KR20200031581A - Thermosponge nanoparticle platform for co-delivery of multi-drugs and its use - Google Patents

Abstract

The present invention relates to a thermosensitive nanosponge platform (TNP) for simultaneous delivery of hydrophilic and hydrophobic drugs and pharmaceutical uses thereof. The TNP according to the present invention can simultaneously deliver a hydrophobic drug and a hydrophilic drug. In addition, the hydrophobic drug can be maintained in the body at a constant concentration for a long time.

Description

Thermosponge nanoparticle platform for co-delivery of multi-drugs and its use}

The present invention relates to a thermosensitive nanosponge platform (TNP) platform for the simultaneous delivery of hydrophilic and hydrophobic drugs and uses thereof.

Cancer cells refer to cells in which normal cells do not die and continue to survive as the number of divisions disappears due to genetic variation. They continue to multiply even when attached to other cells, and even push out normal cells, expanding the area and eventually spreading throughout the body, leading to death. According to estimates from the International Cancer Institute (IARC), cancer is the second most common cause of death in both developing and developed countries, of which it is estimated that about 13% of the world's cancer cases occur in southwest Asia. . In particular, the disease has not been overlooked for more than 10 years as the leading cause of death among Koreans.

Chemotherapy, the main cancer treatment modality, is considered to be a major obstacle in the treatment of clinical cancers, with multi-drug resistance (MDR effect) defined as tumor resistance to the cytotoxic effects of several drugs. In order to overcome this limitation, a treatment method using nanoparticles (NP) has been developed. Among various nanoparticles, nanoparticles capable of delivering two or more different drugs simultaneously can overcome problems such as drug resistance, high cytotoxicity, limitations in clinical application, and insolubility, compared to nanoparticles that can deliver only one drug. In addition, it has the advantage of being able to maximize the efficacy of the drug by creating a synergistic effect.

Prior literature

[Patent Document 1] Republic of Korea Patent Publication No. 2009-0088614 (2009. 08. 20)

The present inventors tried hard to develop a drug delivery system capable of simultaneously delivering two types of drugs (hydrophilic and hydrophobic), and as a result, a biocompatible polymer approved by the US Food and Drug Administration (FDA), PLA (poly-D, L-lactide) , ~ 18kDa, PLA-COOH) and a core part composed of a hydrophobic drug, and a thermosensitive core-shell nano consisting of a Pluronic F127 (12.6kDa) excellent in biocompatibility and biodegradability and a shell part containing a hydrophilic drug. When preparing the particles, it was confirmed that the hydrophilic drug and the hydrophobic drug can be simultaneously delivered to the body, and that the delayed release of the hydrophobic drug is possible after the release of the hydrophilic drug, and the present invention has been completed.

An object of the present invention is to provide a temperature-sensitive nanosponge platform (Thermosponge nanoparticle (TNP) platform) for the simultaneous delivery of hydrophilic and hydrophobic drugs.

The present invention also aims to provide a pharmaceutical composition for the treatment of cancer comprising the temperature-sensitive nano-sponge platform.

The present invention also aims to provide a cosmetic composition comprising the temperature-sensitive nano-sponge platform.

The term “biodegradable” as used herein means that the polymer can be chemically degraded in the body to form a non-toxic compound, the rate of degradation being the same or different from the rate of drug release.

As used herein, the term "biocompatibility" means a substance that interacts with the human body without an undesirable subsequent effect.

The term "sustained release" as used herein refers to the continuous release of a drug or therapeutic agent or any combination thereof over a period of time.

The term "controlled release" as used herein refers to the control of the rate and / or amount of drug or therapeutic agent delivered according to the temperature sensitive nanosponge platform of the present invention. Controlled release can be continuous or discontinuous and / or linear or nonlinear. This can be achieved using one or more types of polymer composition, drug loading, excipients or degradation enhancers, or other modifiers, alone, in combination or continuously administered to provide the desired effect.

The term "drug" as used herein refers to an organic or inorganic compound or substance that has bioactivity and is used or modified for therapeutic use. Protein, oligonucleotide, DNA and gene therapy are included in the broader drug definition.

The term “therapeutic agent” as used herein refers to any that induces the desired pharmacological, immunogenic and / or physiological effect by local and / or systemic action when administered to an organism (human or non-human animal). Means a compound or composition. Thus, the term includes compounds or chemicals that are traditionally considered biopharmaceuticals including drugs, vaccines, and molecules such as proteins, peptides, hormones, nucleic acids, gene constructs, and the like. “Therapeutic agent” includes compounds or compositions used in all major therapeutic fields, including but not limited to: anti-infective agents such as antibiotics and antiviral agents; Analgesic and analgesic combinations; Local and general anesthetics; Anorexia nervosa; Anti-arthritis agents; Anti-asthma; anticonvulsant; Antidepressants; Antihistamines; Anti-inflammatory agents; Antiemetic; Anti-migraine control; Anti-neoplastic agents; Anti-itching agents; Antipsychotics; fever remedy; Anti-spasmodics; Cardiovascular preparations (including calcium channel blockers, β-blockers, β-agonists and antiarrhythmic agents); Antihypertensive drugs; Chemotherapy agents; diuretic; Vasodilators; Central nervous system stimulants; Cough and cold preparations; Decongestants; Diagnostic agents; hormone; Bone stimulants and bone resorption inhibitors; Immunosuppressants; Muscle relaxants; Psychostimulants; sedative; Neurostabilizers; Proteins, peptides and fragments thereof (naturally occurring, chemically synthesized or recombinantly produced); And nucleic acid molecules (ribbon nucleotides (RNA) or deoxyribonucleotides (DNA), gene constructs, expression vectors, plasmids, including polymer forms of two or more nucleotides, double- and single-stranded molecules and supercoiled or condensed molecules, Antisense molecule, etc.).

As used herein, the terms "peptide", "polypeptide", "oligopeptide" and "protein" can be used equally when referring to peptide or protein drugs and are limited to a particular molecular weight, peptide sequence or length, bioactivity or therapeutic field. Does not work.

As used herein, the term "therapeutic effect" refers to any improvement in the disease of a subject, human or animal treated according to the method, and for prophylactic or preventive effects, or for physical investigation, laboratory or mechanical methods. Obtaining any relief in severity of the disease, the signs or symptoms of the disease, disorder or condition that can be detected by. The term “treatment” or “treating” as used herein, unless defined otherwise in connection with a particular disease or condition, means (i) an animal that is prone to disease, disease, and / or illness, but has not yet been diagnosed as ill; or Preventing the occurrence of a disease, disorder or illness in a person; (ii) inhibit the disease, disorder or disease, ie inhibit its progression; And / or (iii) alleviating the disease, disorder or disease, ie causing degeneration of the disease, disease and / or disease.

According to the first embodiment,

The present invention provides simultaneous delivery of a hydrophilic and hydrophobic drug consisting of a core portion comprising an aliphatic polyester and a hydrophobic drug, and a shell portion containing a polymer and a hydrophilic drug surrounding the core part. To provide a temperature-sensitive nano-sponge platform for. 1 is a schematic diagram of a temperature-sensitive nanosponge platform for simultaneous delivery of hydrophilic and hydrophobic drugs according to the present invention.

In the temperature-sensitive nanosponge platform according to the present invention, the weight ratio of the aliphatic polyester in the core portion and the polymer in the shell portion is 1:10 to 1:20.

In the temperature-sensitive nanosponge platform according to the present invention, the aliphatic polyester is poly lactic acid (Poly Lactic Acid, PLA), polyglycolic acid (PGA) or polylactic acid-glycolic acid copolymer (Polylactide-co-Glycolide) , PLGA). The polylactic acid (PLA), polyglycolic acid (PGA), and polylactic acid-glycolic acid copolymer (Polylactide-co-Glycolide, PLGA) are approved for clinical use by the United States Food and Drug Administration (FDA). As a synthetic polymer, it has the advantage of being degraded in vivo and having excellent biocompatibility.

In the temperature-sensitive nano-sponge platform according to the present invention, the polymer in the shell is characterized by being a Pluronic series. Pluronic copolymer is a linear triblock copolymer composed of poly (ethylene oxide) -poly (propylene oxide) -poly (ethylene oxide) as a temperature-sensitive polymer that exhibits sol / gel transition in aqueous solution. For example, the polymer in the shell portion may be Pluronic F127 (poloxamer 407).

In the temperature-sensitive nanosponge platform according to the present invention, the nanosponge has an average particle diameter of 50 to 100 nm. It is possible to efficiently deliver nanoparticles to a desired site while minimizing biostimulation within the above range.

In the temperature-sensitive nano-sponge platform according to the present invention, the drug is nucleic acid, protein, polypeptide, carbohydrate, inorganic substance, antibiotic, anticancer agent, antibacterial agent, steroids, anti-inflammatory analgesic, sex hormone, immunosuppressant, antiviral agent, anesthetic agent, port As an antiemetic, antihistamine, local anesthetic, antiangiogenic agent, vasoactive agent, anticoagulant, immunomodulatory agent, cytotoxic agent, antibody, neurotransmitter, psychoactive agent, oligonucleotide, lipid, cell, tissue, cancer chemotherapy and vaccine It is characterized by being one or more substances selected from the group consisting of.

In the temperature-sensitive nano-sponge platform according to the present invention, the temperature-sensitive nano-sponge platform is characterized in that it is used as a support for molding materials, cosmetic materials or tissue regeneration.

The temperature-sensitive nanosponge platform according to the present invention can simultaneously deliver hydrophobic drugs and hydrophilic drugs. In addition, the hydrophobic drug is encapsulated in the micelle formed by the polymer in the shell portion, is contained in the aliphatic polyester matrix, and is slowly released from the polymer micelle when injected into the body, so it can be maintained for a long time in a certain concentration in the body.

According to the second embodiment,

The present invention is to provide a pharmaceutical composition for cancer treatment comprising the temperature-sensitive nano-sponge platform. 2 is a schematic diagram of an effective cancer treatment process according to the release of hydrophilic and hydrophobic drugs from the temperature-sensitive nanosponge platform according to the present invention.

In the pharmaceutical composition for the treatment of cancer according to the present invention, the cancer is pancreatic cancer, liver cancer, breast cancer, lung cancer, stomach cancer, rectal cancer, gallbladder cancer, ovarian cancer, bladder cancer, colon cancer, lymphoma, brain cancer, uterine cancer, prostate cancer and malignant melanoma Or biliary tract cancer.

In the pharmaceutical composition for the treatment of cancer according to the present invention, the hydrophobic drug is vinblastine, etoposide, alkeran, cytoxan, daunorubicin, hydrrea ), Mithramycin, Mitomycin, Mitoxantrone, Nitrogen Mustard, Velban, Vincristine, Carboplatinum, Idarubi May include, but are not limited to, Idarubicin, Irinotecan, Leustatin, Navelbine, Taxotere, Topotecan adriamycin, Daunomycin or Paclitaxel It is not.

In the pharmaceutical composition for treatment of cancer according to the present invention, the hydrophilic drug is Busulfan, Chlorambucil, Cyclophosphamide, Melphalan, Cisplatin, Ipoh Ifosfamide, cytarabine, 5-fluorouracil (5-FU), methotrexate (MTX), actinomycin-D (Actinomycin-D), bleomycin or gemcitabine It may include, but is not limited to.

In the pharmaceutical composition for the treatment of cancer in the present invention, the pharmaceutical composition is characterized in that it further comprises a pharmaceutically acceptable carrier. The pharmaceutically acceptable carrier includes a carrier and a vehicle commonly used in the pharmaceutical field, specifically, ion exchange resins, alumina, aluminum stearate, lecithin, serum proteins (e.g., human serum albumin), buffer substances ( For example, various phosphates, glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes (e.g. protamine sulfate, disodium hydrogen phosphate, camphorium hydrogen phosphate, sodium chloride and Zinc salt), colloidal silica, magnesium trisilicate, polyvinylpyrrolidone, cellulose-based substrate, polyethylene glycol, sodium carboxymethylcellulose, polyarylate, wax, polyethylene glycol or wool paper, and the like.

According to the third embodiment,

The present invention is to provide a cosmetic composition comprising the temperature-sensitive nano-sponge platform.

In the cosmetic composition according to the present invention, the cosmetic composition is characterized in that it comprises a wrinkle improving agent, skin aging inhibitor, skin whitening agent or anti-hair loss agent.

In the cosmetic composition according to the present invention, the hydrophilic drug is characterized in that it contains arbutin, adenosine, vitamin C, vitamin B3, vitamin B5, vitamin H, acetylglucosamine, madecassoside or selenium aspartate.

In the cosmetic composition according to the present invention, the hydrophobic drug is eugenol, retinol, astaxanthin, caffeine, carnosine acid, catechin, coenzyme-Q10, curcumin, elazinic acid, peric acid, idebenone, isoflavone, linoleic acid, Lipophosphoric acid, lycopene, oleanoic acid, floretine, quercetin, resveratrol, squalane, squalene, tannic acid, vitamin A, vitamin B, vitamin E, vitamin F, cholesterol, phytosphingosine, glycospingolipide, beta-sitosterol, It is characterized by containing lauric acid or lecithin.

In the cosmetic composition according to the present invention, the cosmetic composition is prepared in the form of a flexible lotion, nutrition lotion, nutrition cream, massage cream, essence, eye cream, cleansing cream, cleansing foam, cleansing water, pack, spray or powder It is characterized by.

The temperature-sensitive nano-sponge platform according to the present invention has a wide range of applications in the field of disease treatment because it can treat various diseases as well as cancer treatment. In addition, the whitening and wrinkle improvement functions can be put in one formula to maximize the functionality, so it can be used as a functional beauty care material. In particular, it is expected to be applied to functional cosmetics and therapeutic cosmetics that can improve acne, psoriasis, and atopy.

Figure 1 shows a schematic diagram of a temperature-sensitive nanosponge platform (Thermosponge nanoparticle (TNP) platform) for the simultaneous delivery of hydrophilic and hydrophobic drugs according to the present invention.
Figure 2 shows a schematic diagram of an effective cancer treatment process by the release of hydrophilic and hydrophobic drugs from the temperature sensitive nanosponge platform (TNP platform) according to the present invention.
Figure 3 shows the size of the TNP loaded with a temperature-sensitive nano-sponge platform (TNP platform) and hydrophilic (Lys: lysozyme) and hydrophobic (NR: Nile red) drugs according to Example 3 of the present invention.
Figure 4 shows the dispersion and distribution of the TNP loaded simultaneously with the temperature-sensitive nano-sponge platform (TNP platform) and hydrophilic (Lys: lysozyme) and hydrophobic (NR: Nile red) drugs according to Example 3 of the present invention.
Figure 5 shows the surface charge of the TNP loaded with a temperature-sensitive nano-sponge platform (TNP platform) and hydrophilic (Lys: lysozyme) and hydrophobic (NR: Nile red) drugs according to Example 3 of the present invention.
6 shows the release behavior of hydrophilic and hydrophobic drugs released from TNP according to Example 4 of the present invention.

Hereinafter, various examples are presented to help understanding of the invention. The following examples are only provided to more easily understand the invention, and the protection scope of the invention is not limited to the following examples.

<Example>

Example 1. Preparation of temperature sensitive nano sponge (TNP) platform

The temperature-sensitive nano-sponge is made by nanoprecipitation using PLA (poly-D, L-lactide, ~ 18kDa, PLA-COOH) and Pluronic F127 (12.6kDa) in a weight ratio of 1:20. It was prepared. PLA (10 mg) was dissolved in 1 mL of acetone (99%, sigma aldrich), and after dissolving Pluronic F127 in the reaction solution, the reaction was maintained at room temperature for 2 hours. The reaction solution was slowly dropwise added to 5 mL of tertiary distilled water (DIW) stirred at 530 rpm, and reacted at room temperature for at least 12 hours. Then, ultrafiltration (Amicon Ultra-15 filter) was performed to increase the purity of the nanoparticles.

Example 2. Preparation of a temperature-sensitive nanosponge platform loaded with hydrophilic and hydrophobic drugs

The hydrophobic model drug, Nile red (200 μg), was dissolved in 1 mL of acetone (99%, sigma aldrich), added to PLA (10 mg), and reacted at room temperature for 1 hour. Pluronic F127 was dissolved in the reaction solution and reacted for 2 hours. Then, the reaction solution was slowly dropwise added to 5 mL of tertiary distilled water stirring at 530 rpm, and reacted at room temperature for at least 12 hours. The hydrophilic model drug lysozyme (200 μg / 100 μL DIW) was added to the TNP solution (1 mL) loaded with the hydrophobic drug, and then loaded at 4 ° C. for 2 hours. As the outer layer swells at 4 degrees Celsius due to temperature-sensitive properties, the hydrophilic model drug can be stably loaded without using an organic solvent, and effective loading is possible due to the electrostatic interaction between the negative charge of the PLA core and the positive charge of the lysozyme. . Finally, ultrafiltration (Amicon Ultra-15 filter) was performed to remove unloaded hydrophobic drugs or hydrophilic drugs.

Example 3. Analysis of the size, dispersion and surface charge of the temperature sensitive nano sponge platform

The size, dispersion and surface charge of the TNP platform prepared according to Example 1 and the Nile red and lysozyme prepared according to Example 2 were analyzed using Zetasizer (Nano-ZS, Malvern) equipment. And the results are shown in Figs. 2 to 4, respectively.

Regarding the size of the TNP, the size of the TNP platform (TNP) prepared by the nanoprecipitation method was 75 nm, and the nanoparticles (NR @ TNP) loaded with the hydrophobic drug Nile red (NR) and the hydrophilic / hydrophobic drug The size of the nanoparticles (Lys / NR @ TNP) loaded simultaneously was also found to be similar to the TNP without drug loading at 75 nm. Thus, it was confirmed that drug loading did not affect the physicochemical properties of the nanoparticles (FIG. 2).

In the case of the dispersion value (PDI: polydispersity), the results of comparing the before and after loading of the drug were all less than 0.2, and it was confirmed that the drug loading was stably performed as the distribution also showed almost similar distribution. In addition, as a result of measuring the surface charge, the surface charge value of the TNP itself showed a negative charge of -6 mV, and when the hydrophobic drug was loaded, it showed almost similar values without statistically significant difference (FIG. 4). When the lysozyme, a positively charged hydrophilic model drug, was loaded, it showed a tendency to show a slightly lower surface charge value, but there was no statistically significant difference. That is, it was confirmed that the hydrophilic drug and the hydrophobic drug were stably loaded on the TNP at the same time.

Example 4. Analysis of the release of drugs loaded on a temperature sensitive nanosponge platform

Lysozyme (200 μg) and Nile Red (40 μg) loaded with 1 mL of a temperature-sensitive nano sponge platform into a dialysis tube (Float-A-Lyzer G2 Dialysis Device, 100 kD), then 10 mL of pre-warmed PBS at 100 rpm and 37 ° C. conditions The release experiment was conducted. To confirm the release behavior of the drug by time, 10 ml of release buffer was collected at 20 minutes, 6 hours, 1 day, 2 days, 3 days, 5 days, 7 days, 9 days, 12 days, 15 days, and 17 days ( The experiment was repeated 3 times n = 3). To analyze the release behavior of hydrophilic model drugs, Lysozyme analyzes absorbance using a 96 well plate at a wavelength of 570 nm through a Coomassie assay, and Nile Red uses ex.530 nm / em.570 nm wavelength to analyze the release behavior of hydrophobic model drugs. Fluorescence was analyzed using a 96 well plate (black). At the time after release of the hydrophilic model drug, the amount of Nile red, a hydrophobic model drug remaining in the TNP platform, was quantitatively analyzed by freeze-drying the TNP platform remaining in the Dialysis tube.

As a result, it was confirmed that the hydrophilic Lysozyme was released relatively quickly compared to the hydrophobic Nile red. That is, it has been confirmed that the hydrophobic drug exhibits sustained release behavior, thereby improving the therapeutic efficacy by first releasing the hydrophilic drug when treating the disease, and subsequently, by continuously releasing the hydrophobic drug, a perfect or more effective (synergistic) treatment is possible.

The TNP platform capable of simultaneously loading the hydrophilic drug and the hydrophobic drug according to the present invention can be used for various incurable diseases and diseases treatment fields and beauty care as well as treatment of malignant tumors that are resistant to drugs using different release rates of drugs. Is expected.

Since the specific parts of the present invention have been described in detail above, it is clear that for those skilled in the art, these specific technologies are only preferred embodiments, and the scope of the present invention is not limited thereto. Therefore, the substantial scope of the present invention will be defined by the appended claims and their equivalents.

Claims (12)

i) a core portion comprising poly lactic acid (PLA) and a hydrophobic drug as an aliphatic polyester, and
ii) a polymer surrounding the core portion, which consists of a shell portion containing Pluronic F127 and a hydrophilic drug,
It characterized in that the weight ratio of the aliphatic polyester in the core portion and the polymer in the shell portion is 1:10 to 1:20,
Temperature sensitive nanosponge platform for simultaneous delivery of hydrophilic and hydrophobic drugs.
According to claim 1,
The average particle diameter of the nano-sponge is characterized in that 50 to 100 nm, temperature-sensitive nano-sponge platform.
According to claim 1,
The drugs include nucleic acids, proteins, polypeptides, carbohydrates, inorganic substances, antibiotics, anticancer agents, antibacterial agents, steroids, anti-inflammatory drugs, sex hormones, immunosuppressants, antiviral agents, anesthetics, and antiemetic agents. Group consisting of antihistamines, local anesthetics, antiangiogenic agents, vasoactive agents, anticoagulants, immunomodulators, cytotoxic agents, antibodies, neurotransmitters, psychoactive agents, oligonucleotides, lipids, cells, tissues, cancer chemotherapeutic agents and vaccines Characterized in that at least one material selected from, temperature-sensitive nano-sponge platform.
According to claim 1,
The temperature-sensitive nano-sponge platform is characterized in that it is used as a support for molding materials, cosmetic materials or tissue regeneration, temperature-sensitive nano-sponge platform.
A pharmaceutical composition for the treatment of cancer, comprising the temperature-sensitive nanosponge platform according to claim 1.
The method of claim 5,
The cancer is characterized by being pancreatic cancer, liver cancer, breast cancer, lung cancer, stomach cancer, rectal cancer, gallbladder cancer, ovarian cancer, bladder cancer, colon cancer, lymphoma, brain cancer, uterine cancer, prostate cancer and malignant melanoma or biliary tract cancer. Pharmaceutical composition.
The method of claim 5,
The hydrophobic drugs include vinblastine, etoposide, Alkeran, Cytoxan, Daunorubicin, Hydrea, Mithramycin, Mitomycin ), Mitoxantrone, Nitrogen Mustard, Velban, Vincristine, Carboplatinum, Idarubicin, Irirubican, Irinotecan, Leustatin ( Leustatin), Navelbine (Navelbine), Taxotere (Taxotere), Topotecan (Topotecan) is characterized by adriamycin, daunomycin or paclitaxel, a pharmaceutical composition for the treatment of cancer.
The method of claim 5,
The hydrophilic drug is Busulfan, Chlorambucil, Cyclophosphamide, Melphalan, Cisplatin, Ifosfamide, Cytarabine, 5 -Fluorouracil (5-FU), methotrexate (Methotrexate, MTX), actinomycin-D (Actinomycin-D), characterized in that it contains bleomycin or gemcitabine (gemcitabine), the drug for treating cancer Composition.
i) a core portion comprising poly lactic acid (PLA) and a hydrophobic material as an aliphatic polyester, and
ii) a polymer surrounding the core portion, consisting of a Pluronic F127 and a shell portion comprising a hydrophilic material,
It characterized in that the weight ratio of the aliphatic polyester in the core portion and the polymer in the shell portion is 1:10 to 1:20,
A cosmetic composition comprising a temperature sensitive nanosponge platform for simultaneous delivery of hydrophilic and hydrophobic materials.
The method of claim 9,
The cosmetic composition is characterized in that it comprises a wrinkle improving agent, skin aging inhibitor or skin whitening agent, cosmetic composition.
The method of claim 9,
The hydrophilic material is characterized in that it comprises arbutin, adenosine, vitamin C, vitamin B3, vitamin B5, vitamin H, acetylglucosamine, madecassoside or selenium aspartate.
The method of claim 9,
The hydrophobic substance is eugenol, retinol, astaxanthin, caffeine, carnosine acid, catechin, coenzyme-Q10, curcumin, elazinic acid, peric acid, idebenone, isoflavone, linoleic acid, lipoic acid, lycopene, oleinoic acid, Containing floretine, quercetin, resveratrol, squalane, squalene, tannic acid, vitamin A, vitamin B, vitamin E, vitamin F, cholesterol, phytosphingosine, glycospingolide, beta-sitosterol, lauric acid or lecithin It is characterized in that the cosmetic composition.

Images