KR20190095833A - Phosphatidylcholine-free injectable composition for localized fat reduction without pain and side effect - Google Patents

Abstract

The present invention relates to a phosphatidylcholine-free injectable composition for reducing localized fat without pain and adverse side effects; and a method for preparing the same, wherein the phosphatidylcholine-free injectable composition for reducing localized fat comprises 1 (w/v)% to 6 (w/v)% of glycocholic acid or a salt thereof. An existing injectable composition for reducing localized fat, the DCA single injectable composition, or the PPC injectable preparation containing DCA as a solubilizer, causes inevitable side effects due to necrosis of fat cells, including pain, hematosis, edema, numbness, red spots, swelling, skin hardening, itchiness, nodules, etc. and thus causes pain, discomfort, anxiety, and also, low medication adherence in patients. However, the injectable composition for reducing localized fat of the present invention is safe and stable; exhibits excellent effects in inducing lipolysis and apoptosis; and can selectively induce lysis only in fat cells while the vitality of fibroblasts, muscle cells, and vascular endothelial cells remains high. Thus, the composition of the present invention can free the patients from the aforementioned side effects, thereby improving the quality of life and medication adherence in the patients.

Description

Phosphhatidylcholine-free injectable composition for localized fat reduction without pain and side effect}

The present invention relates to a phosphatidylcholine-free injectable composition for topical fat reduction and a method for preparing the same, and more particularly, to a phosphatidylcholine-containing topical fat-injectable composition for reducing fat, glycocholine acid or a salt thereof. It relates to a topical fat reducing injection composition and a method for preparing the same, which are characterized by including 1 (w / v)% to 6 (w / v)%.

Injection for localized fat reduction is a cosmetic surgery to inject drugs into patients for the purpose of lipolysis, apoptosis and necrosis of fat cells. Locally deposited fats are of particular interest to many. People with unwanted, convex or plump locally deposited fats may appear unattractive and old. These causes may be due to aging, lifestyle or genetic predisposition, and try to improve with exercise therapy and diet to overcome this, but the effect is limited. The current treatment methods for local fat reduction are surgical methods such as liposuction, lipoplasty, liposuction, and non-surgical methods such as medical devices, mesotherapy and off-label lipolysis injection. Surgical methods, however, take weeks or months to heal, and certain people, such as smokers and diabetics, can significantly delay healing, with fatal side effects of 20 deaths per 100,000 people, risk of general anesthesia, excessive bleeding, internal Potential complications and risks include organ damage, bacterial infections, scars, bruising, edema and pain. In the case of non-technical methods to solve this problem, there is a potential risk because safety and efficacy are not secured due to the absence of large-scale clinical trials under the supervision of health authorities. In order to overcome the risks of such surgical methods and the risks of non-surgical methods, the development of new clinically beneficial drugs is necessary. Against this backdrop, non-surgical topical injections for local fat reduction have been studied steadily and a representative case is PPC injection. The main components of PPC injection are phosphatidylcholine (3-sn-Phosphatidylcholine (PPC)) and the solubilizing agent Deoxycholic acid (DCA) composition, which is developed by A Nattermann GmbH, Germany, for the treatment of liver disease. Lipostabil® N iv developed for the prevention and treatment of fat embolism In addition, there are cases of off-label (PPC injection, Lipodissolve, injection-lipolysis, fat-away-injection) in which lipobin® strain, developed as an adjuvant therapy for cirrhosis due to cirrhosis, is administered as a topical fat reducing injection.

PPC has been regarded as a single active ingredient of lipolysis in DCA-incorporated PPC injections.However, in 2004, DCA formulated as a solubilizer dissolves cell membranes and induces cell lysis by inducing a inflammatory response. It has been shown to necrosis the necrosis, and in fact DCA has also been found to be the active ingredient of topical fat reduction (Rotunda AM, Suzuki H, Moy RL, Kolodney M., Detergent effects of sodium deoxycholate are a major feature of an injectable phosphatidylcholine formulation used for localized fat dissolution.Dermatol Surg 30 (7): 1001-8 (2004).

However, the combination of PPC and DCA is not good for pain (78.4%), hematoma (83.8%), erythema (100%), burning sensation (100%), edema (100%), and hardening (66.7%). Responses were reported and DCA alone was reported for pain (100%), bruising (91.9%), erythema (100%), burning sensation (100%), edema (100%), and stiffness (89.2%) (Giovanni Salti). , Phosphatidylcholine and sodum deoxycholate in the treatment of localized fat: A doube-blind, randomized study, Dermatol Surg 34: 60-66, 2007), Pain results (73.6%), hematoma (72.9%), edema (67.8) %), Anesthesia (65.5%), erythema (35.3%), swelling (29.1%), sclerosis (28.3%), pruritus (16.3%), nodules (14.3%) were reported (Humphrey et al, ATX -101 for reduction of submental fat: A Phase III randomized controlled trial, J AM ACAD DERMATOL Vol 75, No. 4, 788-797, 2016). These side effects are due to a mechanism of action where fat cells become necrotic due to a rapid inflammatory reaction, which blocks the oxygen supply to cells after DCA subcutaneous injection, causing blisters and damage to the cell membrane after immediate cell expansion (Duncan, Injectable therapies for localized). fat loss: state of the art, Clin Plastic Surg, 1-13, 2011). DCA also damages musculoskeletal cells as well as adipocytes (Dong-Seok Kim, Phosphatidylcholine induces apoptosis of 3T3-L1 adipocytes, Journal of biomedical science, 18:91, 1-7, 2011).

Due to the side effects mentioned above, all patients seeking topical fat reduction injections complain of pain, discomfort, and anxiety.

When cells die in vivo, there is a significant difference between necrosis and death (apoptosis). Apoptosis is the active death caused by the regulation and expression of several genes and proteins in response to a programmed signal inside the cell. The apoptosomes produced by the process are the cells or macrophages that surround them. It is eliminated by phagocytosis such as macrophage and does not cause inflammation. On the other hand, necrosis is a passive death caused by changes in the external environment and undergoes the process of irregular clumping and swelling of cytoplasm, and finally the degradation of cells. Cell debris are produced and are known to cause inflammation (Earnshaw, WC, Curr. Opin. Cell Biol., 7, pp 337-343, 1995).

Therefore, if topical lipolytic agent acts in the manner of cell necrosis, inflammatory action also affects the surrounding area other than the target site to which the drug is administered, thus affecting the cells that should function normally and die. Putting it all together, pain, hematoma, swelling, numbness, erythema, swelling, sclerosis, pruritus, and nodule, which are present in existing topical liposomal injections, only lipolysis and apoptosis in cells where the drug is acting. It is important to induce high efficiency.

Therefore, the present inventors have been researching to develop injections having excellent local fat reduction ability without side effects such as pain and hematoma, swelling, numbness, erythema, swelling, sclerosis, pruritus and nodules, and are widely used in conventional topical lipolytic injections. Whereas deoxycholic acid acts in a way that induces necrosis of adipocytes, glycocholic acid and its salts reduce adipocytes by pharmacological mechanisms of lipolysis and apoptosis, thus suffering And it was confirmed that having excellent lipolysis and fat cell removal effect without side effects, completed the present invention.

Therefore, an object of the present invention, in the phosphatidylcholine-containing topical fat reducing injection composition, the composition is characterized in that it comprises 1 (w / v) to 6 (w / v)% glycocholic acid or salts thereof And to provide an injection composition for topical fat reduction without side effects.

Another object of the present invention, in the method for producing a topical fat reducing injection composition without pain and side effects, the composition is 1 (w / v)% to 6 (Glycocholic acid (GCA) or salts thereof) It includes w / v)% but does not include phosphatidylcholine, to provide a method for producing a topical fat reducing injection composition without pain and side effects.

In order to achieve the above object, the present invention is a composition for injection of a topical fat reduction containing phosphatidylcholine, wherein the composition comprises 1 (w / v)% to 6 (w / v)% of glycocholine acid or a salt thereof It provides a topical fat reducing injection composition without pain and side effects.

In order to achieve another object of the present invention, the present invention provides a method for producing a topical fat reducing injection composition without pain and side effects, the composition is glycocholic acid (Glycocholic acid, GCA) or salts thereof (w / v)% to 6 (w / v)% but does not include phosphatidylcholine, it provides a method for producing a topical fat reducing injection composition without pain and side effects.

Hereinafter, the present invention will be described in detail.

Terms

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Although any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention, preferred methods and materials are described.

As used herein, each of the following terms has the meaning associated with it in this section.

The articles "a" and "an" are used herein to refer to one or more (ie, at least one) of the grammatical objects of the article. For example, "an element" means one or more elements.

As used herein, when referring to measurable values such as amounts, temporal lengths, etc., "about" is ± 20%, ± 10% from the specified value, since the following variations are appropriate for carrying out the disclosed method. %, ± 5%, ± 1%, or ± 0.1%.

The disease or disorder is "alleviated" if the severity of the symptoms of the disease or disorder, the frequency of such symptoms experienced by the patient, or both are reduced.

In the present invention, the term “phosphatidylcholine” refers to a compound represented by IUPAC name 1,2-diacyl-sn-glycero-3-phosphocholine as phospholipid and is described herein as PPC.

As used herein, the term “bile acid” includes steroid acids (and / or carboxylic acid anions thereof), and salts thereof, which are found in bile of animals (eg, humans). As,

'Deoxycholic acid' is a type of bile salt, IUPAC Name (4R) -4-[(3R, 5R, 8R, 9S, 10S, 12S, 13R, 14S, 17R) -3,12-dihydroxy -10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta [a] phenanthren-17-yl] pentanoic Means a compound represented by acid. Described as DCA in the context of the present invention.

'Gycocholic acid' is a type of bile salt, IUPAC Name 2-[[(4R) -4-[(3R, 5S, 7R, 8R, 9S, 10S, 12S, 13R, 14S, 17R)- 3,7,12-trihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta [a] phenanthren-17-yl] pentanoyl] amino] acetic acid. Described herein as GCA.

'Taurocholic acid' is a type of bile salt. IUPAC Name 2-[[(4R) -4-[(3R, 5S, 7R, 8R, 9S, 10S, 12S, 13R, 14S, 17R) -3,7,12-trihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta [a ] phenanthren-17-yl] pentanoyl] amino] ethanesulfonic acid. Described herein as TCA.

'Cholic acid' is a type of bile salt, IUPAC Name (4R) -4-[(3R, 5S, 7R, 8R, 9S, 10S, 12S, 13R, 14S, 17R) -3,7,12 -trihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta [a] phenanthren-17-yl ] means a compound labeled with pentanoic acid. Described herein as CA.

'Chenodeoxycholic acid' is a type of bile salt, IUPAC Name ((4R) -4-[(3R, 5S, 7R, 8R, 9S, 10S, 13R, 14S, 17R) -3,7 -dihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta [a] phenanthren-17-yl ] pentanoic acid, referred to herein as CDCA.

'Ursodeoxycholic acid' is a type of bile salt, IUPAC Name (4R) -4-[(3R, 5S, 7S, 8R, 9S, 10S, 13R, 14S, 17R) -3,7 -dihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta [a] phenanthren-17-yl ] means a compound labeled with pentanoic acid. Described herein as UDCA.

'Glycodeoxycholic acid' is a type of bile salt, IUPAC Name 2-[[(4R) -4-[(3R, 8R, 9S, 10S, 12S, 13R, 14S, 17R) -3, 12-dihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta [a] phenanthren-17- yl] pentanoyl] amino] acetic acid. Described herein as GDCA.

Taurodeoxycholic acid is a type of bile salt, IUPAC Name 2-[[(4R) -4-[(3R, 5R, 9S, 10S, 12S, 13R, 14S, 17R) -3 , 12-dihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta [a] phenanthren-17 -yl] pentanoyl] amino] ethanesulfonic acid. Described herein as TDCA.

'Hyodeoxycholic acid' is a type of bile salt, IUPAC Name (4R) -4-[(3R, 5R, 6S, 8S, 9S, 10R, 13R, 14S, 17R) -3,6- dihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta [a] phenanthren-17-yl] Means a compound labeled with pentanoic acid. Described herein as HDCA.

'Lithocholic acid' is a type of bile salt, IUPAC Name (4R) -4-[(3R, 5R, 8R, 9S, 10S, 13R, 14S, 17R) -3-hydroxy-10,13- dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta [a] phenanthren-17-yl] pentanoic acid it means. Described herein as LCA.

Taurorsodeoxycholic acid is a type of bile salt, and IUPAC Name 2-[[(4R) -4-[(3R, 5S, 7S, 8R, 9S, 10S, 13R, 14S, 17R ) -3,7-dihydroxy-10,13-dimethyl-2,3,4,5,6,7,8,9,11,12,14,15,16,17-tetradecahydro-1H-cyclopenta [a] phenanthren-17-yl] pentanoyl] amino] ethanesulfonic acid. Described herein as TUDCA.

'Dehydrocholic acid' is a type of bile salt, IUPAC Name (4R) -4-[(5S, 8R, 9S, 10S, 13R, 14S, 17R) -10,13-dimethyl-3,7 , 12-trioxo-1,2,4,5,6,8,9,11,14,15,16,17-dodecahydrocyclopenta [a] phenanthren-17-yl] pentanoic acid. Described herein as DHCA.

The terms “patient”, “subject”, “individual”, etc., are used interchangeably herein and are in vitro or in situ that are compatible with the methods described herein. Refers to any animal, or cell thereof. In certain non-limiting embodiments, the patient, subject or individual is a human.

As used herein, the term "composition" or "pharmaceutical composition" refers to at least one compound of the invention and carriers, stabilizers, diluents, dispersants, suspending agents, thickening agents. A mixture of other chemical components, such as thickening agents, and / or excipients. The pharmaceutical composition facilitates administration of the compound to an organism.

As used herein, the terms "effective amount", "pharmaceutically effective amount" and "therapeutically effective amount" are non-toxic but do not produce the desired biological outcome. Indicates an amount sufficient to provide. The result can be a reduction and / or alleviation of the signs, symptoms, or cause of the disease, or any other desired alteration of the biological system. Appropriate therapeutic amounts in any individual case can be determined by one of skill in the art using routine experimentation.

As used herein, the term "efficacy" refers to the maximum effect (Emax) achieved within the assay.

As used herein, “instructional material” delivers the utility of a compound, composition, vector, or delivery system of the invention in a kit for alleviating the various diseases or disorders referred to herein. Publications, recordings, diagrams, or any other presentation medium that may be used to do so. Alternatively, or alternatively, the indicator material may describe one or more methods for alleviating a disease or disorder in mammalian cells or tissues. The indicator material of the kit of the present invention may be attached to, for example, a container comprising the identified compound, composition, vector, or delivery system of the present invention, or the identified compound, composition, vector, Or with a container containing a delivery system.

Alternatively, the indicator material may be shipped separately from the container with the intention that the indicator material and the compound will be used cooperatively by the recipient.

"Local administration" means administration of a pharmaceutical component to, or around, a muscle or subdermal location of a patient by non-systemic routes. Thus, topical administration excludes administration via systemic routes such as intravenous or oral administration.

"Pharmaceutically acceptable" means a property that is acceptable to a pharmacologist who manufactures from a pharmacological / toxicological point of view and a physical / chemical point of view regarding composition, formulation, stability, patient acceptability and bioavailability. And / or material. “Pharmaceutically acceptable carrier” refers to a medium that does not interfere with the host administered without interfering with the effectiveness of the biological activity of the active ingredient (s).

A "therapeutic" treatment is a treatment administered to a subject who exhibits the pathological signs or symptoms for the purpose of reducing or eliminating the signs or symptoms.

As used herein, the term "treatment" or "treating" means a condition contemplated herein, the symptoms of a condition contemplated herein or the potential to progress to a condition contemplated herein. Therapeutic agents to cure, heal, alleviate, relieve, alter, remedy, ameliorate, improve or influence sex That is, defined as the application or administration of a compound of the invention (alone or in combination with other pharmaceutical agents) to a patient or to tissues or cell lines isolated from the patient (eg for diagnostic or in vivo applications). Application or administration of a therapeutic agent (eg, for diagnostic or in vivo application), and the condition contemplated herein, the symptoms of the condition contemplated herein, or the present It has the potential to evolve into consideration the conditions in the processor. The treatment may be specifically tailored or modified based on knowledge obtained from the field of pharmacology.

A “therapeutically effective amount” is an amount of a compound of the invention that, when administered to a patient, ameliorates the symptoms of the disease. The amount of the compound of the present invention which constitutes a "therapeutically effective amount" may vary depending on the compound, the disease state and its severity, the age of the patient being treated, and the like. A therapeutically effective amount can be routinely determined by one of ordinary skill in the art in view of his knowledge and the present disclosure.

Scope: Throughout this disclosure, various aspects of the invention may be proposed in a range format. It is to be understood that the description in range format is merely for convenience and simplicity and should not be construed as an inflexible limitation on the scope of the invention. Accordingly, the description of a range should be considered to specifically disclose all possible subranges as well as individual numerical values within that range. For example, descriptions in the range 1-6 may include individual values within the range, for example 1, 2, 2.7, 3, 4, 5, 5.3, and 6, as well as 1-3, 1-4. And subranges such as 1 to 5, 2 to 4, 2 to 6, 3 to 6, and the like, should be considered as having specifically disclosed. This applies regardless of the width of the range.

In the present invention, the term '%' means a content of w / v% unless otherwise specified.

Hereinafter, the present invention will be described in detail.

The inventors of the present invention, unlike conventional bile salt solubilizers such as DCA is known to cause necrosis in adipocytes and fibroblasts, glycocholine acid is 1 (w / v)% to 6 (w / v)% The contribution of lipolysis and apoptosis-specific effects at concentrations has been newly identified. That is, in contrast to conventional DCA-containing PPC preparations (PPC-DC preparations) or DCA alone preparations that cause necrosis in adipocytes and other cells in the periphery, the present inventors have found that lipolysis does not occur without adipocyte necrosis. Pharmacological mechanisms of apoptosis and apoptosis resulted in the preparation of a new GCA-only formulation that is fat-free and safe with no side effects. Accordingly, the topical fat reducing injection composition of the present invention containing glycocholine acid or a salt thereof in a specific concentration has an excellent lipolysis effect, and only apoptosis (apoptisos) is applied only to the cells of the topical site to which the drug is administered. Induced by high efficiency, not only the fat cell removal effect is remarkable, but also significantly reduces the side effects and pains such as hematoma, edema, numbness, erythema, swelling, sclerosis, pruritus, nodules in the existing topical reducing injection composition. This is a revolutionary invention that improves patient compliance, improving pain, discomfort and anxiety.

Specifically, the present invention,

Phosphatidylcholine Not included  Topical Fat Reduction Injectables In the composition The composition is Glycocholine acid  Or 1 (w / v)% to 6 (w / v)% of salts thereof, and provides an injection composition for reducing local fat without pain and side effects.

The present invention relates to a non-surgical method, ie, an injection for topical fat reduction that is used alone, without accompanying surgery (eg liposuction, etc.). That is, the composition of the present invention is used for the purpose of reducing subcutaneous fat accumulation in mammals without performing surgical (surgical) treatment.

The composition of the present invention relates to a phospholipid-free composition such as phosphatidyl. That is, the composition of the present invention relates to a composition which does not contain phosphatidylcholine as well as a phospholipid component.

In addition, the composition of the present invention may be a composition that does not include a lipase or a lipase.

The injectable composition of the present invention may be applied to a topical site, and the site is not limited thereto, but is preferably deposited on the abdomen, submandibular, forearm, thigh, waist, hip, under eyes, and brasere line. It can be applied to the fat or lipoma site.

The topical fat reducing injection composition of the present invention is a pharmaceutical composition for the treatment of adipose tissue hyperproliferation or hyperaccumulation disorder (disease), the disorder being of the kind if it is known in the art that the adipose tissue is hyperproliferation or hyperaccumulation. Is not particularly limited but includes, for example, obesity (such as abdominal obesity), lower eyelid prolapse, lipoma, lipodystrophy, or fat accumulation associated with cellulite and the like.

 The method of administering the injectable composition of the present invention is not limited, but may be administered by a method suitable for the patient in view of the severity of the disease, the age, sex and other conditions of the patient, and the like. The route is not particularly limited in the method, but subcutaneous injection, intradermal injection, and the like are preferable.

In the present invention, the pain includes both pain during injection and / or pain after administration. The present invention suffers from conventionally available phosphatidylcholine-deoxycholine acid combinations (PPC-DCA such as lipostavill) or deoxycholineic acid (DCA) single agents (eg Kybella). Contrary to its involvement, it is characterized by significantly reduced pain (substantial painlessness). Such analgesic topical fat reducing injections are disclosed for the first time in the present invention, and in particular, pains caused by adverse reactions (side effects) such as inflammation and sclerosis, which occur secondly after the injection, as well as existing injections during and immediately after the injection. It is characterized by painlessness in the primary pain that is manifested in their own nature.

In the present invention, the term side effect refers to a phosphatidylcholine-deoxycholine acid combination (PPC-DCA, for example, lipostabil) or deoxycholine acid (DCA), which is a known topical or commercially available topical fat reducing injection. Means a side effect of the formulation (for example, Kybella), means a harmful action to the human body other than the main action (in the present invention, the desired fat reduction effect) that the drug expects a therapeutic effect. It is not limited to this, but specifically means any one or more selected from the group consisting of hematoma, edema, numbness, erythema, swelling, sclerosis, pruritus, nodules. The topical fat reducing injectable composition of the present invention is reduced to an extent that local adverse reactions are alleviated and the side effects are substantially absent.

Injectables are those dissolved in glycocholine acid or its salts and other additives, if necessary, in water for injection, filtered through a bacterial filter and aseptically filled and filled in vials, ampoules, or pre-filled syringes in a sterile state. When preparing an injection, water for injection may be used in addition to water to fill the remaining amount. The water for injection is not particularly limited as long as it is distilled water for injection or buffer for injection prepared for dissolving a solid injection or diluting a water-soluble injection, but for example, phosphate buffer solution or sodium dihydrogen phosphate (pH 3.5 to 7.5). NaH ₂ PO ₄ ) -citric acid buffer solution and the like can be used. The phosphate used herein may be in the form of sodium or potassium salts or in the form of anhydrides or hydrates, or in the form of citric acid or anhydrides or hydrates. Examples of water for injection include glucose injection, xylitol injection, D-mannitol injection, fructose injection, physiological saline, dextran 40 injection, dextran 70 injection, amino acid injection, Ringer's solution, lactic acid-ringer's solution, and the like. It is not limited. In the present invention, the water for injection includes all the usual aqueous vehicles used in the field of injection preparation in addition to the above, and the kind of water for injection is not particularly limited, and an excipient component (for example, used in a conventional injection) It also includes an aqueous vehicle that additionally includes sorbitol.

Specifically, the composition for reducing local fat of the present invention is characterized in that it comprises 'glycocholic acid or salt thereof'. The glycocholine acid is a bile acid having a structure of <Formula 1>, and has a molecular weight of about 465.63 g / mol and may be described as GCA or GC herein. The glycocholine acid may be used in the form of a pharmaceutically acceptable salt. As used herein, "pharmaceutically acceptable" means a physiologically acceptable and typically does not cause an allergic or similar reaction when administered to a human, but may be, for example, sodium salt, potassium salt or ammonium salt. have. Preferably, the glycocholine salt of the present invention may be sodium glycocholate (GCNa) represented by, for example, <Formula 2>.

The glycocholic acid or a salt thereof may be extracted and used in the intestine of an animal according to methods known in the art, and may be used commercially or prepared by chemical synthesis known in the art. have.

<Formula 1>

<Formula 2>

In the topical lipolysis injection composition of the present invention, the glycocholine acid is characterized by including 1.0% to 6.0% (w / v) based on the whole composition. If the concentration of glycocholine is less than 1.0% (w / v), there is no lipolytic effect, and when it exceeds 7.0% (w / v), side effects such as pain, swelling, and inflammation may occur severely.

Injectable composition for local fat decomposition of the present invention is characterized in that having a pH 6.0 to pH 8.0, preferably may be adjusted to pH 7.0 to pH 7.8. If it is less than pH 7.0, it is not good in pain, and if it is above pH 7.8, it is not good in pain.

The fat reducing injection composition of the present invention comprising the glycocholic acid (GCA) or salts thereof is excellent in formulation stability at pH 7.0 to pH 7.8 as described above, and the adipocyte lysis injection containing deoxycholine acid is GCA is highly efficient at concentrations of 1.0% to 6.0% (w / v), unlike necrosis, which cause pain in the body and cause side effects such as hematoma, edema, numbness, erythema, swelling, hardening, pruritus and nodules. It induces synergism of lipolysis and apoptosis, alleviates pain and side effects substantially, and shows excellent synergistic effect on fat reduction (pain and edema are 80 Less than%), thus the need for a separate anti-inflammatory or / and analgesic component not necessarily included or used in the composition, or One feature is that Therefore, the composition of the present invention may be characterized in that it does not contain anti-inflammatory agents and analgesics.

On the other hand, the composition of the present invention, based on the total composition of 0.1 (w / v)% to 5 (w / v)% preservative, 0.1 (w / v)% to 10 (w / v)% isotonic agent and It may further comprise one or more substances selected from the group consisting of 0.1 (w / v)% to 5 (w / v)% pH adjusting agent.

The preservative is not limited thereto, but is selected from the group consisting of benzyl alcohol, thimerosal, benzalkonium chloride, phenyl mercury nitrate, and chlorobutanol. It may be one or more. More preferably benzyl alcohol. Benzyl alcohol is a colorless and transparent liquid as one of aromatic alcohols. The concentration of benzyl alcohol included in the injectable composition of the present invention may be preferably 0.1 (w / v)% to 2 (w / v)%.

The isotonic agent serves to properly maintain (modulate) the osmotic pressure when administering the composition of the present invention to the body, and may also exhibit the side effect of further stabilizing GCA in solution. Isotonic agents can be pharmaceutically acceptable sugars, salts, or any combination or mixture thereof, such as glucose as sugar, or sodium chloride, calcium chloride, sodium sulfate, glycerine, propylene glycol, molecular weight as a water-soluble inorganic salt. The polyethylene glycol may be 1000 or less, and more preferably sodium chloride. They can be used in the form of one or two or more combinations. The concentration of the tonicity agent is preferably 0.1 (w / v)% to 5 (w / v)%, and a solution formulation including all of the mixtures according to the type, amount, etc. of the components included in the composition of the present invention. It may be adjusted to an appropriate amount so that the isotonic solution.

The pH adjusting agent of the present invention serves to adjust the pH of the injection, and includes both acidic and basic substances. The acidic materials include, but are not limited to, hydrochloric acid, acetic acid, adipic acid, ascorbic acid, sodium ascorbic acid, sodium ether, malic acid, succinic acid, tartaric acid, fumaric acid, citric acid (citric acid), and the like. The basic materials include inorganic bases (e.g. sodium hydroxide, potassium hydroxide, sodium carbonate, sodium bicarbonate, magnesium carbonate, calcium carbonate, magnesium oxide, ammonia, synthetic hydrotalcite), organic bases (e.g., basic amino acids ( (Eg, lysine, arginine, meglumine, etc.)), and the like. More preferably, the pH adjusting agent of the present invention may be sodium hydroxide or hydrochloric acid. The amount of the pH adjusting agent to be added may vary depending on the type and amount of components constituting the composition of the present invention, preferably 0.1 (w / v)%) to 3 (w / v)%. As described above, the composition of the present invention may be preferably provided in the range of pH 7.0 to pH 7.8, and the type and amount of the pH adjuster according to this may be changed by those skilled in the art according to the specific composition of the solution.

In a preferred embodiment, the topical fat reducing injectable composition of the present invention

1 (w / v)% to 6 (w / v)% of glycocholine acid or salts thereof;

Preservative, Tonicity  And 0.1 or more (w / v)% to 20 (w / v)% of any one or more selected from the group consisting of pH adjusting agents. And

It may be made of a residual amount of water for injection (water), it can be referred to the above bar for the type and composition ratio of the specific components of the composition.

The total effective amount of the injectable composition of the present invention may be administered to a patient in a single dose and may be administered by a fractionated treatment protocol which is administered in multiple doses for a long time. The pharmaceutical composition of the present invention may vary in dosage depending on the amount of fat at the target site, the location of the target site, the type of fat composition and the desired result. For example, it is administered subcutaneously or subcutaneously via subcutaneous injection using a syringe to the target site. The target site can be, for example, from 0.1 cm x 0.1 cm to 5 cm x 5 cm wide. Preferably the preferred total dose of the composition of the invention may be from 0.1 ml to 1.0 ml per cm ² , most preferably from 0.2 ml to 0.5 ml per cm ² . However, since the dosage of the injectable composition is determined in consideration of various factors such as the age, weight, health condition, sex, severity of the disease, diet and excretion rate, as well as the route of administration and the number of treatments of the composition, In view of this, one of ordinary skill in the art will be able to determine the appropriate effective dosage for the particular use of the composition as a therapeutic agent.

In another aspect, the present invention provides a method for producing a topical fat reducing injection composition without pain and side effects,

The composition is Glycocholine acid ( Glycocholic  acid, GCA ) Or salts thereof 1 (w / v)% to 6 (w / v)% but not phosphatidylcholine Characterized by It provides a method for preparing an injection composition for topical fat reduction, free of pain and side effects.

The manufacturing method may be prepared according to the method of mixing, stirring and filtration commonly used in the art, according to the composition of the topical fat reducing injection composition without the pain and side effects of the present invention described above. The kind and composition ratio of the specific component of the said composition are referred to the above-mentioned bar.

In a preferred embodiment, the present invention

(a) based on the total composition Glycocholine acid ( Glycocholic  acid) or its 1% salt (w / v)  Adding to 6 (w / v)% of water for injection and stirring until the solution is transparent;

(b) in step (a) Stirred  In the mixture Tonicity  And adding 0.1 (w / v)% to 15 (w / v)% preservative and stirring; And

(c) adjust the total volume with water (injectable water) after adjusting pH Stirring  Including a step, phosphatidylcholine is provided, characterized in that it provides a method for producing a topical fat reducing injection composition without pain and side effects.

The method for producing a topical fat reducing injection composition of the present invention will be described step by step below.

In step (a), glycolic acid (Glycocholic acid) or a salt thereof is added to the water for injection, and the solution is stirred until the solution becomes substantially transparent. In this case, glycocholine acid or a salt thereof is characterized in that it comprises 1.0% to 6.0% (w / v) based on the whole composition.

Stirring or mixing in the present invention may be by a known stirring means (stirrer), depending on the type or nature of the material to be added to those skilled in the art to improve the working efficiency conditions such as temperature, pressure, time or / and rotation speed It can be different.

In the step (a), administration of the pH adjusting agent may be additionally performed, and may preferably be preceded by glycocholic acid or salts thereof. The adjustment of pH may be performed by adding and stirring one or two or more pH adjusting agents at 0.1 (w / v)% to 5 (w / v)%. Specific types of pH adjusting agents that can be used in the present invention and their inclusion amounts are as described above.

In step (b), 0.1 (w / v)% to 5 (w / v)% of a preservative and 0.1 (w / v)% to 10 (w / v)% of an isotonic agent are added and stirred based on the total composition. . Specific components and concentrations of the preservatives and tonicity agents are as described above in the description of the composition.

In step (c), after adjusting the pH, adjust the total volume with water and mix homogeneously.

The adjustment of the pH may be performed by further adding and stirring one or more mixtures of pH regulators at 0.1 (w / v)% to 5 (w / v)%. Specific types of pH adjusting agents that can be used in the present invention and their inclusion amounts are as described above.

The water may be replaced with water for injection, as described above. In addition, in order to ensure product stability in accordance with the distribution of the injection formulation in this step to prepare a very stable injection by physically or chemically by adjusting the pH using an acid solution or a buffer solution (pH regulator) such as hydrochloric acid and phosphate that can be used as an injection Can be.

The present invention relates to an agent for local fat reduction, which is designed to act in a specific manner of inducing fat cell apoptosis. It has excellent specific effects of lipolysis and apoptosis of adipocytes, without adipocyte necrosis caused by PPC preparations, which can lead to pain, hematoma, edema, numbness, erythema, It is excellent in reducing fat cells without side effects such as swelling, hardening, pruritus and nodules.

1 shows a PPC 5.0% formulation solubilized with a DCA 1.0% formulation and a DCA 2.2%, a single formulation by GCA concentration (GCA 0.5%, GCA 1.0%, GCA 3.0%, GCA 4.5% GCA 6.0%, GCA 7.5%, A comparison of adipocyte viability against GCA 9.0%) is shown.
Figure 2 shows apoptosis (apoptosis) for DCA 1.0% and PPC 5.0% solubilized with DCA 2.2% and GCA concentration alone (GCA 1.0%, GCA 3.0%, GCA 6.0%, GCA 9.0%). ) Induced specificity is shown through the Caspase 3 activity assay.
FIG. 3 shows lipolysis of DCA 1.0% alone and PPC 5.0% solubilized with DCA 2.2%, and GCA concentration alone (GCA 1.0%, GCA 3.0%, GCA 6.0%, GCA 9.0%). Induced specificity is shown through the comparison of Glycerol release.
Figure 4 shows the degree of inflammation induction for the PPC 5.0% formulation solubilized with DCA 1.0% alone and DCA 2.2% and GCA concentration alone (GCA 1.0%, GCA 3.0%, GCA 6.0%, GCA 9.0%). The result measured by ELISA method is shown.
FIG. 5 compares fibroblast viability against a DCA 1.0% formulation and a PPC 5.0% formulation solubilized with DCA 2.2% and a GCA concentration alone formulation (GCA 1.0%, GCA 3.0%, GCA 6.0%, GCA 9.0%). One result is shown.
FIG. 6 compares muscle cell viability for DCA 1.0% alone and PPC 5.0% solubilized with DCA 2.2% and GCA concentration alone (GCA 1.0%, GCA 3.0%, GCA 6.0%, GCA 9.0%). Results are shown.
Figure 7 shows the vascular endothelial cell viability of DCA 1.0% alone and PPC 5.0% solubilized with DCA 2.2% and GCA concentration alone (GCA 1.0%, GCA 3.0%, GCA 6.0%, GCA 9.0%). The comparison result is shown.
Figure 8 shows the result of comparing the degree of edema over time by measuring the plantar thickness by injecting bile acid at each concentration in the rat foot. Figure 8a is a DCA alone composition injection (1.0-7.5%) by concentration, Figure 8b is a CA-only composition injection (1.0-7.5%) by concentration, Figure 8c is a TCA alone composition injection (1.0-7.5%) by concentration, 8d shows the concentration of CDCA alone composition injection (1.0-7.5%) by concentration, FIG. 8e shows the concentration of UDCA alone composition injection (1.0-7.5%) by concentration, 8f shows the concentration of GDCA alone composition injection (1.0-7.5%) by concentration, 8g shows the concentration of TDCA alone injection (1.0-7.5%), Figure 8h shows the concentration of HDCA alone injection (1.0-7.5%), Figure 8i shows the concentration of TUDCA alone injection (1.0-7.5%), Figure 8j DHCA alone composition injection by concentration (1.0 ~ 7.5%), Figure 8k is a graph comparing the injections containing 2.5% each of several types of bile acids (BA) alone to the rat foot, measuring the edema after 3 hours (Use PBS as a control).
9 is injected into the rat paws by injection of DCA 1.0% alone and PPC 5.0% solubilized with DCA 2.2% and GCA concentration alone (GCA 1.0%, GCA 3.0%, GCA 6.0%, GCA 9.0%), Images comparing the extent of tissue lesions over time are shown (three replicates).
FIG. 10 shows DCA 1.0% (FIG. 10A), GCA 1.0% (FIG. 10B), GCA 2.5% (FIG. 10C), GCA 5.0% (FIG. 10D), and GCA 10.0% (FIG. 10E) formulations in rat paws, respectively. 3 hours after injection, the plantar tissue was H & E stained The results of histological comparison of the states of inflammation and edema are shown. 10F shows the results of comprehensive comparison of each experimental group (three replicate experiments).
Figure 11 is administered once each test substance (DCA alone preparations and GCA alone preparations) to a high fat diet-induced obesity fat pad pad, and after 8 days H & E staining subcutaneous fat tissue to reduce the degree of fat cells The histological comparison is shown.
Figure 12 shows the results of comparing the adipocyte viability for DHCA, TUDCA, GCA, TCA by concentrations of edema and inflammation and tissue lesions or low bile salt in the test drug.

Hereinafter, the present invention will be described in detail.

However, the following examples are merely to illustrate the invention, but the content of the present invention is not limited to the following examples.

The composition (Preparation Example) and the comparative compositions (Comparative Example) of the present invention were prepared as follows.

< Production Example > GCA  Sole formulation

Glycocholine acid (GCA) alone preparations were 5 mg (0.5%, Comparative Preparation Example), 10 mg (1%, Preparation Example 1), 15 mg (1.5%, Preparation Example 2), 30 mg (3.0%), respectively, in 1 mL glycocholine acid. , Preparation Example 3), 45mg (4.5%, Preparation Example 4), 60mg (6.0%, Preparation Example 5), 75mg (7.5%, Preparation Example 6) or 90mg (9.0%, Preparation Example 7) The experimental group was prepared, and the remaining material composition was prepared by using 9 mg of benzyl alcohol, 2.2 mg of sodium chloride, and remaining water. Specific manufacturing process is as follows. 1 liter of water for injection (Korea Pharmaceutical Industry Co., Ltd.) was transferred to a sterilized preparation tank, and 500 mL of the water was transferred to a sealed SUS reservoir. Sodium chloride and benzyl alcohol were added and stirred, and after stirring, the remaining water for injection was added to adjust the volume to 1 liter, and hydrochloric acid and sodium hydroxide were added thereto to adjust the pH to 7.2. The agitation process was agitated for 3 hours at a stirring speed of 200 RPM in a condition of light shielding, airtight, 25 degrees Celsius, and the final mixture was filtered through a 0.2 μm filter and then filled and sealed in a vial. Substances used in the preparation include glycocholic acid (New zealand pharm) or sodium glycochocholate (New zealand pharm), benzyl alcohol (Sigma-Aldrich), sodium chloride (Sigma-Aldrich), sodium hydroxide (Sigma-Aldrich). Aldrich) and hydrochloric acid (Sigma-Aldrich).

< Comparative example > Bile acid ( BA A) sole formulation

The bile salt alone formulation was prepared in 10 mL (1%), 25 mg (2.5%), 50 mg (5.0%), 75 mg (7.5%) of each of the bile salts in 1 mL to prepare a test group, the remaining composition of the benzyl alcohol 9 mg, sodium chloride 2.2 mg, the remaining amount of water was prepared as a composition. Specific manufacturing process is as follows. 1 liter of water for injection (Korea Pharmaceutical Industry Co., Ltd.) was transferred to a sterile preparation tank, and 500 mL was transferred to a sealed SUS reservoir, and sodium hydroxide solution was added and stirred to 500 mL of water for injection, and each bile salt was added and stirred. , Sodium chloride and benzyl alcohol were added and stirred. After stirring, the remaining water for injection was added to adjust the volume to 1 liter, and hydrochloric acid and sodium hydroxide were added to adjust the pH to 8.4. The agitation process was agitated for 3 hours at a stirring speed of 200 RPM in a condition of light shielding, airtight, and 25 degrees Celsius, and the final mixture was filled and sealed in a vial after filtration with a 0.2 μm filter. Substances used in the preparation include glycocholic acid (CA, New zealand pharm), deoxycholic acid (DCA, Sigma-Aldrich), sodium deoxycholate (DCNa, New zealand phram), glyco Glycocholic acid (GCA, New zealand phram), Sodium Glycocholate (GCNa, New zealand phram), Taurocholic acid (TCA, Sigma-Aldrich), Sodium taurocholine acid (Sodium Taurocholate, TCNa, New zealand phram), Kenodeoxycholic acid (CDCA, Sigma-Aldrich), Ursodeoxycholic acid (UDCA, Sigma-Aldrich), Glycodeoxycholic acid , GDCA, Sigma-Aldrich), Taurodeoxycholic acid (TDCA, Sigma-Aldrich), Hideoxycholic acid (HDCA, Sigma-Aldrich), Lithocholic acid (Lithocholic acid, LCA, Sigma-Aldrich), Dehydrocholic acid (DHCA, Sigma-Aldrich), Taurusode Soxicol Taurursodeoxycholic acid, TUDCA, Tokyo Chemical Industry, Benzyl Alcohol (Sigma-Aldrich), Sodium Chloride (Sigma-Aldrich), Sodium Hydroxide (Sigma-Aldrich), Hydrochloric Acid (Sigma-Aldrich) to be.

< Experimental Example  1>

GCA  Sole formulation in vitro  Adipocyte Lysis Effect, Death and Degradation

< Experimental Example  1-1> About fat cells Apoptosis ( apoptosis ) Causing effect and lipolysis ( lipolysis ) Effect comparison

One) MTT  assay

Through the MTT assay, the adipocyte death effect of the preparation composition and the comparative composition of the present invention was compared. GCA 0.5%, GCA 1.0%, GCA 1.5%, GCA 3.0%, GCA 4.5%, GCA 6.0%, GCA 7.5%, GCA 9.0% and the like were used as the GCA alone preparation of the present invention, and DCA 1.0% alone and Experiments were performed with 5.0% PPC solubilized with 2.2% DCA (see FIG. 1). In addition, TUDCA 2.5%, TUDCA 5.0%, TUDCA 7.5%, TCA 2.5%, TCA 5.0%, TCA 7.5% was compared in the same way (see Fig. 12). Specific experimental methods are as follows: 3T3-L1 adipocytes were cultured at 70-75% cell confluence. The cells were treated with 100 μml of the experimental material of Preparation Example and Comparative Example, and incubated at 37 ° C. for 0 to 48 hours. The cells were washed twice with PBS, treated with MTT reagent (50 ul), and left at 37 ° C. for 2 hours. After removing the supernatant, the MTT formazan crystals were dissolved by treatment with DMSO, and the absorbance was measured at 540 nm using a microplate reader.

As shown in FIG. 1 , adipocyte lysis was observed in GCA alone, DCA alone, and PPC solubilized with DCA except for 0.5% GCA. But TUDCA TCA group and in 12 were found to not affect the dissolved fat cells.

2) Caspase  3 activity assay

In order to confirm whether cell death in the MTT assay is caused by necrosis or apoptosis, a Caspase 3 activity assay was performed on the preparation composition and the comparative composition of the present invention. Caspase 3 specifically increases when apoptosis occurs. Among the GCA preparations of the present invention, 'GCA 1.0%, GCA 3%, GCA 6.0%, GCA 9.0%' and PPC 5.0% solubilized with control DCA 1.0% and DCA 2.2% were representatively tested. Using Abcam's Caspase 3 Assay Kit (Colorimetric), the experiment was conducted according to the manufacturer's manual. The test method was briefly as follows: Dispense 1X10 ⁵ 3T3-L1 adipocytes into each well and test materials of preparation and comparative examples. 100 μml of each was treated, and incubated for 0 to 48 hours at 37 degrees Celsius. Thereafter, 50 ul of cell lysis buffer (10 mm Tris-HCl, 10 mm NaH 2 PO 4 / NaHPO 4, pH 7.5, 130 mm NaCl, 1% Triton X-100, and 10 mm sodium pyrophosphate) was treated and left at 4 ° C. for 10 minutes. After centrifugation at 1000 X rpm for 1 minute to collect the supernatant, protein quantification was performed by BCA method. 50 ul of reaction buffer (4 mM HEPES, pH 7.5, 10% glycerol, 2 mm dithiothreitol) and 0.5 ul of 4 mM DEVD-p-NA were added to each sample, followed by reaction at 37 ° C for 1 hour. It was then measured at 405 nm wavelength by spectrofluorometry.

As shown in FIG. 2 , Caspase 3 activity was increased in all groups except DCA alone treatment group, and thus, other drugs except DCA alone induced apoptosis in a concentration-dependent manner. In the DCA-only group, Caspase 3 activity decreased rapidly after 48 hours compared to the GCA-only group, indicating that cell death by DCA alone was caused by necrosis. (FIG. 2), which is consistent with previously reported. These results also suggest that the apoptosis effect in DCA + PPC preparations is due to PPC and not DCA.

3) Lipolysis  assay

Through the Lipolysis assay, the lipolysis effect of the composition of the production example and the comparative example of the present invention was compared. Among the GCA alone preparations of the present invention, 'GCA 1.0%, GCA 3.0%, GCA 6.0%, GCA 9.0%' and control DCA 1.0% alone and PPC 5.0% solubilized with DCA 2.2% were representatively tested. Using Abcam's Lipolysis Assay Kit (Colorimetric), the experiment was conducted according to the manufacturer's manual. The test method was as follows: Dispense 1X10 ⁵ 3T3-L1 adipocytes into each well and prepare the test materials of the preparation and comparative examples. Each 100μml treatment, incubated for 0 ~ 48 hours at 37 degrees Celsius to induce Lipolysis. Lipolysis Assay Buffer (137 mM NaCl, 5 mM KCl, 4.2 mM NaHCO3, 1.3 mM CaCl2, 0.5 mM KH2PO4, 0.5 mM MgCl2, 0.5 mM MgSO4, 5 mM Glucose, 20 mM Hepes, pH 7.4), 1 % BSA, 1 μM Isoproterenol) was added and incubated for 20 minutes to a total of 50 ul. After mixing the Glycerol Assay complex (50 ul), it was left for 30 minutes at room temperature and the absorbance was measured at OD570 (using a standard curve for absolute quantification).

As shown in FIG. 3 , glycerol release was increased more than 2 times in all groups except DCA alone treatment group. Lipolysis was also induced in the DCA alone treatment group, but the lipolysis effect of DCA was very low compared to the DCA 1% and GCA 1% treatment groups. In the GCA treatment group, glycerol release was increased depending on GCA content. These results also suggest that the lipolytic effect seen in DCA + PPC preparations depends on PPC rather than DCA.

< Experimental Example  One- 2> inflammatory response  evaluation

Using the TNF ELISA method, the inflammatory effect of the composition of the above-described preparation of the present invention and the comparative composition was compared. In the GCA alone preparations of the present invention, the experiments were representative of 'GCA 1.0%, GCA 3.0%, GCA 6.0%, GCA 9.0%', and PPC 5.0% solubilized with control DCA 1.0% alone and DCA 2.2%. Using the mouse TNF-alpha Quantikine ELISA Kit of R & D systems, the experiment was performed according to the manufacturer's manual. The test method was briefly as follows: Dispense 1X10 ⁵ 3T3-L1 adipocytes into each well, and experiment with the preparation and comparative examples. Each 100 μml of the material was incubated for 0 to 48 hours and 37 degrees. 30 ul of the cell culture solution was reacted overnight at 4 ° C. with a TNF alpha antibody pre-bound culture plate included in the kit. After washing three times with Washing Buffer and reacted with a secondary antibody (1: 500) at room temperature for 1 hour. After washing three times with washing buffer again, a color reaction (substrate reaction) was performed, and the reaction was quickly measured at OD570 after stopping the reaction (using a standard curve for absolute quantification).

The experimental results are shown in FIG. 4 . In the DCA 1% monotherapy and DCA-solubilized PPC groups, TNF alpha was significantly increased compared to the other experimental groups. Especially, the DCA 1% treatment group had a higher TNF- compared to the GCA 9% treatment group. α secretion was observed.

< Experimental Example 1 -3> fibroblasts, Muscle cells , Vascular endothelial cell lysis effect

1) Fibroblasts (CCD- 986sk ; human fibroblast)

Through MTT assay, the fibroblast viability effect of the preparation composition and the comparative example composition of the present invention was compared. Among the GCA alone preparations of the present invention, 'GCA 1.0%, GCA 3.0%, GCA 6.0%, GCA 9.0%', and control DCA 1.0% alone and DCA 5.0% solubilized with 2.2% DCA were representatively tested. Specific experimental methods are as follows: After culturing fibroblasts (CCD-986sk; human fibroblast) to 85% cell confluence, the culture medium (IMDM, 10% FBS, 1% anitibiotics mix) is removed. The cells were mixed and treated with 100 μml of the experimental material of Preparation Example and Comparative Example, respectively, in a new culture medium, and the culture medium was removed at a predetermined treatment time and subjected to MTT assay (measured at 570 nm).

The test results are shown in FIG. PPC preparations solubilized with DCA, DCA alone, and GCA 9.0% alone had high fibroblast lysis potency, whereas GCA 1% to 6% alone had significantly lower fibroblast solubility to selectively reduce fat cells. When applied, local adverse reactions are drastically lowered.

2) Muscle cells ( C2C12 ; mouse myocyte )

Through the MTT assay, the effect of muscle cell viability of the preparation composition and the comparative composition of the present invention was compared. Among the GCA alone preparations of the present invention, 'GCA 1.0%, GCA 3.0%, GCA 6.0%, GCA 9.0%' and control DCA 1.0% alone and DCA solubilized with 2.2% DCA 5.0% were representatively tested. Specific experimental methods are as follows: After culturing muscle cells (C2C12; mouse myocyte) at 80% cell confluence, remove the culture medium (DMEM, 10% FBS, 1% anitibiotics mix). Incubate for 4 days with DMEM containing 2% horse serum. After confirming the elongated shape of more than 80% of the cells, 100 μml of each of the preparations of the preparation and comparative examples were mixed and treated with a new culture medium, and the culture medium was removed at a predetermined treatment time, followed by MTT assay (measured at 570 nm). To be implemented.

The test results are shown in FIG. 6 . PPC preparations solubilized with DCA, DCA alone, and GCA 9.0% alone showed high muscle cell solubility, whereas GCA 1% to 6% alone showed significantly lower muscle cell solubility, resulting in selective reduction of fat cells. When applied, local adverse reactions are drastically lowered.

3) vascular endothelial cells ( HUVEC ; human endothelium)

Through the MTT assay, the effect of muscle cell viability of the preparation composition and the comparative composition of the present invention was compared. Among the GCA alone preparations of the present invention, 'GCA 1.0%, GCA 3.0%, GCA 6.0%, GCA 9.0%', and control DCA 1.0% alone and PPC 5.0% solubilized with DCA 2.2% were representatively tested. Specific test methods are as follows: 1% gelatin is applied to the culture dish and left to stand at room temperature for 1 day. After culturing up to 80% of the cell fullness in the coated culture dish remove the culture medium (EGM-Plus, 10% FBS, 1% antibiotics mix). The cells were mixed and treated with 100 μml of the experimental material of Preparation Example and Comparative Example, respectively, in a new culture medium, and the culture medium was removed at a predetermined treatment time and subjected to MTT assay (measured at 570 nm).

The test results shown in FIG. PPC preparations solubilized with DCA, DCA alone, and GCA 9.0% alone had high muscle cell solubility, whereas GCA 1% to 6% alone had significantly lower vascular endothelial cell solubility, resulting in selective reduction of adipocytes. Local adverse reactions are drastically lowered in human application.

< Experimental Example  2>

GCA  Of single agent in vivo   Local reduction  Effectiveness and side effects control assessment

< Experimental Example  2- 1> general  Evaluation of adverse events (inflammation and edema) in mice

1) Sole thickness measurement

Six-week-old males (weight: 170-200 g) of Sprague Dolly (rat), 0.1 ml of each test drug of 1.0%, 2.5%, 5.0%, and 7.5% of bile acids and DCA 1.0% alone as a control. Subcutaneous injection into the soles of the hind feet. After the drug injection, the thickness of the sole was measured by calipers every 1, 2 and 3 hours, and the measurer was carried out in an unknown state (blind test). Baldabak thickness measurement results with time for each experimental group is shown in Figure 8a to 8k . In addition, in order to observe the tissue lesion of the administration site when measuring the thickness of the plantar photograph (using a 4 × 4 cm scale) was taken, which is shown in FIG.

As seen in FIGS . 8A- 8K , All of the bile acids except TUDCA and DHCA showed edema in concentration-dependent edema, and bile acids with lower edema level than DCA 1% and concentrations were 1.0% to 2.5% CA, 1.0% to 5.0% TCA, 1.0% to 6.0% GCA. , UDCA 1.0%, GDCA 1.0%, TDCA 1.0%, HDCA 1.0%. Tissue lesions at the site of administration were moderately to severe except for TUDCA and DHCA, with no or mild tissue lesions at concentrations below TCA 5.0% and GCA 6.0%. In addition , as shown in FIG. 9 , in the rats' soles, the side effects of severe erythema and inflammation were visually confirmed in the group administered with PPC 5.0% formulation solubilized with DCA 1.0% alone and DCA 2.2%. Hardly any tissue lesion was observed with the naked eye.

2) H & E staining histological examination

6-week-old male (weight: 170-200 g) of Sprague Dolly (rat) with 0.1 ml of GCA 1.0%, GCA 2.5%, GCA 5.0%, GCA 10.0% alone and control DCA 1.0% alone Subcutaneous injection into the soles of the hind feet. After 3 hours of administration, the soles were cut and the skins of the soles were cut out and fixed with 10% formalin aqueous solution. It was stained with hematoxylin & eosin solution to observe morphological changes. The observation results are shown in Figs. 10A, 10B, 10C, 10D, 10E and 10F .

As comprehensively compared in FIG. 10F , inflammatory cells were significantly reduced in the GCA 5.0% or less administration group of the present invention compared to the DCA administration group. Through the above experimental results and the present experimental results, the side effects due to adipocyte necrosis (necrosis) occurs in the existing DCA-use preparations, whereas the GCA 6% or less of the present invention has an adipocyte-specific apoptosis-specific inflammatory response It was confirmed that fat was reduced almost without this.

< Experimental Example  2-2> On high paper  Fat Reduction Effect and Inflammation Assessment in an Induced Mouse Obesity Model

C57BL / 6 rats (4 weeks old) were fed a high-fat diet (Research diet, 60% kcal lipid) for 12 weeks, resulting in high obesity. The high fat diet-induced obese mouse inguinal pads were treated with saline (negative control), DCA alone (DCA 1%, positive control) or GCA 1.0%, GCA 2.5%, GCA 5.0%, GCA 10.0%. Administration was once. Each test substance was performed in a subcutaneous injection of 0.2 mL into the abdomen. Eight days after the first infusion was completed, mice were sacrificed. The abdomen of the sacrificed mice was excised and subcutaneous fat was quickly removed and soaked in 4% formaldehyde solution and fixed. After fixation and washing with water and dehydration, the solution was treated with a paraffin solution to form a paraffin block, cut into 4μm thicknesses, stained with hematoxylin and eosin and observed under an optical microscope. . The results are shown in Fig.

As shown in Figure 11 , in the DCA alone (DCA 1%) administration group, granulocytes gathered together with traces of adipocyte necrosis, and an inflammatory response appeared. On the other hand, in the GCA 6% or less administration group of the present invention, the inflammatory response was less than that of the control group, and fat cells were killed.

As described above, the present invention relates to a phosphatidylcholine-free injectable composition for topical fat reduction and a method for preparing the same, which has no pain and no side effects, and more particularly, to a phosphatidylcholine-containing topical fat injectable composition. It relates to a topical fat reducing injection composition and a method for producing the same, characterized in that it comprises 1 (w / v)% to 6 (w / v)% of glycocholine acid or a salt thereof.

Fat loss caused by adipocyte necrosis in conventional deoxycholine acid (DCA) single agents or phosphatidylcholine-deoxycholine acid combinations known as topical topical fat injections can cause pain, edema, inflammation, fibrosis, erythema, Local adverse reactions such as hematoma and nerve injury have been reported. However, the injection composition for topical lipolysis provided by the present invention induces lipolysis and fat cell death (apoptosis) to reduce local fat, and the fat cell removal effect is remarkable. Side effects such as pain, swelling, inflammation, fibrosis, erythema, hematoma, and nerve damage in the injection composition are remarkably alleviated, and it is highly industrially available because it dramatically improves patients' low compliance with the pain and swelling during and after administration. .

Claims (14)

In the composition for injection of topical fat containing no phosphatidylcholine,
The composition is a topical fat reduction injection composition without pain and side effects, characterized in that containing 1 (w / v) to 6 (w / v)% glycocholic acid or salts thereof.
According to claim 1, wherein the topical fat reduction is a topical fat reduction injection composition without pain and side effects, characterized in that by the method of lipolysis (lipolysis) and apoptosis of fat cells (apoptosis).
The method of claim 1, wherein the side effects are any one or more selected from the group consisting of hematoma, edema, numbness, erythema, swelling, sclerosis, pruritus and nodules, the injection composition for topical fat reduction without pain and side effects.
According to claim 1, wherein the composition has a pH 6.0 to pH 8.0 injection composition for topical fat reduction free from pain and side effects.
According to claim 1, wherein the salt is a sodium salt, potassium salt or ammonium salt, injection composition for topical fat reduction without pain and side effects, characterized in that
The composition of claim 1, wherein the composition further comprises benzyl alcohol.
The group of claim 1, wherein the topical fat consists of fat accumulation associated with a mammal's abdomen, under eyes, under jaw, under arm, hip, thigh, calf, back, thigh, braline and lipoma. Injectable composition for reducing local fat without pain and side effects, characterized in that selected from.
In the method for producing a topical fat reducing injection composition without pain and side effects,
The composition comprises 1 (w / v)% to 6 (w / v)% of glycocholic acid (GCA) or salts thereof, but does not include phosphatidylcholine, and has no pain and no side effects. A method of making a reducing injectable composition.
The method of claim 8, wherein the reduction of topical fat is performed by a method of lipolysis and apoptosis of fat cells.
10. The method of claim 8, wherein the side effect is any one or more selected from the group consisting of edema, swelling, nerve damage, nodules, sclerosis, numbness, erythema and pruritus How to manufacture.
10. The method of claim 8, wherein the composition has a pH of 6.0 to 8.0.
The method of claim 8, wherein the salt is sodium, potassium, or ammonium salt.
10. The method of claim 8, wherein the composition further comprises benzyl alcohol.
9. The group of claim 8, wherein the topical fat comprises a accumulation of fat associated with a mammal's abdomen, under eyes, under jaw, under arm, hip, thigh, calf, back, thigh, braline and lipoma. Method for producing an injection composition for reducing fat without pain and side effects, characterized in that selected from.

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