Lipid nanoparticle composition comprising sodium DNA

Abstract

The present invention relates to a lipid nanoparticle composition comprising sodium DNA, and more particularly, to a lipid nanoparticle composition comprising: sodium DNA as an active ingredient; the sodium bis (lauramide glutamine) lysine is used as an amphiphilic surfactant; the distearoyl phosphatidylcholine is used as an auxiliary surfactant; cholesterol as a stabilizer; the auxiliary stabilizer is selected from the group consisting of PEG-120 stearate, polyglycerol-4 decanoate, a poly (ethylene glycol)-poly (epsilon-caprolactone) copolymer (PEG-PCL copolymer) and a mixture of the PEG-120 stearate, the polyglycerol-4 decanoate and the poly (ethylene glycol)-poly (epsilon-caprolactone) copolymer; and a solvent.

Description

Technical Field

[0001] The present invention relates to a lipid nanoparticle composition comprising sodium DNA, and more particularly, to a lipid nanoparticle composition comprising the following components: sodium DNA as an active ingredient; sodium dilauramidoglutamide lysine as an amphiphilic surfactant; distearoylphosphatidylcholine as an auxiliary surfactant; cholesterol as a stabilizer; an auxiliary stabilizer selected from PEG-120 stearate, polyglyceryl-4 caprate, poly(ethylene glycol)-poly(ε-caprolactone) copolymer (PEG-PCL copolymer) and mixtures thereof; and a solvent. Background Art

[0002] Lipid nanoparticles (LNPs) are particulate nanocarriers composed of a lipid membrane that can encapsulate insoluble or unstable substances. Lipid nanoparticles have attracted attention as an attractive delivery system due to their ease of preparation, reduced immune response, multi-capacity functionality (high encapsulation efficiency), larger payload, and design flexibility.

[0003] For example, messenger ribonucleic acid (mRNA) for expressing therapeutic proteins can be used to treat a variety of diseases. The therapeutic uses of mRNA are as follows: (1) protein replacement, which restores the function of a single protein in rare monogenic diseases; (2) cell reprogramming, in which mRNA can regulate cell behavior by expressing transcription or growth factors; (3) immunotherapy, in which mRNA-encoded transcripts trigger specific immune responses against target cells such as therapeutic antibodies, all of which are often hampered by inefficient nucleic acid delivery. Lipid nanoparticles have been widely studied as carriers of such mRNA. Lipid nanoparticles have many advantages over previous lipid-based delivery systems, including: i) efficient nucleic acid encapsulation and efficient transfection, ii) improved tissue permeability for delivery of therapeutic agents, and iii) low cytotoxicity and immunogenicity.

[0004] In addition, sodium deoxyribonucleic acid (DNA) is the sodium salt of DNA, which is the sodium salt of DNA extracted and refined from salmon, commonly known as polydeoxyribonucleotide (PDRN). Sodium DNA is a tissue regeneration substance similar to human DNA, with high stability, present in cells, and has the effect of stimulating physiological regeneration and metabolic activities. Sodium DNA helps to promote growth factors, so it can be expected to have effects such as skin regeneration and relieving inflammation, so it has attracted much attention as a functional cosmetic raw material. As an example of the use of this sodium DNA, Korean Patent Application Publication No. 10-2020-0066420 discloses a skin anti-aging cosmetic composition using sodium DNA and astaxanthin as active ingredients.

[0005] [Prior art literature]

[0006] [Patent Document]

[0007] Korean Patent Application Publication No. 10-2020-0066420 Summary of the invention

[0008] Technical issues to be solved

[0009] The technical problem of the present invention is to provide a novel transdermal delivery system, which can enable sodium DNA as an active substance to penetrate through the skin efficiently in a state of improved stability, thereby improving its bioavailability.

[0010] Technical Solution

[0011] In order to solve the above technical problems, the present invention provides a lipid nanoparticle composition, which comprises: sodium DNA as an active ingredient; sodium di(laurylamideglutamine)lysine as an amphiphilic surfactant; distearoylphosphatidylcholine as an auxiliary surfactant; cholesterol as a stabilizer; an auxiliary stabilizer selected from PEG-120 stearate, polyglycerol-4 caprate, poly(ethylene glycol)-poly(ε-caprolactone) copolymer (PEG-PCL copolymer) and a mixture thereof; and a solvent.

[0012] In addition, the present invention provides a cosmetic composition comprising the lipid nanoparticle composition.

[0013] Hereinafter, the present invention will be described in more detail.

[0014] According to one aspect of the present invention, a lipid nanoparticle composition is provided, which comprises: sodium DNA as an active ingredient; sodium di(laurylamideglutamine)lysine as an amphiphilic surfactant; distearoylphosphatidylcholine as an auxiliary surfactant; cholesterol as a stabilizer; an auxiliary stabilizer selected from PEG-120 stearate, polyglycerol-4 caprate, poly(ethylene glycol)-poly(ε-caprolactone) copolymer (PEG-PCL copolymer) and a mixture thereof; and a solvent.

[0015] The lipid nanoparticle composition of the present invention comprises sodium DNA (sodium deoxyribonucleic acid) as an active ingredient. In a specific embodiment according to the present invention, the lipid nanoparticle composition may comprise sodium DNA as an active ingredient at a content of 0.005-1% by weight, 0.01-0.5% by weight, or 0.04-0.1% by weight. In the present invention, when the content of sodium DNA is less than 0.005% by weight, the effect of the active ingredient may be weakened, and when the content of sodium DNA exceeds 1% by weight, the stability of the lipid nanoparticles may be problematic.

[0016] The lipid nanoparticle composition of the present invention comprises sodium di(laurylamide glutamine) lysine as an amphipathic surfactant. Sodium di(laurylamide glutamine) lysine is an amphipathic surfactant having a tripeptide structure of glutamic acid-lysine-glutamic acid. In addition, the sodium di(laurylamide glutamine) lysine is a gemini form having more than two hydrophilic groups and more than two hydrophobic groups, has high solubility in water, and has a low critical micelle concentration (CMC) and a high ability to reduce surface tension. In another embodiment according to the present invention, the lipid nanoparticle composition may comprise sodium di(laurylamide glutamine) lysine as an amphipathic surfactant in an amount of 0.001-2% by weight, 0.005-1% by weight, or 0.01-0.2% by weight. In the present invention, when the content of sodium di(laurylamideglutamine)lysine is less than 0.001 wt %, the formation of lipid nanoparticles may be problematic, and when the content of sodium di(laurylamideglutamine)lysine exceeds 2 wt %, the stability of lipid nanoparticles may be problematic.

[0017] The lipid nanoparticle composition of the present invention comprises distearoylphosphatidylcholine (1,2-distearoylphosphatidylcholine) as an auxiliary surfactant. s-Glycerol-3-phosphocholine (1,2-distearoyl- s -glycero-3-phosphocholine), DSPC). Distearoylphosphatidylcholine is a phosphatidylcholine as a kind of phospholipid, which is a natural component of cell membrane. In another specific embodiment according to the present invention, the lipid nanoparticle composition may contain distearoylphosphatidylcholine as an auxiliary surfactant in an amount of 0.0001-1% by weight, 0.0005-0.5% by weight or 0.001-0.1% by weight. In the present invention, when the content of distearoylphosphatidylcholine is less than 0.0001% by weight, there may be problems with the formation of lipid nanoparticles, and when the content of distearoylphosphatidylcholine exceeds 1% by weight, there may be problems with the stability of lipid nanoparticles.

[0018] The lipid nanoparticle composition of the present invention comprises cholesterol as a stabilizer. In the present invention, cholesterol plays a role in enhancing the stability of the lipid membrane structure. In another specific embodiment according to the present invention, the lipid nanoparticle composition may comprise cholesterol as a stabilizer in an amount of 0.001-2% by weight, 0.005-1% by weight or 0.01-0.2% by weight. In the present invention, when the content of cholesterol is less than 0.001% by weight, the stabilizing effect brought by cholesterol may become weak, and when the content of cholesterol exceeds 2% by weight, there may be problems with the formation of lipid nanoparticles.

[0019] The auxiliary stabilizer included in the lipid nanoparticle composition of the present invention is selected from PEG-120 stearate, polyglycerol-4 caprate, poly(ethylene glycol)-poly(ε-caprolactone) copolymer (PEG-PCL copolymer) and mixtures thereof. PEG-120 stearate is a polyethylene glycol ester (ester) form of glycerol stearate. Polyglycerol-4 caprate is an ester form of capric acid and polyglycerol-4. Poly(ethylene glycol)-poly(ε-caprolactone) copolymer (PEG-PCL copolymer) is an amphiphilic block copolymer that can be prepared by polymerizing polyethylene glycol and polycaprolactone and is biodegradable. In another specific embodiment according to the present invention, the auxiliary stabilizer can be a mixture of PEG-120 stearate and polyglycerol-4 caprate. In another specific embodiment according to the present invention, the lipid nanoparticle composition can contain an auxiliary stabilizer in an amount of 0.0001-1% by weight, 0.0003-0.5% by weight or 0.0005-0.1% by weight. In the present invention, when the content of the auxiliary stabilizer is less than 0.0001 wt % or exceeds 1 wt %, the formation of lipid nanoparticles may be problematic.

[0020] The lipid nanoparticle composition of the present invention comprises a solvent. In another specific embodiment according to the present invention, the solvent may be selected from alcohol, water and mixtures thereof. In another specific embodiment according to the present invention, the solvent may be a mixture of alcohol and water. In another specific embodiment according to the present invention, the alcohol may be ethanol. In another specific embodiment according to the present invention, the lipid nanoparticle composition may comprise a solvent in an amount of 93-99.99% by weight, 96.5-99.95% by weight or 99.3-99.9% by weight. In the present invention, when the content of the solvent is less than 93% by weight or exceeds 99.99% by weight, there may be problems with the formation of lipid nanoparticles.

[0021] In another specific embodiment according to the present invention, the lipid nanoparticle composition can be prepared by thin-film hydration, reverse phase evaporation, emulsion or microfluidics. In another specific embodiment according to the present invention, the lipid nanoparticle composition can be prepared by microfluidics. Microfluidics can use organic solvents such as ethanol and phospholipids in microfluidic chips to prepare lipid-based nanocarriers. Microfluidics can control particle size and the surface of the membrane by adjusting production process conditions such as the speed of the fluid, the type and content of the lipid. Microfluidics can be carried out in a continuous process and can have the advantage of high reproducibility.

[0022] According to another aspect of the present invention, a cosmetic composition comprising the lipid nanoparticle composition is provided. In another specific embodiment of the present invention, the cosmetic composition may contain the lipid nanoparticle composition at a content of 1-50% by weight. In the present invention, when the cosmetic composition contains less than 1% by weight of the lipid nanoparticle composition, the effect of DNA sodium as an active ingredient may be minimal. If the content of the lipid nanoparticle composition exceeds 50% by weight, it may be economically undesirable because the effect of DNA sodium cannot be expected to increase in proportion to its addition amount.

[0023] Beneficial Effects

[0024] According to the present invention, by configuring sodium DNA as an active ingredient into lipid nanoparticles, sodium DNA can be efficiently delivered into the skin in a very stable state, thereby showing high bioavailability and also providing excellent effects in skin regeneration and relief of inflammation. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1aTo take a photo of the appearance of the lipid nanoparticles prepared in Example 1, Figure 1b This is a photograph taken to show the appearance of the lipid nanoparticles prepared in Example 2.

[0026] Figure 2a To measure the particle distribution of the lipid nanoparticles prepared in Example 1, Figure 2b The results of measuring the particle distribution of the lipid nanoparticles prepared in Example 2 are shown in FIG.

[0027] Figure 3a The results of measuring the zeta potential of the lipid nanoparticles prepared in Example 1 are as follows: Figure 3b The results of measuring the ζ-potential of the lipid nanoparticles prepared in Example 2 are shown in FIG.

[0028] Figure 4a To measure the results of Fourier transform infrared spectroscopy (FT-IR) of the lipid nanoparticles prepared in Example 1, Figure 4b The results of Fourier transform infrared spectroscopy (FT-IR) of the lipid nanoparticles prepared in Example 2 are shown.

[0029] Figure 5a The results of measuring the high temperature stability of samples L1W3, L1W4, L1W5 and L1W6 of Example 1 using Turbiscan are as follows: Figure 5b The results are obtained by measuring the high temperature stability of samples L1W4(D), L1W5(D), L1W6(D) and L1W7(D) of Example 2 using Turbiscan.

[0030] Figure 6 The results of measuring the ultraviolet absorption spectrum of the lipid nanoparticles prepared in Example 2.

[0031] Figure 7 The lipid nanoparticles prepared in Example 2 were magnified and photographed using a cryo-electron microscope (freeze-fracture scanning electron microscope ×12000). DETAILED DESCRIPTION

[0032] Example 1: Preparation of lipid nanoparticles without active substances (L1W1 to L1W10)

[0033] First, prepare lipid solution and aqueous solution respectively.Use microfluidic device (Liposome Synthesis System, NEO NANOTECH, South Korea) to prepare lipid nanoparticles, which can control the speed (rpm) of the drive motor by the feedback function between the flow sensor and the drive motor.Prepared lipid solution and aqueous solution are placed in a microfluidic chip, and lipid nanoparticles of L1W1 to L1W10 are prepared by adjusting the flow rate to have the composition of Table 1 below.At this time, the flow rate can be controlled from a minimum of 0.0015mL / min to a maximum of 18mL / min.The appearance of the prepared lipid nanoparticles is photographed and shown in Figure 1a middle.

[0034] [Table 1] Preparation of lipid solution and aqueous solution

[0035]

[0036] Example 2: Preparation of Lipid Nanoparticles Containing Sodium DNA [L1W1(D) to L1W10(D)]

[0037] A lipid solution and an aqueous solution were prepared separately. Lipid nanoparticles were prepared using the same microfluidic device as in Example 1. In the microchip, lipid nanoparticles of L1W1(D) to L1W10(D) were prepared by adjusting the flow rate of each solution so as to have the composition shown in Table 2 below. The appearance of the prepared lipid nanoparticles was photographed and shown in Figure 1b middle.

[0038] [Table 2]

[0039]

[0040] Experimental Example 1: Measurement of Particle Distribution

[0041] In order to confirm the particle distribution of the lipid nanoparticles prepared in Example 1 and Example 2, the lipid nanoparticle solution was diluted by 10%, and then 10 mL was put into the ELS-Z cell of Photal (Photal Co., Japan) and measured. The results are shown in Figure 2a and Figure 2b In. From Figure 2a and Figure 2b It can be seen that the difference in composition leads to different particle distribution due to the adjustment of flow rate.

[0042] Experimental Example 2: Measurement of zeta potential

[0043] In order to confirm the surface charge of the lipid nanoparticles prepared in Example 1 and Example 2, the lipid nanoparticle solution was diluted by 10%, and then 1 mL was placed in a Photal ELS-Z (Photal, Japan) cell and the ζ-potential was measured. The results are shown in Figure 3a and Figure 3b In. From Figure 3a and Figure 3b It can be seen that due to the adjustment of the flow rate, the difference in composition leads to different surface charges of the particles.

[0044] Experimental Example 3: Measurement of Fourier-transform infrared (FT-IR) spectroscopy

[0045] In order to confirm the composition of the lipid nanoparticles prepared in Example 1 and Example 2, FT-IR was measured using a Nicolet Summit FTIR spectrometer (Thermo Fisher Scientific, USA), and the results are shown in Figure 4a and Figure 4b As the proportion of aqueous solution increases, it can be confirmed that the lipid-related peak decreases and the water-related peak increases.

[0046] Experimental Example 4: Turbiscan Measurement

[0047] In order to confirm the high temperature stability of the lipid nanoparticles prepared in Example 1, the particle size and ζ-potential values ​​of L1W3, L1W4, L1W5 and L1W6 samples were relatively stable, and the results were measured using Turbiscan LAB (Formulaction, France). Figure 5a The analysis conditions were 40°C and 50 scans within 5 hours. It can be confirmed that L1W4 in Example 1 is the most stable. The high temperature stability of samples L1W4 (D), L1W5 (D), L1W6 (D) and L1W7 (D) in the lipid nanoparticles prepared in Example 2 was measured and the results are shown in Figure 5b In Example 2, it was confirmed that L1W6(D) was the most stable.

[0048] Experimental Example 5: Measurement of Ultraviolet (UV) Absorption Spectrum

[0049] In order to confirm whether the lipid nanoparticles prepared in Example 2 contain DNA sodium, the absorption spectrum was measured using an ultraviolet spectrometer (UV Spectrometer) (Thermo Fisher Scientific, USA), and the results are shown in Figure 6As the proportion of aqueous solution increases, the generation of DNA sodium peak can be observed, and a peak shift occurs.

[0050] Experimental Example 6: Cryo-electron microscopy

[0051] The DNA sodium-lipid nanoparticles prepared in Example 2 were photographed at a magnification of ×12000 using a cryo-electron microscope (JEM 1010, JEOL, Japan). Figure 7 ).from Figure 7 It can be seen that the lipid nanoparticles are well formed and have uniform size.

Claims

1. A lipid nanoparticle composition comprising: Sodium DNA as the active ingredient; Sodium di(laurylamideglutamine)lysine as an amphiphilic surfactant; distearoylphosphatidylcholine as a cosurfactant; cholesterol as a stabilizer; an auxiliary stabilizer selected from PEG-120 stearate, polyglyceryl-4 caprate, poly(ethylene glycol)-poly(ε-caprolactone) copolymer (PEG-PCL copolymer), and mixtures thereof; and Solvent.

2. The lipid nanoparticle composition according to claim 1, characterized in that The lipid nanoparticle composition comprises: 0.005-1 wt % of sodium DNA, 0.001-2 wt % of sodium di(laurylamideglutamine)lysine, 0.0001-1 wt % of distearoylphosphatidylcholine, 0.001-2 wt % of cholesterol, 0.0001-1 wt % of an auxiliary stabilizer and 93-99.99 wt % of a solvent.

3. The lipid nanoparticle composition according to claim 2, characterized in that The lipid nanoparticle composition comprises 0.01-0.5 wt % of sodium DNA, 0.005-1 wt % of sodium di(laurylamideglutamine)lysine, 0.0005-0.5 wt % of distearoylphosphatidylcholine, 0.005-1 wt % of cholesterol, 0.0003-0.5 wt % of an auxiliary stabilizer and 96.5-99.95 wt % of a solvent.

4. The lipid nanoparticle composition according to claim 3, characterized in that The lipid nanoparticle composition comprises: 0.04-0.1 wt % of sodium DNA, 0.01-0.2 wt % of sodium di(laurylamideglutamine)lysine, 0.001-0.1 wt % of distearoylphosphatidylcholine, 0.01-0.2 wt % of cholesterol, 0.0005-0.1 wt % of an auxiliary stabilizer and 99.3-99.9 wt % of a solvent.

5. The lipid nanoparticle composition according to claim 1, characterized in that The lipid nanoparticle composition is prepared by a thin film hydration method, a reverse phase evaporation method, an emulsion method or a microfluidics method.

6. The lipid nanoparticle composition according to claim 5, characterized in that The lipid nanoparticle composition is prepared by a microfluidics method.

7. The lipid nanoparticle composition according to claim 1, characterized in that The auxiliary stabilizer is a mixture of PEG-120 stearate and polyglyceryl-4 caprate.

8. The lipid nanoparticle composition according to claim 1, characterized in that The solvent is selected from alcohol, water and a mixture thereof.

9. The lipid nanoparticle composition according to claim 8, characterized in that The alcohol is ethanol.

10. A cosmetic composition comprising the lipid nanoparticle composition according to any one of claims 1 to 9.

11. The cosmetic composition according to claim 10, characterized in that: The content of the lipid nanoparticle composition is 1-50% by weight.

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