WO2021137656A1 - Novel formulation for local injection - Google Patents

Abstract

The present invention relates to a drug delivery composition for local injection and a method of preparing same. By using a polyacrylate polymer, a polymethacrylate polymer, or a copolymer thereof, dissolved in an organic solvent, and more specifically, a composite polymer containing an Eudragit E grade and an Eudragit RS or RL grade mixed in a uniform ratio, the present invention efficiently achieves all of the features required in local injectable formulations, the features being: the ability to maintain a solution state pre-injection; a rapid sol-gel transition immediately after injection; and a high biodegradation rate of the gel thus formed. The present invention is applicable to sustained-release injection formulations capable of sustaining a drug effect for several days to several months by being administered by a subcutaneous or intramuscular route, and can be advantageously used as an excellent intratumoral injection formulation that concentrates pharmacological effects, reduces side effects, and greatly improves patient administration convenience by localizing anticancer drugs, which have been systemically administered through intravenous injection, specifically to a tumor site.

Description

Novel topical injectable formulations

The present invention relates to a formulation for local injection comprising a polyacrylate polymer, a polymethacrylate polymer, or a copolymer thereof, for example, a composite polymer in which EUDRAGIT E grade and EUDRAGIT RS or RL grade are mixed in a constant content ratio. It relates to a formulation for intratumoral injection comprising.

Long-acting parenteral formulations last from one week to several months with a single administration, and are advantageous in terms of patient convenience. Technologies that enable long-acting parenterals include microspheres, liposomes, prodrugs and in situ gels (Aijaz A. Sheikh 2016).

Anticancer drugs are generally circulated throughout the body through intravenous injection, but are accompanied by various side effects such as hair loss, nausea, vomiting, skin rash, and decreased bone marrow function. In addition, some anticancer drugs are rapidly eliminated from the blood, so there are cases where a therapeutically effective amount cannot be delivered to the tumor lesion. Therefore, targeted drug delivery systems and local drug delivery systems that are specifically delivered to cancer cells were studied. In topical treatment, intratumoral (IT) injection is a method of injecting anticancer drugs directly into solid tumors (Zheng, Gao et al. 2017). This has the advantage of avoiding systemic side effects and delivering pharmacological components directly to the tumor to increase treatment efficiency, and can be used to reduce the size of a tumor before surgical resection, and it can also be used in areas where surgery is impossible. It can also be used for anti-cancer treatment. Some anticancer drugs have very low water solubility, which can be overcome by intratumoral injection because systemic circulation through veins is difficult in this case. Intratumoral injection can also reduce dosage and frequency of administration (Fakhari and Anand Subramony 2015).

5-FU is an anticancer agent that has been used for over 40 years and has been used for solid cancers such as colorectal cancer, breast cancer and liver cancer (Lee, Beumer et al. 2016), but when administered systemically through intravenous injection, nausea, vomiting, hair loss, bone marrow Various side effects, including inhibition and hand-foot syndrome, appear (Nurgali, Jagoe et al. 2018). When 5-FU is converted to a specific nucleotide, an anticancer effect appears. Thymidine phosphorylase (TP) is an enzyme that converts 5-FU to fluorodioxyuridine, which in turn is converted to its active metabolite, fluorodioxyuridine monophosphate. The mechanism of action of 5-FU is inhibition of thymidylate synthase (TS), a rate limiting enzyme in pyrimidine nucleotide synthesis (Longley, Harkin et al. 2003). Deoxyuridine monophosphate is converted via TS to deoxythymidine monophosphate (dTMP). Inhibition of TS ultimately leads to inhibition of DNA synthesis and repair.

An in situ gel drug delivery system was applied to the formulation design of intratumoral injections. Injection of a solution in which a drug is dissolved is easily released out of the tumor tissue and the drug is not delivered well. Therefore, for direct injection into the tumor tissue, the formulation must form a reservoir and general injection of low viscosity must be used. A suitable system for this is an in situ gel drug delivery system. In situ gel refers to a drug delivery system that forms a gel in situ when applied to the site of administration (Sarada K 2014). When applied to the site of administration, it becomes a gel by photopolymerization, chemical crosslinking, physical crosslinking, pH-sensitivity, temperature sensitivity and solvent exchange (Kempe and Mader 2012). The formulation is designed using solvent exchange technology, which can be prepared simply by mixing an organic solvent, a polymer and a drug, and this simple process is one of the advantages. Upon injection, the organic solvent diffuses out of the injection site and body fluid flows into the formulation (Thakur, McMillan et al. 2014). As a result of this solvent exchange, polymer precipitation occurs at the injection site, and the injected drug is captured by this polymer matrix and released when the polymer matrix is decomposed. The most important thing in the solvent exchange-based situ gel is which polymer is used (Parent, Nouvel et al. 2013), which should be soluble in a specific organic solvent and insoluble in water, as well as biocompatible and biodegradable.

Numerous papers and patent documents are referenced throughout this specification and their citations are indicated. The disclosure contents of the cited papers and patent documents are incorporated herein by reference in their entirety to more clearly describe the level of the technical field to which the present invention pertains and the content of the present invention.

The present inventors have made intensive research efforts to develop an effective formulation for local injection that can more concentrate the pharmacological effect and reduce side effects by locally acting the drug specifically on the lesion when it is administered systemically through intravenous injection. As a result, when a drug delivery vehicle is prepared using the polymer of Formula 1 dissolved in an organic solvent, it exists in a solution of low viscosity that can be injected, and then rapidly sol-gel immediately after injection into the lesion site. transition) occurs, so that the release rate of the drug is appropriately controlled, and the formed gel exhibits excellent biodegradability that almost all of it disappears in the body within about 1 month, so it can be used as an efficient drug delivery system for local injection. By discovering that , the present invention has been completed.

Accordingly, an object of the present invention is to provide a drug delivery composition for local injection.

Another object of the present invention is to provide a composition for preventing or treating solid cancer.

Another object of the present invention is to provide a method for preparing a drug-loaded formulation for local injection.

Other objects and advantages of the present invention will become more apparent from the following detailed description of the invention, claims and drawings.

According to one aspect of the present invention, there is provided a drug delivery composition for local injection comprising (a) an organic solvent and (b) a polymer represented by the following formula (1) dissolved in the organic solvent:

Formula 1

In Formula 1, R ₁ to R ₃ are each independently hydrogen or C ₁ to C ₃ alkyl, R ₄ is or N(R ₅ ) ₂ or N(R ₅ ) ₃ ⁺ , and R ₅ is C ₁ to C ₃ alkyl, and n is an integer from 100 to 3,000.

The present inventors have made intensive research efforts to develop an effective formulation for local injection that can more concentrate the pharmacological effect and reduce side effects by locally acting the drug specifically on the lesion when it is administered systemically through intravenous injection. As a result, when a drug delivery vehicle is prepared using the polymer of Formula 1 dissolved in an organic solvent, it exists in a solution of low viscosity that can be injected, and then rapidly sol-gel immediately after injection into the lesion site. transition) occurs so that the release rate of the drug is appropriately controlled, and the formed gel exhibits excellent biodegradability that almost all of it disappears in vivo within about 1 month, making it an efficient local injection drug delivery system. found that it can be used.

As used herein, the term “topical injection” is a concept relative to systemic administration in which a pharmacological component affects the whole body through the circulatory system, and a subcutaneous injection or intramuscular injection in which the pharmacological component is applied only to a part of the body of a subject. , including ventricular injection, intratumoral injection, and all injection types except systemic administration.

As used herein, the term “polymer” refers to a synthetic or natural high molecular compound in which the same or different types of monomers are continuously combined. Thus, polymers include homopolymers (polymers in which one type of monomer is polymerized) and interpolymers prepared by the polymerization of at least two different monomers, and interpolymers include copolymers (polymers prepared from two different monomers). polymers) and polymers prepared from more than two different monomers.

As used herein, the term “alkyl” refers to a straight-chain or pulverized saturated hydrocarbon group, C ₁ -C ₃ alkyl refers to an alkyl group having an alkyl unit having 1 to 3 carbon atoms, and when C ₁ -C ₃ alkyl is substituted, a substituent carbon number is not included.

According to a specific embodiment of the present invention, R ₁ to R ₃ in Formula 1 are each independently hydrogen or C ₁ alkyl, and R ₅ is C ₁ alkyl.

More specifically, R ₁ and R ₂ are each independently hydrogen or C ₁ alkyl, R ₃ is C ₁ or C ₂ alkyl, and R ₅ is C ₁ alkyl.

As used herein, the term “organic solvent” refers to an organic molecule capable of at least partially dissolving other molecules (solute) and existing in a liquid state at room temperature. Examples of the organic solvent include hydrocarbon solvents such as n-heptane, n-hexane, n-heptane, n-octane, paraffin, cyclohexane and methylcyclohexane; aromatic organic solvents such as benzene, toluene, o-xylene, m-xylene, and p-xylene; alcohol organic solvents such as methanol, ethanol, 1-propanol, 1-butanol, 1-octanol, benzyl alcohol, phenol, ethylene glycol and m-cresol; Various organic solvents known in the art including, but not limited to, nitrogenous organic solvents such as N,N,-dimethylformamide, N,N,-dimethylacetamide, acetonitrile, and N-methyl-2-pyrrolidone Solvents may be used. Specifically, the organic solvent used in the present invention is at least one solvent selected from the group consisting of ethanol, methanol, and N-methyl pyrrolidone (NMP), and more specifically, NMP.

According to a specific embodiment of the present invention, the polymer is included in the composition in an amount of 25 to 35 w/w% based on the total composition. More specifically, it is included in 27 to 33 w/w%, and most in volume it is included in 29 to 31 w/w%.

According to a specific embodiment of the present invention, the polymer represented by Formula 1 is (i) R ₁ to R ₃ are C ₁ alkyl, R ₄ is —N(R ₅ ) ₂ , and R ₅ is C ₁ alkyl. a first polymer; (ii) R ₁ and R ₂ is a C ₁ alkyl, R ₃ is a C ₂ alkyl, R ₄ is ^{_{-N (R 5) 3 +,}} R 5 is a mixture of the second polymer is C ₁ alkyl.

More specifically, in the composition, the first polymer and the second polymer are present in a content ratio of 4:6 to 6:4.

The first polymer in which R ₁ to R ₃ is C ₁ alkyl(methyl) and R ₄ is —N(CH ₃ ) ₂ is a polyacrylate polymer of EUDRAGIT E grade, R ₁ and R ₂ are C ₁ alkyl(methyl) and R ₄ is —N(CH ₃ ) ₃ ⁺ , the second polymer is a polyacrylate polymer of EUDRAGIT RS grade. According to the present invention, in the local injection drug delivery composition of the present invention, the polymer of Formula 1 is contained in an amount of about 30 w/w% in the total composition, and the polymer is mixed with EUDRAGIT E grade and EUDRAGIT RS grade at about 1:1. The formulation in Table 1 below is formulation No. 8, and it was confirmed that the formulation showed excellent properties in sol-gel transfer, gel biocompatibility and biodegradability, water diffusion, and drug release.

According to a specific embodiment of the present invention, the polymer represented by Formula 1 is (i) R ₁ to R ₃ are C ₁ alkyl, R ₄ is —N(R ₅ ) ₂ , and R ₅ is C ₁ alkyl. a first polymer; (ii) R ₁ is hydrogen, R ₂ and R ₃ are C ₁ alkyl, R ₄ is ^{_{-N (R 5) 3 +,}} R 5 is a mixture of a third polymer of C ₁ alkyl.

More specifically, in the composition, the first polymer and the third polymer are present in a content ratio of 4:6 to 6:4.

The third polymer in which R ₁ is hydrogen, R ₂ and R ₃ are C ₁ alkyl(methyl) and R ₄ is —N(CH ₃ ) ₃ ⁺ is a polyacrylate polymer of EUDRAGIT RL grade. The formulation in which the polymer of Formula 1 is contained in an amount of about 30w/w% in the total composition, and EUDRAGIT E grade and EUDRAGIT RL grade are mixed in about 1:1 to the polymer is the formulation No. 9 in Table 1 below, No. 9 The formulation also shows excellent properties in terms of sol-gel transition, gel biocompatibility and biodegradability, water diffusion and drug release.

As used herein, the term “biocompatibility” refers to a property that does not cause short-term or long-term side effects when administered in vivo and in contact with cells, tissues or body fluids of organs, specifically, causes tissue necrosis by contact with living tissues or blood. This includes not only tissue compatibility and blood compatibility that does not coagulate blood, but also biodegradability, which disappears after a certain period of time after administration in vivo.

As used herein, the term “biodegradability” refers to the property of being naturally decomposed when exposed to a physiological solution of pH 6-8, specifically, the lapse of time by body fluids, degrading enzymes, microorganisms, etc. in vivo. It means a property that can be decomposed according to The polymer of the present invention showed excellent biodegradability, degrading up to about 80% of the initial injection weight 4 weeks after injection in vivo , which is a very important property as a formulation component regardless of whether EUDRAGIT is non-toxic.

According to another aspect of the present invention, the present invention provides a composition for preventing or treating solid cancer comprising the above-described local injection drug delivery composition and the anticancer composition for solid cancer as an active ingredient.

When the pharmacological component loaded by the drug delivery system of the present invention is an anticancer composition for solid cancer, "local injection" of the present invention becomes "intratumoral injection". Intratumoral injection has a disadvantage in that the target carcinoma is limited in that it is an invasive dosage form. However, by injecting an anticancer drug directly into a solid cancer, the systemic side effects of intravenous injection can be avoided as well as impeding drug arrival due to an immune response in the blood. There is an advantage that direct contact between the cancer tissue and the drug is possible without it.

As used herein, the term “prevention” refers to inhibiting the occurrence of a disease or disease in a subject who has never been diagnosed with a disease or disease, but is likely to have the disease or disease. When the present invention is applied to a benign tumor tissue or a tissue in which whether it is malignant or benign, the composition of the present invention may be expressed as “a composition for preventing cancer”.

As used herein, the term “treatment” refers to (a) inhibiting the development of a disease, disorder or condition; (b) alleviation of the disease, condition or condition; or (c) eliminating the disease, condition or symptom. When the composition of the present invention is injected intratumorally into a solid cancer site of a subject, it serves to inhibit the growth and proliferation of a tumor by direct delivery of an anticancer component, or to eliminate or alleviate symptoms caused by a tumor. Accordingly, the composition of the present invention may be a cancer treatment composition by itself, or may be administered together with other pharmacological ingredients to be applied as a therapeutic adjuvant for cancer. Accordingly, as used herein, the term “treatment” or “therapeutic agent” includes the meaning of “therapeutic adjuvant” or “therapeutic adjuvant”.

As used herein, the term “administration” or “administering” refers to directly administering a therapeutically effective amount of the composition of the present invention to a subject so that the same amount is formed in the subject's body.

In the present invention, the term “therapeutically effective amount” refers to the content of the composition in which the pharmacological component in the composition is sufficient to provide a therapeutic or prophylactic effect to an individual to whom the pharmaceutical composition of the present invention is to be administered. prophylactically effective amount”.

As used herein, the term “subject” includes, without limitation, humans, mice, rats, guinea pigs, dogs, cats, horses, cattle, pigs, monkeys, chimpanzees, baboons or rhesus monkeys. Specifically, the subject of the present invention is a human.

According to a specific embodiment of the present invention, the solid cancer that can be prevented or treated with the composition of the present invention is colorectal cancer, esophageal cancer, stomach cancer, pancreatic cancer, breast cancer, cervical cancer, ovarian cancer, lung cancer, malignancy, tongue cancer and liver cancer from the group consisting of is chosen More specifically, the solid cancer is colorectal cancer.

According to a specific embodiment of the present invention, the anti-cancer composition is 5-FU (5-fluorouracil) or a pharmaceutically acceptable salt thereof.

As used herein, the term “pharmaceutically acceptable salt” includes salts derived from pharmaceutically acceptable inorganic acids, organic acids, or bases. Examples of suitable acids include hydrochloric acid, hydrobromic acid, sulfuric acid, nitric acid, perchloric acid, fumaric acid, maleic acid, phosphoric acid, glycolic acid, lactic acid, salicylic acid, succinic acid, toluene-p-sulfonic acid, tartaric acid, acetic acid, trifluoroacetic acid, citric acid, methane sulfonic acid, formic acid, benzoic acid, malonic acid, naphthalene-2-sulfonic acid, benzenesulfonic acid, and the like. Salts derived from suitable bases may include alkali metals such as sodium, alkaline earth metals such as magnesium, and ammonium and the like.

The composition of the present invention is a pharmaceutical composition, and includes a pharmaceutically acceptable carrier. Pharmaceutically acceptable carriers included in the pharmaceutical composition of the present invention are commonly used in formulation, and include lactose, dextrose, sucrose, sorbitol, mannitol, starch, acacia gum, calcium phosphate, alginate, gelatin, calcium silicate, microcrystalline cellulose, polyvinylpyrrolidone, cellulose, water, methyl cellulose, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil, and the like. no. The pharmaceutical composition of the present invention may further include a lubricant, a wetting agent, an emulsifier, a suspending agent, a preservative, and the like, in addition to the above components. Suitable pharmaceutically acceptable carriers and agents are described in detail in Remington's Pharmaceutical Sciences (19th ed., 1995).

Since the pharmaceutical composition of the present invention is administered intratumorally, in consideration of factors such as the administration mode, the nature and volume of the target tumor, the patient's age, weight, sex, pathological condition, food, administration time, excretion rate, and response sensitivity Dosage can be determined. The dosage of the pharmaceutical composition of the present invention is, for example, in the range of 0.001-100 mg/kg on an adult basis.

The pharmaceutical composition of the present invention is prepared in unit dosage form by formulating using a pharmaceutically acceptable carrier and/or excipient according to a method that can be easily carried out by a person of ordinary skill in the art to which the present invention pertains. or may be prepared by incorporation into a multi-dose container. In this case, the formulation may additionally include a dispersing agent or a stabilizing agent.

According to another aspect of the present invention, the present invention provides for the prevention of solid cancer comprising administering to a subject a pharmaceutical composition comprising the above-described local injection drug delivery composition and anticancer composition for solid cancer as an active ingredient or a method of treatment.

According to another aspect of the present invention, the present invention provides a method for preparing a drug-loaded formulation for local injection, comprising the steps of:

(a) adding a polymer represented by Formula 1 to an organic solvent:

Formula 1

In Formula 1, R ₁ to R ₃ are each independently hydrogen or C ₁ to C ₃ alkyl, R ₄ is or N(R ₅ ) ₂ or N(R ₅ ) ₃ ⁺ , and R ₅ is C ₁ to C ₃ alkyl, n is an integer from 100 to 3,000;

(b) stirring until the organic solvent to which the polymer represented by Formula 1 is added becomes transparent;

(c) adding the drug to the stirred mixture.

Since the organic solvent and the polymer of Formula 1 used in the present invention have already been described above, description thereof will be omitted to avoid excessive overlap.

The features and advantages of the present invention are summarized as follows:

(a) The present invention provides a drug delivery composition for local injection and a method for preparing the same.

(b) the present invention is a polyacrylate polymer, a polymethacrylate polymer or a copolymer thereof dissolved in an organic solvent, specifically, EUDRAGIT E grade and EUDRAGIT RS or RL grade by using a composite polymer mixed in a constant content ratio. maintenance of solution prior to injection; rapid sol-gel transition immediately after injection; and a high biodegradation rate of the gel formed, which efficiently achieves all the characteristics required for a formulation for topical injection.

(c) The present invention is applicable to a sustained-release injection formulation that can sustain the drug effect for several days to several months by subcutaneous or intramuscular injection, and has a pharmacological effect by local action of an anticancer agent administered systemically through intravenous injection, specifically to the tumor site. It can be usefully used as an excellent intratumoral injection formulation that concentrates, reduces side effects, and greatly improves patient administration convenience.

1 is a diagram showing the mechanism of the solvent exchange-based situ gel.

Figure 2 is a diagram showing a schematic diagram of a long-acting intratumoral injection.

3 is a diagram showing the structure of EUDRAGIT E, RS, and RL grades.

4 is a diagram showing a schematic schematic diagram of a water diffusion test.

5 is a diagram showing the results of sol-gel transition according to EUDRAGIT grade and concentration.

6 is a diagram showing the sol-gel transition results of formulations having two EUDRAGIT grades of L and S.

7 is a diagram showing the weight reduction rate in in vitro gel decomposition.

8 is a diagram showing the moisture diffusion characteristics of each formulation according to EUDRAGIT grade and concentration.

9 is a diagram showing the release characteristics of 5-FU for 12 hours in an in situ gel.

10 is a diagram showing the release characteristics of 5-FU for 10 days in an in situ gel.

11 is a diagram showing the in vivo sol-gel transition in the designed formulation.

12 is a diagram showing in vivo gel degradation with time.

Hereinafter, the present invention will be described in more detail through examples. These examples are only for illustrating the present invention in more detail, and it will be apparent to those skilled in the art that the scope of the present invention is not limited by these examples according to the gist of the present invention. .

Example

Experimental method

experimental material

5-FU was purchased from Sigma Aldrich (Saint Louis, MO, USA). EUDRAGIT E 100, E PO, RL 100, RL PO, RS 100, RS PO, L100 and S100 are from Evonik industries® (Essen, Germany). NMP (N-methyl pyrrolidone) was purchased from DC Chemical Co., Ltd (Seoul, South Korea). Sodium phosphate dibasic, anhydrous and sodium phosphate monobasic were purchased from Samchun Chemicals (Gyeonggi-do, South Korea). Sodium 1-heptanesulfonate was purchased from Tokyo Chemical Industry Co., Ltd (Tokyo, Japan). Purified water was Milli-Q (Milli-Q Reference, Millipore®, Molsheim, France) quality, and other reagents were of analytical grade.

Preparation of drug-loaded sols

A 20 mL vial was prepared and added to the vial after weighing the NMP. EUDRAGIT polymer was weighed and added to the vial containing NMP. A transparent solution was obtained by uniformly mixing by stirring at room temperature for several hours using a magnetic bar. At this time, the polymer not dissolved during stirring was sonicated for 30 minutes with an ultrasonic grinder and stirred again to obtain a transparent solution. Through this process, the polymer was finally dissolved in an organic solvent, the drug was weighed, added, and stirred for several hours to prepare a transparent final formulation in which the drug particles were visible. The contents of the prepared formulations are summarized in Table 1.

Content of 5-FU long-acting intratumoral injection formulation

number	One	2	3	4	5	6	7	8	9	10	11
NMP	8	8	8	7	7	7	8.5	7	7	8.5	8.5
E 100	2			3			1.5	1.5	1.5
RS 100		2			3			1.5
RL 100			2			3			1.5
L 100										1.5
S 100											1.5
Sum	10	10	10	10	10	10	10	10	10	10	10
5-FU	0.2	0.2	0.2	0.2	0.2	0.2	0.2	0.2	0.2	0.2	0.2
total	10.2	10.2	10.2	10.2	10.2	10.2	10.2	10.2	10.2	10.2	10.2

Evaluation of sol-gel transition

Since the pH inside the tumor is about 7.2 (Zhang, Lin et al. 2010), pH 7.2 PBS was prepared as a medium for sol-gel transfer. Sodium phosphate monobasic and sodium phosphate dibasic were weighed and dissolved in distilled water to prepare a pH 7.2 buffer (pH was measured with a pH meter). 10 mL of pH 7.2 PBS was added to a 20 mL vial using a pipette. The formulation to be used in the above-described sol-gel transfer process was prepared, and a 1 mL syringe and 20 gauge needle were prepared. 0.5 mL of the prepared formulation into a 20 mL vial containing pH 7.2 PBS was injected with a syringe, and gel formation was observed immediately after injection. A solvent exchange reaction was observed around the injected solution, confirming formulation-specific differences in how well and rapidly the gel was formed.

Gel degradation measurement

Solutions of 4 formulations (No. 4, 7, 8 and 9) were prepared, respectively (n=3). After weighing the empty syringe and the prepared sol-filled syringe, the injected weight was calculated. 45 mL of pH 7.2 PBS was added to a 50 mL conical tube. In vitro degradation of the prepared gel was measured by injecting the sample into 45 mL pH7.2 PBS. The medium buffer was refreshed daily. Each sample was incubated in a shaking bath at 37°C and 50 rpm for 2 weeks. Then, the weight loss rate was calculated after drying the sample with hot air of 70° C. or higher for 72 hours (n=3).

moisture diffusion test

The experiments were designed assuming that the rate of water diffusion during the solvent exchange reaction was related to the rate of gel formation. Formulations 4, 7, 8, and 9 prepared in a glass tube having a diameter of 5.6 mm were filled at the same height. The glass tube filled with the formulation was fixed so as not to be tilted, and 1 mL of pH 7.2 PBS was dropped with a pipette ( FIG. 4 ). The moisture diffusion distance was observed by changing the transparency from the transparent state to the opaque state. The water diffusion rate was measured immediately after dropping 1 mL of pH 7.2 PBS and after 6, 12, and 24 hours.

drug release test

There are various methods for dissolving the injection, such as the dialysis sac method, the reverse dialysis bag method, the method using the device 2, the method using the device 4, the membraneless method, etc. (Gray, Cady et al. 2018) . Since the formulation designed in the present invention easily forms a gel immediately after injection, a membrane-free dissolution method can be used as a dissolution method for an in situ gel-based long-acting parenteral agent (Janagam, Wang et al. 2016). In the dissolution test of long-acting parenteral preparations, accelerated conditions were applied due to problems due to drug stability, loss of release medium, microbial growth and long-term experiments. There are a number of variables such as pH, aeration medium and agitation (Shen and Burgess 2012). Accelerated conditions were established taking into account the drug release mechanism of the formulation. Since the polymer used in the formulation is dissolved in EtOH, the release medium was prepared using EtOH. A pH 7.2 PBS was prepared using sodium phosphate monobasic and sodium phosphate dibasic, and the PBS and EtOH were mixed in a ratio of 8:2 to prepare a release medium. When the final release medium was ready, 45 mL was added to a 50 mL conical tube. Then 0.25 mL of formulations 4, 7, 8 and 9 were injected with a 20 gauge needle to form a gel. The conical tube containing the formed gel was transferred directly to the shaking water bath. Shaking bath conditions were set at 25° C. in consideration of drug stability, and stirring rpm was set at 50. Sampling times were set to 30 minutes, 1 hour, 4 hours, 8 hours, 12 hours, 24 hours, 2 days, 4 days, 6 days, 8 days, and 10 days. The HPLC system for analyzing 5-FU was a 1200 Infinity series (Agilent technology, Santa Clara, USA). Chromatographic conditions were as follows; C18 reversed phase column, 3.9 mm x 300 mm (Waters, Leinster, Ireland); Mobile phase, 70% 0.005 M S-1 HSA solution at a flow rate of 2 mL/min. All solutions used for analysis are HPLC grade. The injection volume was 20 μl and the UV detection wavelength was 254 nm. The retention time is about 1.73 min.

in vivo test

in vivo gel formation

To confirm whether the designed formulation forms a gel in vivo, 10-week-old male SD rats were prepared. After removing the hair from the back of SD rats and injecting 0.7 mL of formulations 4 and 8 without drug loading , gel formation in vivo was observed.

in vivo Gel degradation

In order to confirm whether the formulation designed in the present invention has biodegradability, an in vivo gel degradation experiment was performed using 10-week-old male SD rats (Gao, Deng et al. 2013). After depilating the back of SD rats and injecting 0.7 mL of formulations 4 and 8 without drug loading into 4 rats, the rats were sacrificed 1 day, 1 week, 2 weeks and 4 weeks later and the gel was removed from the body. was collected. By measuring the weight of each gel, the amount of decomposition of the gel over time was investigated.

Experiment result

Evaluation of sol-gel transition

As a result of injecting the formulation into pH 7.2 PBS and measuring the sol-gel transition, immediate gel formation was confirmed in the formulations of EUDRAGIT E grade 30% (w/w) and EUDRAGIT RS grade 30% (w/w). However, even in the EUDRAGIT RL 30% (w/w) formulation, a gel was formed but rapidly disintegrated ( FIG. 5 ). Comparing the gel-forming ability according to grade, EUDRAGIT E grade was the best, followed by EUDRAGIT RS grade and EUDRAGIT RL grade, respectively. Of the formulations with 20% (w/w) polymer ratio, only 20% of EUDRAGIT E grade formed a gel. The formulations of EUDRAGIT E grade 15% and RS grade 15% and EUDRAGIT E grade 15% and RL grade 15% formulations immediately formed a gel ( FIG. 6 ). In long-acting parenteral formulations, rapid sol-gel transition is a very important feature. The initial burst appears to occur when gel formation is not instantaneous. However, when a large amount of polymer is used so that gelation occurs faster and better, there is a problem that the injectable viscosity is exceeded. In this experiment, the sol-gel transition according to polymer grade, concentration and combination of the two grades was investigated. The gel formed well when the polymer concentration was 30% (w/w). EUDRAGIT E grades, unlike EUDRAGIT RS and RL grades, have dimethylamine functionality instead of trimethylammonium. This means that EUDRAGIT E grade is insoluble in water and more prone to gel formation. Compared to EUDRAGIT RS and EUDRAGIT RL grades, EUDRAGIT RS grades form a gel while retaining their appearance, whereas EUDRAGIT RL grades do not retain their shape over time. The EUDRAGIT RS grade has an ethyl group and the EUDRAGIT RL grade has a methyl group, which appears to be more soluble in water because the EUDRAGIT RL grade is more hydrophilic. When the polymer concentration was 20%, only EUDRAGIT E grade formed a gel matrix that retained its shape. EUDRAGIT RS and EUDRAGIT RL grades showed polymer precipitation but did not retain the morphology of the gel matrix. Based on these results, it was found that EUDRAGIT E grade had the highest gel-forming ability. Each formulation was made in combination with EUDRAGIT E grade 15% based on EUDRAGIT RL grade 15% and EUDRAGIR RS grade 15%. In this formulation, the shape of the gel matrix was well maintained. The EUDRAGIT L and S grade sol-gal transitions initially formed a thin precipitation layer but dissolved over time in PBS, pH 7.2. Of course, the L and S grades are soluble at pH 6 and above 7, respectively. However, at low pH they form gels that can be applied to pH-dependent systems.

Gel degradation measurement

The degree of degradation of the formed gel under in vitro conditions was measured. Since the degree of degradation was expected to affect the dissolution tendency, we sought to investigate the different degradation progressions depending on the formulation. A comparison of EUDRAGIT E grade 30% (formulation 4) and EUDRAGIT E grade 15% (formulation 7) showed that the EUDRAGIT E grade 15% formulation showed a higher degree of degradation. Comparing EUDRAGIT E grade 30%, EUDRAGIT E grade 15% and RS grade 15% (formulation 8) with EUDRAGIT E grade 15% and RL grade 15% (formulation 9), EUDRAGIT E grade 15% and RL grade 15% It was the highest with 93.4%, followed by EUDRAGIT E grade 30% with 81.5%, and EUDRAGIT E grade 15% and RS grade 15% with 70.6%. Based on these results, the dissolution rates were 15% EUDRAGIT E grade and 15% RL grade; EUDRAGIT E grade 30%; It could be predicted that the order would be EUDRAGIT E grade 15% and RS grade 15%, and the dissolution test results were consistent with the prediction ( FIG. 7 ).

moisture diffusion test

The present inventors attempted to compare the rate of gel barrier formation in the sol-gel transition by comparing the degree of water diffusion by the polymer. Assuming that the polymer barrier will be formed quickly if water diffusion is fast, drug entrapment will be good and the initial burst will be low. As soon as the pH 7.2 buffer was added dropwise, the water diffusion ranged from 1.8 mm to 2.3 mm, showing no significant difference ( FIG. 8 ). After 6 hours, there was a small change in Formulation 4 and a slight increase in Formulations 7, 8 and 9. Formulations 8 and 9 diffused moisture for 12 hours and significantly increased. There was no significant difference between the 24 hour observation and the 12 hour observation. Formulations 8 and 9 showed relatively high water diffusion, while Formulations 4 and 7 showed low water diffusion. EUDRAGIT, E grades have an uncharged dimethylamine functional group, whereas RL and RS have a charged trimethylammonium functional group, showing a high affinity for water. EUDRAGIT RL grades and RS grades exhibit relatively high water diffusion rates, while EUDRAGIT E grades exhibit low diffusion rates. The difference between the EUDRAGIT RL grade and the RS grade is not large, but looking at the chemical structure, the EUDRAGIT RL grade has a methyl group and is more hydrophilic than the EUDRAGIT RS grade having an ethyl group. The degree of hydrophilicity according to the chemical structure is predicted in the order of EUDRAGIT RL grade, RS grade and E grade, and moisture diffusion is also high in the same order. A logical inference is that if the polymer barrier is rapidly formed due to high moisture diffusion, the first burst release will be suppressed and the dissolution sequence will be EUDRAGIT E grade, RS grade, RL grade. However, as a result of the dissolution test, the dissolution occurred rapidly in the order of EUDRAGIT RL grade, E grade, and RS grade during the first 12 hours. The first of two ways to explain these results is barrier penetration. In other words, the high hydrophilicity of the polymer allows the drug to flow out of the polymer barrier, but EUDRAGIT RL grade as well as EUDRAGIT RS grade have hydrophilicity, so this assumption is not reasonable in situations where the dissolution rate is slower than EUDRAGIT E grade. Second, the decomposition of the part in contact with the water may have accelerated and the drug may have flowed out.

drug release test

Due to drug stability, loss of release medium and bacterial growth, etc., in vitro release experiments were performed under accelerated conditions. There are several variables such as temperature, pH, release medium and agitation. In this experiment, the release medium was the most important factor in drug release, and the accelerated conditions were established. The polymer used in the formulation is insoluble in water and is soluble in certain organic solvents. In the case of EtOH, since the polymer was well dissolved, a release medium was prepared while controlling the ratio of EtOH. If the amount of EtOH is too small, the accelerating effect is low, and if it is too high, the initial burst release is too large or the gel formation itself is greatly affected. Therefore, in order to establish optimal conditions, it was attempted to determine the composition of the release medium by preliminarily conducting a dissolution experiment in a solvent made of EtOH. When 20% EtOH was used, there was no problem in gel formation and 70% of the drug was dissolved after 7 days. As a result of the experiment after establishing the conditions, it was confirmed that significant differences appeared for each grade. First, comparing the EUDRAGIT E grade 15% rate formulation (Formulation 7) and 30% rate formulation (Formulation 4), the dissolution pattern was lower in the 30% rate formulation and higher in the 15% rate formulation. When these results are reviewed over time, the dissolution results approximately doubled during the first 4 hours, and then the gap narrowed over time, but dissolution was slower than in the 30% formulation until the final sampling time of 10 days. Next, as a result of comparing territorial water patterns according to polymer grades, EUDRAGIT E grade 15% and RL grade 15% (formulation 9) showed the fastest dissolution pattern, and EUDRAGIT E grade 15% and RS grade 15% (formulation 8) had the most Dissolution was slow. Similar to the dissolution pattern according to the polymer concentration, the deviation was large during the first 4 hours and the pattern remained constant thereafter ( FIGS. 9 and 10 ). The combination of EUDRAGIT RL grade resulted in fast dissolution and the combination of EUDRAGIT RS grade resulted in slow dissolution. In the previous in vitro gel degradation experiment, the degree of gel degradation was shown to be different in the order of EUDRAGIT RL grade, E grade and RS grade. Furthermore, the main factor determining drug dissolution is how well the gel is decomposed after formation, and the ability of the drug to penetrate the formed gel matrix is not an important factor. Accordingly, when the polymer concentration is lowered, drug release is accelerated, and when it is increased, drug release is slowed. In addition, drug release is fast when EUDRAGIT RL grade is used in the formulation, whereas drug release is slow when EUDRAGIT RS grade is used. This is the basis for the development of a system that can control the drug release pattern as desired. It can be applied not only to 5-FU intratumoral formulations, but also to other drugs applied to formulations for subcutaneous or intramuscular injection for systemic circulation. The drug release pattern can also be adjusted to suit the pharmacokinetic properties of each drug.

in vivo test

in vivo gel formation

As a result of the in vivo sol-gel transfer experiment of each formulation, it was confirmed that the formulation maintained its shape for a short time in vivo upon injection, and the appearance of the gel was revealed a few seconds after injection ( FIG. 11 ).

in vivo Gel degradation

EUDRAGIT is widely used in oral formulations. L and S grades are mainly used for enteric coating of oral tablets, E grades are mainly used for film coating and RL and RS are applied for sustained release. However, it has never been used in injectables and has never been approved by the FDA for injectable use. EUDRAGIT is known to be non-toxic, but its biodegradability is unknown. Therefore, in the development of long-acting parenteral formulations using EUDRAGIT, confirmation of biodegradability of polymers is a very important issue. Furthermore, biodegradability is an essential condition for the development of IM and SC injections for systemic circulation. The biodegradability of the gel matrix was investigated through subcutaneous injection into SD rats, and EUDRAGIT E grade 30% formulations and EUDRAGIT E grade 15% and RS grade 15% formulations were used. After 1 week, the weight of the gel decreased by 20%, after 2 weeks the weight of the gel decreased by 50%, and after 4 weeks by 80% ( FIG. 12 ). This is the result showing the biodegradability of EUDRAGIT.

As described above in detail a specific part of the present invention, for those of ordinary skill in the art, this specific description is only a preferred embodiment, and it is clear that the scope of the present invention is not limited thereto. Accordingly, the substantial scope of the present invention will be defined by the appended claims and their equivalents.

Claims (15)

1. A drug delivery system composition for local injection comprising (a) an organic solvent and (b) a polymer represented by the following formula (1) dissolved in the organic solvent:

   Formula 1

   

   In Formula 1, R ₁ to R ₃ are each independently hydrogen or C ₁ to C ₃ alkyl, R ₄ is or N(R ₅ ) ₂ or N(R ₅ ) ₃ ⁺ , and R ₅ is C ₁ to C ₃ alkyl, and n is an integer from 100 to 3,000.
2. The composition of claim 1, wherein R ₁ to R ₃ are each independently hydrogen, C ₁ alkyl, or C ₂ alkyl, and R ₅ is C ₁ alkyl.
3. The composition of claim 2, wherein R ₁ and R ₂ are each independently hydrogen or C ₁ alkyl, R ₃ is C ₁ or C ₂ alkyl, and R ₅ is C ₁ alkyl.
4. The composition of claim 1, wherein the organic solvent is at least one solvent selected from the group consisting of ethanol, methanol, and N-methyl pyrrolidone (NMP).
5. The composition according to claim 1, wherein the polymer is contained in an amount of 25 to 35 w/w% based on the total composition.
6. The first polymer according to claim 1, wherein the polymer represented by Formula 1 is (i) R ₁ to R ₃ are C ₁ alkyl, R ₄ is —N(R ₅ ) ₂ , and R ₅ is C ₁ alkyl. Wow; (ii) R ₁ and R ₂ is a C ₁ alkyl, R ₃ is a C ₂ alkyl, R ₄ is -N (R _{5) 3} ⁺ a, R ₅ is characterized in that the mixture of the second polymer is C ₁ alkyl A composition comprising
7. The composition according to claim 6, wherein the first polymer and the second polymer are present in a content ratio of 4:6 to 6:4 in the composition.
8. The first polymer according to claim 1, wherein the polymer represented by Formula 1 is (i) R ₁ to R ₃ are C ₁ alkyl, R ₄ is —N(R ₅ ) ₂ , and R ₅ is C ₁ alkyl. Wow; (ii) and R ₁ is hydrogen, R ₂ and R ₃ is a C ₁ alkyl, R ₄ is ^{_{-N (R 5) 3 +,}} R 5 is characterized in that the mixture of a third polymer of C ₁ alkyl composition.
9. The composition according to claim 8, wherein the first polymer and the third polymer are present in a content ratio of 4:6 to 6:4 in the composition.
10. 10. A composition for preventing or treating solid cancer, comprising the drug delivery composition for local injection of any one of claims 1 to 9 and the anticancer composition for solid cancer as an active ingredient.
11. 11. The composition of claim 10, wherein the solid cancer is selected from the group consisting of colon cancer, esophageal cancer, stomach cancer, pancreatic cancer, breast cancer, cervical cancer, ovarian cancer, lung cancer, jaw cancer, tongue cancer and liver cancer.
12. The composition of claim 10, wherein the anticancer composition is 5-FU (5-fluorouracil) or a pharmaceutically acceptable salt thereof.
13. A method for preparing a drug-loaded formulation for local injection comprising the steps of:

    (a) adding a polymer represented by Formula 1 to an organic solvent:

    Formula 1

    

    In Formula 1, R ₁ to R ₃ are each independently hydrogen or C ₁ to C ₃ alkyl, R ₄ is or N(R ₅ ) ₂ or N(R ₅ ) ₃ ⁺ , and R ₅ is C ₁ to C ₃ alkyl, n is an integer from 100 to 3,000;

    (b) stirring until the organic solvent to which the polymer represented by Formula 1 is added becomes transparent;

    (c) adding the drug to the stirred mixture.
14. The method of claim 13, wherein the organic solvent is at least one solvent selected from the group consisting of ethanol, methanol, and N-methyl pyrrolidone (NMP).
15. 14. The method according to claim 13, wherein step (a) is carried out by adding the polymer such that the polymer is included in an amount of 25 to 35 w/w% with respect to the total composition.

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