Pharmaceutical composition for preventing or treating corovirus infective diseases, containing cell permeable peptide-

Abstract

The present invention provides the use of a PNA oligomer comprising a modified PNA for the prevention or treatment of coronavirus infectious diseases. The PNA oligomer of the present invention can be delivered into a virus-infected cell without a delivery vehicle and inhibits viral proliferation, and thus does not have side effects caused by a delivery vehicle for increasing cell permeability of a conventional drug, and can be used as a target-specific drug due to its high binding affinity. Virus proliferation can be inhibited by binding to a target even if there is a mismatch. Therefore, it is expected that the PNA oligomer will be used as an effective coronavirus therapeutic agent that can respond to rapidly mutated viral infectious diseases.

Description

TECHNICAL FIELD

[0001] The present invention relates to a pharmaceutical composition for preventing or treating a coronavirus infectious disease, the pharmaceutical composition comprising, as an active ingredient, a peptide-nucleic acid oligomer targeting the genomic positive (+) and / or negative (-) strand RNA of a coronavirus. BACKGROUND

[0002] Coronaviruses generally cause endemic infections in animals (e.g., birds and mammals), but they can also cross the animal-human species barrier, causing zoonotic infections in humans. Coronaviruses cause a variety of diseases in mammals, birds, and humans, including respiratory, gastrointestinal, and central nervous system infections, and over the past 50 years, a variety of coronaviruses causing diseases in humans and animals have emerged. Coronaviruses also cause a variety of diseases in domestic animals, particularly those affecting agriculture, and after acute respiratory infections were first reported in chickens raised as domestic animals in North America in the late 1920s, Edward C. Kendall, Malcolm Bynoe, and David Tyrrell identified coronaviruses as one of the causes of the common cold in animals in 1961 and named them Coronavirus B814. After many studies, coronaviruses were first discovered in the United Kingdom in 1965 as one of the causes of respiratory infections in humans, and virologist June Almeida and physician David Tyrrell named them coronaviruses in 1967 and published this discovery in the journal Nature under the name of coronaviruses in 1968. Since the 1970s, they have been studied for about 20 years using a model of beta coronaviruses, murine hepatitis virus (MHV), and human alpha coronavirus HCoV-229E. Human circulating coronaviruses (HCoV) HCoV-229E, HCoV-NL63, HCoV-OC43, and HCoV-HKU1 generally cause mild upper respiratory tract disease and are overall associated with 10 to 30% of common cold patients. Since the outbreak of severe acute respiratory syndrome (SARS) in 2002, coronaviruses that were previously considered only as one of the causes of the common cold have formed a variety of mutations of Middle East respiratory syndrome (MERS), as well as SARS-CoV-2, which now causes severe acute respiratory syndrome type 2 (COVID-19), causing new infectious diseases such as severe infectious pneumonia in humans.

[0003] Coronaviruses (CoV) are spherical, enveloped RNA viruses, about 80 nm to 125 nm in diameter, whose envelope is derived from the host cell's plasma membrane. The viral genome is present in a helical capsid formed by nucleocapsid proteins (N) and is protected by the envelope. The envelope has three prominent viral structural proteins: spike protein (S), membrane protein (M), and envelope protein (E), and includes the nucleocapsid protein inside the membrane.

[0004] The coronavirus genome is non-segmented, single-stranded, positive-sense RNA, 26 to 32 kilobases in size, making it the largest genome among all RNA viruses.

[0005] Meanwhile, antiviral drugs treat viral infectious diseases by weakening or eliminating the effects of viruses that invade the human body. Antiviral drugs are classified into methods of directly recognizing and attacking viral proteins, methods of inhibiting each stage of the viral life cycle to prevent the viral replication process, methods of enhancing immunity, etc.

[0006] An antisense oligonucleotide (ASO) that specifically binds to the viral genome can inhibit the synthesis of viral proteins and inhibit their proliferation, but the ASO consisting of DNA and / or RNA has low stability and is easily degraded by nucleases present in the body, limiting its therapeutic use, and there is also a problem in that a delivery vehicle is required to introduce it into cells.

[0007] The present inventors have conducted extensive research to develop a coronavirus infectious disease therapeutic agent using a peptide-nucleic acid derivative that can move into cells without a delivery vehicle and has high stability, and as a result, the present invention has been completed. SUMMARY

[0008] TECHNICAL PROBLEM

[0009] The technical problem to be achieved by the present invention is to provide a PNA oligomer comprising a modified PNA derivative for preventing and / or treating a coronavirus infectious disease.

[0010] However, the technical problem to be achieved by the present invention is not limited to the above-mentioned problem, and other problems not mentioned can be clearly understood by those skilled in the art from the following description.

[0011] TECHNICAL SOLUTION

[0012] To solve the above-mentioned problems, the present invention provides a pharmaceutical composition for preventing or treating a coronavirus infectious disease, the pharmaceutical composition comprising a peptide nucleic acid (PNA) oligomer represented by the following Chemical Formula 1 or a pharmaceutically acceptable salt thereof as an active ingredient.

[0013] [Chemical Formula 1]

[0014]

[0015] In the present application, the PNA oligomer can be complementarily bound to a part of the genome (+) RNA or genome (-) RNA of the coronavirus, and the length is 10-mer to 30-mer.

[0016] Specifically, in Chemical Formula 1, n is an integer between 10 and 30;

[0017] X is one selected from the group consisting of hydrogen [H], formyl [H-C(=O)-], aminocarbonyl [NH2-C(=O)-], substituted or unsubstituted alkyl, substituted or unsubstituted aryl, substituted or unsubstituted alkylacyl, substituted or unsubstituted arylacyl, substituted or unsubstituted alkoxycarbonyl, substituted or unsubstituted aryloxycarbonyl, substituted or unsubstituted alkylaminocarbonyl, substituted or unsubstituted arylaminocarbonyl, substituted or unsubstituted alkylsulfonyl, and substituted or unsubstituted arylsulfonyl;

[0018] Z is one selected from the group consisting of hydroxyl [-OH], substituted or unsubstituted alkoxy, substituted or unsubstituted aryloxy, amino [-NH2], substituted or unsubstituted alkylamino, substituted or unsubstituted arylamino, substituted or unsubstituted alkyl, and substituted or unsubstituted aryl;

[0019] B1, B2,..., B n-1 and B n are each independently selected from natural nucleotides of adenine (A), guanine (G), thymine (T), or cytosine (C) and unnatural nucleotides;

[0020] one or more of B1 to B n are modified PNA, and specifically, one or more of B1 to B n are each independently selected from the group consisting of modified PNA of Chemical Formula 2 to Chemical Formula 4 below;

[0021] [Chemical Formula 2]

[0022]

[0023] [Chemical Formula 3]

[0024]

[0025] [Chemical Formula 4]

[0026]

[0027] In Chemical Formula 2 to Chemical Formula 4, j, k, l, and m are each independently an integer between 1 and 16.

[0028] Meanwhile, the pharmaceutical composition of the present application does not include a delivery vehicle for intracellular delivery of the PNA oligomer.

[0029] In one embodiment of the present application, in Chemical Formula 1, X can be Fmoc [(9-fluorenyl) methoxy] carbonyl], Fethoc [2-(9-fluorenyl) ethyl-1-oxyl] carbonyl], acetyl (Ac), benzoyl, or hydrogen, and more specifically, can be Fethoc-.

[0030] In another embodiment of the present application, in Chemical Formula 1, X can be Fethoc, Z can be NH2, j can be 2, k can be 1, l can be 5, and m can be 6.

[0031] In still another embodiment of the present application, the coronavirus can be one or more selected from the group consisting of human coronavirus 229E (HCoV-229E), human coronavirus OC43 (HCoV-OC43), severe acute respiratory syndrome coronavirus (SARS-CoV), human coronavirus NL63 (HCoV-NL63), human coronavirus HKU1 (HCoV-HKU1), Middle East respiratory syndrome coronavirus (MERS-CoV), and severe acute respiratory syndrome coronavirus type 2 (SARS-Cov-2).

[0032] Since the PNA oligomers complementary to HCoV-OC43 RNA in Table 1 below were constructed and the virus proliferation inhibitory effect thereof was confirmed in the present application, in still another embodiment of the present application, the PNA oligomer can be one or more selected from the group consisting of the PNA oligomers in Table 1, and the coronavirus can be HCoV-OC43, but the target virus is not limited thereto.

[0033] Since the PNA oligomers complementary to SARS-CoV-2 RNA in Table 2 below were constructed and the virus proliferation inhibitory effect thereof was confirmed in the present application, in still another embodiment of the present application, the PNA oligomer can be one or more selected from the group consisting of the PNA oligomers in Table 2, and the coronavirus can be SARS-CoV-2, but the target virus is not limited thereto.

[0034] Meanwhile, since the PNA oligomer was randomly selected from the PNA oligomers of Table 1 to confirm whether it can be applied to inhibit the proliferation of the SARS-CoV-2 virus, and the result confirmed that the selected PNA oligomer forms a complementary binding region including a partial mismatch with the SARS-CoV-2 genome and inhibits the expression of the SARS-CoV-2 protein, in still another embodiment of the present application, the PNA oligomer can have 80% or more complementarity to a part of the genome (+) RNA or genome (-) RNA of the coronavirus.

[0035] Further, the present application provides a method of preventing or treating a coronavirus infectious disease and a method of inhibiting the proliferation of a coronavirus, the method including administering the above-described PNA oligomer to an individual.

[0036] In one embodiment of the present application, the individual can be a mammal infected with a coronavirus or at risk of exposure to a coronavirus.

[0037] In another embodiment of the present application, each of the methods can further include detecting a coronavirus in a biological sample isolated from the individual after the PNA oligomer is administered.

[0038] Further, the present application provides a method of inhibiting the proliferation of a coronavirus, the method including treating a cell with the above-described PNA oligomer.

[0039] In one embodiment of the present application, the cell can be a mammalian cell.

[0040] In another embodiment of the present application, the cell can be a coronavirus-infected cell.

[0041] In still another embodiment of the present application, the method can further include lysing the cell after the PNA oligomer treatment and detecting coronavirus genetic material.

[0042] In still another embodiment of the present application, the method can be used to separately prepare a cell not treated with the PNA oligomer as a control group to confirm the effect of the PNA oligomer treatment on the inhibition of the proliferation of the coronavirus, and the cell of the control group can be the same cell line as the cell treated with the PNA oligomer.

[0043] In still another embodiment of the present application, the method can further include lysing the cell treated with the PNA oligomer and the cell of the same cell line not treated with the PNA oligomer, respectively, and quantifying the level of the coronavirus genetic material.

[0044] Further, the present application provides the use of the above-described PNA oligomer for the preparation of a medicament for preventing or treating a coronavirus.

[0045] Beneficial effects

[0046] The present application provides the use of a PNA oligomer comprising a modified PNA for preventing or treating a coronavirus infectious disease. The PNA oligomer of the present application can be delivered into a virus-infected cell without a delivery vehicle and inhibit viral proliferation, and thus has no side effects caused by a delivery vehicle for increasing the cellular penetration of a conventional drug, and can inhibit viral proliferation by binding to a target even in the presence of a mismatch due to its high binding affinity to the target. Therefore, it is expected that the PNA oligomer will be used as an effective coronavirus therapeutic agent that can respond to rapidly mutating virus infectious diseases. BRIEF DESCRIPTION OF DRAWINGS

[0047] Figures la to lc Some chromatogram results confirming that the designed PNA oligomer was constructed as intended are shown. Specifically, Figure la C 18 -reverse phase UPLC reverse phase chromatogram results, Figure lb C 18 -reverse phase UPLC reverse phase chromatogram results, and Figure lc C 18 ESI-TOF mass spectrum results of PNA123 after RP preparative HPLC.

[0048] Figure 2 Results of titer analysis of Paxlovid in SARS-CoV-2 virus are shown.

[0049] Figures 3a to 3i and Figures 4a to 4k Results of titer analysis of PNA oligomers and combination drugs thereof in SARS-CoV-2 virus are shown.

[0050] Figure 5 Results of Western blot analysis confirming the effect of inhibiting SARS-CoV-2 virus proliferation using a PNA oligomer complementary to a part of HCoV-OC43 RNA are shown. DETAILED DESCRIPTION

[0051] Coronaviruses are single-stranded RNA viruses that synthesize genomic positive (+) RNA into genomic negative (-) RNA using an RNA polymerase to construct double-stranded RNA. The present inventors intended to block the replication process of coronaviruses using an oligonucleotide targeting the coronavirus genome.

[0052] However, since oligonucleotides composed of DNA and / or RNA have low stability and are easily degraded by nucleases present in organisms, there are limitations in their use for treating viral infectious diseases. To overcome this, various types of unnatural oligonucleotides have been developed, and the present inventors have developed a new peptide nucleic acid (PNA) in previous research.

[0053] PNA is a polypeptide having a backbone of N-(2-aminoethyl)glycine [Aeg] units, invented by Nielsen et al. PNA can bind to a complementary nucleic acid like DNA and RNA, and the binding affinity of RNA-PNA is stronger than that of RNA-DNA or RNA-RNA. In addition, since PNA is very different in structure from DNA, it is not recognized by hepatobiliary transporters that recognize DNA.

[0054] The PNA monomer has structure, and in the PNA monomer structure, B is a nucleobase, which can be a natural or unnatural nucleobase, and examples of the natural or unnatural nucleobase in the present application are as follows.

[0055]

[0056]

[0057] In previous research, the present inventors constructed PNA using a "modified nucleobase" in which a cationic lipid was covalently linked, and confirmed that PNA containing the modified nucleobase has high cell membrane permeability, thereby developing PNA having increased cell permeability (PCT / KR2009 / 001256). However, there was no research at all on whether the modified PNA can be used to treat viral infectious diseases.

[0058] The modified nucleobase used to construct PNA having increased cell permeability in the present invention is as follows.

[0059]

[0060]

[0061]

[0062] The modified nucleobase belongs to a non-natural nucleobase, and in the present application document, the term "modified nucleobase" is used interchangeably with the term "non-natural nucleobase" unless it is intended to be distinguished from a non-natural nucleobase other than the modified nucleobase. Also, in the present application document, when it is necessary to distinguish a PNA comprising a modified nucleotide from a PNA not comprising the modified nucleotide, the PNA comprising the modified nucleotide is referred to as a "modified PNA".

[0063] The present inventors constructed a PNA oligomer of the following Chemical Formula 1 in which the above PNA monomers are covalently linked.

[0064] [Chemical Formula 1]

[0065]

[0066] The PNA oligomer of Chemical Formula 1 is designed to be able to complementarily bind to a part of the coronavirus genome RNA of a coronavirus, and the complementary binding region is set to 10 or more.

[0067] Accordingly, in Chemical Formula 1, n is an integer between 10 and 30. This literally means that n is an integer selected from 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, and 30.

[0068] In the present application document, the "complementary binding region" refers to a continuous region including the start and end of the bases forming base pairs between the coronavirus genome RNA and the PNA oligomer. At the same time, since the modified PNA has a high binding affinity to the target gene, even if there is a mismatch in the complementary binding region, the PNA can bind to the target and inhibit viral replication. Accordingly, in the present application document, "complementary binding" includes complete complementary binding as well as partial complementary binding having 20% or less of mismatches within the range in which the PNA derivative can maintain the binding affinity to the target RNA. For example, when n = 10 in Chemical Formula 1, the PNA oligomer of Chemical Formula 1 can include 1 or 2 mismatches in the complementary binding region with a part of the coronavirus genome RNA, when n = 20, the PNA oligomer of Chemical Formula 1 can include 1 or 4 mismatches in the complementary binding region with a part of the coronavirus genome RNA, and when n = 30, the PNA oligomer of Chemical Formula 1 can include 1 or 6 mismatches in the complementary binding region with a part of the coronavirus genome RNA. In this case, the mismatches can be continuous and / or discontinuous.

[0069] In Chemical Formula 1, X is one selected from the group consisting of hydrogen [H], formyl [H-C(=0)-], aminocarbonyl [NH2-C(=0)-], a substituted or unsubstituted alkyl group, a substituted or unsubstituted aryl group, a substituted or unsubstituted alkylacyl group, a substituted or unsubstituted arylacyl group, a substituted or unsubstituted alkoxycarbonyl group, a substituted or unsubstituted aryloxycarbonyl group, a substituted or unsubstituted alkylaminocarbonyl group, a substituted or unsubstituted arylaminocarbonyl group, a substituted or unsubstituted alkylsulfonyl group, and a substituted or unsubstituted arylsulfonyl group.

[0070] In Chemical Formula 1, Z is one selected from the group consisting of hydroxy [-OH], a substituted or unsubstituted alkoxy group, a substituted or unsubstituted aryloxy group, amino [-NH2], a substituted or unsubstituted alkylamino group, a substituted or unsubstituted arylamino group, a substituted or unsubstituted alkyl group, and a substituted or unsubstituted aryl group.

[0071] Substituents for describing the PNA derivative of Chemical Formula 1 are described below.

[0072] Examples of the substituted or unsubstituted alkyl group are as follows.

[0073] Examples of unsubstituted alkyl radicals

[0074]

[0075] Examples of substituted alkyl radicals

[0076]

[0077] Examples of the substituted or unsubstituted alkylacyl group are as follows.

[0078] Examples of unsubstituted alkyl acyl radicals

[0079]

[0080] Examples of substituted alkyl acyl radicals

[0081]

[0082] Examples of the substituted or unsubstituted arylacyl group are as follows.

[0083] Examples of substituted or unsubstituted aryl acyl radicals

[0084]

[0085] Examples of the substituted or unsubstituted alkylamino and substituted or unsubstituted arylamino groups are as follows.

[0086] Examples of substituted alkyl or aryl amino radicals

[0087]

[0088] Examples of substituted or unsubstituted aryl groups are as follows.

[0089] Examples of substituted or unsubstituted aryl radicals

[0090]

[0091] Examples of substituted or unsubstituted alkylsulfonyl and substituted or unsubstituted arylsulfonyl groups are as follows.

[0092] Examples of substituted or unsubstituted alkyl sulfonyl or aryl sulfonyl radicals

[0093]

[0094] Examples of substituted or unsubstituted alkylphosphonyl and substituted or unsubstituted arylphosphonyl groups are as follows.

[0095] Examples of substituted or unsubstituted alkyl phosphonyl or aryl phosphonyl radicals

[0096]

[0097] Examples of substituted or unsubstituted alkoxycarbonyl groups are as follows.

[0098] Examples of substituted or unsubstituted alkoxycarbonyl radicals

[0099]

[0100] Examples of substituted or unsubstituted aryloxycarbonyl groups are as follows.

[0101] Examples of substituted or unsubstituted aryloxycarbonyl radicals

[0102]

[0103] Examples of substituted or unsubstituted alkylaminocarbonyl groups are as follows.

[0104] Examples of substituted or unsubstituted alkyl aminocarbonyl radicals

[0105]

[0106] Examples of substituted or unsubstituted arylaminocarbonyl groups are as follows.

[0107] Examples of substituted or unsubstituted aryl aminocarbonyl radicals

[0108]

[0109] In addition, the N-terminal and C-terminal substituents of Chemical Formula 1 can include the following substituents.

[0110]

[0111] The abbreviations of the substituents of the N-terminal and C-terminal are specifically described as follows: "Fmoc-" is an abbreviation of "[(9-fluorenyl)methoxy]carbonyl-"; "Fethoc-" is an abbreviation of "[2-(9-fluorenyl)ethyl-l-oxyl]carbonyl"; "Ac-" is an abbreviation of "acetyl-"; "benzoyl-" is an abbreviation of "benzoyl-"; "Piv-" is an abbreviation of "pivaloyl-"; "Me-" is an abbreviation of "methyl-"; "n-Propyl-" is an abbreviation of "1-(n-propyl)-"; "H-" is an abbreviation of "hydrogen-" group; "p-Toluenesulfonyl" is an abbreviation of "(4-methylphenyl)-l-sulfonyl-"; "-Lys-" is an abbreviation of the amino acid residue "lysine"; "-Val-" is an abbreviation of the amino acid residue "valine"; "-Leu-" is an abbreviation of the amino acid residue "leucine"; "-Arg-" is an abbreviation of the amino acid residue "arginine"; "-Gly-" is an abbreviation of the amino acid residue "glycine"; "[N-(2-Phenylethyl)amino]carbonyl-" is an abbreviation of "[N-l-(2-phenylethyl)amino]carbonyl-"; "Benzyl-" is an abbreviation of "1-(phenyl)methyl-"; "Phenyl-" is an abbreviation of "phenyl-"; "Me-" is an abbreviation of "methyl-"; "-HEX-" is an abbreviation of "6-amino-l-hexanoyl-"; "FAM-" is an abbreviation of "5- or 6-fluorescein carbonyl- (isomer mixture)"; and "-NH2" is an abbreviation of an unsubstituted "-amino" group.

[0112] The examples of the substituents of the N-terminal and C-terminal of the compound of Chemical Formula 1 in the present application are intended to show only the diversity of the allowable substituents, and do not limit the scope of the present application. In particular, for the binding to the target RNA, the sequence of the PNA oligomer is more important than the substituents of the N-terminal and / or C-terminal of the PNA oligomer.

[0113] In addition, in Chemical Formula 1, B1, B2,..., B n-1 and B n are each independently selected from the group consisting of natural nucleobases of adenine (A), guanine (G), thymine (T), or cytosine (C) and unnatural nucleobases, and one or more of B1to B n include modified nucleobases.

[0114] The inventors constructed 20-mer PNA oligomers complementary to the target region based on the HCoV-OC43 RNA sequence provided by NCBI (NCBI Reference Sequence: NC_006213) (Table 1).

[0115] [Table 1]

[0116]

[0117] Furthermore, the inventors constructed 20-mer PNA oligomers complementary to the target region based on the SARS-CoV-2 RNA sequence provided by NCBI (NCBI Reference Sequence: NC_045512) (Table 2).

[0118] [Table 2]

[0119]

[0120] In the PNA sequences of Tables 1 and 2 above, X (A, G, T or C) is each a natural nucleobase, X(p) or X(pOq) is a modified nucleobase, Fethoc- is [2-(9-fluorenyl)ethyl-1- oxy]carbonyl], and the substituent at the N-terminus, in addition to Fethoc-, can also be Fmoc- [(9-fluorenyl)methoxy]carbonyl]-, acetyl (Ac), benzoyl or hydrogen, the structures of which are as follows.

[0121]

[0122] .

[0123] At the same time, the PNA derivatives in the following Table 3 target sequences that are conserved in multiple coronaviruses. Specifically, PNA 35 targets a sequence common to HCoV-OC43 and SARS-CoV-2, and PNA 53 targets a sequence common to HCoV-OC43, SARS-CoV-2 and SARS-CoV.

[0124] [Table 3]

[0125]

[0126] The present inventors designed and constructed the PNA oligomers of Tables 1 and 2, performed mass spectrometry analysis, and compared the expected mass value of the PNA oligomer designed at the time of design with the mass value of the constructed PNA, thereby confirming that the designed PNA oligomer was constructed (see Example 2-1 and Example 3-1).

[0127] Next, the present inventors confirmed whether the constructed PNA oligomers can inhibit viral proliferation. First, to confirm whether the constructed PNA oligomers targeting HcoV-OC43 RNA can bind to viral genomic RNA and efficiently inhibit replication and translation thereof, qRT-PCR was performed to confirm the level of viral genes in the viral culture solution treated with the PNA oligomers. In this case, it was also confirmed whether enhanced viral RNA replication inhibitory effect can be obtained by the combination of various PNA oligomers. To compare the above efficacy, a commercially available drug PF07321332 was used as a control. As a result, it was confirmed that the majority of the constructed PNA oligomers can inhibit the replication of HcoV-OC43 viral RNA, and in addition, the combination of PNA oligomers showed enhanced viral proliferation inhibitory activity (see Example 2-2).

[0128] Next, it was confirmed whether the constructed PNA oligomers targeting SARS-CoV-2 RNA can bind to viral genomic RNA and efficiently inhibit replication thereof. In this case, it was also confirmed whether enhanced viral RNA replication and translation inhibitory effect can be obtained by the combination of various PNA oligomers. As a result of the qRT-PCR experiment, it can be confirmed that, as with the PNA oligomers targeting HcoV-OC43, the PNA oligomers targeting SARS-CoV-2 effectively inhibited the replication of the target viral genes, and in particular, the combination of various PNA oligomers more effectively inhibited viral proliferation. In addition, 20 drugs that efficiently inhibited viral proliferation were selected based on the qRT-PCR results and a titer analysis was performed, and as a result, it was confirmed that 19 drugs showed a viral proliferation inhibition rate of 90% or more at 1000 nM.

[0129] Meanwhile, the present inventors intended to confirm whether the designed and constructed PNA oligomers can be universally used for coronaviruses. It was assumed that the modified PNA can maintain the binding affinity to the target even when there is a mismatch in the complementary binding region, thereby inhibiting viral proliferation, and as a result of confirming the viral protein expression level by treating the SARS-CoV-2 culture solution with a PNA oligomer having a sequence 100% complementary to a part of the HcoV-OC43 RNA, it can be confirmed that the PNA oligomers targeting HcoV-OC43 work even on the SARS-CoV-2 genome containing some mismatches and inhibit the protein expression thereof.

[0130] In summary, the present inventors constructed an oligomer capable of complementary binding to a part of the coronavirus genome using a modified PNA, which enabled the oligomer to penetrate into virus-infected cells without a delivery vehicle and inhibit virus proliferation, and in addition, since the PNA oligomer functions even when a mismatch is included in the complementary binding region, the present invention can be provided as a viral therapeutic agent that can be universally used even when the coronavirus mutates.

[0131] The compounds of Formula I of the present invention do not require a separate invasive agent to facilitate systemic delivery to achieve the intended therapeutic or biological activity. In the present application, an invasive agent is described as a "delivery vehicle" for enhancing cell penetration. When the compounds of Formula I are typically dissolved in a phosphate buffered solution (PBS) or a salt solution and then administered systemically, they exhibit the desired therapeutic (analgesic) or biological activity in target tissues, primarily neural cells. The compounds of the present invention do not require a separate strong invasive agent to achieve systemic therapeutic activity.

[0132] Meanwhile, in vivo administration of oligonucleotides for treating diseases has various difficulties due to decomposition by enzymes in blood, intercombination with substances in blood, and non-specific delivery to cells. In order to overcome these difficulties, attempts have been made to improve the delivery ability by using nuclease-resistant analogs or linking structures such as PEG, and this has led to the development of viral vector-based and non-viral delivery systems. However, viral vector-based delivery systems have problems such as immunogenicity caused by the body's immune system, unwanted genomic integration, size limitations of the nucleic acid to be loaded, difficulties in repeated administration, risks of complications, and high production costs, etc. Meanwhile, in the case of non-viral-based delivery systems (e.g., polymers, lipids, and lipid nanoparticles (LNP)) that encapsulate and deliver nucleic acids, lipid-based structures utilizing cholesterol or PEG and polymer-based structures (e.g., PLGA) are being used, but these structures have toxicity problems and are limited in that they can only be delivered to specific organs. In addition, even in the case of non-natural oligonucleotides for improving stability, there is a problem of requiring a delivery vehicle to penetrate the cell membrane.

[0133] The compounds of Formula I of the present invention do not require a separate delivery vehicle to achieve cell membrane penetration, and thus can inhibit the replication of intracellular coronaviruses even when administered systemically. Therefore, the compounds of the present invention can be used to safely treat symptoms associated with coronavirus infectious diseases.

[0134] The compounds of Formula I of the present invention can be used with pharmaceutically acceptable acids or bases, and pharmaceutically acceptable acids or bases include, but are not limited to, sodium hydroxide, potassium hydroxide, hydrochloric acid, methanesulfonic acid, citric acid, trifluoroacetic acid, etc.

[0135] In this invention, the compound of formula I or a pharmaceutically acceptable salt may be administered together with a pharmaceutically acceptable excipient, and the excipient includes, but is not limited to, citric acid, hydrochloric acid, tartaric acid, stearic acid, polyethylene glycol, polypropylene glycol, ethanol, isopropanol, sodium bicarbonate, distilled water, preservatives, etc.

[0136] [Embodiments of the Invention]

[0137] Because this invention can be modified in various forms and includes various exemplary embodiments, specific exemplary embodiments will be shown in the accompanying drawings and described in detail below. However, this description is not intended to limit the invention to the specific exemplary embodiments, and it should be understood that all changes, equivalents, and substitutions within the spirit and scope of the invention are included in this invention. In describing the invention, detailed descriptions of relevant prior art will be omitted where it is determined that such detailed descriptions may obscure the essential points of the invention.

[0138] Example

[0139] Example 1. General method for synthesizing PNA oligomers

[0140] PNA monomers with modified nucleobases were synthesized using the method disclosed in related technology [PCT / KR 2009 / 001256] or a slightly modified method thereof. Using Fmoc monomers with modified nucleobases, i.e., PNA monomers protected with [{(9-fluorenyl)methoxy}carbonyl]- and Fmoc monomers with native nucleobases protected with Fmoc, PNA oligomers of formula (I) were synthesized in a solid phase using the method disclosed in related technology [US6,133,444; WO96 / 40685] or a slightly modified method thereof, as shown in [Reaction Scheme 1]. H-Rink Amide-ChemMatrix resin, purchased from PCAS BioMatrix Inc. (Quebec, Canada), was typically used as the solid phase for synthesis. The synthesized PNA oligomers were identified by TOF LC / MS and analyzed using C 18 - Separation and analysis were performed by reversed-phase UPLC. (Distilled water / acetonitrile or distilled water / methanol, 0.1% TFA) Figure la and Figure lb [These are HPLC chromatograms of "PNA123" before and after HPLC purification, and] Figure lcFigure 1 is an ESI / TOF / MS spectrum of "PNA123" purified by HPLC, and these are provided for the purpose of describing the PNA oligomers of the present invention, and are not intended to limit the scope of the present invention.

[0141] [Reaction Scheme 1] is a schematic of a typical process of solid phase synthesis, and each reaction process is briefly provided as follows.

[0142] [Activation of H-Rink-ChemMatrix resin] In the case where the Fmoc protecting group is attached to the amine of the resin, the suspension (where 0.94 mmol (about 200 mg of resin) and 5 mL of 20% piperidine / dimethylformamide (DMF) are mixed) is vortexed in a libra tube for 5 minutes, and then filtered and the DeFmoc solution is removed. When the resin is washed with 5 mL of dichloromethane (MC), 5 mL of DMF, and 5 mL of DMF for 30 seconds each, the amine of the resin is ready to react with the Fmoc-protected PNA monomer.

[0143] [Reaction Scheme 1]

[0144]

[0145] [Fmoc removal (DeFmoc)] In the case where the Fmoc protecting group is attached to the amine of the resin, the suspension (where 0.94 mmol (about 200 mg of resin) and 5 mL of 20% piperidine / dimethylformamide (DMF) are mixed) is vortexed in a libra tube for 5 minutes, and then filtered and the DeFmoc solution is removed. When the resin is washed with 5 mL of dichloromethane (MC), 5 mL of DMF, and 5 mL of DMF for 30 seconds each, the amine of the resin is ready to react with the Fmoc-protected PNA monomer.

[0146] [Coupling with Fmoc-PNA monomer] The amine of the resin is coupled with the Fmoc-PNA monomer by the following method. 0.235 mmol of the PNA monomer, 0.235 mmol of HBTU, 0.2585 mmol of DIEA, etc. are dissolved in 5 mL of anhydrous DMF, and then the solution is added to the resin after 2 minutes. After the resin suspension is vortexed for 1 hour, the solution is filtered and removed, and the resin is washed with 5 mL of MC, 5 mL of DMF, 5 mL of MC, 5 mL of DMF, and 5 mL of DMF for 30 seconds each.

[0147] [Capping] After the coupling reaction, the uncoupled amine is capped by reacting in 5 mL of a capping solution (5% acetic anhydride and 6% of a 2,6-dimethylpyridine DMF solution) for 5 minutes. After the capping solution is filtered and removed, the resin is washed with 5 mL of MC, 5 mL of DMF, and 5 mL of DMF for 30 seconds each.

[0148] [Introduction of the “Fethoc” group to the N-terminus] The “Fethoc” group was introduced to the N-terminal amine of the resin under basic coupling conditions by reaction with “Fethoc-Osu” [CAS number 179337-69-0, C 20 H 17 The chemical structure of NO5, MW 351.36, is as follows.

[0149]

[0150] [Isolation from the resin] The PNA oligomer bound to the resin was isolated from the resin by reaction in 6 mL of cleavage solution (2% triisopropylsilane and 2% trifluoroacetic acid in water) for 3 hours. The resin was filtered off and the filtrate was concentrated under low pressure. The solid obtained by treating the residue with diethyl ether was obtained by filtration and then purified by reverse phase HPLC.

[0151] [HPLC analysis and purification] The crude PNA oligomer product isolated from the resin was analyzed and purified by C 18 - Reverse phase HPLC purification using water / acetonitrile or water / methanol containing 0.1% TFA as eluent. Figure la and Figure lb are C 18 - Examples of reverse phase UPLC chromatograms.

[0152] Synthesis examples of PNA derivatives of formula I

[0153] By partial modification of the above method, PNA oligomers complementary to the coronavirus genomic RNA were synthesized. These PNA oligomers targeting coronavirus RNA are provided to illustrate the content of the present invention and are not intended to limit the scope of the invention.

[0154] Table 1 and Table 2 above show PNA oligomers targeting HcoV-OC43 RNA or SARS-CoV-2 RNA.

[0155] Table 4 below shows PNA oligomers among the constructed PNA oligomers that are fully complementary to HcoV-OC43 RNA and Table 5 shows PNA oligomers among the constructed PNA oligomers that are fully complementary to SARS-CoV-2 RNA.

[0156] [Table 4]

[0157]

[0158] [Table 5]

[0159]

[0160] Example 2. Analysis of PNA oligomers targeting HCoV-OC43 RNA

[0161] 2-1. Mass structure analysis

[0162] Table 6 below shows the results of the mass structure analysis of the constructed PNA oligomers targeting HCoV-OC43.

[0163] [Table 6]

[0164]

[0165] a) Theoretical value, b) Observed value.

[0166] 2-2. Evaluation of the antiviral efficacy of PNA oligomers using qRT-PCR

[0167] The efficacy of the drugs based on the constructed PNA oligomers was evaluated to determine how well the drugs could block viral gene replication.

[0168] In addition, to confirm whether the antiviral efficacy is enhanced during treatment with a combination of PNA oligomers, experiments were also performed using a drug in which PNA 46 and PNA 8 were combined (Combi 1).

[0169] To analyze the level of viral RNA in the viral particles present in the viral culture solution to determine how well the constructed PNA oligomer-based drugs inhibit viral genetic replication, qRT-PCR was performed.

[0170] Specifically, HCT8 cells cultured in 6-well plates one day before drug screening were infected with HCoV-OC43 at a dose of 500 TCID50 using a serum-free RPMI medium. At the same time of infection, the cells were also treated with 1 µM of PNA oligomers, and 30 minutes later, the cells were washed twice with serum-free RPMI, and then the medium was replaced with RPMI-complete medium (containing 10% FBS and 1% penicillin / streptomycin), and the cells were treated with 1 µM of PNA oligomers. On day 3 after infection, viral RNA was extracted from 140 µL of cell supernatant using the QIAmp Viral RNA Mini Kit manufactured by Qiagen. Viral extraction was performed according to the kit manual. The viral titer of the extracted viral RNA was measured by real-time PCR using the One step TB green kit manufactured by Takara. The primers used are as follows. HCoV-OC43 primers; nucleoprotein (NP) gene (5'-AGCAACCAGGCTGATGTCAATACC-3', 5'-AGCAGACCTTCCTGAGCCTTCAAT-3'). The real-time PCR conditions are as follows. Temperature phase 1: reverse transcription 42°C for 5 minutes, 95°C for 10 seconds, 1 cycle. Phase 2: PCR reaction 95°C for 5 seconds, 60°C for 34 seconds, 35 cycles. Phase 3: melting curve analysis 95°C for 15 seconds, 65°C for 60 seconds, 95°C for 15 seconds.

[0171] The results of the evaluation of the antiviral efficacy of the PNA oligomers targeting HCoV-OC43 and their combinations through qRT-PCR-based viral RNA level analysis are shown in Table 7 below.

[0172] [Table 7]

[0173]

[0174] Example 3. Analysis of PNA oligomers targeting SARS-CoV-2 RNA

[0175] 3-1. Mass structure analysis and target binding affinity confirmation

[0176] Next, quality structure analysis was performed to confirm whether the designed PNA oligomers targeting SARS-CoV-2 were constructed as intended, and Tm analysis was performed to confirm the binding affinity of each PNA oligomer to the target. Each PNA oligomer designed in Table 2 above exhibited a quality value consistent with the predicted value at the time of design and the actually measured value, indicating that all oligomers were constructed as intended, and some data are provided in Table 8 below. At the same time, Table 9 shows some results of Tm measurement experiments to confirm the binding affinity of PNA oligomers to the target. Since 20-mer PNA oligomers showed high binding affinity even at 90°C or higher, Tm values were measured using 10-mer at the N-terminus and 10-mer at the C-terminus of PNA to compare the relative binding affinity of each PNA oligomer.

[0177] The Tm value of the duplex of 10-mer DNA complementary to PNA was so high that it was difficult to reliably measure the value in an aqueous buffer solution because the aqueous buffer solution tended to boil during Tm measurement. Therefore, the Tm value was measured using a UV / Vis spectrophotometer as follows. In a 15 mL polypropylene Falcon tube, 4 μM of PNA oligomer and 4 μM of 10-mer DNA at the N-terminus were mixed in a buffer solution (pH 7.16, 10 mM sodium phosphate, 100 mM NaCl), then incubated at 90°C for 1 minute and slowly cooled to room temperature. The solution was transferred to a 3 mL quartz cuvette, sealed well, and mounted on an Agilent Cary100 UV / Visible spectrophotometer. While increasing the temperature of the cuvette at a rate of 0.5°C or 1.0°C per minute, the change in absorbance at 260 nm was measured, and the temperature at which the rate of change in absorbance was maximum, i.e., the inflection point, was determined as the Tm value between PNA oligomer and DNA. The DNA used for Tm measurement was purchased from BIONEER (www.bioneer.com, Korea, Daejeon) and used without separate purification.

[0178] [Table 8]

[0179]

[0180] [Table 9]

[0181]

[0182] 3-2. Evaluation of antiviral efficacy

[0183] The efficacy of a drug based on the constructed PNA oligomer was evaluated to determine how well the drug could block viral protein production and gene replication.

[0184] In addition, to confirm whether the antiviral efficacy was enhanced during combination therapy with multiple PNA oligomers, experiments were conducted in which the drugs were combined as shown in Table 10 below (Multi).

[0185] [Table 10]

[0186]

[0187] 3-2-1. qRT-PCR

[0188] To analyze the viral RNA levels in viral particles present in viral culture medium and to determine how well the constructed PNA-based oligomer-based drug inhibits viral genetic replication, infectious viral genes were quantified by qRT-PCR in viral culture medium treated with a test substance (drug) at a concentration of 1000 nM.

[0189] For this purpose, Vero cells cultured in 12-well plates the previous day were pre-incubated for 2 hours in cell culture medium containing a predetermined concentration (1000 nM) of the evaluation drug, then infected with the virus, and the cell culture medium was treated with the same concentration of the evaluation drug. Twenty-four hours post-infection, viral RNA was extracted from the viral culture medium and quantified by qPR-PCR to compare and analyze antiviral efficacy, thereby selecting therapeutic agents. Paxlovid, used as a positive control, was treated in the same manner.

[0190] The screening results are shown in Table 11 below.

[0191] It can be confirmed that among the 50 PNA and Multi drugs, the 20 drugs shown in bold in Table 11 have a viral inhibition rate equal to or higher than Paxlovid's 96.03%.

[0192] [Table 11]

[0193]

[0194] 3-2-2. Titer Analysis

[0195] The 50% inhibitory concentration (IC50) was calculated for the 20 drugs that showed excellent viral replication inhibition efficacy in Example 3-2-1.

[0196] For this purpose, Vero cells were cultured in 12-well plates and pre-incubated for 2 hours in cell culture fluid containing each concentration of the evaluation drug. Thereafter, the cells were infected with the virus and the same concentration of the evaluation drug was added to the cell culture fluid of each well. After 24 hours, the viral RNA was extracted from the virus culture fluid and quantified by qPR-PCR to analyze the antiviral efficacy. The treatment concentrations of the test substances used in the experiment were set as shown in Table 12 below, and a 2-fold dilution was used.

[0197] [Table 12]

[0198]

[0199] The results of the three repeated experiments are shown in Table 13 and Figures 2 to 3b below.

[0200] [Table 13]

[0201]

[0202] The IC50 of Paxlovid used as a positive control in each experiment was confirmed to be 316.6, 328.3, and 464.4 nM, respectively Figure 2 ).

[0203] As a result of deducing the IC50, it was again confirmed that 19 out of 20 had a viral inhibition rate of 90% or more at 1000 nM. In addition, it has been confirmed that when the concentration of the test substance is less than 250 nM, the inhibition rate does not exceed 50% or the antiviral efficacy disappears Figures 3a to 3i and Figures 4a to 4k ).

[0204] Multi 18 drug is the only drug confirmed to have a viral inhibition rate of less than 90% at 1000 nM. In addition, since Multi 28 and Multi 29 do not show a dose-dependent pattern, the IC50 is very low or not derived, i.e., 61.54 nM and not definite, respectively. Therefore, the IC50 for re-verification was not derived from these three substances.

[0205] The results of deducing the IC50 for 17 substances except for the excluded 3 substances, the IC50 range of the test substances was confirmed to be 215.1 to 690.1 nM (Table 9). Among them, the IC50 of Multi 27, Multi 36, and Multi 26 was confirmed to be the best. As a single drug, PNA 123 was also confirmed to show a better IC50 than Paxlovid.

[0206] Example 4. Inhibition of proliferation of SARS-CoV-2 by PNA oligomers targeting HCoV-OC43 RNA

[0207] To confirm whether viral replication can be inhibited even when there is a mismatch in the complementary binding region between the PNA oligomer and the target gene, it was confirmed whether the PNA oligomer targeting the HCoV-OC43 RNA inhibits protein translation of the SARS-CoV-2 virus. Accordingly, SARS-CoV-2 virus culture solution was treated with three types of PNA oligomers (PNA69, 53, 46) designed to be complementary to a portion of the HCoV-OC43 RNA, and Western blotting was performed to confirm the expression level of the N protein of the virus.

[0208] Specifically, for drug screening, HCT8 cells cultured in a 6-well plate the previous day were treated with 1 μM PNA derivatives, while replacing the culture medium with fresh culture medium. Thereafter, Myc-NSP14 was transfected using Lipofectamin 2000 according to the instructions. Forty-eight hours after transfection, the cells were collected, lysed with M-PER lysis solution, and centrifuged, and then a Western blot sample was prepared from the supernatant using 4x sample buffer. Analysis was performed on a 12% SDS-PAGE gel. The antibodies used were anti-myc (cell signaling technology) and anti-β actin (Merck). Detection was performed using an ECL detection reagent (AbClon), visualization and analysis were performed using an Amersham ImageQuant 800, and the results are shown in FIG. 6. Figure 5 .

[0209] Although the embodiments have been described with limited drawings as described above, various technical modifications and variations can be applied by those skilled in the art from the above description. For example, even if the described techniques are performed in an order different from the above-described order, and / or the described components such as systems, structures, devices, and circuits are combined in a different manner from the above-described manner, or the components are replaced or substituted by other components or their equivalents, appropriate results can be achieved.

[0210] Therefore, other implementations, other embodiments, and equivalents of the claims also belong to the scope of the claims to be described below.

[0211] [Industrial applicability]

[0212] The modified PNA oligomer of the present application can be used as an effective coronavirus therapeutic agent capable of responding to rapidly mutating viral infectious diseases.

Claims

1. A pharmaceutical composition for the prevention or treatment of coronavirus infectious diseases, said pharmaceutical composition comprising, as an active ingredient, a peptide nucleic acid (PNA) oligomer represented by the following chemical formula 1, or a pharmaceutically acceptable salt thereof. in, The PNA oligomers are capable of binding complementaryly to a portion of the coronavirus's genomic (+) RNA or genomic (-) RNA: [Chemical Formula 1] In the formula, n is an integer between 10 and 30; X is selected from the group consisting of: hydrogen [H], formyl [HC(=O)-], aminocarbonyl [NH2-C(=O)-], substituted or unsubstituted alkyl, substituted or unsubstituted aryl, substituted or unsubstituted alkyl acyl, substituted or unsubstituted aryl acyl, substituted or unsubstituted alkoxy carbonyl, substituted or unsubstituted aryloxy carbonyl, substituted or unsubstituted alkylamino carbonyl, substituted or unsubstituted arylamino carbonyl, substituted or unsubstituted alkylsulfonyl and substituted or unsubstituted arylsulfonyl; Z is selected from the group consisting of: hydroxyl [-OH], substituted or unsubstituted alkoxy, substituted or unsubstituted aryloxy, amino [-NH2], substituted or unsubstituted alkylamino, substituted or unsubstituted arylamino, substituted or unsubstituted alkyl and substituted or unsubstituted aryl. B1, B2, ..., B n-1 and B n Each is an independent natural or non-natural nucleotide selected from adenine (A), guanine (G), thymine (T), or cytosine (C); B1 to B n One or more of them are independently selected from the group consisting of modified non-natural nucleobases of chemical formulas 2 to 4 below; [Chemical Formula 2] [Chemical Formula 3] [Chemical Formula 4] In chemical formulas 2 to 4, j, k, l, and m are each an integer between 1 and 16.

2. The pharmaceutical composition according to claim 1, wherein, The pharmaceutical composition does not contain a delivery medium for intracellular penetration of the PNA oligomers.

3. The pharmaceutical composition according to claim 1, wherein, X is: Fmoc, i.e., [(9-fluorenyl)methoxy]carbonyl], Fethoc, i.e., [2-(9-fluorenyl)ethyl-1-oxy]carbonyl], acetyl (Ac), benzoyl or hydrogen.

4. The pharmaceutical composition according to claim 1, wherein, X is Fethoc, i.e., [2-(9-fluorenyl)ethyl-1-oxy]carbonyl. Z is NH2. j is 2. k is 1, l is 5, and m is 6.

5. The pharmaceutical composition according to claim 1, wherein, The coronavirus is selected from one or more of the group consisting of human coronavirus 229E (HCoV-229E), human coronavirus OC43 (HCoV-OC43), severe acute respiratory syndrome coronavirus (SARS-CoV), human coronavirus NL63 (HCoV-NL63), human coronavirus HKU1 (HCoV-HKU1), Middle East respiratory syndrome coronavirus (MERS-CoV), and severe acute respiratory syndrome coronavirus type 2 (SARS-Cov-2).

6. The pharmaceutical composition according to claim 1, wherein, The coronavirus in question is human coronavirus OC43 (HCoV-OC43), and The PNA oligomers are selected from one or more of the PNA oligomers listed in Table 1.

7. The pharmaceutical composition according to claim 1, wherein, The coronavirus in question is type 2 Severe Acute Respiratory Syndrome Coronavirus (SARS-CoV-2), and The PNA oligomers are selected from one or more of the PNA oligomers listed in Table 2.

8. The pharmaceutical composition according to claim 7, wherein, The pharmaceutical composition comprises PNA 123 as shown in Table 2, namely [Fethoc-A(5)GC-C(1O2)CT-G(6)TA-TA(5)C-GA(5)C-ATC(1O2)-AG-NH2].

9. The pharmaceutical composition according to claim 1, wherein, The coronavirus is selected from one or more of the group consisting of human coronavirus OC43 (HCoV-OC43) and severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2), and The PNA oligomers are selected from one or more of the PNA oligomers listed in Table 3.

10. The pharmaceutical composition according to claim 1, wherein, The PNA oligomer has 80% or higher complementarity to a portion of the coronavirus’s genomic (+) RNA or genomic (-) RNA.

11. The pharmaceutical composition according to claim 1, wherein, The PNA oligomers have 90% or higher complementarity to a portion of the coronavirus’s genomic (+) RNA or genomic (-) RNA.

12. A method for preventing or treating infectious diseases caused by coronaviruses, the method comprising administering to an individual a peptide nucleic acid (PNA) oligomer represented by the following chemical formula 1, or a pharmaceutically acceptable salt thereof. in, The PNA oligomers are capable of binding complementaryly to a portion of the coronavirus's genomic (+) RNA or genomic (-) RNA: [Chemical Formula 1] In the formula, n is an integer between 10 and 30; X is selected from the group consisting of: hydrogen [H], formyl [HC(=O)-], aminocarbonyl [NH2-C(=O)-], substituted or unsubstituted alkyl, substituted or unsubstituted aryl, substituted or unsubstituted alkyl acyl, substituted or unsubstituted aryl acyl, substituted or unsubstituted alkoxy carbonyl, substituted or unsubstituted aryloxy carbonyl, substituted or unsubstituted alkylamino carbonyl, substituted or unsubstituted arylamino carbonyl, substituted or unsubstituted alkylsulfonyl and substituted or unsubstituted arylsulfonyl; Z is selected from the group consisting of: hydroxyl [-OH], substituted or unsubstituted alkoxy, substituted or unsubstituted aryloxy, amino [-NH2], substituted or unsubstituted alkylamino, substituted or unsubstituted arylamino, substituted or unsubstituted alkyl and substituted or unsubstituted aryl. B1, B2, ..., B n-1 and B n Each is an independent natural or non-natural nucleotide selected from adenine (A), guanine (G), thymine (T), or cytosine (C); B1 to B n One or more of them are independently selected from the group consisting of modified non-natural nucleobases of chemical formulas 2 to 4 below; [Chemical Formula 2] [Chemical Formula 3] [Chemical Formula 4] In chemical formulas 2 to 4, j, k, l, and m are each an integer between 1 and 16.

13. A method for inhibiting coronavirus replication, said method comprising administering to an individual a peptide nucleic acid (PNA) oligomer represented by the following chemical formula 1 or a pharmaceutically acceptable salt thereof. in, The PNA oligomers are capable of binding complementaryly to a portion of the coronavirus's genomic (+) RNA or genomic (-) RNA: [Chemical Formula 1] In the formula, n is an integer between 10 and 30; X is selected from the group consisting of: hydrogen [H], formyl [HC(=O)-], aminocarbonyl [NH2-C(=O)-], substituted or unsubstituted alkyl, substituted or unsubstituted aryl, substituted or unsubstituted alkyl acyl, substituted or unsubstituted aryl acyl, substituted or unsubstituted alkoxy carbonyl, substituted or unsubstituted aryloxy carbonyl, substituted or unsubstituted alkylamino carbonyl, substituted or unsubstituted arylamino carbonyl, substituted or unsubstituted alkylsulfonyl and substituted or unsubstituted arylsulfonyl; Z is selected from the group consisting of: hydroxyl [-OH], substituted or unsubstituted alkoxy, substituted or unsubstituted aryloxy, amino [-NH2], substituted or unsubstituted alkylamino, substituted or unsubstituted arylamino, substituted or unsubstituted alkyl and substituted or unsubstituted aryl. B1, B2, ..., B n-1 and B n Each is an independent natural or non-natural nucleotide selected from adenine (A), guanine (G), thymine (T), or cytosine (C); B1 to B n One or more of them are independently selected from the group consisting of modified non-natural nucleobases of chemical formulas 2 to 4 below; [Chemical Formula 2] [Chemical Formula 3] [Chemical Formula 4] In chemical formulas 2 to 4, j, k, l, and m are each an integer between 1 and 16.

14. Use of a peptide nucleic acid (PNA) oligomer represented by the following chemical formula 1, or a pharmaceutically acceptable salt thereof, in the preparation of a medicament for the prevention or treatment of coronavirus infectious diseases. in, The PNA oligomers are capable of binding complementaryly to a portion of the coronavirus's genomic (+) RNA or genomic (-) RNA: [Chemical Formula 1] In the formula, n is an integer between 10 and 30; X is selected from the group consisting of: hydrogen [H], formyl [HC(=O)-], aminocarbonyl [NH2-C(=O)-], substituted or unsubstituted alkyl, substituted or unsubstituted aryl, substituted or unsubstituted alkyl acyl, substituted or unsubstituted aryl acyl, substituted or unsubstituted alkoxy carbonyl, substituted or unsubstituted aryloxy carbonyl, substituted or unsubstituted alkylamino carbonyl, substituted or unsubstituted arylamino carbonyl, substituted or unsubstituted alkylsulfonyl and substituted or unsubstituted arylsulfonyl; Z is selected from the group consisting of: hydroxyl [-OH], substituted or unsubstituted alkoxy, substituted or unsubstituted aryloxy, amino [-NH2], substituted or unsubstituted alkylamino, substituted or unsubstituted arylamino, substituted or unsubstituted alkyl and substituted or unsubstituted aryl. B1, B2, ..., B n-1 and B n Each is an independent natural or non-natural nucleotide selected from adenine (A), guanine (G), thymine (T), or cytosine (C); B1 to B n One or more of them are independently selected from the group consisting of modified non-natural nucleobases of chemical formulas 2 to 4 below; [Chemical Formula 2] [Chemical Formula 3] [Chemical Formula 4] In chemical formulas 2 to 4, j, k, l, and m are each an integer between 1 and 16.

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