Cell-penetrating peptide, conjugate containing same, and composition containing same

Abstract

To provide a cell-penetrating peptide.SOLUTION: The present invention employs a conjugate of a cell-penetrating carrier peptide and an active ingredient, the carrier peptide comprising a specific amino acid sequence.SELECTED DRAWING: None

Description

[Technical field]

[0001] The present invention relates to a cell-penetrating peptide derived from telomerase, a conjugate of the cell-penetrating peptide with an active ingredient, and a composition containing the conjugate. [Background technology]

[0002] Although low molecular weight substances, nucleic acids, proteins, nanoparticles, etc. have great potential as molecular level therapeutic substances, their use is limited due to their resistance to tissue penetration and cell membrane penetration. The development of a system for transporting the substances into cells has become an issue in molecular therapy. In the case of low molecular weight substances, nucleic acids, and nanoparticles, intracellular transport has been made possible by various reagents, electroporation, heat shock, etc., but in the case of proteins, transporting them into cells while maintaining their activity is a very difficult problem, and no clue has been found to solve the problem. Meanwhile, in the 1980s, in the process of research into the penetration of HIV cell membranes, it was revealed that the HIV-TAT protein, which consists of a specific 11 amino acid sequence, plays an important role in the process of delivery into cells, and since the 1990s, research into intracellular delivery of proteins has been actively conducted.

[0003] Telomeres are genetic material that are repeated at the ends of chromosomes and are known to prevent damage to the chromosome or binding to other chromosomes. Each time a cell divides, the length of the telomere shortens slightly, but after a certain number of cell divisions, the telomere becomes very short and the cell stops dividing and dies. On the other hand, it is known that lengthening the telomere can extend the lifespan of a cell. For example, in cancer cells, an enzyme called telomerase is secreted to prevent the telomeres from shortening, allowing the cancer cells to continue to grow without dying. Summary of the Invention [Problem to be solved by the invention]

[0004] One aspect of the present invention provides novel peptides.

[0005] One aspect of the present invention provides polynucleotides encoding the novel peptides.

[0006] One aspect of the present invention provides a cell-penetrating peptide.

[0007] One aspect of the present invention provides peptides useful as intracellular active ingredient delivery vehicles.

[0008] Another aspect of the present invention is to provide a peptide that is useful as an intracellular active ingredient delivery vehicle, particularly for localized delivery to mitochondria.

[0009] Another aspect of the present invention is to provide a peptide useful for locally delivering to mitochondria an active ingredient for ameliorating, preventing or treating a mitochondrial-related disease or disorder.

[0010] One aspect of the present invention provides a conjugate in which a cell-penetrating peptide is conjugated with an active ingredient.

[0011] One aspect of the present invention provides a composition comprising a conjugate of a cell-penetrating peptide and an active ingredient.

[0012] One aspect of the present invention provides a pharmaceutical composition comprising a conjugate of a cell-penetrating peptide and an active ingredient.

[0013] One aspect of the present invention provides a functional cosmetic composition comprising a conjugate of a cell-penetrating peptide and an active ingredient.

[0014] One aspect of the present invention provides a health food composition comprising a conjugate of a cell-penetrating peptide and an active ingredient.

[0015] One aspect of the present invention provides an imaging agent comprising a conjugate of a cell-penetrating peptide and an active ingredient. [Means for solving the problem]

[0016] A conjugate according to one aspect of the present invention is a conjugate of a cell-permeable carrier peptide and an active ingredient, wherein the carrier peptide is a peptide comprising any one or more of SEQ ID NO:2 to SEQ ID NO:178, a peptide having more than 80% sequence identity to the peptide sequence, or a fragment thereof, and the peptide or fragment having more than 80% sequence identity is a conjugate that retains the cell permeability of any one of the peptides of SEQ ID NO:2 to SEQ ID NO:178.

[0017] In the conjugate according to another aspect of the present invention, the fragment is also a fragment consisting of three or more amino acids.

[0018] In a conjugate according to another aspect of the invention, the delivery peptide is also a peptide consisting of 30 amino acids or less.

[0019] In a conjugate according to another aspect of the invention, the delivery peptide is a peptide having any one of the sequences of SEQ ID NO: 2 to SEQ ID NO: 178, or a peptide having greater than 80% sequence identity with said peptide sequences.

[0020] An imaging agent according to one aspect of the present invention is also an imaging agent comprising any one of the conjugates described above.

[0021] In the contrast agent according to another aspect of the present invention, the contrast material is also for imaging cells.

[0022] In the contrast agent according to another aspect of the present invention, the cells are also stem cells.

[0023] A composition according to one aspect of the invention may comprise any one of the conjugates described above.

[0024] In the composition according to another aspect of the present invention, the active ingredient is an active ingredient for treating or preventing a disease, and the composition is also a pharmaceutical composition.

[0025] In a composition according to another aspect of the present invention, the active ingredient is an active ingredient of a functional cosmetic product, and the composition is also a cosmetic composition.

[0026] In the composition according to another aspect of the present invention, the active ingredient is an active ingredient of a functional health food, and the composition is also a health food composition.

[0027] A method according to one aspect of the present invention comprises administering any one of the above-mentioned conjugates to a subject in need thereof as a method for delivering an active ingredient into a cell, wherein the transport peptide is a cell-penetrating peptide that delivers the active ingredient into the cell, and the peptides and fragments having greater than 80% sequence homology also possess the cell permeability of any one of the peptides of SEQ ID NO: 2 to SEQ ID NO: 178.

[0028] A method for intracellular delivery of an active ingredient according to another aspect of the present invention also provides for localized delivery of the active ingredient to intracellular mitochondria.

[0029] A cell-penetrating peptide according to one aspect of the present invention is a peptide comprising any one of SEQ ID NO: 2 to SEQ ID NO: 178, or a fragment thereof.

[0030] A polynucleotide according to one aspect of the present invention is a polynucleotide encoding the above-mentioned cell-penetrating peptide.

[0031] A vector according to one aspect of the present invention is a vector comprising the polynucleotide.

[0032] A transformed cell according to one aspect of the present invention is also a transformed cell containing the above vector. Effect of the Invention

[0033] By using the peptide according to the present invention or a conjugate of the peptide and an active ingredient, the active ingredient that is difficult to penetrate into cells can be easily delivered into cells. This enhances the efficacy of the active ingredient, and as a result, the dosage can be reduced, minimizing side effects caused by drug administration and improving treatment efficiency. In particular, by locally delivering the active ingredient to mitochondria, the efficacy of improving, preventing or treating mitochondria-related diseases or disorders can be improved. Even in the case of cosmetics, excellent effects can be achieved with only a very small amount of active ingredient. By using a conjugate with a contrast material, it can be used as a contrast agent for monitoring the cell transplantation process or transplanted cells in cell therapy. In particular, it can be effectively used as a contrast agent for stem cells injected into a living body. [Brief description of the drawings]

[0034] [Figure 1] This is a diagram showing the number of cells uptaken by HeLa cell line after binding FITC to peptide pep1 of SEQ ID NO: 1 and analyzing it with a flow cytometer (FACS). The control group was treated with FITC only. [Diagram 2] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Diagram 3] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 4]FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Diagram 5] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 6] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 7] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 8] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 9] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 10] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 11]FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 12] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 13] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 14] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 15] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 16] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 17] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 18]FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 19] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 20] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 21] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 22] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 23] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 24] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Diagram 25]FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 26] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 27] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 28] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Figure 29] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a HeLa cell line. The results of analysis using a flow cytometer (FACS) showed the number of cells uptaken. The control group was treated with FITC only. [Diagram 30] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was shown as a result of analysis using a flow cytometer (FACS). The control group was treated with FITC only. [Diagram 31] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was shown as a result of analysis using a flow cytometer (FACS). The control group was treated with FITC only. [Diagram 32]FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Diagram 33] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Diagram 34] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Diagram 35] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Diagram 36] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 37] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 38] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 39]FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was shown as a result of analysis using a flow cytometer (FACS). The control group was treated with FITC only. [Diagram 40] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was shown as a result of analysis using a flow cytometer (FACS). The control group was treated with FITC only. [Diagram 41] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was shown as a result of analysis using a flow cytometer (FACS). The control group was treated with FITC only. [Diagram 42] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was shown as a result of analysis using a flow cytometer (FACS). The control group was treated with FITC only. [Diagram 43] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was shown as a result of analysis using a flow cytometer (FACS). The control group was treated with FITC only. [Diagram 44] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was shown as a result of analysis using a flow cytometer (FACS). The control group was treated with FITC only. [Diagram 45] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was shown as a result of analysis using a flow cytometer (FACS). The control group was treated with FITC only. [Figure 46]FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was shown as a result of analysis using a flow cytometer (FACS). The control group was treated with FITC only. [Figure 47] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was shown as a result of analysis using a flow cytometer (FACS). The control group was treated with FITC only. [Figure 48] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was shown as a result of analysis using a flow cytometer (FACS). The control group was treated with FITC only. [Figure 49] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was shown as a result of analysis using a flow cytometer (FACS). The control group was treated with FITC only. [Figure 50] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was shown as a result of analysis using a flow cytometer (FACS). The control group was treated with FITC only. [Figure 51] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a Hur7 cell line. The number of cells uptaken was shown as a result of analysis using a flow cytometer (FACS). The control group was treated with FITC only. [Figure 52] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 53]FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 54] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 55] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 56] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 57] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 58] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 59]FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 60] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 61] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 62] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 63] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 64] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 65]FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 66] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 67] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 68] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 69] FITC was bound to peptides of SEQ ID NO: 2 to SEQ ID NO: 178, and then each was applied to a human T lymphocyte cell line (Jurkat), and the number of cells uptaken was analyzed by a flow cytometer (FACS). The control group was treated with FITC only. [Figure 70] FITC was bound to peptide pep1 of SEQ ID NO: 1, and then treated with HeLa cells and cultured, and the cell viability and toxicity were measured. [Figure 71] FITC was bound to peptides of SEQ ID NO: 1 to SEQ ID NO: 178, and then each was applied to a HeLa cell line and cultured, after which the cell viability and toxicity were measured. [Figure 72]FITC was bound to peptides of SEQ ID NO: 1 to SEQ ID NO: 178, and then each was applied to a HeLa cell line and cultured, after which the cell viability and toxicity were measured. [Figure 73] FITC was bound to peptides of SEQ ID NO: 1 to SEQ ID NO: 178, and then each was applied to a HeLa cell line and cultured, after which the cell viability and toxicity were measured. [Figure 74] FITC was bound to peptides of SEQ ID NO: 1 to SEQ ID NO: 178, and then each was applied to a HeLa cell line and cultured, after which the cell viability and toxicity were measured. [Figure 75] FITC was bound to peptides of SEQ ID NO: 1 to SEQ ID NO: 178, and then each was applied to a HeLa cell line and cultured, after which the cell viability and toxicity were measured. [Figure 76] FITC was bound to peptides of SEQ ID NO: 1 to SEQ ID NO: 178, and then each was applied to a HeLa cell line and cultured, after which the cell viability and toxicity were measured. [Figure 77] FITC was bound to peptides of SEQ ID NO: 1 to SEQ ID NO: 178, and then each was applied to a HeLa cell line and cultured, after which the cell viability and toxicity were measured. [Figure 78] FITC was bound to peptides of SEQ ID NO: 1 to SEQ ID NO: 178, and then each was applied to a HeLa cell line and cultured, after which the cell viability and toxicity were measured. [Figure 79] FITC was bound to peptides of SEQ ID NO: 1 to SEQ ID NO: 178, and then each was applied to a HeLa cell line and cultured, after which the cell viability and toxicity were measured. [Figure 80] FITC was bound to peptides of SEQ ID NO: 1 to SEQ ID NO: 178, and then each was applied to a HeLa cell line and cultured, after which the cell viability and toxicity were measured. [Figure 81] FITC was bound to peptides of SEQ ID NO: 1 to SEQ ID NO: 178, and then each was applied to a HeLa cell line and cultured, after which the cell viability and toxicity were measured. [Figure 82] FITC was bound to peptides of SEQ ID NO: 1 to SEQ ID NO: 178, and then each was applied to a HeLa cell line and cultured, after which the cell viability and toxicity were measured. [Figure 83] FITC was bound to peptides of SEQ ID NO: 1 to SEQ ID NO: 178, and then each was applied to a HeLa cell line and cultured, after which the cell viability and toxicity were measured. [Figure 84] FITC was bound to peptides of SEQ ID NO: 1 to SEQ ID NO: 178, and then each was applied to a HeLa cell line and cultured, after which the cell viability and toxicity were measured. [Figure 85] FITC was bound to peptides of SEQ ID NO: 1 to SEQ ID NO: 178, and then each was applied to a HeLa cell line and cultured, after which the cell viability and toxicity were measured. [Figure 86] FITC was bound to peptides of SEQ ID NO: 1 to SEQ ID NO: 178, and then each was applied to a HeLa cell line and cultured, after which the cell viability and toxicity were measured. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0035] Proteins, nucleic acids, peptides, and viruses have great potential as therapeutic substances, but their use has been very limited because they are large at the molecular level and cannot penetrate tissues and cell membranes. In addition, even if the substance is small, it is often unable to penetrate the lipid bilayer of the cell membrane due to its nature or structure. Therefore, there have been attempts to move proteins, nucleic acids, peptides, and viruses into cells using electroporation or heat shock, but it has been difficult to move the substance into cells without damaging the cell membrane and while maintaining the activity of the substance. In this context, it has been revealed that the trans-activating transcriptional activator (TAT) protein derived from the human immunodeficiency virus (HIV) can act as a cell penetrating peptide that can move large active substances into cells, and research on this has been actively conducted. Specifically, unlike TAT protein, which is known to cause intracellular toxicity, research has been conducted on a substance that does not cause toxicity in vivo and can more effectively transport macromolecules such as proteins, nucleic acids, peptides, or viruses into target cells. Through continuous research, the present inventors have discovered that a peptide derived from telomerase has little cytotoxicity and excellent effects as a cell-penetrating peptide.

[0036] The peptides described in SEQ ID NO:1 to SEQ ID NO:178 are as shown in Tables 1 to 5 below. SEQ ID NO:179 is the sequence of human telomerase full-length protein. SEQ ID NO:1 is a peptide derived from telomerase and is a peptide composed of 16 amino acids. The peptides described in SEQ ID NO:2 to SEQ ID NO:77 are peptides containing the sequence of SEQ ID NO:1. The peptides described in SEQ ID NO:78 to SEQ ID NO:178 are fragments of the peptide of SEQ ID NO:1. The "name" in Table 1 below is given to distinguish the peptides. In another aspect of the present invention, one or more of the peptides described in SEQ ID NO:1 to SEQ ID NO:178 include "synthetic peptides" synthesized by selecting peptides at the corresponding positions among peptides contained in telomerase. In this specification, "pep" is a term that collectively refers to a peptide having the sequence of SEQ ID NO:1 or any one of SEQ ID NO:2 to SEQ ID NO:178, or a peptide having a sequence homology of more than 80% to the sequence, or a fragment of the sequence.

[0037] [Table 1]

[0038] [Table 2]

[0039] [Table 3]

[0040] [Table 4]

[0041] [Table 5]

[0042] One aspect of the present invention provides a polynucleotide encoding a peptide comprising SEQ ID NO: 2 to SEQ ID NO: 178, or a peptide having a sequence homology of more than 80% to said peptide sequence, or a fragment thereof. The polynucleotide can be used to mass-produce the peptide. For example, the peptide can be mass-produced by introducing a vector comprising a polynucleotide encoding the peptide into a host cell and culturing the cell.

[0043] The peptides disclosed herein may include peptides having greater than 80%, greater than 85%, greater than 90%, greater than 95%, greater than 96%, greater than 97%, greater than 98%, greater than 99% sequence identity, and may include peptides comprising any one or more of SEQ ID NO:2 through SEQ ID NO:178 or fragments thereof, as well as peptides with one or more amino acid changes, two or more amino acids, three or more amino acids, four or more amino acids, five or more amino acids, six or more amino acids, or seven or more amino acids changes.

[0044] In one aspect of the invention, the amino acid changes are of a nature that alter the physicochemical properties of the peptide, for example, amino acid changes that improve the thermostability, alter the substrate specificity, or change the pH optimum of the peptide.

[0045] As used herein, "amino acid" includes not only the 22 standard amino acids naturally incorporated into peptides, but also D-isomers and modified amino acids. Thus, in one aspect of the present invention, a peptide may also include a D-amino acid. Meanwhile, in another aspect of the present invention, a peptide may include non-standard amino acids that have been post-translationally modified. Examples of post-translational modifications include phosphorylation, glycosylation, acylation (including, for example, acetylation, myristoylation, and palmitoylation), alkylation, carboxylation, hydroxylation, glycation, biotinylation, ubiquitinylation, changes in chemical properties (e.g., beta-elimination deimidization, deamidation), and structural changes (e.g., formation of disulfide bridges). It also includes amino acid changes, such as changes at the amino group, carboxyl group or side chain, that occur due to chemical reactions that occur during conjugation with a crosslinker to form a peptide conjugate.

[0046] The peptides disclosed herein are also wild-type peptides identified and isolated from natural sources. On the other hand, the peptides disclosed herein are also artificial variants that contain an amino acid sequence in which one or more amino acids are replaced, deleted and / or inserted compared to a peptide that is a fragment of SEQ ID NO: 2-178. Amino acid changes in wild-type polypeptides, as well as in artificial variants, include conservative amino acid substitutions that do not significantly affect protein folding and / or activity. Examples of conservative substitutions are within the group of basic amino acids (arginine, lysine and histidine), acidic amino acids (glutamic acid and aspartic acid), polar amino acids (glutamine and asparagine), hydrophobic amino acids (leucine, isoleucine, valine and methionine), aromatic amino acids (phenylalanine, tryptophan and tyrosine), and small amino acids (glycine, alanine, serine and threonine). In general, amino acid substitutions that do not alter specific activity are known in the art. The most commonly occurring exchanges are Ala / Ser, Val / Ile, Asp / Glu, Thr / Ser, Ala / Gly, Ala / Thr, Ser / Asn, Ala / Val, Ser / Gly, Tyr / Phe, Ala / Pro, Lys / Arg, Asp / Asn, Leu / Ile, Leu / Val, Ala / Glu, and Asp / Gly, and their opposites. Other examples of conservative substitutions are set forth in Table 6 below. [Table 6]

[0047] Substantial modifications in the biological properties of the peptide are made by choosing substitutions that differ substantially in (a) their effect on maintaining the structure of the polypeptide backbone in the region of the substitution, e.g., a sheet or helical conformation, (b) their effect on maintaining the charge or hydrophobicity of the molecule at the target site, or (c) their effect on maintaining the bulk of the side chain. Natural residues are divided into the following groups based on common side chain properties: (1) Hydrophobic: norleucine, met, ala, val, leu, ile; (2) Neutral hydrophilic: cys, ser, thr; (3) Acidic: asp, glu; (4) Basic: asn, gln, his, lys, arg; (5) residues that affect chain orientation: gly, pro; and (6) Aromatics: trp, tyr, phe.

[0048] Non-conservative substitutions are made by exchanging a member of one of the classes for another. Any cysteine ​​residue that is not involved in maintaining the proper conformation of the peptide is generally replaced with serine to improve the oxidative stability of the molecule and prevent aberrant cross-linking. Conversely, cysteine ​​bonds can be added to the peptide to improve its stability.

[0049] Another type of amino acid variant of the peptides is one that alters the glycosylation pattern of the antibody, by which is meant the deletion of one or more carbohydrate residues found in the peptide and / or the addition of one or more glycosylation sites that are not present in the peptide.

[0050] Glycosylation of peptides is typically either N-linked or O-linked. N-linked refers to the attachment of the carbohydrate residue to the side chain of an asparagine residue. The tripeptide sequences asparagine-X-serine and asparagine-X-threonine, where X is any amino acid except proline, are the recognition sequences for enzymatic attachment of the carbohydrate residue to the asparagine side chain. Thus, the presence of one of these tripeptide sequences in a polypeptide creates a potential glycosylation site. O-linked glycosylation refers to the attachment of one of the sugars N-acetylgalactosamine, galactose, or xylose to a hydroxyamino acid. Most commonly, this refers to attachment to serine or threonine, although 5-hydroxyproline or 5-hydroxylysine can also be used.

[0051] Glycosylation sites may be added to the peptide by altering the amino acid sequence such that it contains one or more of the above-mentioned tripeptide sequences (for N-linked glycosylation sites).Such alterations may also be made by adding or substituting one or more serine or threonine residues to the sequence of the original antibody (for O-linked glycosylation sites).

[0052] One aspect of the present invention provides a cell-penetrating peptide, which is a peptide having any one of the sequences of SEQ ID NO:2 to SEQ ID NO:178, or a fragment thereof, or a peptide having more than 80% sequence homology with the peptide sequence. One aspect of the present invention provides a pharmaceutical composition for delivering one or more active ingredients, which comprises a peptide having SEQ ID NO:2 to SEQ ID NO:178, or a fragment thereof, or a peptide having more than 80% sequence homology with the peptide sequence. A peptide having SEQ ID NO:2 to SEQ ID NO:178, or a fragment thereof, or a peptide having more than 80% sequence homology with the peptide sequence, acts as a safe yet excellent cell-penetrating peptide, and can bind to a drug and effectively deliver the drug into cells.

[0053] One aspect of the present invention provides a conjugate in which a peptide having any one of the sequences of SEQ ID NO: 2 to SEQ ID NO: 178, or a peptide that is a fragment thereof, or a peptide having a sequence homology of more than 80% with the peptide sequence, is conjugated with an active ingredient to be transferred. In one aspect of the present invention, the active ingredient is one or more selected from proteins, nucleic acids, peptides, lipids, glycolipids, minerals, sugars, contrast agents, drugs, and chemical compounds. In one aspect of the present invention, the active ingredient is also a peptide. In one aspect of the present invention, the peptide that is the active ingredient is also a cytokine, an antibody, an antibody fragment, a therapeutic enzyme, a soluble receptor, or a ligand.

[0054] As used herein, the term "cell penetrating peptide (CPP)" refers to a peptide capable of transporting a cargo into a cell in vitro and / or in vivo. As used herein, the term "cargo" refers to any substance that can be bound to a cell penetrating peptide and transported into a cell, including, for example, any substance for which it is desired to increase the cell penetration efficiency, specifically, active substances of drugs, cosmetics, or health foods, more specifically, substances that are not easily transported into a cell via a general route, more specifically, proteins, nucleic acids, peptides, minerals, glucose, sugars, nanoparticles, biological preparations, viruses, contrast materials, or other chemical substances, but is not limited thereto. As used herein, the term "drug" refers to a broad concept that includes substances for alleviating, preventing, treating, or diagnosing a disease, injury, or a specific symptom.

[0055] As used herein, the term "carrier peptide" refers to a peptide that binds to an active ingredient and transports the active ingredient to a desired site.

[0056] In one aspect of the present invention, the protein or peptide to be transferred includes, but is not limited to, one or more of hormones, hormone analogs, enzymes, enzyme inhibitors, signal transduction proteins (or peptides), antibodies, and vaccines. In one aspect of the present invention, the nucleic acid may be a naturally occurring or artificial DNA molecule or RNA molecule, single-stranded or double-stranded. The nucleic acid molecule may be one or more nucleic acid molecules of the same type (e.g., having the same nucleoside sequence) or another type of nucleic acid molecule. The nucleic acid molecule may include, but is not limited to, one or more of DNA, cDNA, decoy DNA, RNA, siRNA, miRNA, shRNA, stRNA, snoRNA, snRNA, PNA, antisense oligomers, plasmids, and other modified nucleic acids. In one aspect of the present invention, the virus may include the entire virus or a viral core containing the nucleic acid of the virus. In one aspect of the present invention, the chemical is a broad concept that includes chemicals that can function as drugs, and includes natural or synthetic chemicals.

[0057] In one aspect of the present invention, the drug delivered into a cell by a cell-penetrating peptide may further comprise a drug delivery vehicle such as a liposome, a micelle, a nanoparticle, a magnetic particle or a quantum dot.

[0058] As used herein, the term "imaging agent" is a broad term that includes all agents used in medical imaging to image structures or fluids in vivo. Suitable imaging agents include, but are not limited to, radiopaque contrast agents, paramagnetic contrast agents, superparamagnetic contrast agents, computed tomography (CT) contrast agents, and other imaging agents. For example, radiopaque contrast agents (for X-ray imaging) include inorganic and organic iodine compounds (e.g., diatrizoic acid), radiopaque metals and their salts (e.g., silver, gold, platinum, etc.), and other radiopaque compounds (e.g., calcium salts, barium salts such as barium sulfate, tantalum, and tantalum oxide). Suitable paramagnetic contrast agents (for MR imaging) include gadolinium diethylene triaminepentaacetic acid (Gd-DTPA). acid and its derivatives, and other complexes of gadolinium, manganese, iron, dysprosium, copper, europium, erbium, chromium, nickel, and cobalt, such as 1,4,7,10-tetraazacyclododecane-N,N',N",N'''-tetraacetic acid (DOTA), ethylenediaminetetraacetic acid (EDTA), 1,4,7,10-tetraazacyclododecane-N,-N',N"-triacetic acid (D03A), 1,4,7-triazacyclononane-N,N',N"-triacetic acid (NOTA), 1,4,8,10-tetraazacyclotetradecane-N,N',N",N'''-tetraacetic acid (TETA), hydroxybenzylethylene-diaminediacetic acid (HBED), and the like. Suitable superparamagnetic contrast materials (for MR imaging) include magnetite, superparamagnetic iron oxide (SPIO), ultrasmall superparamagnetic iron oxide (USPIO), and monocrystailine iron oxide.Other suitable contrast agents include iodized and non-iodinated, ionic and non-ionic CT contrast agents, as well as contrast agents such as spin-labels, or other diagnostically effective agents.

[0059] Other examples of imaging agents include marker genes that code for proteins that are readily detectable when expressed in a cell, including, but not limited to, β-galactosidase, green fluorescent protein, blue fluorescent protein, luciferase, etc. A variety of labels are available, such as radionuclides, fluorescers, enzymes, enzyme substrates, enzyme cofactors, enzyme inhibitors, and ligands (particularly haptens).

[0060] In one embodiment of the present invention, the imaging material is ferrocenecarboxylic acid, which is represented by the following formula 2: The structure of ferrocene is shown in formula 1: [ka] [ka]

[0061] In one embodiment of the present invention, the conjugate of the cell-penetrating peptide and the contrast agent is ferrocenecarboxylic acid-pep1 represented by the following formula 3: [ka]

[0062] In one aspect of the invention, the peptide or composition is fused with one or more detectable labels. The label may also be a compound that generates, directly or indirectly, a detectable compound or signal in a chemical, physical, or enzymatic reaction. Labeling and subsequent detection is accomplished by methods known in the art (e.g., Sambrook, J., and Russel, DW (2001); Lottspeich, F., and Zorbas H. (1998), Bioanalytik, Spektrum Akademischer Verlag, Heidelberg / Berlin, Germany). The labels include, but are not limited to, fluorescent labels, enzyme labels, chromogenic labels, luminescent labels, radiolabels, haptens, biotin, metal complexes, metals, and gold colloids. All such types of labels are well known in the art and are commercially available from a variety of suppliers.

[0063] In one aspect of the present invention, a cargo can be directly bound to a peptide. In another aspect of the present invention, a cargo can be bound to a peptide via a number of binding methods, including covalent or non-covalent binding. The cargo can be bound, for example, to the N-terminus or C-terminus of a peptide according to one aspect of the present invention. For example, a cargo can be bound to a peptide via a disulfide bond or a covalent bond that binds the cargo to the alpha amine (α-amine) of a peptide N-glutamate (E) or the amine of a C-terminal lysine (K) residue. Alternatively, a cargo can be bound to a peptide via a non-covalent bond in which one of the peptide and the cargo envelops the other in a form, including, for example, a capsule.

[0064] In another aspect of the present invention, the peptide and the transfer target can be linked via a linker. For example, the transfer target can be linked to the peptide by introducing a linker such as 6-hydrazinopyridine-3-carboxylic acid linker to the α-amine of peptide N-glutamate or the amine of the C-terminal lysine residue, and then binding the transfer target to the linker.

[0065] In yet another aspect of the present invention, when the target of transfer is DNA or RNA, an SH group (thiol group) is introduced into the peptide and a maleimide group is introduced into the DNA or RNA, and then the SH group of the peptide is bonded to the maleimide group of the DNA or RNA, thereby binding the target of transfer to the peptide.

[0066] In yet another aspect of the present invention, when the target of transfer is a protein or peptide, the target of transfer can be linked to the target of transfer in the form of a fusion protein of the target of transfer and the peptide by binding the target of transfer to the DNA expressing the target of transfer and then expressing the binding. A specific example of binding by a fusion protein is as follows: when preparing a primer for producing a fusion protein, a nucleotide coding for the target of transfer is added before the nucleotide expressing the target of transfer, and the resulting nucleotide is inserted into a vector, such as a pET vector, as a restriction enzyme, and then transformed into a cell, such as BL-21(DE3), for expression. At this time, the fusion protein can be effectively expressed by treating with an expression inducer, such as IPTG (isopropyl-1-thio-β-D-galactopyranoside). The expressed fusion protein can then be purified by a method such as His tag purification, dialyzed with PBS, placed in a kit, and centrifuged at, for example, 2,000 to 4,000 rpm for 5 to 20 minutes to concentrate.

[0067] In one aspect of the invention, the delivery peptide can be conjugated to a dye or a fluorescent substance, specifically, fluorescein isothiocyanate (FITC) or green fluorescent protein (GFP). In one aspect of the invention, FITC binds to the amino group (NH3 + When the peptide does not have Lys at the terminus, the peptide can be linked to FITC via a linker containing Lys.

[0068] A peptide having any one or more of SEQ ID NO: 2 to SEQ ID NO: 178 disclosed herein, or a peptide fragment thereof, or a peptide having a sequence homology of more than 80% with the peptide sequence, which functions as a transport peptide, can be bound to a target for transport at a molar ratio of 1:1, but can also be bound at other molar ratios. For example, the molar ratio of CPP:transport target can be 2:1 or more. Specifically, it can be 2:1 or more, 3:1 or more, 4:1 or more, 5:1 or more, 6:1 or more, 7:1 or more, 8:1 or more, 9:1 or more, or 10:1 or more. This means that a plurality of transport peptide molecules can be bound to one target for transport molecule. A plurality of transport peptide molecules are linked to each other in series or parallel. Linked in series means that they are linked to each other at the terminal amino acid sites of the transport peptide, and linked in parallel means that they are linked to each other at the parts other than the terminal amino acid of the transport peptide. Conversely, the molar ratio of transport peptide:transport target can be 1:2 or more. This means that a plurality of molecules to be transferred can be bound to one delivery peptide molecule. For example, the molar ratio of delivery peptide to delivery target can be 1:2. Specifically, the molar ratio is 1:2 or more, 1:3 or more, 1:4 or more, 1:5 or more, 1:6 or more, 1:7 or more, 1:8 or more, 1:9 or more, or 1:10 or more.

[0069] Since the transport pathway of a peptide bound to fluorescein isothiocyanate can be easily identified, the delivery peptide according to one aspect of the present invention is utilized for cell imaging or intracellular drug delivery pathway tracing.

[0070] One aspect of the present invention provides the use of a peptide comprising any one or more of SEQ ID NO:2 to SEQ ID NO:178, or a fragment thereof, or a peptide having greater than 80% sequence identity to said peptide sequence, as a drug delivery vehicle for delivering one or more active ingredients.

[0071] One aspect of the present invention provides a method for delivering a drug into a cell of a subject, comprising applying to the subject a composition comprising a drug and a peptide comprising any one or more of SEQ ID NO:2 to SEQ ID NO:178, or a peptide that is a fragment thereof, or a peptide having greater than 80% sequence identity to the peptide sequence.

[0072] One aspect of the present invention provides a method for tracking the delivery pathway of a drug applied to a subject, comprising applying to the subject a peptide comprising any one or more of SEQ ID NO:2 to SEQ ID NO:178, or a fragment thereof, or a peptide having greater than 80% sequence identity to the peptide sequence, and a contrast agent.

[0073] One aspect of the present invention provides a method for tracking the delivery pathway of a drug applied to a subject, comprising administering to the subject a conjugate of a peptide comprising any one or more of SEQ ID NOs: 2 to 178, or a fragment thereof, or a peptide having greater than 80% sequence homology to the peptide sequence, and a contrast agent.

[0074] One aspect of the present invention provides a kit for intracellular drug delivery to a subject, comprising a composition comprising a peptide comprising any one or more of SEQ ID NO:2 to SEQ ID NO:178, or a peptide that is a fragment thereof, or a peptide having greater than 80% sequence homology to the peptide sequence, and a conjugate of a drug to be delivered, and instructions disclosing one or more of the dosage, route of administration, frequency of administration and indications of the composition.

[0075] One aspect of the present invention provides a cosmetic or food composition comprising an active ingredient and a peptide comprising any one or more of SEQ ID NO:2 to SEQ ID NO:178, or a peptide which is a fragment thereof, or a peptide having greater than 80% sequence identity with said peptide sequence. Another aspect of the present invention provides a cosmetic or food composition comprising a conjugate of an active ingredient and a peptide comprising any one or more of SEQ ID NO:2 to SEQ ID NO:178, or a peptide which is a fragment thereof, or a peptide having greater than 80% sequence identity with said peptide sequence.

[0076] One aspect of the present invention provides a pharmaceutical, cosmetic or food composition that comprises a peptide comprising any one or more of SEQ ID NO:2 to SEQ ID NO:178, or a peptide that is a fragment thereof, or a peptide having more than 80% sequence homology to the peptide sequence, and a conjugate of an active ingredient, and has excellent effects of delivering the active ingredient into cells.

[0077] Mitochondria are the central organelles for energy metabolism in nucleated cells and were the first intracellular organelles to be linked to human disease (Luft R, Ikkos D, Palmieri G, Ernster L, Afzelius B: A case of severe hypermetabolism of nonthyroid origin with a defect in the maintenance of mitochondrial respiratory control: a correlated clinical, biochemical, and morphological study, J Clin Invest 41: 1776-804, 1962).

[0078] Mitochondria play an important role in regulating cellular energy metabolism and apoptosis, and therefore act as the main target of various therapeutic drugs. In addition, the organelle is involved in regulating intracellular calcium concentration, and the mitochondrial respiratory chain acts as an important electron transport system for energy production and causes the production of reactive oxygen species. As a result, mitochondrial dysfunction is closely related to adult diseases such as diabetes, cardiomyopathy, infertility, blindness, kidney / liver disease, and stroke (Modica-Napolitano KS, Singh KK: April mitochondria as targets for detection and treatment of cancer, Expert Rev Mol Med 11: 1-19, 2002). At the same time, it has been suggested that mutations in mitochondrial genes may be involved in the pathogenesis of aging, degenerative neurological diseases, and cancer diseases.

[0079] According to one aspect of the present invention, there is provided a mitochondrial targeting active ingredient delivery system, comprising any one of the conjugates described above, a carrier peptide that is a peptide that moves locally to intracellular mitochondria and performs localized mitochondrial delivery of the active ingredient, and a peptide or a fragment thereof having a sequence homology of more than 80% is also a mitochondrial targeting active ingredient delivery system having the mitochondrial targeting ability of the corresponding peptide of any one of SEQ ID NO: 2 to SEQ ID NO: 178.

[0080] Furthermore, according to one aspect of the present invention, a composition for regulating mitochondrial activity is provided, which comprises any one of the conjugates described above, and the transport peptide is a peptide that transports locally to intracellular mitochondria and performs local mitochondrial delivery of the active ingredient, and the peptide and fragment thereof having more than 80% sequence homology is also a composition for regulating mitochondrial activity that possesses the mitochondrial targeting ability of the corresponding peptide of any one of SEQ ID NO: 2 to SEQ ID NO: 178.

[0081] In a composition for regulating mitochondrial activity according to one aspect of the present invention, the composition is a pharmaceutical composition for treating, preventing, inhibiting the progression of, or alleviating symptoms of a mitochondrial-related disease or disorder, and the active ingredient is also an ingredient that exhibits the function of treating, preventing, inhibiting the progression of, or alleviating symptoms of a mitochondrial-related disease or disorder.

[0082] "Mitochondrial associated diseases" referred to herein include, but are not limited to, Huntington's disease; amyotrophic lateral sclerosis; MELAS (mitochondrial encephalomyopathy with lactic academia and stroke-like episodes); MERRF (myoclonus, epilepsy, and myopathy with ragged red fibers); NARP / MILS (neurogenic muscular weakness, ataxia, retinitis pigmentosa / maternally inherited Leigh syndrome); LHON (Lebers hereditary optic neuropathy); KSS (Kearns-Sayre syndrome); PMPS (Pearson marrow-pancreas syndrome); syndrome;Pearson bone marrow-pancreatic syndrome;chronic progressive external opthalnoplegia (CPEO);Reye's syndrome;Alpers syndrome;multiple mtDNA deficiency syndromes;mtDNA wasting syndromes;complex I defects;complex II (SDH) defects;complex III defects;cytochrome c oxidase (COX, complex IV) defects;complex V defects;athenin nucleotide transporter (ANT) defects;pyruvate dehydrogenase (PDH) defects;ethylmalonic aciduria with lactic acidemia;3-methylglutaconic aciduria with lactic acidemia;refractory epilepsy with a decline during infection;Asperger's syndrome with a decline during infection;autism with a decline during infection;attention deficit hyperactivity disorder (ADHD);cerebral palsy with a decline during infection;dyslexia with a decline during infection;maternally inherited thrombocytopenia;leukemia;MNGIE (mitochondrial myopathy, peripheral and autonomic neuropathy, gastrointestinal dysfunction, and epilepsy); MARIAHS syndrome (mitochondrial ataxia, recrudescent infection, aphasia, hypouricemia / hypomyelination, seizure and dicarboxylic acid aciduria mitochondrial ataxia, recurrent infections, aphasia, hypouricemia / hypomyelinosis, seizures, and dicarboxyliuria); ND6 dystonia; cyclic vomiting syndrome with decline during infection; 3-hydroxyisobutyric aciduria with lactic acidemia, diabetes mellitus with lactic acidemia; uridine-responsive neurological syndrome (URNS); familial bilateral striatal necrosis (FBSN); aminoglycoside-associated hearing loss; attenuated cardiomyopathy; splenic lymphoma; Wolfram syndrome; multiple mitochondrial DNA defects syndrome; and renal tubular acidosis / diabetes / ataxia syndrome.

[0083] In another aspect, the present invention provides a nucleic acid molecule encoding the polypeptide, the base sequence of which is, for example, GAA GCG CGC CCG GCG CTG CTG ACC AGC CGC CTG CGC TTT ATT CCG AAA. The nucleic acid molecule is introduced into the host cell by techniques known to those skilled in the art, such as calcium phosphate, liposome, electroporation, transformation by contacting the cell with a virus, or by microinjection directly into the cell. The host cell may be a higher eukaryotic cell, such as a mammalian cell, or a lower eukaryotic cell, such as a yeast cell, or a prokaryotic cell, such as a bacterial cell. Examples of prokaryotic hosts suitable for transformation include Escherichia coli, Bacillus subtilis, Salmonella typhimurium, Pseudomonas, Streptomyces, and Mycobacteria.

[0084] The vector containing the nucleic acid molecule is generally a recombinant expression vector and may include an origin of replication and a selectable marker (e.g., dihydrofolate reductase for eukaryotic cell culture, or neomycin resistance, or tetracycline resistance or ampicillin resistance in E. coli, or the S. cerevisiae TRP1 gene) to allow for transformation of the host cell, and a promoter to control transcription of the protein coding sequence. Useful expression vectors that may be used include, for example, SV40; derivatives of pcDNA; known bacterial plasmids such as col E1, pCR1, pBR322, pMal-C2, pET, pGEX (Smith, et al., Gene 67: 31-40 (1988)); plasmids such as pMB9 and its derivative RP4; phage DNA, such as numerous derivatives of phage I, such as NM989; phage DNA, such as M13, and filamentous single-stranded phage DNA; yeast plasmids, e.g., phage DNAb, or vectors derived from combinations of plasmids and phage DNA modified to use expression control sequences. Mammalian expression vectors include an origin of replication, a suitable promoter and enhancer. They may also include necessary ribosome binding sites, polyadenylation sites, splice donor and splice acceptor sites, transcription termination sequences, and 5' flanking nontranscribed sequences. Mammalian expression vectors include vectors containing inducible promoters, such as the dihydrofolate reductase promoter, any expression vector containing a DHFR expression cassette, or a DHFR / methotrexate coamplification vector such as pED (Randal J, Kaufman, 1991, Randal J. Kaufman, Current Protocols in Molecular Biology, 16, 12 (1991)).Alternatively, glutamine synthetase / methionine sulfoximine co-amplification vectors, such as pEE14 (Celltech), vectors directing episomal expression under the control of Epstein-Barr virus (EBV) or nuclear antigen (EBNA), such as pREP4 (Invitrogen), pCEP4 (Invitrogen), pMEP4 (Invitrogen), pREP8 (Invitrogen), pREP9 (Invitrogen) and pEBVHis (Invitrogen), may be used. Selectable mammalian expression vectors include Rc / CMV (Invitrogen), pRc / RSV (Invitrogen), etc. Vaccinia virus mammalian expression vectors for use in the present invention include pSC11, pMJ601, pTKgptF1S, etc.

[0085] Yeast expression systems that can be used in the present invention include the non-fusion pYES2 vector (Invitrogen), the fusion pYESHisA,B,C (Invitrogen), and the pRS vector.

[0086] The vector can be introduced into various mammalian cells, particularly human-derived cells, bacteria, yeast, fungi, insects, nematodes and plant cells. Examples of suitable cells include VERO cells, HELA cells, such as ATCC No. CCL2, CHO cell lines, such as ATCC No. CCL61, COS cells, such as COS-7 cells and ATCC No. CRL1650 cells, W138, BHK, HepG2, 3T3, such as ATCC No. CRL6361, A549, PC12, K562 cells, 293 cells, Sf9 cells, such as ATCC No. CRL1711 and Cv1 cells, such as ATCC No. CCL70.

[0087] Other suitable cells for use in the present invention include prokaryotic host cell strains such as Escherichia coli (eg, strains DH5-α), Bacillus subtilis, Salmonella typhimurium, or strains belonging to the genera Pseudomonas, Streptomyces and Staphylococcus.

[0088] The composition according to one aspect of the present invention may contain a peptide having any one or more of SEQ ID NOs: 2 to 178, or a peptide that is a fragment thereof, or a peptide having a sequence homology of more than 80% to the peptide sequence, in an amount of 0.1 μg / mg to 1 mg / mg, specifically 1 μg / mg to 0.5 mg / mg, more specifically 10 μg / mg to 0.1 mg / mg. When contained within the above range, it is not only suitable for exhibiting the intended effect of the present invention, but also satisfies both the stability and safety of the composition, and it is also suitable to contain within the above range in terms of cost-effectiveness.

[0089] The compositions according to one aspect of the present invention may be applied to all animals including humans, dogs, chickens, pigs, cows, sheep, guinea pigs or monkeys.

[0090] Pharmaceutical compositions according to one aspect of the present invention may be administered orally, rectally, transdermally, intravenously, intramuscularly, intraperitoneally, intraosseously, intrathecally, or subcutaneously, and the like.

[0091] Dosage forms for oral administration include, but are not limited to, tablets, pills, soft or hard capsules, granules, powders, liquids or emulsions, while dosage forms for parenteral administration include, but are not limited to, injections, drops, lotions, ointments, gels, creams, suspensions, emulsions, suppositories, patches or sprays.

[0092] The pharmaceutical composition according to one aspect of the present invention may contain additives such as diluents, excipients, lubricants, binders, disintegrants, buffers, dispersants, surfactants, colorants, flavors, or sweeteners, if necessary. The pharmaceutical composition according to one aspect of the present invention is manufactured by a method commonly used in the art.

[0093] The active ingredient of the pharmaceutical composition according to one aspect of the present invention varies depending on the age, sex, weight, pathological condition and its severity, administration route, or the prescriber's judgment. Determining the dosage based on such factors is within the level of one skilled in the art, and the daily dosage may be, for example, but is not limited to, 0.1 μg / kg / day to 1 g / kg / day, specifically 1 μg / kg / day to 10 mg / kg / day, more specifically 10 μg / kg / day to 1 mg / kg / day, and even more specifically 50 μg / kg / day to 100 μg / kg / day. The pharmaceutical composition according to one aspect of the present invention is administered once to three times a day, but is not limited thereto.

[0094] The cosmetic composition according to one aspect of the present invention may be provided in any form suitable for topical application, such as a solution, an emulsion obtained by dispersing an oily phase in an aqueous phase, an emulsion obtained by dispersing an aqueous phase in an oily phase, a suspension, a solid, a gel, a powder, a paste, a foam or an aerosol. Such formulations may be prepared by methods commonly known in the art.

[0095] The cosmetic composition according to one aspect of the present invention may contain other ingredients that can provide a synergistic effect to the main effect, as long as the main effect is not impaired. The cosmetic composition according to one aspect of the present invention may further contain a moisturizer, an emollient, a surfactant, an ultraviolet absorber, a preservative, a bactericide, an antioxidant, a pH adjuster, an organic or inorganic pigment, a fragrance, a cooling agent, or an antiperspirant. The amount of the above ingredients can be easily selected by a person skilled in the art within a range that does not impair the object and effect of the present invention, and the amount is 0.01 to 5 wt%, specifically 0.01 to 3 wt%, based on the total weight of the cosmetic composition.

[0096] The dosage form of the food composition according to one aspect of the present invention is not particularly limited, and may be, for example, a tablet, granule, powder, liquid, solid preparation, etc. In addition to the active ingredient, each dosage form may be appropriately selected and mixed by a person skilled in the art according to the dosage form or purpose of use, and when used simultaneously with other ingredients, a synergistic effect occurs.

[0097] Determining the dosage of the active ingredient is within the level of a person skilled in the art, and the daily dosage may be, for example, but not limited to, 1 μg / kg / day to 10 mg / kg / day, more specifically 10 μg / kg / day to 1 mg / kg / day, and even more specifically 50 μg / kg / day to 100 μg / kg / day, and may vary depending on various factors such as the age, health condition, and complications of the subject to be administered.

[0098] The terminology used herein is intended only for the purpose of describing particular embodiments and is not intended to be limiting of the present invention. The use of omitted numbers before a noun does not denote a limitation of quantity, but rather indicates that there are one or more of the referenced noun item. The terms "including," "having," and "containing" are to be construed as open ended (i.e., meaning "including, but not limited to").

[0099] When referring to a range of values, it is merely an easy way to refer to each separate value falling within the range individually, and unless expressly stated otherwise, each separate value is incorporated herein just as if it were individually referred to in the specification. All range end values ​​are included within the range and are independently combinable.

[0100] All methods referred to herein may be performed in any suitable manner unless otherwise specified or clearly contradicted by context. The use of one and all examples, or exemplary language (e.g., "such as"), is merely to better describe the invention and does not limit the scope of the invention, unless otherwise included in the claims. No language in the specification should be construed as rendering any non-claimed element essential to the practice of the invention. Unless otherwise defined, technical and scientific terms used herein have the meanings commonly understood by those of ordinary skill in the art to which the invention pertains.

[0101] The preferred embodiments of the present invention include the best mode known to the inventor for carrying out the invention. Variations of the preferred embodiments will become apparent to those skilled in the art upon reading the preceding description. The inventors expect that those skilled in the art will take advantage of such variations and expect that the invention will be practiced in ways other than as described herein. Accordingly, the present invention includes equivalents and all modifications of the subject matter recited in the appended claims as permitted by the patent laws. Moreover, any combination of the above-described elements, within all possible variations, is encompassed by the present invention unless expressly stated to the contrary herein or clearly contradicted by context. Although the present invention has been specifically shown and described with reference to illustrative embodiments, it will be readily apparent to those skilled in the art that various changes in form and detail may be made therein without departing from the spirit and scope of the invention as defined by the appended claims. EXAMPLES

[0102] Example 1: Synthesis of peptides Peptides of SEQ ID NO:2 to SEQ ID NO:178 were prepared by a conventional solid phase peptide synthesis method. Specifically, the peptides were synthesized by coupling amino acids one by one from the C-terminus via Fmoc solid phase peptide synthesis (SPPS) using ASP48S (Peptron, Inc., Daejeon, Republic of Korea). The peptides were used with the first amino acid at the C-terminus attached to the resin as follows. For example,

[0103] NH2-Lys(Boc)-2-chloro-Trityl Resin NH2-Ala-2-chloro-Trityl Resin NH2-Arg(Pbf)-2-chloro-Trityl Resin

[0104] All amino acid raw materials used in peptide synthesis were protected at the N-terminus with Fmoc, and the residues were protected with Trt, Boc, t-Bu (t-butylester), Pbf (2,2,4,6,7-pentamethyl dihydro-benzofuran-5-sulfonyl), etc., all of which can be removed with acid. For example,

[0105] Fmoc-Ala-OH, Fmoc-Arg(Pbf)-OH, Fmoc-Glu(OtBu)-OH, Fmoc-Pro-OH, Fmoc-Leu-OH, Fmoc-Ile-OH, Fmoc-Phe-OH, Fmoc-Ser(tBu)-OH, Fmoc-Thr(tBu)-OH, F moc-Lys(Boc)-OH, Fmoc-Gln(Trt)-OH, Fmoc-Trp(Boc)-OH, Fmoc-Met-OH, Fmoc-Asn(Trt)-OH, Fmoc-Tyr(tBu)-OH, Fmoc-Ahx-OH, Trt-Mercaptoaceticacid

[0106] The coupling reagents used were HBTU [2-(1H-Benzotriazole-1-yl)-1,1,3,3-tetamethylaminium hexafluorophosphate] / HOBt [N-Hydroxxybenzotriazole] / NMM [4-Methylmorpholine]. Fmoc removal was performed using 20% ​​piperidine in DMF. The cleavage cocktail [TFA (trifluoroacetic acid) / TIS (triisopropylsilane) / EDT (ethanedithiol) / H2O = 92.5 / 2.5 / 2.5 / 2.5] was used to separate the synthesized peptide from the resin and remove the protecting groups of the residues.

[0107] Each peptide was synthesized by reacting the amino acid with a protecting group bound to the starting amino acid on the solid support, washing with a solvent, and repeating the deprotection process. The synthesized peptide was cut off from the resin, purified by HPLC, and freeze-dried after confirming the synthesis by MS.

[0108] Regarding the peptide synthesis method, the specific synthesis process will be described below using PEP 1 of SEQ ID NO: 1 as an example.

[0109] 1) Coupling The amino acid (8 equivalents) protected by NH2-Lys(Boc)-2-chloro-Trityl Resin and the coupling reagents HBTU (8 equivalents) / HOBt (8 equivalents) / NMM (16 equivalents) were dissolved in DMF and added, and the reaction was allowed to proceed at room temperature for 2 hours. The reaction was then washed with DMF, MeOH, and DMF, in that order.

[0110] 2) Fmoc deprotection 20% piperidine in DMF was added, and the reaction was carried out at room temperature for 5 minutes twice, and the mixture was washed with DMF, MeOH, and DMF in that order.

[0111] 3) Reactions 1 and 2 were repeated to produce the peptide backbone NH2-E(OtBu)-AR(Pbf)-PALLT(tBu)-S(tBu)-R(Pbf)LR(Pbf)-FIPK(Boc)-2-chloro-Trityl Resin).

[0112] 4) Cleavage: A cleavage cocktail was added to the completed peptide resin to separate the peptide from the resin.

[0113] 5) Cooling diethyl ether is added to the resulting mixture, which is then centrifuged to precipitate the resulting peptide.

[0114] 6) After purification by Prep-HPLC, the molecular weight was confirmed by LC / MS, and the product was frozen and made into a powder.

[0115] Example 2: Preparation of CPP and FITC conjugate (1) Preparation of FITC-CPP conjugate Conjugates of peptides of SEQ ID NO: 2 to SEQ ID NO: 178 conjugated with FITC were prepared as follows. For example, a conjugate of pep1 of SEQ ID NO: 1 and FITC, i.e., FITC-linker-pep1, was prepared as follows.

[0116] FITC was reacted with the peptide backbone NH2-linker-E(OtBu)-AR(Pbf)-PALLT(tBu)-S(tBu)-R(Pbf)LR(Pbf)-FIPK(Boc)-2-chloro-trityl resin obtained in Example 1. Specifically, fluorescein-5-isothiocyanate (FITC) (8 equivalents) and N,N-diisopropylethylamine (DIPEA) (16 equivalents) were dissolved in DMF and added, and the reaction was carried out at room temperature for 2 hours, followed by washing with DMF, MeOH, and DMF in that order. As a result, FITC-linker-E(OtBu)-AR(Pbf)-PALLT(tBu)-S(tBu)-R(Pbf)LR(Pbf)-FIPK(Boc)-2-chloro-trityl resin was obtained. Here, the linker was 6-aminohexanoic acid (Ahx). TFA / TIS / H2O=95 / 2.5 / 2.5 was added to the peptide resin after the synthesis was completed, and the conjugate was separated from the resin. Cooling diethyl ether was added to the resulting mixture, and the resulting conjugate was precipitated by centrifugation. Then, it was purified by Prep-HPLC, and the purity was confirmed by analytical HPLC, and the molecular weight was confirmed by LC / MS. It was confirmed that the obtained substance was FITC-pep1 through molecular weight confirmation. Then, it was lyophilized. Conjugates of peptides of SEQ ID NO:2 to SEQ ID NO:178 conjugated with FITC were also prepared, along with pep1 of SEQ ID NO:1.

[0117] (2) Preparation of CPP-FITC conjugate As in Example 1, a peptide backbone (NH2-E(OtBu)-AR(Pbf)-PALLT(tBu)-S(tBu)-R(Pbf)LR(Pbf)-FIPK(Dde)-2-chloro-trityl resin) was prepared. In order to selectively introduce FITC into the C-term of the prepared peptide backbone, the N-term was protected with Boc. Then, di-tert-butyl dicarbonate (30 equivalents) and DIPEA (30 equivalents) were dissolved in DMF and added, and the mixture was reacted at room temperature for 2 hours and washed with DMF, MeOH, and DMF in that order. As a result, Boc-E(OtBu)-AR(Pbf)-PALLT(tBu)-S(tBu)-R(Pbf)LR(Pbf)-FIPK(Dde)-2-chloro-trityl resin was obtained. Then, to attach FITC to the C-term K residue, the Dde protecting group of the C-term Lys residue was removed by treatment with hydrazine in 2% DMF. As a result, Boc-E(OtBu)-AR(Pbf)-PALLT(tBu)-S(tBu)-R(Pbf)LR(Pbf)-FIPK(NH2)-2-chloro-trityl resin was obtained. Then, FITC (8 equivalents) and DIPEA (16 equivalents) were dissolved in DMF and added, and the reaction was carried out at room temperature for 2 hours, and the resin was washed with DMF, MeOH, and DMF in that order. As a result, Boc-E(OtBu)-AR(Pbf)-PALLT(tBu)-S(tBu)-R(Pbf)LR(Pbf)-FIPK(FITC)-2-chloro-trityl resin was obtained. The peptide was separated from the resin by adding TFA / TIS / H2O=95 / 2.5 / 2.5 to the peptide resin after the synthesis was completed. Cooling diethyl ether was added to the resulting mixture, and the resulting peptide was precipitated by centrifugation. It was then purified by Prep-HPLC, and its purity was confirmed by analytical HPLC, and its molecular weight was confirmed by LC / MS. As a result, it was confirmed that the obtained substance was pep1-FITC. It was then dried.

[0118] Peptide-FITC conjugates of SEQ ID NO:2 to SEQ ID NO:178 were also prepared in the same manner as pep1-FITC.

[0119] Example 3: Cell-penetrating ability experiment of ferrocenecarboxylic-CPP conjugates 1. Preparation of ferrocenecarboxylic-CPP conjugates For coupling, the amino acid (8 equivalents) protected by NH2-Lys(Dde)-2-chloro-trityl resin and the coupling reagent HBTU (8 equivalents) / HOBt (8 equivalents) / NMM (16 equivalents) were dissolved in DMF and added, and the mixture was reacted at room temperature for 2 hours and washed with DMF, MeOH, and DMF in that order. Then, for Fmoc deprotection, 20% piperidine in DMF was added, and the mixture was reacted twice at room temperature for 5 minutes and washed with DMF, MeOH, and DMF in that order. The above reaction was repeated to prepare the peptide backbone (NH2-E(OtBu)-AR(Pbf)-PALLT(tBu)-S(tBu)-R(Pbf)LR(Pbf)-FIPK(Dde)-2-chloro-trityl resin). This was treated with hydrazine in 2% DMF to remove the Dde residue protecting group of the C-terminal Lys. Then, ferrocenecarboxylic acid (Sigma Aldrich cat.#46264, 16 equivalents) and coupling reagent HBTU (16 equivalents) / HOBt (16 equivalents) / NMM (32 equivalents) were dissolved in DMF and added, and the mixture was reacted at room temperature for 2 hours, and washed with DMF, MeOH, and DMF in that order. TFA / TIS / H2O=95 / 2.5 / 2.5 was added to the peptide resin after synthesis was completed, and the peptide was separated from the resin. Cooling diethyl ether was added to the resulting mixture, and the resulting peptide was precipitated by centrifugation. This was purified by HPLC, confirmed by MS, and lyophilized.

[0120] Example 4: Cell-penetrating ability experiment of pep-FITC conjugate (1) Cell permeability test in HeLa cell line Cell line culture HeLa cell line obtained from ATCC was cultured in minimum essential medium (MEM) supplemented with 10% fetal bovine serum (Invitrogen, USA), Earle's salts, non-essential amino acids, sodium pyruvate, 100 μg / ml penicillin, and 100 units / ml streptomycin at 37°C in a 5% CO2 incubator.

[0121] flow cytometry The cell line was treated with the peptide of the present invention, pep (CPP), represented by SEQ ID NO: 2 to 178, and analysis was performed using a flow cytometry and confocal microscopy to compare the uptake level with that of the control.

[0122] The cell lines were distributed into 6-well plates, and the medium was supplemented with 10% fetal bovine serum (Invitrogen, USA), 100 μg / ml penicillin, and 100 units / ml streptomycin, and cultured at 37°C in a 5% CO2 incubator for 12 hours. After washing with PBS, the cells were starvated in MEM (minimum essential medium) for 1 hour. Each carrier peptide was treated with 20 μM and cultured at 37°C for 1 hour. The washing process with PBS was repeated three times, and the carrier peptide attached to the outside of the cells was removed by treating with Trypsin-EDTA at 37°C for 10 minutes. The cells were harvested with chilled PBS, and the washing process was repeated three times via centrifugation. The cells were then suspended in 0.5 ml PBS containing 4% paraformaldehyde, and fluorescence was analyzed using a FACS Calibur (Becton Dickinson). The cellular uptake patterns of cells not treated with the delivery peptide (control group) and various conjugated peptides were compared and analyzed using MFI (mean fluorescence intensity).

[0123] The results are shown in Figures 2 to 29. Figure 1 shows the results for pep1 of SEQ ID NO: 1, which was carried out for reference. The analysis results shown in Figures 2 to 29 are shown in detail in Table 7 below.

[0124] [Table 7]

[0125] [Table 8]

[0126] [Table 9]

[0127] (2) Cell penetration test in Huh7 cell line Cell line culture A suspension cell line of human hepatocellular carcinoma cell line Huh7 (purchased from ATCC (American Type Cell Culture)) was used. The Huh7 cell line uses RPMI 1640 medium. The medium was supplemented with 10% fetal bovine serum (Invitrogen, USA), 100 μg / ml penicillin, and 100 units / streptomycin, and cultured at 37°C in a 5% CO2 incubator.

[0128] Cell permeability screening via flow cytometry To examine the cell-penetrating ability of the peptides, flow cytometry was performed on the Huh7 cell line treated with the peptides of SEQ ID NO: 2 to SEQ ID NO: 178. The analysis method was as described in the Hela cell line analysis in (1) above. The analysis results are shown in Figures 30 to 51.

[0129] (3) Cell penetration test in human T lymphocyte cell lines Cell line culture A suspension cell line of the human T-lymphocyte cell line Jurkat (human T-cell leukemia cell line) was used (purchased from ATCC (American Type Cell Culture)). The Jurkat cell line was grown in RPMI 1640 medium. The medium was supplemented with 10% fetal bovine serum (Invitrogen, USA), 100 μg / ml penicillin, and 100 units / ml streptomycin, and cultured at 37°C in a 5% CO2 incubator.

[0130] Human lymphocyte isolation was performed by drawing blood (50 ml) from a healthy subject, and then using Biocoll Separating Solution (Biochrom AG, Berlin, Germany), recovering the PBMC (peripheral blood mononuclear cells) layer and lymphocytes.

[0131] Cell penetration analysis of human T lymphocyte cell lines To examine the cell-penetrating ability of the peptides, flow cytometry was performed on human T lymphocyte cell lines treated with peptides of SEQ ID NO: 2 to SEQ ID NO: 178. The analysis method was as described in the HeLa cell line analysis in (1) above. The analysis results are shown in Figures 52 to 69.

[0132] (4) Cell viability and toxicity analysis Meanwhile, the HeLa cell line cultured in Example 4(1) was dispensed into a 96-well plate, and the medium was supplemented with 10% fetal bovine serum (Invitrogen, USA), 100 μg / ml penicillin, and 100 units / ml streptomycin, and cultured at 37°C in a 5% CO2 incubator for 12 hours. After washing with PBS, the cells were starvated in MEM (minimum essential medium) for 1 hour. After treating with 20 μM of each delivery peptide and culturing at 37°C for 24 hours, the cell viability and toxicity were analyzed using the MTT assay method. The results are shown in Figures 70 to 86.

Claims

1. A conjugate of a cell-permeable delivery peptide and an active ingredient, wherein the delivery peptide is a peptide consisting of any one of the amino acid sequences of SEQ ID NO:146, SEQ ID NO:148 to SEQ ID NO:150, SEQ ID NO:106, SEQ ID NO:108, SEQ ID NO:111 to SEQ ID NO:113, SEQ ID NO:115, SEQ ID NO:116, SEQ ID NO:125, SEQ ID NO:126, SEQ ID NO:131, SEQ ID NO:133 to SEQ ID NO:136, SEQ ID NO:142, and SEQ ID NO:

143.

2. The conjugate of claim 1 , wherein the active ingredient is one or more selected from the group consisting of proteins, nucleic acids, peptides, lipids, glycolipids, minerals, sugars, nanoparticles, biological agents, contrast agents, drugs, and chemical substances.

3. The conjugate of claim 1 , wherein the delivery peptide and the active ingredient are linked by a covalent bond, optionally mediated by a linker.

4. The conjugate of claim 1 , wherein the delivery peptide and the active ingredient are linked by a non-covalent bond.

5. The conjugate of claim 2 , wherein the active ingredient is a protein or peptide.

6. The conjugate of claim 5 , wherein the active ingredient is a cytokine, an antibody, an antibody fragment, a therapeutic enzyme, a soluble receptor or a ligand.

7. 3. The conjugate of claim 2, wherein the contrast agent is selected from the group consisting of a radiopaque contrast agent, a paramagnetic contrast agent, a superparamagnetic contrast agent, and a CT contrast agent.

8. The conjugate of claim 2 , wherein the imaging agent is iron-based.

9. The conjugate of claim 2 , wherein the imaging agent is ferrocene carboxylic acid.

10. A contrast agent comprising a conjugate according to any one of claims 1 to 9.

11. The imaging agent according to claim 10, which is for imaging cells.

12. The method of claim 11 , wherein the cells are stem cells.

13. A composition comprising as an active ingredient a conjugate according to any one of claims 1 to 9.

14. The composition of claim 13, wherein the active ingredient is a functional cosmetic active ingredient and the composition is a cosmetic composition.

15. A cell-permeable transport peptide, the peptide consisting of any one of the amino acid sequences of SEQ ID NO:146, SEQ ID NO:148 to SEQ ID NO:150, SEQ ID NO:106, SEQ ID NO:108, SEQ ID NO:111 to SEQ ID NO:113, SEQ ID NO:115, SEQ ID NO:116, SEQ ID NO:125, SEQ ID NO:126, SEQ ID NO:131, SEQ ID NO:133 to SEQ ID NO:136, SEQ ID NO:142, and SEQ ID NO:

143.

16. A polynucleotide encoding the peptide of claim 15.

17. A vector comprising the polynucleotide of claim 16.

18. A transformed cell comprising the vector of claim 17.

19. A composition for delivering an active ingredient into a cell, comprising a conjugate of a cell-permeable transport peptide and an active ingredient, wherein the transport peptide is a peptide consisting of any one of the amino acid sequences of SEQ ID NO:146 to SEQ ID NO:151, SEQ ID NO:106 to SEQ ID NO:108, SEQ ID NO:111 to SEQ ID NO:116, SEQ ID NO:122 to SEQ ID NO:126, SEQ ID NO:131 to SEQ ID NO:136, SEQ ID NO:140, and SEQ ID NO:142 to SEQ ID NO:

144.

20. Use of a peptide consisting of any one of the amino acid sequences of SEQ ID NO:146 to SEQ ID NO:151, SEQ ID NO:106 to SEQ ID NO:108, SEQ ID NO:111 to SEQ ID NO:116, SEQ ID NO:122 to SEQ ID NO:126, SEQ ID NO:131 to SEQ ID NO:136, SEQ ID NO:140, and SEQ ID NO:142 to SEQ ID NO:144, for the manufacture of a pharmaceutical for delivering an active ingredient into a cell.

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