Novel antiviral peptides

Abstract

The invention relates to an antiviral peptide and application thereof. More specifically, the present disclosure relates to a composition and method of treatment for treating viral diseases using antiviral peptides.

Description

Technical Field

[0001] The present invention relates to an antiviral peptide. Some embodiments of the present application provide antiviral agents having therapeutic uses against viral diseases. Background Art

[0002] Common mechanisms of action of known antiviral peptides include inhibiting viral membrane protein function, inhibiting the amplification of viral RNA in the nucleus of infected cells, inhibiting infection by disrupting the viral envelope, etc. In the case of enveloped viruses, the envelope has a lipid bilayer structure, so inactivation of the envelope means loss of viral infectivity. Summary of the Invention

[0003] Technical Problem

[0004] The present application provides an antiviral peptide not derived from known antiviral peptides.

[0005] Technical Solution

[0006] The present application provides a novel antiviral peptide. In this case, the amino acid sequence of the antiviral peptide is derived from the amino acid sequence of the Swiss-Prot sequence database.

[0007] Advantageous Effects

[0008] One object of the present application is to provide an antiviral peptide.

[0009] Another object of the present application is to provide the use of an antiviral peptide for the prevention or treatment of viral diseases.

[0010] Another object of the present application is to provide a pharmaceutical composition containing an antiviral peptide for the prevention or treatment of viral diseases.

[0011] Another object of the present application is to provide a method for treating viral diseases using an antiviral peptide.

[0012] Another object of the present application is to provide a method for screening a group of antiviral peptide candidates.

[0013] Another object of the present application is to provide a method for selecting an antiviral peptide. Brief Description of the Drawings

[0014] Figure 1Shows the predicted positions of amino acid residues when the peptide with the amino acid sequence DWLRIIWDWVCSVVSDFK (SEQ ID NO: 301) has an α-helical structure. In SEQ ID NO: 301, the residue aspartic acid (Asp, D) at the first position is designated as the N-terminus, and the residue lysine (Lys, K) at the last position is designated as the C-terminus. In this case, starting from D at the N-terminus, connect the lines in the order of the next amino acid residue W after D, and then the amino acid residue L, and the position where the line finally connects is the C-terminus K. Figure 1 Shows that in the single structure of the peptide, the hydrophilic region and the hydrophobic region are separated, and each region includes 9 amino acid residues. In this case, the types of amino acid residues included in the hydrophilic region are tryptophan (Trp, W), valine (Val, V), leucine (Leu, L), phenylalanine (Phe, F), and isoleucine (Ile, I), all of which are hydrophilic amino acid residues. The types of amino acid residues included in the hydrophobic region consist of the hydrophobic amino acid residues lysine (Lys, K), arginine (Arg, R), serine (Ser, S), and aspartic acid (Asp, D), as well as the hydrophilic amino acid residues cysteine (Cys, C) and isoleucine (Ile, I), and 7 of the 9 amino acid residues included in the hydrophobic region are hydrophobic amino acid residues. In Figure 1 Among the amino acid residues shown, the amino acid residues with a square pattern are hydrophilic amino acid residues, and the amino acid residues with other patterns are hydrophobic amino acid residues, so it is easier to identify the characteristics of the amino acid residues distributed in each region.

[0015] Figure 2 Shows an example of a structure in which gaps can occur in the viral phospholipid bilayer due to the small size of the virus.

[0016] Figure 3 Shows the results of confirming the liposome leakage efficacy of peptides SP-001 to SP-013.

[0017] Figure 4 Shows the results of confirming the liposome leakage efficacy of peptides SP-014 to SP-026.

[0018] Figure 5 Shows the results of confirming the liposome leakage efficacy of peptides SP-027 to SP-039.

[0019] Figure 6 Shows the results of confirming the liposome leakage efficacy of peptides SP-040 to SP-052.

[0020] Figure 7 Shows the results of confirming the liposome leakage efficacy of peptides SP-053 to SP-065.

[0021] Figure 8 Shows the results of confirming the liposome leakage efficacy of peptides SP-066 to SP-078.

[0022] Figure 9 Shows the results of confirming the liposome leakage efficacy of peptides SP-079 to SP-091.

[0023] Figure 10 Shows the results of confirming the liposome leakage efficacy of peptides SP-092 to SP-104.

[0024] Figure 11 Shows the results of confirming the liposome leakage efficacy of peptides SP-105 to SP-117.

[0025] Figure 12 Shows the results of confirming the liposome leakage efficacy of peptides SP-118 to SP-130.

[0026] Figure 13 Shows the results of confirming the liposome leakage efficacy of peptides SP-131 to SP-143.

[0027] Figure 14 Shows the results of confirming the liposome leakage efficacy of peptides SP-144 to SP-156.

[0028] Figure 15 Shows the results of confirming the liposome leakage efficacy of peptides SP-157 to SP-169.

[0029] Figure 16 Shows the results of confirming the liposome leakage efficacy of peptides SP-170 to SP-182.

[0030] Figure 17 Shows the results of confirming the liposome leakage efficacy of peptides SP-183 to SP-195.

[0031] Figure 18 Shows the results of confirming the liposome leakage efficacy of peptides SP-196 to SP-208.

[0032] Figure 19 Shows the results of confirming the liposome leakage efficacy of peptides SP-209 to SP-221.

[0033] Figure 20 Shows the results of confirming the liposome leakage efficacy of peptides SP-222 to SP-234.

[0034] Figure 21 Shows the results of confirming the liposome leakage efficacy of peptides SP-235 to SP-247.

[0035] Figure 22 showed the results of confirming the liposome leakage efficacy of SP-248 to SP-260 peptides.

[0036] Figure 23 showed the results of confirming the liposome leakage efficacy of SP-261 to SP-273 peptides.

[0037] Figure 24 showed the results of confirming the liposome leakage efficacy of SP-274 to SP-286 peptides.

[0038] Figure 25 showed the results of confirming the liposome leakage efficacy of SP-287 to SP-300 peptides.

[0039] Figures 3 to 25 The shown graph showed the results of measuring the change in fluorescence level over time in each well, with the horizontal axis representing time and the vertical axis representing the fluorescence level. In this case, the data of each graph can be analyzed to show that the peptides treated in the wells showing high fluorescence levels have excellent efficacy in damaging (or disrupting) the viral lipid membrane.

[0040] Figure 26 showed the results of confirming the liposome rupture efficacy of SP-001, SP-003, and SP-101 peptides. Specifically, Figure 26 showed the changes in fluorescence signals observed immediately (0 minutes), 2 minutes, 4 minutes, 6 minutes, 8 minutes, and 10 minutes after the cover slips attached with liposomes were treated with SP-001, SP-003, and SP-101 peptides at a constant rate.

[0041] Figure 27 showed the graph of fluorescence signals measured at 20-second intervals when the cover slips attached with liposomes were treated with SP-001 peptide at a constant rate. In this case, the vertical axis of the graph represents the normalized fluorescence signal obtained by converting the fluorescence value immediately after peptide treatment (0 seconds) to 1.

[0042] Figure 28 showed the graph of fluorescence signals measured at 20-second intervals when the cover slips attached with liposomes were treated with SP-003 peptide at a constant rate. In this case, the vertical axis of the graph represents the normalized fluorescence signal obtained by converting the fluorescence value immediately after peptide treatment (0 seconds) to 1.

[0043] Figure 29 showed the graph of fluorescence signals measured at 20-second intervals when the cover slips attached with liposomes were treated with SP-101 peptide at a constant rate. In this case, the vertical axis of the graph represents the normalized fluorescence signal obtained by converting the fluorescence value immediately after peptide treatment (0 seconds) to 1.

[0044] Figure 30 Shows the results of measuring the amount of remaining dengue virus RNA that was not degraded by ribonuclease after treating the dengue virus with 50 peptides respectively. In this case, the vertical axis of the graph represents the increase in the Ct value after peptide treatment relative to the Ct value of the negative control group (ΔCt).

[0045] Figure 31 Shows the Ct values of the viral RNA measured after incubating the dengue virus incubated with a specific concentration of peptides (30 kinds) with cells.

[0046] Figure 32 Shows the IC 50 value calculated based on the Ct values of the viral RNA measured after incubating the dengue virus incubated with various concentrations of SP-049 peptide with cells.

[0047] Figure 33 Shows the IC 50 value calculated based on the Ct values of the viral RNA measured after incubating the dengue virus incubated with various concentrations of SP-001 peptide with cells.

[0048] Figure 34 Shows the IC 50 value calculated based on the Ct values of the viral RNA measured after incubating the dengue virus incubated with various concentrations of SP-046 peptide with cells.

[0049] Figure 35 Shows the IC 50 value calculated based on the Ct values of the viral RNA measured after incubating the dengue virus incubated with various concentrations of SP-143 peptide with cells.

[0050] Figure 36 Shows the IC 50 value calculated based on the Ct values of the viral RNA measured after incubating the dengue virus incubated with various concentrations of SP-003 peptide with cells.

[0051] Figure 37 Shows the IC 50 value calculated based on the Ct values of the viral RNA measured after incubating the dengue virus incubated with various concentrations of SP-004 peptide with cells.

[0052] Figure 38 Shows the IC 50 value calculated based on the Ct values of the viral RNA measured after incubating the dengue virus incubated with various concentrations of SP-101 peptide with cells.

[0053] Figure 39Shows the IC calculated based on the Ct values of viral RNA measured after incubation of dengue virus with cells incubated with various concentrations of SP-185 peptide 50 values.

[0054] Figure 40 Shows the inhibition rate and IC calculated based on the Ct values of viral RNA measured after incubation of dengue virus with cells incubated with various concentrations of SP-191 peptide 50 values.

[0055] Figure 41 Shows the neutralization ability and IC calculated based on the percentage of infected cells measured after incubation of SARS-CoV2 virus with cells incubated with various concentrations of SP-049 peptide 50 values.

[0056] Figure 42 Shows the neutralization ability and IC calculated based on the percentage of infected cells measured after incubation of SARS-CoV2 virus with cells incubated with various concentrations of SP-116 peptide 50 values.

[0057] Figure 43 Shows the neutralization ability and IC calculated based on the percentage of infected cells measured after incubation of SARS-CoV2 virus with cells incubated with various concentrations of SP-151 peptide 50 values.

[0058] Figure 44 Shows the neutralization ability and IC calculated based on the percentage of infected cells measured after incubation of SARS-CoV2 virus with cells incubated with various concentrations of SP-046 peptide 50 values.

[0059] Figure 45 Shows the neutralization ability and IC calculated based on the percentage of infected cells measured after incubation of SARS-CoV2 virus with cells incubated with various concentrations of SP-004 peptide 50 values.

[0060] Figure 46 Shows the neutralization ability and IC calculated based on the percentage of infected cells measured after incubation of SARS-CoV2 virus with cells incubated with various concentrations of SP-003 peptide 50 values.

[0061] Figure 47 Shows the neutralization ability and IC calculated based on the percentage of infected cells measured after incubation of SARS-CoV2 virus with cells incubated with various concentrations of SP-101 peptide 50 values.

[0062] Figure 48 Shows the neutralization ability and IC 50 value calculated based on the percentage of infected cells measured after incubation of SARS-CoV2 virus with cells incubated with various concentrations of SP-171 peptide.

[0063] Figure 49 Shows the neutralization ability and IC 50 value calculated based on the percentage of infected cells measured after incubation of SARS-CoV2 virus with cells incubated with various concentrations of SP-110 peptide.

[0064] Figure 50 Shows the neutralization ability and IC 50 value calculated based on the percentage of infected cells measured after incubation of SARS-CoV2 virus with cells incubated with various concentrations of SP-114 peptide.

[0065] Figure 51 Shows the neutralization ability and IC 50 value calculated based on the percentage of infected cells measured after incubation of SARS-CoV2 virus with cells incubated with various concentrations of SP-143 peptide.

[0066] Figure 52 Shows the neutralization ability and IC 50 value calculated based on the percentage of infected cells measured after incubation of SARS-CoV2 virus with cells incubated with various concentrations of SP-221 peptide.

[0067] Figure 53 Shows a graph of the fluorescence signal measured at 20-second intervals when a coverslip attached with liposomes was treated with SP-049 peptide at a constant rate. In this case, the vertical axis of the graph represents the normalized fluorescence signal obtained by converting the fluorescence value immediately after peptide treatment (0 second) to 1. Figure 53 Shows the results of liposomes with various sizes (50 nm, 150 nm, and 250 nm), and shows that the normalized fluorescence intensity of smaller liposomes decreases faster than that of larger liposomes. Detailed Description

[0068] Some embodiments of the present application provide an antiviral peptide.

[0069] Some embodiments of the present application provide an antiviral peptide, the amino acid sequence of which is selected from the following sequences:

[0070] SEQ ID NO:1, SEQ ID NO:3 to 5, SEQ ID NO:8 to 23, SEQ ID NO:26, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:33SEQ ID NO: 35, SEQ ID NO: 36, SEQ ID NO: 38 to 40, SEQ ID NO: 43, SEQ ID NO: 45 to 52, SEQ ID NO: 55, SEQ ID NO: 58, SEQ ID NO: 59, SEQ ID NO: 62, SEQ ID NO: 64 to 73, SEQ ID NO: 75, SEQ ID NO: 78 to 87, SEQ ID NO: 90 to 93, SEQ ID NO: 97 to 102, SEQ ID NO: 104 to 106, SEQ ID NO: 109 to 112, SEQ ID NO: 114 to 116, SEQ ID NO: 118 to 123, SEQ ID NO: 126, SEQ ID NO: 127, SEQ ID NO: 130 to 132, SEQ ID NO: 136, SEQ ID NO: 138 to 147, SEQ ID NO: 151 to 155, SEQ ID NO: 162, SEQ ID NO: 164, SEQ ID NO: 166 to 168, SEQ ID NO: 170 to 174, SEQ ID NO: 176, SEQ ID NO: 180 to 182, SEQ ID NO: 184 to 186, SEQ ID NO: 188, SEQ ID NO: 189, SEQ ID NO: 191, SEQ ID NO: 192, SEQ ID NO: 194 to 196, SEQ ID NO: 198, SEQ ID NO: 201, SEQ ID NO: 204, SEQ ID NO: 206, SEQ ID NO: 208 to 211, SEQ ID NO: 215, SEQ ID NO: 218, SEQ ID NO: 219, SEQ ID NO: 221, SEQ ID NO: 224, SEQ ID NO: 225, SEQ ID NO: 228, SEQ ID NO: 229, SEQ ID NO: 231, SEQ ID NO: 232, SEQ ID NO: 236, SEQ ID NO: 237, SEQ ID NO: 240, SEQ ID NO: 242 to 246, SEQ ID NO: 248 to 254, SEQ ID NO: 256 to 258, SEQ ID NO: 260 to 270, SEQ ID NO: 272, SEQ ID NO: 273, SEQ ID NO: 275 to 288, SEQ ID NO: 290, SEQ ID NO: 291, SEQ ID NO: 293 to 296, SEQ ID NO: 298 and SEQ ID NO: 299.,

[0071] In certain embodiments, the antiviral peptide may be characterized as an amphiphilic peptide.

[0072] In certain embodiments, the antiviral peptide may be characterized as being able to cause damage and / or disruption of the viral lipid bilayer.

[0073] In such cases, the virus may be characterized as belonging to the Bunyaviridae, Coronaviridae, Filoviridae, Flaviviridae, Hepadnaviridae, Herpesviridae, Orthomyxoviridae, Poxviridae, Rhabdoviridae, Retroviridae, Togaviridae or Herpesviridae.

[0074] In certain embodiments, the antiviral peptide may be characterized as being able to form an α-helical structure.

[0075] Some embodiments of the present application provide a pharmaceutical composition for treating viral diseases, comprising a therapeutically effective amount of an antiviral peptide.

[0076] In such cases, the viral disease may be a disease caused by viral infection.

[0077] In certain embodiments, the pharmaceutical composition further comprises a pharmaceutically acceptable carrier and / or a pharmaceutically acceptable adjuvant.

[0078] In certain embodiments, the viral disease may be a respiratory disease caused by viral infection or coronavirus disease 2019 (COVID-19).

[0079] Some embodiments of the present application provide a method for treating viral diseases, comprising:

[0080] administering to a subject infected with a virus a pharmaceutical composition comprising a therapeutically effective amount of an antiviral peptide.

[0081] Some embodiments of the present application provide the use of an antiviral peptide for treating viral diseases.

[0082] Some embodiments of the present application provide the use of an antiviral peptide in the preparation of a medicament for treating viral diseases.

[0083] Some embodiments of the present application provide a method for screening a group of antiviral peptide candidates, comprising selecting peptides that meet the following conditions:

[0084] 1) The peptide consists of 18 amino acid residues;

[0085] 2) The peptide does not contain proline residues;

[0086] 3) The peptide contains at least 6 polar amino acid residues;

[0087] 4) The peptide contains 4 to 6 charged amino acid residues;

[0088] 5) The peptide has a hydrophobic moment (μH) value of 0.5 to 0.8 measured by the HeliQuest software;

[0089] 6) The peptide has a hydrophobicity (H) value of 0.7 to 0.9 measured by the HeliQuest software; and

[0090] 7) The peptide has a net charge of -2 to 0 measured by the HeliQuest software.

[0091] In certain embodiments, the polar amino acid residues can be characterized by aspartic acid (Asp, D), glutamic acid (Glu, E), lysine (Lys, K), arginine (Arg, R), histidine (His, H), serine (Ser, S), threonine (Thr, T), asparagine (Asn, N), glutamine (Gln, Q), and glycine (Gly, G), and the charged amino acid residues can be characterized by aspartic acid (Asp, D), glutamic acid (Glu, E), lysine (Lys, K), and arginine (Arg, R).

[0092] In certain embodiments, the method can be characterized by selecting a group of candidates for antiviral peptides from the amino acid sequences disclosed in Swiss-Prot sequence data.

[0093] Some embodiments of the present application provide a method for screening antiviral peptides, including:

[0094] Verifying the antiviral efficacy of the peptides selected by the method of screening a group of candidates for antiviral peptides.

[0095] In certain embodiments, verifying the antiviral efficacy of the selected peptides can be characterized by including at least one selected from the following:

[0096] Mixing liposomes with the selected peptides and confirming the damage or destruction of the liposomes; or

[0097] Mixing viruses with the selected peptides, and confirming the damage or destruction of the viral phospholipid bilayer, or confirming the reduction or inactivation of the virus's ability to infect cells.

[0098] In certain embodiments, the size of the liposomes can be characterized by 5 nm to 1500 nm.

[0099] In certain embodiments, the size of the liposomes can be characterized by 20 nm to 400 nm.

[0100] Embodiments of the present invention

[0101] In the following, the content of the present invention will be described in more detail by way of specific exemplary embodiments and examples with reference to the accompanying drawings. It should be noted that the accompanying drawings include some exemplary embodiments of the present invention, but not all exemplary embodiments. The content of the present invention disclosed in this specification can be implemented in various ways and is not limited to the specific exemplary embodiments described herein. These embodiments should be understood as being provided to meet the legal requirements applicable to this specification. Those of ordinary skill in the art to which the invention disclosed herein pertains will be able to conceive of many modifications and other exemplary embodiments of the content of the present invention disclosed herein. Therefore, it should be understood that the content of the present invention disclosed herein is not limited to the specific exemplary embodiments described herein, and its modifications and other exemplary embodiments are also within the scope of the claims.

[0102] Definition of Terms

[0103] The definitions of the terms used herein are as follows.

[0104] About

[0105] As used herein, the term "about" refers to an amount, level, value, quantity, frequency, percentage, dimension, size, content, weight or length that varies by 30%, 25%, 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2% or 1% relative to a reference amount, level, value, quantity, frequency, percentage, dimension, size, content, weight or length.

[0106] Amino Acid Sequence Label

[0107] Unless otherwise specified, when describing the sequence of a peptide in this specification, the single-letter symbols or three-letter symbols of amino acids are used, and it is described in the direction from the N-terminus to the C-terminus. For example, when expressed as RNVP, it refers to a peptide in which arginine, asparagine, valine and proline are sequentially linked in the direction from the N-terminus to the C-terminus. As another example, when expressed as Thr-Leu-Lys, it refers to a peptide in which threonine, leucine and lysine are sequentially linked in the direction from the N-terminus to the C-terminus. In the case of amino acids that cannot be represented by single-letter symbols, other letters are used to describe these amino acids, and they will be described by additional explanations.

[0108] The labeling method for each amino acid is as follows: alanine (Ala, A); arginine (Arg, R); asparagine (Asn, N); aspartic acid (Asp, D); cysteine (Cys, C); glutamic acid (Glu, E); glutamine (Gln, Q); glycine (Gly, G); histidine (His, H); isoleucine (Ile, I); leucine (Leu, L); lysine (Lys, K); methionine (Met, M); phenylalanine (Phe, F); proline (Pro, P); serine (Ser, S); threonine (Thr, T); tryptophan (Trp, W); tyrosine (Tyr, Y); and valine (Val, V).

[0109] Nucleic Acid Sequence Label

[0110] The symbols A, T, C, G, and U used in this specification will be interpreted to have the meanings understood by those skilled in the art. Depending on the context and technology, they can be appropriately interpreted as bases, nucleosides, or nucleotides on DNA or RNA. For example, when referring to bases, they can be respectively interpreted as adenine (A), thymine (T), cytosine (C), guanine (G), or uracil (U) itself, and when referring to nucleosides, they can be respectively interpreted as adenosine (A), thymidine (T), cytidine (C), guanosine (G), or uridine (U), and when referring to nucleotides in a sequence, they should be respectively interpreted as nucleotides including each of the above nucleosides.

[0111] In this specification, the symbol N can be appropriately interpreted as a base, nucleoside, or nucleotide on DNA or RNA depending on the context and technology. For example, when referring to bases, it can be respectively interpreted as any one of adenine (A), thymine (T), cytosine (C), guanine (G), and uracil (U), and when referring to nucleosides, each can be interpreted as any one of adenosine (A), thymidine (T), cytidine (C), guanosine (G), and uridine (U), and when referring to nucleotides in a sequence, they should be respectively interpreted as nucleotides including each of the above nucleosides.

[0112] Antiviral

[0113] As used herein, the term "antiviral" refers to the property of damaging or destroying the outer membrane of a virus, thereby reducing or inactivating the ability of the virus to infect cells. For example, an "antiviral peptide" refers to a peptide that can damage or destroy the outer membrane of a virus, thereby reducing or inactivating the ability of the virus to infect cells.

[0114] Background Art - Known Peptides

[0115] HCV is an enveloped single-stranded positive-sense RNA virus. The length of the single-stranded HCV RNA genome is approximately 9,500 nucleotides and it has a single open reading frame (ORF) encoding a single large protein of approximately 3,000 amino acids. In infected cells, this protein is cleaved at different sites by cellular and viral proteases to generate structural and non-structural proteins.

[0116] In the case of HCV, the production of mature non-structural proteins (NS2, NS3, NS4A, NS4B, NS5A, and NS5B) is affected by two viral proteases. The first viral protease is a metalloprotease located in NS2 that cis-cleaves the NS2-NS3 junction. The second viral protease is a serine protease contained within the N-terminal region of NS3 (hereinafter referred to as the NS3 protease) that mediates all subsequent cleavages downstream of NS3, cis-cleaving at the NS3-NS4A cleavage site and trans-cleaving at the remaining NS4A-NS4B, NS4B-NS5A, and NS5A-NS5B sites. The NS4A protein performs various functions by acting as a cofactor for the NS3 protease and possibly contributing to the membrane localization of NS3 and other viral replicase components.

[0117] Meanwhile, NS5A is present in the non-structural proteins of HCV. As a result of examining the amino acid sequence of NS5A by various studies, it was confirmed that it has an amphipathic α-helical structure at the NH2 terminus. This amphipathic α-helix is required to mediate the membrane binding of NS5A, and the protein is known to be anchored by inserting the α-helix into the plane of the membrane. In particular, disruption of the amphipathic α-helix of NS5A has been shown to disrupt the membrane-binding ability of NS5A and interrupt HCV RNA replication.

[0118] Cheng, Guofeng et al. identified antiviral peptides from the NS5A protein of HCV (Cheng, Guofeng et al., "Avirocidal phamic-helical peptide that inhibites hepatitis C virus infection in vitro.", Proceedings of the National Academy of Sciences 105.8 (2008): 3088-3093). Specifically, Cheng, Guofeng et al. confirmed that an amphipathic α-helical peptide (C5A) derived from the membrane-anchored domain of the hepatitis C virus (HCV) NS5A protein has antiviral activity against HCV in vitro. C5A has the same amino acid sequence as amino acid sequence numbers 3 to 20 near the N-terminus of the NS5A protein. Since C5A is derived from the membrane-anchored domain of NS5A, it is predicted to be a planar amphipathic α-helix, and it can destabilize HCV virions by permeabilizing the envelope of HCV virions. In addition, Cheng, Guofeng et al. compared the antiviral activities of C5A analogs derived from various HCV genotypes. Specifically, the antiviral activities of C5A analogs derived from HCV 1a, 1b, 2a, 3a, 4a, 5a, and 6a were compared, and it was confirmed that C5A derived from HCV 1a and C5A analogs derived from HCV 3a (e.g., DWLRIIWDWVCSVVSDFK (SEQ ID NO: 301)) exhibited relatively high antiviral activities (Table 1 of Cheng, Guofeng et al.).

[0119] 1. Antiviral Peptides of the Present Disclosure

[0120] 1.1 Overview - Antiviral Peptides of the Present Disclosure

[0121] According to one aspect of the present disclosure, antiviral peptides are disclosed. The antiviral peptides are amphipathic peptides having hydrophilic (polar, hydrophilic) amino acids and hydrophobic (non-polar, hydrophobic) amino acids. In this case, the antiviral peptides are selected from known amino acid sequences.

[0122] Details regarding the structures, characteristics, etc. that the antiviral peptides of the present disclosure may have will be described below.

[0123] 1.2 Length of Antiviral Peptides

[0124] The antiviral peptides of the present disclosure are composed of 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35 or 36 amino acid residues. In other words, the antiviral peptides are 12-mers, 13-mers, 14-mers, 15-mers, 16-mers, 17-mers, 18-mers, 19-mers, 20-mers, 21-mers, 22-mers, 23-mers, 24-mers, 25-mers, 26-mers, 27-mers, 28-mers, 29-mers, 30-mers, 31-mers, 32-mers, 33-mers, 34-mers, 35-mers or 36-mers peptides.

[0125] As an example, the number of amino acid residues constituting the antiviral peptide can be between any two values selected in the previous sentence. For example, the antiviral peptide can be composed of 16 to 27 amino acid residues.

[0126] Preferably, the number of amino acid residues constituting the antiviral peptide can be 18. In other words, the antiviral peptide can be composed of 18 amino acid residues. In other words, the antiviral peptide can be an 18-mer peptide.

[0127] 1.3 Amino Acid Residues of Antiviral Peptides

[0128] The antiviral peptides of the present disclosure include hydrophilic amino acid residues and hydrophobic amino acid residues. Here, the hydrophilic amino acid residues can include arginine (Arg, R), asparagine (Asn, N), aspartic acid (Asp, D), cysteine (Cys, C), glutamic acid (Glu, E), glutamine (Gln, Q), glycine (Gly, G), histidine (His, H), lysine (Lys, K), serine (Ser, S) and / or threonine (Thr, T). The hydrophobic amino acid residues can include alanine (Ala, A), isoleucine (Ile, I), leucine (Leu, L), methionine (Met, M), proline (Pro, P), valine (Val, V), phenylalanine (Phe, F), tyrosine (Tyr, Y) and / or tryptophan (Trp, W).

[0129] In some embodiments, the number of hydrophilic amino acid residues and hydrophobic amino acid residues contained in the antiviral peptide can be the same. In some other embodiments, the antiviral peptide can include 1, 2, 3, 4 or 5 more hydrophilic amino acid residues than hydrophobic amino acid residues. In some other embodiments, the antiviral peptide can include 1, 2, 3, 4 or 5 more hydrophobic amino acid residues than hydrophilic amino acid residues. In a preferred embodiment, the antiviral peptide can include 4 more hydrophobic amino acid residues than hydrophilic amino acid residues.

[0130] In some embodiments, when the antiviral peptide is an 18-mer peptide, the antiviral peptide may include 7 hydrophilic amino acid residues and 11 hydrophobic amino acid residues. In some other embodiments, when the antiviral peptide is an 18-mer peptide, the antiviral peptide may include 8 hydrophilic amino acid residues and 10 hydrophobic amino acid residues. In some other embodiments, when the antiviral peptide is an 18-mer peptide, the antiviral peptide may include 9 hydrophilic amino acid residues and 9 hydrophobic amino acid residues. In some other embodiments, when the antiviral peptide is an 18-mer peptide, the antiviral peptide may include 10 hydrophilic amino acid residues and 8 hydrophobic amino acid residues. In some other embodiments, when the antiviral peptide is an 18-mer peptide, the antiviral peptide may include 11 hydrophilic amino acid residues and 7 hydrophobic amino acid residues.

[0131] 1.4 Structure of Antiviral Peptides - α-Helical Structure

[0132] When reacting with a lipid membrane, the antiviral peptides of the present disclosure can form an α-helical structure. In this case, the reaction of the antiviral peptide with the lipid membrane may refer to the situation where the antiviral peptide contacts the lipid membrane (specifically, the phospholipid bilayer).

[0133] If the antiviral peptide has an α-helical structure, when viewed from the top down along the axis of the α-helical structure, the positions of the amino acid residues can be schematically represented as Figure 1 shown. Figure 1 One of the known antiviral peptides is illustrated to describe the α-helical structure of the antiviral peptide, and Figure 1 the amino acid sequence of the peptide shown in is N-terminus - DWLRIIWDWVCSVVSDFK (SEQ ID NO: 301) - C-terminus. When the peptide has an α-helical structure that rotates around a virtual first axis and the peptide is viewed from a direction parallel to the virtual first axis, each amino acid can be observed as Figure 1 shown. In this case, when a virtual plane including the virtual first axis is carefully selected (the virtual plane is shown by the dotted line in Figure 1 ), the peptide can be divided into a hydrophilic region and a hydrophobic region by the virtual plane. In this case, each of the two regions divided by the virtual plane includes 9 amino acid residues.

[0134] In the present specification, the hydrophilic region may refer to the region among the two regions divided by the virtual plane that contains a plurality of hydrophilic amino acid residues, and the hydrophobic region may refer to the other region that is not the hydrophilic region among the two divided regions.

[0135] In some embodiments, the hydrophilic region may contain only hydrophilic amino acid residues and no hydrophobic amino acid residues. That is, the hydrophilic region may contain 9 hydrophilic amino acid residues.

[0136] In some other embodiments, the hydrophilic region may comprise 8 hydrophilic amino acid residues and 1 hydrophobic amino acid residue.

[0137] In some other embodiments, the hydrophilic region may comprise 7 hydrophilic amino acid residues and 2 hydrophobic amino acid residues.

[0138] In some other embodiments, the hydrophilic region may comprise 6 hydrophilic amino acid residues and 3 hydrophobic amino acid residues.

[0139] In some embodiments, the hydrophobic region may comprise only hydrophobic amino acid residues and no hydrophilic amino acid residues. That is to say, the hydrophobic region may comprise 9 hydrophobic amino acid residues.

[0140] In some other embodiments, the hydrophobic region may comprise 8 hydrophobic amino acid residues and 1 hydrophilic amino acid residue.

[0141] In some other embodiments, the hydrophobic region may comprise 7 hydrophobic amino acid residues and 2 hydrophilic amino acid residues.

[0142] In some other embodiments, the hydrophobic region may comprise 6 hydrophobic amino acid residues and 3 hydrophilic amino acid residues.

[0143] In an embodiment, each hydrophilic amino acid or hydrophilic amino acid residue may be any one selected from arginine (Arg, R), asparagine (Asn, N), aspartic acid (Asp, D), cysteine (Cys, C), glutamic acid (Glu, E), glutamine (Gln, Q), glycine (Gly, G), histidine (His, H), lysine (Lys, K), serine (Ser, S), and threonine (Thr, T).

[0144] In an embodiment, each hydrophobic amino acid or hydrophobic amino acid residue may be any one selected from alanine (Ala, A), isoleucine (Ile, I), leucine (Leu, L), methionine (Met, M), valine (Val, V), phenylalanine (Phe, F), tyrosine (Tyr, Y), and tryptophan (Trp, W).

[0145] 1.5 Characteristics of Antiviral Peptides - Amphiphilicity

[0146] The antiviral peptides of the present disclosure are amphiphilic peptides. Specifically, the antiviral peptides do not have amphiphilic characteristics when having a linear structure, but have amphiphilic characteristics when having an α-helical structure. This is because, due to the characteristics of the amino acid sequence of the antiviral peptides, when the antiviral peptides have a linear structure, there is no region in the single structure where hydrophilic or hydrophobic amino acid residues are concentratedly distributed, but when the antiviral peptides have an α-helical structure, there is a region in the single structure where hydrophilic or hydrophobic amino acid residues are concentratedly distributed. That is to say, when the antiviral peptides form an α-helical structure, due to the interaction with the lipid membrane or other reasons, the antiviral peptides have amphiphilic characteristics.

[0147] The amphiphilic characteristic means that when the peptide is considered to have a three-dimensional structure such as an α-helical structure, in the three-dimensional structure, one side of the peptide exhibits hydrophilic (polar, hydrophilic) properties, while the other side exhibits hydrophobic (non-polar, hydrophobic) properties. That is to say, when the peptide has amphiphilicity, it means that the peptide has both hydrophobicity and hydrophilicity.

[0148] The amphiphilic characteristic of the antiviral peptides can be described by the distribution of hydrophilic amino acid residues and hydrophobic amino acid residues in the three-dimensional helical structure of the antiviral peptides, as described in the section "1.4 Structure of Antiviral Peptides - α-Helical Structure" above. Specifically, in any one of the two regions divided by a virtual plane in the α-helical structure, there are more hydrophilic amino acid residues distributed than hydrophobic amino acid residues, while in the other region, there are more hydrophobic amino acid residues distributed than hydrophilic amino acid residues, so that the antiviral peptides with an α-helical structure have both hydrophilicity and hydrophobicity in the single structure.

[0149] The amphiphilic characteristic is considered to be one of the reasons for the efficacy of the antiviral peptides in acting on the phospholipid bilayer.

[0150] As an example, the HeliQuest software can be used to measure the amphiphilic characteristics of antiviral peptides. In some embodiments, when the sequence of an antiviral peptide is input into the HeliQuest software, a hydrophobic moment (μH) value of any one of 0.5 to 0.8 can be output. That is, the antiviral peptide can have any one of the hydrophobic moment values of 0.5 to 0.8 (hydrophobic moment values calculated by the HeliQuest software). In some embodiments, when the sequence of an antiviral peptide is input into the HeliQuest software, a hydrophobicity (H) value of any one of 0.7 to 0.9 can be output. That is, the antiviral peptide can have any one of the hydrophobic values of 0.7 to 0.9 (hydrophobic values calculated by the HeliQuest software). In some embodiments, when the sequence of an antiviral peptide is input into the HeliQuest software, a net charge of any one of the range from -2 to 0 can be output. That is, the antiviral peptide can have a net charge of any one of the range from -2 to 0 (net charge calculated by the HeliQuest software).

[0151] 1.6 Sequences of Antiviral Peptides

[0152] The amino acid sequences of the antiviral peptides disclosed in the present disclosure may be selected from SEQ ID NO:1, SEQ ID NO:3 to 5, SEQ ID NO:8 to 23, SEQ ID NO:26, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:35, SEQ ID NO:36, SEQ ID NO:38 to 40, SEQ ID NO:43, SEQ ID NO:45 to 52, SEQ ID NO:55, SEQ ID NO:58, SEQ ID NO:59, SEQ ID NO:62, SEQ ID NO:64 to 73, SEQ ID NO:75, SEQ ID NO:78 to 87, SEQ ID NO:90 to 93, SEQ ID NO:97 to 102, SEQ ID NO:104 to 106, SEQ ID NO:109 to 112, SEQ ID NO:114 to 116, SEQ ID NO:118 to 123, SEQ ID NO:126, SEQ ID NO:127, SEQ ID NO:130 to 132, SEQ ID NO:136, SEQ ID NO:138 to 147, SEQ ID NO:151 to 155, SEQ ID NO:162, SEQ ID NO:164, SEQ ID NO:166 to 168, SEQ ID NO:170 to 174, SEQ ID NO:176, SEQ ID NO:180 to 182, SEQ ID NO:184 to 186, SEQ ID NO:188, SEQ ID NO:189, SEQ ID NO:191, SEQ ID NO:192, SEQ ID NO:194 to 196, SEQ ID NO:198, SEQ ID NO:201, SEQ ID NO:204, SEQ ID NO:206, SEQ ID NO:208 to 211, SEQ ID NO:215, SEQ ID NO:218, SEQ ID NO:219, SEQ ID NO:221, SEQ ID NO:224, SEQ ID NO:225, SEQ ID NO:228, SEQ ID NO:229, SEQ ID NO:231, SEQ ID NO:232, SEQ ID NO:236, SEQ ID NO:237, SEQ ID NO:240, SEQ ID NO:242 to 246, SEQ ID NO:248 to 254, SEQ ID NO:256 to 258, SEQ ID NO:260 to 270, SEQAny one of ID NO:272, SEQ ID NO:273, SEQ ID NO:275 to 288, SEQ ID NO:290, SEQ ID NO:291, SEQ ID NO:293 to 296, SEQ ID NO:298, and SEQ ID NO:299, or a sequence having 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99% or higher homology (sequence identity) thereto.

[0153] In certain embodiments, the amino acid sequence of the antiviral peptide can be any one of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:20, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:46, SEQ ID NO:48, SEQ ID NO:49, SEQ ID NO:52, SEQ ID NO:59, SEQ ID NO:78, SEQ ID NO:82, SEQ ID NO:87, SEQ ID NO:90, SEQ ID NO:91, SEQ ID NO:98, SEQ ID NO:99, SEQ ID NO:101, SEQ ID NO:106, SEQ ID NO:110, SEQ ID NO:112, SEQ ID NO:114, SEQ ID NO:115, SEQ ID NO:116, SEQ ID NO:123, SEQ ID NO:126, SEQ ID NO:131, SEQ ID NO:132, SEQ ID NO:139, SEQ ID NO:141, SEQ ID NO:143, SEQ ID NO:144, SEQ ID NO:151, SEQ ID NO:162, SEQ ID NO:167, SEQ ID NO:171, SEQ ID NO:189, SEQ ID NO:221, SEQ ID NO:228, SEQ ID NO:242, SEQ ID NO:245, SEQ ID NO:246, SEQ ID NO:254, SEQ ID NO:256, SEQ ID NO:264, SEQ ID NO:283, SEQ ID NO:290, and SEQ ID NO:296, or a sequence having 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99% or higher homology (sequence identity) thereto.

[0154] In certain embodiments, the amino acid sequence of the antiviral peptide can be any one selected from SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:20, SEQ ID NO:46, SEQ ID NO:48, SEQ ID NO:49, SEQ ID NO:59, SEQ ID NO:87, SEQ ID NO:90, SEQ ID NO:91, SEQ ID NO:98, SEQ ID NO:101, SEQ ID NO:110, SEQ ID NO:114, SEQ ID NO:116, SEQ ID NO:126, SEQ ID NO:132, SEQ ID NO:141, SEQ ID NO:143, SEQ ID NO:144, SEQ ID NO:151, SEQ ID NO:221, SEQ ID NO:246, SEQ ID NO:256, SEQ ID NO:264, SEQ ID NO:283, SEQ ID NO:290, and SEQ ID NO:296, or a sequence having 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99% or higher homology (sequence identity) thereto.

[0155] In certain embodiments, the amino acid sequence of the antiviral peptide can be any one selected from SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:46, SEQ ID NO:49, SEQ ID NO:101, SEQ ID NO:143, SEQ ID NO:185, and SEQ ID NO:191, or a sequence having 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99% or higher homology (sequence identity) thereto.

[0156] In certain embodiments, the amino acid sequence of the antiviral peptide can be any one selected from SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:46, SEQ ID NO:49, SEQ ID NO:101, SEQ ID NO:110, SEQ ID NO:114, SEQ ID NO:116, SEQ ID NO:143, SEQ ID NO:151, SEQ ID NO:171, and SEQ ID NO:221, or a sequence having 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99% or higher homology (sequence identity) thereto.

[0157] In certain embodiments, the amino acid sequence of the antiviral peptide can be any one selected from SEQ ID NO:1, SEQ ID NO:3, and SEQ ID NO:101, or a sequence having 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99% or higher homology (sequence identity) thereto.

[0158] In certain embodiments, the amino acid sequence of the antiviral peptide can be SEQ ID NO:49 or a sequence having 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, or 99% or higher homology (sequence identity) thereto.

[0159] Meanwhile, the antiviral effects of the peptides selected by the method of selecting antiviral peptide candidates were confirmed through Experimental Examples 3 to 7, as described below.

[0160] 1.7 Method for producing antiviral peptide

[0161] The antiviral peptides disclosed in the present disclosure can be produced by known peptide synthesis methods. For example, the antiviral peptides can be synthesized by the peptide synthesis method of general solid-phase peptide synthesis (SPPS). As a specific example, the antiviral peptides of the present disclosure can be produced by the method disclosed in Experimental Example 2 of the present application.

[0162] 2. Method for screening antiviral peptides

[0163] 2.1 Method for Screening a Group of Antiviral Peptide Candidates

[0164] The inventors of the present application screened novel antiviral peptide candidates based on the characteristics of a peptide having the amino acid sequence DWLRIIWDWVCSVVSDFK (SEQ ID NO:301) derived from HCV genotype 3a.

[0165] Thus, as an aspect of the present disclosure, a method for screening a group of candidates for antiviral peptides is disclosed. The detailed method of the screening method will be described below.

[0166] 2.1.1 Peptide conditions

[0167] The method for screening a group of candidates for antiviral peptides according to the present disclosure includes selecting, finding, and / or searching for peptides that satisfy at least one of the following conditions:

[0168] 1) The peptide consists of 12 to 36 amino acid residues (or the length of the peptide is from 12-mer to 36-mer);

[0169] 2) The peptide does not contain proline residues;

[0170] 3) The peptide contains at least 6 polar amino acid residues;

[0171] 4) The peptide contains 4 to 6 charged amino acid residues;

[0172] 5) The peptide has a hydrophobic moment (μH) value of 0.5 to 0.8;

[0173] 6) The peptide has a hydrophobicity (H) value of 0.7 to 0.9; and

[0174] 7) The peptide has a net charge of -2 to 0.

[0175] In some embodiments, the polar amino acid residues may be aspartic acid (Asp, D), glutamic acid (Glu, E), lysine (Lys, K), arginine (Arg, R), histidine (His, H), serine (Ser, S), threonine (Thr, T), asparagine (Asn, N), glutamine (Gln, Q), and glycine (Gly, G).

[0176] In some embodiments, the charged amino acid residues may be aspartic acid (Asp, D), glutamic acid (Glu, E), lysine (Lys, K), and arginine (Arg, R).

[0177] In some embodiments, the hydrophobic moment (μH) value of the peptide may refer to the value measured by the HeliQuest software.

[0178] In some embodiments, the hydrophobicity (H) value may refer to the value measured by the HeliQuest software.

[0179] In some embodiments, the net charge of the peptide may refer to the value measured by the HeliQuest software.

[0180] In some embodiments, a method for screening a group of antiviral peptide candidates may include selecting, finding, and / or searching for peptides that meet the following conditions:

[0181] The number of amino acid residues constituting the peptide is 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, or 36, or falls within a range defined by any two of these values (e.g., 18 to 27).

[0182] In some embodiments, a method for screening a group of antiviral peptide candidates may include selecting, finding, and / or searching for peptides that meet the following conditions:

[0183] Among the amino acid residues constituting the peptide, the number of charged amino acid residues is 4, 5, or 6, or falls within a range defined by any two of these values (e.g., 4 to 6).

[0184] In some embodiments, a method for screening a group of candidates for antiviral peptides may include selecting, finding, and / or searching for peptides that meet the following conditions:

[0185] The hydrophobic moment (μH) value of the hydrophobic moment of the peptide is about 0.50, about 0.51, about 0.52, about 0.53, about 0.54, about 0.55, about 0.56, about 0.57, about 0.58, about 0.59, about 0.60, about 0.61, about 0.62, about 0.63, about 0.64, about 0.65, about 0.66, about 0.67, about 0.68, about 0.69, about 0.70, about 0.71, about 0.72, about 0.73, about 0.74, about 0.75, about 0.76, about 0.77, about 0.78, about 0.79, or about 0.80, or falls within a range defined by any two of these values (e.g., 0.50 to 0.80).

[0186] In some embodiments, a method for screening a group of candidates for antiviral peptides may include selecting, finding, and / or searching for peptides that meet the following conditions:

[0187] The hydrophobicity (H) value of the peptide is about 0.70, about 0.71, about 0.72, about 0.73, about 0.74, about 0.75, about 0.76, about 0.77, about 0.78, about 0.79, about 0.80, about 0.81, about 0.82, about 0.83, about 0.84, about 0.85, about 0.86, about 0.87, about 0.88, about 0.89, or about 0.90, or falls within a range defined by any two of these values (e.g., 0.70 to 0.90).

[0188] In some embodiments, a method for screening a group of candidates for antiviral peptides may include selecting, finding, and / or searching for peptides that meet the following conditions:

[0189] The net charge of the peptide is about -2.0, about -1.9, about -1.8, about -1.7, about -1.6, about -1.5, about -1.4, about -1.3, about -1.2, about -1.1, about -1.0, about -0.9, about -0.8, about -0.7, about -0.6, about -0.5, about -0.4, about -0.3, about -0.2, about -0.1, or about 0.0, or falls within a range defined by any two of these values (e.g., -2.0 to 0.0).

[0190] In a specific embodiment, a method for screening a group of candidates for antiviral peptides includes selecting, finding, and / or searching for peptides that meet at least one of the following conditions:

[0191] 1) The peptide consists of 18 amino acid residues (in other words, the length of the peptide is an 18-mer);

[0192] 2) The peptide does not contain proline residues;

[0193] 3) The peptide contains at least 6 polar amino acid residues, where the polar amino acid residues are aspartic acid (Asp, D), glutamic acid (Glu, E), lysine (Lys, K), arginine (Arg, R), histidine (His, H), serine (Ser, S), threonine (Thr, T), asparagine (Asn, N), glutamine (Gln, Q), and glycine (Gly, G);

[0194] 4) The peptide contains 4 to 6 charged amino acid residues, where the charged amino acid residues are aspartic acid (Asp, D), glutamic acid (Glu, E), lysine (Lys, K), and arginine (Arg, R);

[0195] 5) The peptide has a hydrophobic moment (μH) value of 0.5 to 0.8 measured by HeliQuest software;

[0196] 6) The peptide has a hydrophobicity (H) value of 0.7 to 0.9 measured by HeliQuest software; and

[0197] 7) The peptide has a net charge of -2 to 0 measured by HeliQuest software.

[0198] 2.1.2 Use of the database

[0199] The method for screening a candidate group of antiviral peptides according to the present disclosure may include selecting, searching for, and / or hunting for peptides that meet the conditions disclosed in the "2.1.1 Peptide Conditions" section of the present disclosure from the amino acid sequences disclosed in a known database:

[0200] In certain embodiments, the known database may be the SwissProt database (or Swiss-Prot sequence data).

[0201] In certain embodiments, the amino acid sequences disclosed in the known database may be the amino acid sequences of proteins derived from human or non-human animals (such as rats, mice, or yeast).

[0202] 2.1.3 Examples of the candidate group for screening antiviral peptides

[0203] The present inventors have selected peptides represented by any of the amino acid sequences of SEQ ID NO: 1 to 300 as a candidate group of antiviral peptides by the method for screening a candidate group of antiviral peptides, which method includes selecting peptides that meet the following conditions from the amino acid sequences disclosed in Swiss-Prot sequence data.

[0204] 2.2 Method for Selecting Antiviral Peptides

[0205] According to one aspect of the present disclosure, a method for selecting an antiviral peptide is disclosed. The method for selecting an antiviral peptide includes selecting an antiviral peptide from the peptides selected by the method for screening a candidate group of antiviral peptides disclosed in the "2.1 Screening method for candidate group of antiviral peptides" section of the present invention. In this case, selecting an antiviral peptide refers to the process of measuring, confirming, determining, and / or experimenting on the antiviral effect of the selected peptide.

[0206] In this case, the process of measuring, confirming, determining, and / or experimenting on the antiviral effect may include at least one selected from the following:

[0207] Mixing a liposome (or a solution containing a liposome) with the selected peptide (or a solution containing the selected peptide), and confirming the damage or destruction of the liposome; or

[0208] Mixing a virus (or a solution containing a virus) with the selected peptide (or a solution containing the selected peptide), and confirming the damage or destruction of the viral phospholipid bilayer, or confirming the reduction or inactivation of the ability of the virus to infect cells.

[0209] In some embodiments, the size of the liposome (or the distance between one phosphate group moiety and the farthest phosphate group moiety in the phospholipids constituting the liposome) can be 5 nm, 6 nm, 7 nm, 8 nm, 9 nm, 10 nm, 20 nm, 30 nm, 40 nm, 50 nm, 60 nm, 70 nm, 80 nm, 90 nm, 100 nm, 150 nm, 200 nm, 250 nm, 300 nm, 350 nm, 400 nm, 500 nm, 600 nm, 700 nm, 800 nm, 900 nm, 1000 nm, 1100 nm, 1200 nm, 1300 nm, 1400 nm or 1500 nm, or a value between any two of these values (e.g., 55 nm), or can fall within a range defined by any two of these numerical values (e.g., 5 nm to 1500 nm).

[0210] In certain embodiments, the size of the liposome (or the distance between one phosphate group moiety and the farthest phosphate group moiety in the phospholipids constituting the liposome) can be from 20 nm to 400 nm.

[0211] In certain embodiments, the size of the liposome (or the distance between one phosphate group moiety and the farthest phosphate group moiety in the phospholipids constituting the liposome) can be 55 nm, 150 nm or 250 nm.

[0212] In certain embodiments, the virus can belong to the family Bunyaviridae, Coronaviridae, Filoviridae, Flaviviridae, Hepadnaviridae, Herpesviridae, Orthomyxoviridae, Poxviridae, Rhabdoviridae, Retroviridae, Togaviridae, or Herpesviridae. For example, the virus can be at least one virus selected from the following: Sin Nombre Hantavirus belonging to the family Bunyaviridae; coronaviruses belonging to the family Coronaviridae and involved in various acute respiratory syndromes; Ebola virus and Marburg virus belonging to the family Filoviridae; West Nile virus, yellow fever virus, dengue virus, and hepatitis C virus belonging to the family Flaviviridae; hepatitis B virus belonging to the family Hepadnaviridae; herpes simplex virus type 1 and herpes simplex virus type 2 belonging to the family Herpesviridae; influenza virus belonging to the family Orthomyxoviridae; variola virus, vaccinia virus, molluscum contagiosum virus, and monkeypox virus belonging to the family Poxviridae; rabies virus belonging to the family Rhabdoviridae; human immunodeficiency virus (HIV) belonging to the family Retroviridae; chikungunya virus belonging to the family Togaviridae; pseudorabies virus and human herpesvirus (HHV) belonging to the family Herpesviridae; and influenza virus belonging to the family Orthomyxoviridae.

[0213] In certain embodiments, the virus can be dengue virus or SARS-CoV-2 virus.

[0214] The present inventor has selected: a peptide represented by any one of the amino acid sequences selected from SEQ ID NO:1, SEQ ID NO:3 to 5, SEQ ID NO:8 to 23, SEQ ID NO:26, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:35, SEQ ID NO:36, SEQ ID NO:38 to 40, SEQ ID NO:43, SEQ ID NO:45 to 52, SEQ ID NO:55, SEQ ID NO:58, SEQ ID NO:59, SEQ ID NO:62, SEQ ID NO:64 to 73, SEQ ID NO:75, SEQ ID NO:78 to 87, SEQ ID NO:90 to 93, SEQ ID NO:97 to 102, SEQ ID NO:104 to 106, SEQ ID NO:109 to 112, SEQ ID NO:114 to 116, SEQ ID NO:118 to 123, SEQ ID NO:126, SEQ ID NO:127, SEQ ID NO:130 to 132, SEQ ID NO:136, SEQ ID NO:138 to 147, SEQ ID NO:151 to 155, SEQ ID NO:162, SEQ ID NO:164, SEQ ID NO:166 to 168, SEQ ID NO:170 to 174, SEQ ID NO:176, SEQ ID NO:180 to 182, SEQ ID NO:184 to 186, SEQ ID NO:188, SEQ ID NO:189, SEQ ID NO:191, SEQ ID NO:192, SEQ ID NO:194 to 196, SEQ ID NO:198, SEQ ID NO:201, SEQ ID NO:204, SEQ ID NO:206, SEQ ID NO:208 to 211, SEQ ID NO:215, SEQ ID NO:218, SEQ ID NO:219, SEQ ID NO:221, SEQ ID NO:224, SEQ ID NO:225, SEQ ID NO:228, SEQ ID NO:229, SEQ ID NO:231, SEQ ID NO:232, SEQ ID NO:236, SEQ ID NO:237, SEQ ID NO:240, SEQ ID NO:242 to 246, SEQ ID NO:248 to 254, SEQ ID NO:256 to 258, SEQ IDNO: 260 to 270, SEQ ID NO: 272, SEQ ID NO: 273, SEQ ID NO: 275 to 288, SEQ ID NO: 290, SEQ ID NO: 291, SEQ ID NO: 293 to 296, SEQ ID NO: 298 and SEQ ID NO: 299; a peptide represented by any amino acid sequence of SEQ ID NO: 1 to 300, as an antiviral peptide.

[0215] 3. Function of the antiviral peptide of the present disclosure

[0216] 3.1 Overview - Action of Antiviral Peptides

[0217] The antiviral peptide of the present disclosure causes damage to the outer membrane (or phospholipid bilayer) of the virus.

[0218] The antiviral peptide of the present disclosure can cause disruption of the outer membrane (or phospholipid bilayer) of the virus. Specifically, the antiviral peptide can cause disruption of the viral phospholipid bilayer by causing damage to the viral phospholipid bilayer.

[0219] The antiviral peptide of the present disclosure can cause a decrease or inactivation of the ability of the virus to infect cells. Specifically, the antiviral peptide can cause a decrease or inactivation of the ability of the virus to infect cells by causing damage (or disruption) to the phospholipid bilayer.

[0220] The antiviral peptide of the present disclosure can cause a decrease or inactivation of the ability of the virus to proliferate (or replicate). Specifically, the antiviral peptide can cause a decrease or inactivation of the ability of the virus to proliferate (or replicate) by causing damage (or disruption) to the phospholipid bilayer.

[0221] 3.2 Influence of Antiviral Peptides on Damage (or Disruption) of the Viral Outer Membrane

[0222] The antiviral peptide of the present disclosure causes damage to the outer membrane (or phospholipid bilayer) of the virus. In addition, the antiviral peptide can cause disruption of the outer membrane (or phospholipid bilayer) of the virus by causing damage to the outer membrane (or phospholipid bilayer) of the virus.

[0223] Here, the damage (or disruption) of the antiviral peptide to the outer membrane (or phospholipid bilayer) of the virus can be caused by the insertion of the antiviral peptide into the phospholipid bilayer of the virus. Those skilled in the art studying the antiviral peptides disclosed in this specification understand that the mechanism by which the antiviral peptide inserts into the viral phospholipid bilayer to damage (or disrupt) the viral phospholipid bilayer is related to the structural characteristics according to the virus size. A detailed explanation thereof is as follows.

[0224] The size of the virus enclosed by the outer membrane is much smaller than that of the cell. Due to this size difference, the phospholipid bilayer of the virus outer membrane has structural features that distinguish it from the phospholipid bilayer of the cell outer membrane. To explain in detail the structural features of the phospholipid bilayer of the virus outer membrane, for the convenience of calculation, it is assumed that the shapes of the virus and the cell are perfect spheres, and the differences are compared by the formula for calculating the surface area of a sphere.

[0225] First, the phospholipids that make up the phospholipid bilayer of the outer membrane can be divided into an inner layer and an outer layer. Here, the inner layer refers to the phospholipid layer in which the phosphate group part in the phospholipid bilayer faces the inside of the cell. The outer layer refers to the phospholipid layer in which the phosphate group part in the phospholipid bilayer faces the outside of the cell.

[0226] When the distance between any one phosphate part and the farthest phosphate part in the outer layer is called the diameter (=2R) and the length of the fatty acid part of the phospholipid is called a (assuming that the phosphate group part of the phospholipid has no volume and length),

[0227] (Surface area composed of phosphate group parts in the outer layer)=4×π×R 3

[0228] (Surface area composed of phosphate group parts in the inner layer)=4×π×(R - 2a) 3

[0229] (Surface area ratio)=(Surface area composed of phosphate group parts in the inner layer) / (Surface area composed of phosphate group parts in the outer layer)=(R - 2a) 3 / R 3 =(1-(2a / R)) 3 )。

[0230] In this case, the length of the fatty acid (a) is usually a fixed value, and the value of R varies greatly according to the size of the virus or the cell. Considering this, as the value of R increases, the surface area ratio approaches 1, and as the value of R decreases, the surface area ratio decreases. In other words, since the difference in the surface area ratio between the inner and outer layers of the virus with a low R value is larger than that of the cell with a high R value, the virus has a structure with a wider spacing between the phosphate groups on the outer layer (see Figure 2 )。

[0231] That is to say, the virus phospholipid bilayer can have a structure with gaps that do not exist in the cell or are larger than those in the cell. Considering these structural differences, it is expected that the antiviral peptides of the present disclosure will be able to insert into the virus phospholipid bilayer more easily than the phospholipid bilayer of the cell. In this case, the experimental results show that the antiviral peptides inserted into the virus phospholipid bilayer can cause damage or destruction of the phospholipid bilayer, but there is no exact known mechanism.

[0232] Thus, it is expected that the antiviral peptides of the present disclosure will only cause damage or disruption to the viral phospholipid bilayer and will not have a significant impact on cells. As described above, the reason is that the virus has a smaller total size (or diameter), so the outer membrane has a greater curvature than that of cells.

[0233] Through the above mechanism, the antiviral peptides of the present disclosure can not only cause damage or disruption to the phospholipid bilayer of a specific virus, but also cause general damage or disruption to the phospholipid bilayer of viruses with a phospholipid bilayer outer membrane.

[0234] It can be confirmed by the experimental methods and results disclosed in Experimental Examples 3 to 5 that the antiviral peptides of the present disclosure can cause damage or disruption to the viral phospholipid bilayer.

[0235] In addition, through the experiment using liposomes of different sizes in Experimental Example 4.2, it can be seen that the antiviral peptides of the present disclosure cause a high level of damage (or impairment) to the phospholipid bilayer of small-diameter liposomes. These results indicate that due to the size difference between cells and viruses, the antiviral peptides of the present disclosure can cause damage (or disruption) to the phospholipid bilayer of viruses other than cells.

[0236] 3.3 Target Viruses

[0237] In the present disclosure, the virus whose phospholipid bilayer is damaged, disrupted, etc. by the antiviral peptide is a virus with a phospholipid bilayer envelope (or membrane).

[0238] The virus can be a virus belonging to the Bunyaviridae, Coronaviridae, Filoviridae, Flaviviridae, Hepadnaviridae, Herpesviridae, Orthomyxoviridae, Poxviridae, Rhabdoviridae, Retroviridae, Togaviridae or Herpesviridae. For example, the virus can be at least one virus selected from the following: Sin Nombre virus of the genus Hantavirus belonging to the Bunyaviridae; coronaviruses belonging to the Coronaviridae and involved in various acute respiratory syndromes; Ebola virus and Marburg virus belonging to the Filoviridae; West Nile virus, yellow fever virus, dengue virus and hepatitis C virus belonging to the Flaviviridae; hepatitis B virus belonging to the Hepadnaviridae; herpes simplex virus type 1 and herpes simplex virus type 2 belonging to the Herpesviridae; influenza virus belonging to the Orthomyxoviridae; smallpox virus, vaccinia virus, molluscum contagiosum virus and monkeypox virus belonging to the Poxviridae; rabies virus belonging to the Rhabdoviridae; human immunodeficiency virus (HIV) belonging to the Retroviridae; Chikungunya virus belonging to the Togaviridae; pseudorabies virus and human herpesvirus (HHV) belonging to the Herpesviridae; and influenza virus belonging to the Orthomyxoviridae.

[0239] In certain embodiments, the virus can be dengue virus or SARS-CoV-2 virus.

[0240] 4. Use of the antiviral peptides of the present disclosure

[0241] 4.1 Overview of the Use of Antiviral Peptides of the Present Disclosure

[0242] According to one aspect of the present disclosure, the antiviral peptides of the present disclosure can be used to cause disruption or damage to the lipid bilayer of a virus. The disruption or damage to the lipid bilayer reduces the infectivity of the virus, which can be expected to have a prophylactic or therapeutic effect on virus infection (or related diseases). That is, the antiviral peptides can be used as effective therapeutic agents for virus infection (or related diseases) by inducing disruption or damage to the lipid bilayer. Here, the virus is a virus with an envelope (or membrane) having a phospholipid bilayer.

[0243] Various embodiments of the use of the antiviral peptides in combination with the treatment of virus infection (or related diseases) will be described below.

[0244] 4.2 Target diseases

[0245] The target diseases for which the antiviral peptides of the present disclosure can be used for therapeutic purposes are virus infection (or related diseases).

[0246] In some embodiments, the virus that is the target of lipid bilayer disruption (or damage) or treatment with the antiviral peptides can be a virus belonging to the following families: Bunyaviridae, Coronaviridae, Filoviridae, Flaviviridae, Hepadnaviridae, Herpesviridae, Orthomyxoviridae, Poxviridae, Rhabdoviridae, Retroviridae, Togaviridae or Herpesviridae. For example, the virus can be at least one virus selected from the following: Sin Nombre virus of the genus Hantavirus belonging to the Bunyaviridae; coronaviruses belonging to the Coronaviridae and involved in various acute respiratory syndromes; Ebola virus and Marburg virus belonging to the Filoviridae; West Nile virus, yellow fever virus, dengue virus and hepatitis C virus belonging to the Flaviviridae; hepatitis B virus belonging to the Hepadnaviridae; herpes simplex virus type 1 and herpes simplex virus type 2 belonging to the Herpesviridae; influenza virus belonging to the Orthomyxoviridae; variola virus, vaccinia virus, molluscum contagiosum virus and monkeypox virus belonging to the Poxviridae; rabies virus belonging to the Rhabdoviridae; human immunodeficiency virus (HIV) belonging to the Retroviridae; Chikungunya virus belonging to the Togaviridae; pseudorabies virus and human herpesvirus (HHV) belonging to the Herpesviridae; and influenza virus belonging to the Orthomyxoviridae.

[0247] In certain embodiments, the virus can be dengue virus or SARS-CoV-2 virus.

[0248] In certain embodiments, the target disease can be a respiratory disease caused by virus infection or coronavirus disease 2019 (COVID-19).

[0249] 4.3 Pharmaceutical Compositions Containing Antiviral Peptides

[0250] 4.3.1 Overview of Pharmaceutical Compositions

[0251] According to one aspect of the present disclosure, a pharmaceutical composition for treating or preventing viral diseases is disclosed. The pharmaceutical composition comprises a therapeutically effective amount of an antiviral peptide. In some instances, the pharmaceutical composition may further comprise a pharmaceutically acceptable carrier and / or adjuvant. Here, the pharmaceutically acceptable carrier can be used to appropriately formulate the pharmaceutical composition for treating or preventing viral diseases.

[0252] The antiviral peptide is the antiviral peptide detailedly disclosed in the section "1. Antiviral Peptides of the Present Disclosure" of this specification.

[0253] The viral disease refers to a disease caused by viral infection disclosed in the section "4.2 Target Diseases" of the present disclosure.

[0254] 4.3.2 Examples of Pharmaceutical Composition Formulations

[0255] As a method for formulating the pharmaceutical composition of the present disclosure, any known peptide formulation method can be used.

[0256] As an embodiment, the pharmaceutical composition can be formulated into lozenges, troches, tablets, aqueous suspensions, oily suspensions, prepared powders, granules, emulsions, hard capsules, soft capsules, syrups or elixirs. As another embodiment, the pharmaceutical composition can be formulated into injections, suppositories, powder aerosols for inhalation, aerosol sprays, ointments, dusting powders, oils or creams. As another embodiment, the pharmaceutical composition can be formulated into an injection. Specifically, a therapeutically effective amount of the antiviral peptide is mixed with a stabilizer or buffer in water to prepare a solution or suspension, and the solution or suspension can be formulated into a unit dosage form in an ampoule or vial. As another embodiment, the pharmaceutical composition can be formulated into an aerosol by mixing a propellant, etc. with additives to prepare a water-dispersible concentrate or wet powder. As another embodiment, when the pharmaceutical composition is formulated for transdermal use, a therapeutically effective amount of the antiviral peptide can be added to animal fats, vegetable fats, waxes, paraffins, starches, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonite, silica, talc, zinc oxide, etc. as a carrier to prepare ointments, creams, dusting powders, oils, topical skin preparations, etc.

[0257] 4.3.3 Examples of Pharmaceutically Acceptable Carriers

[0258] Pharmaceutically acceptable carriers are commonly used in formulations and include saline, sterile water, Ringer's solution, buffered saline, cyclodextrin, glucose solution, maltodextrin solution, glycerol, ethanol, liposomes, etc., but are not limited thereto, and may also include other typical additives, such as antioxidants and buffers, if needed. In addition, the pharmaceutical composition can be formulated into an injectable preparation, such as an aqueous solution, suspension and emulsion, pill, capsule, granule or tablet, by additionally adding a diluent, dispersant, surfactant, binder, lubricant, etc. Regarding suitable pharmaceutically acceptable carriers and formulations, the pharmaceutical composition can preferably be formulated according to the methods disclosed in "Remington's Pharmaceutical Sciences" (19th Edition, 1995) for each component. As an embodiment, the pharmaceutical composition can include, as a pharmaceutically acceptable carrier, binders such as lactose, sucrose, sorbitol, mannitol, starch, amylopectin, cellulose or gelatin; excipients such as dicalcium phosphate; disintegrants such as corn starch or sweet potato starch; lubricants such as magnesium stearate, calcium stearate, sodium stearyl fumarate or polyethylene glycol wax; sweeteners; flavoring agents; syrups; liquid carriers such as fatty oils; sterile aqueous solutions; propylene glycol; polyethylene glycol; injectable esters such as ethyl oleate; suspending agents; emulsions; freeze-dried preparations; topical preparations; stabilizers; buffers; animal oils; vegetable oils; waxes; paraffin wax; starch; tragacanth; cellulose derivatives; polyethylene glycol; silicone; bentonite; silica; talc; zinc oxide or any suitable combination thereof.

[0259] 4.4 Treatment Methods Using Antiviral Peptides

[0260] 4.4.1 Overview of the treatment method

[0261] According to one aspect of the present disclosure, a method for treating a viral disease is disclosed. The treatment method includes administering a pharmaceutical composition for treating a viral disease to a subject using a suitable administration method, a suitable dosage regimen and a suitable dose. In this case, administering to the subject can refer to an individual infected with a virus. In this case, the subject to be administered can be a human or a non-human animal.

[0262] The pharmaceutical composition for treating a viral disease is the pharmaceutical composition specifically disclosed in the "4.3. Pharmaceutical composition containing antiviral peptides" section of the present disclosure.

[0263] A viral disease refers to a disease caused by infection with a virus disclosed in the "4.2 Target diseases" section of the present disclosure.

[0264] 4.4.2 Examples of administration methods

[0265] The treatment methods of the present disclosure may involve administering a suitable formulation of a therapeutically effective antiviral peptide to a subject using a suitable administration method. In an embodiment, the treatment method may be achieved by administering a suitable formulation of a therapeutically effective antiviral peptide by one of the following methods: oral administration, parenteral administration, intravenous administration, intraperitoneal administration, intramuscular administration, transdermal administration, and subcutaneous administration.

[0266] 4.4.3 Examples of Dosages

[0267] The treatment methods of the present disclosure may involve administering a suitable formulation of a therapeutically effective antiviral peptide to a subject using a suitable administration method. As an embodiment, the dosage may be about 0.01 mg to 1000 mg of antiviral peptide / kg of subject body weight.

[0268] 4.4.4 Examples of Administration Cycles

[0269] The treatment methods of the present disclosure may involve administering a suitable formulation of a therapeutically effective antiviral peptide to a subject using a suitable administration method. As an embodiment, the pharmaceutical composition containing an appropriate dosage of the antiviral peptide may be administered once a day. As another embodiment, the administration cycle may be administering the pharmaceutical composition containing an appropriate dosage of the antiviral peptide at least twice a day. As another embodiment, the administration cycle may be administering the pharmaceutical composition containing an appropriate dosage of the antiviral peptide at intervals of 1 hour, 2 hours, 6 hours, 12 hours, 24 hours, 2 days, 3 days, 1 week, 2 weeks, 1 month, and / or 3 months.

[0270] 4.5 Therapeutic Use of Antiviral Peptides

[0271] According to one aspect of the present disclosure, the use of an antiviral peptide for the treatment of viral diseases is disclosed. In this case, the antiviral peptide refers to the antiviral peptide disclosed in the present disclosure. In this case, the viral disease refers to a disease caused by a viral infection disclosed in the "4.2 Target Diseases" section of the present disclosure.

[0272] 4.6 Use of Antiviral Peptides in the Preparation of Therapeutic Agents

[0273] According to one aspect of the present disclosure, the use of an antiviral peptide in the preparation of a drug for the treatment of viral diseases is disclosed. In this case, the antiviral peptide refers to the antiviral peptide disclosed in the present disclosure. In this case, the viral disease refers to a disease caused by a viral infection disclosed in the "4.2 Target Diseases" section of the present disclosure.

[0274] Possible Embodiments of the Present Invention

[0275] In the following text, possible embodiments of the present invention provided in this specification will be listed. The following embodiments provided in this section are merely examples of the present invention. Therefore, the invention provided in this specification should not be construed as being limited to the following embodiments. The brief descriptions numbered with the embodiments are also for the convenience of distinguishing the embodiments and should not be construed as a limitation on the invention disclosed in this specification.

[0276] Antiviral Peptides

[0277] Embodiment 1. Antiviral peptide 1

[0278] An antiviral peptide, which is represented by the amino acid sequence of any one of the following amino acid sequences or an amino acid sequence having 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98 or 99% or higher homology (sequence identity) thereto:

[0279] SEQ ID NO:1, SEQ ID NO:3 to 5, SEQ ID NO:8 to 23, SEQ ID NO:26, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:35, SEQ ID NO:36, SEQ ID NO:38 to 40, SEQ ID NO:43, SEQ ID NO:45 to 52, SEQ ID NO:55, SEQ ID NO:58, SEQ ID NO:59, SEQ ID NO:62, SEQ ID NO:64 to 73, SEQ ID NO:75, SEQ ID NO:78 to 87, SEQ ID NO:90 to 93, SEQ ID NO:97 to 102, SEQ ID NO:104 to 106, SEQ ID NO:109 to 112, SEQ ID NO:114 to 116, SEQ ID NO:118 to 123, SEQ ID NO:126, SEQ ID NO:127, SEQ ID NO:130 to 132, SEQ ID NO:136, SEQ ID NO:138 to 147, SEQ ID NO:151 to 155, SEQ ID NO:162, SEQ ID NO:164, SEQ ID NO:166 to 168, SEQ ID NO:170 to 174, SEQ ID NO:176, SEQ ID NO:180 to 182, SEQ ID NO:184 to 186, SEQ ID NO:188, SEQ ID NO:189, SEQ ID NO:191, SEQ ID NO:192, SEQ ID NO:194 to 196, SEQ ID NO:198, SEQ ID NO:201, SEQ ID NO:204, SEQ ID NO:206, SEQ ID NO:208 to 211, SEQ ID NO:215, SEQ ID NO:218, SEQ ID NO:219, SEQ ID NO:221, SEQ ID NO:224, SEQ ID NO:225, SEQ ID NO:228, SEQ ID NO:229, SEQ ID NO:231, SEQ ID NO:232, SEQ ID NO:236, SEQ ID NO:237, SEQ ID NO:240, SEQ ID NO:242 to 246, SEQ ID NO:248 to 254, SEQ ID NO:256 to 258, SEQ ID NO:260 to 270, SEQ ID NO:272, SEQ IDNO: 273, SEQ ID NO: 275 to 288, SEQ ID NO: 290, SEQ ID NO: 291, SEQ ID NO: 293 to 296, SEQ ID NO: 298, and SEQ ID NO: 299.

[0280] Embodiment 2. Antiviral Peptide 2

[0281] The antiviral peptide of Embodiment 1, wherein the amino acid sequence of the antiviral peptide is the amino acid sequence of any one of the following amino acid sequences:

[0282] SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 20, SEQ ID NO: 29, SEQ ID NO: 30, SEQ ID NO: 46, SEQ ID NO: 48, SEQ ID NO: 49, SEQ ID NO: 52, SEQ ID NO: 59, SEQ ID NO: 78, SEQ ID NO: 82, SEQ ID NO: 87, SEQ ID NO: 90, SEQ ID NO: 91, SEQ ID NO: 98, SEQ ID NO: 99, SEQ ID NO: 101, SEQ ID NO: 106, SEQ ID NO: 110, SEQ ID NO: 112, SEQ ID NO: 114, SEQ ID NO: 115, SEQ ID NO: 116, SEQ ID NO: 123, SEQ ID NO: 126, SEQ ID NO: 131, SEQ ID NO: 132, SEQ ID NO: 139, SEQ ID NO: 141, SEQ ID NO: 143, SEQ ID NO: 144, SEQ ID NO: 151, SEQ ID NO: 162, SEQ ID NO: 167, SEQ ID NO: 171, SEQ ID NO: 189, SEQ ID NO: 221, SEQ ID NO: 228, SEQ ID NO: 242, SEQ ID NO: 245, SEQ ID NO: 246, SEQ ID NO: 254, SEQ ID NO: 256, SEQ ID NO: 264, SEQ ID NO: 283, SEQ ID NO: 290, and SEQ ID NO: 296.

[0283] Embodiment 3. Antiviral Peptide 3

[0284] The antiviral peptide of Embodiment 1, wherein the amino acid sequence of the antiviral peptide is the amino acid sequence of any one of the following amino acid sequences:

[0285] SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:20, SEQ ID NO:46, SEQ ID NO:48, SEQ ID NO:49, SEQ ID NO:59, SEQ ID NO:87, SEQ ID NO:90, SEQ ID NO:91, SEQ ID NO:98, SEQ ID NO:101, SEQ ID NO:110, SEQ ID NO:114, SEQ ID NO:116, SEQ ID NO:126, SEQ ID NO:132, SEQ ID NO:141, SEQ ID NO:143, SEQ ID NO:144, SEQ ID NO:151, SEQ ID NO:221, SEQ ID NO:246, SEQ ID NO:256, SEQ ID NO:264, SEQ ID NO:283, SEQ ID NO:290, and SEQ ID NO:296.

[0286] Embodiment 4. Antiviral peptide 4

[0287] The antiviral peptide of Embodiment 1, wherein the amino acid sequence of the antiviral peptide is the amino acid sequence of any one of the following amino acid sequences:

[0288] SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:46, SEQ ID NO:49, SEQ ID NO:101, SEQ ID NO:143, SEQ ID NO:185, and SEQ ID NO:191.

[0289] Embodiment 5. Antiviral peptide 5

[0290] The antiviral peptide of Embodiment 1, wherein the amino acid sequence of the antiviral peptide is the amino acid sequence of any one of the following amino acid sequences:

[0291] SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:46, SEQ ID NO:49, SEQ ID NO:101, SEQ ID NO:110, SEQ ID NO:114, SEQ ID NO:116, SEQ ID NO:143, SEQ ID NO:151, SEQ ID NO:171, and SEQ ID NO:221.

[0292] Structure and Characteristics of Antiviral Peptides

[0293] Embodiment 6. Structure of Antiviral Peptide 1

[0294] The antiviral peptide according to any one of Embodiments 1 to 5, wherein the antiviral peptide is capable of forming an α-helical structure.

[0295] Embodiment 7. Structure of Antiviral Peptide 2

[0296] The antiviral peptide according to any one of Embodiments 1 to 6, wherein when the antiviral peptide reacts with a lipid membrane, the antiviral peptide is capable of forming an α-helical structure.

[0297] Embodiment 8. Characteristics of Antiviral Peptide 1

[0298] The antiviral peptide according to any one of Embodiments 1 - 7, wherein the antiviral peptide is an amphiphilic peptide.

[0299] Embodiment 9. Characteristics of Antiviral Peptide 2

[0300] The antiviral peptide according to any one of Embodiments 1 to 8, wherein when the antiviral peptide forms an α-helical structure, the antiviral peptide has amphiphilic characteristics.

[0301] Action of Antiviral Peptides

[0302] Embodiment 10. Function of Antiviral Peptide 1

[0303] The antiviral peptide according to any one of Embodiments 1 to 9, wherein the antiviral peptide is capable of causing damage to the outer membrane (or lipid bilayer) of the virus.

[0304] Embodiment 11. Function of Antiviral Peptide 2

[0305] The antiviral peptide according to any one of Embodiments 1 to 10, wherein the antiviral peptide is capable of causing disruption of the outer membrane (or lipid bilayer) of the virus.

[0306] Embodiment 12. Function of Antiviral Peptide 3

[0307] The antiviral peptide according to any one of Embodiments 1 to 11, wherein the antiviral peptide can reduce or inactivate the ability of a virus to infect cells.

[0308] Embodiment 13. Function of antiviral peptide 4

[0309] The antiviral peptide according to any one of Embodiments 1 to 12, wherein the antiviral peptide can reduce or inactivate the ability of a virus to proliferate (or replicate).

[0310] Embodiment 14. Function of antiviral peptide 5

[0311] The antiviral peptide according to any one of Embodiments 10 to 13, wherein the virus belongs to the Bunyaviridae, Coronaviridae, Filoviridae, Flaviviridae, Hepadnaviridae, Herpesviridae, Orthomyxoviridae, Poxviridae, Rhabdoviridae, Retroviridae, Togaviridae or Herpesviridae.

[0312] Use of Antiviral Peptides

[0313] Embodiment 15. Pharmaceutical composition containing antiviral peptide

[0314] A pharmaceutical composition for treating viral diseases, comprising a therapeutically effective amount of an antiviral peptide,

[0315] wherein the antiviral peptide is the antiviral peptide according to any one of Embodiments 1 to 14, and

[0316] the viral disease is caused by virus infection.

[0317] Embodiment 16. Pharmaceutical composition containing antiviral peptide

[0318] The pharmaceutical composition according to Embodiment 15, wherein the pharmaceutical composition further comprises a pharmaceutically acceptable carrier and / or a pharmaceutically acceptable adjuvant.

[0319] Embodiment 17. Pharmaceutical composition containing antiviral peptide

[0320] The pharmaceutical composition according to any one of Embodiments 15 and 16, wherein the viral disease is caused by virus infection with a virus belonging to the Bunyaviridae, Coronaviridae, Filoviridae, Flaviviridae, Hepadnaviridae, Herpesviridae, Orthomyxoviridae, Poxviridae, Rhabdoviridae, Retroviridae, Togaviridae or Herpesviridae.

[0321] Embodiment 18. Therapeutic method using antiviral peptide

[0322] A method for treating viral diseases, comprising:

[0323] Administering to a subject infected with a virus a pharmaceutical composition comprising a therapeutically effective amount of an antiviral peptide,

[0324] wherein the antiviral peptide is the antiviral peptide according to any one of Embodiments 1 to 14.

[0325] Embodiment 19. Therapeutic use of antiviral peptides

[0326] Use of the antiviral peptide according to any one of Embodiments 1 to 14 for the treatment of viral diseases.

[0327] Embodiment 20. Use of antiviral peptides in the preparation of therapeutic agents

[0328] Use of the antiviral peptide according to any one of Embodiments 1 - 14 in the preparation of a medicament for the treatment of viral diseases.

[0329] Method for Selecting Antiviral Peptides

[0330] Embodiment 21. Method for selecting antiviral peptide 1

[0331] A method for screening a group of antiviral peptide candidates, comprising selecting, finding, and / or searching for a peptide that meets at least one of the following conditions:

[0332] 1) The peptide consists of 18 amino acid residues;

[0333] 2) The peptide does not contain proline residues;

[0334] 3) The peptide contains at least 6 polar amino acid residues;

[0335] 4) The peptide contains 4 to 6 charged amino acid residues;

[0336] 5) The peptide has a hydrophobic moment (μH) value of 0.5 to 0.8 measured by HeliQuest software;

[0337] 6) The peptide has a hydrophobicity (H) value of 0.7 to 0.9 measured by HeliQuest software; and

[0338] 7) The peptide has a net charge of -2 to 0 measured by HeliQuest software.

[0339] Embodiment 22. Method for selecting antiviral peptide 2

[0340] The method according to Embodiment 21, further comprising a process of verifying the antiviral efficacy of the selected peptide.

[0341] Embodiment 23. Method for selecting antiviral peptide 3

[0342] The method according to Embodiment 22, wherein verifying the antiviral efficacy of the selected peptide includes at least one selected from the following:

[0343] Mixing the liposome with the selected peptide and confirming the damage or destruction of the liposome; or

[0344] Mixing the virus with the selected peptide and confirming the damage or destruction of the viral phospholipid bilayer, or confirming the reduction or inactivation of the virus's ability to infect cells.

[0345] Embodiment 24. Method for selecting antiviral peptide 4

[0346] The method according to Embodiment 23, wherein the size of the liposome is 5 nm to 1500 nm.

[0347] Hereinafter, the invention provided in this specification will be described in more detail with reference to the following experimental examples. These experimental examples are only for illustrative purposes of the content disclosed in this specification, and it is obvious to those skilled in the art that the scope of the content disclosed in this specification should not be construed as being limited by these experimental examples.

[0348] Experimental Example 1. Selection of peptides according to the embodiments of the present disclosure

[0349] The inventors selected 300 peptides from the peptides disclosed in the Swiss-Prot sequence data, and these peptides 1) consist of 18 amino acid residues, 2) do not contain proline residues among the 18 amino acid residues, 3) contain at least 6 polar amino acid residues among the 18 amino acid residues, 4) contain 4 to 6 charged amino acid residues among the 18 amino acid residues, 5) have a hydrophobic moment (μH) value of 0.5 to 0.8 measured by the HeliQuest software, 6) have a hydrophobicity (H) value of 0.7 to 0.9 measured by the HeliQuest software; and 7) have a net charge of -2 to 0 measured by the HeliQuest software. In this case, the polar amino acid residues are composed of aspartic acid (Asp, D), glutamic acid (Glu, E), lysine (Lys, K), arginine (Arg, R), histidine (His, H), serine (Ser, S), threonine (Thr, T), asparagine (Asn, N), glutamine (Gln, Q), and glycine (Gly, G), and the charged amino acid residues are composed of aspartic acid (Asp, D), glutamic acid (Glu, E), lysine (Lys, K), and arginine (Arg, R).

[0350] Information on the amino acid sequences, hydrophobicity values, and hydrophobic moment values of the selected 300 peptides (SP-001 to SP-300) is disclosed in Table 1 below.

[0351] [Table 1]

[0352]

[0353]

[0354]

[0355]

[0356]

[0357]

[0358]

[0359]

[0360]

[0361] Experimental Example 2. Preparation of Peptides According to Embodiments of the Present Disclosure

[0362] The selected peptides SP-001 to SP-300 were synthesized by ANYGEN. Each peptide was synthesized by the general solid-phase peptide synthesis (SPPS) method.

[0363] The Fmoc protecting group was removed by reacting with 20% piperidine in DMF for 10 minutes (twice), and coupling was carried out using Fmoc amino acid (6 equivalents), HOBT (6 equivalents), HBTU (6 equivalents), and DIPEA (12 equivalents). At each step, the resin was washed with DMF and methanol (repeated twice).

[0364] The synthesized crude peptide was reacted with a mixture of TFA / EDT / anisole / TIS / DW (volume 90 / 2.5 / 2.5 / 2.5) for 2 hours to remove the resin and amino acid protecting groups. Ether was added to the resulting mixed solution, and then centrifuged to recover the peptide, which was then lyophilized.

[0365] The crude peptide was dissolved in DW and purified by reverse-phase HPLC using a C18 reverse-phase column. In this case, separation was carried out using a water-acetonitrile linear gradient method (acetonitrile concentration 10 to 75% (v / v)) with 0.1% (v / v) trifluoroacetic acid. Thereafter, the purified fraction was lyophilized.

[0366] Experimental Example 3. Confirmation of the Liposome Leakage Efficacy of Peptides According to Embodiments of the Present Disclosure

[0367] The inventors of the present application attempted to use a liposome solution encapsulating a fluorescent substance to confirm the antiviral efficacy (or lipid membrane disruption efficacy) of the peptide prepared in Experimental Example 2 (specifically, utilizing the property that the fluorescence level to be measured is relatively low when the fluorescent substance is encapsulated in the liposome, while the fluorescence level to be measured is relatively high when the fluorescent substance leaks out of the liposome).

[0368] As a detailed experimental method for confirming the efficacy, a solution containing the peptide prepared in Experimental Example 2 was treated with a liposome solution encapsulating a fluorescent substance, and then the change in the fluorescence level was observed. In this case, data showing an increase in the fluorescence level indicates that the fluorescent substance has leaked out of the liposome, and such leakage indicates damage or disruption of the liposome (specifically, the lipid membrane).

[0369] The detailed experimental method and results are shown in the following Experimental Examples 3.1 and 3.2.

[0370] Experimental Example 3.1 Preparation of Liposomes Encapsulating 6-Carboxyfluorescein

[0371] 2.5 μmol of 1-palmitoyl-2-oleoyl-glycero-3-phosphocholine (POPC) was formed into a film by completely removing the organic solvent in a rotary evaporator for 16 hours, and the fluorescent substance 6-carboxyfluorescein (6CF) was prepared at a concentration of 100 mM by dissolving it in a buffer solution (10 mM Tris, 150 mM NaCl, pH 7.5). After dissolving the formed POPC film in 1 mL of the 100 mM 6CF solution to prepare a POPC solution with a concentration of 2.5 mM, it was repeatedly frozen and thawed 7 times by liquid nitrogen and a constant temperature water bath at 60 °C.

[0372] The POPC solution was made into liposomes by a microextruder. The extruder was equipped with a 1 μm filter membrane and the POPC solution was passed through the extruder 15 times, and then the extruder was equipped with a 30 nm filter and the solution was passed through the extruder 15 times, thereby preparing liposomes with a uniform size (55 nm). The remaining fluorescent substance not encapsulated in the liposomes was removed using a PD-10 desalting column.

[0373] Experimental Example 3.2 Liposome Leakage Assay

[0374] Peptides SP-001 to SP-300 were separately dissolved in DMSO at a concentration of 200 μM and diluted to a concentration of 500 nM with a buffer solution. Also, a liposome solution encapsulating a fluorescent substance was diluted to a POPC concentration of 20 μM in the buffer solution. The diluted peptides, 0.06% Triton X-100 as a positive control (PC), and the buffer solution as a negative control (N.C.) were each dispensed in triplicate at 50 μL into three wells of a 96-well black plate. In this case, the positive control 0.06% Triton X-100 is a solution containing 0.06% of the surfactant Triton X-100, which is known to disrupt lipid membranes.

[0375] Thereafter, immediately after dispensing 50 μL of the diluted liposome solution into each well, the fluorescence value (RFU) was measured at 1-minute intervals for 20 minutes using a plate reader at an excitation wavelength of 492 nm and an emission wavelength of 517 nm.

[0376] After measuring fluorescence for 20 minutes, 50 μL of 0.06% Triton X-100 was added to all wells, and then fluorescence was measured again. In this case, 0.06% Triton X-100 was added to all wells after 20 minutes, and fluorescence was measured again to confirm 1) that the increase in fluorescence level over time in each well was due to the disruption of liposomes, and 2) that there was no significant difference in the total amount of fluorescent substance contained in each well.

[0377] The measured fluorescence levels were graphed and shown in Figures 3 to 25 .

[0378] According to Figures 3 to 25From the results shown, it can be seen that due to SP-001, SP-003 to SP-005, SP-008 to SP-023, SP-026, SP-029, SP-030, SP-033, SP-035, SP-036, SP-038 to SP-040, SP-043, SP-045 to SP-052, SP-055, SP-058, SP-059, SP-062, SP-064 to SP-073, SP-075, SP-078 to SP-087, SP-090 to SP-093, SP-097 to SP-102, SP-104 to SP-106, SP-109 to SP-112, SP-114 to SP-116, SP-118 to SP-123, SP-126, SP-127, SP-130 to SP-132, SP-136, SP-138 to SP-147, SP-151 to SP-155, SP-162, SP-164, SP-166 to SP-168, SP-170 to SP-174, SP-176, SP-180 to SP-182, SP-184 to SP-186, SP-188, SP-189, SP-191, SP-192, SP-194 to SP-196, SP-198, SP-201, SP-204, SP-206, SP-208 to SP-211, SP-215, SP-218, SP-219, SP-221, SP-224, SP-225, SP-228, SP-229, SP-231, SP-232, SP-236, SP-237, SP-240, SP-242 to SP-246, SP-248 to SP-254, SP-256 to SP-258, SP-260 to SP-270, SP-272, SP-273, SP-275 to SP-288, SP-290, SP-291, SP-293 to SP-296, SP-298 and SP-299, the fluorescent substance encapsulated in the liposome leaks from the liposome. These results indicate that the antiviral peptides of the present disclosure can cause damage (or disruption) to the lipid bilayer of the virus.

[0379] Experimental Example 4. Liposome disruption efficacy of the peptides according to the embodiments of the present disclosure

[0380] Experimental Example 4.1 Experiments on Liposomes of Specific Sizes

[0381] The present inventors attempted to confirm the antiviral activity (or lipid membrane disruption activity) of the peptides prepared in Experimental Example 2 using liposomes in which the lipid membrane was labeled with a fluorescent substance.

[0382] As a detailed experimental method for confirming the efficacy, when the present inventors treated a coverslip attached with liposomes having a lipid membrane labeled with a fluorescent substance with a solution containing the peptide prepared in Experimental Example 2 at a constant rate, a change in the fluorescence level was observed.

[0383] In this case, the data showing that the fluorescence level decreases over time indicate that the lipid membrane of the liposome is disrupted and the disrupted lipid membrane is removed during the washing process. In other words, from the results of this experiment, it can be confirmed whether the peptide prepared in Experimental Example 2 can cause liposome rupture.

[0384] The detailed experimental method and results are shown in Experimental Example 4.1.1 and Experimental Example 4.1.2 below.

[0385] Experimental Example 4.1.1 Preparation of Rhodamine Fluorescently Labeled Liposomes

[0386] 2.49 μmol of 1-palmitoyl-2-oleoyl-glycero-3-phosphocholine (POPC), 2.51 nmol of 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[biotinyl(polyethylene glycol)-2000] (DSPE-PEG(2000) biotin), and 17.58 nmol of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine-N-(lissamine rhodamine B sulfonyl) (18:1 LissRhod PE) were mixed for 16 hours and the organic solvent was completely removed in a rotary evaporator for 16 hours to form a film. In this case, biotin is used to bind to streptavidin on the plate, and rhodamine is used as the fluorescent substance.

[0387] After dissolving the phospholipid solution forming the film in 1 mL of a buffer solution (10 mM Tris, 150 mM NaCl, pH 7.5) to prepare a phospholipid solution with a concentration of 2.5 mM, it was repeatedly frozen and thawed 7 times using liquid nitrogen and a constant temperature water bath at 60 °C. The phospholipid solution was passed through a small extruder equipped with a 30 nm filter membrane 15 times to prepare liposomes with a uniform size (55 nm).

[0388] Experimental Example 4.1.2 TIRF Microscopy Measurement

[0389] A biotin-coated coverslip was attached to a channel-type adhesive slide (IBIDI, catalog number 80608), treated with 100 μl of 10 μg / mL neutravidin diluted with a buffer solution, and then allowed to react for 10 minutes. The unreacted neutravidin was removed, and after washing the coverslip 5 times, the prepared liposomes were diluted to a phospholipid concentration of 0.03 μM. After treating and incubating with 100 μl of the diluted liposomes for 10 minutes, the liposomes were attached to the coverslip. Then the unattached liposomes were removed and then washed 5 times.

[0390] Thereafter, SP-001, SP-003, and SP-101 peptides were separately dissolved in DMSO at a concentration of 200 μM and diluted with buffer to prepare solutions at a concentration of 100 nM. Using a peristaltic pump, the diluted peptides were delivered to a coverslip with attached liposomes at a rate of 150 μL / min, and under a total internal reflection fluorescence microscope (TIRF microscope), the change in the fluorescence signal of individual liposomes was measured at 20-second intervals in the TRITC channel for 25 minutes. The measurement was repeated three times, and for each measurement, the change in the fluorescence signal of 200 to 300 liposomes was measured. The fluorescence value at 0 seconds was converted to 1, and this change was calculated and graphed.

[0391] The change in the observed fluorescence signal is as Figure 26 shown, and the graph showing the change in the fluorescence signal is as Figures 27 to 29 shown. Figure 27 The results of the SP-001 peptide are shown. Figure 28 The results of the SP-003 peptide are shown. Figure 29 The results of the SP-101 peptide are shown.

[0392] Figures 26 to 29 The experimental results in

[0393] Experimental Example 4.2 Experiments on Liposomes of Various Sizes

[0394] showed that the lipid membranes of liposomes labeled with a fluorescent substance were disrupted by SP-001, SP-003, and SP-101 peptides, resulting in a decrease in the fluorescence value. These results indicate that the antiviral peptides of the present disclosure have antiviral efficacy (or lipid membrane disruption efficacy).

[0395] Therefore, the present inventors conducted the same experiment as in Example 4.1, in which liposomes of various sizes were used. The experimental method and results are as follows.

[0396] Experimental Example 4.2.1 Preparation of Rhodamine Fluorescently Labeled Liposomes

[0397] Mix 2.49 μmol of 1-palmitoyl-2-oleoyl-glycero-3-phosphocholine (POPC), 2.51 nmol of 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[biotinyl(polyethylene glycol)-2000] (DSPE-PEG(2000) biotin), and 17.58 nmol of 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine-N-(lissamine rhodamine B sulfonyl) (18:1 LissRhod PE) for 16 hours, and form a film by completely removing the organic solvent in a rotary evaporator for 16 hours. After dissolving the phospholipid solution forming the film in 1 mL of buffer solution (10 mM Tris, 150 mM NaCl, pH 7.5) to prepare a phospholipid solution with a concentration of 2.5 mM, repeat freezing and thawing 7 times through liquid nitrogen and a constant temperature water bath at 60 °C.

[0398] Pass the phospholipid solution through a microextruder to prepare liposomes with three sizes. The phospholipid solution is passed through an extruder equipped with a 30 nm filter membrane 15 times to prepare liposomes with a uniform size (55 nm), through an extruder equipped with a 200 nm filter membrane 11 times to prepare liposomes with a uniform size (150 nm), and through an extruder equipped with a 400 nm filter membrane 15 times to prepare liposomes with a uniform size (250 nm).

[0399] Experimental Example 4.2.2 TIRF Microscope Measurement

[0400] Attach a biotin-coated coverslip to a channel-type adhesive slide (IBIDI, catalog number 80608), treat it with 100 μl of 10 μg / mL neutral avidin diluted with buffer solution, and then allow it to react for 10 minutes. Remove the unreacted neutral avidin. After washing the coverslip 5 times, dilute the prepared liposomes to a phospholipid concentration of 0.03 μM. After treating and incubating with 100 μl of diluted liposomes for 10 minutes, attach the liposomes to the coverslip. Then remove the unattached liposomes and wash 5 times.

[0401] Thereafter, dissolve the SP-049 peptide prepared in Experimental Example 2 in DMSO at a concentration of 200 μM, and dilute it with buffer solution to prepare a peptide with a diluted concentration of 100 nM. Using a peristaltic pump, deliver the diluted peptide to the coverslip with attached liposomes at a rate of 150 μL / min, and measure the change in the fluorescence signal of individual liposomes at 20-second intervals in the TRITC channel under a total internal reflection fluorescence microscope (TIRF microscope) for 25 minutes. Repeat the measurement three times, and for the fluorescence signal changes of 200 to 300 liposomes for each measurement, convert the fluorescence value at 0 seconds to 1, calculate and graph this change.

[0402] The graph showing the observed changes in fluorescence signals is as shown in Figure 53 as follows. Figure 53 The results of the SP-049 peptide are shown.

[0403] Figure 53 The experimental results in show that the lipid membrane of the liposome labeled with a fluorescent substance is disrupted by the SP-049 peptide, resulting in a decrease in the fluorescence value. These results indicate that the antiviral peptide of the present disclosure has antiviral efficacy (or lipid membrane disruption efficacy).

[0404] Figure 53 The experimental results in show that for liposome particles with the smallest diameter (maximum curvature), the peptide has the greatest phospholipid bilayer disrupting effect, and for liposome particles with the largest diameter (minimum curvature), the peptide has the smallest phospholipid bilayer disrupting effect.

[0405] Therefore, the results of this experiment indicate that the phospholipid bilayer disrupting effect of the antiviral peptide of the present disclosure depends on the curvature of the lipid membrane. The greater the curvature of the target particle with a phospholipid bilayer (the smaller the diameter of the target particle), the greater the phospholipid bilayer disrupting effect of the antiviral peptide of the present disclosure. These results are considered due to the following phenomenon: the greater the curvature of the phospholipid bilayer of the particle, the greater the stress applied to the lipid membrane, and when the antiviral peptide of the present disclosure is inserted into the phospholipid bilayer structure under conditions of greater stress, the structure of the lipid membrane is easily disrupted.

[0406] These results mean that the antiviral peptide of the present disclosure is very effective in damaging (or disrupting) the phospholipid bilayer of the virus (referring to the size or diameter of the virus is usually about 20 nm to 200 nm), but has an insignificant effect on the phospholipid bilayer of other cells (referring to the size or diameter of animal cells is usually about 10 to 30 μm).

[0407] Experimental Example 5. RNase Digestion Assay of Peptides According to Embodiments of the Present Disclosure

[0408] The present inventors attempted to use a nuclease that can enter the virus through pores in the lipid membrane of the virus and degrade RNA to confirm the antiviral efficacy (or lipid membrane disruption efficacy) of the peptide prepared in Experimental Example 2.

[0409] As an experimental method for confirming the efficacy, the present inventors treated dengue virus with the peptide, incubated the treated dengue virus with micrococcal nuclease (NEB, catalog number M0247S), and then confirmed the RNA level of the virus. In this case, data showing a low level of viral RNA means that the lipid membrane of the virus is disrupted and a large amount of RNA is degraded by the RNase that enters the virus. In other words, the lower the viral RNA level, the better the antiviral efficacy of the peptide.

[0410] The detailed experimental methods and results are as follows.

[0411] At 1×107 Dengue virus stored at a concentration of pfu / ml was diluted to a concentration of 4×10 5 pfu / ml in a buffer solution (50 mM Tris-HCl, 5 mM CaCl2). Thereafter, 50 selected peptides were each dissolved in DMSO at a concentration of 200 μM and diluted with the buffer to prepare peptide solutions at concentrations of 2 μM and 1 μM. The diluted peptide solutions, 2% Triton X-100 solution as a positive control, and buffer solution as a negative control were each aliquoted in triplicate at 50 μL into three microtubes (at each concentration), and the diluted virus solution (or control solution) was mixed at a ratio of 1:1 (v / v), and then incubated at 37 °C for 1 hour. After incubation, each microtube was treated with 14 μg BSA and 960 gel units of micrococcal nuclease (NEB, catalog number M0247S) to a final volume of 140 μL and incubated at 37 °C for 1 hour. In this case, micrococcal nuclease (NEB, catalog number M0247S) was used to degrade the RNA of the virus. In other words, when the lipid membrane of the virus is disrupted, the RNA of the virus will be degraded by the micrococcal nuclease (NEB, catalog number M0247S) that enters the virus. Conversely, when the lipid membrane of the virus is not disrupted, the micrococcal nuclease (NEB, catalog number M0247S) will not enter the virus, and thus the RNA of the virus will not be degraded.

[0412] Viral RNA was extracted from the incubated mixed solution using the QIAamp Viral RNA Kit (Qiagen; catalog number 52906), and each Ct value was confirmed by RT-qPCR. Using the One Step PrimeScript TM III RT-qPCR Mix (TaqMan) (Takara; catalog number #RR600A) and using the forward primer 5'-GAAAGACCAGAGATCCTGCTGTCT-3' (SEQ ID NO:302), reverse primer 5'-ACCATTCCATTTTCTGGCGTT-3' (SEQ ID NO:303), probe 5'-FAM-AGCATCATTCCAGGCAC-3BHQ1-3' (AGCATCATTCCAGGCAC (SEQ ID NO:304)) for RT-qPCR, and the confirmation was carried out by: 42 °C for 30 minutes, 95 °C for 5 minutes, 95 °C for 30 seconds, and then 60 °C for 60 seconds, repeating 45 cycles.

[0413] The results were analyzed by calculating the increase in Ct value (ΔCt) of the Ct value after peptide treatment compared to the Ct value of the negative control. Each value was obtained from two replicate experiments (in duplicate). In this case, a high ΔCt value means that the lipid membrane of the virus was disrupted and a large amount of RNA was degraded by RNase that entered the virus. In other words, the higher the ΔCt value, the better the antiviral efficacy of the peptide.

[0414] The experimental results are shown in Table 2 and Figure 30 as follows.

[0415] The experimental results showed that the peptides SP-116, SP-046, SP-049, SP-110, SP-001, SP-143, SP-141, SP-003, SP-151, SP-114, SP-101, SP-059, SP-126, SP-296, SP-004, SP-020, SP-132, SP-264, SP-087, SP-090, SP-290, SP-283, SP-221, SP-048, SP-091, SP-144, SP-171, SP-246, SP-098, SP-256, SP-228, SP-242, SP-078, SP-254, SP-030, SP-139, SP-029, SP-052, SP-115, SP-106, SP-131, SP-162, SP-123, SP-245, SP-167, SP-099, SP-082, SP-189, SP-112, and SP-005 disrupted the lipid membrane of the virus and the RNA of the virus was degraded by RNase that entered the virus.

[0416] In other words, it was shown that the antiviral peptides of the present disclosure have antiviral efficacy (or lipid membrane disruption efficacy).

[0417] [Table 2]

[0418]

[0419]

[0420] Experimental Example 6. Confirmation of the antiviral efficacy (against dengue virus) of the peptides according to the embodiments of the present disclosure

[0421] The present inventors attempted to confirm the antiviral efficacy (or lipid membrane disruption efficacy) of the peptides prepared in Experimental Example 2. For this purpose, after co-culturing the virus and the peptide, the cells were treated with the culture solution, and the RNA level of the virus cultured with the cells was measured. In this case, the lower the measured RNA level, the better the antiviral efficacy of the peptide.

[0422] The detailed experimental methods and results are shown in the following Experimental Examples 6.1 and 6.2.

[0423] Experimental Example 6.1 Evaluation at Specific Concentrations

[0424] Vero cells were seeded into 96-well plates at a density of 1.5×10 4 cells per well. The seeded cells were cultured overnight.

[0425] Thirty selected peptides were each dissolved in DMSO at a concentration of 200 μM and serially diluted 2-fold in MEM medium to prepare peptide solutions diluted to concentrations of 120 nM and 60 nM.

[0426] When treating the cells with dengue virus, the dengue virus was diluted to an MOI of 0.03 to prepare a diluted virus solution. The diluted peptide solution and the diluted virus solution were mixed at a ratio of 1:1 (v / v), and then incubated at 37 °C for 1 hour to prepare a mixed solution of virus and peptide.

[0427] The seeded cells were treated with 200 μL of each prepared mixed solution of virus and peptide and cultured at 37 °C and 5% CO2 for 1 hour. After removing the mixed solution, 200 μL of MEM medium containing 10% FBS was added, and the cells were cultured at 37 °C and 5% CO2 for 2 days.

[0428] Viral RNA was extracted from the incubated mixed solution using the QIAamp Viral RNA Kit (Qiagen; catalog number 52906), and each Ct value was confirmed by RT-qPCR. Using the One Step PrimeScript TM III RT-qPCR Mix (TaqMan) (Takara; catalog number #RR600A) and using the forward primer 5'-GAAAGACCAGAGATCCTGCTGTCT-3' (SEQ ID NO:302), reverse primer 5'-ACCATTCCATTTTCTGGCGTT-3' (SEQ ID NO:303), probe 5'-FAM-AGCATCATTCCAGGCAC-3BHQ1-3' (AGCATCATTCCAGGCAC (SEQ ID NO:304)) for RT-qPCR, and the confirmation was carried out by: 42 °C for 30 minutes, 95 °C for 5 minutes, 95 °C for 30 seconds, then 60 °C for 60 seconds, repeating 45 cycles.

[0429] The results were analyzed by calculating the increase in Ct value (ΔCt) of the Ct value after peptide treatment compared to the Ct value of the negative control. Each value was obtained from two replicate experiments (in duplicate).

[0430] The experimental results are shown in Table 3 andFigure 31 as shown in

[0431] The experimental results show that peptides SP-049, SP-143, SP-046, SP-116, SP-114, SP-110, SP-283, SP-151, SP-221, SP-256, SP-003, SP-048, SP-004, SP-101, SP-141, SP-290, SP-098, SP-132, SP-246, SP-090, SP-020, SP-059, SP-001, SP-087, SP-091, SP-144, SP-264, SP-296, and SP-126 attack the virus (e.g., disrupt the lipid membrane of the virus), thereby reducing the amount of virus in infected cells compared to the negative control.

[0432] In other words, it is shown that the antiviral peptides of the present disclosure have antiviral efficacy (or lipid membrane disruption efficacy).

[0433] [Table 3]

[0434]

[0435]

[0436] Experimental Example 6.2 Derivation of IC50 Value

[0437] Vero cells were seeded into a 96-well plate at a density of 1.5×10 4 cells per well. The seeded cells were cultured overnight.

[0438] Nine selected peptides were each dissolved in DMSO at a concentration of 4 mM and serially diluted 2-fold in MEM medium to prepare solutions diluted to 8 concentrations.

[0439] Peptide SP-049 was prepared such that the maximum concentration was 125 nM and the minimum concentration was 0.98 nM (concentrations: 125 nM; 62.5 nM; 31.25 nM; 15.62 nM; 7.81 nM; 3.91 nM; 1.95 nM; and 0.98 nM).

[0440] Peptides SP-046 and SP-143 were prepared such that the maximum concentration was 250 nM and the minimum concentration was 1.95 nM (concentrations: 250 nM; 125 nM; 62.5 nM; 31.25 nM; 15.62 nM; 7.81 nM; 3.91 nM; and 1.95 nM).

[0441] Prepare SP-001, SP-003, SP-004, and SP-101 peptides such that the maximum concentration is 2000 nM and the minimum concentration is 15.63 nM (concentrations: 2000 nM, 1000 nM, 500 nM, 250 nM, 125 nM, 62.5 nM, 31.25 nM, and 15.63 nM).

[0442] Prepare SP-185 and SP-191 peptides such that the maximum concentration is 10000 nM and the minimum concentration is 78.13 nM (concentrations: 10000 nM; 5000 nM; 2500 nM; 1250 nM; 625 nM; 312.5 nM; 156.25 nM; and 78.13 nM).

[0443] When treating cells with dengue virus, dilute the dengue virus to an MOI of 0.03 to prepare a diluted virus solution. Mix the diluted peptide solution and the diluted virus solution at a ratio of 1:1 (v / v), and then incubate at 37 °C for 1 hour to prepare a mixed solution of virus and peptide. Treat the inoculated cells with 200 μL of each prepared mixed solution of virus and peptide, and culture at 37 °C and 5% CO2 for 1 hour. After removing the mixed solution, add 200 μL of MEM medium containing 10% FBS, and culture the cells at 37 °C and 5% CO2 for 2 days.

[0444] Extract viral RNA from the incubated mixed solution using the QIAamp Viral RNA Kit (Qiagen; catalog number 52906), and confirm each Ct value by RT-qPCR. Use the one-step PrimeScript TM III RT-qPCR Mix (TaqMan) (Takara; catalog number #RR600A) and perform RT-qPCR using the forward primer 5'-GAAAGACCAGAGATCCTGCTGTCT-3' (SEQ ID NO:302), reverse primer 5'-ACCATTCCATTTTCTGGCGTT-3' (SEQ ID NO:303), probe 5'-FAM-AGCATCATTCCAGGCAC-3BHQ1-3' (AGCATCATTCCAGGCAC (SEQ ID NO:304)), and confirm by: 42 °C for 30 minutes, 95 °C for 5 minutes, 95 °C for 30 seconds, then 60 °C for 60 seconds, repeating for 45 cycles.

[0445] The present inventors converted the average Ct value of the wells infected with dengue virus without peptide treatment in the same orifice plate into 100% infection rate, and made a standard curve therefrom. The inhibition rate (%) of each well was calculated according to the standard curve formula. Each value was obtained through three repeated experiments (in triplicate), and the IC50 value was obtained using GraphPad Prism 5 software. It can be determined that the lower the IC50 value, the better the antiviral effect.

[0446] The IC50 (nM) value of SP-049 peptide was 3.85.

[0447] The IC50 (nM) value of SP-001 peptide was 22.60.

[0448] The IC50 (nM) value of SP-046 peptide was 17.68.

[0449] The IC50 (nM) value of SP-143 peptide was 16.09.

[0450] The IC50 (nM) value of SP-003 peptide was 21.99.

[0451] The IC50 (nM) value of SP-004 peptide was 206.6.

[0452] The IC50 (nM) value of SP-101 peptide was 100.1.

[0453] The IC50 (nM) value of SP-185 peptide was 518.9.

[0454] The IC50 (nM) value of SP-191 peptide was 776.2.

[0455] The experimental results are shown in Tables 4 to 12 and Figures 32 to 40 as shown below.

[0456] The experimental results showed that SP-049, SP-046, SP-143, SP-001, SP-003, SP-004, SP-101, SP-185 and SP-191 peptides attacked the virus (e.g., disrupted the lipid membrane of the virus), thereby reducing the amount of virus in the infected cells compared to the negative control.

[0457] In other words, it was shown that the antiviral peptides of the present disclosure had antiviral efficacy (or lipid membrane disruption efficacy).

[0458] [Table 4]

[0459]

[0460] [Table 5]

[0461]

[0462] [Table 6]

[0463]

[0464] [Table 7]

[0465]

[0466] [Table 8]

[0467]

[0468] [Table 9]

[0469]

[0470] [Table 10]

[0471]

[0472] [Table 11]

[0473]

[0474] [Table 12]

[0475]

[0476] Experimental Example 7. Confirmation of the Antiviral Efficacy (SARS-CoV2 Virus) of Peptides According to Embodiments of the Present Disclosure

[0477] The present inventors attempted to confirm the antiviral efficacy (or lipid membrane disruption efficacy) of the peptide prepared in Experimental Example 2. For this purpose, after co-culturing the virus and the peptide, the cells were treated with the culture solution and cultured, and then the proportion of infected cells was measured. In this case, the higher the measured proportion of normal cells not infected with the virus, the better the antiviral efficacy of the peptide.

[0478] The detailed experimental methods and results are as follows.

[0479] Vero cells were seeded into a 96-well plate at a density of 1.5×10 4 cells per well. The seeded cells were cultured overnight.

[0480] Twelve selected peptides were each dissolved in DMSO at a concentration of 4 mM, diluted 1 / 100 with DPBS, and then serially diluted 2-fold to prepare peptide solutions with a maximum concentration of 40 μM and a minimum concentration of 0.08 μM (concentrations: 40 μM; 20 μM; 10 μM; 5 μM; 2.5 μM; 1.25 μM; 0.63 μM; 0.31 μM; 0.16 μM; and 0.08 μM). The diluted peptide solutions were mixed with 200 TCID50 of SARS-CoV-2 virus at a ratio of 1:1 (v / v), and then incubated at 37 °C for 30 minutes to prepare mixed solutions. 40 μL of each concentration of cells was seeded into four wells, treated with the mixed solutions, and cultured at 37 °C and 5% CO2 for 4 days. After culturing, the cells were fixed with 4% paraformaldehyde and stained with crystal violet solution, and the neutralization activity (%) was calculated by observing the cytopathic effect (CPE). The neutralization activity was calculated by normalizing with a positive control (mock) and a negative control (0.5% DMSO), and the 50% inhibitory concentration (IC50) was calculated.

[0481] The IC50 (μM) value of SP-049 peptide was 1.8.

[0482] The IC50 (μM) value of SP-116 peptide was 4.1.

[0483] The IC50 (μM) value of SP-151 peptide was 5.0.

[0484] The IC50 (μM) value of SP-046 peptide was 7.9.

[0485] The IC50 (μM) value of SP-004 peptide was 8.9.

[0486] The IC50 (μM) value of SP-003 peptide was 10.3.

[0487] The IC50 (μM) value of SP-101 peptide was 11.6.

[0488] The IC50 (μM) value of SP-171 peptide was 18.7.

[0489] The IC50 (μM) value of SP-110 peptide was 19.5.

[0490] The IC50 (μM) value of SP-114 peptide was 19.5.

[0491] The IC50 (μM) value of SP-143 peptide was 19.5.

[0492] The IC50 (μM) value of SP-221 peptide was 19.5.

[0493] The experimental results are as Figures 41 to 52 shown.

[0494] The experimental results show that peptides SP-049, SP-116, SP-151, SP-046, SP-004, SP-003, SP-101, SP-171, SP-110, SP-114, SP-143, and SP-221 attack the virus (e.g., disrupt the lipid membrane of the virus), thereby reducing the level of virus-infected cells.

[0495] In other words, it is shown that the antiviral peptides of the present disclosure have antiviral efficacy (or lipid membrane disruption efficacy).

Claims

1. An antiviral peptide having an amino acid sequence selected from the group consisting of the following sequences: SEQ ID NO:1, SEQ ID NO:3 to 5, SEQ ID NO:8 to 23, SEQ ID NO:26, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:33, SEQ ID NO:35, SEQ ID NO:36, SEQ ID NO:38 to 40, SEQ ID NO:43, SEQ ID NO:45 to 52, SEQ ID NO:55, SEQ ID NO:58, SEQ ID NO:59, SEQ ID NO:62, SEQ ID NO:64 to 73, SEQ ID NO:75, SEQ ID NO:78 to 87, SEQ ID NO:90 to 93, SEQ ID NO:97 to 102, SEQ ID NO:104 to 106, SEQ ID NO:109 to 112, SEQ ID NO:114 to 116, SEQ ID NO:118 to 123, SEQ ID NO:126, SEQ ID NO:127, SEQ ID NO:130 to 132, SEQ ID NO:136, SEQ ID NO:138 to 147, SEQ ID NO:151 to 155, SEQ ID NO:162, SEQ ID NO:164, SEQ ID NO:166 to 168, SEQ ID NO:170 to 174, SEQ ID NO:176, SEQ ID NO:180 to 182, SEQ ID NO:184 to 186, SEQ ID NO:188, SEQ ID NO:189, SEQ ID NO:191, SEQ ID NO:192, SEQ ID NO:194 to 196, SEQ ID NO:198, SEQ ID NO:201, SEQ ID NO:204, SEQ ID NO:206, SEQ ID NO:208 to 211, SEQ ID NO:215, SEQ ID NO:218, SEQ ID NO:219, SEQ ID NO:221, SEQ ID NO:224, SEQ ID NO:225, SEQ ID NO:228, SEQ ID NO:229, SEQ ID NO:231, SEQ ID NO:232, SEQ ID NO:236, SEQ ID NO:237, SEQ ID NO:240, SEQ ID NO:242 to 246, SEQ ID NO:248 to 254, SEQ ID NO:256 to 258, SEQ ID NO:260 to 270, SEQ ID NO:272, SEQ IDNO: 273, SEQ ID NO: 275 to 288, SEQ ID NO: 290, SEQ ID NO: 291, SEQ ID NO: 293 to 296, SEQ ID NO: 298, and SEQ ID NO:

299.

2. The antiviral peptide according to claim 1, Among them, wherein the amino acid sequence of the antiviral peptide is selected from the following sequences: SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:20, SEQ ID NO:29, SEQ ID NO:30, SEQ ID NO:46, SEQ ID NO:48, SEQ ID NO:49, SEQ ID NO:52, SEQ IDNO:59, SEQ ID NO:78, SEQ ID NO:82, SEQ ID NO:87, SEQ ID NO:90, SEQ ID NO:91, SEQ IDNO:98, SEQ ID NO:99, SEQ ID NO:101, SEQ ID NO:106, SEQ ID NO:110, SEQ ID NO:112, SEQ ID NO:114, SEQ IDNO:115, SEQ ID NO:116, SEQ ID NO:123, SEQ ID NO:126, SEQ IDNO:131, SEQ ID NO:132, SEQ ID NO:139, SEQ ID NO:141, SEQ ID NO:143, SEQ ID NO:144, SEQ ID NO:151, SEQ ID NO:162, SEQ ID NO:167, SEQ ID NO:171, SEQ ID NO:189, SEQ IDNO:221, SEQ ID NO:228, SEQ ID NO:242, SEQ ID NO:245, SEQ ID NO:246, SEQ ID NO:254, SEQ ID NO:256, SEQ ID NO:264, SEQ ID NO:283, SEQ ID NO:290 and SEQ ID NO:

296.

3. The antiviral peptide according to claim 1, Among them, wherein the amino acid sequence of the antiviral peptide is selected from the following sequences: SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:20, SEQ ID NO:46, SEQ ID NO:48, SEQ ID NO:49, SEQ ID NO:59, SEQ ID NO:87, SEQ ID NO:90, SEQ ID NO:91, SEQ ID NO:98, SEQ ID NO:101, SEQ ID NO:110, SEQ ID NO:114, SEQ ID NO:116, SEQ ID NO:126, SEQ ID NO:132, SEQ ID NO:141, SEQ ID NO:143, SEQ ID NO:144, SEQ ID NO:151, SEQ ID NO:221, SEQ ID NO:246, SEQ ID NO:256, SEQ ID NO:264, SEQ ID NO:283, SEQ ID NO:290 and SEQ ID NO:

296.

4. The antiviral peptide according to claim 1, Among them, wherein the amino acid sequence of the antiviral peptide is selected from the following sequences: SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:46, SEQ ID NO:49, SEQ ID NO:101, SEQ ID NO:143, SEQ ID NO:185 and SEQ ID NO:

191.

5. The antiviral peptide according to claim 1, Among them, wherein the amino acid sequence of the antiviral peptide is selected from the following sequences: SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:46, SEQ ID NO:49, SEQ ID NO:101, SEQ ID NO:110, SEQ ID NO:114, SEQ ID NO:116, SEQ ID NO:143, SEQ ID NO:151, SEQ ID NO:171 and SEQ ID NO:

221.

6. The antiviral peptide according to claim 1, Among them, wherein the antiviral peptide is an amphiphilic peptide.

7. The antiviral peptide according to claim 1, Among them, wherein the antiviral peptide is capable of causing damage and / or disruption of the lipid bilayer of the virus.

8. The antiviral peptide according to claim 7, Among them, The virus belongs to the family Bunyaviridae, Coronaviridae, Filoviridae, Flaviviridae, Hepadnaviridae, Herpesviridae, Orthomyxoviridae, Poxviridae, Rhabdoviridae, Retroviridae, Togaviridae or Herpesviridae.

9. The antiviral peptide according to claim 1, Among them, The antiviral peptide is capable of forming an α-helical structure.

10. A pharmaceutical composition for treating viral diseases, comprising a therapeutically effective amount of an antiviral peptide, Among them, The antiviral peptide is the antiviral peptide according to any one of claims 1-9, and wherein the viral disease is caused by a viral infection.

11. The pharmaceutical composition according to claim 10, Among them, The pharmaceutical composition further comprises a pharmaceutically acceptable carrier and / or a pharmaceutically acceptable adjuvant.

12. The pharmaceutical composition according to claim 10, Among them, The viral disease is a respiratory disease caused by a viral infection or coronavirus disease 2019 (COVID-19).

13. A method for treating viral diseases, comprising: administering to a subject infected with a virus a pharmaceutical composition comprising a therapeutically effective amount of an antiviral peptide, wherein the antiviral peptide is the antiviral peptide according to any one of claims 1-9.

14. Use of the antiviral peptide according to any one of claims 1-9 for treating viral diseases.

15. Use of the antiviral peptide according to any one of claims 1-9 in the preparation of a drug for treating viral diseases.

16. A method for screening a group of candidates for antiviral peptides, comprising selecting peptides that meet the following conditions: 1) The peptide consists of 18 amino acid residues; 2) The peptide does not contain proline residues; 3) The peptide contains at least 6 polar amino acid residues; 4) The peptide contains 4 to 6 charged amino acid residues; 5) The peptide has a hydrophobic moment (μH) value of 0.5 to 0.8 measured by HeliQuest software; 6) The peptide has a hydrophobicity (H) value of 0.7 to 0.9 measured by HeliQuest software; and 7) The peptide has a net charge of -2 to 0 measured by HeliQuest software.

17. The method according to claim 16, Among them, The polar amino acid residues are aspartic acid (Asp, D), glutamic acid (Glu, E), lysine (Lys, K), arginine (Arg, R), histidine (His, H), serine (Ser, S), threonine (Thr, T), asparagine (Asn, N), glutamine (Gln, Q) and glycine (Gly, G). Among them, the charged amino acid residues are aspartic acid (Asp, D), glutamic acid (Glu, E), lysine (Lys, K) and arginine (Arg, R).

18. The method according to claim 16 Among them, The method is characterized in that a group of candidates for antiviral peptides is selected from the amino acid sequences disclosed in Swiss-Prot sequence data.

19. A method for selecting antiviral peptides, comprising:[[]]END]] Verifying the antiviral efficacy of the peptides selected by the method for screening a group of candidates for antiviral peptides according to claim 16.

20. The method according to claim 19 Among them, Verifying the antiviral efficacy of the selected peptides includes at least one selected from the following:[[]]END]] Mixing liposomes with the selected peptides and confirming the damage or disruption of the liposomes; or Mixing a virus with the selected peptides, and confirming the damage or disruption of the phospholipid bilayer of the virus, or confirming the reduction or inactivation of the infectivity of the virus to cells.

21. The method according to claim 20 Among them, The size of the liposomes is from 5 nm to 1500 nm.

22. The method according to claim 21 Among them, The size of the liposomes is from 20 nm to 400 nm.

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