Lipid nanoparticle composition containing sodium DNA

Abstract

The present invention relates to a lipid nanoparticle composition comprising sodium DNA, and more specifically, to a lipid nanoparticle composition comprising sodium DNA as an active ingredient; sodium dilauramidoglutamide lysine as an amphiphilic surfactant; distearoylphosphatidylcholine as a co-surfactant; cholesterol as a stabilizer; a co-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.

Description

[Technical Field]

[0001] The present invention relates to a lipid nanoparticle composition comprising sodium DNA, and more specifically, to a lipid nanoparticle composition comprising sodium DNA as an active ingredient; sodium dilauramidoglutamide lysine as an amphiphilic surfactant; distearoylphosphatidylcholine as a co-surfactant; cholesterol as a stabilizer; a co-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 technology]

[0002] Lipid nanoparticles (LNPs) are particulate nanodeliveries composed of lipid membranes that can encapsulate poorly soluble or unstable substances. Lipid nanoparticles have attracted attention as an attractive delivery system due to their ease of fabrication, reduced immune response, high encapsulation efficiency, large payload capacity, and design flexibility.

[0003] For example, the use of messenger ribonucleic acid (mRNA) to express therapeutic proteins has the potential to treat a variety of diseases. Therapeutic applications of mRNA include: (1) protein replacement to restore the function of a single protein in rare monogenic diseases; (2) cellular reprogramming, in which mRNA modulates cellular behavior by expressing transcription or growth factors; and (3) immunotherapy. While transcripts encoded by mRNA trigger specific immune responses against target cells, such as therapeutic antibodies, these responses are often hindered by inefficient nucleic acid delivery. The potential of lipid nanoparticles as delivery vehicles for such mRNA has been extensively studied. Compared to conventional lipid-based delivery systems, lipid nanoparticles offer many advantages, including i) highly efficient nucleic acid encapsulation and efficient transfection, ii) improved tissue penetration for therapeutic drug delivery, and iii) low cytotoxicity and immunogenicity.

[0004] Meanwhile, DNA sodium (deoxyribonucleic acid) is the sodium salt of DNA. DNA sodium, commonly called polydeoxyribonucleotide (PDRN), is a sodium salt of DNA extracted and purified from salmon gonad tissue. DNA sodium is a tissue regeneration substance similar to human DNA, highly stable, and presents within cells, stimulating physiological regeneration and metabolic activity. Because DNA sodium contributes to promoting growth factors, it is expected to have effects such as skin regeneration and inflammation relief, and is attracting attention as a functional cosmetic ingredient. Patent Document 1 discloses an example of the use of DNA sodium, namely, a skin anti-aging cosmetic composition containing DNA sodium and astaxanthin as active ingredients. [Prior art documents] [Patent documents]

[0005] [Patent Document 1] Korean Patent Application Publication No. 10-2020-0066420 Summary of the Invention [Problem to be solved by the invention]

[0006] The technical objective of the present invention is to provide a novel transdermal delivery system that can efficiently permeate sodium DNA as an active ingredient through the skin while improving its stability, thereby increasing its bioavailability. [Means for solving the problem]

[0007] In order to solve the above technical problems, the present invention provides a lipid nanoparticle composition comprising sodium DNA as an active ingredient; sodium dilauramidoglutamide lysine as an amphiphilic surfactant; distearoylphosphatidylcholine as a co-surfactant; cholesterol as a stabilizer; a co-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.

[0008] The present invention also provides a cosmetic composition comprising the lipid nanoparticle composition. [Effects of the Invention]

[0009] According to the present invention, by formulating the active ingredient, DNA sodium, into lipid nanoparticles, DNA sodium can be efficiently delivered into the skin in a very stable state, thereby exhibiting high bioavailability and excellent effects on skin regeneration and inflammation relief. [Brief explanation of the drawings]

[0010] [Figure 1A] 1 is a photograph showing the properties of lipid nanoparticles produced in Example 1. [Figure 1B] 1 is a photograph showing the properties of lipid nanoparticles produced in Example 2. [Figure 2A] 1 shows the results of measuring the particle distribution of the lipid nanoparticles produced in Example 1. [Figure 2B]1 shows the results of measuring the particle distribution of the lipid nanoparticles produced in Example 2. [Figure 3A] 1 shows the results of measuring the zeta potential of the lipid nanoparticles produced in Example 1. [Figure 3B] 1 shows the results of measuring the zeta potential of the lipid nanoparticles produced in Example 2. [Figure 4A] 1 shows the results of measuring the Fourier transform infrared spectroscopy (FT-IR) spectrum of the lipid nanoparticles produced in Example 1. [Figure 4B] 1 shows the results of measuring the Fourier transform infrared spectroscopy (FT-IR) spectrum of the lipid nanoparticles produced in Example 2. [Figure 5A] 1 shows the results of measuring the high temperature stability of samples L1W3, L1W4, L1W5, and L1W6 of Example 1 using Turbiscan. [Figure 5B] 1 shows the results of measuring the high temperature stability of samples L1W4(D), L1W5(D), L1W6(D), and L1W7(D) of Example 2 using Turbiscan. [Figure 6] 1 shows the results of measuring the ultraviolet absorption spectrum of the lipid nanoparticles produced in Example 2. [Figure 7] 1 is a magnified photograph (12,000x magnification) of the lipid nanoparticles produced in Example 2 taken by freeze-fracture scanning electron microscopy. DETAILED DESCRIPTION OF THE INVENTION

[0011] The present invention will be described in detail below. According to one aspect of the present invention, there is provided a lipid nanoparticle composition comprising sodium DNA as an active ingredient; sodium dilauramidoglutamide lysine as an amphiphilic surfactant; distearoylphosphatidylcholine as a co-surfactant; cholesterol as a stabilizer; a co-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.

[0012] The lipid nanoparticle composition of the present invention contains sodium deoxyribonucleic acid (DNA) as an active ingredient. In one embodiment of the present invention, the lipid nanoparticle composition may contain sodium DNA as an active ingredient in an amount of 0.005 to 1 wt %, 0.01 to 0.5 wt %, or 0.04 to 0.1 wt %. In the present invention, if the content of sodium DNA is less than 0.005 wt %, the effect of the active ingredient may be weakened, and if it exceeds 1 wt %, problems may arise with the stability of the lipid nanoparticles.

[0013] The lipid nanoparticle composition of the present invention contains sodium dilauramidoglutamide lysine as an amphiphilic surfactant. Sodium dilauramidoglutamide lysine is an amphiphilic surfactant with a tripeptide structure of glutamic acid-lysine-glutamic acid. Furthermore, sodium dilauramidoglutamide lysine is a gemini surfactant having two or more hydrophilic groups and two or more hydrophobic groups, and has high solubility in water, a low CMC (critical micelle concentration), and a high ability to reduce surface tension. In another embodiment of the present invention, the lipid nanoparticle composition may contain sodium dilauramidoglutamide lysine as an amphiphilic surfactant in an amount of 0.001 to 2 wt %, 0.005 to 1 wt %, or 0.01 to 0.2 wt %. In the present invention, if the content of dilauramidoglutamide lysine sodium is less than 0.001% by weight, problems may arise in the formation of lipid nanoparticles, and if it exceeds 2% by weight, problems may arise in the stability of the lipid nanoparticles.

[0014] The lipid nanoparticle composition of the present invention contains distearoylphosphatidylcholine (1,2-distearoyl-sn-glycero-3-phosphocholine, DSPC) as a co-surfactant. Distearoylphosphatidylcholine is a phosphatidylcholine, a type of phospholipid, and is a natural component of cell membranes. In another embodiment of the present invention, the lipid nanoparticle composition may contain distearoylphosphatidylcholine as a co-surfactant in an amount of 0.0001 to 1 wt %, 0.0005 to 0.5 wt %, or 0.001 to 0.1 wt %. In the present invention, if the content of distearoylphosphatidylcholine is less than 0.0001 wt %, problems may occur in the formation of lipid nanoparticles, while if it exceeds 1 wt %, problems may occur in the stability of the lipid nanoparticles.

[0015] The lipid nanoparticle composition of the present invention contains cholesterol as a stabilizer. In the present invention, cholesterol plays a role in enhancing the stability of the lipid membrane structure. In another embodiment according to the present invention, the lipid nanoparticle composition may contain cholesterol as a stabilizer in an amount of 0.001 to 2 wt %, 0.005 to 1 wt %, or 0.01 to 0.2 wt %. In the present invention, if the cholesterol content is less than 0.001 wt %, the stabilizing effect of cholesterol may be weak, and if it exceeds 2 wt %, problems may occur in the formation of lipid nanoparticles.

[0016] The lipid nanoparticle composition of the present invention contains a co-stabilizer selected from PEG-120 stearate, polyglyceryl-4 caprate, poly(ethylene glycol)-poly(ε-caprolactone) copolymer (PEG-PCL copolymer), and mixtures thereof. PEG-120 stearate is a polyethylene glycol ester of glyceryl stearate. Polyglyceryl-4 caprate is an ester of capric acid and polyglycerin-4. Poly(ethylene glycol)-poly(ε-caprolactone) copolymer (PEG-PCL copolymer) is an amphiphilic block copolymer obtained by polymerizing polyethylene glycol and polycaprolactone, and is biodegradable. In another embodiment of the present invention, the co-stabilizer may be a mixture of PEG-120 stearate and polyglyceryl-4 caprate. In another embodiment of the present invention, the lipid nanoparticle composition may contain the co-stabilizer in an amount of 0.0001 to 1 wt %, 0.0003 to 0.5 wt %, or 0.0005 to 0.1 wt %. In the present invention, if the content of the auxiliary stabilizer is less than 0.0001% by weight or more than 1% by weight, problems may arise in the formation of lipid nanoparticles.

[0017] The lipid nanoparticle composition of the present invention contains a solvent. In another embodiment of the present invention, the solvent can be selected from alcohol, water, and a mixture thereof. In another embodiment of the present invention, the solvent may be a mixture of alcohol and water. In another embodiment of the present invention, the alcohol may be ethanol. In another embodiment of the present invention, the lipid nanoparticle composition may contain the solvent in an amount of 93 to 99.99 wt %, 96.5 to 99.95 wt %, or 99.3 to 99.9 wt %. In the present invention, if the solvent content is less than 93 wt % or more than 99.99 wt %, problems may occur in the formation of lipid nanoparticles.

[0018] In another embodiment of the present invention, the lipid nanoparticle composition can be produced by thin film hydration, reverse phase evaporation, emulsion, or microfluidics. In another embodiment of the present invention, the lipid nanoparticle composition may be produced by a microfluidics method. The microfluidics method can produce lipid-based nanodeliveries using an organic solvent such as ethanol and phospholipids in a microfluidic chip. The microfluidics method may control particle size and membrane surface by adjusting production process conditions such as fluid velocity, lipid type, and lipid content. The microfluidics method has the advantage of being a continuous process and highly reproducible.

[0019] Another aspect of the present invention provides a cosmetic composition containing the lipid nanoparticle composition. In another embodiment of the present invention, the cosmetic composition may contain the lipid nanoparticle composition in an amount of 1 to 50 wt %. In the present invention, if the lipid nanoparticle composition is contained in the cosmetic composition at less than 1 wt %, the effect of the active ingredient, sodium DNA, may be weak. If the lipid nanoparticle composition is contained in the cosmetic composition at more than 50 wt %, the effect of the active ingredient cannot be expected to increase commensurate with the amount added, which may be economically undesirable. [Example]

[0020] Example 1: Preparation of lipid nanoparticles containing no active ingredient (L1W1 to L1W10) First, lipid solutions and aqueous solutions were prepared separately. Lipid nanoparticles were produced using a microfluidic device (liposome synthesis system, NEO NANOTECH, Korea), which controls the rotation speed of the drive motor via a feedback function between the flow sensor and the drive motor. The lipid solutions and aqueous solutions prepared above were fed into a microfluidic chip with the compositions shown in Table 1 below, and the flow rates were adjusted to produce lipid nanoparticles L1W1 to L1W10. The flow rates were controllable from a minimum of 0.0015 mL / min to a maximum of 18 mL / min. The appearance of the produced lipid nanoparticles was photographed and shown in Figure 1A.

[0021] [Table 1]

[0022] Example 2: Preparation of lipid nanoparticles containing sodium DNA [L1W1(D) to L1W10(D)] Lipid solutions and aqueous solutions were prepared separately. Lipid nanoparticles were prepared using the same microfluidic device as in Example 1. The flow rates of each solution were adjusted within the microchip to prepare lipid nanoparticles L1W1(D) to L1W10(D) with the compositions shown in Table 2 below. Photographs of the appearance of the prepared lipid nanoparticles were taken and are shown in Figure 1B.

[0023] [Table 2]

[0024] Experimental Example 1: Measurement of particle size distribution To confirm the particle size distribution of the lipid nanoparticles produced in Examples 1 and 2, the lipid nanoparticle solution was diluted to 10% and measured in a 10 mL Photal ELS-Z (Photal, Japan) cell. The results are shown in Figures 2A and 2B. As shown in Figures 2A and 2B, the particle size distribution differs depending on the composition due to flow rate adjustment.

[0025] Experimental Example 2: Zeta Potential Measurement To confirm the surface charge of the lipid nanoparticles prepared in Examples 1 and 2, the lipid nanoparticle solution was diluted to 10% and placed in a 1 mL Photal ELS-Z (Photal, Japan) cell to measure the zeta potential. The results are shown in Figures 3A and 3B. As shown in Figures 3A and 3B, the surface charge of the particles differs depending on the composition obtained by adjusting the flow rate.

[0026] Experimental Example 3: Fourier Transform Infrared Spectroscopy (FT-IR) To confirm the composition of the lipid nanoparticles prepared in Examples 1 and 2, FT-IR was measured using a Nicolet Summit FTIR spectrometer (ThermoFisherScientific, USA). The results are shown in Figures 4A and 4B. It was confirmed that as the ratio of aqueous solution increased, the lipid-related peak decreased and the aqueous solution-related peak increased.

[0027] Experimental example 4: Turbiscan measurement Among the lipid nanoparticles prepared in Example 1, the L1W3, L1W4, L1W5, and L1W6 samples, which had relatively stable particle sizes and zeta potentials, were measured for their high-temperature stability using a TurbiscanLAB (Formulaction, France). The results are shown in Figure 5A. The analysis conditions were a temperature of 40°C, 5 hours, and 50 scans. In Example 1, L1W4 was confirmed to be the most stable. Among the lipid nanoparticles prepared in Example 2, the L1W4(D), L1W5(D), L1W6(D), and L1W7(D) samples were measured for high-temperature stability, and the results are shown in Figure 5B. In Example 2, L1W6(D) was confirmed to be the most stable.

[0028] Experimental Example 5: Measurement of UV absorption spectrum To confirm whether the lipid nanoparticles prepared in Example 2 contained sodium DNA, the absorption spectrum was measured using a UV spectrometer (ThermoFisherScientific, USA), and the results are shown in Figure 6. As the aqueous solution ratio increased, a peak corresponding to sodium DNA was generated, and a peak shift was observed.

[0029] Experimental Example 6: Cryo-electron microscopy The sodium DNA-lipid nanoparticles prepared in Example 2 were photographed at 12,000x magnification using a cryo-electron microscope (JEM1010, JEOL, Japan) (Figure 7). Figure 7 shows that the lipid nanoparticles were well formed and of uniform size.

Claims

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

2. The lipid nanoparticle composition of claim 1, comprising 0.005 to 1% by weight of sodium DNA, 0.001 to 2% by weight of sodium dilauramidoglutamide lysine, 0.0001 to 1% by weight of distearoylphosphatidylcholine, 0.001 to 2% by weight of cholesterol, 0.0001 to 1% by weight of a co-stabilizer, and 93 to 99.99% by weight of a solvent.

3. The lipid nanoparticle composition according to claim 2, characterized in that it comprises 0.01 to 0.5% by weight of sodium DNA, 0.005 to 1% by weight of sodium dilauramidoglutamide lysine, 0.0005 to 0.5% by weight of distearoylphosphatidylcholine, 0.005 to 1% by weight of cholesterol, 0.0003 to 0.5% by weight of a co-stabilizer, and 96.5 to 99.95% by weight of a solvent.

4. The lipid nanoparticle composition according to claim 3, comprising 0.04 to 0.1% by weight of sodium DNA, 0.01 to 0.2% by weight of sodium dilauramidoglutamide lysine, 0.001 to 0.1% by weight of distearoylphosphatidylcholine, 0.01 to 0.2% by weight of cholesterol, 0.0005 to 0.1% by weight of a co-stabilizer, and 99.3 to 99.9% by weight of a solvent.

5. The lipid nanoparticle composition of claim 1, which is produced by thin film hydration, reverse phase evaporation, emulsion, or microfluidics.

6. The lipid nanoparticle composition of claim 5, which is produced by a microfluidic method.

7. The lipid nanoparticle composition of claim 1, wherein the auxiliary stabilizer is a mixture of PEG-120 stearate and polyglyceryl-4 caprate.

8. The lipid nanoparticle composition of claim 1, wherein the solvent is selected from alcohol, water, and mixtures thereof.

9. The lipid nanoparticle composition of claim 8, wherein 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 it contains 1 to 50 wt % of the lipid nanoparticle composition.

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