Peptide inhibiting formation of snare complex and cosmetic composition comprising same

Abstract

The present invention relates to: a peptide that exhibits a similar effect to Botox by binding to SNAP -25 or VAMP to inhibit the formation of a snare complex; and a cosmetic composition comprising same.

Description

Peptide that inhibits the formation of a SNARE complex and a cosmetic composition comprising the same

[0001] The present invention relates to a peptide that exhibits efficacy similar to botox by binding to SNAP-25 or VAMP and inhibiting the formation of the SNAP complex, and to a cosmetic composition comprising the same.

[0002] The SNARE complex, comprised of VAMP, Syntaxin, and SNAP-25, promotes the release of the neurotransmitter acetylcholine, which in turn causes muscle contraction and the formation of wrinkles. Specifically, acetylcholine repeatedly moves within muscles, exerting continuous pressure on the skin. This pressure alters collagen structure and destroys elastin, causing muscle cell contraction and the formation of wrinkles.

[0003] Accordingly, a technology is being widely developed to prevent the formation of the SNARE complex and suppress the secretion of acetylcholine to provide a wrinkle improvement effect. An example is botox described in the following patent document. The botox decomposes SNAP-25 to suppress the secretion of acetylcholine, thereby inhibiting muscle contraction and providing a temporary wrinkle improvement effect.

[0004] <Patent Document>

[0005] Patent Document: Patent No. 10-1506204 (registered on March 20, 2015) "Drug composition with enhanced botulinum toxin function, including fat-dissolving function."

[0006] However, the above botox has safety issues depending on the dosage, so it must be administered by a specialist, which causes inconvenience in use.

[0007] The present invention has been devised to solve the above problems.

[0008] The purpose of the present invention is to provide a peptide that temporarily provides a wrinkle improvement effect by reducing the physical binding of SNAP-25 and VAMP, thereby inhibiting the formation of SNAP complexes and thereby inhibiting acetylcholine secretion, and a cosmetic composition using the same.

[0009] In order to achieve the above-mentioned purpose, the present invention is implemented by an embodiment having the following configuration.

[0010] According to one embodiment of the present invention, the peptide according to the present invention is characterized in that it consists of any one of the amino acid sequences of SEQ ID NOs: 1 to 7.

[0011] According to another embodiment of the present invention, the peptide according to the present invention is characterized by inhibiting the formation of a SNARE complex.

[0012] According to another embodiment of the present invention, it is characterized by being used for improving wrinkles according to the present invention.

[0013] According to another embodiment of the present invention, the peptide according to the present invention is characterized in that it temporarily provides a wrinkle improvement effect by inhibiting the formation of SNAP complex by reducing the physical binding of SNAP-25 and VAMP, thereby inhibiting the secretion of acetylcholine.

[0014] According to another embodiment of the present invention, the cosmetic composition according to the present invention is characterized in that it comprises a peptide consisting of any one of the amino acid sequences of SEQ ID NOs: 1 to 7.

[0015] According to another embodiment of the present invention, a composition for improving skin wrinkles according to the present invention is characterized in that it comprises a peptide consisting of any one of the amino acid sequences of SEQ ID NOs: 1 to 7.

[0016] According to another embodiment of the present invention, the cosmetic composition according to the present invention is characterized in that it is used for improving skin wrinkles.

[0017] The present invention can obtain the following effects through the above-described examples.

[0018] The present invention can produce a temporary wrinkle improvement effect similar to botox by inhibiting the formation of a SNARE complex using a peptide consisting of any one of the amino acid sequences of SEQ ID NOs: 1 to 7.

[0019] Figure 1 is a graph showing the effect of a peptide according to one embodiment of the present invention on the physical binding of SNAP-25 and VAMP.

[0020] Hereinafter, a peptide that inhibits the formation of a SNARE complex according to the present invention and a cosmetic composition comprising the same will be described in detail with reference to the attached drawings. Unless otherwise defined, all terms in this specification have the same general meaning as those terms understood by a person skilled in the art to which the present invention pertains, and if there is a conflict with the meaning of a term used in this specification, the definitions used in this specification shall apply. In addition, detailed descriptions of well-known functions and structures that may unnecessarily obscure the gist of the present invention will be omitted. Throughout the specification, when a part is said to "include" a certain component, this does not mean that other components can be included, but rather that other components can be included, unless specifically stated otherwise.

[0021]

[0022] One embodiment of the present invention relates to a peptide that inhibits the formation of a SNARE complex, wherein the peptide is characterized by having an amino acid sequence of SEQ ID NO: 1 (Glu-Met-Gln), 2 (Ala-Glu-Lys-Asn-Leu), 3 (Glu-Lys-Asn), 4 (Lys-Asn-Leu), 5 (Asn-Leu-Lys-Met), 6 (Met-Ile-Ile-Leu) or 7 (Gly-Val-Ile). The peptide can provide a wrinkle improvement effect by inhibiting the formation of a SNARE complex. Specifically, the peptide can temporarily provide a wrinkle improvement effect by inhibiting the secretion of acetylcholine by reducing the physical binding between SNAP-25 and VAMP to inhibit the formation of the SNARE complex. The peptide can be used in a cosmetic composition, a therapeutic composition, etc.

[0023]

[0024] Another embodiment of the present invention relates to a cosmetic composition comprising a peptide comprising an amino acid sequence of any one of SEQ ID NOs: 1 to 7, wherein the cosmetic composition is capable of improving skin wrinkles. Various known ingredients that can provide formulation stability and additional skin condition improvement effects may be added to the cosmetic composition.

[0025]

[0026] Hereinafter, the present invention will be described in more detail through examples. However, these examples are provided solely to illustrate the present invention in more detail and are not intended to limit the scope of the present invention.

[0027]

[0028] <Example 1> Synthesis of peptides

[0029]

[0030] Peptides of sequence numbers 1 to 7 listed in Table 1 below were synthesized by the FMOC solid-phase method using an automated synthesizer (PeptrEx-R48, Peptron, Korea). The synthesized peptides were purified and analyzed by reverse-phase high-performance liquid chromatography (Prominence LC-20AB, Shimadzu, Japan) using a C18 analytical RP column (Shiseido capcell pak), and identified using a mass spectrometer (HP 1100 Series LC / MSD, Hewlett-Packard, USA).

[0031]

[0032]

[0033] <Example 2> Confirmation of whether the peptide of Example 1 reduces the physical binding between VAMP and SNAP-25

[0034]

[0035] 1. Botulinum toxin A inhibits muscle contraction by decomposing SNAP-25, a component of the SNARE complex, thereby inhibiting the secretion of acetylcholine, resulting in a temporary wrinkle improvement effect. In order to confirm whether the peptide of Example 1 has a similar function to Botox by inhibiting the binding of SNAP-25, a component protein of the SNARE complex, and VAMP, the effect on the physical binding of SNAP-25 and VAMP was evaluated using the in situ PLA (proximity ligation assay) method.

[0036] 2. Specifically, 4.5 × 10 in each well of a 24-well microplate 4SH-SY5Y cells (human neuroblastoma cells) were seeded in MEM serum medium and cultured for 24 hours in a 37℃, 5% CO2 incubator, and then treated with each peptide of Example 1 and incubated for 24 hours under the same conditions. The concentration of each peptide in the medium was 10 μM, and untreated cells were used as a control group. Afterwards, the cells in each well were washed with PBS and fixed by treating with 2% formaldehyde for 15 minutes, and then treated with 0.1% TritonX-100 for 5 minutes to increase antibody permeability into the cells. Anti-VAMP-1 / 2 / 3 polyclonal antibodies (Santa Cruz, USA) and anti-SNAP-25 monoclonal antibodies (Santa Cruz, USA) were added, and PLA probes were added using an in situ PLA kit (Sigma-Aldrich), followed by hybridization, ligation, amplification, and mounting steps.

[0037] 3. Afterwards, the luminescence signals (PLA signals) detected in each cell were measured using a fluorescence microscope (Logosbio / CelenaS, Korea) to quantify the physical interaction between SNAP-25 and VAMP antibodies, and the results are shown in Figure 1.

[0038] 4. Referring to Figure 1, it can be confirmed that when Botox-Pep 1 to 7 of Example 1 are treated, the physical binding of SNAP-25 and VAMP is reduced compared to the untreated control group. That is, Botox-Pep 1 to 7 help temporarily improve skin wrinkles by reducing the physical binding of SNAP-25 and VAMP and thereby inhibiting muscle contraction.

[0039]

[0040] <Example 3> Evaluation of the effect of the peptide of Example 1 on acetylcholine secretion in nerve cells

[0041] 1. When a SNARE complex is formed, acetylcholine, a neurotransmitter, is secreted and is exported to muscles, causing wrinkles. Therefore, in order to confirm whether the peptide of Example 1 inhibits the secretion of acetylcholine from nerve cells, the amount of acetylcholine secretion was measured using an acetylcholine assay kit.

[0042] 2. Specifically, SH-SY5Y cells were seeded at 1×10 per well. 6 After preparation, each peptide of Example 1 was treated and incubated under the same conditions for 24 hours. The concentration of each peptide in the medium was 10 μM, and untreated cells were used as a control group. After that, the SH-SY5Y cells were washed three times with PBS, and the cells were lysed using an Acetylcholine assay kit (abcam, USA), and then treated with a reaction solution containing a choline probe, and the amount of intracellular acetylcholine was measured using a microplate reader. The relative acetylcholine amount of the experimental group was calculated based on the amount of acetylcholine in the control group (100%) and is shown in Table 2.

[0043] 3. Looking at Table 2, it can be confirmed that the amount of intracellular acetylcholine is reduced when Botox-Pep 1 to 7 of Example 1 are treated, so it can be inferred that the peptide manufactured in Example 1 inhibits the formation of a SNARE complex that causes acetylcholine to be secreted, and it can also be inferred that since the secretion of acetylcholine is inhibited, it can provide a temporary wrinkle improvement effect.

[0044]

[0045]

[0046] <Example 4> Confirmation of the wrinkle improvement effect through skin patch test of the cosmetic composition containing the peptide of Example 1

[0047]

[0048] 1. A cosmetic composition was prepared by adding each of the peptides manufactured in Example 1 to a commercially available ampoule base at a concentration of 50 ppm.

[0049] 2. Nine adults in their 20s to 60s were instructed to apply the same amount of the above cosmetic composition to the wrinkle area under the eyes every morning and evening for two weeks. The degree of wrinkles under the eyes before and after use was evaluated using a skin diagnostic device (JNAUS, Korea). The results are shown in Table 3. As a control group, Argireline® peptide (Lubrizol, USA), known as a botulinum toxin component, was used.

[0050] 3. As shown in Table 3, it can be seen that the cosmetic compositions containing each of the peptides manufactured in Example 1 have an effect of improving wrinkles under the eyes. That is, it can be seen that Botox-Pep 1 to 7 can be used in cosmetic compositions to help temporarily improve wrinkles.

[0051]

[0052]

[0053] In the above, the applicant has described preferred embodiments of the present invention, but such embodiments are only examples of implementing the technical idea of ​​the present invention, and any change or modification that implements the technical idea of ​​the present invention should be interpreted as falling within the scope of the present invention.

Claims

1. A peptide consisting of any one of the amino acid sequences of sequence numbers 1 to 7.

2. In paragraph 1, The above peptide is a peptide characterized in that it inhibits the formation of a SNARE complex.

3. In paragraph 1, The above peptide is characterized as being used for improving wrinkles.

4. In paragraph 1, The above peptide is characterized in that it temporarily provides a wrinkle improvement effect by inhibiting the formation of SNAP complex by reducing the physical binding of SNAP-25 and VAMP, thereby inhibiting the secretion of acetylcholine.

5. A cosmetic composition characterized by comprising a peptide composed of any one of the amino acid sequences of sequence numbers 1 to 7.

6. A composition for improving skin wrinkles, characterized in that it comprises a peptide consisting of any one of the amino acid sequences of sequence numbers 1 to 7.

7. In paragraph 5, A cosmetic composition characterized in that the above cosmetic composition is used for improving skin wrinkles.

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