglutide or tirzepatide, symptoms such as vomiting, nausea, hypoglycemia, and headache may occur. Therefore, when using semaglutide or tirzepatide as the active ingredient in a depot composition, it is very important to prevent excessive release of the active ingredient from the microspheres of the depot composition.

[0062] In the present invention, by using the depot composition described below, an early controlled release effect can be secured. That is, the dosage of semaglutide or tirzepatide can be controlled, thereby providing a novel treatment for the aforementioned semaglutide or tirzepatide.

[0063] In one specific example, when formulated as a depot composition, the content of semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof may be 5 to 25 wt%, respectively, based on the total weight of the formulation.

[0064] In one specific embodiment, the pharmaceutical composition of the present invention exhibits an AUC of semaglutide or tirzepatide when administered to a subject. t / C max It can be more than 100 hours.

[0065] In the present invention, AUC means the area under the plasma concentration-time curve, which is a measure of total bioavailability, and AUC t (AUC last ) means the area under the plasma concentration-time curve until the measurement time, and C max (Peak blood concentration) refers to the highest blood concentration of the administered drug when administered in vivo. Furthermore, all of these values ​​are pharmacokinetic parameters for a single dose.

[0066] The pharmaceutical combination according to the present invention has the above AUC t / C max By indicating the value, the blood concentration of the drug can be maintained at a constant level while being therapeutically effective and not excessively high when administered. Through this, the side effects commonly seen due to the increatin hormone (GLP-1 receptor agonist) of semaglutide or tirzepatide, such as gastrointestinal side effects such as nausea, vomiting, constipation, and diarrhea, insomnia, dizziness, dysgeusia, asthenia, cholelithiasis, cholecystitis, pancreatitis, dehydration, tachycardia, or acute renal failure, can be significantly reduced.

[0067]

[0068] The present invention also provides a depot composition, wherein the depot composition comprises microspheres comprising semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof as an active ingredient.

[0069] The above microspheres may be O / W2 type microspheres including an oil layer (O layer) and a second aqueous layer (W2 layer), or may be W1 / O / W2 type microspheres including a first aqueous layer (W1 layer), an oil layer (O layer), and a second aqueous layer (W2 layer).

[0070] The depot composition of the present invention has the advantage of excellent suspending ability while controlling initial release by including O / W2 type microparticles or W1 / O / W2 type microparticles.

[0071] In this specification, for the convenience of explanation regarding two or more aqueous layers or aqueous layer solutions, the aqueous layer existing inside the oil layer of microparticles or the aqueous layer solution for forming it may be referred to as the first aqueous layer (W1 layer) or the first aqueous layer solution, and the aqueous layer formed outside the oil layer of microparticles or the aqueous layer solution for forming it may be referred to as the second aqueous layer (W2 layer) or the second aqueous layer solution, respectively. In this case, the aqueous layer (W layer) or the aqueous layer solution in the O / W type microparticles is understood to have a configuration corresponding to the second aqueous layer (W2 layer) or the second aqueous layer solution in the W1 / O / W2 type microparticles. Therefore, the O / W emulsion may also be referred to as an O / W2 emulsion in the present invention. In the following description, references to the second aqueous layer (W2 layer) or the second aqueous layer solution should be understood to have a description corresponding to the aqueous layer or the aqueous layer solution for the O / W type microparticles.

[0072] In the present invention, the oil layer (O layer) in the O / W2 type microparticles or the first aqueous layer (W1 layer) in the W1 / O / W2 type microparticles contains an active ingredient. The active ingredient may include semaglutide or a pharmaceutically acceptable salt thereof as an API; or tirzepatide or a pharmaceutically acceptable salt thereof. The depot composition according to the present invention can be administered into the body and serve to release the active ingredient into the body over a certain period of time. The active ingredient is contained inside the microparticles and is protected from the microparticles, and can remain in the body for a certain period of time.

[0073] In one specific example, the content of semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof may be 5 to 25 wt%, respectively, based on the total weight of the microparticles.

[0074] In the present invention, the oil layer (O layer) in the O / W2 type microspheres or the oil layer (O layer) in the W1 / O / W2 type microspheres comprises a biodegradable polymer. The biodegradable polymer has the property of being degraded in vivo. In the present invention, microspheres are formed, and as the polymer is gradually degraded in the body, the active ingredient contained therein can be gradually released. In other words, the biodegradable polymer can protect the drug inside during the drug release period and play a role in controlling long-term drug release.

[0075] The biodegradable polymer may be any polymer approved by the pharmaceutical safety authorities of each country for use in injectable preparations, and may include, for example, a polymer selected from the group consisting of polylactide (PLA), polyglycolide (PGA), and poly(lactide-co-glycolide) (PLGA).

[0076] The above biodegradable polymer may have an inherent viscosity of 0.35 to 0.65 dL / g, or 0.50 to 0.55 dL / g. If the inherent viscosity is less than 0.35 dL / g, the drug may be released faster than the desired period of time, and if the inherent viscosity is more than 0.65 dL / g, the drug may be released slower than the desired period of time, making it difficult to obtain a sufficient drug effect. The inherent viscosity may be measured according to the viscosity measurement method for poly(lactide-co-glycolide) (PLGA) provided by the manufacturer.

[0077] The content of the biodegradable polymer may be 75 to 95 wt% based on the total weight of the microparticles.

[0078] In the present invention, the oil layer (O layer) in the O / W2 type microparticles or the oil layer (O layer) in the W1 / O / W2 type microparticles may additionally include a plasticizer.

[0079] Plasticizers are used to improve the flexibility and brittleness of biodegradable polymers. These plasticizers help the organic solvent to quickly escape into the water phase and be removed before the polymer solidifies during the process of heat-drying the microspheres, which are then manufactured using an underwater drying method and then vaporized and removed. Consequently, the amount of organic solvent remaining in the final microspheres can be minimized.

[0080] In one specific example, the plasticizer may be any one approved by the drug safety authorities of each country as a pharmaceutical additive without limitation. For example, the plasticizer may include at least one selected from the group consisting of glycerin (glycerol), concentrated glycerin, sorbitol, propylene glycol, poloxamer polymers, phthalate polymers such as dimethyl-, diethyl-, and dibutyl phthalate, citrate derivatives such as tributyl-, triethyl-, or acetyl citrate, triacetin, and castor oil. The poloxamer may be poloxamer 188, poloxamer 407, and the like, and the citrate derivative may be triethyl citrate (TEC), and the like.

[0081] In one specific example, the plasticizer may be included in a concentration of 0.1 to 10.0% (w / v) with respect to the oil layer (O layer). Specifically, when triethyl citrate is used as the plasticizer, the concentration may be 0.5 to 10% (w / v), or 1 to 8% (w / v), with respect to the oil layer (O layer), and when poloxamer is used, the concentration may be 0.1 to 5.0% (w / v), or 0.1 to 3.0% (w / v) with respect to the oil layer (O layer).

[0082] In the present invention, the second aqueous layer (W2 layer) in the O / W2 type microparticles or W1 / O / W2 type microparticles contains calcium chloride.

[0083] Typically, microspheres are manufactured through a homogenization process, and the solvent used to dissolve the polymer is vaporized during the process, solidifying the microspheres. In the present invention, during the solidification process, calcium chloride comes into contact with the polymer on the surface of the microspheres, and a gelation phenomenon occurs at the interface due to hydrophobic ion pairing, coating the surface of the microspheres to form pore-free microspheres. Through this, the calcium chloride can control the initial release of the active ingredient.

[0084] The above calcium chloride may be included in the second aqueous layer (W2 layer) at a concentration of 0.1 to 5.0% (w / v), 0.1 to 4.0% (w / v), 0.1 to 3.0% (w / v), or 0.1 to 1.5% (w / v).

[0085] The depot composition of the present invention may additionally include a pharmaceutically acceptable carrier or excipient, etc. However, preferably, the depot composition of the present invention may not include a stabilizer, pH regulator, or oxidizing agent.

[0086]

[0087] In addition, the present invention relates to a method for producing the above-described depot composition.

[0088] The depot composition according to the present invention can be prepared by a single emulsification method in which an oil phase solution is added to an aqueous phase solution and emulsified to create an O / W emulsion. Alternatively, it can be prepared by a double emulsification method in which a water-oil phase emulsion is added to an aqueous phase solution and emulsified to create a water-oil phase emulsion.

[0089] In one specific embodiment, the depot composition comprises a step of preparing an oil phase (O layer) solution or mixing a first water phase 1 (W1 layer) solution and the oil phase (O layer) solution to prepare a W1 / O emulsion (hereinafter, step S1); and

[0090] It can be manufactured through a step (hereinafter, step S2) of manufacturing an O / W2 emulsion or a W1 / O / W2 emulsion by adding the above oil phase solution or W1 / O emulsion to a second water phase 2 (W2 layer) solution.

[0091] In addition, the manufacturing method of the present invention may additionally include a step of heating and drying the manufactured O / W2 emulsion or W1 / O / W2 emulsion to vaporize it. This step may be performed at a temperature below the glass transition temperature (Tg) of the biodegradable polymer when using a plasticizer described below.

[0092] In addition, the manufacturing method of the present invention may additionally include a step of drying and / or filtering the manufactured O / W2 emulsion or W1 / O / W2 emulsion and then centrifuging to recover microspheres.

[0093] In the present invention, step S1 is a step of preparing an oil solution or preparing a W1 / O emulsion.

[0094] In one specific embodiment, the oil-based solution may include a fat-soluble solvent. The fat-soluble solvent may be any carrier or solvent commonly used in the art and pharmaceutically acceptable. For example, dichloromethane may be used as the fat-soluble solvent, but is not limited thereto.

[0095] In one specific embodiment, the solvent of the first aqueous layer solution may be any carrier or solvent commonly used in the art and pharmaceutically acceptable. Examples include, but are not limited to, polyvinyl alcohol (PVA) or sodium acetate.

[0096] In one specific embodiment, the first aqueous layer solution in the oily solution or W1 / O emulsion comprises an active ingredient, wherein the active ingredient is a GLP-1 receptor agonist, which may be semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof. The content of the GLP-1 receptor agonist may be 5 to 25 wt%, and specifically, the content of semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof may be 5 to 25 wt%, respectively.

[0097] In one specific embodiment, the oily solution comprises a biodegradable polymer.

[0098] The biodegradable polymers mentioned above can be used without limitation as long as they are approved by the drug safety authorities of each country for use in injectable preparations, and for example, a polymer selected from the group consisting of polylactide (PLA), polyglycolide (PGA), and poly(lactide-co-glycolide) (PLGA) can be used.

[0099] The content of the biodegradable polymer may be 75 to 95 wt% based on the total weight of the microparticles.

[0100] In the present invention, the oil solution may additionally contain a plasticizer.

[0101] As the plasticizer, for example, one or more selected from the group consisting of glycerin (glycerol), concentrated glycerin, sorbitol, propylene glycol, poloxamer series polymers, phthalate polymers such as dimethyl-, diethyl-, and dibutyl phthalate, citrate derivatives such as tributyl-, triethyl-, or acetyl citrate, triacetin, and castor oil can be used. The poloxamer can be poloxamer 188, poloxamer 407, etc., and the citrate derivative can be triethyl citrate (TEC), etc.

[0102] In one specific example, the plasticizer may be included in a concentration of 0.1 to 10.0% (w / v) relative to the entire oil layer solution. Specifically, when triethyl citrate is used as the plasticizer, the concentration may be 0.5 to 10.0% (w / v) or 1 to 8% (w / v) relative to the oil layer solution, and when poloxamer is used, the concentration may be 0.1 to 5.0% (w / v) or 0.1 to 3.0% (w / v) relative to the oil layer solution.

[0103] In one specific example, a W1 / O emulsion can be prepared by mixing an oil phase with a first aqueous layer solution and stirring using a homogenizer. The stirring speed and time can be varied depending on the emulsion formation conditions and the amount of sample.

[0104] In the present invention, step S2 is a step of preparing an O / W2 emulsion or a W1 / O / W2 emulsion by adding the oil phase solution or W1 / O emulsion prepared / manufactured in step S1 to a second aqueous phase solution.

[0105] In one specific embodiment, the solvent of the second aqueous layer solution may be any carrier or solvent commonly used in the art and pharmaceutically acceptable. Examples include, but are not limited to, polyvinyl alcohol (PVA) or sodium acetate. The second aqueous layer solution may additionally include an osmotic pressure regulator to control the osmotic pressure of the solution during the emulsion preparation process.

[0106] Additionally, the solvent may further include methylcellulose, polyvinylpyrrolidone, carboxymethylcellulose, lecithin, gelatin, polyoxyethylene, oleoxyethylene sorbitan fatty acid ester, polyoxyethylene palmitate derivatives, and mixtures thereof.

[0107] In one specific example, the second aqueous layer solution comprises calcium chloride. Calcium chloride significantly enhances the suspension capacity of the depot composition, in addition to its initial release suppression effect, which suppresses excessive release of the active ingredient within the microparticles, and thus can be effectively used as a depot agent.

[0108] The calcium chloride may be included in a concentration of 0.1 to 5.0% (w / v), 0.1 to 4.0% (w / v), 0.1 to 3.0% (w / v), or 0.1 to 1.5% (w / v) relative to the entire second aqueous layer solution.

[0109] In one specific example, an O / W2 emulsion or a W1 / O / W2 emulsion can be prepared by stirring with a homogenizer. The stirring speed and time can be varied depending on the emulsion formation conditions and the amount of sample.

[0110]

[0111] The advantages and features of the present invention, and the methods for achieving them, will become clearer with reference to the embodiments described in detail below. However, the present invention is not limited to the embodiments disclosed below, but may be implemented in various different forms. These embodiments are provided solely to ensure that the disclosure of the present invention is complete and to fully inform those skilled in the art of the scope of the invention. The present invention is defined solely by the scope of the claims.

[0112]

[0113] Example

[0114] [Examples 1 and 2] Preparation of microspheres containing calcium chloride

[0115] In Example 1, 1 g of semaglutide as an API and in Example 2, 1 g of tirzepatide as an API were dissolved in 4 mL of water to prepare a first aqueous layer (W1 layer) solution. An oil layer (O layer) solution was prepared by dissolving the polymer in the content (g) according to Tables 2 and 3 in 60 mL of dichloromethane (Dichloromethane, Honeywell). The W1 layer solution and the O layer solution were mixed and stirred with a homogenizer at 8,000 rpm for 2 minutes to prepare a W1 / O emulsion.

[0116] Next, calcium chloride (CaCl2) according to the concentrations corresponding to Tables 2 and 3 was dissolved in 5 L of 1 wt% polyvinyl alcohol (PVA; Gohsenol EG-40PW, Nippon Gohsei) solution to prepare a second aqueous layer (W2 layer) solution. The W1 / O emulsion was injected into the W2 layer solution at a rate of 18 ml / min while stirring at 8,000 rpm with a homogenizer to prepare a W1 / O / W2 emulsion.

[0117] The prepared W1 / O / W2 emulsion was heated from 15°C to 40°C for 1 hour and then maintained for 3 hours to volatilize. Thereafter, the temperature was reduced to 25°C. After filtration through a sieve with a mesh size of 75 μm, microspheres were collected by centrifugation. The collected microspheres were redispersed in distilled water and centrifuged to wash the microsphere surface three times. The washed microspheres were freeze-dried to obtain drug-encapsulated microspheres.

[0118] Hereinafter, depending on the type of API used, when semaglutide (Example 1) is used, it can be expressed as semaglutide microspheres (depot), and when tirzepatide (Example 2) is used, it can be expressed as tirzepatide microspheres (depot).

[0119]

[0120] [Comparative Example 1] Preparation of microspheres without calcium chloride

[0121] Microspheres were prepared in the same manner as in Examples 1 and 2, except that calcium chloride was not used. (Table 1)

[0122]

[0123] The main components of the microspheres manufactured in Examples 1 and 2 and Comparative Example 1 are listed in Tables 1, 2 and 3.

[0124]

[0125]

[0126] Classification W1 Phase (g) O Phase (g) W2 Phase (%(w / v)) Amount / API Polymer Type Polymer Amount CaCl2 Addition Concentration Comparative Example 1-11 / Semaglutide PLGA 504H90.00 Comparative Example 1-21 / Semaglutide PLGA 504H40.00 Comparative Example 1-31 / Semaglutide PLGA 502H40.00 Comparative Example 1-41 / Semaglutide PLGA 752H40.00 Comparative Example 1-51 / Semaglutide PLA 202H40.00 Comparative Example 1-61 / Tirzepatide PLGA 504H90.00

[0127]

[0128] Classification W1 Phase (g) O Phase (g) W2 Phase (%(w / v)) Amount / API Polymer Type Polymer Amount CaCl2 Addition Concentration Example 1-11 / Semaglutide PLGA 504H9 0.25 Example 1-21 / Semaglutide PLGA 504H9 0.50 Example 1-31 / Semaglutide PLGA 504H9 0.75 Example 1-41 / Semaglutide PLGA 504H9 1.00 Example 1-51 / Semaglutide PLGA 504H9 1.25 Example 1-61 / Semaglutide PLGA 504H9 1.50 Example 1-71 / Semaglutide PLGA 504H9 3.00 Example 1-81 / Semaglutide PLGA 504H60.25 Example 1-91 / Semaglutide PLGA 504H60.50 Example 1-101 / Semaglutide PLGA 504H60.75 Example 1-111 / Semaglutide PLGA 504H40.25 Example 1-121 / Semaglutide PLGA 504H40.50 Example 1-131 / Semaglutide PLGA 504H40.75 Example 1-141 / Semaglutide PLGA 504H41.50 Example 1-151 / Semaglutide PLGA 502H40.50 Example 1-161 / Semaglutide PLGA 752H40.50 Example 1-171 / Semaglutide PLA 202H40.50 Example 1-181 / Semaglutide PLA 202S40.50 Example 1-191 / Semaglutide PLGA 504 H: 502 H = 2: 141.50 Example 1-201 / Semaglutide PLGA 504 H: 752 H = 1: 140.50

[0129]

[0130] Classification W1 Phase (g) O Phase (g) W2 Phase (%(w / v)) Amount / API Polymer Type Polymer Amount CaCl2 Addition Concentration Example 2-11 / Tirzepatide PLGA 504H4 0.50 Example 2-21 / Tirzepatide PLGA 504H4 1.50 Example 2-31 / Tirzepatide PLGA 504H6 0.50 Example 2-41 / Tirzepatide PLGA 504H6 1.50 Example 2-51 / Tirzepatide PLGA 504H9 0.50 Example 2-61 / Tirzepatide PLGA 504H9 1.50 Example 2-71 / Tirzepatide PLGA 504H9 3.00

[0131]

[0132] [Comparative Example 2] Preparation of microspheres containing cationic salts other than calcium chloride

[0133] Microspheres were prepared in the same manner as in Example 1, except that sodium chloride (Comparative Examples 2-1 to 2-7, Comparative Examples 2-10 to 2-12), zinc chloride (Comparative Example 2-8), or magnesium chloride (Comparative Example 2-9) was used instead of calcium chloride as the cationic salt.

[0134] Additionally, microspheres were prepared in the same manner as in Example 2, except that sodium chloride (Comparative Example 2-13) was used instead of calcium chloride as the cationic salt.

[0135] The main components of the manufactured microspheres are listed in Table 4.

[0136]

[0137] Classification W1 Phase (mg) O Phase (mg) W2 Phase (%(w / v)) Amount / API Polymer Type Polymer Amount Salt Type Concentration Comparative Example 1-11 / Semaglutide PLGA 504H9--Comparative Example 1-61 / Tirzepatide PLGA 504H9--Comparative Example 2-11 / Semaglutide PLGA 504H9 NaCl 0.25 Comparative Example 2-21 / Semaglutide PLGA 504H9 NaCl 0.50 Comparative Example 2-31 / Semaglutide PLGA 504H9 NaCl 0.75 Comparative Example 2-41 / Semaglutide PLGA 504H9 NaCl 1.00 Comparative Example 2-51 / Semaglutide PLGA 504H9 NaCl 1.25 Comparative Example 2-61 / Semaglutide PLGA 504H9NaCl1.50Comparative Example 2-71 / Semaglutide PLGA 504H9NaCl3.00Comparative Example 2-81 / Semaglutide PLGA 504H6ZnCl20.50Comparative Example 2-91 / Semaglutide PLGA 504H6MgCl20.50Comparative Example 2-101 / Semaglutide PLGA 504H4NaCl0.25Comparative Example 2-111 / Semaglutide PLGA 504H4NaCl0.50Comparative Example 2-121 / Semaglutide PLGA 504H4NaCl0.75Comparative Example 2-131 / Tirzepatide PLGA 504H9NaCl3.00

[0138]

[0139] [Example 3 and Comparative Example 3] Preparation of microspheres containing plasticizer

[0140] A W1 / O / W2 emulsion was prepared by adding a plasticizer to the oil layer solution, and microspheres were prepared in the same manner as in Example 1 or 2.

[0141] In the case of Comparative Example 3-5, the manufactured W1 / O / W2 emulsion was heated to a temperature higher than the glass transition temperature (Tg) of the biodegradable polymer (PLGA 504H) for 1 hour and then maintained for 3 hours. In the case of Comparative Example 3-2, 100% polyvinyl alcohol (PVA) solution was added during volatilization, in the case of Comparative Example 3-3, volatilization was performed in a tank with a baffle added to the volatilization tank, and in the case of Comparative Example 3-4, 10% EtOH was added relative to the second aqueous layer (W2) during volatilization. In the cases of Comparative Examples 3-5, 3-7, 3-8, and 3-10, 3.5% EtOH relative to the oil phase was added as a cosolvent, and in the cases of Comparative Examples 3-6 and 3-9, 0.6% Tween 80 relative to the oil phase was added as a surfactant.

[0142] Additionally, in the case of Comparative Example 3-10, 3.5% EtOH was added as a co-solvent compared to the oil phase, and microspheres were manufactured in the same manner as in Example 2.

[0143] The main components of the microspheres manufactured in Example 3 and Comparative Example 3 are listed in Tables 5 and 6.

[0144]

[0145] Classification W1 Phase (g) O Phase (g) W2 Phase (%(w / v)) Process Addition Amount / API Polymer Type Polymer Good Solvent (%) Surfactant Addition Concentration (%(w / v)) CaCl2 or NaCl Addition Concentration - Comparative Example 1-11 / Semaglutide PLGA 504H9----Example 1-61 / Semaglutide PLGA 504H9--1.50 % CaCl2-Example 1-71 / Semaglutide PLGA 504H9--3.00 % CaCl2-Comparative Example 2-71 / Semaglutide PLGA 504H9--3.00 % NaCl-Example 1-141 / Semaglutide PLGA 504H4--1.50 % CaCl2-Comparative Example 3-11 / Semaglutide PLGA 504H9--1.50%CaCl2Volatilization temperature maintained above TgComparative example 3-21 / Semaglutide PLGA 504H9--3.00%CaCl2Add 100% of water phase during volatilizationComparative example 3-31 / Semaglutide PLGA 504H9--3.00%CaCl2Using tank with baffleComparative example 3-41 / Semaglutide PLGA 504H9--3.00%CaCl2Aqueous layer (W2)Cosolvent application (10% EtOH)Comparative example 3-51 / Semaglutide PLGA 504H93.5%EtOH-3.00%CaCl2-Comparative example 3-61 / Semaglutide PLGA 504H9-0.6% Tween 803.00%CaCl2-Comparative example 3-71 / Semaglutide PLGA 504H 9 3.5% EtOH - 3.00 % NaCl - Comparative Example 1-61 / Tirzepatide PLGA 504H 9 ---- Example 2-71 / Tirzepatide PLGA 504H 9 -- 3.0 % CaCl 2 - Comparative Example 3-81 / Tirzepatide PLGA 504H 9 3.5% EtOH - 3.0 % CaCl 2 - Comparative Example 3-91 / Tirzepatide PLGA 504H 9 - 0.6% Tween 8 0 3.0 % CaCl 2 - Comparative Example 3-101 / Tirzepatide PLGA 504H 9 3.5% EtOH ---

[0146]

[0147] Classification W1 Phase (g) O Phase (g) W2 Phase (%(w / v)) Amount / API Polymer Type Polymer Amphoteric Concentration (%(w / v)) CaCl2 or NaCl Addition Concentration Example 3-11 / Semaglutide PLGA 504H 9 7.0% Triethylcitrate 3.00 % CaCl2 Example 3-21 / Semaglutide PLGA 504H 9 2.5% Triethylcitrate 3.00 % CaCl2 Example 3-31 / Semaglutide PLGA 504H 9 1.1% Triethylcitrate 3.00 % CaCl2 Example 3-41 / Semaglutide PLGA 504H 9 0.3% Poloxamer 188 3.00 % CaCl2 Example 3-51 / Semaglutide PLGA 504H90.3%Poloxamer 4073.00%CaCl2Example 3-61 / Semaglutide PLGA 504H93.5%Triethylcitrate 3.00%NaClExample 3-71 / Semaglutide PLGA 504H93.0%Poloxamer 1883.00%NaClExample 3-81 / Semaglutide PLGA 504H90.3%Poloxamer 1883.00%NaClExample 3-91 / Semaglutide PLGA 504H43.5%Triethylcitrate 1.50%CaCl2Example 3-101 / Semaglutide PLGA 504H42.5%Triethylcitrate 1.50%CaCl2Example 3-111 / Semaglutide PLGA 504H4 1.1% Triethylcitrate 1.50% CaCl2 Example 3-121 / Semaglutide PLGA 504H4 0.3% Poloxamer 188 1.50% CaCl2 Example 3-131 / Semaglutide PLGA 504H4 0.3% Poloxamer 407 1.50% CaCl2 Example 3-141 / Semaglutide PLGA 504H9 2.5% Triethylcitrate-Example 3-151 / Semaglutide PLGA 504H9 0.3% Poloxamer 188-Example 3-161 / Tirzepatide PLGA 504H9 2.5% Triethylcitrate3.00 %CaCl2Example 3-171 / Tirzepatide PLGA 504H90.3%Poloxamer 1883.00 %CaCl2Example 3-181 / Tirzepatide PLGA 504H92.5%Triethylcitrate-Example 3-191 / Tirzepatide PLGA 504H90.3%Poloxamer 188-.

[0148]

[0149] [Reference] Test method

[0150] 1. Content evaluation

[0151] 5 mg of lyophilized microspheres as semaglutide or tirzepatide were placed in a 100 mL volumetric flask, dissolved in 50 mL of acetonitrile, adjusted to the mark with DW, filtered through a 0.45 syringe filter, and the amount of semaglutide or tirzepatide was quantified by high-performance liquid chromatography according to the following analysis conditions.

[0152]

[0153] Semaglutide analysis conditions Column Aegispak C8 (4.6 x 150 mm, 5.0 μm) or column of equivalent specifications Mobile phase MP A - 10 mM potassium dihydrogen phosphate solution (pH 2.0 by phosphoric acid) MP B - Acetonitrile Flow rate 1 mL / min Detector UV, 215 nm Gradient condition Time (min) MP A (%) MP B (%) 0 8 0 2 0 1 0 3 0 7 0 1 0 1 8 0 2 0 2 0 8 0 2 0 Tirzepatide analysis conditions Column Aegispak C18 (4.6 x 250 mm, 5.0 μm) or column of equivalent specifications Mobile phase MP A - 0.1 % Trifluoroacetic Acid in Water MP B - 0.1 % Trifluoroacetic Acid in Acetonitrile Flow rate 1 mL / min Detector UV, 220 nm Gradient condition Time (min) MP A (%)MP B (%)06040304555

[0154]

[0155] 2. Emission test

[0156] 10 mg of lyophilized microspheres as semaglutide or tirzepatide were dispersed in 10 mL of release solution (prepared by dissolving 1.42 g of di-sodium hydrogen phosphate dihydrate and 14 g of propylene glycol in 5 L and adjusting the pH to 7.4 with 1 N HCl) and a release test was performed using a Rotator (PTR-25) at 25 rpm and 37°C. The supernatant and microspheres were separated by centrifugation, and the amount of semaglutide or tirzepatide present in the supernatant was quantified by high-performance liquid chromatography. The analysis conditions were the same as those for the content analysis method.

[0157]

[0158] 3. SEM measurement

[0159] Approximately 10 mg of microspheres were fixed to an aluminum stub and coated with platinum for 3 minutes under a vacuum of 0.1 torr and a high voltage (10 kV). The microspheres were then mounted on a SEM powder and the surface of the microspheres was observed using an image analysis program.

[0160]

[0161] 4. Particle size measurement

[0162] The size of the microspheres was measured using a Malvern Mastersizer. Approximately 100 mg of the microspheres were suspended in 5 ml of distilled water, the suspension was placed in a dispersion device, and the size was measured after dispersion for 1 minute at 2,800 rpm and 50% ultrasound.

[0163]

[0164] 5. Measurement of osmotic pressure

[0165] 0.1 ml of the W2 layer was taken and placed in Osmotech's single-sample micro-osmometer to measure the osmotic pressure.

[0166]

[0167] 6. Plasma analysis

[0168] The concentration of test substances in plasma was analyzed using LC-MS / MS at the QbestBio Analysis Center. Plasma separation was performed at 12,000 rpm for approximately 2 minutes at 4°C. Pharmacokinetic analysis was performed non-compartmentally at the QbestBio Non-Clinical Evaluation Center using Phoenix® WinNonlin® software (Ver. 8.3, Certara). The LC-MS / MS analysis method for plasma samples is as follows.

[0169]

[0170] Analytical Conditions Column ACE C18 (2.1 x 50 mm, 3.0 μm), ACE Column Temperature 40 ºC Mobile Phase A 10 mM ammonium acetate in distilled water Mobile Phase B 0.1% formic acid in acetonitrile Mobile Phase Conditions Time (min) Flow (ml / min) AB 0 0.45 5 45 1.5 0.45 9 5 2.5 0.45 9 5 2.5 1 0.45 5 45 0.45 5 45

[0171]

[0172] 7. Measurement of MC residual solvent

[0173] 100 mg of the sample was dissolved in 1 mL of DMF, and the residual solvent was measured using Headspace-GC under the following analysis conditions.

[0174]

[0175] Analysis Conditions Column DB-624, 0.53 mm × 30 m, 3 μm, Agilent Technologies Column Temperature Maintain at 40°C for 5 minutes, then increase to 240°C at 20°C per minute for 10 minutes and maintain at 240°C for 5 minutes Flow Rate 1.2 mL / min Headspace Conditions Injection Volume 1 mL Equilibrium Temperature (°C) 80°C Equilibration Time (min) 30 min

[0176]

[0177] 8. DSC measurement

[0178] DSC measurements were performed in a differential scanning calorimeter. 5–10 mg of each microsphere was placed in an aluminum sample pan and measured from 25°C to 250°C at 5°C / min.

[0179]

[0180] 9. FTIR measurement

[0181] FTIR measurements were performed using a Nicolet iS5TM instrument from Thermofisher Scientific. To utilize attenuated total reflectance (ATR), approximately 2 mg of sample was applied to a diamond laminate plate, and the wavelength was 600 to 4,000 cm. -1 In , the resolution is 4 cm -1 At intervals, analysis was performed using a single beam.

[0182]

[0183] [Experimental Example 1] Confirmation of the physical properties and initial release control effect of the depot composition according to the calcium chloride content.

[0184] Tables 10 and 11 below show the results of measuring the physical properties and initial release effects of the depot compositions manufactured in the examples and comparative examples.

[0185]

[0186] Classification W2 phase CaCl2 concentration (w / v %) Osmolarity (mOsm) Residual solvent (ppm) Long-term release Day 1 (%) Comparative Example 1-10.005 33.25 15.30 Comparative Example 1-20.005 N.D 5 4.95 Comparative Example 1-30.005 N.D 5 9.11 Comparative Example 1-40.005 N.D 4 4.31 Comparative Example 1-50.005 N.D 7 1.27 Comparative Example 1-60.005 68.5 35.15 Example 1-10.25 50 N.D 7.96 Example 1-20.50 140 N.D 4.39 Example 1-30.75 180 N.D 3.11 Example 1-61.50 36 1279.70.13 Example 1-73.00 743 1183.000.00 Example 1-80.25 50 N.D9.08 Example 1-90.50 1402.60 6.00 Example 1-100.75 18021.612.69 Example 1-110.25 50 N.D35.79 Example 1-130.75 180 N.D28.72 Example 1-141.50 36 1 N.D18.55 Example 1-150.50 140 N.D29.39 Example 1-160.50 140 N.D24.50 Example 1-170.50 140N.D49.07 Example 1-180.50 140N.D34.58 Example 1-190.50 140N.D31.42 Example 1-200.50 140N.D19.22 Example 2-10.50 140N.D48.53 Example 2-21.50 361N.D10.39 Example 2-30.50 140N.D26.41 Example 2-41.50 361N.D8.43 Example 2-50.50 140N.D6.22 Example 2-61.50 361N.D0.00 Example 2-73.00 70 1483.80.00

[0187]

[0188] Classification W2 phase CaCl2 concentration (w / v %) Long-term release Day 1 (%) Rat PK test C max (ng / mL) Comparative Example 1-10.00 15.30 162 1.35 Example 1-30.75 3.11 263.20 Example 1-61.50 0.13 130.80 Comparative Example 1-60.00 35.15 1814.05 Example 2-61.50 0.00 183.50

[0189]

[0190] As shown in Table 10 above, it can be confirmed that the initial release of the depot composition is controlled when producing microspheres using calcium chloride. Furthermore, it can be confirmed that the initial release (first day of extended release) decreases proportionally depending on the amount of calcium chloride.

[0191] The long-term release on the first day of Comparative Example 1-1, which does not contain calcium chloride, was 15.3%, whereas the long-term release on the first day of Example 1-6, which contains 1.5% calcium chloride, was 0.13%, confirming that rapid initial release was controlled. Through this, it can be confirmed that the effect of reducing initial release is achieved even when the ratio of drug to polymer changes (Examples 1-8 to 1-10 and 1-11 to 1-14).

[0192] Meanwhile, in the case of Example 2-6 to which tirzepatide was applied, it can be confirmed that the initial release reduction effect is applied equally through comparison with Comparative Example 1-6 (Comparative Example 1-6, Example 2-6).

[0193] In addition, it can be confirmed that the initial release control effect is present even when the molecular weight of the polymer (PLGA) is different, as in Example 1-15 and Comparative Example 1-3, when the type of polymer (change in the ratio of polylactic acid / polyglycolic acid in PLGA) is different, as in Example 1-16 and Comparative Example 1-4, and when polylactide polymer (PLA) is used, as in Examples 1-17 and 18 and Comparative Example 1-5.

[0194] In addition, as shown in Table 11, the maximum plasma concentration (C) in animal experiments (administration of test drug in rat PK test) max ) can also be seen to decrease proportionally depending on the amount of calcium chloride.

[0195] As described in the present invention, when calcium chloride is used at a concentration of 0 to 3.0% in the preparation of a depot composition, it can be confirmed that a depot composition with low residual solvent is possible. However, when other cationic salts such as sodium chloride, magnesium chloride, or zinc chloride are used, it can be confirmed that the residual solvent has a high value of up to 2,200 ppm (Comparative Example 2-6). This may raise safety concerns when considering the allowable residual solvent amount standards for use in pharmaceuticals.

[0196] Figures 1a to 1c are SEM photographs of microspheres manufactured according to Example 1, where 1a is an SEM photograph of microspheres manufactured according to Example 1-3, 1b is an SEM photograph of microspheres manufactured according to Example 1-9, and 1c is an SEM photograph of microspheres manufactured according to Example 1-13. In addition, Figure 2 shows an SEM photograph of microspheres manufactured according to Comparative Example 1-1, and Figure 3 shows an SEM photograph of microspheres manufactured according to Comparative Example 2-7.

[0197] Figure 4a shows the animal PK profiles of semaglutide microspheres manufactured according to Comparative Example 1-1, Example 1-3, Example 1-5, and Comparative Example 2-7 of the present invention. In addition, Figure 4b shows the animal PK profiles of tirzepatide microspheres manufactured according to Comparative Example 1-6 and Example 2-6 of the present invention.

[0198] As shown in the above drawing, Comparative Examples 1-1 and 2-7, which did not use calcium chloride, had many pores on the surface of the microspheres, and thus, it was confirmed that the initial release of the API was also high. It was also confirmed that the initial release was reduced not only for semaglutide but also for tirzepatide.

[0199] Typically, microspheres are manufactured through a homogenization process, and the solvent used to dissolve the polymer is vaporized during the process, thereby solidifying the microspheres. In the present invention, during the solidification process, calcium chloride comes into contact with the polymer on the surface of the microspheres, and a gelation phenomenon occurs at the interface due to hydrophobic ion pairing, thereby coating the surface of the microspheres, thereby forming pore-free microspheres.

[0200] In particular, when calcium chloride is included in the W2 layer, it can have an initial release suppression effect of API while maintaining good properties. Preferably, when calcium chloride is included at a concentration of 0.25 to 3.0%, a depot composition with low residual solvent and suppressed initial release can be prepared.

[0201]

[0202] [Experimental Example 2] Confirmation of physical properties and initial release control effects according to the type of cationic salt.

[0203] Tables 12 and 13 below show the results of measuring the physical properties and initial release effect of the depot composition manufactured in Comparative Example 2.

[0204]

[0205] Distinctive salt type W2 phase Salt concentration (w / v %) Osmolarity (mOsm) Residual solvent (ppm) Long-term release Day 1 (%) Comparative example 1-1 NaCl 0.00 5 3 3.25 15.30 Comparative example 2-1 NaCl 0.25 9 2 1 1 8 1.03 2 1.86 Comparative example 2-4 NaCl 1.00 3 0 0 1 3 8 9.8 2 2.37 Comparative example 2-6 NaCl 1.5 0 5 6 8 2 20 0.00 2 4.46 Comparative example 2-7 NaCl 3.00 1 0 0 1 5 0 6.8 2 2.85 Comparative example 2-8 ZnCl 2 0.5 0 1 5 1 1 4 3 6.2 0 4 4.49 Comparative example 2-9 MgCl 2 0.5 0 1 6 5 1 2 0 9.4 5 2.87 Comparative example 2-11NaCl0.501811506.8569.10Comparative example 2-13NaCl3.0010211906.429.10

[0206]

[0207] Classification W2 phase NaCl concentration (w / v %) Long-term release Day 1 (%) Rat PK test Cmax (ng / mL) Comparative Example 1-10.00 15.30 1,621.35 Comparative Example 2-73.00 22.85 1,145.75

[0208] As shown in Table 12, when a depot composition was prepared using a cationic salt other than calcium chloride, it was confirmed that, unlike calcium chloride, there was no effect on the control of the initial release depending on the amount of salt. In addition, as shown in Table 13, the maximum plasma concentration (C) in animal experiments (administration of test drug in rat PK test) max ) also confirmed that the initial release control effect was minimal. In addition, the residual solvent exceeded the allowable amount standard (dichloromethane 600 ppm or less), making human administration impossible due to patient safety concerns.

[0209] When sodium chloride is used as a cationic salt, unlike calcium chloride, hydrophobic ion pairing gelation does not appear on the surface of the microspheres, so the initial release control effect is thought to be minimal. When comparing Example 1-6 and Comparative Example 2-6, which have the same cationic salt concentration of 1.5%, the osmotic pressure of Comparative Example 2-6 was 568 mOsm compared to the osmotic pressure of Example 1-6 (361 mOsm), and although they had an osmotic pressure that was equal or greater, the initial release was evaluated to be 0.13% and 24.46%, respectively. Based on these results, it can be confirmed that the initial release effect is not related to the osmotic pressure. In addition, in Comparative Example 2-6, a high concentration of MC, a toxic solvent, remains due to the osmotic pressure, which may cause safety issues.

[0210] In addition, when magnesium chloride or zinc chloride was used as a cationic salt, it was confirmed that the properties of the microspheres were poor and the yield was low, making formulation difficult. Compared to the 6.00% long-term release result on the first day of Example 1-9, the long-term release results on the first day of Comparative Example 2-8 (using magnesium chloride) and Comparative Example 2-9 (using zinc chloride) were 44.49% and 52.87%, respectively, confirming that the initial release in long-term release was not controlled in other polyvalent salts. In other words, it was confirmed that the initial release control effect of the drug was minimal in the cationic salt.

[0211] Through these test results, it can be confirmed that only when calcium chloride is used as a cationic salt as in the present invention, the initial release control effect for the active ingredient can be achieved within the allowable amount of residual solvent.

[0212] Looking at the osmotic pressure measurement results of the W2 layer of the examples and comparative examples, it is believed that the API early release control effect by calcium chloride is not related to osmotic pressure. In addition, since the W2 layer according to the cationic salt used in the examples and comparative examples was confirmed to have a pH between 5.6 and 5.8, it can be confirmed that the early release control effect is not due to a change in pH. In other words, it can be confirmed that the present invention has an API early release control effect through gelation caused by pairing of hydrophobic ions, which is a new mechanism different from osmotic pressure or pH.

[0213]

[0214] [Experimental Example 3] Confirmation of physical property effects according to the use and type of plasticizer.

[0215] Table 14 below shows the results of measuring the physical properties of the depot compositions manufactured in Example 3 and Comparative Example 3.

[0216]

[0217] ClassificationHigh molecular weight salt typeW2 phaseSalt concentration(w / v%)Process additionOil phasePlasticizer / additive concentration(w / v%)Residual solvent(ppm)Comparative example 1-19CaCl20.00--33.25Example 1-69CaCl21.50--279.7Example 1-79CaCl23.00--1,183.0Comparative example 2-74NaCl3.00--1,506.8Example 1-144CaCl21.50--NDComparative example 3-19CaCl21.50Maintain above volatilization temperature Tg-98.0Comparative example 3-29CaCl23.00Add 100% of water phase during volatilization-1,198.0Comparative example 3-39CaCl23.00Tank with baffle Use-1,184.0Comparative Example 3-49NaCl3.00Aqueous layer (W2) co-solvent application (10% EtOH)-1,099.0Comparative Example 3-59CaCl23.00-3.5% EtOHN.DComparative Example 3-69CaCl23.00-0.6%Tween 80126.0Comparative Example 3-79NaCl3.00-3.5% EtOH117.0Comparative Example 1-69CaCl20.00--68.5Example 2-79CaCl23.00--483.8Comparative Example 3-89CaCl23.00-3.5% EtOH228.1Comparative Example 3-99CaCl20.00-0.6% Tween 8055.8Comparative Example 3-109CaCl20.00-3.5%EtOH70.5Example 3-19CaCl23.00-3.5%TriethylcitrateN.DExample 3-29CaCl23.00-2.5%TriethylcitrateN.DExample 3-39CaCl23.00-1.1%TriethylcitrateN.DExample 3-49CaCl23.00-3.0%Poloxamer 188N.DExample 3-59CaCl23.00-0.3%Poloxamer 188N.DExample 3-69CaCl23.00-0.3%Poloxamer 407N.DExample 3-79NaCl3.00-3.5%TriethylcitrateN.DExample 3-89NaCl3.00-0.3%Poloxamer 188N.DExample 3-94CaCl21.50-3.5%TriethylcitrateN.DExample 3-104CaCl21.50-2.5%TriethylcitrateN.Example 3-114CaCl 2 1.50-1.1%TriethylcitrateN.D Example 3-124CaCl 2 1.50-0.3%Poloxamer 188N.D Example 3-134CaCl 2 1.50-0.3%Poloxamer 407N.D Example 3-149CaCl 2 0.00-2.5%TriethylcitrateN.D Example 3-159CaCl 2 0.00-0.3%Poloxamer 188N.D Example 3-169CaCl 2 3.00-2.5%TriethylcitrateN.D Example 3-179CaCl 2 3.00-0.3%Poloxamer 188N.D Example 3-189CaCl20.00-2.5%TriethylcitrateN.DExample 3-199CaCl20.00-0.3%Poloxamer 188N.D.

[0218]

[0219] As shown in Table 14, when a depot composition is prepared using a plasticizer, residual solvent can be controlled depending on the use and type of plasticizer. This demonstrates that the use of a plasticizer can significantly remove residual solvent, thereby addressing patient safety concerns.

[0220] It was confirmed that the residual solvent can be significantly reduced by applying a plasticizer even when changing to sodium chloride instead of calcium chloride, or when there is no calcium chloride, or when the type of drug is changed (semaglutide and tirzepapatide).

[0221] When the volatilization temperature is adjusted to a temperature higher than the glass transition temperature (Tg) of the biodegradable polymer (Comparative Example 3-1), the residual solvent is reduced, but it is difficult to effectively reduce the residual solvent, and the bioactive substance may be denatured by heat. In addition, when the temperature is raised above the Tg temperature of the polymer, there is a disadvantage in that the properties of the fine particles deteriorate. There is a method to reduce the residual solvent by increasing the water phase during the volatilization process (Comparative Example 3-2) or installing a baffle in the volatilization tank (Comparative Example 3-3), but in the results of this comparative example, the residual solvent rather increased or it was difficult to significantly reduce the residual solvent. In addition, it was confirmed that it was difficult to significantly reduce the residual solvent when a cosolvent was applied to the water phase (Comparative Example 3-4). In addition, it was difficult to reduce the residual solvent even when applying the solvent EtOH (Comparative Example 3-5, Comparative Example 3-7, Comparative Example 3-8, Comparative Example 3-10) or applying the surfactant Tween 80 (Comparative Example 3-6, Comparative Example 3-9).

[0222] When the plasticizer according to the present invention is used and the oil phase (oil phase) contains the plasticizer in the corresponding amount, the residual solvent can be controlled to the ND level.

[0223]

[0224] [Experimental Example 4] DSC and FTIR measurements of the depo-form compositions of the examples and comparative examples

[0225] Table 15 below shows the results of measuring DSC and FT-IR of the depot compositions manufactured in the examples and comparative examples.

[0226] In addition, FIGS. 5a to e show DSC results of microspheres manufactured according to 504H polymer (a) and Comparative Examples 1-1 (b), 2-7 (c), and Examples 1-3 (d), 1-6 (e) of the present invention.

[0227]

[0228] Differential salt type W2 phase salt concentration (w / v %) Osmotic pressure (mOsm) C=OFT-IRWavenummer (cm-1 )DSCTg(°C)504H polymer---1650.861537.5652.93Comparative Example 1-1-0.0051651.571538.2151.62Comparative Example 2-7NaCl3.0010001657.741546.9650.67Example 1-3CaCl20.751801657.721543.6353.60Example 1-6CaCl21.503611660.211539.5052.20

[0229]

[0230] As shown in Table 15 and Fig. 5 mentioned above, when comparing the case where the polymer alone was measured (Fig. 5a) with the case where no cationic salt was applied (Comparative Example 1-1, Fig. 5b), the change in the Tg value was minimal. When calcium chloride was used ((Example 1-3, Fig. 5d), (Example 1-6, Fig. 5e)), the Tg value increased somewhat, but the difference was minimal, and when sodium chloride was used (Comparative Example 2-7, Fig. 5c), the Tg value decreased somewhat, but the difference was minimal.

[0231] In addition, FIGS. 6a to 6e show the FT-IR results of microspheres manufactured according to 504H polymer (a) and Comparative Examples 1-1 (b), 2-7 (c), and Examples 1-3 (d), 1-6 (e) of the present invention.

[0232] In the above Fig. 6, when the polymer alone was measured (Fig. 6a) and when no cationic salt was used (Comparative Example 1, Fig. 6b), a change was observed near the FT-IR peak of the carboxyl group of the polymer, and when the cationic salt was used, a change was observed in the peak near the C=O peak. In particular, when calcium chloride was used ((Example 1-3, Fig. 6d), Example 1-6, Fig. 6e)), the higher the concentration, the higher the FT-IR peak of Comparative Example 1-1 at 1650.86 cm -1 are 1657.72 cm each -1 and 1660.21 cm -1 The peak shifted to 1537.56 cm -1 are 1543.63 cm each -1and 1539.50 cm -1 The peak shifted to 1657.74 cm even in the case of sodium chloride, which showed a minimal effect in the initial emission (Comparative Example 2-7, Fig. 6c). -1 and 1546.96 cm -1 As the peak shifted, the change in the peak was confirmed to be minimal despite the application of a high concentration.

[0233] Summarizing the above results, when calcium chloride is applied, DSC results remain unchanged, while FT-IR peak shifts are observed. This suggests that gelation occurs on the polymer surface due to a physical phenomenon called hydrophobic ion pairing caused by calcium chloride, resulting in a coating effect.

[0234]

[0235] [Experimental Example 5] Blood Glucose and HbA1c Measurement

[0236] Diabetic db / db mice were obtained at 5-6 weeks of age and allowed to drink water and feed ad libitum from Day 0 after an acclimatization period. Groups were separated based on body weight, and body weight and feed intake were measured once every two days until the end of the experiment. After subcutaneous drug administration, blood glucose levels were measured 13 times in total on Day 0 (before administration), 1 h, 2 h, 4 h, 6 h, 1, 2, 3, 5, 7, 14, 21, and 28 without fasting, and the measurement results are shown in Figures 7a and 7b.

[0237] In Fig. 7, G1 vehicle represents a case in which no experimental substance was administered, G2 represents a case in which a control drug (Ozempic) was administered at 0.06 mpk daily for 4 weeks, G3 represents a case in which the semaglutide depot composition according to the present invention (Example 1-12) was administered once at 6.72 mpk, and G4 represents a case in which the semaglutide depot composition according to the present invention (Example 1-12) was administered once at 16.8 mpk.

[0238] As shown in Fig. 7a, it can be confirmed that the G3 and G4 groups using the depot composition according to the present invention have a blood sugar lowering effect equivalent to or greater than the control drug. As a result of checking the blood sugar concentration for 4 weeks after administration, compared to the G1 (vehicle) group, the G2 (control drug) group showed a blood sugar lowering rate of 12.9%, and the G3 and G4 groups showed a blood sugar lowering rate of 18.4% and 13.5%, respectively, without the side effect of rapid blood sugar lowering.

[0239] In addition, as shown in Fig. 7b, it can be confirmed that the G3 and G4 groups using the depot composition according to the present invention have a blood glucose alkaloid reduction effect at a level equivalent to that of the control drug. Comparing the results 4 weeks after administration and before administration, the G3 and G4 groups showed a decrease of -18.00% and -18.79%, respectively, while the control drug showed a decrease of -18.49%, securing similar results.

[0240]

[0241] [Experimental Example 6] Body weight measurement

[0242] From the time of acquisition (Day 0) of 7-week-old mice, they were fed a 60% kcal% high-fat diet ad libitum for 22 weeks to induce obesity, except for the control group. The body weight and feed intake of each individual were measured twice a week until the time of necropsy. The amount of food consumed per cage per day from the start of the test to the week before necropsy was measured by dividing it by the number of mice. At the 18th week of induction, the mice were divided into groups based on body weight. After subcutaneous administration of the drugs of Example 1-12, blood glucose levels were measured weekly for 4 weeks. During the test period, body weight and feed intake were measured using the same scale on the same day of the week and at the same time to minimize errors in each measurement.

[0243] The measurement results are shown in Fig. 8. In Fig. 8, G1 represents a normal control rat (negative control group) that did not consume a high-fat diet, G2 vehicle represents a group that consumed a high-fat diet and was not administered the test substance, G3 represents a group that was administered the control drug at 0.12 mpk daily, G4 represents a group that was administered the semaglutide depot composition (Example 1-12) once at 10.08 mpk, and G5 represents a group that was administered the semaglutide depot composition (Example 1-12) once at 20.16 mpk.

[0244] As shown in Figure 8, weight loss effects comparable to or greater than those of existing drugs were observed at 10.08 mpk and 20.16 mpk. (Group G3: 29.2% weight loss compared to the vehicle group, Group G4: 23.0% weight loss compared to the vehicle group, Group G5: 29.4% weight loss compared to the vehicle group)

[0245]

[0246] The microspheres of the depot composition of the present invention contain calcium chloride in their aqueous layer, thereby controlling the release of excessive amounts of the active ingredient contained within the microspheres during the initial injection of the depot. Furthermore, they exhibit excellent suspending properties and significantly lower residual solvent levels, allowing users to consistently and safely obtain the effects of the active ingredient even when administered as an injection.

[0247] Additionally, the dosage of semaglutide and / or tirzepatide can be adjusted through the depot formulation, thereby providing improved medical therapy of semaglutide and / or tirzepatide.

Claims

1. A pharmaceutical composition for preventing or treating obesity or diabetes, comprising semaglutide or a pharmaceutically acceptable salt thereof or tirzepatide or a pharmaceutically acceptable salt thereof as an active ingredient, The dosage of GLP-1 receptor agonists for the prevention or treatment of obesity is 12 to 96 mg / 4 weeks, A pharmaceutical composition comprising a GLP-1 receptor agonist for the prevention or treatment of diabetes, the dosage being 4 to 80 mg / 4 weeks.

2. In paragraph 1, A pharmaceutical composition comprising semaglutide or a pharmaceutically acceptable salt thereof for the prevention or treatment of obesity; or tirzepatide or a pharmaceutically acceptable salt thereof, wherein the dosage is 28 to 48 mg / 4 weeks.

3. In paragraph 1, A pharmaceutical composition comprising semaglutide or a pharmaceutically acceptable salt thereof for the prevention or treatment of diabetes; or tirzepatide or a pharmaceutically acceptable salt thereof, wherein the dosage is 12 to 32 mg / 4 weeks.

4. In paragraph 1, A pharmaceutical composition comprising semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof, which is administered to a subject as a microneedle formulation or a depot formulation.

5. In paragraph 4, A pharmaceutical composition comprising 5 to 25 parts by weight of semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof, per 100 parts by weight of each preparation.

6. In paragraph 1, When administering a pharmaceutical composition to a subject, the AUC of semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof t / C max A pharmaceutical composition having a shelf life of 100 hours or more.

7. O / W2 type microspheres including an oil layer (O layer) and a second aqueous layer (W2 layer), or W1 / O / W2 type microspheres including a first aqueous layer (W1 layer), an oil layer (O layer), and a second aqueous layer (W2 layer), In the above O / W2 type microspheres, the oil layer (O layer) or the first aqueous layer (W1 layer) in the above W1 / O / W2 type microspheres comprises semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof. A depot composition, wherein the oil layer (O layer) of the O / W2 type microparticles or W1 / O / W2 type microparticles comprises a biodegradable polymer, and the second aqueous layer (W2 layer) comprises calcium chloride.

8. In paragraph 7, A depot composition, wherein the biodegradable polymer comprises a polymer selected from the group consisting of polylactide (PLA), polyglycolide (PGA) and poly(lactide-co-glycolide) (PLGA).

9. In paragraph 7, A depot composition comprising calcium chloride at a concentration of 0.1 to 5.0% (w / v) for the second water layer (W2 layer).

10. In paragraph 7, A depot composition wherein the oil layer (O layer) in the O / W2 type microspheres or W1 / O / W2 type microspheres additionally contains a plasticizer.

11. In paragraph 10, A depot composition, wherein the plasticizer is at least one selected from the group consisting of glycerin (glycerol), concentrated glycerin, sorbitol, propylene glycol, poloxamer series polymers, phthalate polymers, citrate derivatives, triacetin, and castor oil.

12. A step of preparing an oil phase (O layer) solution or mixing a first water phase 1 (W1 layer) solution and the oil phase (O layer) solution to prepare a W1 / O emulsion; and It includes a step of preparing an O / W2 emulsion or a W1 / O / W2 emulsion by adding the above oil layer (O layer) solution or W1 / O emulsion to a second water phase 2 (W2) solution, In the above O / W2 emulsion, the oil layer (O layer) solution or in the above W1 / O / W2 emulsion, the first water layer (W1 layer) solution comprises semaglutide or a pharmaceutically acceptable salt thereof; or tirzepatide or a pharmaceutically acceptable salt thereof; A method for producing a depot composition, wherein the oil layer (O layer) solution in the O / W2 emulsion or W1 / O / W2 emulsion contains a biodegradable polymer, and the second aqueous layer (W2 layer) solution contains calcium chloride.

13. In paragraph 12, A method for producing a depot composition, wherein calcium chloride is included in a concentration of 0.1 to 5.0% (w / v) relative to the entire second aqueous layer (W2 layer) solution.

14. In paragraph 12, A method for producing a depot composition, wherein the oil layer (O layer) in the O / W2 type microspheres or W1 / O / W2 type microspheres additionally contains a plasticizer.

15. In paragraph 12, A method for producing a depot composition, further comprising a step of drying and / or filtering the produced O / W2 emulsion or W1 / O / W2 emulsion and then centrifuging it to recover microspheres.

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